Primer on the Autonomic Nervous System, Third Edition

PRIMER ON THE AUTONOMIC NERVOUS SYSTEM THIRD EDITION

PRIMER ON THE AUTONOMIC NERVOUS SYSTEM THIRD EDITION Editor In Chief

David Robertson Vanderbilt University

Editors

Italo Biaggioni Vanderbilt University

Geoffrey Burnstock

University College Medical School

Phillip A. Low

Mayo College of Medicine

Julian F.R. Paton University of Bristol

AMSTERDAM • BOSTON • HEIDELBERG • LONDON NEW YORK • OXFORD • PARIS • SAN DIEGO SAN FRANCISCO • SINGAPORE • SYDNEY • TOKYO Academic Press is an imprint of Elsevier

Academic Press is an imprint of Elsevier 32 Jamestown Road, London NW1 7BY, UK 225 Wyman Street, Waltham, MA 02451, USA 525 B Street, Suite 1800, San Diego, CA 92101-4495, USA First edition 1996 Second edition 2004 Third edition 2012 Copyright © 2012 Elsevier Inc. All rights reserved Except the figures of chapter 76 for which the author retains copyright Cover Image: The human autonomic nervous system, which in large part lies deep within the body, is readily accessible through biopsies of the skin. In the cover image, using the cholinergic marker, vasoactive intestinal peptide (shown in red), there is clear visualization of the sympathetic cholinergic innervation running between the endothelial and basal lamina layers of cutaneous blood vessels in a skin biopsy of the distal thigh. The blood vessels are visualized with the endothelial vascular marker CD31 (an endothelial vascular marker that highlights blood vessels – shown in green) and the vascular basal lamina with collagen type IV (a marker that highlights the basal lamina layer of the extracellular matrix – shown in blue). Image courtesy of Ningshan Wang, Christopher Gibbons and Roy Freeman. No part of this publication may be reproduced, stored in a retrieval system or transmitted in any form or by any means electronic, mechanical, photocopying, recording or otherwise without the prior written permission of the publisher. Permissions may be sought directly from Elsevier’s Science & Technology Rights Department in Oxford, UK: phone (44) (0) 1865 843830; fax (44) (0) 1865 853333; email: [email protected]. Alternatively, visit the Science and Technology Books website at www.elsevierdirect.com/rights for further information Notice No responsibility is assumed by the publisher for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions or ideas contained in the material herein. Because of rapid advances in the medical sciences, in particular, independent verification of diagnoses and drug dosages should be made British Library Cataloguing-in-Publication Data A catalogue record for this book is available from the British Library Library of Congress Cataloging-in-Publication Data A catalog record for this book is available from the Library of Congress ISBN : 978-0-12-386525-0 For information on all Academic Press publications visit our website at elsevierdirect.com Typeset by MPS Limited, a Macmillan Company, Chennai, India www.macmillansolutions.com Printed and bound in United States of America 12

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Preface

The Primer on the Autonomic Nervous System aims to provide a concise and accessible overview of autonomic neuroscience for students, scientists, and clinicians. In spite of its compact size, its 144 chapters draw on the expertise of more than 250 scientists and clinicians. We thank the American Autonomic Society for its continued interest and moral support of this project. We especially express appreciation to our contributors, who, along with the editors, prepared their chapters without compensation in order to keep the cost of the Primer within the reach of students and trainees. We are delighted with the enthusiastic reception of the first and second English editions, and the Japanese edition of the Primer, which has sold more copies than any previous text on autonomic neuroscience. With this edition we welcome Julian F.R. Paton as a new editor. The third edition of the Primer would not have been possible without Mrs. Sonja Campbell, whose efficiency

and wisdom, combined with her mastery of lucid English prose, facilitated the preparation of this substantially enlarged edition. We also thank Mica Haley and Melissa Turner at Academic Press, who kept all of us on track and on schedule. In earlier editions, readers were encouraged to email their criticisms and advice for improving the text. We thank the many of you who took time to do just that. Several new sections and a number of clarifications based on these suggestions have been implemented. If you have comments or advice for improving future editions please send them to [email protected].

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David Robertson Italo Biaggioni Geoffrey Burnstock Phillip A. Low Julian F.R. Paton

List of Contributors

Ana P.L. Abdala School of Physiology and Pharmacology, Bristol Heart Institute, University of Bristol, Bristol, UK David H. Adams Guggenheim Pavillion, New York, NY, USA Marlies Alvarenga Baker IDI Heart and Diabetes Institute, Melbourne, Australia Amy C. Arnold Division of Clinical Pharmacology, Vanderbilt University School of Medicine, Nashville, TN, USA Felicia B. Axelrod Professor, Pediatrics and Neurology, New York University School of Medicine, New York, NY 10016, USA Franca Barbic Neuroscience Research Association, Bolognini Hospital, Seriate (Bg), Italy Peter J. Barnes Department of Thoracic Medicine, National Heart and Lung Institute, London, UK Deborah Bauer Departments of Pediatrics and Pharmacology, Children’s Hospital of Philadelphia, University of Pennsylvania, Philadelphia, PA 19104, USA Christopher Bell Department of Health and Exercise Science, Colorado State University, Fort Collins, CO, USA Eduardo E. Benarroch Department of Neurology, Mayo Clinic, Rochester, MN, USA Elizabeth M. Berry-Kravis Professor of Pediatrics, Neurology, and Biochemistry at Rush University Medical Center, Chicago, IL 60612, USA Luciano Bernardi Clinica Medica 2 – Dipartimento Medicina Interna, IRCCS S. Matteo, Universita’ di Pavia, 27100 Pavia, Italy Italo Biaggioni Professor of Medicine and Pharmacology, Vanderbilt University, Nashville, TN 37212, USA Lori Birder University of Pittsburgh School of Medicine, Departments of Medicine and Pharmacology, Pittsburgh PA 15261, USA Virginia L. Brooks Department of Physiology and Pharmacology, Oregon Health and Science University, Portland, OR 97239, USA Joan Heller Brown Department of Pharmacology, University of California, San Diego School of Medicine, La Jolla, CA, USA Geoffrey Burnstock Autonomic Neuroscience Centre, University College Medical School, London NW3 2PF, UK Michael Camilleri Mayo Clinic, Rochester, MN, USA J. Preston Campbell Vanderbilt University Medical Center, Nashville, TN 37232, USA Robert M. Carey Division of Endocrinology, University of Virginia Health Systems, Charlottesville, VA, USA Marc G. Caron Department of Cell Biology, Duke University Medical Center, Durham, NC 27710, USA Calvin Carter Department of Neurology, University of Iowa, College of Medicine, Iowa City, IA, USA Priscila A. Cassaglia Department of Physiology and Pharmacology, Oregon Health and Science University, Portland, OR 97239, USA Javier G. Castillo Resident Physician, Department of Cardiothoracic Surgery, The Mount Sinai School of Medicine, New York, NY, USA

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00149-9

Mark W. Chapleau Departments of Internal Medicine, and Molecular Physiology and Biophysics, University of Iowa and Veterans Affairs Medical Center, Iowa City, IA, USA Nisha Charkoudian Department of Anesthesiology and Department of Physiology, and Biomedical Engineering, Mayo Clinic, Rochester, MN, USA P. David Charles Movement Disorders Clinic, Medical Center South, Vanderbilt University, Nashville, TN, USA Gisela Chelimsky Department of Pediatrics, Rainbow Babies and Children’s Hospital, and University Hospitals Case Medical Center, Cleveland, OH 44106, USA Thomas Chelimsky Department of Neurology, University Hospitals Case Medical Center, Cleveland, OH 44106, USA Pei-Wen Cheng Department of Medical Education and Research, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan, ROC Gilles Clément International Space University, Strasbourg, France Pietro Cortelli Department of Neurological Sciences, Alma Mater Studiorum, University of Bologna, 40123 Bologna, Italy Allen W. Cowley Department of Physiology, Medical College of Wisconsin, Milwaukee, WI 53226, USA Leslie Crews Department of Pathology, University of California, San Diego/La Jolla, CA, USA Stephen N. Davis Chair, Internal Medicine, University of Maryland, Baltimore, MD, USA Thomas L. Davis Clinical Research Center, Vanderbilt University, Nashville, TN, USA William C. de Groat University of Pittsburgh School of Medicine, Departments of Medicine and Pharmacology, Pittsburgh, PA 15261, USA Vincent G. DeMarco University of Missouri, Diabetes and Cardiovascular Center, and the Harry S. Truman VA Medical Center, Columbia, MO, USA André Diedrich Autonomic Dysfunction Center, Department of Medicine and Department of Biomedical Engineering, Vanderbilt University School of Medicine, Nashville, TN, USA Donald J. DiPette Departments of Medicine (DJD), and Cell Biology and Anatomy (SCS), University of South Carolina School of Medicine, Columbia, SC 29208, USA Debra I. Diz Professor and Director, Hypertension and Vascular Research Center, Wake Forest University School of Medicine, Winston-Salem, NC 27157-1032, USA Marcus J. Drake FRCS(Urol)Bristol Urological Institute, Bristol, UK Rachel C. Drew Heart and Vascular Institute, Penn State College of Medicine, Milton S. Hershey Medical Center, Hershey, PA, USA Matthias Dütsch Department of Neurology, University of Erlangen-Nuremberg, D-91054 Erlangen, Germany, and Department of Neurology, Rummelsberg Hospital, D-90592 Schwarzenbruck, Germany Graeme Eisenhofer Department of Medicine, and Institute of Clinical Chemistry and Laboratory Medicine, University Hospital Carl Gustav Carus, Dresden, Dresden, Germany

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© 2012 Elsevier Inc. All rights reserved.

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LIst of ContrIbutors

Florent Elefteriou Assistant Professor and Director Elect, Vanderbilt Center for Bone Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA Fernando Elijovich Professor of Medicine, Texas A&M HSC, College of Medicine and Center for Neuroscience, USA Brett A. English Division of Allergy, Pulmonary and Critical Care Medicine, Drug Discovery, Vanderbilt University Medical Center, Nashville, TN, USA Murray Esler Baker IDI Heart and Diabetes Institute, Melbourne, Australia John Y. Fang Assistant Professor, Department of Neurology, Vanderbilt University, Nashville, TN, USA and Staff Physician, Neurology Service, Tennessee Valley Healthcare System, Nashville, TN, USA Robert D. Fealey Department of Neurology, Mayo Clinic, Rochester, MN, USA Stanley Fernandez Department of Medicine, State University of New York at Buffalo, Buffalo, NY 14215, USA Gregory D. Fink Michigan State University, Department of Pharmacology and Toxicology, East Lansing, MI 48840, USA John S. Floras University Health Network and Mount Sinai Hospital Department of Medicine, University of Toronto, Toronto, Ont., Canada Roy Freeman Center for Autonomic and Peripheral Nerve Disorders, Beth Israel Deaconess Medical Center, Boston, MA 02215, USA Qi Fu Institute for Exercise and Environmental Medicine, Texas Health Presbyterian Hospital Dallas, The University of Texas, Southwestern Medical Center at Dallas, Dallas, TX, USA Liang-Wu Fu Department of Medicine, School of Medicine, University of California, Irvine, Irvine, CA 92697-4075, USA Raffaello Furlan Internal Medicine, Bolognini Hospital, Seriate (Bg), University of Milan, Milan, Italy Alfredo Gamboa Autonomic Dysfunction Center, Vanderbilt University, Nashville, TN, USA Emily M. Garland Division of Clinical Pharmacology, Medical Center North, Vanderbilt University, Nashville, TN, USA Christopher H. Gibbons Center for Autonomic and Peripheral Nerve Disorders, Beth Israel Deaconess Medical Center, Boston MA 02215, USA Michael P. Gilbey Department of Physiology, University College London, London, UK Janice L. Gilden Professor of Medicine, Rosalind Franklin University of Medicine and Science, James A. Lovell Federal Health Care Center, North Chicago, and Saints Mary and Elizabeth Medical Center, Chicago, IL, USA Sid Gilman Department of Neurology, 300 N. Ingalls St. 3D15, Ann Arbor, MI, USA David S. Goldstein National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA Diego A. Golombek Universidad Nacional de Quilmes/ CONICET, Buenos Aires, Argentina Robert M. Graham Victor Chang Cardiac Research Institute, Darlinghurst, Sydney, NSW 2010, Australia Guido Grassi Università Milano-Bicocca, Ospedale San Gerardo, Monza (Milan), Milan, Italy, andIstituto Auxologico Italiano, Milan, Italy

Mark D. Grier Department of Pharmacology, Vanderbilt University, Nashville, TN, USA Jan T. Groothuis Department of Physiology, Radboud University Nijmegen Medical Centre, 6500 HB Nijmegen, The Netherlands, and, Department of Rehabilitation, St Maartenskliniek, 6500 GM Nijmegen, The Netherlands Blair P. Grubb Recanati Autonomic Dysfunction Center, Tel Aviv University, Faculty of Medicine, Tel-Aviv 64239, Israel Maureen K. Hahn Department of Medicine and Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA Julian P.J. Halcox Professor of Cardiology, Cardiff University School of Medicine, Wales Heart Research Institute, Cardiff, CF14 4XN, UK Robert W. Hamill Department of Neurology, University of Vermont, College of Medicine, Burlington, VT, USA Kenneth R. Hande Division of Medical Oncology, Preston Research Building, Nashville, TN, USA Yadollah Harati Department of Neurology, Baylor College of Medicine, Houston, TX, USA David G. Harrison Department of Psychiatry, Vanderbilt University School of Medicine, Nashville, TN, USA Emma C. Hart Department of Anesthesiology, Mayo Clinic, Rochester, MN, USA Jacqui Hastings Baker IDI Heart and Diabetes Institute, Melbourne, Australia Luke A. Henderson Department of Anatomy and Histology, University of Sydney, Sydney, Australia Max J. Hilz Department of Neurology, University of ErlangenNuremberg, D-91054 Erlangen, Germany, and Departments of Neurology, Medicine, Psychiatry, New York University School of Medicine, New York, NY 10016, USA Robert Hoeldtke Division of Endocrinology, West Virginia University, Morgantown, WV, USA Shung Tai Ho Graduate Institute of Medical Science, National Defense Medical Center, Taipei, Taiwan, ROC Peter Hunter Auckland Bioengineering Institute, University of Auckland, New Zealand Keith Hyland Department of Neurochemistry, Medical Neurogenetics, Atlanta, GA, USA Lauren Hyland Department of Neurochemistry, Medical Neurogenetics, Atlanta, GA, USA Shahram Izadyar Department of Neurology, Baylor College of Medicine, Houston, TX, USA Joseph L. Izzo Department of Medicine, State University of New York at Buffalo, Buffalo, NY 14215, USA Edwin K. Jackson University of Pittsburgh School of Medicine, Department of Pharmacology and Chemical Biology, Pittsburgh, PA 15219, USA Giris Jacob Head of Medicine F, Recanati Autonomic Dysfunction Center, Tel Aviv (Sourasky) Medical Center, Tel Aviv University, Faculty of Medicine, Tel-Aviv 64239, Israel Wilfrid Jänig Physiologisches Institut, Christian-AlbrechtsUniversität zu Kiel, Kiel, Germany Megan S. Johnson University of Missouri, Diabetes and Cardiovascular Center, Columbia, MO, USA Carrie K. Jones Department of Pharmacology, and Center for Neuroscience Drug Discovery, Vanderbilt University Medical Center, Nashville, TN, USA James F.X. Jones School of Medicine and Medical Sciences, University College Dublin, Ireland

I. INTRODUCTION

LIst of ContrIbutors

Karen M. Joos Vanderbilt Eye Institute, Vanderbilt University Medical Center, Nashville, TN, USA Jens Jordan Institute of Clinical Pharmacology, Hannover Medical School, Hannover, Germany Jens Jordan Institute of Clinical Pharmacology, Hannover Medical School, 30625 Hannover, Germany Michael J. Joyner Department of Anesthesiology, Mayo Clinic, Rochester, MN, USA Stephen G. Kaler Program in Molecular Medicine, NICHD, Bethesda, Maryland 20892-1853, USA Sergey Kasparov Professor of Molecular Physiology, University of Bristol, Bristol, BS8 1TD, UK Horacio Kaufmann Professor, Neurology, Pediatrics and Medicine, New York University School of Medicine, New York, NY 10016, USA Horacio Kaufmann New York University School of Medicine, New York, NY 10016, USA David Kaye Baker IDI Heart and Diabetes Institute, Melbourne, Australia Ramesh K. Khurana Division of Neurology, Union Memorial Hospital, Baltimore, MD, USA Chun-Hyung Kim Department of Psychiatry, Harvard Medical School, Boston, MA, USA Kwang-Soo Kim Department of Psychiatry, Harvard Medical School, Boston, MA, USA Kazuto Kobayashi Department of Molecular Genetics, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima 960-1295, Japan Nancy L. Kuntz Associate Professor of Pediatrics, Northwestern University Feinberg School of Medicine, Center for Autonomic Medicine in Pediatrics at CMH, Chicago, IL 60614, USA Tomas Konecny Assistant Professor of Medicine, Department of Cardiovascular Diseases and Internal Medicine, Mayo Clinic, Rochester, MN, USA, and ICRC – Department of Cardiovascular Diseases, St Anne’s University Hospital Brno, Brno, Czech Republic Andrew Kontak Division of Cardiology/Hypertension Section, University of Texas Southwestern Medical Center, Dallas, TX, USA Cheryl L. Laffer Associate Professor of Medicine, Texas A&M HSC College of Medicine, USA Andre H. Lagrange Assistant Professor of Neurology, Epilepsy Division, Vanderbilt University, Nashville, TN, USA Nora Laiken Department of Pharmacology, University of California, San Diego School of Medicine, La Jolla, CA, USA Gavin Lambert Baker IDI Heart and Diabetes Institute, Melbourne, Australia Jacques W.M. Lenders Department of Internal Medicine, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands, and Department of Medicine, University Hospital Carl Gustav Carus Dresden, Dresden, Germany Benjamin D. Levine Institute for Exercise and Environmental Medicine, Texas Health Presbyterian Hospital Dallas, The University of Texas, Southwestern Medical Center at Dallas, Dallas, TX, USA Lewis A. Lipsitz Institute for Aging Research, Hebrew Senior Life; Division of Gerontology, Beth Israel Deaconess Medical Center; Harvard Medical School, Boston, MA, USA Julian H. Lombard Department of Physiology, Medical College of Wisconsin, Milwaukee, WI 53226, USA

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John C. Longhurst Departments of Medicine, Physiology and Biophysics, Pharmacology and Biomedical Engineering, University of California, Irvine, Irvine, CA 92697-4075, USA David A. Low Autonomic and Neurovascular Medicine Unit, Faculty of Medicine, Imperial College London at St Mary’s Hospital London, WZ1NY, UK Phillip A. Low Department of Neurology, Mayo Foundation, Rochester, MN 55905, USA Chih Cherng Lu, MD,MS Department of Anaesthesiology, Departments of Tri-Service General Hospital/National Defense Medical Center, Taipei, Taiwan, ROC James M. Luther Division of Clinical Pharmacology, Departments of Medicine and Pharmacology, Vanderbilt University, Nashville, TN, USA Vaughan G. Macefield Professor of Integrative Physiology, School of Medicine, University of Western Sydney, NSW 1797, Australia Belinda H. McCully Department of Physiology and Pharmacology, Oregon Health and Science University, Portland, OR 97239, USA James G. McLeod Department of Medicine, University of Sydney, Sydney, Australia William M. Manger New York University Medical Center, and National Hypertension Association, New York, NY, USA Tadaaki Mano Tokai Central Hospital, Kakamigahara, Gifu, Japan Paul J. Marvar Department of Psychiatry, and Center of Behavioral Sciences, Vanderbilt University School of Medicine, Nashville, TN, USA Eliezer Masliah University of California-San Diego, La Jolla, CA 92093-0624, USA Christopher J. Mathias Autonomic and Neurovascular Medicine Unit, Faculty of Medicine, Imperial College London at St Mary’s Hospital London, WZ1NY, UK and Autonomic Unit, National Hospital for Neurology and Neurosurgery, Queen Square/Institute of Neurology, University College London, London, UK Mark R. Melson Vanderbilt Eye Institute, Vanderbilt University Medical Center, Nashville, TN, USA Douglas F. Milam Department of Urologic Surgery, Vanderbilt University, Nashville, TN, USA Marion C. Mohl Victor Chang Cardiac Research Institute, Darlinghurst, Sydney, NSW, 2010, Australia Yaroslav I. Molkov Department of Neurobiology and Anatomy, Drexel University College of Medicine, Philadelphia, PA, USA Margaret Morris Baker IDI Heart and Diabetes Institute, Melbourne, Australia Shaun F. Morrison Department of Neurological Surgery, Oregon Health and Science University, Portland, OR 97239, USA Toshiharu Nagatsu Department of Pharmacology, School of Medicine, Fujita Health University, Toyoake 470-1192, Japan Charles D. Nichols Department of Pharmacology, LSU Health Sciences Center, New Orleans, LA, USA Lucy Norcliffe-Kaufmann Instructor, Physiology and Neuroscience, New York University School of Medicine, New York, NY 10016, USA Vera Novak Division of Gerontology, Beth Israel Deaconess Medical Center;, Harvard Medical School, Boston, MA, USA Luis E. Okamoto Department of Medicine, Division of Clinical Pharmacology, and the Autonomic Dysfunction Center, Vanderbilt University School of Medicine, Nashville, TN, USA

I. INTRODUCTION

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John W. Osborn University of Minnesota, Department of Integrative Biology and Physiology, Minneapolis, MN 55455, USA Brian A. Parsons Bristol Urological Institute, Southmead Hospital, Bristol, UK Julian F.R. Paton School of Physiology and Pharmacology, Bristol Heart Institute, University of Bristol, Bristol, BS8 1TD, UK Pallavi P. Patwari Assistant Professor of Pediatrics, Northwestern University Feinberg School of Medicine, and Assistant Director, Center for Autonomic Medicine in Pediatrics at CMH, Chicago, IL 60614, USA Cecile L. Phan Department of Neurology, Baylor College of Medicine, Houston, TX, USA Fenna T. Phibbs Department of Neurology, Vanderbilt University Medical Center, Nashville, TN, USA Nanduri R. Prabhakar Institute for Integrative Physiology, and Center for Systems Biology of O2 Sensing, Biological Sciences Division, University of Chicago, IL, USA Amanda C. Peltier Department of Neurology, Division of Neuromuscular, Vanderbilt University, Nashville, TN, USA Sean M. Peterson Department of Cell Biology, Duke University Medical Center, Durham, NC 27710, USA Anthony E Pickering School of Physiology and Pharmacology, Bristol Heart Institute, University of Bristol, Bristol, BS8 1TD, UK J. Howard Pratt Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, USA Kamal Rahmouni University of Iowa, Cardiovascular Center, Iowa City, IA, USA Satish R. Raj Autonomic Dysfunction Unit, Division of Clinical Pharmacology, Departments of Medicine and Pharmacology, Vanderbilt University, Nashville, TN, USA Casey M. Rand Center for Autonomic Medicine in Pediatrics at CMH, Chicago, IL 60614, USA Heinz Reichmann Department of Neurology, University Hospital Carl Gustav Carus Dresden, Dresden, Germany Jeff Richards Baker IDI Heart and Diabetes Institute, Melbourne, Australia L.Jackson Roberts Division of Clinical Pharmacology, Robinson Research Building, Vanderbilt University, Nashville, TN, USA David W. Robertson Vanderbilt University, Nashville, TN, USA Rose Marie Robertson Vanderbilt University, Nashville, TN, USA Michael Robinson Departments of Pediatrics and Pharmacology, Children’s Hospital of Philadelphia, University of Pennsylvania, Philadelphia, PA 19104, USA Ilya A. Rybak Department of Neurobiology and Anatomy, Drexel University College of Medicine, Philadelphia, PA, USA Elaine Sanders-Bush Department of Pharmacology, Vanderbilt University, Nashville, TN, USA Paola Sandroni Deptartment of Neurology, Mayo Clinic, Rochester, MN, USA Kyoko Sato Department of Cardiovascular Control, Kochi Medical School, Nankoku, Japan Takayuki Sato Department of Cardiovascular Control, Kochi Medical School, Japan Irwin J. Schatz John A. Burns School of Medicine, University of Hawaii at Manoa, Department of Medicine, Honolulu, HI, USA

Ernesto L. Schiffrin Department of Medicine, Sir Mortimer B. Davis-Jewish General Hospital, and Lady Davis Institute for Medical Research, McGill University, Montreal, Que., Canada. Ronald Schondorf Department of Neurology, Sir Mortimer B. Davis Jewish General Hospital, Montreal, QC, Canada Rosemary Schwarz Baker IDI Heart and Diabetes Institute, Melbourne, Australia Gino Seravalle Istituto Auxologico Italiano, Milan, Italy. Robert E. Shapiro Department of Neurology, University of Vermont College of Medicine, Burlington, VT, USA Cyndya Shibao Department of Medicine, Division of Clinical Pharmacology, and the Autonomic Dysfunction Center,Vanderbilt University School of Medicine, Nashville, TN, USA Virend Somers Professor of Medicine, Department of Cardiovascular Diseases and Internal Medicine, Mayo Clinic, Rochester, MN, USA Michaela Stampfer Movement Disorders Section, Department of Neurology, University Hospital, Innsbruck, Austria C.Michael Stein Division of Clinical Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA Sylvia Stemberger Division of Clinical Neurobiology, Innsbruck Medical University, Innsbruck, Austria Julian Stewart New York Medical College, Hawthorne, NY, USA Lawrence I. Sinoway Pennsylvania State University College of Medicine, Milton S. Hershey Medical Center, Hershey, PA, USA James R. Sowers University of Missouri, Diabetes and Cardiovascular Center, and the Harry S. Truman VA Medical Center, Columbia, MO, USA Sirisha Srikakarlapudi Department of Medicine, State University of New York at Buffalo, Buffalo, NY 14215, USA Kenji Sunagawa Department of Cardiovascular Medicine, Kyushu University Graduate School of Medical Sciences, Japan Scott C. Supowit Departments of Medicine (DJD), and Cell Biology and Anatomy (SCS), University of South Carolina School of Medicine, Columbia, SC 29208, USA Palmer Taylor Department of Pharmacology, University of California, La Jolla, CA, USA Jane Thompson Baker IDI Heart and Diabetes Institute, Melbourne, Australia Roland D. Thijs Department of Neurology and Clinical Neurophysiology, Leiden University Medical Centre, 2300 RC Leiden, The Netherlands and Department of Neurology, Dutch Epilepsy Clinics Foundation, 2300 RC Hoofddorp, the Netherlands Rhian M Touyz Kidney Research Centre, Ottawa Hospital Research Institute, University of Ottawa, Ottawa, Ont., Canada Daniel Tranel Department of Neurology, University of Iowa, College of Medicine, Iowa City, IA, USA Subbulaxmi Trikudanathan Endocrinology, Diabetes and Hypertension Division, Brigham and Women’s Hospital, and, Harvard Medical School, Boston, MA, USA Ching-Jiunn Tseng Department of Medical Education and Research, Kaohsiung Veterans’ General Hospital, Kaohsiung, Taiwan, ROC Che-Se Tung Department of Physiology, National Defense Medical Center, Taipei, Taiwan, ROC Kiren Ubhi Department of Neurosciences, University of California, San Diego/La Jolla, CA, USA

I. INTRODUCTION

LIst of ContrIbutors

Nikhil Urs Department of Cell Biology, Duke University, Durham, NC 27710, USA Joseph G. Verbalis Georgetown University, Washington, DC 20007, USA Steven Vernino Department of Neurology, UT Southwestern Medical Center, Dallas, TX 75390-9036, USA Ronald G. Victor Hypertension Center, The Heart Institute, Cedars-Sinai Medical Center, Los Angeles, CA, USA Margaret A. Vizzard University of Vermont College of Medicine, Burlington, VT, USA Wanpen Vongpatanasin Division of Cardiology/Hypertension Section, University of Texas Southwestern Medical Center, Dallas, TX, USA B. Gunnar Wallin Institute of Neuroscience and Physiology, Sahlgrenska Academy at Göteborg University, S-41345 Göteborg, Sweden Tobias Wang Zoophysiology, Department of Biological Sciences, Aarhus University, Denmark Qin Wang Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, AL, USA Andrew A. Webster Professor and Chair, Department of Pharmaceutical Sciences, Belmont University School of Pharmacy, Nashville, TN, USA

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Debra E. Weese-Mayer Professor of Pediatrics, Northwestern University Feinberg School of Medicine, and Director, Center for Autonomic Medicine in Pediatrics, at Children’s Memorial Hospital (CMH), Chicago, IL 60614, USA Gregor K. Wenning Movement Disorders Section, Department of Neurology, University Hospital, Innsbruck, Austria Adam Whaley-Connell University of Missouri, Diabetes and Cardiovascular Center, and the Harry S. Truman VA Medical Center, Columbia, MO, USA Wouter Wieling Department of Internal Medicine, Academic Medical Centre, 1105 AZ Amsterdam, The Netherlands Gordon H. Williams Endocrinology, Diabetes and Hypertension Division, Brigham and Women’s Hospital, and, Harvard Medical School, Boston, MA, USA Scott Wood Universities Space Research Association, Houston, TX, USA Michael G. Ziegler UCSD Medical Center, San Diego, CA, USA Daniel B. Zoccal Department of Physiological Sciences, Center of Biological Sciences, Federal University of Santa Catarina, Florianópolis, Santa Catarina, Brazil

I. INTRODUCTION

C H A P T E R

1 Development and Differentiation of Autonomic Neurons Chun-Hyung Kim, Kwang-Soo Kim increasing the sympathetic outflow to the heart and other viscera, the parasympathetic system is responsible for the basal autonomic functions such as heart rate and respiration under normal conditions. The enteric system regulates peristalsis of the gut wall and modulates the activity of the secretary glands. During the last decade, exciting progress has been made with regard to the molecular mechanisms underlying the development of the ANS. Among many different aspects, this review will focus primarily on the transcriptional regulatory code underlying the development and neurotransmitter identity determination of the ANS.

An intricate network of extracellular signals and nuclear transcription factors orchestrates the specification of numerous neuronal phenotypes during development of the vertebrate nervous system. During the last decade or so, impressive progress has been achieved in identifying the extracellular signaling molecules and key transcription factors that critically govern the development and fate determination of the autonomic nervous system (ANS). In particular, the so-called “transcriptional regulatory code” underlying the development and differentiation of the ANS has been elucidated; several key fatedetermining transcription factors such as Mash1, Phox2, AP2 and GATA3, have been identified to be responsible for development of the autonomic nervous system and the neurotransmitter identity specification. One important emerging feature is that those key transcription factors regulate not only development, but also final properties of differentiated neurons such as neurotransmitter identity. In line with this concept, those factors directly or indirectly regulate the expression of both cell type-specific markers as well as pan-neuronal markers. Second, these transcription factors function in an intricate regulatory cascade, starting from key signaling molecules such as bone morphogenic proteins. Finally, as evidenced by the study of dopamine β-hydroxylase gene regulation, multitudes of cell type-specific factors (e.g., Phox2a and 2b) and general transcription factors (e.g., CREB and Sp1) co-operatively regulate the expression of cell type-specific marker genes. This new molecular information will facilitate our understanding of the function of the autonomic nervous system in the normal, as well as in the diseased brain.

THE ANS IS DERIVED FROM NEURAL CREST CELLS During the early developmental period of vertebrate embryo, the neural tube and notochord are formed from the ectodermal and mesodermal layers, respectively. At this time, neural crest cells originate from the dorsolateral edge of the neural plate, and are generated at the junction of the neural tube and the ectoderm (Fig. 1.1). Interactions between the non-neural ectodermal layer and the neural plate critically influence the formation of neural crest cells at their interface [1]. Upon formation, neural crest cells migrate along specific routes to diverse destinations and differentiate into a variety of cell types that include all neurons and glial cells of the peripheral nervous system and the neurons of the gastric mucosal plexi. In addition, some other cell types such as smooth muscle cells, pigment cells, and chromatophores are known to arise from neural crest cells. Much progress has recently been achieved in the identification of signaling molecules and downstream transcription factors that control lineage determination and differentiation of neural crest cells [2,3].

INTRODUCTION The autonomic nervous system (ANS; also called the autonomic division or the autonomic motor system) is one of two major divisions of the peripheral nervous system. The ANS has three major subdivisions that are spatially segregated: the sympathetic, the parasympathetic, and the enteric nervous systems. While the sympathetic system controls the fight-or-flight reactions during emergencies by

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00001-9

SIGNALING MOLECULES REGULATE THE DEVELOPMENTAL PROCESSES OF THE ANS Recent studies have demonstrated that various signaling molecules, e.g., members of the bone morphogenic

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1. DEvElOPmENT AND DIffERENTIATION Of AUTONOmIC NEURONs

development, while its robust activation opposes, even in the presence of BMP-2, SA cell development and the expression of SA lineage-determining genes [9]. Finally, during the last decade, numerous studies demonstrated that different neurotrophic factors such as NGF, GDNF, NT3, and/or their receptor signaling, critically regulate the survival and development of all three divisions of the ANS [10,11].

Neural plate Ectodermis Mesoderm

TRANSCRIPTIONAL REGULATORY CODE UNDERLYING THE DEVELOPMENT AND PHENOTYPIC SPECIFICATION OF THE ANS Notochord

Neural crest

Neural tube

Somite Dorsal aorta

FIGURE 1.1 All cell types of the ANS are derived from neural crest cells. Neural crest cells are formed at the dorsal neural tube and migrate along diverse routes. Depending on their specific routes and interactions with the target tissues, they differentiate into a variety of cell types including pigment cells, different types of neurons and glia of the ANS, and parts of the adrenal gland.

protein (BMP) family, Wnt, sonic hedgehog, and fibroblast growth factor, play critical roles in the early formation of neural crest cells as well as for final determination of neuronal identity [1–5]. These signals are often provided from neighboring tissues during migration of neural crest cells. For instance, grafting and ablation experiments in chick embryos demonstrate that the notochord is necessary but not sufficient to induce adrenergic phenotypes of neural crest-derived sympathetic ganglia [6,7]. A possible candidate for notochord-derived molecule(s) is sonic hedgehog [7]. In addition, a series of elegant experiments established that BMP family members, expressed from the dorsal aorta, play a crucial role in the differentiation and fate determination of the sympathetic nervous system [8 and references therein]. It is of note that these extracellular signaling molecules seem to work in concert with intracellular signals such as cAMP. Consistent with this, using neural crest cell culture, induction of differentiation and neurotransmitter phenotypes by BMP is enhanced by cAMP-elevating agents [4,9]. Interestingly, it appears that cAMP signaling acts as a bimodal regulator of sympathoadrenal (SA) cell development in neural crest cultures because its moderate activation promotes SA cell

Various transcription factors are thought to trigger a regulatory cascade by inducing the expression of downstream transcription factors, which eventually activate or repress the final target genes [4,12]. The regulatory cascade controlling the ANS’s noradrenergic neurotransmitter phenotype has been extensively studied, leading to the identification and functional characterization of critical transcription factors (Fig. 1.2). For example, the basic helix-loop-helix (bHLH) factor, Mammalian achaete-scute homolog-1 (Mash1, and the chicken homolog Cash1) induced by BMPs is the first transcription factor shown to be essential for noradrenergic neuron development. Downstream of Mash1 lies the homeodomain transcription factor Phox2a which is a critical regulator of noradrenergic cell lineage development. A closely related transcription factor Phox2b is also induced by BMPs independently of Mash1 and is another essential regulator of noradrenergic neuron development. In addition, GATA2/3 and dHand play critical roles for noradrenergic neuron development. Recent work from our laboratory showed that AP2β may critically regulate NA neuron development in the ANS. Specific functional roles of these key transcription factors in the development of the ANS have been identified as shown below (Fig. 1.2).

Mash1 (also called Cash1) Mash1, a basic helix-loop-helix (bHLH) protein, is the first transcription factor shown to be essential for development of the ANS. In Mash1/ mice, virtually all noradrenaline (NA) neurons of the nervous system are affected, suggesting that Mash1 is a critical factor for determining the NA fate. Mash1 appears to relay BMP molecules’ signals for sympathetic development. Consistent with this idea, Mash1 expression was induced by BMPs in neural crest cultures and it was largely diminished in sympathetic ganglia following inhibition of BMPs function [13]. In Mash1-inactivated mouse embryos, neural crest cells migrated to the vicinity of the dorsal aorta, but did not develop into mature sympathetic neurons, as evidenced by the lack of expression of tyrosine hydroxylase (TH) and dopamine β-hydroxylase (DBH), as well as the absence of pan-neuronal markers [14]. In addition,

I. INTRODUCTION

TRANsCRIPTIONAl REgUlATORy CODE UNDERlyINg THE DEvElOPmENT AND PHENOTyPIC sPECIfICATION Of THE ANs

cAMP

whereas Phox2a expression is regulated by both Mash1 and Phox2b (Fig. 1.2). Recent promoter studies showed that Phox2a and 2b are able to directly activate the DBH promoter by interacting with multiple sequence motifs residing in the 5 flanking region (see below). Collectively, Phox2a and 2b appear to regulate both the development and neurotransmitter identity of sympathetic neurons and of other noradrenergic neurons.

BMPs

? Phox2b MASH1/CASH1

PKA

AP2 β

PNMT

CREB

GATA3

TH Neurotransmitter identity

Phox2a

DBH

5

c-Ret

GATA3

Survival

FIGURE 1.2 Diagrams depicting the regulatory network of the NA phenotype determination and maintenance. The diagram shows the possible regulatory interactions in the cascade of development and NA phenotype expression of ANS neurons. Thick arrows indicate likely direct regulation and thin arrows indicate direct or indirect regulation. BMPs are secreted from the dorsal aorta and activate the expression of MASH1/CASH1 and Phox2b. Phox2b then activates the expression of Phox2a. Phox2a and/or Phox2b in turn activate neurotransmitterspecifying genes TH and DBH. Direct action of Phox2a/2b on DBH transcription has been demonstrated. The transcription factor CREB has been shown to interact with the 5 promoter and directly activates transcription of the TH and DBH genes. The cAMP signaling pathway appears to regulate NA phenotype determination in concert with Phox2a/2b. The zinc finger factor GATA3, presumably downstream of Phox2b, also appears to be required for expression of NA-specific genes. Phox2a and 2b also regulate the expression of pan-neuronal genes including the GDNF receptor c-Ret, which may relate to their role in formation and/or survival of ANS neurons.

Mash1 directly or indirectly affects the expression of Phox2a, another key transcription factor important for DBH gene expression in a noradrenergic neuron-specific manner [15–17]. Thus, Mash1 is the first transcription factor known to control both differentiation and maintenance of noradrenergic neurons [18].

Phox2 Genes Phox2a and Phox2b are two closely related homeodomain transcription factors that are expressed in virtually all neurons that transiently or permanently express the noradrenergic neurotransmitter phenotype [19]. Gene inactivation studies have demonstrated that Phox2a and/or 2b are essential for proper development of all three divisions of the ANS and some noradrenergic containing structures of the CNS. For instance, in both Phox2a/ and 2b/ mouse brain, the major noradrenergic population in the locus coeruleus does not form, strongly suggesting that both genes are required for its development [20,21]. In contrast, only Phox2b seems to be required for the development of sympathetic neurons. In Phox2a/ mice, sympathetic neuron development is largely normal. Interestingly, however, both genes are able to induce sympathetic neuron-like phenotype when ectopically expressed in chick embryos [22]. During sympathetic development, Phox2b expression is induced by BMP molecules independently of Mash1,

The zinc finger transcription factor GATA-3 is a master regulator of type 2 T helper cell development. Interestingly, in GATA-3/ mice, noradrenaline deficiency is a proximal cause of embryonic lethality suggesting that GATA-3 is involved in the specification of noradrenergic neurotransmitter phenotype during noradrenergic neuron development [23]. Forced expression of GATA-3 in primary neural crest stem cell (NCSC) culture and developing chick embryos demonstrated that GATA-3 is able to increase the number of sympathoadrenergic neurons among NCSC culture, and induce ectopic expression of noradrenergic marker genes (TH and DBH) in developing chick embryos [24]. Furthermore, both TH and DBH promoters are robustly transactivated by GATA-3 via specific upstream subdomains encompassing binding motifs for transcription factors CREB, Sp1, and AP4. Protein–protein interaction assays showed that GATA-3 is able to physically interact with these transcription factors in vitro as well as in vivo [25]. Taken together, it is likely that GATA3 plays a critical role for specification of the NA phenotype via novel and distinct protein-protein interactions in both CNS and ANS development.

AP2(Activator Protein 2)β AP2 is a retinoic acid-inducible and developmentally regulated transcription factor with the basic helix-span-helix domain recognizing the palindromic 5-GCCNNNGGC-3 motif or its related GC-rich sequences. AP-2 family has 5 members, AP-2α, AP-2β, AP-2γ, AP-2δ, and AP-2, which show different spatiotemporal expression patterns during development. Based on its expression pattern in the neural crest, it was assumed that AP-2α regulates differentiation of neural crest-derived cells. However, our recent works demonstrated that AP2β is expressed in sympathetic ganglia of developing chick and mouse embryos and facilitates sympathoadrenergic differentiation in neural crest stem cells, while AP2α dramatically increases melanocytes at the expense of sympathoadrenergic cells [26,27]. Lossof-function studies of AP2β/ mouse revealed that DBH expression was significantly reduced in sympathetic ganglia as well as LC, indicating distinctive roles in neural crest differentiation and noradrenergic neurotransmitter specification in both central and peripheral nervous systems. Furthermore, these studies showed that AP2β directly controls the epinephrine phenotype by activating the

I. INTRODUCTION

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1. DEvElOPmENT AND DIffERENTIATION Of AUTONOmIC NEURONs

phenylethanolamine-N-methyl-transferase (PNMT) gene expression (Fig. 1.2).

Other Transcription Factors Although the above transcription factors are the most promising candidates, additional factors are emerging as being important for the ANS’ development and phenotype specification. For instance, dHand is another bHLH transcription factor whose expression is induced by BMPs and is dependent on Mash1. Based on its specific expression in sympathetic neurons, dHand may directly contribute to the ANS development. However, analysis of dHand function was hampered because knockout mice die before sympathetic neuron development. It is worthwhile to note that transcriptional regulation of the ANS and phenotype identity may require the combinatorial action of cell type-specific factors (e.g., Phox2a and 2b) and general transcription factors. Such examples may include cAMP response element binding protein and Sp1, which are required for transcriptional activity of both the TH and DBH genes (see below).

NEUROTRANSMITTER PHENOTYPES OF THE ANS Among the various phenotypes of a particular neuron, neurotransmitter identity is an essential feature because it determines the nature of the chemical neurotransmission a given neuron will mediate, and influences its specific connectivity with target cells. For specification of neurotransmitter identity, given neurons should express relevant genes encoding the biosynthetic enzymes and cofactors, as well as the specific reuptake protein(s). In addition, expression of these genes needs to be matched with the appropriate receptors of the target tissues. Therefore, expression of particular neurotransmitter phenotypes should be coordinated with the differentiation and phenotype specification of the target tissues. Recently, molecular mechanisms underlying the specification of neurotransmitters in the nervous system have been investigated extensively and key signaling molecules and transcriptional factors have been identified. Among these, specification of the noradrenaline (NA) phenotype of the sympathetic nervous system and the CNS is well characterized. Therefore, we will focus our discussion on the molecular characterization of the NA phenotype determination and its phenotypic switch to the cholinergic phenotype.

NA Phenotype NA is a major neurotransmitter of the ANS, especially in sympathetic neurons, and fundamentally mediates the function of the ANS. Consistent with this, a rare human disease called the dopamine β-hydroxylase deficient disease, in which NA is undetectable, was identified to be associated with severe autonomic function failure [28].

NA is one of the catecholamine neurotransmitters that are synthesized from tyrosine by three consecutive enzymatic steps. While tyrosine hydroxylase is responsible for the first step of catecholamine biosynthesis, converting tyrosine to L-dopa, and is expressed in all catecholamine neurons, dopamine β-hydroxylase (DBH) is responsible for conversion of dopamine to NA and is specifically expressed in NA neurons. Thus, DBH is a hallmark protein of NA neurons and the control mechanism of its expression is an essential feature of the development of NA neurons.

Control Mechanism of DBH Gene Expression is Closely Related to the ANS Development Numerous investigators using both in vivo transgenic mouse approaches and in vitro cell culture systems have studied DBH gene regulation. As schematically summarized in Figure 1.3, the 5 1.1 kb region upstream of the DBH gene promoter has three functional domains that can drive reporter gene expression in a NA cell type-specific manner. More detailed deletional and sitedirected mutational analyses indicate that as little as 486 bp of the upstream sequence of the human DBH gene can direct expression of a reporter gene in a cell-specific manner [29]. While the distal region spanning 486 to 263 bp appears to have a cell-specific silencer function, the proximal part spanning 262 to 1 bp is essential for high-level and cell-specific DBH promoter activity. In this 262 bp proximal area, four protein-binding regions (domains I to IV), initially identified by DNase I footprinting analysis, were found to encompass functionally important, multiple cis-regulatory elements [29], including the cAMP response element (CRE), YY1, AP2, Sp1, and core motifs of homeodomain (HD) binding sites. Site-directed mutagenesis of each sequence motif has revealed that these multiple cis-acting elements synergistically and/ or co-operatively regulate the transcriptional activity of the DBH gene [16]. Among these, two ATTA-containing motifs in domain IV and another motif in domain II were identified to be NA-specific cis-acting motifs in that their mutation diminished the DBH promoter function only in NA cell lines. More recently, another NA-specific cisregulatory element was identified between domain II and III (Fig. 1.3). Interestingly, analysis of DNA-protein interactions on the DBH promoter demonstrated that all of these four NA-specific cis-regulatory elements are Phox2binding sites [15]. Taken together, this experimental evidence establishes that the DBH gene is an immediate downstream target of Phox2 proteins.

Mutations of DBH Gene are Closely Associated with the Autonomic Disorder, Orthostatic Hypotension DBH deficiency is a rare congenital disorder, first described in 1986 [28,30]. DBH deficiency is a severe autonomic disorder exhibiting sympathetic noradrenergic

I. INTRODUCTION

NEUROTRANsmITTER PHENOTyPEs Of THE ANs DBH deficiency

Normal

nucleus

nucleus

ER X

L-tyrosine

L-tyrosine

L-DOPA

L-DOPA

dopamine

dopamine

Golgi

LDCV

FIGURE 1.3 Effects of pathogenic mutations on DBH synthesis and trafficking. In normal, DBH enzymes (circle) are translated in the ER, transported to the Golgi, and then packaged into the LDCVs. In LDCV, DBH converts dopamine (triangle) to noradrenaline (rectangle). Noradrenaline are released by exocytosis upon stimulation of the nerve terminal and readily detectable in plasma. In DBH deficiency, DBH proteins (S-shape) encoded by missense mutations are retained in the ER and fail to be targeted to the LDCV due to their misfolding. As the result, noradrenaline is not synthesized in LDCV, leading to absence of noradrenaline in plasma. Instead, dopamine is dramatically increased.

failure and adrenomedullary failure but intact vagal and sympathetic cholinergic function [31]. DBH deficient patients exhibit severe deficits in autonomic regulation of cardiovascular function predisposing them to orthostatic hypotension. These patients display characteristic perturbations in the level of catecholamines: undetectable noradrenaline and its metabolites, and highly elevated dopamine and its metabolites. Given the fundamental role of noradrenaline in the nervous system, the report of adult patients with undetectable noradrenaline is both surprising and interesting. The report of frequent miscarriages and spontaneous abortions in mothers of known DBH deficiency cases suggests the interesting possibility that there could be many more undiagnosed fetal and neonatal deaths resulting from DBH deficiency and that those adult patients are lucky survivors [30]. In line with this, a DBH knock out mouse study showed less than 5% live births [32]. Mortality appeared to be due to cardiovascular failure caused by DBH deficiency in utero, which is reminiscent of tyrosine hydroxylase null mice. We, and others, have recently reported a mutation in the splice donor site of the first exon-intron junction and several missense mutations associated with the DBH deficiency syndrome in the DBH gene of these patients [33–35]. The mutation in the splice donor site (IVS1  2T→C) of DBH resulted in abnormal mRNA splicing and generated a transcript containing a premature stop codon as well as

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a normal transcript. Interestingly, all missense mutations so far identified in DBH deficient patients are subtle amino acid substitutions resulting in defective protein trafficking and mislocation, probably due to protein misfolding (Fig. 1.3) [35]. One of the well-known chemical chaperones, glycerol, could partially rescue defective secretion of DBH mutant proteins, suggesting the possibility that DBH deficiency disease could be more fundamentally treated with pharmacological chaperones [35].

Cholinergic Phenotype and the Switch of Neurotransmitter Phenotypes by Target Cell Interactions Another major neurotransmitter of the ANS is acetylcholine, and neurons generating this neurotransmitter are designated cholinergic. Among sympathetic neurons, the number of cholinergic neurons is much less than NA neurons. While differentiation and cholinergic specification are extensively investigated in central motor neurons [4], development of cholinergic ANS neurons is not well characterized. Therefore, it is of great interest to understand if key transcription factors such as HB9 and MNR2, likewise play key roles in determining the cholinergic phenotype during development of the ANS. Interestingly, it is well described that upon contacting developing sweat glands, the NA phenotype of sympathetic axons switches to the cholinergic phenotype [10]. The putative cholinergic-inducing factor secreted from sweat glands remains to be defined although leukemia inhibitory factor and ciliary neurotrophic factor are candidates. Consistent with the observation that TH and DBH expression remains even after neurotransmitter switch from the NA to the cholinergic phenotype, a recent study reported that levels of the TH cofactor, tetrahydrobiopterin (BH4), dropped significantly during the switch [36]. Immunoreactivity for the BH4-synthesizing GTP cyclohydrolase became undetectable in sweat gland neurons during this phenotypic switch, suggesting that suppression of cofactor expression underlies the neurotransmitter switch during development.

Acknowledgements This work was supported by NIH grants (MH048866, MH087903, and NS070577).

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[4] Edlund T, Jessell TM. Progression from extrinsic to intrinsic signaling in cell fate specification: a view from the nervous system. Cell 1999;96:211–24. [5] Wilson SI, Rydstrom A, Trimborn T, Willert K, Nusse R, et al. The status of Wnt signalling regulates neural and epidermal fates in the chick embryo. Nature 2001;411:325–30. [6] Stern CD, Artinger KB, Bronner-Fraser M. Tissue interactions affecting the migration and differentiation of neural crest cells in the chick embryo. Development 1991;113:207–16. [7] Ernsberger U, Rohrer H. The development of the noradrenergic transmitter phenotype in postganglionic sympathetic neurons. Neurochem Res 1996;21:823–9. [8] Schneider C, Wicht H, Enderich J, Wegner M, Rohrer H. Bone morphogenetic proteins are required in vivo for the generation of sympathetic neurons. Neuron 1999;24:861–70. [9] Bilodeau ML, Boulineau T, Hullinger RL, Andrisani OM. Cyclic AMP signaling functions as a bimodal switch in sympathoadrenal cell development in cultured primary neural crest cells. Mol Cell Biol 2000;20:3004–14. [10] Francis NJ, Landis SC. Cellular and molecular determinants of sympathetic neuron development. Annu Rev Neurosci 1999;22:541–66. [11] Taraviras S, Pachnis V. Development of the mammalian enteric nervous system. Curr Opin Genet Dev 1999;9:321–7. [12] Goridis C, Brunet JF. Transcriptional control of neurotransmitter phenotype. Curr Opin Neurobiol 1999;9:47–53. [13] Goridis C, Rohrer H. Specification of catecholaminergic and serotonergic neurons. Nat Rev Neurosci 2002;3:531–41. [14] Guillemot F, Lo LC, Johnson JE, Auerbach A, Anderson DJ, et al. Mammalian achaete-scute homolog 1 is required for the early development of olfactory and autonomic neurons. Cell 1993;75:463–76. [15] Seo H, Hong SJ, Guo S, Kim HS, Kim CH, et al. A direct role of the homeodomain proteins Phox2a/2b in noradrenaline neurotransmitter identity determination. J Neurochem 2002;80:905–16. [16] Kim HS, Seo H, Yang C, Brunet JF, Kim KS. Noradrenergic-specific transcription of the dopamine beta-hydroxylase gene requires synergy of multiple cis-acting elements including at least two Phox2abinding sites. J Neurosci 1998;18:8247–60. [17] Yang C, Kim HS, Seo H, Kim CH, Brunet JF, et al. Paired-like homeodomain proteins, Phox2a and Phox2b, are responsible for noradrenergic cell-specific transcription of the dopamine betahydroxylase gene. J Neurochem 1998;71:1813–26. [18] Hirsch MR, Tiveron MC, Guillemot F, Brunet JF, Goridis C. Control of noradrenergic differentiation and Phox2a expression by MASH1 in the central and peripheral nervous system. Development 1998;125:599–608. [19] Brunet JF, Pattyn A. Phox2 genes - from patterning to connectivity. Curr Opin Genet Dev 2002;12:435–40. [20] Morin X, Cremer H, Hirsch MR, Kapur RP, Goridis C, et al. Defects in sensory and autonomic ganglia and absence of locus coeruleus in mice deficient for the homeobox gene Phox2a. Neuron 1997;18:411–23. [21] Pattyn A, Goridis C, Brunet JF. Specification of the central noradrenergic phenotype by the homeobox gene Phox2b. Mol Cell Neurosci 2000;15:235–43.

[22] Stanke M, Junghans D, Geissen M, Goridis C, Ernsberger U, et al. The Phox2 homeodomain proteins are sufficient to promote the development of sympathetic neurons. Development 1999;126:4087–94. [23] Lim KC, Lakshmanan G, Crawford SE, Gu Y, Grosveld F, et al. Gata3 loss leads to embryonic lethality due to noradrenaline deficiency of the sympathetic nervous system. Nat Genet 2000;25:209–12. [24] Hong SJ, Huh Y, Chae H, Hong S, Lardaro T, et al. GATA-3 regulates the transcriptional activity of tyrosine hydroxylase by interacting with CREB. J Neurochem 2006;98:773–81. [25] Hong SJ, Choi HJ, Hong S, Huh Y, Chae H, et al. Transcription factor GATA-3 regulates the transcriptional activity of dopamine betahydroxylase by interacting with Sp1 and AP4. Neurochem Res 2008;33:1821–31. [26] Hong SJ, Lardaro T, Oh MS, Huh Y, Ding Y, et al. Regulation of the noradrenaline neurotransmitter phenotype by the transcription factor AP-2beta. J Biol Chem 2008;283:16860–16867. [27] Hong SJ, Huh YH, Leung A, Choi HJ, Ding Y, et al. Transcription factor AP-2b regulates the neurotransmitter phenotype and maturation of chromaffin cells. Mol Cell Neurosci 2011;46:245–51. [28] Robertson D, Goldberg MR, Onrot J, Hollister AS, Wiley R, et al. Isolated failure of autonomic noradrenergic neurotransmission. Evidence for impaired beta-hydroxylation of dopamine. N Engl J Med 1986;314:1494–7. [29] Seo H, Yang C, Kim HS, Kim KS. Multiple protein factors interact with the cis-regulatory elements of the proximal promoter in a cellspecific manner and regulate transcription of the dopamine betahydroxylase gene. J Neurosci 1996;16:4102–12. [30] Man in’t Veld AJ,, Boomsma F, Moleman P, Schalekamp MA. Congenital dopamine-beta-hydroxylase deficiency. A novel orthostatic syndrome. Lancet 1987;1:183–8. [31] Biaggioni I, Goldstein DS, Atkinson T, Robertson D. Dopaminebeta-hydroxylase deficiency in humans. Neurology 1990;40:370–3. [32] Thomas SA, Matsumoto AM, Palmiter RD. Noradrenaline is essential for mouse fetal development. Nature 1995;374:643–6. [33] Kim CH, Zabetian CP, Cubells JF, Cho S, Biaggioni I, et al. Mutations in the dopamine beta-hydroxylase gene are associated with human norepinephrine deficiency. Am J Med Genet 2002;108:140–7. [34] Deinum J, Steenbergen-Spanjers GC, Jansen M, Boomsma F, Lenders JW, et al. DBH gene variants that cause low plasma dopamine beta hydroxylase with or without a severe orthostatic syndrome. J Med Genet 2004;41:e38. [35] Kim CH, Leung A, Huh YH, Yang E, Kim DJ, et al. Norepinephrine deficiency is caused by combined abnormal mRNA processing and defective protein trafficking of dopamine b-hydroxylase. J Biol Chem 2011;286:9196–9204. [36] Habecker BA, Klein MG, Sundgren NC, Li W, Woodward WR. Developmental regulation of neurotransmitter phenotype through tetrahydrobiopterin. J Neurosci 2002;22:9445–52.

I. INTRODUCTION

C H A P T E R

2 Central Autonomic Control Eduardo E. Benarroch Central control of the sympathetic and parasympathetic outputs involves several interconnected areas distributed throughout the neuraxis. This central autonomic network has a critical role in moment-to-moment control of visceral function, homeostasis, and adaptation to internal or external challenges. The functions of the central autonomic network are organized in four hierarchical levels that are closely interconnected: spinal, bulbopontine, pontomesencephalic and forebrain levels (Fig. 2.1). The spinal level mediates segmental sympathetic or sacral parasympathetic reflexes and is engaged in stimulus-specific patterned responses under the influence of the other levels. The bulbopontine (lower brainstem) level is involved in reflex control of circulation, respiration, gastrointestinal function, and micturition. The pontomesencephalic (upper brainstem) level integrates autonomic control with pain modulation and integrated behavioral responses to stress. The forebrain level includes the hypothalamus, which is involved in integrated control of autonomic and endocrine responses for homeostasis and adaptation, and components of the anterior limbic circuit, including the insula, anterior cingulate cortex, and amygdala, which are involved in integration of bodily sensation with emotional and goal-related autonomic responses.

sympathetic and parasympathetic outputs, primarily via a relay in the lateral hypothalamus [4,5].

Anterior Cingulate Cortex The anterior cingulate cortex is interconnected with the anterior insula and is subdivided into ventral (affective) and dorsal (cognitive) regions [6]. The ventral anterior cingulate is part of the brain “default mode network” whereas the dorsal anterior cingulate is a component of the frontoparietal attention networks. The ventral anterior cingulate cortex includes subcallosal and precallosal portions that have extensive connections with the insula, prefrontal cortex, amygdala, hypothalamus, and brain stem. Via these projections, the anterior cingulate cortex controls sympathetic and parasympathetic functions [7].

Amygdala The amygdala provides affective or emotional value to incoming sensory information [8] and has multiple downstream targets that participate in the autonomic and neuroendocrine response to stress [9]. The central nucleus of the amygdala (CeA), both directly and via the bed nucleus of the stria terminalis, has a major role in integration of the stress responses, particularly fear responses, via its widespread connections with the hypothalamus and brainstem, particularly the periaqueductal gray and the medullary reticular formation [10].

FOREBRAIN COMPONENTS The forebrain regions involved in the control of autonomic functions include the insular cortex, anterior cingulate cortex, amygdala, and several areas of the hypothalamus.

Hypothalamus The hypothalamus integrates autonomic and endocrine responses necessary for homeostasis and adaptation. It acts as a visceromotor pattern generator that initiates specific patterns of autonomic and endocrine responses according to the stimulus, such as hypoglycemia, changes in blood temperature or osmolarity, or external stressors [2,11]. The preoptic-hypothalamic area is functionally subdivided into three functional zones, periventricular, medial, and lateral [11]: The periventricular zone includes the suprachiasmatic nucleus (the circadian pacemaker), and several areas involved in neuroendocrine control via the pituitary gland. The medial zone includes the medial

Insular Cortex The insular cortex is the primary interoceptive cortex and integrates visceral, pain and temperature sensations [1,2]. The dorsal insula has a viscerotropic organization [3] and receives inputs from gustatory, visceral, muscle, and skin receptors via the thalamus. The dorsal insula projects to the right anterior insula, which, via its connections with neocortical association and limbic areas, integrates these interoceptive inputs with emotional and cognitive processing to conveys the conscious experience of bodily sensation [1]. The insula is also a visceromotor area controlling both the

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00002-0

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© 2012 Elsevier Inc. All rights reserved.

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2. CEnTRAl AuTonomIC ConTRol

FIGURE 2.1 Central autonomic control areas and levels of interaction of autonomic control.

preoptic area, paraventricular nucleus (PVN) and dorsomedial nucleus (DMH), which orchestrate coordinated autonomic and endocrine outputs for thermoregulation, osmoregulation, and stress responses [11–14]. The lateral zone includes nuclei that control sleep, arousal and motivated behavior [2]. The main autonomic outputs of the hypothalamus originate from the PVN, DMH, and lateral hypothalamic area [2]. The PVN contains different neuronal populations that are differentially activated during stress responses [12]. They include magnocellular neurons that release arginine-vasopressin (AVP) to the general circulation; neurons that release corticotropin releasing hormone and activate the adrenocortical axis, and neurons that project to autonomic nuclei of the brainstem and spinal cord [12]. Via these outputs, the PVN modulates stress responses, food and sodium intake, glucose metabolism, and cardiovascular, renal, gastrointestinal, and respiratory functions. The DMH participates in stress responses [13]; thermoregulation [14,15] and cardiovascular control [16].

Hypocretin/orexin neurons of the posterior lateral hypothalamus contribute to control autonomic output in the setting of arousal, feeding and reward driven behaviors [2].

BRAINSTEM COMPONENTS The brainstem areas controlling autonomic output include the periaqueductal gray matter of the midbrain (PAG), the parabrachial nucleus (PBN) and several medullary regions, including the nucleus of the solitary tract (NTS), ventrolateral reticular formation of the medulla, and medullary raphe (Fig. 2.1).

Periaqueductal Gray The PAG is the interface between the forebrain and the lower brain stem and has a major role in integrated autonomic and somatic responses to stress, pain modulation,

I. INTRODUCTION

AuTonomIC ouTPuT of THE CEnTRAl nERvous sysTEm

and other adaptive functions. It consists of different longitudinal columns that, via their different spinal, brainstem, and cortical connections, participate in cardiovascular responses associated with pain modulation [17]; coordination of the micturition reflex [18]; and control of respiration.

Parabrachial Complex and Adjacent Regions of the Pons The PBN is a major relay center that receives converging visceral, nociceptive and thermoreceptive inputs from the spinal cord and conveys this information to the hypothalamus, amygdala, and thalamus [2]. The PBN also participates in the control of respiratory, cardiovascular, and gastrointestinal functions. The dorsal pontine tegmentum also contains the pontine micturition center (PMC), also referred to as Barrington nucleus or M- (for medial) region, which is critical for the coordination of the micturition reflex and participates in the control of the function of the lower gastrointestinal tract and sexual organs [18].

Nucleus of the Solitary Tract The NTS is the first relay station of taste and visceral afferent information and includes several subnuclei with a viscerotropic organization. The rostral portion of the NTS receives taste inputs; the intermediate portion receives gastrointestinal afferents; and the caudal portion receives baroreceptor, cardiac, chemoreceptor, and pulmonary afferents [19]. The NTS relays this information, either directly or via the PBN, to rostral brainstem and forebrain areas [2,3]. The NTS is also the first central relay for all medullary reflexes controlling cardiovascular function (baroreflex and cardiac reflexes) [20], respiration (carotid chemoreflex and pulmonary mechanoreflexes), and gastrointestinal motility [21].

Rostral Ventrolateral Medulla The rostral ventrolateral medulla (RVLM), including the C1 group of epinephrine-containing neurons, is a key area for regulation of arterial blood pressure [20,21]. Glutamatergic neurons of the RVLM project directly and provide tonic excitation to sympathetic preganglionic neurons controlling cardiac output and total peripheral resistance [15]. The RVLM mediates all reflexes controlling arterial blood pressure, including the baroreflex, cardiopulmonary reflexes, and chemoreflexes [20,22]. The sympathoexcitatory RVLM neurons receive and integrate a large variety of inputs from the brainstem and forebrain [22]. These include inhibitory signals from baroreceptorsensitive neurons of the NTS and mediated via inhibitory gamma-aminobutyric acid (GABA) ergic neurons of the caudal ventrolateral medulla [22]. The RVLM also receives several inputs from the hypothalamus, including the PVN.

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Caudal Ventrolateral Medulla The caudal ventrolateral medulla contains GABAergic neurons that maintain a tonic inhibitory control on the RVLM and relay the inhibitory inputs from the NTS mediating the sympathoinhibitory component of the arterial baroreflex. Electrical stimulation studies indicate that the caudal medulla also contains pressor regions [23]. The caudal ventrolateral medulla also contains the A1 group of neurons that provide noradrenergic innervation to the hypothalamus and are a component of a reflex pathway that triggers AVP release in response to hypovolemia or hypotension [24].

Ventromedial Medulla and Caudal Raphe The rostral ventromedial medulla including the caudal raphe nuclei, has an important role in thermoregulation [15] pain modulation [17], and control of automatic ventilation [25]. One group of medullary raphe neurons initiate sympathetic responses to cold via input to preganglionic sympathetic neurons that activate skin vasoconstriction and non-shivering thermogenesis in the brown adipose tissue [26].

AUTONOMIC OUTPUT OF THE CENTRAL NERVOUS SYSTEM Sympathetic Preganglionic Units The sympathetic output is critical for maintenance of arterial pressure, thermoregulation, and redistribution of regional blood flow during stress and exercise. The sympathetic output originates from sympathetic preganglionic neurons located in the thoracolumbar spinal cord at the T1 to -L2 segments, primarily in the intermediolateral cell column. These neurons are organized into functionally separate units that innervate selective subpopulations of sympathetic ganglion neurons and receive distinct segmental afferent inputs triggering segmental somato- and viscerosympathetic reflexes [27]. Different preganglionic sympathetic units are recruited in a coordinated fashion by premotor neurons in the brainstem and hypothalamus to initiate different patterns of responses to specific internal or external stressors, such as postural changes, exercise, hypoglycemia, dehydration, exposure to heat or cold, or stress [2]. The main sources of premotor sympathetic innervation are the RVLM, medullary raphe, A5 noradrenergic group of the pons; PVN, and lateral hypothalamic area [4,28].

Parasympathetic Outputs In contrast to the sympathetic system, which mediates stimulus-specific patterns of responses affecting multiple effectors; the parasympathetic system mediates reflexes activated in an organ-specific fashion.

I. INTRODUCTION

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Vagal Output The vagal output originates from preganglionic neurons located in the dorsal motor nucleus of the vagus (DMV) and in the ventrolateral portion of the nucleus ambiguus (NAmb) in the medulla. The DMV contains most of the vagal preganglionic parasympathetic neurons that are organized in a viscerotropic fashion [29] and innervate the local ganglia in the respiratory tract, enteric nervous system (ENS), liver and pancreas. The DMV receives inputs from the NTS and mediates all vago-vagal reflexes controlling gastrointestinal motility and secretion [21]. Vagal preganglionic neurons located in the ventrolateral portion of the NAmb provide the primary control of the heart via the cardiac ganglia [30]. These cardiovagal outputs inhibit the automatism of the sinoatrial node exerting a beat-to-beat control of the heart rate. The cardiovagal NAMb neurons are activated by the NTS during the baroreflex and are inhibited during inspiration. Sacral Parasympathetic Output The sacral preganglionic output originates in neurons located in the lateral gray matter at the S2-S4 segments of the sacral spinal cord [31]. These neurons are critical for normal micturition, defecation, and sexual organs; this involves their coordinated interactions with both lumbar sympathetic neurons located at T12-L2 levels and somatic motor neurons of the Onuf nucleus at the S2-S4 levels innervating the external urinary sphincter and pelvic floor.

References [1] Craig AD. Interoception: the sense of the physiological condition of the body. Curr Opin Neurobiol 2003;13:500–5. [2] Saper CB. The central autonomic nervous system: conscious visceral perception and autonomic pattern generation. Annu Rev Neurosci 2002;25:433–69. [3] Cechetto DF. Central representation of visceral function. Fed Proc 1987;46:17–23. [4] Loewy AD. Descending pathways to the sympathetic preganglionic neurons. Prog Brain Res 1982;57:267–77. [5] Westerhaus MJ, Loewy AD. Central representation of the sympathetic nervous system. 2001 [6] Vogt BA, Vogt L, Farber NB, Bash E, et al. Architecture and neurocytology of monkey cingulate gyrus. J Comp Neurol 2005;485:218–39. [7] Verberne AJ, Owens NC. Cortical modulation of the cardiovascular system. Prog Neurobiol 1998;54:149–68. [8] LeDoux J. The amygdala. Curr Biol 2007;17:R868–874. [9] Ulrich-Lai YM, Herman JP. Neural regulation of endocrine and autonomic stress responses. Nat Rev Neurosci 2009;10:397–409. [10] Davis M. The role of the amygdala in fear and anxiety. Annu Rev Neurosci 1992;15:353–75. [11] Thompson RH, Swanson LW. Structural characterization of a hypothalamic visceromotor pattern generator network. Brain Res Brain Res Rev 2003;41:153–202.

[12] Sawchenko PE, Li HY, Ericsson A. Circuits and mechanisms governing hypothalamic responses to stress: a tale of two paradigms. Prog Brain Res 2000;122:61–78. [13] Dimicco JA, Zaretsky DV. The dorsomedial hypothalamus: a new player in thermoregulation. Am. J Physiol Regul Integr Comp Physiol 2007;292:R47–63. [14] Yoshida K, Li X, Cano G, et al. Parallel preoptic pathways for thermoregulation. J Neurosci 2009;29:11954–11964. [15] Morrison SF. RVLM and raphe differentially regulate sympathetic outflows to splanchnic and brown adipose tissue. Am J Physiol 1999;276:R962–973. [16] Dampney RA, Horiuchi J, McDowall LM. Hypothalamic mechanisms coordinating cardiorespiratory function during exercise and defensive behavior. Auton Neurosci 2008. [17] Bandler R, Keay KA, Floyd N, et al. Central circuits mediating patterned autonomic activity during active vs. passive emotional coping. Brain Res Bull 2000;53:95–104. [18] Holstege G. Micturition and the soul. J Comp Neurol 2005;493:15–20. [19] Jean A. The nucleus tractus solitarius: neuroanatomic, neurochemical and functional aspects. Arch Int Physiol Biochim Biophys 1991;99:A3–52. [20] Dampney RA, Horiuchi J. Functional organisation of central cardiovascular pathways: studies using c-fos gene expression. Prog Neurobiol 2003;71:359–84. [21] Travagli RA, Hermann GE, Browning KN, Rogers C, et al. Brainstem circuits regulating gastric function. Annu Rev Physiol 2006;68:279–305. [22] Guyenet PG. The sympathetic control of blood pressure. Nat Rev Neurosci 2006;7:335–46. [23] Goodchild AK, Moon EA. Maps of cardiovascular and respiratory regions of rat ventral medulla: focus on the caudal medulla. J Chem Neuroanat 2009;38:209–21. [24] Lightman SL, Todd K, Everitt BJ. Ascending noradrenergic projections from the brainstem: evidence for a major role in the regulation of blood pressure and vasopressin secretion. Exp Brain Res 1984;55:145–51. [25] Corcoran AE, Hodges MR, Wu Y, Wang W, Wylie CJ, Deneris ES, Richerson S, et al. Medullary serotonin neurons and central CO2 chemoreception. Respir. Physiol Neurobiol 2009;168:49–58. [26] Morrison SF, Nakamura K, Madden CJ. Central control of thermogenesis in mammals. Exp Physiol 2008;93:773–97. [27] Janig W, Habler HJ. Neurophysiological analysis of target-related sympathetic pathways – from animal to human: similarities and differences. Acta Physiol Scand 2003;177:255–74. [28] Strack AM, Sawyer WB, Hughes JH, Platt KB, Lavey AD, et al. A general pattern of CNS innervation of the sympathetic outflow demonstrated by transneuronal pseudorabies viral infections. Brain Res. 1989;491:156–62. [29] Huang XF, Tork I, Paxinos G. Dorsal motor nucleus of the vagus nerve: a cyto- and chemoarchitectonic study in the human. J Comp Neurol 1993;330:158–82. [30] Hopkins DA, Armour JA. Brainstem cells of origin of physiologically identified cardiopulmonary nerves in the rhesus monkey (Macaca mulatta). J Auton Nerv Syst 1998;68:21–32. [31] Birder L, de Groat W, Mills I, Morrison J, Thor K, Drake M, et al. Neural control of the lower urinary tract: peripheral and spinal mechanisms. Neurourol Urodyn 29:128–39.

I. INTRODUCTION

C H A P T E R

3 Imaging of Brainstem Sites Involved in Cardiovascular Control Vaughan G. Macefield, Luke A. Henderson The medulla is phylogenetically the oldest structure of the brain, containing dense clusters of neurons (nuclei) that coordinate many homeostatically critical functions, such as respiration, heart rate and blood pressure. Much of what we know of the functions of the medulla we have learnt from experiments in anesthetized or decerebrate animals. And, although its small size makes it difficult to study its function in awake humans, it is possible to use non-invasive neuroimaging approaches to delineate some of its nuclei. We shall limit our discussion to the control of arterial pressure, in which most of the work has been carried out. The maintenance of blood pressure within a relatively narrow physiological range is essential to ensure adequate perfusion of vital organs such as the brain. A primary determinant of arterial pressure is the level of sympathetically-mediated vasoconstriction within skeletal muscle. Muscle sympathetic nerve activity (MSNA) occurs as pulse-synchronous bursts, the incidence and magnitude of which are inversely correlated to diastolic pressure. The beat-to-beat control of arterial pressure is governed by the baroreflex – a classic negative-feedback loop in which increases in blood pressure, detected by the arterial baroreceptors (located in the carotid sinus and aortic arch), are corrected by withdrawal of muscle vasoconstrictor drive as well as decreases in heart rate and stroke volume. This short-term regulation of blood pressure survives removal of structures rostral to the medulla yet is abolished by transection of the spinal cord immediately below the medulla, indicating that the medulla contains structures critical for the generation and maintenance of vasomotor tone.

produce increases in activity in cutaneous sympathetic neurons yet no increase in blood pressure [1]. It is known from intracellular recordings from RVLM neurons that they exhibit an irregular tonic firing that decreases with increases in blood pressure and increases with decreases in blood pressure; it is also known that this tonic activity is modified by excitatory and inhibitory inputs, but that in the absence of such inputs this tonic activity continues [2]. The current understanding of the baroreflex circuitry defined primarily in anesthetized animal preparations – is that primary afferent axons from the baroreceptors project to the caudal region of the nucleus tractus solitarius (NTS), where they synapse onto second-order neurons, which in turn send excitatory (glutamatergic) projections onto GABAregic neurons within the region of the caudal ventrolateral medulla (CVLM). These CVLM GABAergic neurons synapse directly onto excitatory neurons within the RVLM and serve to inhibit the spontaneous activity of RVLM premotor sympathetic neurons. Nucleus ambiguus and the dorsal motor nucleus of the vagus also receive glutamatergic projections from NTS [3], activating vagal cardiac efferents and slowing the heart. In addition to these components of the basic baroreflex arc, it is known that other brainstem regions, such as the caudal pressor area (CPA), located in the most caudal part of the ventrolateral medulla, can also influence baroreflex activity by altering the activity in these baroreflex medullary nuclei [2]. Studies in conscious animals, using c-fos expression as a marker of neuronal activation, have confirmed the operation of the NTS-CVLM-RVLM serial pathway during maneuvers that increase or decrease arterial pressure [4,5].

IDENTIFICATION OF MEDULLARY CARDIOVASCULAR NUCLEI IN EXPERIMENTAL ANIMALS

IDENTIFICATION OF MEDULLARY CARDIOVASCULAR NUCLEI IN HUMANS

The circuitry responsible for the baroreflex has been examined in detail in anesthetized experimental animals. Early work had shown that excitation within discrete regions of the rostral ventrolateral medulla (RVLM) could activate muscle vasoconstrictor neurons and increase blood pressure, while activation within other areas could

Non-invasive neuroimaging has become a powerful tool in defining the functions of various regions in the human brain, but relatively little work has been done in cardiovascular control (for review see [6]). Most of the recent studies in this area use functional magnetic resonance imaging (fMRI), in which changes in blood-oxygen

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FIGURE 3.1 Significant BOLD signal intensity changes in the medulla and cerebellum, obtained from the brainstem-specific scans (n  15) overlaid onto an average T2-weighted anatomical image set. Ventral is at the base of each image. The hot and cool color scales (coded for t-value) indicate regional signal increases and decreases, respectively. Slice positions (MNI space) are indicted in the top right of each image. To the right are three graphs showing the mean (SEM) percent changes in signal intensity (SI) over time for three significant clusters. Vertical grey boxes indicate each of the three inspiratory-capacity apnea periods. From Macefield et al. (2006) [8].

level dependent (BOLD) signal intensity are used as a proxy marker of neuronal activity: increases in signal intensity reflect increases in neuronal activity, while decreases reflect deactivations. Such approaches have been used in the anesthetized cat to identify changes in signal intensity within the medullary regions described above [7], but relatively few studies have applied fMRI to investigate the role of the brainstem in human cardiovascular control. One of the first investigations to do so used the Valsalva maneuver – a forced expiratory effort which evokes a stereotypical pattern of autonomic changes. The virtue in using this maneuver is that the changes in heart rate and arterial pressure are well known, with the increase in arterial pressure being brought about by an increase in muscle sympathetic nerve activity (MSNA). Significant changes in BOLD signal intensity occurred in the amygdala, hippocampus, insular and lateral frontal cortices, as well as in the dorsal pons, dorsal medulla, lentiform nucleus, and fastigial and dentate nuclei of the cerebellum [7]. This was one of the first studies to have attempted to identify the functional roles of the human brainstem in autonomic control, but the fact that the Valsalva maneuver is a complex, volitionally generated task makes it difficult to tease out elements related to motor planning, execution, sensory feedback and autonomic changes. A simpler maneuver, the inspiratory-capacity apnea – a maximal inspiratory

FIGURE 3.2 Left panel, axial sections of the brainstem during a series of inspiratory-capacity apneas, showing bilateral activation of the dorsolateral medulla. Data from Macefield et al. (2006) [8]. Right panel, axial sections of the human brainstem showing a high density of AngII binding in RVLM. Data reproduced with permission from Allen et al. (1988) [9]. See color plate at back of the book.

breath-hold – evokes a sustained increase in MSNA that is believed to be due primarily to unloading of the low-pressure baroreceptors. During the static phase of the maneuver, in which the inspiratory pump muscles are quiescent and only the laryngeal constrictors are active, intrathoracic pressure changes because of the elastic recoil of the lungs and chest wall against the closed glottis. This is different to the Valsalva maneuver, in which the diaphragm, abdominal and intercostal muscles are co-activated in a sustained effort that increases intra-abdominal and intrathoracic pressures. Like the Valsalva maneuver,

I. INTRODUCTION

IdEnTIfICATIon of mEdullARy CARdIovASCulAR nuClEI In HumAnS

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FIGURE 3.3 Significant increases (warm color scale) and decreases (cool color scale) in BOL1D signal intensity within the brainstem correlated with MSNA total burst activity in the resting state; raw fMRI data from seven experiments. Equivalent histological sections are shown on the right. Figure from Macefield and Henderson (2010) [12].

the inspiratory-capacity apnea evokes discrete changes in BOLD signal intensity over multiple brain regions. Significant increases in BOLD signal intensity occur bilaterally in the lateral prefrontal cortex, anterior insular cortex, anterior cingulate cortex, dorsomedial hypothalamus and deep cerebellar nuclei; decreases in signal intensity occur in the hippocampus, posterior cingulate cortex and the cerebellar cortex [8]. As shown in the sagital and axial sections of the human brainstem in Figure 3.1, bilateral increases in activity also occur in the rostral lateral medulla, with decreases occurring in the dorsomedial and caudal lateral medulla. We believe the increases in the rostral lateral medulla reflect increases in activity of the RVLM. In humans, the RVLM and CVLM are displaced dorsally by the large inferior olivary nuclei [9]. The functional and anatomical localization of the human RVLM is illustrated in Figure 3.2: the left panel shows a bilateral increase in signal intensity during a maximal inspiratory breath-hold; the right panel shows the anatomical identification of the human RVLM, based on histochemical identification [9]. While McAllen and colleagues have functionally identified the medullary raphé nucleus during whole-body cooling [10], until recently no human fMRI studies had functionally identified NTS. As indicated in the right panel of Figure 3.2, the solitary tract nucleus is located dorsal and medial to the RVLM. We believe the decreases in signal intensity in the dorsomedial medulla reflect decreases in NTS, with the decreases in the caudal lateral medulla reflecting

decreases in neuronal activity in the CVLM. These responses can be explained by (i) the reduction of baroreceptor inputs to the NTS; (ii) a consequent reduction in activity of CVLM; and hence (iii) a reduction in inhibitory drive to the RVLM. The withdrawal of inhibitory drive to RVLM then allows this nucleus to increase its activity and thereby bring about an increase in MSNA. Sustained increases in MSNA can also be induced by static handgrip exercise and post-exercise ischemia: accumulation of the metabolites of contraction activates metaboreceptors within the muscle and induces a reflex increase in MSNA and arterial pressure. It is known that metaboreceptors project to the NTS, which sends excitatory projections to the RVLM without an intervening inhibitory synapse: increases in BOLD signal intensity occurred in discrete regions of the human medulla corresponding to the NTS and RVLM, but did not include CVLM [11]. A recent development has been concurrent microneurography and fMRI. By recording spontaneous bursts of MSNA and correlating these to spontaneous fluctuations in BOLD signal intensity it has been possible to functionally identify regions of the brainstem involved in the generation of spontaneous MSNA [12]. Analysis of the temporal coupling between BOLD signal intensity and nerve signal intensity revealed sites in which the two signals covaried, but these sites were limited to the medulla. The advantage of this approach is that maneuvers, with all of the associated confounds, need not be performed. As shown in Figure 3.3, increases in MSNA were associated

I. INTRODUCTION

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with bilateral increases in signal intensity in the rostral lateral medulla that corresponds to the human equivalent of the RVLM. There was also a unilateral (left) increase in activity in the area that corresponds to the caudal pressor area (CPA). Reciprocal decreases in signal intensity occurred in the dorsomedial region of NTS and in CVLM, though on average the latter appeared to be limited to the right side. Although not illustrated here, increases in signal intensity also occurred in the medullary raphé and in the region that includes the dorsal motor nucleus of the vagus. The use of this approach promises to increase our understanding of the functional organization of the medulla and changes in sympathetic outflow that are associated with many pathophysiological states.

References [1] McAllen RM, May CN, Shafton AD. Functional anatomy of sympathetic premotor cell groups in the medulla. Clin Exp Hypertens 1995;17:209–21. [2] Dampney RA, Polson JW, Potts PD, Hirooka Y, Horiuchi J. Functional organization of brain pathways subserving the baroreceptor reflex: studies in conscious animals using immediate early gene expression. Cell Mol Neurobiol 2003;23:597–616. [3] Wang J, Irnaten M, Neff RA, Venkatesan P, Evans C, Loewy AD, et al. Synaptic and neurotransmitter activation of cardiac vagal neurons in the nucleus ambiguus. Ann N Y Acad Sci 2001;940:237–46. [4] Minson JB, Llewellyn-Smith IJ, Arnolda LF, Pilowsky PM, Chalmers JP. C-fos expression in central neurons mediating the arterial baroreceptor reflex. Clin Exp Hypertens 1997;19:631–43.

[5] Horiuchi J, Killinger S, Dampney RA. Contribution to sympathetic vasomotor tone of tonic glutamatergic inputs to neurons in the RVLM. Am J Physiol 2004;287:R1335–43. [6] Critchley HD, Nagai Y, Gray MA, Mathias CJ. Dissecting axes of autonomic control in humans: insights from neuroimaging. Auton Neurosci. 2011;161:34–42. [7] Henderson LA, Richard CA, Macey PM, Runquist ML, Yu PL, Galons JP, et al. Functional magnetic resonance signal changes in neural structures to baroreceptor reflex activation. J Appl Physiol 2004;96:693–703. [8] Macefield VG, Gandevia SC, Henderson LA. Neural sites involved in the sustained increase in muscle sympathetic nerve activity induced by inspiratory-capacity apnea – a fMRI study. J Appl Physiol 2006;100:266–73. [9] Allen AM, Moeller I, Jenkins TA, Zhuo J, Aldred GP, Chai SY, Mendelsohn FA. Angiotensin receptors in the nervous system. Brain Res Bull 1998;47:17–28. [10] McAllen RM, Farrell M, Johnson JM, Trevaks D, Cole L, McKinley MJ, et al. Human medullary responses to cooling and rewarming the skin: a functional MRI study. Proc Nat Acad Sci USA 2006;103:809–13. [11] Sander M, Macefield VG, Henderson LA. Cortical and brainstem changes in neural activity during static handgrip and post-exercise ischemia in humans. J Appl Physiol 2010;108:1691–700. [12] Macefield VG, Henderson LA. Real-time imaging of the medullary circuitry involved in the generation of spontaneous muscle sympathetic nerve activity in awake subjects. Human Brain Mapp 2010;31:539–59.

I. INTRODUCTION

C H A P T E R

4 Peripheral Autonomic Nervous System Robert W. Hamill, Robert E. Shapiro, Margaret A. Vizzard are paired structures that are located bilaterally along the vertebral column. They extend from the superior cervical ganglia (SCG), located rostrally at the bifurcation of the internal carotid arteries, to ganglia located in the sacral region. All told, there are three cervical ganglia (the SCG, middle cervical ganglion, and inferior cervical ganglion, which is usually termed the cervicothoracic or stellate ganglion because it is a fused structure combining the inferior cervical and first thoracic paravertebral ganglia), eleven thoracic ganglia, four lumbar ganglia, and four to five sacral ganglia. More caudally, two paravertebral ganglia join to become the ganglion impar. Prevertebral ganglia are midline structures located anterior to the aorta and vertebral column, and are represented by the celiac ganglia, aortico-renal ganglia, and the superior and inferior mesenteric ganglia. Previsceral, or terminal ganglia, are small collections of sympathetic ganglia located close to target structures; they are also referred to as short noradrenergic neurons since their axons cover limited distances. Generally, the preganglionic fibers are relatively short and the postganglionic fibers are quite long in the SNS. The axons of these postsynaptic neurons are generally unmyelinated and of small diameter (5 μm). The target organs of sympathetic neurons include smooth muscle and cardiac muscle, glandular structures, and parenchymal organs (e.g., liver, kidney, bladder, reproductive organs, muscles, etc. (see Fig. 4.1)) as well as other cutaneous structures. The spinal cells of origin for the presynaptic input to sympathetic peripheral ganglia are located from the first thoracic to the second lumbar level of the cord, although minor variations exist. The principal neurons generally have been viewed as located in the lateral horn of the spinal gray matter (intermediolateral cell column-IML), but four major groups of autonomic neurons exist: intermediolateralis pars principalis (ILP), intermediolateralis pars funicularis (ILF), nucleus intercalatus spinalis (IC) and the central autonomic nucleus (CAN) or dorsal commissural nucleus (DCN) (anatomical nomenclature is nucleus intercalatus pars paraependymalis (ICPE)). For paravertebral ganglia, 85–90% of the presynaptic fibers originate from cell bodies in ILP or ILF. Prevertebral ganglia and terminal ganglia receive a larger proportion of preganglionic terminals from the CAN/DCN. The nucleus intercalatus also contributes preganglionic fibers, but the exact

The autonomic nervous system (ANS) is structurally and functionally positioned to interface between the internal and external milieu, coordinating bodily functions to ensure homeostasis (cardiovascular and respiratory control, thermal regulation, gastrointestinal motility, urinary and bowel excretory functions, reproduction, and metabolic and endocrine physiology), and adaptive responses to stress (flight or fight response). Thus, the ANS has the daunting task of ensuring the survival as well as the procreation of the species. These complex roles require complex responses, and depend upon the integration of behavioral and physiological responses that are coordinated centrally and peripherally. In 1898, Langley, a Cambridge University physiologist coined the term “autonomic nervous system” and identified three separate components (sympathetic, parasympathetic, and enteric). The following section of the Primer will focus on the first two aspects of the peripheral ANS: sympathetic nervous system (SNS) including the adrenal medulla; and parasympathetic nervous system (PNS). The following précis will address the neuroanatomy of the SNS, adrenal medulla, and PNS, and then present a more detailed, albeit brief, review of the functional neuroanatomy, physiology and pharmacology of the peripheral autonomic nervous system. Importantly, the ANS’s role at multiple interfaces in normal and abnormal physiology is emerging as a key mediator of pathophysiology in a range of complex disorders (anxiety and panic, chronic fatigue syndrome, regional pain syndromes, autonomic failure) and a critical substrate underpinning the field of neurocardiology. The following information will serve as a framework from which to view the complexity of the ANS as revealed in the more detailed descriptions that follow.

SYMPATHETIC NERVOUS SYSTEM (SEE FIG. 4.1) The SNS is organized at a spinal and peripheral level such that cell bodies within the thoracolumbar segments of the spinal cord provide preganglionic efferent innervation to sympathetic neurons that reside in ganglia dispersed in three arrangements: paravertebral, prevertebral, and previsceral or terminal ganglia. Paravertebral ganglia

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4. PERIPHERAl AuToNomIC NERvouS SySTEm

Parasympathetic Division

Brainstem & Spinal Cord

Sympathetic Division

Target Organs

Trunk

Trunk

Target Organs

Oliary Ganglion

III

Nucleus Ambigours Superior Cervical Ganglion

Parotid Gland

Middle Cervical Ganglion Stellate

Heart

Ganglion

Respiratory Tract & Lungs

Small Intestine Adrenal Medulla Small Intestine

Celiac Ganglion

Greater Splanchnic Nerve Lesser Splanchnic Nerve Superior Mesenteric Ganglion

Colon Pelvin Colon & Rectum Genitourinary Tract

Interior Mesenteric Ganglion

C1 C2 C3 C4 C5 C6 C7 C8 T1 T2 T3 T4 T5 T6 T7 T8 T9 T10 T11 T12 L1 L2 L3 L4 L5 S1 S2 S3 S4 S5

Submandibular Ganglion

Sweat Glands, Hair Follicles, Blood Vessels

Lacrimal & Submandibular Glands

Stomach

VII IX X X

Edinger-Westphal Nucleus Superior Salivatory Nucleus Inferior Salivatory Nucleus Dorsal Motor Nucleus

Eye

Pterygopalatine Ganglion

Otic Ganglion

Eye Lacrimal & Palatine Glands Submandibular & Sublingual Glands Parotid Gland

Respiratory Tract & Lungs Gastrointestinal Tract, Liver & Pancreas Heart Nasopharynx Esophagus

Pelvic Colon & Rectum Pelvic Nerve

Genitourinary Tract

Hypogastric Ganglion

FIGURE 4.1 Schematic diagram of the sympathetic and parasympathetic divisions of the peripheral autonomic nervous system. The paravertebral chain of the sympathetic division is illustrated on both sides of the spinal outflow in order to demonstrate the full range of target structures innervated. Although the innervation pattern is diagrammatically illustrated to be direct connects between preganglionic outflow and postganglionic neurons, there is overlap of innervation such that more than one spinal segment provides innervation to neurons within the ganglia.

extent of these is not fully understood and is probably limited. These spinal autonomic nuclei receive substantial supraspinal input from multiple transmitter systems located at multiple levels of the neuraxis; diencephalon (hypothalamus) and brainstem (raphe, locus coeruleus, reticular formation and ventral lateral medulla) provide the largest input and the pattern of innervation viewed in horizontal sections reveals a ladder like arrangement of the distribution of nerve terminals [1]. Without detailing the source or course of specific systems, it is important to point out that the following different neurotransmitter systems impinge on preganglionic neurons within this ladder like structure: monoamines – epinephrine, norepinephrine (NE), serotonin; neuropeptides – substance P, thyrotropin-release hormone (TRH), met-enkephalin, vasopressin, oxytocin, and neuropeptide Y (NPY); amino acids – glutamate, GABA, and glycine. Undoubtedly, others exist and more will be found. It is apparent that dysfunction of these supraspinal systems, or alterations of these

neurotransmitters by disease or pharmacological agents will alter the spinal control of peripheral ganglia and result in clinical dysfunction. The outflow from the spinal cord to peripheral ganglia is segmentally organized with some overlap. Retrograde tracing studies indicate that there is a rostral-caudal gradient: SCG receives innervation from spinal segments T1–T3; stellate ganglia – T1–T6; adrenal gland – T5–T11; celiac and superior mesenteric ganglia – T5–T12; inferior mesenteric and hypogastric ganglion from L1–L2. These presynaptic fibers, which are small in diameter (2–5 μm) and thinly myelinated, exit the ventral roots via the white rami communicantes to join the paravertebral chain either directly innervating their respective ganglion at the same level or traveling along the chain to innervate a target ganglion many levels away (Fig. 4.2). The distribution of postsynaptic fibers also follow a regional pattern with the head, face and neck receiving innervation from the cervical ganglia (spinal segments T1–4), the upper limb and

I. INTRODUCTION

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SymPATHETIC NERvouS SySTEm

Parasympathetic Nervous System

Sympathetic Nervous System Thoracolumbar Spinal Cord

Sacral Spinal Cord

Spinal ganglion Afferent autonomic fiber Vasomotor Sudomotor Pilomotor fibers

Preganglionic fiber Postganglionic fiber

Gray communicating ramus (post ganglionic) White communicating ramus (preganglionic)

Paravertebral ganglion

Splanchnic nerve

Descending colon Rectum Urinary Bladder Sexual organs

Pelvic splanchnic nerve Prevertebral ganglion Pelvic ganglion

Postganglionic visceral nerves to smooth muscle of inner organs & visceral blood vessels

FIGURE 4.2 Schematic illustration of the segmental spinal arrangement of the sympathetic and parasympathetic nervous system. Although segmental interactions exist, they are polysynaptic operating via interneurons; the primary input to spinal preganglionic neurons is supraspinal originating from brainstem structures (not shown).

thorax from the stellate and upper thoracic ganglia (spinal segments T1–8); the lower trunk and abdomen from lower thoracic ganglia (spinal segments T4–12), and the pelvic region and lower limbs from lumbar and sacral ganglia (spinal segments T10–L2). More recently, with the introduction of trans-neuronal tracing techniques, using pseudorabies virus (PRV), it has been possible to inject ganglia in the periphery and examine the trans-synaptic passage of PRV. Thus, supraspinal neurons projecting to the specific sets of preganglionic neurons that innervate the peripheral ganglia injected may be examined. Interestingly, a surprisingly common set of central pathways influencing the thoracolumbar sympathetic outflow were labeled. For instance, following injections in either the SCG, stellate or celiac ganglia, or the adrenal gland, the following five brain areas are labeled: ventromedial and rostral ventrolateral medulla; caudal raphe nuclei; A5 noradrenergic cell group; and the paraventricular nucleus of the hypothalamus [2]. Apparently, these central loci must share regulatory functions that are coordinated through similar pathways of thoracolumbar outflow. These same studies indicate that other brain areas are only labeled from specific ganglia; thus, site specific central control exists as well. Of additional interest is that numerous, small interneurons in Rexed laminae VII and X of the spinal cord were labeled providing structural support

for the observation that spinal intersegmental and intrasegmental autonomic interactions (including autonomic reflexes) exist. It is apparent that the structural organizational of the sympathetic nervous system permits the integration and dissemination of responses depending on demand. Multiple supraspinal descending pathways provide a dense innervation of all four major autonomic cell groups in the spinal cord, but clearly specific topographic responses exist as well. In turn, each preganglionic neuron innervates anywhere from 4 to 20 postganglionic sites, and each spinal outflow level may reach multiple peripheral ganglia which in turn supply multiple targets, permitting additional dispersion of sympathetic responses when indicated. At each thoracic level there are an estimated 5000 preganglionic neurons (these counts have generally been limited to the cells located in the ILP). Since preganglionic output to prevertebral ganglia originates from more medially placed cell bodies, it is conceivable that a greater number of neurons at certain segmental levels contribute to the output. Thus, a given spinal segment has a powerful base to influence greater than 100,000 postganglionic neurons. Although original thinking suggested that responses were “all-or-none and widespread”, anatomical studies have continued to reveal subtleties of structural arrangements that indicate that the system is not only poised for

I. INTRODUCTION

20

4. PERIPHERAl AuToNomIC NERvouS SySTEm

generalized activation of the peripheral sympathetic nervous system, but is also able to exert control of relatively specific sites and functions. The postganglionic fibers in the SNS travel quite lengthy paths to arrive at target organs. For instance, fibers from the SCG traverse the extracranial and intracranial vasculature to reach such targets as the lacrimal glands, parotid glands, pineal gland, and pupils. Fibers from the stellate ganglia course through the brachial plexus to reach vascular and cutaneous targets in the upper limb and hand. Within the abdomen, axons originating from the paravertebral ganglia supply the viscera as well as the mesenteric vasculature. Lumbar and sacral paravertebral ganglia course distally along peripheral nerves and blood vessels to reach the distal vasculature and cutaneous structures in the feet. In humans, the innervation to the leg requires a sympathetic axon to be 50 cm long and with an estimated overall diameter of 1.2 μm the axonal volume is approximately 565,000 μm3. This axonal cytoskeleton and its metabolic requirements are supported by a perikaryon of about 30 μm with a somal volume of 14,000 μm3. With this structural architecture to maintain, these neurons are vulnerable to various metabolic and structural insults. Although most preganglionic fibers have a relatively short course to their ganglion targets, the upper thoracic preganglionic fibers travel relatively longer distances to reach the stellate and SCG, and preganglionic fibers to the adrenal medulla and prevertebral ganglia course through the paravertebral chain reaching these visceral targets as the splanchnic nerves. Along the course, fiber systems may be interrupted, resulting in local autonomic dysfunction. For example, the Horner’s syndrome results from lesion of either preganglionic fibers to the SCG or the postganglionic axons which leave the SCG to innervate Müller’s muscle of the upper eyelid, pupillodilator muscles, facial vasculature, and sudomotor structures of the face (see Fig. 4.1). The autonomic neuroeffector junction is generally a poorly defined synaptic structure lacking the pre- and postjunctional specializations that are observed in the central nervous system or skeletal muscle motor end plates. The unmyelinated, highly branched postganglionic fibers become beaded with varicosities as they approach their targets. The varicosities are not static; they move along as structures with a diameter of 0.5 to 2 μm, and a length of approximately 1.0 μm. The number of varicosities varies from 10,000/mm3 to over 2 million per mm3 depending on the target being innervated. The varicosities are packed with mitochondria and vesicles containing various transmitters, and are at varying distances from their target organs. For instance, for smooth muscle targets, this distance varies from 20 nm in the vas deferens to 1–2 μm in large arteries. In a sense the release of transmitter is accomplished en passage as the impulse travels along an autonomic axon. The lack of a restrictive synaptic arrangement permits the released NT to diffuse various distances along a target organ and activate multiple receptors,

again expanding the overall effect of sympathetic activation. Between 100 and 1000 vesicles exist in each varicosity in noradrenergic fibers. Traditional teaching suggests that vesicle characteristics indicate the transmitter system: small granular vesicles are noradrenergic; small agranular vesicles are cholinergic; large granular vesicles are peptidergic. However, exceptions to these correlations exist. The principal neuronal phenotype in peripheral sympathetic ganglia is the noradrenergic neuron which is generally multipolar in character with synapses mainly located more on dendrites than somata. Depending on which ganglia are examined, studies indicate that from 80–95% of ganglion cells will stain positively for tyrosine hydroxylase, the rate-limiting enzyme in catecholamine biosynthesis, or have positive catecholamine fluorescence. The remaining cells have a mixture of transmitters, or are postganglionic cholinergic cells (the sudomotor and periosteal components of sympathetic function). Within sympathetic ganglia there is a small group of small intensely fluorescent (SIF) neurons. The transmitters identified in SIF cells include dopamine, epinephrine, or serotonin. As will be described later, the original concept that preganglionic neurons in the SNS are cholinergic and postganglionic neurons are noradrenergic has given way to new information that a whole array of molecules (cholinergic, catecholaminergic, monoaminergic, peptidergic, “non-cholinergic, non-adrenergic”, and gaseous) appear to be involved in neurotransmission either as agents themselves or as neuromodulators (vide infra).

SYMPATHOADRENAL AXIS AND THE ADRENAL GLAND Interactions between the adrenal cortex and adrenal medulla constitute a critical link between the autonomic and endocrine systems. The adrenal cortex is largely regulated by the hypothalamic-pituitary-adrenocortical axis, whereas the adrenal medulla is primarily under neural control. Both adrenal cortex and medulla, respond to stress and metabolic aberrations. The coordinated response of elevated plasma cortisol and catecholamines during stress indicate that central limbic and hypothalamic centers exert combined influences to ensure the needed neurohumoral adaptations. The interdependence of these two components of the adrenal gland arise early in development: migrating sympathoblasts destined for the adrenal medulla require the presence of the cortical tissue to change their developmental fate from neurons to that of chromaffin cells. These cells, named because they exhibit brown color when treated with “chrome salts”, do not develop neural processes but instead serve an endocrine function by releasing their neurohumors (epinephrine, norepinephrine, and neuropeptides) into the blood stream. During adulthood, the presence of the cortex is critical for maintaining the levels of epinephrine since the induction of the enzyme

I. INTRODUCTION

PARASymPATHETIC NERvouS SySTEm

phenylethanolamine-n-methyltransferase is dependent on local levels of cortisol. Although traditional teaching emphasizes the preganglionic cholinergic splanchnic innervation of the adrenal medulla, there is also evidence that postganglionic sympathetic fibers, vagal afferents, and other sensory afferents are present. Tracing studies indicate that dye placed within the adrenal medulla is transported retrogradely within spinal preganglionic sympathetic neurons in a somewhat bell shaped distribution from approximately T2–L1 with the predominant innervation originating from T7–T10. Neuronal cell bodies are primarily within the nucleus intermediolateralis pars principalis (ILP) with the pars funicularis and pars intercalatus providing a relatively small portion of the innervation. The exiting nerve roots pass through the sympathetic chain, join to form the greater splanchnic nerve, and distribute themselves beneath the adrenal capsule and within the medulla. A small number of nerve cells are labeled in ganglia within the sympathetic chain suggesting that postganglionic sympathetic fibers innervate the gland. Whether these terminals are labeled as they pass along blood vessels within the gland or whether they innervate medullary or cortical cells is not fully resolved. Also, at least in the guinea pig, tracing studies indicate that the parasympathetic system may contribute a small efferent innervation to the gland since neurons in the dorsal motor nucleus of the vagus are labeled after injections in the medulla. Also, cell bodies within the dorsal root ganglia and vagal sensory ganglia (nodose) are also labeled after tracer studies of the adrenal medulla indicating an afferent innervation as well. Lastly, although not a prominent innervation pattern, there appears to be an intrinsic innervation that arises from ganglion cells sparsely populating the subcapsular, cortical and medullary regions of the gland. The innervation pattern of the adrenal medulla is thus more complex than the traditionally listed thoracolumbar preganglionic cholinergic outflow, although the major adaptive responses depend on the preganglionic cholinergic innervation since surgical section of these nerves, or pharmacological blockade with cholinergic antagonists preclude the induction of tyrosine hydroxylase and appropriate release of catecholamines following various stress paradigms. Morphological studies of the adrenal medulla have revealed the presence of two basic types of granules in chromaffin cells. A diffuse spherical granule contains the predominant monoamine secreted by medullary cells, epinephrine, whereas eccentrically located dense core granules contain norepinephrine. As indicated above for ganglion neurons, chromaffin cells of the adrenal medulla also cocontain other molecules. For instance, the opioid molecules are well represented: enkephalin is co-contained in vesicles with the monoamines. The signaling cascade responsible for enhancing the synthesis and release of these neurohormonal agents is complicated and includes preganglionic innervation, steroid hormones (e.g., glucocorticoids), and growth factors (e.g., nerve growth factor, NGF).

21

PARASYMPATHETIC NERVOUS SYSTEM (SEE FIG. 4.1) The craniosacral outflow is the source of central neuronal pathways providing the efferent innervation of peripheral ganglia of the parasympathetic nervous system (PNS). The cranial nerves involved include cranial nerves III, VII, IX, and X, and the sacral outflow is largely restricted to sacral cord levels 2, 3, and 4. As indicated for the SNS, the preganglionic innervation is largely cholinergic with these terminals releasing acetylcholine (ACh) at the ganglion synapses. In contrast to the SNS, the major transmitter postsynaptically is also ACh. These cholinergic neurons also co-contain other transmitter substances: preganglionic neurons contain enkephalins, and ganglionic cholinergic neurons frequently contain vasoactive intestinal peptide (VIP) and/or NPY. The parasympathetic fibers in cranial nerve III originate in the Edinger–Westphal nuclei of the midbrain and travel in the periphery of the nerve (where they are subject to dysfunction secondary to nerve compression), exiting along with the nerve to the inferior oblique to supply the ciliary ganglion. Second order postganglionic fibers exit in the ciliary nerves and supply the pupiloconstrictor fibers of the iris and the ciliary muscle where their combined action permits the near response, including accommodation. The salivatory nuclei, located near the pontomedullary junction, provide the preganglionic parasympathetic innervation for cranial nerves VII and IX. The superior salivatory nucleus sends preganglionic fibers, which leave the facial nerve at the level of the geniculate ganglion (non-parasympathetic sensory ganglion), to form the greater superficial petrosal nerve to the pterygopalatine (sphenopalatine) ganglia that provides postganglionic secretomotor and vasodilator fibers to the lacrimal glands via the maxillary nerve. Other preganglionic fibers in the facial nerve continue and subsequently leave via the chorda tympani to join the lingual nerve, eventually synapsing in the submandibular ganglion. Postsynaptic cholinergic fibers supply the sublingual and submandibular glands. Postganglionic fibers from the pterygopalatine and submandibular ganglia also supply glands and vasculature in the mucosa of the sinuses, palate, and nasopharynx. The inferior salivatory nucleus sends preganglionic fibers via the glossopharyngeal nerve (cranial nerve IX) to the otic ganglion that in turn relays postganglionic fibers to the parotid gland via the auriculotemporal nerve. The preganglionic fibers in cranial nerve IX branch from the nerve at the jugular foramen and contribute to the tympanic plexus and form the lesser superficial petrosal nerve. This nerve exits the intracranial compartment via the foramen ovale along with the third division of the trigeminal nerve to reach the otic ganglion. The most caudal cranial nerve participating in the preganglionic parasympathetic system is the vagus nerve (cranial nerve X). The dorsal motor nucleus of the vagus is located in the medulla and sends preganglionic fibers to innervate essentially all organ systems within the chest

I. INTRODUCTION

22

4. PERIPHERAl AuToNomIC NERvouS SySTEm

and abdomen including the gastrointestinal tract as far as the left colonic flexure (splenic flexure). Also, the nucleus ambiguus supplies preganglionic fibers to the vagus and these fibers are believed to be involved mostly with regulating visceral smooth muscle whereas the dorsal motor vagus neurons may be secretomotor in nature. The glossopharyngeal and vagus nerves also contain a substantial number of afferent fibers (in the vagus, afferent fibers may exceed the efferent fiber system by a ratio of 9 to 1) so that a sensory component related to autonomic control exists within cranial nerve IX and X. These afferents provide a critical component of the baroreceptor reflex arc relaying information regarding the systemic blood pressure to central cardiovascular areas in the nucleus tractus solitarii and other medullary centers involved in blood pressure and heart rate control. The parasympathetic cell bodies in the spinal cord are located in the sacral parasympathetic nucleus of second, third, and fourth sacral segments (see Fig. 4.2). These neurons send preganglionic nerve fibers via the pelvic nerve to ganglia located close to or within the pelvic viscera. Postganglionic fibers are relatively short, in contradistinction to their length in the SNS, and supply cholinergic terminals to structures involved in excretory (bladder and bowel) and reproductive (fallopian tubes and uterus, prostate, seminal vesicles, vas deferens, and erectile tissue) functions. Of interest, the pelvic ganglia involved in some of these functions appear to be mixed ganglia (especially in rodents) where sympathetic and parasympathetic neurons are components of the same pelvic ganglion. They appear to receive their traditional preganglionic input, but may have local interconnections that are not fully revealed by current studies. As indicated below, there is clear evidence that the cholinergic postganglionic neurons in the pelvic ganglion in the rodent co-contain vasoactive intestinal peptide and nitric oxide as two other neuroactive compounds. These neurons are believed to be integrally involved in sexual functions in the male, permitting the development and maintenance of potency. The exact regulatory factors controlling the synthesis and release of these transmitter molecules, and the specific receptor systems involved remain to be fully explored.

THE CONCEPT OF PLURICHEMICAL TRANSMISSION AND CHEMICAL CODING The notion of the presence of multiple transmitters and a chemical coding system of autonomic neurons is now firmly established. Originally it was posited that principal neurons were only noradrenergic (contained NE), but over the last two decades it has become clear that within a single neuron multiple transmitter systems may exist, and that within a given ganglion the variety and pattern of neurotransmitters may be quite extensive (see Table 4.1). Also, the composition of neurotransmitters (NT) may change depending on the location of the ganglia: paravertebral

TABLE 4.1 Neurotransmitter Phenotypes in Autonomic Neurons Autonomic Neurons Sympathetic neurons Paravertebral ganglia Prevertebral ganglia Terminal ganglia (previsceral ganglia) Parasympathetic neurons Major parasympathetic ganglia Ciliary Sphenopalatine Otic Submandibular/sublingual Pelvic ganglia Terminal parasympathetic ganglia (previsceral ganglia) Enteric neurons Myenteric plexus (Auerbach’s) Submucosal plexus (Meisner’s)

Transmitter Characteristics (not all inclusive) NE, CCK, somatostatin, SP, Enk, Ach VIP, 5-HT, NPY, DYN1-8, DYN1-17

ACh, VIP, SP, CAs-SIF, NPY, NO

GABA, ACh, VIP, 5-HT SP, Enk, SRIF, motilinlike peptide, bombesinlike peptide

Enteric ganglia Chromaffin cells of adrenal medulla Paraganglia-chromaffin

E, NE, Enk, NPY, APUD

SIF cells, ganglia

ganglia tend to have fewer transmitters whereas prevertebral and terminal ganglia may have various NT, although as noted for the guinea pig SGC (Fig. 4.3), some paravertebral ganglia have a broad array of transmitters. The exact co-location and functions of these multiple transmitters are not fully understood, but some general principles exist. Neuropeptide NPY is probably the most prominent peptide in sympathetic ganglia and is highly co-localized with NE. The sudomotor component of ganglia is dependent upon a population of cells, which are cholinergic in character [contain ACh as NT], and the most frequent peptide colocalized with ACh is vasoactive intestinal peptide (VIP). The distribution of these cholinergic cells varies: in paravertebral ganglia they may represent 10–15% of the neuronal population whereas they represent 1% of the neurons in prevertebral ganglia. NE  NPY, and ACh  VIP are believed to be released together, but some degree of activity associated-chemical coding exists. That is, at lower levels of activation NE is preferentially released, whereas higher levels of stimulation result in NPY being released. Both agents have vasoconstrictor properties and are integral in cardiovascular control, especially in the maintenance of blood pressure. Of course, the eventual action and effect of a transmitter rests with the receptor system that is activated (vide infra). Purinergic neurotransmission expands the co-transmission motif. Presynaptic and postsynaptic mechanisms exist: the purine nucleotide adenosine triphosphate (ATP) is in high concentration in sympathetic synaptic vesicles and following release ATP is catabolized to adenosine

I. INTRODUCTION

vISCERAl AffERENT NEuRoNS ANd AuToNomIC NERvouS SySTEm

23

FIGURE 4.3 Chemical coding and target organization. Chemical coding of sympathetic neurons projecting from the superior cervical ganglion to various targets in the head of guinea-pigs. Each population of neurons has a specific combination of neuropeptides. Note that all neurons containing a form of dynorphin A (DYN1-8 or DYN1-17) also contain dynorphin B and neo-endorphin. No neuropeptides have been found in neurons projecting to secretory tissue in the salivary or lacrimal glands. Neurons with similar peptide combinations also occur in other paravertebral ganglia of guinea-pigs, except that the salivary secretomotor neurons are absent. Conversely, the paravertebral ganglia have many non-noradrenergic vasodilatory neurons containing prodynorphin-derived peptides, VIP and NPY. AVAs, arterio-venous anastomoses; NA, noradrenaline; NPY, neuropeptide Y; ACh, acetylcholine; CGRP, calcitonin gene-related peptide; SP, substance P; VIP, vasoactive intestinal polypeptide (see Elfvin, Lindh and Hokfelt, 1993 [11]; Jänig, 2006 [12]).

moieties. There are at least eight receptors (four purinergic and four adenosinergic) that serve to translate purinergic and adenosinergic effects (vasoconstriction and vasodilatation) to vascular beds via endothelium-dependent and endothelium-independent mechanisms. Chemical coding also reveals that ganglion neurons with specific transmitter molecules innervate specific targets or receive specific afferent inputs. Apparently, anterograde and retrograde transsynaptic information appears to determine the transmitter phenotype of the neuron. Thus, studies of neuronal circuitry indicate that pathwayspecific combinations determine the presence and combinations of specific peptides within autonomic neurons. This is particularly so in the prevertebral ganglia, but studies in the guinea pig SCG demonstrate that the transmitter molecules will vary depending on the target organ supplied (see Fig. 4.3). Although all principal cells portrayed are noradrenergic in character (as indicated by NA), the neuropeptides co-contained in neurons vary depending on whether the targets are secretory (salivary and lacrimal glands), vascular (small vs. large arteries, arterioles vs. arterio-venous anastomoses), pupil or skin. Detailed pictures of these chemically coded circuits are beginning to emerge from studies of paravertebral (SCG), prevertebral (superior mesenteric), and previsceral (pelvic) ganglia. This phenomenon pertains to both the SNS and PNS. Preganglionic sympathetic neurons in the spinal cord have traditionally been viewed as cholinergic neurons.

More recently, it has been recognized that neuronal cell bodies in the cat ILP may contain a variety of transmitters including enkephalin, neurotensin, somatostatin, and substance P. In rodents, VIP and calcitonin gene-related pepitde (CGRP) containing neurons have also been localized to the ILP by immunocytochemistry. Also, preganglionic fibers in the sacral parasympathetic outflow co-contain enkephalin. It is apparent that neurotransmission is plurichemical in both preganglionic and postganglionic-fiber systems.

VISCERAL AFFERENT NEURONS AND AUTONOMIC NERVOUS SYSTEM Visceral afferent neurons are polymodal and are excited by physical (distention, contraction), chemical and thermal events. These stimuli are transmitted to the spinal cord and brainstem neurons resulting in target organ regulation, reflexes and sensation. In rats, vagal afferent neurons arising in the stomach may terminate within the medullary nucleus tractus solitarius in proximity to dendrites from dorsal motor nucleus efferent neurons sending axons back to the stomach [3]. Spinal primary afferent neurons that innervate viscera have multiple functions (afferent, efferent and trophic functions) [4] and contain a diverse assortment of neuropeptides including calcitonin generelated peptide (CGRP), substance P, vasoactive intestinal

I. INTRODUCTION

24

4. PERIPHERAl AuToNomIC NERvouS SySTEm

polypeptide (VIP) and pituitary adenylate cyclase activating polypeptide (PACAP) [5]. For example, bladder afferent neurons travel in the hypogastric and pelvic nerves, and their cell bodies are located in dorsal root ganglia (DRG) at spinal segments T11–L2 and S2–S4 in humans and L1–L2 and L6–S1 in rats [5,6]. Bladder afferent fibers consist of lightly myelinated Aδ fibers and unmyelinated C-fibers. Sensation of bladder filling is conveyed by Aδ fibers, the most important mechanoreceptors of the bladder. C-fibers are normally “silent”, but they do respond to chemical or noxious stimuli, including extreme bladder pressure [5,6].

FUNCTIONAL NEUROANATOMY AND BIOCHEMICAL PHARMACOLOGY The peripheral ANS is well structured to provide the physiological responses critical for homeostasis and acute adaptations to stressful, perhaps life-threatening, circumstances. As outlined in Figure 4.1 and Table 4.2, multiple organ systems respond to neurotransmitters released from autonomic endings, and circulating catecholamines released from the adrenal medulla. Traditional teaching is that the effects of activation of the SNS and PNS are generally antagonistic; this is still largely the case. However, viewed more specifically, the relationships between these two major components of the ANS are far from simple. For instance, not all organs receive an equal number of both sets of fibers, and in some situations, both SNS and PNS effects are similar. It is important to remember that receptor systems, including the signal transduction components, on the target organs are the critical molecular proteins that determine the actual effects of ligand-receptor interactions on the cell membrane. Thus, when the SNS is stimulated with its capability to produce a widespread response, a host of receptor systems are activated to effect the necessary and desired change. An example of these responses includes the following aspects of SNS activation: dilatation of the pupil; slight increase in glandular secretions; bronchodilatation; increased heart rate and force of contraction; decreased gastrointestinal tract motility; decreased function of the reproductive organs; and mobilization of energy substrates to meet demands. The receptor systems mediating these responses include α1A, α1B,, α1D,, α2A, α2B, α2C, β1, β2 and β3 receptors. α1 receptors have subtypeselective distributions, second messenger systems and functions. Activation of these receptors occurs following interaction with NE and variously results in contraction of smooth muscle in the vasculature and iris, and relaxation of smooth muscle in the gut. α1 receptors exert a limited positive inotropic effect on the heart, as well as mediate salivary gland secretion and contraction of the prostate gland. α2 receptors also have discrete subtype-specific localizations and functions in brain, peripheral nerve, and target tissues. α2 receptors serve as autoreceptors on sympathetic nerve terminals and inhibit the release of NE as

TABLE 4.2 Autonomic Nervous System functions Organ

Sympathetic Nervous System

Parasympathetic Nervous System

Eye Pupil Ciliary muscle

Dilatation Relax (far vision)

Constriction Constrict (near vision)

Lacrimal gland

Slight secretion

Secretion

Parotid gland

Slight secretion

Secretion

Submandibular gland

Slight secretion

Secretion

Heart

Increased rate Positive inotropism

Slowed rate Negative inotropism

Lungs

Bronchodilation

Bronchodilation

Gastrointestinal tract

Decreased motility

Increased motility

Kidney

Decreased output

None

Bladder

Relax detrusor Contract sphincter

Contract detrusor Relax sphincter

Penis

Ejaculation

Erection

Sweat glands

Secretion

Palmar sweating

Piloerection muscles

Contraction

None

Blood vessels Arterioles

Constriction

None

Constriction or dilatation Glycogenolysis

None

Muscle Arterioles Metabolism

None

part of a negative feed back loop. α2 receptors also can act as constrictors of both arterial and venous vascular smooth muscle. Furthermore, these receptors play important roles in nociception and also mediate metabolic and endocrine changes such as inhibition of lipolysis in adipose tissue and reduction of insulin release from the pancreas. β receptors (β1) provide positive inotropic and chronotropic effects on the heart and stimulate renin release from the kidney. β2 receptors relax smooth muscle of the bronchi and pelvic organs as well as the vascular structures of the gut and skeletal muscle. β2 receptors located in liver and skeletal muscle elicit activation of glycogenolysis and gluconeogenesis. This receptor system is particularly activated by epinephrine rather than norepinephrine. The parasympathetic nervous system is poised for more focal responses, but some effects may be quite broad, particularly with the wide-ranging innervation of the vagus nerve. Activation of parasympathetic pathways lead to pupillary constriction, substantial secretion from lacrimal and salivary glands, slowed cardiac rate and negative inotropism, bronchoconstriction, enhanced gastrointestinal motility and contraction of the detrusor muscle of the bladder. In contrast to the SNS, the PNS does not appear to influence metabolic or endocrine processes in any major way. However, recent evidence indicates that preganglionic vagal fibers originating in the dorsal motor nucleus innervate postganglionic parasympathetic ganglia in the

I. INTRODUCTION

STRESS ANd AuToNomIC dySfuNCTIoN

pancreas, and appear to influence exocrine and endocrine function. Thus, as new data emerge, the integrated roles of the SNS and PNS will continue to expand. An understanding of the receptor systems mediating the responses to PNS activation is incomplete. Preganglionic cholinergic receptors, which exist in the SNS as well as PNS, are nicotinic in character whereas postganglionic cholinergic receptors are muscarinic. Recently, molecular cloning studies have revealed multiple subtypes of both sets of receptors. The subtypes of muscarinic receptors (mAChRs), M(1)-M(5), are more fully understood (at least for the first three subtypes): M1 receptors are excitatory to neurons in ganglia and lead to noradrenaline release in sympathetic neurons; M2 receptors mediate the bradycardia and decreased contractility of the heart following vagal activation; M3 stimulation leads to contraction of smooth muscle and enhanced secretion from glandular tissues. Nicotinic acetylcholine receptor subtypes (nAChRs) are widely expressed in the mammalian CNS and PNS, playing a central role in autonomic transmission. Neuronal nicotinic acetylcholine receptors (nAChR) are composed of 12 subunits (α2–α10, β2–β4) but very little was known, until recently, about their physiological roles. The repertoire of nicotinic subunits in autonomic ganglia includes α3, α5, α7, β2 and β4 subunits. In the periphery, nicotinic receptors mediate vital excitatory fast synaptic cholinergic transmission at both the neuromuscular junction and ganglia. However, there is only a limited understanding of subunit actions in the presynaptic and postsynaptic components of the peripheral ANS. Functional deletions of subunit by gene knockout in animals are beginning to overcome these limitations.

STRESS AND AUTONOMIC DYSFUNCTION A prominent role for stress in the pathophysiology of cardiovascular disease and presentation of clinical pain states, including functional gastrointestinal disorders have been well documented. In addition, anxiety and stress may generate and worsen urinary symptoms and functional urinary disorders [7], such as interstitial cystitis (IC)/bladder pain syndrome (BPS). The majority of IC/BPS patients report symptom exacerbation by stress, and clinical studies have shown that acute stress increases bladder pain and urgency in IC/BPS patients [7,8]. Clinical populations afflicted with bladder disorders have a greater incidence of comorbid anxiety disorders than populations without bladder dysfunction. Both anxiety and bladder dysfunction can be caused and/or exacerbated by exposure to environmental stressors in human clinical populations and animal models. A number of ongoing studies from various laboratories are evaluating whether autonomic nervous system pathology and stress/anxiety share overlapping neuronal circuitry. The complexity of autonomic control and the range of mechanisms available to peripheral sympathetic and

25

parasympathetic neurons and their targets are expanded by presynaptic gaseous molecules (nitric oxide [NO]) and postsynaptic endothelium released peptides. Also, central nervous system centers involved in autonomic control contain nitric oxide synthase and evidence suggests that NO may mediate sympatho-inhibition or sympathoexcitation depending on the nuclear groups involved. Peripherally, NO exerts tonic vasodilation and mediates acetylcholine-induced vasodilation, as well as mediating catecholamine release and action. Endothelins, released abluminally by endothelial cells, bind to endothelial and adrenoceptors on vascular smooth muscle and endothelial cells and appear to regulate sympathetic terminals as well. The recent discoveries that a number of neurotrophic factors (neurotrophins) and their receptors are part and parcel of the development and integrity of the SNS and PNS expand the horizons regarding how anatomical pathways are established and maintained, and adapt to intrinsic and extrinsic demands. Because of its relatively simple anatomical architecture, the peripheral ANS continues to serve as a model system to understand neuronal development, structural and functional linkages among neurons and their targets, and the integrative role(s) these “little brains” (6000–30,000 neurons) serve. More recently, observations of the clinical phenotype and neuropathology of Parkinson Disease (PD) suggest that the peripheral ANS may serve as a model to study human neurodegenerative disease since peripheral and preganglionic, including medullary preganglionic vagal (DMV) neurons, may be involved at the earliest stages of the disease [9], before the onset of the well-known motor system dysfunctions related to the basal ganglia. A neuropathological cascade that originates in peripheral autonomic nerves may gradually and progressively ‘spread’ to involve central nervous system structures with the eventual involvement of the classically described neurons of the locus, coeruleus, substantia nigra and more rostral structures. Recent investigations in animal models of PD have taken advantage of the relative simplicity of the peripheral ANS to explore pathophysiological mechanism of how α-synuclein might initiate the premotor clinical dysfunction and presumed underlying alteration is neurotransmitter biochemistry and neurophysiology of peripheral autonomic neurons [10]. As the structure-function relationships and molecular pharmacology of the peripheral ANS are clarified, and more understandings accrue regarding their involvement in disease processes, new approaches to understanding ANS functions in health and disease and treatments of its maladies will undoubtedly emerge. The following sections of this Primer will expand on these issues.

References [1] Romagnano MA, Hamill RW. Spinal sympathetic pathway: an enkephalin ladder. Science 1984;225:737–9. [2] Strack AM, Sawyer WG, Hughes JH, Platt KB, Loewy AD. A general pattern of CNS innervation of the sympathetic outflow

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[3] [4]

[5]

[6] [7]

[8] [9] [10]

[11] [12]

4. PERIPHERAl AuToNomIC NERvouS SySTEm

demonstrated by transneuronal pseudorabies viral infections. Brain Res 1989;491:156–62. Shapiro RE, Miselis RR. The central organization of the vagus nerve innervating the stomach of the rat. J Comp Neurol 1985;238:473–88. Jänig W, Habler HJ. Specificity in the organization of the autonomic nervous system: a basis for precise neural regulation of homeostatic and protective body functions. Prog Brain Res 2000;122:351–67. Arms L, Vizzard MA. Neuropeptides in lower urinary tract function. In: Urinary tract, handbook of experimental pharmacology 202. Berlin Heidelberg: Springer-Verlag; 2011. Fowler CJ, Griffiths D, de Groat WC. The neural control of micturition. Nat Rev Neurosci 2008;9(6):453–66. Baldoni F, Ercolani M, Baldaro B, Trombini G. Stressful events and psychological symptoms in patients with functional urinary disorders. Percept Mot Skills 1995;80:605–6. Koziol JA, Clark DC, Gittes RF, Tan EM. The natural history of interstitial cystitis: a survey of 374 patients. J Urol 1993;149:465–9. Braak H, Del Tredici K. Invited Article: Nervous system pathology in sporadic Parkinson disease. Neurology 2008;70:1916–25. Hamill RW, Girard B, Tompkins JD, Galli JR, Parsons RL, Kershen RT, et al. Synucleinopathy model: Autonomic plasticity in the major pelvic ganglion (MPG) in mice with human alpha-synuclein overexpression. Developmental Neurobiol, in revision. Elfvin L-G, Lindh B, Hökfelt TJ. The chemical neuroanatomy of sympathetic ganglia. Ann Rev Neurosci 1993;16:471–507. Jänig W. The Integrative Action of the Autonomic Nervous System Neurobiology of homeostasis. Cambridge: Cambridge University Press; 2006.

Further Reading Baloh RH, Enomoto H, Johnson Jr. EM, Milbrandt J. The GDNF family ligands and receptors-implications for neural development. Curr Opin Neurobiol 2000;10:103–10. Burnstock G, Milner P. Structural and chemical organization of the autonomic nervous system with special reference to non-adrenergic, non-cholinergic transmission. In: Banister R, Mathias CJ, editors. Autonomic failure (4th edn.). New York: Oxford University Press; 1999. p. 63. Chowdhary S, Townend JN. Role of nitric oxide in the regulation of cardiovascular autonomic control. Clin Sci (Lond) 1999;97(July):5–17. Dinner DS, editor. The autonomic nervous system. (Review articles) J Clin Neurophysiol, 10; 1993 1–82 Gibbins I. Chemical neuroanatomy of sympathetic ganglia. In: McLachlan EM, editor. Autonomic ganglia. Luxembourg: Harwood Academic Publishers; 1995. p. 73–121. Goldstein DS. The autonomic nervous system in health and disease. New York: Marcel Dekker, Inc; 2001. 23–135 Schober A, Unsicker K. Growth and neurotrophic factors regulating development and maintenance of sympathetic preganglionic neurons. Int Rev Cytol 2001;205:37–76. Zansinger J. Role of nitric oxide in the neural control of cardiovascular function. Cardiovasc Res 1999;43:639–49.

I. INTRODUCTION

C H A P T E R

5 Cotransmission Geoffrey Burnstock

EARLY STUDIES

TABLE 5.1 Cotransmitters in the Peripheral and Central Nervous System

For many years, understanding of neurotransmission incorporated the concept that one neuron releases only a single transmitter, known as “Dale’s Principle”. This idea arose from a widely adopted misinterpretation of Dale’s suggestion in 1935 that the same neurotransmitter was stored in and released from all terminals of a single neuron, a suggestion which did not specifically preclude the possibility that more than one transmitter may be associated with the same neuron. Early hints that nerves might release more than one transmitter began in the 1950s with evidence for the involvement of both noradrenaline (NA) and acetylcholine (ACh) in sympathetic transmission. Koelle identified acetylcholinesterase in some adrenergic neurons in 1955, while Burn and Rand introduced the concept of a “cholinergic” link in adrenergic transmission in 1959. Another line of evidence provided by Hillarp concerned the coexistence of adenosine 5-triphosphate (ATP) with catecholamines, first in adrenal chromaffin cells and later in sympathetic nerves. Inconsistencies in the single transmitter hypothesis provided by these and other studies, including those concerned with invertebrate neurotransmission, from the early literature were rationalized in an article by Burnstock in 1976 [1] with the provocative title: “Do some nerve cells release more than one transmitter?” Today, it is widely accepted that cotransmission is an integral feature of neurotransmission. A role for ATP as a cotransmitter in sympathetic, parasympathetic, sensory-motor and enteric non-adrenergic, non-cholinergic (NANC) inhibitory nerves was supported by research from Burnstock and colleagues, while Hökfelt and colleagues focused on the colocalization, vesicular storage and release of peptides from both peripheral and central nerves (see [2–4]). Furness and Costa introduced the concept of “chemical coding” to describe the combination of potential neurotransmitters found in enteric nerves and this concept has since been applied to other nerve types, in both peripheral and central nervous systems (CNS) [5]. Colocalized substances are not necessarily cotransmitters, they can (especially peptides) act as pre- and/or postjunctional neuromodulators of the release and actions of the principal cotransmitters. The proportions of cotransmitters vary considerably between species and organs, and show

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00005-6

PERIPHERAL NERVOUS SYSTEM Sympathetic nerves

ATP  NA  NPY

Parasympathetic nerves

ATP  ACh  VIP

Sensory-motor

ATP  CGRP  SP

NANC enteric nerves

ATP  NO  VIP

Motor nerves (in early development)

ATP  Ach

CENTRAL NERVOUS SYSTEM Cortex, caudate nucleus

ATP  Ach

Hypothalamus, locus ceruleus

ATP  NA

Hypothalamus, dorsal horn, retina

ATP  GABA

Mesolimbic system

ATP  DA

Hippocampus, dorsal horn

ATP  glutamate

Compiled from [4]. ACh, acetylcholine; ATP, adenosine 5-triphosphate; CGRP, calcitonin gene-related peptide; DA, dopamine; GABA, γ-amino butyric acid; NA, noradrenaline; NO, nitric oxide; NPY, neuropeptide Y; SP, substance P; VIP, vasoactive intestinal peptide.

plasticity of expression during development and in pathological conditions. In general, classical transmitters are contained in small synaptic vesicles, whereas peptides are stored in large granular (dense-cored) vesicles (LGVs), although small molecule transmitters are sometimes stored together with peptides in LGVs. Pharmacological studies of pre- and postjunctional neuromodulation provide evidence, which is complementary to the concept of cotransmission. Evidence for ATP being a cotransmitter with established neurotransmitters in the CNS as well as in the periphery has been reported (see Table 5.1).

SYMPATHETIC NERVES There is substantial evidence to show that NA, ATP and neuropeptide Y (NPY) are cotransmitters in sympathetic nerves, having differentially important roles as transmitters and neuromodulators depending on the tissue, the species, and on the parameters of stimulation.

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© 2012 Elsevier Inc. All rights reserved.

28

5. CoTRANSmISSIoN

The first indication that ATP might be released from sympathetic nerves was the demonstration that stimulation of sympathetic nerves by Su et al. in 1971 [6] led to release of tritium from taenia coli preincubated in [3H]adenosine (which is taken up and converted to [3H]ATP). Later Sol Langer and colleagues suggested that the substantial residual NANC response of the cat nictitating membrane observed after depletion of NA by reserpine was due to the release of ATP remaining in the sympathetic nerves. Most of the early studies establishing the model of cotransmission of NA and ATP were made by Dave Westfall and colleagues on the vas deferens, a tissue with a high density of sympathetic nerves. In retrospect, there was a good indication that the excitatory junction potentials (EJPs) recorded in the guinea pig vas deferens when the electrophysiology of sympathetic nerve smooth muscle transmission was first described by Burnstock and Holman in 1960, were due to ATP released as a cotransmitter from sympathetic nerves, rather than to NA. It was puzzling at the time that adrenoceptor antagonists failed to block the EJPs, although guanethidine, a drug that prevents the release of sympathetic transmitters, was effective. It was over 20 years later that NANC EJPs were shown to be blocked by desensitization of the ATP (P2) receptors by α,β-methylene ATP (α,β-meATP) and mimicked by ATP [7]. After destruction of sympathetic nerves with 6-hydroxydopamine, purinergic nerve-mediated responses were abolished. ATP is co-stored with NA in small and large vesicles. Differential prejunctional modulation of the release of NA and ATP by various agents has been shown in the vas deferens, perhaps indicating that NA and ATP are stored in different vesicles. Cotransmission of NA and ATP in perivascular sympathetic nerves supplying the rabbit aorta, portal vein and saphenous, pulmonary and mesenteric arteries, and the dog basilar artery has been demonstrated (see [8]). Electrophysiological studies have shown that in a number of vessels the electrical response to stimulation of perivascular sympathetic nerves is biphasic; an initial fast, transient depolarization or EJP of the vascular smooth muscle is followed by a slow, prolonged depolarization. The EJP and slow depolarization are mimicked by the effects of ATP and NA, respectively. Considerable variation exists in the proportions of NA and ATP utilized by sympathetic nerves. For example, in guinea pig submucosal arterioles both vasoconstriction and EJPs, evoked in response to electrical stimulation of sympathetic nerves, are mediated exclusively by ATP, with NA assuming the role of a neuromodulator, by acting through prejunctional α2-adrenoceptors to depress transmitter release. At the other extreme, in rat renal arteries the purinergic component is relatively small. It has also been noted that the purinergic component is optimal with short bursts of low frequency stimulation, whereas longer durations of higher frequency favor adrenergic transmission. NPY has been found to be present in LGV in most sympathetic nerves. The release of NPY, as well as NA and ATP, in response to electrical stimulation of sympathetic nerve terminals is prevented by guanethidine. The major role of

FIGURE 5.1 Schematic of sympathetic cotransmission. ATP and NA released from small granular vesicles (SGV) act on P2X and α1 receptors on smooth muscle, respectively. ATP acting on inotropic P2X receptors evokes excitatory junction potentials (EJPs), increase in intracellular calcium ([Ca2]i) and fast contraction; while occupation of metabotropic α1 adrenoceptors leads to production of inositol triphosphate (InsP3), increase in [Ca2]i and slow contraction. Neuropeptide Y (NPY) stored in large granular vesicles (LGV) acts on release both as a prejunctional inhibitory modulator of release of ATP and NA and as a postjunctional modulatory potentiator of the actions of ATP and NA. Nucleotidases are released from nerve varicosities, and are also present as ectonucleotidases. (Reproduced from [14] with permission from Elsevier.)

NPY in the vasculature, and in the vas deferens, appears to be that of a pre- and/or postjunctional modulator of sympathetic transmission, since it has little direct postjunctional action or causes contraction only at high concentrations (see Fig. 5.1). Direct vasoconstrictor actions of NPY have, however, been demonstrated in some vessels. At the prejunctional level, NPY has potent inhibitory effects, reducing the release of NA and ATP from sympathetic nerves. Postjunctionally, NPY generally acts to enhance the actions of sympathetic nerve stimulation, NA and ATP. Although 5-hydroxytryptamine (5-HT) immunofluorescent nerves have been localized in a number of vessels, for the most part 5-HT is not synthesized and stored in separate nerves, but is taken up, stored in, and released as a “false transmitter” from sympathetic nerves. Enkephalins have been shown to coexist with NA in cell bodies and fibers of some postganglionic sympathetic neurons. The functional significance of sympathetic coexistence of opioids is likely to be related to their prejunctional inhibitory effects on sympathetic transmission.

PARASYMPATHETIC NERVES The classical evidence for cotransmission of ACh and vasoactive intestinal polypeptide (VIP) in certain

I. INTRODUCTION

ENTERIC ANd CARdIAC NERvES

postganglionic parasympathetic neurons comes from pharmacological studies performed by Lundberg in 1981 on cat salivary glands (see [9]). ACh and VIP are released from the same parasympathetic nerve terminals in response to transmural nerve stimulation. During low frequency stimulation ACh is released to cause an increase in salivary secretion from acinar cells and also to elicit some minor dilatation of blood vessels in the gland. VIP is preferentially released at high frequencies to cause marked vasodilatation of blood vessels and, while it has no direct effect on acinar cells, it acts as a neuromodulator to enhance both the postjunctional effect of ACh on acinar cell secretion and the release of ACh from nerve varicosities via prejunctional receptors. Vasodilator nerves to the uterine arteries in the guinea pig contain immunoreactivity to VIP, which coexists with dynorphin, NPY and somatostatin. NPY-like immunoreactivity has been reported in some of the choline acetyltransferase-/VIP-containing neurons of the parasympathetic ciliary, sphenopalatine, otic and pterygopalatine ganglia with targets including the iris and cerebral vessels. Autonomic control of penile erection, involving relaxation of the smooth muscle of the corpus cavernosum as well as dilatation of other penile vascular beds, has traditionally been attributed to the vasodilator effects of ACh and VIP released from parasympathetic nerves. More recently, nitric oxide (NO) released from nerves arising from nerves in the pelvic ganglia, have been claimed to play a role in smooth muscle relaxation leading to penile erection. NO synthase (NOS)-containing fibers, shown by lesion studies to arise from parasympathetic cell bodies in the sphenopalatine ganglia, have been localized in the adventitia of cerebral arteries and many of these also contain VIP. A functional role for perivascular neuronal NO in cerebral arteries has been identified in studies showing that stimulation of adventitial nerve fibers causes vascular relaxation, which is attenuated by inhibitors of NOS. Parasympathetic nerves supplying the urinary bladder utilize ACh and ATP as cotransmitters, in variable proportions in different species, and by analogy with sympathetic nerves, ATP again acts through P2X ionotropic receptors to produce EJPs and fast contraction, while the slow component of the response is mediated by a metabotropic receptor, in this case muscarinic (see [10]). There is also evidence for parasympathetic, purinergic cotransmission to resistance vessels in the heart and airways.

SENSORY-MOTOR NERVES The neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP) are the principal transmitters of primary afferent nerves and have been shown to coexist in the same terminals. Furthermore, with the use of colloidal gold particles of different sizes, they have been shown to coexist in the same large granular vesicles. The motor (efferent) function of sensory nerves has been

29

demonstrated in rat mesenteric arteries where evidence exists for a role for CGRP as the mediator of vasodilatation following release from sensory motor nerves. In contrast, SP is not co-released with CGRP by electrical stimulation and SP has little or no vasodilator action on rat mesenteric arteries. While it is possible that SP released from nerves supplying the microvasculature could produce vasodilatation via SP receptors on endothelial cells, it is most unlikely to reach the endothelium without degradation in larger blood vessels. It may be that the role of the coexisting SP is either trophic or sensory (and not motor). Other peptide and non-peptide substances including neurokinin A, somatostatin, VIP and ATP have been described in capsaicin-sensitive sensory neurons. Unmyelinated sensory neurons containing cholecystokinin (CCK)/CGRP/dynorphin/SP have been shown to project to cutaneous arterioles in guinea pig skin. Neurons from the same ganglia which contain CCK/CGRP/SP, innovate arterioles of skeletal muscle, CGRP/dynorphin/ SP nerve fibers mostly supply the pelvic viscera, and CGRP/SP fibers run mainly to the heart, large arteries, and veins. There is also evidence for a sensory role for ATP and it has been proposed that ATP may coexist in sensory nerve terminals with SP and CGRP (Fig. 5.2).

ENTERIC AND CARDIAC NERVES Intrinsic neurons exist in most of the major organs of the body. Many of these are part of the parasympathetic nervous system, but certainly in the gut and perhaps also in the heart and airways, some of these intrinsic neurons are derived from neural crest tissue, which differs from that which forms the sympathetic and parasympathetic systems and appear to participate in local reflex actions independent of the CNS. The enteric nervous system contains several hundred million neurons located in the myenteric plexus between muscle coats and the submucous plexus. The chemical coding of these nerves has been examined in detail. A subpopulation of these intramural enteric nerves provides NANC inhibitory innervation of the gastrointestinal smooth muscle. It seems likely that three major cotransmitters are released from these nerves. ATP produces fast inhibitory junction potentials, NO also produces inhibitory potentials but with a slower time course, while VIP produces slow tonic relaxations. The proportions of these three transmitters varies considerably in different regions of the gut and in different species; for example, in some sphincters the NANC inhibitory nerves utilize largely VIP, in others largely NO, while in non-sphincteric regions of the intestine ATP is more prominent. In recent papers, evidence has suggested that ACh and ATP are fast excitatory cotransmitters to myenteric neurons and that there may be colocalization of ACh, ATP and 5-HT in enteric Dogiel Type I/S neurons. Detailed studies have allowed a very complete mapping of the complex neuronal markers and

I. INTRODUCTION

30

5. CoTRANSmISSIoN

FIGURE 5.2 Schematic hypotheses of the neurogenic basis of reflex vasodilatation of skin vessels. One suggestion is that substance P (a putative transmitter in sensory nerve endings in the spinal cord) is released from sensory nerve collaterals in the skin; another that ATP is the transmitter released from these collaterals. Both substances are powerful vasodilators and can release histamine from mast cells. ATP is also a potent inducer of prostaglandin synthesis. Another alternative, although less likely, is that a peripherally placed purinergic neuron is interposed between the sensory nerve collateral and the effector system. (This hypothesis incorporates early histologic reports of neurons in the skin and provides an alternative or additional explanation for the initiation of the vasodilator reflex by nicotine and its block by hexamethonium.) (Reproduced from [15], with permission.)

projections of enteric neurons. Several peptidergic substances, including NPY, VIP, enkephalin, somatostatin, peptide histidine isoleucine, galanin, SP and CGRP have been identified in enteric neurons, often coexisting (up to five peptides in the same neuron) with the neurotransmitters NA, ACh, 5-HT, NO and ATP (see Table 5.2). Studies of intrinsic cardiac neurons in culture have shown that subpopulations of intrinsic nerves in the atrial and intra-atrial septum contain and/or release cotransmitters, including ATP, NO, NPY, ACh and 5-HT. Many of these nerves project to the coronary microvasculature and produce potent vasomotor actions. NO together with ATP have been shown to be the mediators of NANC vasodilatation of the rabbit portal vein.

PHYSIOLOGICAL SIGNIFICANCE OF COTRANSMISSION In general, cotransmission offers greater physiological events by peripheral mechanisms rather than the all-ornone control by messages coming from the CNS, that has been the dominant view for many years (see [11]).

Cotransmitters with Different Firing Patterns Although single presynaptic action potentials release small molecule neurotransmitters, trains of impulses are

needed to release neuropeptides. For sympathetic and parasympathetic cotransmission, release of ATP is favored at low frequency stimulation, whereas NA and ACh are released at higher frequencies. There are instances where more than one fast cotransmitter is released (e.g. glutamate and ATP) together with one or more peptides.

Different Cotransmitters act on Different Postjunctional Cells Neurons using multiple transmitters may project to two or more targets. For example, ACh released at low frequency stimulation from parasympathetic nerves supplying salivary glands acts on acinar cells to produce secretion and a minor dilatation of vessels, whereas, at higher frequency stimulation, its cotransmitter VIP causes powerful vasodilatation of vessels in the glands and postjunctional enhancement of ACh-induced saliva secretion.

Neuromodulation A cotransmitter can feed back on presynaptic receptors that increase or decrease its own release or that of its cotransmitter(s). For example, ATP released as a cotransmitter with glutamate from primary afferent fibers in lamina II of the spinal cord can act on prejunctional P2X3 receptors to facilitate the release of its cotransmitter, glutamate,

I. INTRODUCTION

31

PHySIologICAl SIgNIfICANCE of CoTRANSmISSIoN

TABLE 5.2 Types of Neurons in the Enteric Nervous System Proportion

Chemical Coding

Function/Comments

Excitatory circular muscle motor neurons

12%

Short: ChAT/TK/ENK/GABA Long: ChAT/TK/ENK/NFP

To all regions, primary transmitter ACh, cotransmitter TK

Inhibitory circular muscle motor neurons

16%

Short: NOS/VIP/PACAP/ENK/ NPY/GABA Long: NOS/VIP/PACAP/ Dynorphin/BN/NFP

Several cotransmitters with varying prominence: NO, ATP, VIP, PACAP

Excitatory longitudinal muscle motor neurons

25%

ChAT/Calretinin/TK

Primary transmitter ACh, cotransmitter TK

Inhibitory longitudinal muscle motor neurons

~2%

NOS/VIP/GABA

Several cotransmitters with varying prominence: NO, ATP, VIP, PACAP

Ascending interneurons (local reflex)

5%

ChAT/Calretinin/TK

Primary transmitter ACh

Descending interneurons (local reflex)

5%

ChAT/NOS/VIP  BN  NPY

Primary transmitter ACh, ATP may be a cotransmitter

Descending interneurons (secretomotor reflex)

2%

ChAT/5-HT

Primary transmitter ACh, 5-HT (at NK3 receptors)

Descending interneurons (migrating myoelectric complex)

4%

ChAT/SOM

Primary transmitter ACh

Myenteric intrinsic primary afferent (primary sensory) neurons

26%

ChAT/Calbindin/TK/NK3 receptor

Primary transmitter TK

Intestinofugal neurons

1%

ChAT/BN/VIP/CCK/ENK

Primary transmitter ACh

Motor neurons to gut endocrine cells

N/A

N/A

For example, myenteric neurons innervating gastrin cells. Neurons of this type may be in submucosal ganglia

Non-cholinergic secretomotor/vasodilator neurons

45%

VIP/GAL

Primary transmitter VIP. A small proportion of these have cell bodies in myenteric ganglia

Cholinergic secretomotor/vasodilator neurons

15%

ChAT/Calretinin/Dynorphin

Primary transmitter ACh

MYENTERIC NEURONS

SUBMUCOSAL NEURONS

*Also listed are three types of motor neuron that are found in other parts of the tubular digestive tract, marked by asterisks. Reproduced from [13], with permission from Elsevier. ACh, acetylcholine; ATP, adenosine 5-triphosphate; BN, bombesin; CCK, cholecystokinin; CGRP, calcitonin gene-related peptide; ChAT, choline acetyltransferase; DA, dopamine; ENK, enkephalin; GABA, γ-amino butyric acid; GAL, galanin; 5-HT, 5-hydroxytryptamine; NA, noradrenaline; NFP, neurofilament protein; NK, neurokinin; NOS, nitric oxide synthase; NPY, neuropeptide Y; PACAP, pituitary adenylyl cyclase activating peptide; SOM, somatostatin; SP, substance P; TK, tachykinin; VIP, vasoactive intestinal peptide.

whereas adenosine resulting from ectoenzymatic breakdown of ATP acts on presynaptic P1 receptors to inhibit glutamate release. Both NA and ATP can prejunctionally modulate sympathetic transmission, NA via prejunctional α2-adrenoceptors and ATP via P1 receptors following breakdown to adenosine. Modulation of cotransmitter release and presynaptic action by other agents also occurs and might provide a new level of synaptic flexibility, in which individual neurons utilize more than one transmitter but retain independent control over their synaptic activity.

Synergism There are an increasing number of reports of the synergistic actions of cotransmitters. ATP and NA released from sympathetic nerves have synergistic actions on

smooth muscle of vas deferens and blood vessels, and ATP released with ACh from motoneurons facilitates the nicotinic actions of ACh at the skeletal neuromuscular junction. Co-operativity between receptors for ATP and N-methyl-D-aspartate (NMDA) in induction of long-term potentiation in hippocampal CA1 neurons has also been demonstrated. Thyrotropin-releasing hormone and 5-HT have been reported to have synergistic actions in spinal cord neurons. The mechanisms underlying cotransmitter synergism are not well understood. However, it has been suggested that postjunctional synergism between the responses of vas deferens to NA and ATP is caused by the ability of NA to potentiate the contractile responses to ATP by sensitizing smooth muscle cells to Ca2 via an inhibitory action on myosin light chain phosphatase, an action mediated by protein kinase C.

I. INTRODUCTION

32

5. CoTRANSmISSIoN

Negative Cross-Talk Co-application of nicotinic and P2X receptor agonists produces less than the additive responses predicted by independent receptor activation. Inhibitory interactions between 5-HT3 and P2X receptors have been described in submucosal and myenteric neurons.

Trophic Factors Some co-stored and co-released substances can act as long-term (trophic) factors, as well as neurotransmitters. For example, ATP can act on P2 receptors, or P1 (adenosine) receptors after ectoenzymatic breakdown, to promote vascular cell proliferation, motility, differentiation or death. NPY released from sympathetic nerves has cardiovascular trophic effects in end-stage renal disease. There is growing evidence that neurotrophic factors might be synthesized, stored and released from nerve terminals together with fast neurotransmitters.

Excitatory and Inhibitory Cotransmitters Although cotransmitters generally have similar actions on postjunctional cells, there are some examples of cotransmitters having opposite actions. For example, in the mammalian uterus, one or other cotransmitter dominates depending on the hormonal and/or tonic status of the postjunctional muscle cells. Brain-derived neurotrophic factor (BDNF) increases the release of ACh and reduces NA release from sympathetic nerves to cause a rapid shift from excitatory to inhibitory transmission.

False Cotransmitters For example, it has been known for some time that sympathetic nerves take up 5-HT, which can then be released as a “false transmitter”, rather than a genuine “cotransmitter”. A “false transmitter” is a substance actively taken up and subsequently released by a neuron that does not synthesize it.

Coexisting Peptides Acting as Neuromodulators In general, most neuropeptides act as neuromodulators rather than neurotransmitters. For example, NPY released from sympathetic nerves acts as a pre- and postjunctional modulator of ATP and NA release and postjunctional actions.

COTRANSMITTER PLASTICITY Cotransmitter plasticity occurs during development and ageing, following trauma or surgery and after chronic exposure to drugs, as well as in disease (see [12]). There were some outstanding early studies of the factors

influencing cotransmitter expression in sympathetic nerves. Evidence was presented that cholinergic differentiation in sympathetic neurons is promoted by neurotrophic factors from three different protein families (glial cell line-derived neurotrophic factor, neurotrophin 3 and ciliary neurotrophic factor), whereas noradrenergic differentiation is promoted by nerve growth factor. In another study, BDNF was claimed to switch sympathetic neurotransmission to the heart from an adrenergic excitation to cholinergic inhibition; it was also shown that the action of BDNF was mediated by the P75 neurotrophic receptor. Histamine, galanin and γ-aminobutyric acid (GABA) acting as cotransmitters in neurons of the tuberomammillary nucleus (hypothalamus) have independent control mechanisms. CGRP-like immunoreactivity was found earlier than SP-like immunoreactivity in cerebrovascular nerves during development, and increased in old age, while the density of SP-like immunoreactive fibers did not change. NA and NPY also show different expression in cerebrovascular nerves during development. Changes in chemical coding of myenteric neurons in ulcerative colitis have been reported, with a shift from cholinergic to more SP-mediated cotransmission. In interstitial cystitis and outflow obstruction, there is a substantial increase in ATP as the cotransmitter with ACh in parasympathetic nerves supplying the human bladder, and there is increase in ATP as a cotransmitter with NA in sympathetic nerves in spontaneous hypertensive rats (see [4]).

CONCLUDING COMMENTS It has been particularly difficult to establish cotransmitter roles for the many peptides found in nerves, partly because specific receptors and physiological roles have not been established for some of these and partly because of the lack of selective antagonists. In some enteric neurons, up to five neuropeptides have been identified. However, it is important to distinguish between neuromodulator, neurotransmitter and neurotrophic roles for released peptides or indeed as yet unrecognized roles. It is becoming clear that ATP is a primitive signalling molecule that has been retained as a cotransmitter in every nerve type in both the peripheral and central nervous systems, although the relative role of ATP varies considerably in different species and pathophysiological conditions. ATP appears to become a more prominent cotransmitter in stress and inflammatory conditions. Most nerves contain and release ATP as a fast cotransmitter together with classical transmitters such as ACh, NA, glutamate, GABA and one or more peptides. Now that cotransmission is recognized as a universal mechanism, it is recommended that the terms “adrenergic”, “cholinergic”, “peptidergic”, “purinergic”, “aminergic” and “nitrergic” should not be used when nerves are described, although adrenergic, cholinergic, peptidergic, purinergic, aminergic or nitrergic transmission is still meaningful.

I. INTRODUCTION

CoNCludINg CommENTS

References [1] Burnstock G. Do some nerve cells release more than one transmitter? Neuroscience 1976;1:239–48. [2] Kupfermann I. Functional studies of cotransmission. Physiol Rev 1991;71:683–732. [3] Merighi A. Costorage and coexistence of neuropeptides in the mammalian CNS. Prog Neurobiol 2002;66:161–90. [4] Burnstock G. Physiology and pathophysiology of purinergic neurotransmission. Physiol Rev 2007;87:659–797. [5] Furness JB, Morris JL, Gibbins JL, Costa M. Chemical coding of neurons and plurichemical transmission. Ann Rev Pharmacol Toxicol 1989;29:289–306. [6] Su C, Bevan JA, Burnstock G. [3H]adenosine triphosphate: release during stimulation of enteric nerves. Science 1971;173:337–9. [7] Sneddon P, Burnstock G. Inhibition of excitatory junction potentials in guinea-pig vas deferens by α,β-methylene-ATP: further evidence for ATP and noradrenaline as cotransmitters. Eur J Pharmacol 1984;100:85–90. [8] Burnstock G. Noradrenaline and ATP as cotransmitters in sympathetic nerves. Neurochem Int 1990;17:357–68.

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[9] Lundberg JM. Pharmacology of cotransmission in the autonomic nervous system: integrative aspects on amines, neuropeptides, adenosine triphosphate, amino acids and nitric oxide. Pharmacol Rev 1996;48:113–78. [10] Burnstock G. Cotransmission with particular emphasis on the involvement of ATP. In: Fuxe K, Hökfelt T, Olson L, Ottoson D, Dahlström A, Björklund A, editors. Molecular Mechanisms of Neuronal Communication. A Tribute to Nils-Ake Hillarp. Oxford: Pergamon Press; 1996. p. 67–87. [11] Burnstock G. Cotransmission. Curr Opin Pharmacol 2004;4:47–52. [12] Burnstock G. Co-transmission. The fifth Heymans memorial lecture – Ghent, February 17, 1990. Arch Int Pharmacodyn Ther 1990;304:7–33. [13] Furness JB. Types of neurons in the enteric nervous system. J Auton Nerv Syst 2000;81:87–96. [14] Burnstock G, Verkhratsky A. Vas deferens - a model used to establish sympathetic cotransmission. Trends Pharmacol Sci 2010;31:131–9. [15] Burnstock G. Autonomic neuroeffector junctions - reflex vasodilatation of the skin. J Invest Dermatol 1977;69:47–57.

I. INTRODUCTION

C H A P T E R

6 Noradrenergic Neurotransmission David S. Goldstein The main chemical transmitter of the sympathetic nervous system mediating regulation of the circulation is the catecholamine norepinephrine (NE). Sympathetic stimulation releases NE, and binding of NE to adrenoceptors on cardiovascular smooth muscle cells causes the cells to contract. Sympathoneural NE therefore satisfies the main criteria defining a neurotransmitter – a chemical released from nerve terminals by electrical action potentials that interacts with specific receptors on nearby structures to produce specific physiological responses. Unlike most other neurotransmitters, NE can be measured in human plasma. This capability, coupled with the ability to assay simultaneously levels of NE and other compounds related to synthesis and disposition of NE, provides the bases for clinical neurochemical biomarkers of sympathetic innervation and function and for metabolomic approaches to diagnose several disorders. Such neurochemical measures often are more sensitive than physiological measures in clinical laboratory assessments of patients with autonomic dysfunction.

fibers in the ansae subclaviae pass along the dorsal surface of the pulmonary artery into the plexus that supplies the left main coronary artery. In primates, cardiac sympathetic nerves originate about equally from the superior, middle, and inferior cervical (stellate) ganglia. The right sympathetic chain generally projects to the anterior left ventricle and the left to the posterior left ventricle. Sympathetic innervation of the sinus and atrioventricular nodes also has a degree of sidedness, the right sympathetics projecting more to the sinus node and the left to the atrioventricular node. This sidedness means that left stellate stimulation produces relatively little sinus tachycardia. Epicardial sympathetic nerves provide the main source of noradrenergic terminals in the myocardium. Sympathetic nerves travel with the coronary arteries in the epicardium before penetrating into the myocardium, whereas vagal nerves penetrate the myocardium after crossing the atrioventricular groove and then course distally in the subendocardium. Post-ganglionic noradrenergic fibers reach all parts of the heart. The sinus and atrioventricular nodes and the atria receive the densest innervation, the ventricles less dense innervation, and the Purkinje fibers the least. Sympathetic and vagal afferents follow similar intracardiac routes to those of the efferents. The heart contains relatively high NE concentrations compared to other organs. In humans, the left ventricular myocardial NE concentration is roughly 10 pmoles per mg wet weight, with substantial inter-individual variability, corresponding to about 10 times the plasma concentration. The coronary arteries possess sympathetic noradrenergic innervation, but assessing the physiological role of this innervation has proven difficult, because several interacting factors complicate neural control of the coronary vasculature. Alterations in myocardial metabolism and systemic hemodynamics change coronary blood flow, coronary vasomotion in response to sympathetic stimulation depends on the functional integrity of the endothelium, and coronary arteries appear to possess less dense innervation than do other arteries. The body’s myriad arterioles largely determine total resistance to blood flow in the body and therefore contribute importantly to blood pressure. Sympathetic nerves enmesh blood vessels in lattice-like networks in the adventitial outer surface that extend inward to the adventitial-medial border. The concentration of sympathetic

CATECHOLAMINES, AND CATECHOLS The endogenous catecholamines are NE, dopamine (DA), and epinephrine (EPI, synonymous with adrenaline). Catecholamines are catechols, which are chemicals that have adjacent hydroxyl groups on a benzene ring. Catechol itself does not exist in the human body, but compounds that contain catechol as part of their molecular structure are called catechols. Human plasma normally contains six catechols – the catecholamines, L-DOPA (the amino acid precursor of the catecholamines), and two metabolites of the catecholamines, dihydroxyphenylglycol (DHPG, the main neuronal metabolite of NE), and dihydroxyphenylacetic acid (DOPAC, the main neuronal metabolite of DA). As will be seen, particular patterns of levels of catechols characterize different disorders of catecholamine systems.

NORADRENERGIC INNERVATION OF THE CARDIOVASCULAR SYSTEM Sympathetic nerves to the heart travel via the ansae subclaviae, branches of the left and right stellate ganglia. The

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00006-8

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nerves increases as the arterial caliber decreases, so that small arteries and arterioles, the smallest nutrient vessels possessing smooth muscle cells, have the most intense innervation. The architectural association between sympathetic nerves and the vessels that determine peripheral resistance has enticed cardiovascular researchers particularly in the area of autonomic regulation for many years. Sympathetic vascular innervation varies widely among vascular beds, with dense innervation of resistance vessels in the gut, kidney, skeletal muscle, and skin. Sympathetic stimulation in these beds produces profound vasoconstriction, whereas stimulation in the coronary, cerebral, and bronchial beds elicits weaker constrictor responses, consistent teleologically with the “goal” of preserving blood flow to vital organs in emergencies.

NOREPINEPHRINE: THE MAIN SYMPATHETIC CARDIOVASCULAR NEUROTRANSMITTER Norepinephrine Synthesis Catecholamine biosynthesis begins with uptake of the amino acid tyrosine (TYR) into the cytoplasm of sympathetic neurons, adrenomedullary cells, possibly para-aortic enterochromaffin cells, and specific centers in the brain. Tyrosine hydroxylase (TH) catalyzes the conversion of TYR to DOPA. This is the enzymatic rate-limiting step in catecholamine synthesis. The enzyme is stereospecific, and tetrahydrobiopterin, ionized iron, and molecular oxygen regulate TH activity. Dihydropteridine reductase (DHPR) catalyzes the reduction of dihydropterin, produced during the hydroxylation of TYR. Since the reduced pteridine, tetrahydrobiopterin, is a key co-factor for TH, DHPR deficiency decreases the amount of TYR hydroxylation for a given amount of TH enzyme. Both phenylalanine hydroxylase and TH require tetrahydrobiopterin as a co-factor. DHPR deficiency therefore also inhibits phenylalanine metabolism and presents clinically as an atypical form of phenylketonuria. Multiple and complex mechanisms contribute to TH activation. Short-term mechanisms include feedback inhibition by DOPA and catecholamines and phosphorylation of the enzyme, the latter depending on membrane depolarization, contractile elements, and receptors. Long-term mechanisms include changes in TH synthesis. During stress-induced sympathetic stimulation, acceleration of catecholamine synthesis in sympathetic nerves helps to maintain tissue stores of NE. Even with diminished stores after prolonged sympathoneural activation, increased nerve traffic can maintain extracellular fluid levels of the transmitter. L-aromatic-amino-acid decarboxylase (LAAAD, also called DOPA decarboxylase) catalyzes conversion of DOPA to DA. Many types of tissues contain this enzyme – especially the kidneys, gut, liver, and brain. LAAAD activity depends on pyridoxal phosphate (vitamin B6).

Although LAAAD metabolizes most of the DOPA formed in catecholamine-synthesizing tissues, some of the DOPA enters the circulation unchanged. This provides the basis for using plasma DOPA levels to examine catecholamine synthesis. LAAAD inhibitors include carbidopa and benserazide. Both are catechols that do not readily penetrate the blood–brain barrier. By inhibiting conversion of DOPA to DA in the periphery, carbidopa and benserazide enhance the efficacy of L-DOPA and reduce its side effects in the treatment for Parkinson disease. LAAAD blockade increases DOPA levels and decreases levels of DOPAC, the deaminated metabolite of DA. Rates of increase in extracellular fluid DOPA levels and of decrease in DOPAC levels after acute LAAAD inhibition provide in vivo indices of TH activity. Dopamine-β-hydroxylase (DBH) catalyzes conversion of DA to NE. DBH is localized to vesicles in cells that synthesize catecholamines, such as noradrenergic neurons and chromaffin cells. Treatment with reserpine, which blocks translocation of amines from the axonal cytoplasm into vesicles, prevents conversion of DA to NE in sympathetic nerves. DBH contains – and its activity depends on – copper. Because of this dependence, children with Menkes disease, a rare, X-linked recessive inherited disorder of copper metabolism, have neurochemical evidence of concurrently increased catecholamine biosynthesis and decreased conversion of DA to NE, with high plasma ratios of DOPA:DHPG. Patients with congenital absence of DBH have virtually undetectable levels of both NE and DHPG and high levels of DA and DOPAC. DBH activity also requires ascorbic acid (vitamin C), which provides electrons for the hydroxylation. Phenylethanolamine-N-methyltransferase (PNMT) catalyzes the conversion of NE to EPI in the cytoplasm of chromaffin cells and some brainstem neurons.

Norepinephrine Storage Vesicles generated near the Golgi apparatus in cell bodies travel by axonal transport to the nerve terminals. Noradrenergic vesicles may also form by endocytosis within the axons. Since reserpine eliminates the electrondense cores of the small but not the large vesicles, the cores of the small vesicles may represent NE, whereas the electron-dense cores of the large vesicles may represent additional components. Cores of both types of vesicle contain adenosine triphosphate (ATP). The vesicles also contain at least three types of polypeptides: chromogranin A, an acidic glycoprotein; enkephalins; and neuropeptide Y (NPY). Extracellular fluid levels of each of these compounds have been considered as indices of exocytosis. Vesicles in sympathetic nerves actively remove and trap axoplasmic amines via the type 2 vesicular monoamine transporter (VMAT). Adrenomedullary chromaffin cells express both types of VMAT. Vesicular uptake favors l- over d-NE, Mg and ATP accelerate the uptake, and reserpine effectively and irreversibly blocks it. VMAT

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proteins resemble the neuronal uptake carriers of the cell membrane. Neurotransmitter specificity appears to depend on different transporters in the cell membrane rather than on different vesicular transporters. Tissue NE stores are maintained by a balance of synthesis and loss. Under resting conditions, the main determinant of loss of NE from the tissue is net leakage of NE from the vesicles into the axoplasm, with subsequent enzymatic breakdown of the axoplasmic NE. This loss is balanced by ongoing synthesis of DA in the neuronal cytosol, vesicular uptake of cytosolic DA, and conversion of DA to NE catalyzed by DBH. Sympathetic neuroimaging using radio-iodinated metaiodobenzylguanidine, positron-emitting analogs of sympathomimetic amines, and 6-[18F]fluorodopamine depends on uptake of the imaging agents into the axoplasm via the cell membrane norepinephrine transporter (NET) and subsequent translocation into vesicles via the VMAT. Visualization of sympathetic innervation in organs such as the heart, therefore, results in radiolabeling of the vesicles in noradrenergic nerves.

Norepinephrine Release Adrenomedullary chromaffin cells, much easier to study than sympathetic nerves, have provided the most commonly used model for studying mechanisms of catecholamine release. Agonist occupation of nicotinic acetylcholine receptors releases catecholamines from the cells. Since nicotinic receptors mediate ganglionic neurotransmission, researchers have presumed that findings in adrenomedullary cells apply to post-ganglionic sympathetic noradrenergic neurons. Low plasma levels of NE in patients with autoimmune autonomic ganglionopathy from circulating antibodies to the neuronal nicotinic receptor support this view. According to the exocytotic theory of NE release, acetylcholine depolarizes terminal membranes by increasing membrane permeability to sodium. The increased intracellular sodium levels directly or indirectly enhance transmembrane influx of calcium, via voltage-gated calcium channels. The increased cytoplasmic calcium concentration evokes a cascade of biomechanical events resulting in fusion of the vesicular and axoplasmic membranes. Vesicle poration causes the interior of the vesicle to exchange with the extracellular compartment, and the soluble contents of the vesicles diffuse into the extracellular space. As predicted from this model, manipulations besides application of acetylcholine that depolarize the cell, such as electrical stimulation or increased K concentrations in the extracellular fluid, also activate voltage-gated calcium channels and trigger exocytosis. During cellular activation, simultaneous, stoichiometric release of soluble vesicular contents – ATP, enkephalins, chromogranins, and DBH – without similar release of cytoplasmic macromolecules, has provided biochemical support for the exocytosis theory.

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At least two storage pools of NE seem to exist in sympathetic nerve terminals – a small, readily releasable pool of newly-synthesized NE and a large reserve pool in longterm storage. Because of conversion of DA to NE within vesicles, plasma DA responses during sympathetic stimulation may reflect release from the readily releasable pool. Sympathetic nerve endings can also release NE by calcium-independent, non-exocytotic mechanisms. One such mechanism is reverse transport through the neuronal uptake carrier. The indirectly acting sympathomimetic amine, tyramine, releases NE non-exocytotically, since tyramine releases NE independently of calcium and does not release DBH. Myocardial ischemic hypoxia also evokes calcium-independent release of NE. As noted above, sympathetic stimulation releases other compounds besides NE. Some of these compounds may function as co-transmitters. ATP, adenosine, NPY, acetylcholine, DA, and EPI have received the most attention. Pharmacological stimulation of a large variety of receptors on noradrenergic terminals affects the amount of NE released during cellular activation. Compounds inhibiting NE release include acetylcholine, gamma-aminobutyric acid (GABA), prostaglandins of the E series, opioids, adenosine, and NE itself. Compounds enhancing NE release include angiotensin II, acetylcholine (at nicotinic receptors), ACTH, GABA (at GABAA receptors), and EPI (via stimulation of pre-synaptic β2-adrenoceptors). In general, whether at physiological concentrations these compounds exert modulatory effects on endogenous NE release remains unproven, especially in humans. An exception, however, is inhibitory presynaptic modulation by NE itself, via autoreceptors on sympathetic nerves. This modulatory action appears to vary with the vascular bed under study, being prominent in skeletal muscle beds such as the forearm, relatively weak in the kidneys, and virtually absent in the adrenals. In addition to local feedback control of NE release, reflexive “long-distance” feedback pathways via high- and low-pressure baroreceptors elicit reflexive changes in sympathoneural impulse activity. Alterations in receptor numbers or of intracellular biomechanical events after receptor activation also affect responses to agonists. These factors may, therefore, regulate NE release by trans-synaptic local and reflexive long-distance mechanisms. NE released from sympathetic nerve terminals acts mainly locally, with only a small proportion of released NE reaching the bloodstream. One must, therefore, keep in mind the indirect and distant relationship between plasma NE levels and sympathetic nerve activity in interpreting plasma NE levels in response to stressors, pathophysiologic situations, and drugs.

Norepinephrine Disposition NE is inactivated mainly by uptake into cells, with subsequent intracellular metabolism or storage (Fig. 6.1). Reuptake into nerve terminals – Uptake-1 – via the cell

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VMA

Exo

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3-OMeTyramine

AR

HVA

DOPET

DOPEGAL

ALDH

DOPAC

AR

DHPG

COMT

NMN

ALDH

MHPG

DHMA COMT

HVA DOPAC

MHPG

DHPG

FIGURE 6.1 Diagram of steps in norepinephrine biosynthesis, release, cellular uptake, and metabolism. Abbreviations: ADH, alcohol dehydrogenase; ALDH, aldehyde dehydrogenase; AR, aldose/aldehyde reductase; COMT, catechol-O-methyltransferase; DA, dopamine; DBH, dopamine-β-hydroxylase; DHMA, dihydroxymandelic acid; DHPG, dihydroxyphenylglycol; DOPA, dihydroxyphenylalanine; DOPAC, dihydroxyphenylacetic acid; DOPEGAL, dihydroxyphenylglycolaldehdye; Exo, exocytosis; HVA, homovanillic acid; LAAAD, L-aromatic-amino-acid decarboxylase; MAO, monoamine oxidase; MHPG, methoxyhydroxyphenylglycol; 3-MT, 3-methoxytyrosine; 3-OMdTyramine, 3-methoxytyramine; NEc, cytoplasmic norepinephrine; NEv, vesicular norepinephrine; NET, cell membrane norepinephrine transporter; NMN, normetanephrine; TH, tyrosine hydroxylase; TYR, tyrosine; VMA, vanillylmandelic acid; VMAT, vesicular monoamine transporter.

membrane NET is the predominant means of terminating the actions of released NE. Uptake-1 is energy-requiring and can transport catecholamines against large concentration gradients. The only common structural feature of all known substrates for Uptake-1 is an aromatic amine, with the ionizable nitrogen moiety not incorporated in the aromatic system. Uptake-1 does not require a catechol nucleus. Alkylation of the primary amino group decreases the effectiveness of the transport, explaining why sympathetic nerves take up NE more efficiently than they do EPI and why they do not take up isoproterenol, an extensively alkylated catecholamine, at all. Methylation of the phenolic hydroxyl groups also markedly decreases susceptibility to Uptake-1, and so sympathetic nerves do not take up O-methylated catecholamine metabolites such as normetanephrine. Many drugs or in vitro conditions inhibit Uptake-1, including cocaine, tricyclics such as desipramine, NET blockers such as reboxetine, low extracellular Na concentrations, and Li(Fig. 6.2). Neuronal uptake absolutely requires intracellular K and extracellular Na. Transport does not directly require ATP; however, maintaining ionic gradients across cell membranes depends on ATP, and the carrier uses the energy expended in maintaining the transmembrane Na gradient to co-transport amines with Na.

There are distinct cell membrane transporters for NE and DA. The human NET protein includes 12–13 hydrophobic and therefore membrane-spanning domains, a structure that differs substantially from that of adrenoceptors and other receptors coupled with G-proteins but resembles that of transporters for DA, GABA, serotonin, and vesicular monoamines. Non-neuronal cells remove NE actively by a process called Uptake-2, which is characterized by the ability to transport isoproterenol, susceptibility to blockade by O-methylated catecholamines, corticosteroids, and β-haloalkylamines, and absence of susceptibility to blockade by cocaine and tricyclics. In contrast with Uptake-1, Uptake-2 functions independently of extracellular Na. The Uptake-2 carrier has little if any stereoselectivity and has low affinity and specificity for catecholamines. For instance, extraneuronal cells remove imidazolines such as clonidine by Uptake-2. Whereas reverse transport via the Uptake-1 carrier requires special experimental conditions, one can readily demonstrate reverse transport via the Uptake-2 carrier. Because of reverse transport, during infusion of a catecholamine at a high rate, the catecholamine can accumulate in extraneuronal cells, with re-entry of the catecholamine into the extracellular fluid via the Uptake-2 carrier after the infusion ends.

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NOREPINEPHRINE: THE MAIN SyMPATHETIC CARDIOvASCuLAR NEuROTRANSMITTER

SNS

Uptake-1

PLASMA

NE

FIGURE 6.2 Diagram of sympathoneural, hepatic, adrenomedullary, and gut contributions to plasma levels of norepinephrine and its metabolites. Additional abbreviations: DHPG-S, DHPG sulfate; MHPG-S, MHPG sulfate; MN-S, MN sulfate; mPST, monoamine-preferring phenolsulfotransferase; NMN-S, NMN sulfate.

NE NE

MAO

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GUT CELL

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COMT MAO AD

VMA

VMA DHPG-S MHPG-S

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ADRENOMEDULLARY CELL

mPST

mPST

NMN

Norepinephrine Metabolism NE taken up into the axoplasm by the Uptake-1 transporter is subject to two fates, translocation into storage vesicles and deamination by monoamine oxidase (MAO). The combination of enzymatic breakdown and vesicular uptake constitute an intraneuronal “sink,” which keeps axoplasmic NE concentrations very low. Reserpine prevents the conservative recycling of NE and depletes NE stores. After reserpine injection, plasma DHPG levels increase rapidly, reflecting marked net leakage of NE from vesicular stores, and then decline to very low levels, reflecting decreased vesicular uptake of DA and decreased conversion of DA to NE. MAO catalyzes the oxidative deamination of DA to form DOPAC and of NE to form DHPG. Because of the efficient uptake and reuptake of catecholamines into the axoplasm of catecholamine neurons, and the rapid exchange of amines between the vesicles and axoplasm, the neuronal pool of MAO, located in the outer mitochondrial membrane, figures prominently in the overall functioning of catecholamine systems. Two isozymes of MAO, MAO-A and MAO-B, have been described. Clorgyline blocks MAO-A, and deprenyl and rasagiline block MAOB. MAO-A predominates in neural tissue, whereas both subtypes exist in non-neuronal tissue. Inhibitors of MAO-A potentiate pressor effects of tyramine, whereas inhibitors of MAO-B do not. NE and EPI are substrates for MAO-A, and DA is a substrate for both MAO-A and MAO-B. The immediate products of the deamination are short-lived aldehydes. For DA, the aldehyde intermediate,

dihydroxyphenylacetaldehyde (DOPAL) is converted rapidly to DOPAC by aldehyde dehydrogenase; for NE, the aldehyde intermediate is converted mainly to DHPG by an aldehyde/aldose reductase. The formation of the aldehydes reduces a flavine component of the enzyme. The reduced enzyme reacts with molecular oxygen, regenerating the enzyme but also producing hydrogen peroxide. Catechol-O-methyltransferase (COMT) in non-neuronal cells catalyzes the O-methylation of DHPG to form methoxyhydroxyphenylglycol (MHPG) and of DOPAC to form homovanillic acid (HVA). In the liver MHPG is converted vanillylmandelic acid (VMA), and in splanchnic organs catecholamines (especially DA) and their glycol metabolites are extensively sulfoconjugated. The main end-products of NE metabolism in plasma, therefore, are MHPG, conjugated MHPG, and VMA. Although MAO inhibitors are effective anti-depressants, a phenomenon known as the “cheese effect” limits their clinical use. In patients taking MAO inhibitors, administration of sympathomimetic amines such as in many nonprescription decongestants or ingestion of foods such as aged cheese, wine, or meat, which contain tyramine, can produce paroxysmal hypertension. Since tyramine and other sympathomimetic amines displace NE from sympathetic vesicles into the axoplasm, blockade of MAO in this setting causes non-exocytotic NE release, stimulating cardiovascular smooth muscle cells and producing intense vasoconstriction and hypertension. COMT catalyzes the conversion of NE to normetanephrine (NMN) and EPI to metanephrine (MN). Uptake-2 and COMT probably act in series to remove and degrade

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circulating catecholamines. The methyl group donor for the reaction is S-adenosyl methionine. COMT is expressed by non-neuronal cells and adrenomedullary chromaffin cells but not by sympathetic neurons. Ongoing production of NMN and MN in such cells explains the high sensitivity of plasma levels of free (unconjugated) metanephrines in the diagnosis of pheochromocytomas, which are chromaffin cell tumors.

BIOMARKERS OF SYMPATHETIC NORADRENERGIC INNERVATION AND FUNCTION Simultaneous measurement of plasma levels of NE and its metabolites provides valuable information about sympathetic noradrenergic innervation and function. In general, sympathetic noradrenergic stimulation produces similar absolute increments in plasma NE and DHPG levels. In contrast, decreased Uptake-1 activity increases plasma levels of plasma NE more than of DHPG. Under resting conditions plasma DHPG is determined mainly by net leakage of NE from vesicles into the cytosol – an ongoing passive process. Thus, plasma DHPG is related indirectly to NE stores in sympathetic nerves. Because loss of sympathetic noradrenergic terminals can compensatorily increase nerve traffic, NE release and plasma NE levels may be maintained until denervation is advanced. Accordingly, plasma DHPG provides a more sensitive index of sympathetic noradrenergic denervation than does plasma NE. Assessments of effects of neuropharmacologic probes such as tyramine, desipramine, atomoxetine, yohimbine, clonidine, trimethaphan, and isoproterenol are often included in clinical studies of sympathetic noradrenergic function. In general, physiological measurements such as of blood pressure, heart rate, or local vascular resistances are not as sensitive as neurochemical measurements in detecting and characterizing sympathetic noradrenergic dysfunction. This is because patients with autonomic failure typically also have baroreflex failure, and patients with sympathetic noradrenergic denervation typically also have augmented cardiovascular responses to agonists at adrenoceptors, complicating the relationship between NE release and physiological dependent measures.

NE AND METABOLOMICS Patterns of plasma levels of catechols yield metabolomic information reflecting abnormal enzymatic activities that can be relevant to the diagnosis of several disorders. Because of the dependence of DBH on copper, all patients with Menkes disease, an X-linked recessive disorder of a copper-ATPase, have high plasma DOPAC:DHPG and DA:NE ratios. Familial dysautonomia and DBH deficiency also entail elevated ratios of DA metabolites to NE

metabolites. Due to the vesicular localization of DBH, the same abnormal pattern characterizes low VMAT-2 activity. Decreased TH activity, from reduced synthesis either of the enzyme itself or of the required co-factor tetrahydrobiopterin, results in low levels of all the endogenous catechols. LAAAD deficiency is associated with high DOPA:NE, DOPA:DHPG, and DOPA:DOPAC ratios. Decreased Uptake-1 activity, such as by hypofunctional mutation of the gene encoding the NET, results in relatively larger increments in plasma NE than in plasma DHPG during sympathetic noradrenergic stimulation. Decreased MAO-A activity from treatment with an MAO inhibitor or from mutation of the X-linked gene encoding MAO-A manifests as high NE:DHPG and DA:DOPAC ratios. Since DHPG exiting sympathetic noradrenergic neurons is extensively O-methylated by COMT, low COMT activity is associated with increased DHPG:MHPG ratios. Theoretically, decreased alcohol dehydrogenase activity should be associated with elevated MHPG:VMA ratios.

DIFFERENTIAL NORADRENERGIC VS. ADRENERGIC ACTIVATION IN STRESS Different stressors can elicit different patterns of sympathoneural outflows and therefore differential NE release in the various vascular beds. This patterning redistributes blood flows. Local sympathoneural release of NE also markedly affects cardiac function and glandular activity. The adjustments usually are not sensed, and the organism usually does not feel distressed. Examples of situations involving prominent changes in sympathoneural outflows include orthostasis, mild exercise, post-prandial hemodynamic changes, mild changes in environmental temperature, and performance of non-distressing locomotor tasks. In contrast, in response to perceived global threats whether from external physical or internal psychological or metabolic stimuli – especially when the organism senses an inability to cope with those stimuli – increased neural outflow to the adrenal medulla elicits catecholamine secretion into the adrenal venous drainage. In humans, the predominant catecholamine in the adrenal venous drainage is EPI. EPI rapidly reaches all cells of the body (with the exception of most of the brain), producing a wide variety of hormonal effects at low blood concentrations. One can comprehend all of the many effects of EPI in terms of countering acute threats to survival that mammals have faced throughout their evolution, such as sudden lack of metabolic fuels, trauma with hemorrhage, and antagonistic confrontations. Even mild hypoglycemia elicits marked increases in plasma levels of EPI, in contrast with relatively small increases in levels of NE. Distress accompanies all these situations, the experience undoubtedly fostering the long-term survival of the individual and the species by motivating avoidance learning and producing signs that are understood instinctively by other members of the species.

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DIffERENTIAL NORADRENERGIC vS. ADRENERGIC ACTIvATION IN STRESS

Across a variety of stressors, increases in adrenomedullary activity as indicated by elevated plasma EPI levels correlate more closely with increases in pituitaryadrenocortical activity as indicated by elevated plasma levels of corticotropin than with increases in sympathoneural activity as indicated by elevated plasma levels of NE. Thus, insulin-induced hypoglycemia produces large increases in plasma EPI and ACTH levels and rather mild NE responses, whereas cold exposure produces increases in plasma NE levels, with little or no increases in EPI or ACTH. These findings call into question the long held concept of a unitary sympathoadrenal system activated in a stereotypical manner during stress.

Further reading Eisenhofer G, Kopin IJ, Goldstein DS. Catecholamine metabolism: A contemporary view with implications for physiology and medicine. Pharmacol Rev 2004;56:331–49.

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Goldstein DS. The autonomic nervous system in health and disease. New York: Marcel Dekker, Inc. 2001. Goldstein DS. Adrenaline and the inner world: An introduction to scientific integrative medicine. Baltimore: The Johns Hopkins University Press; 2006. Goldstein DS. Catecholamines 101. Clin Autonomic Res 2010;20:331–52. Goldstein D.S. Neurocardiology: Therapeutic implications for cardiovascular disease. Cardiovasc Ther 2010 Nov 25. doi: 10.1111/j.17555922.2010.00244.x. [Epub ahead of print] Goldstein DS, Holmes C, Kaler SG. Relative efficiencies of plasma catechol levels and ratios for neonatal diagnosis of Menkes disease. Neurochem Res 2009;34:1464–8. Goldstein DS, Holmes C. Neuronal source of plasma dopamine. Clin Chem 2008;54:1864–71. Goldstein DS, Kopin IJ. Adrenomedullary, adrenocortical, and sympathoneural responses to stressors: A meta-analysis. Endocr Regul 2008;42:111–9. Lenders JWM, Pacak K, Walther MM, Linehan WM, Mannelli M, Friberg P, et al. Biochemical diagnosis of pheochromocytoma: which test is best? JAMA 2002;287:1427–34.

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C H A P T E R

7 Tyrosine Hydroxylase Kazuto Kobayashi, Toshiharu Nagatsu INTRODUCTION

approximately 8.5 kb (GenBank Accession Number: BC104967). Alternative splicing from a single gene produces four kinds of mRNAs encoding TH isoforms (hTH1-4) that are different in amino acid sequences in the N-terminal region (see Fig. 7.1A) [see ref. 1 for detail]. The hTH1-4 isoforms are distinguishable by the combination of insertion/deletion of the 12-bp sequence (4 amino acids) in the 3-terminal portion of exon 1 and the 81-bp sequence (27 amino acids) corresponding to exon 2. The hTH1 isoform contains the shortest amino acid sequence (497 amino acids), and the hTH2, hTH3, and hTH4 isoforms contain additional sequences of 4, 27, and 31 amino acids, respectively, between the 30th and 31st amino acids in the hTH1 isoform. TH activity is regulated by phosphorylation mainly at serines 19, 31, and 40 in the N-terminal region (hTH1). In particular, the TH isoforms appear to be differentially modified by extracellular signal-regulated protein kinase (ERK) [3]. ERK phosphorylates serine 31 in the hTH1 isoform, whereas ERK phosphorylation of the corresponding site in the hTH2-4 isoforms is less efficient. The phosphorylation of serine 19 increases the rate of phosphorylation on serine 44 in the hTH2 isoform (corresponding to serine 40 in the hTH1) more greatly than phosphorylation of the site in the hTH1 isoform. Dumas et al. [4] subsequently demonstrated the generation of more isoforms encoding human TH through skipping exon 3 in the adrenal medulla (Fig. 7.1B). These isoform are overexpressed in the tissue of patients with progressive supranuclear palsy, a neurodegenerative disease mostly affecting the basal ganglia, cerebral cortex, and brain stem. The isoforms lack 74 amino acids containing the two major phosphorylation sites (serines 31 and 40). In addition, Ohye et al. [5] report a new splicing variant in the adrenal medulla that encodes TH isoform lacking exon 4 sequence, resulting in truncation of the C-terminal region of the protein. This splicing seems to cause inactivation of TH function. These recent data support that alternative splicing produces the functional diversity of TH isoforms, which may be involved in the complex mechanisms that regulate catecholamine metabolism. Altered metabolism related to TH gene expression may underlie the pathophysiology of some neurological and neurodegenerative diseases.

Tyrosine hydroxylase (TH; tyrosine 3-monooxygenase) (EC 1.14.16.2) catalyzes the conversion of L-tyrosine to L-3,4dihydroxyphenylalanine (L-DOPA), which is the initial and rate-limiting step in the biosynthetic pathway of catecholamines including dopamine, noradrenaline, and adrenaline [reviewed in ref. 1]. These catecholamines play important roles in a variety of physiological and behavioral functions in the nervous and endocrine systems. TH activity is regulated through different mechanisms at the levels of transcription of the gene and posttranslational modification of the protein. Gene targeting technique of the mouse genome reveals that TH function is essential for animal development and survival as well as brain function [reviewed in ref. 2]. Disruption of the TH gene (homozygous mutation) causes the lethality at the embryonic and neonatal stages because of alterations in the cardiovascular system. Reduction of TH activity in the mice heterozygous for the mutation impairs retention of long-term memory at the adult stage. Recent advances in molecular biological study of the human TH gene have shown that a number of TH isoforms are generated by alternative splicing from a single gene, suggesting a complex mechanism that generates the diversity of TH isoforms in the normal and pathological conditions. Human genetic studies have found mutations that lead to TH deficiency, showing an infantile onset, progressive hypokinetic-rigidity with dystonia or a complex encephalopathy with neonatal onset. In addition, the use of TH gene function has provided a useful approach for gene therapy trials of Parkinson’s disease. In the present chapter, we describe recent advances in molecular biological and genetic studies related to human TH gene. We recapitulate the mechanism that generates the diversity of human TH isoforms, the implication of TH mutations in congenital diseases, and the update of gene therapy trials for neurological disease by the use of TH gene.

DIVERSITY OF HUMAN TH GENE EXPRESSION The human TH gene is encoded on chromosome 11p15.5 and composed of 14 exons, spanning

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7. TyRosInE HydRoxylAsE

FIGURE 7.1 Schematic illustration of alternative splicing from a single TH gene that generates the diversity of isoforms. (A) Alternative splicing events producing four kinds of mRNAs encoding TH isoforms (hTH1-4). These mRNAs are distinguishable by the combination of insertion/deletion of the 12-bp sequence in the 3-terminal portion of exon 1 and the 81-bp sequence corresponding to exon 2. (The data are cited from ref. 1.) (B) Splicing events producing the mRNAs lacking exon 3. These mRNAs correspond to hTH1, hTH3 and hTH4 mRNAs deleting exon 3. (The data are cited from ref. 4.)

CONGENITAL DISEASE ASSOCIATED WITH TH MUTATION

GENE THERAPY WITH TH GENE FUNCTION

TH deficiency is an autosomal recessive disorder associated with the mutations in the TH gene (see Fig. 7.2). The clinical phenotype of the disease shows an infantile onset, progressive L-DOPA-responsive dystonia or a progressive encephalopathy with L-DOPA-nonresponsive dystonia. This deficiency can be diagnosed by measuring the levels of catecholamine metabolites such as homovanillic acid and 3-methoxy-4-hydroxyphenylethylene glycol in cerebrospinal fluid and direct sequencing of the TH gene. So far, the deficiency is reported in fewer than 40 patients worldwide. TH mutations in the deficiency have been extensively studied, and the clinical, biochemical, and genetic data are summarized in ref. [6]. The majority of TH mutations (~95%) were missense mutations that lead to amino acid substitutions in the protein-coding region. Some of these mutations are clarified to result in partial loss of enzymatic activity in the bacterial expression system. In addition, deletion of a single nucleotide in the coding region is reported that generates a truncated form of TH protein. Single nucleotide substitutions in the cyclic AMP response element (CRE) in the promoter region of TH gene are also found in some cases. Since the CRE in the TH promoter is known to be important for both basal and inducible transcriptional activity, the mutations in the CRE may down-regulate the transcription of TH gene, affecting the TH expression level. The relationship between the mutation and clinical phenotype does not appear to be fully understood.

Parkinson’s disease is an age-related neurodegenerative disease caused by the progressive loss of dopamine neurons in the ventral midbrain and the consequent reduction of the dopamine level. The patients exhibit a range of clinical phenotypes with the most common motor deficits such as the resting tremor, rigidity, akinesia, and postural instability. The standard procedure to treat the disease is pharmacotherapy by oral administration of L-DOPA, but many patients gradually develop L-DOPA-induced dyskinesia and motor fluctuation. Gene therapy trials offer alternative, complementary approaches for clinical application of Parkinson’s disease [7]. One potential strategy for gene therapy is to produce dopamine in striatal cells through gene transfer of enzymes related to dopamine biosynthesis. Muramatsu et al. [8] report gene therapy trial of a non-human primate model for Parkinson’s disease by adeno-associated viral vectors that carry the genes encoding TH, GTP cyclohydrolase I (GCH1), and aromatic L-amino acid decarboxylase (AADC). GCH1 is the rate-limiting enzyme in the biosynthesis of the cofactor for TH, tetrahydrobiopterin (BH4), and AADC converts L-DOPA to dopamine. Ectopic expression of these three enzymes successfully achieves the increased production and release of dopamine, and thus improves behavioral deficits in the parkinsonian model. Gene therapy for continuous L-DOPA delivery is also a potentially useful therapeutic strategy for the disease. Björklund et al. [9] report that gene transfer of TH and GCH1 genes into striatal cells

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ConClusIon

47

FIGURE 7.2 Overview of the mutations in TH deficiency. The exon/intron structure of the human TH gene is shown. The closed boxes indicate the protein-coding regions and the open boxes indicate 5- and 3-untranslated regions. Three mutations (71C  T, 70G  A, and 69T A) are located in the cyclic AMP response element in the TH promoter (the nucleotides 67 to 74 upstream of the initiation codon). The 295delC and 296delT mutations cause the frame shift in the protein-coding region. The 1198-24T  A mutation in the putative branchpoint sequence in intron 11 results in alternative splicing that produces aberrant mRNAs. Other mutations generate amino acid substitutions in the protein-coding region. (The data are cited from ref. 6.)

of the parkinsonian model rats with adeno-associated viral vectors results in ectopic L-DOPA production, showing the optimal ratio of the amount of TH and GCH1 enzymes for the in vivo DOPA production. The elevated level in BH4 by GCH1 expression appears to contribute to the stabilization of TH protein. A subsequent study with positron emission tomography imaging demonstrates that the straital transfer with TH and GCH1 genes enables behavioral recovery correlated with correction of dopamine transmission in the striatum [10]. These gene therapy trials aiming at the delivery of dopamine or L-DOPA provide a reasonable, promising strategy for clinical application in the disease.

CONCLUSION In the present chapter, we presented a short review of recent advances of molecular biological and genetic studies of human TH gene. The functional diversity of TH isoforms is generated by alternative splicing from a single gene. This diversity appears to mainly affect the regulatory mechanism of TH activity through phosphorylation of the protein. The different regulatory mechanism may be implicated in the control of catecholamine metabolism. Mutations in the coding or promoter region of the human TH gene exist in congenital TH deficiency with the infantile onset, progressive hypokinetic-rigidity with dystonia or a complex encephalopathy with neonatal onset. Future genetic study will address the question whether mutations in the TH gene are involved in the neuropsychiatric diseases. Gene therapy trials with the use of TH gene function aiming at the in vivo delivery of dopamine or L-DOPA indeed show the recovery of behavior and brain dopamine metabolism in animal models for Parkinson’s disease. This strategy will generate a viable procedure for clinical application in the disease.

The progress on the TH gene studies provides a clearer understanding of catecholamine function in the pathophysiology in neurological and neurodegenerative diseases and a novel strategy for the management of these diseases.

References [1] Nagatsu T. The catecholamine system in health and diseases – Relation to tyrosine 3-monooxygenase and other catecholamine-synthesizing enzymes. Proc Jpn Acad Ser B Phys Biol Sci 2006;82:388–415. [2] Kobayashi K, Nagatsu T. Molecular genetics of tyrosine 3-monooxygenase and inherited diseases. Biochem Biophys Res Commun 2005;338:267–70. [3] Lehman IT, Bobrovskaya L, Gordon SL, Dunkley PR, Dickson PW. Differential regulation of the human tyrosine hydroxylase isoforms via hierarchical phosphorylation. J Biol Chem 2006;281:17644–17651. [4] Dumas S, Hir HL, Bodeau-Péan S, Hirsch E, Thermes C, Mallet J. New species of human tyrosine hydroxylase mRNA are produced in variable amounts in adrenal medulla and are overexpressed in progressive supranuclear palsy. J Neurochem 1996;67:19–25. [5] Ohye T, Ichinose H, Yoshizawa T, Kanazawa I, Nagatsu T. A new splicing variant for human tyrosine hydroxylase in the adrenal medulla. Neurosci Lett 2001;312:157–60. [6] Willemsen MA, Verbeek MM, Kamsteeg E-J, de Rijk-van Andel JF, et al. Tyrosine hydroxylase deficiency: a treatable disorder of brain catecholamine biosynthesis. Brain 2010;133:1810–22. [7] Feng LR, Maguire-Zeiss KA. Gene therapy in Parkinson’s disease: rationale and current status. CNS Drugs 2010;24:177–92. [8] Muramatsu S, Fujimoto K, Ikeguchi K, Shizuma N, et al. Behavioral recovery in a primate model of Parkinson’s disease by triple transduction of striatal cells with adeno-associated viral vectors expressing dopamine-synthesizing enzymes. Hum Gene Ther 2002;13:345–54. [9] Björklund T, Hall H, Breysse N, Soneson C, Carlsson T, Mandel RJ, et al. Optimization of continuous in vivo DOPA production and studies on ectopic DA synthesis using rAAV5 vectors in Parkinsonian rats. J Neurochem 2009;111:355–67. [10] Leriche L, Björklund T, Breysse N, Besret L, Grégoire M-C, Carlsson T, et al. Positron emission tomography imaging demonstrates correlation between behavioral recovery and correction of dopamine neurotransmission after gene therapy. J Neurosci 2009;29:1544–53.

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C H A P T E R

8 Antidepressant-Sensitive Norepinephrine Transporters: Structure and Regulation Maureen K. Hahn Chemical signaling at central and peripheral noradrenergic synapses is terminated through reuptake of released norepinephrine (NE) [1]. The protein executing this activity, the antidepressant-sensitive NE transporter (NET), has been cloned from multiple species including man [2]. The human NET (hNET) gene encodes a 617 amino acid polypeptide. A high resolution structure of a SLC6 bacterial family member, the leucine transporter (LeuTAa), from Aquifex aeolicus, was recently achieved, confirming the structure of this family as twelve transmembrane domain proteins with intracellular NH2 and COOH termini (Fig. 8.1) [3]. Expression of the original hNET isolate is sufficient to confer antidepressant-sensitive NE transport [4]. During biosynthesis, hNET protein is N-glycosylated at sites on a large extracellular loop between transmembrane domains (TMDs) 3 and 4, and the protein is subsequently trafficked to the plasma membrane [5]. Although hNET glycosylation is not known to be regulated, this modification enhances protein stability and transport activity [6]. Once inserted in the plasma membrane, hNET is exposed to the transmembrane sodium gradient that the transporter utilizes to bring about NE uptake. Extracellular chloride also facilitates uptake of NE

and intracellular potassium may offer further stimulation, though evidence that potassium is countertransported is lacking. Additionally, hNET supports NE-gated channel states that may allow NE to translocate hNET at high rates when synaptic concentrations are elevated [7]. To take advantage of this possibility, NETs would need to be enriched at synaptic sites of release. Consistent with this, studies with NET-specific antibodies reveal NET localized to the plasma membrane of synaptic boutons [8]. Additional hNET splice variants have been identified at the mRNA level, predicting additional NET species with altered COOH termini [9]. Whereas related, though not identical, mRNA variants have been identified in rat brain and bovine adrenal, evidence of splice variant protein expression is currently lacking [10,11]. Recent heterologous expression studies incorporating these variants reveal significantly disrupted maturation and surface trafficking, raising doubt as to their functional relevance, at least as monomers [12]. In this regard, although a single hNET cDNA can confer NE transport activity, multiple hNET proteins may assemble together as an oligomeric complex. Indeed, dimer or higher order complexes have been documented for the closely related dopamine transporter (DAT) and serotonin transporter (SERT) and the presence of oligomers of hNET would offer an explanation for genetic NET deficiency exhibited by subjects with a mutation in one of two hNET alleles [13–15]. Mouse models of genetic NET deficiency support a contribution of transporter expression to presynaptic NE homeostasis, extracellular NE clearance and psychostimulant action [16]. Studies conducted over the past few years have revealed that NET and related transporters are subject to rapid regulation including changes in transporter surface expression as well as intrinsic transport activity [17,18]. This regulatory potential is believed to involve both transporter phosphorylation and the regulated associations of accessory proteins. Thus, NET has been found to complex with the catalytic subunit of protein phosphatase 2A (PP2Ac), the SNARE protein syntaxin 1A as well as the scaffolding protein PICK1 [18–20]. The NH2 terminus of hNET supports syntaxin 1A associations whereas the transporter’s COOH terminal PDZ domain is required for PICK1 binding. Protein kinase C activators disrupt Syntaxin and PP2Ac associations,

NE, Na+, Cl–

Extracellular

Intracellular P P

FIGURE 8.1 Norepinephrine transporters. Depicted is the 12transmembrane topology predicted for the NET protein, bearing three N-glysosylated residues on the second extracellular loop. The cytoplasmic NH2 and COOH termini bear multiple consensus sites for Ser/Thr phosphorylation. Norepinephrine (NE) is depicted as cotransported with Na and Cl which provide the energy for uptake of catecholamine into the presynaptic terminal.

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8. ANTIdEPRESSANT-SENSITIvE NoREPINEPHRINE TRANSPoRTERS: STRuCTuRE ANd REgulATIoN

NE

[3] TAPs dissociate

[4]

Phosphorylation NT

[5]

Recycling

Internalization

[6]

Dephosphorylation

[7]

FIGURE 8.2 Norepinephrine transporter regulation. Illustrated is a cycle for the regulated trafficking of NET proteins. NETs are depicted in complex with multiple associated proteins that may stabilize the transporter at the plasma membrane. Stimuli linked to receptor activation by other neurotransmitters, hormones or cytosolic second messengers influence the stability of the NET associated protein complex and enhance NET phosphorylation. In parallel with, or as a result of such stimulation, NET proteins internalize leaving reduced presynaptic NE transport capacity. NETs can recycle to the plasma membrane, a process that likely occurs at basal rates but which may be linked to regulatory stimuli. TAP, transporter-associated proteins.

stimulation that also leads to NET internalization. PP2Ac also appears to be required for alterations in hNET intrinsic activity triggered by insulin and MAP-kinase linked pathways [17]. Possibly, trafficking and intrinsic function are coregulated through kinase-dependent mechanisms. Insulin signaling to promote NET internalization, an effect mediated through protein kinase B (Akt), occurs in both brain and peripheral neurons in vivo, thus providing a potential link between diseases of metabolism and neurobehavioral disorders [2]. Although these are early days in molecular studies of NET regulation, it is already clear that NE clearance capacity is likely to involve multiple regulatory proteins that localize, stabilize and activate NET (Fig. 8.2). They may also influence the tendency of NET to support catecholamine efflux when ion gradients are perturbed as they can be in ischemic insults. Finally, successful assimilation of these proteins into a regulatory model of NE inactivation should offer new opportunities to manipulate NET in autonomic and mental disorders, such as depression, where altered noradrenergic signaling has been recognized.

Acknowledgements

[8]

[9]

[10]

[11]

[12]

[13]

[14]

[15]

[16]

[17]

[18]

M.K. Hahn gratefully acknowledges the NIMH for their support of studies on NET genetics and regulation. [19]

References [1] Iversen LL. Uptake processes for biogenic amines. In: Iversen LL, editor. Handbook of Psychopharmacology (third ed.). New York: Prenum Press; 1978. p. 381–442. [2] Robertson SD, Matthies HJ, Owens WA, Sathananthan V, Christianson NS, Kennedy JP, et al. Insulin reveals Akt signaling as

[20]

a novel regulator of norepinephrine transporter trafficking and norepinephrine homeostasis. J Neurosci 2010;30(34):11305–11316. Yamashita A, Singh SK, Kawate T, Jin Y, Gouaux E, et al. Crystal structure of a bacterial homologue of Na/Cl -dependent neurotransmitter transporters. Nature 2005;437(7056):215–23. Pacholczyk T, Blakely RD, Amara SG. Expression cloning of a cocaine- and antidepressant-sensitive human noradrenaline transporter. Nature 1991;350(6316):350–4. Melikian HE, McDonald JK, Gu H, Rudnick G, Moore KR, Blakely RD, et al. Human norepinephrine transporter. Biosynthetic studies using a site-directed polyclonal antibody. J Biol Chem 1994;269(16):12290–12297. Melikian HE, Ramamoorthy S, Tate CG, Blakely RD. Inability to N-glycosylate the human norepinephrine transporter reduces protein stability, surface trafficking, and transport activity but not ligand recognition. Mol Pharmacol 1996;50(2):266–76. Galli A, Blakely RD, DeFelice LJ. Patch-clamp and amperometric recordings from norepinephrine transporters: channel activity and voltage-dependent uptake [see comments]. Proc Natl Acad Sci USA 1998;95(22):13260–13265. Matthies HJ, Han Q, Shields A, Wright J, Moore JL, Winder DG, et al. Subcellular Localization of the antidepressant-sensitive norepinephrine transporter. BMC Neurosci 2009;10(1):65. Pörzgen P, Bönisch H, Hammermann R, Brüss M. The human noradrenaline transporter gene contains multiple polyadenylation sites and two alternatively spliced C-terminal exons. Biochim Biophys Acta 1998;1398(3):365–70. Burton LD, Kippenberger AG, Lingen B, Bruss M, Bonisch H, Christie DL, et al. A variant of the bovine noradrenaline transporter reveals the importance of the C-terminal region for correct targeting to the membrane and functional expression. Biochem J 1998; 330(Pt 2):909–14. Kitayama S, Ikeda T, Mitsuhata C, Sato T, Morita K, Dohi T, et al. Dominant negative isoform of rat norepinephrine transporter produced by alternative RNA splicing. J Biol Chem 1999; 274(16):10731–10736. Bauman PA, Blakely RD. Determinants within the C-terminus of the human norepinephrine transporter dictate transporter trafficking, stability, and activity. Arch. Biochem Biophys 2002;404(1):80–91. Hastrup H, Karlin A, Javitch JA. Symmetrical dimer of the human dopamine transporter revealed by cross-linking Cys-306 at the extracellular end of the sixth transmembrane segment. Proc Natl Acad Sci U S A 2001;98(18):10055–10060. Kilic F, Rudnick G. Oligomerization of serotonin transporter and its functional consequences. Proc Natl Acad Sci USA 2000;97(7):3106–11. Shannon JR, Flattem NL, Jordan J, Jacob G, Black BK, Biaggioni I, et al. Clues to the origin of orthostatic intolerance: a genetic defect in the cocaine- and anti-depressant sensitive norepinephrine transporter. N Eng J Med 2000;342(8):541–9. Xu F, Gainetdinov RR, Wetsel WC, Jones SR, Bohn LM, Miller GW. et al. Mice lacking the norepinephrine transporter are supersensitive to psychostimulants. Nat Neurosci 2000;3(5):465–71. Apparsundaram S, Sung U, Price RD, Blakely RD. Traffickingdependent and -independent pathways of neurotransmitter transporter regulation differentially involving p38 mitogen-activated protein kinase revealed in studies of insulin modulation of norepinephrine transport in SK-N-SH Cells. J Pharmacol Exp Ther 2001;299(2):666–77. Bauman HJ, Apparsundaram S, Ramamoorthy S, Wadzinski BE, Vaughan RA, Blakely RD, et al. Cocaine and antidepressant-sensitive biogenic amine transporters exist in regulated complexes with protein phosphatase 2A. J Neurosci 2000;20(20):7571–8. Sung U, Apparsundaram S, Galli A, Kahlig K, Savchenko V, Schroeter S, et al. A regulated interaction of syntaxin 1A with the antidepressant-sensitive norepinephrine transporter establishes catecholamine clearance capacity. J Neurosci 2003;23(5):1697–709. Torres GE, Yao W, Mohn AR, Quan H, Kim K, Levey AI, et al. Functional interaction between monoamine plasma membrane transporters and the synaptic PDZ domain-containing protein PICK1. Neuron 2001;30(1):121–34.

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9 α1-Adrenergic Receptors Marion C. Mohl, Robert M. Graham

a1-ADRENORECEPTOR SUBTYPES

gene family in the human genome – α1-ARs are long, single chain, integral membrane proteins. Like rhodopsin and β-ARs, α1-ARs contain seven transmembrane (TM)spanning α-helical domains linked by three intracellular and three extracellular loops [1]. In addition, the juxtamembranous portion of the C-terminal tail likely forms an eighth α-helix that lies parallel to the plain of the membrane and is importantly involved in receptor signaling – a domain structure found in all class A GPCRs for which highresolutions structures are available, but not in the recently determined structure of the CXCR4 chemokine receptor [2]. As with all GPCRs, the N-terminus of α1-AR subtypes is located extracellularly and the C-terminus, intracellularly (Fig. 9.1). The ligand-binding pocket is formed by the clustering of the seven α-helical domains to form a water accessible region for agonist to bind, and is located in the outer (extracellular) one third of the membranespanning domain. Residues of the intracellular (cytoplasmic) domains, particularly the third intracellular loop, mediate specific interactions with their cognate G proteins and, thus, are involved in receptor-engagement with its signaling and regulatory pathways.

Three subtypes have been identified by molecular cloning in several species including humans. They are classified as the α1A (previously the α1A/c)-, the α1B- and α1D (previously the α1a or α1a/d)-ARs (for a detailed consideration of α1-AR subtype-classification, see ref. 1). For the α1A-AR, four splice variants (α1A-1, α1A-2, α1A-3, α1A-4) have been identified, and are expressed at different levels in various tissues, including the liver, heart and prostate. However, the functional significance of these splice variants is unclear, since they all display similar ligandbinding and functional activity when expressed in heterologous cell systems. The characteristics of the various α1-AR subtypes are shown in Table 9.1. In addition to these subtypes, a putative fourth subtype, the α1L-AR, which displays low affinity for prazosin and other α1-antagonists, has been identified by functional but not by molecular studies. This subtype is postulated to mediate contraction of prostate smooth muscle, but it remains unclear if it is a distinct subtype or a merely a functional variant of one of the cloned α1-subtypes, such as the α1A splice variants, since the latter all display some of the characteristics of an α1L-subtype when expressed in heterologous cell systems. Another molecular variant of the α1A-AR is a coding region polymorphism that involves an arginine to cysteine (Arg492Cys) substitution in an arginine-rich region of the C-terminal tail. This polymorphic receptor is found with higher frequency in African-Americans, but it is not associated with essential hypertension. In addition, it is unclear if it alters receptor regulation or produces any other functional effects.

LIGAND BINDING AND ACTIVATION OF a1-ADRENORECEPTORS Binding of catecholamines to α1-AR involves an ionic interaction between the basic aliphatic nitrogen atom common to all sympathomimetic amines and an aspartate (Asp125 in the hamster α1B-AR) in the third transmembranespanning segment (TMIII) [3]. In the ground state, this TMIII aspartate forms a salt bridge with a lysine residue (Lys331 in the α1B-adrenergic receptor) in TMVII. Activation of α1-ARs, likely involves disruption of this ionic interaction. This is due to competition between the protonated amine of catecholamines and the TMVII lysine, which is just favored by the slightly more basic pKa of the protonated amine (pKa 11.0) versus the lysine (pKa 10.5). Agonistbinding to α1-ARs also involves an H-bond interactions between the meta hydroxyl group of catecholamines and a serine residue (Ser188, α1A-receptor numbering) in TMV,

STRUCTURE OF a1-ADRENORECEPTORS α1-ARs are integral membrane glycoproteins and members of the biogenic amine or class A family (also includes α2- and β-ARs, as well as the light-activated photoreceptor, rhodopsin) of G-protein-coupled receptors (GPCRs) [1]. Like other members of the GPCR-superfamily – the largest family of membrane receptors and possibly the largest

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9. α1-ADRENERGIC RECEPTORS

TABLE 9.1 Characteristics of the α1A-Adrenoreceptor Subtypes Receptor Subtype α1A

α1B

α1D

68

80

≈65

Amino acids

431–501

515–520

561–572

Gylcosylation sites (N-terminus)

3

4

2

Phosphorylation sites

PKA

PKA

–

Genomic organization Introns

1

1

2

2

2

8

5

20 p13

5-Methylurapidil, ()niguldipine, oxymetazoline, A-61603, SNAP5089, KMD-3213, RS17053

AH11110A, L-765,314

()Norepinephrine, BMY 7378, SKF105854

–

Prazosin, phentolamine, benoxathian, abanoquil, terazosin, doxazosin, tamulosin, phenylephrine, methoxamine, cirazoline

–

Prototypic tissues

Rat kidney and submaxillary gland, rabbit liver, human heart and liver

Rat spleen, liver, and heart

Rat aorta, lung, and cerebral cortex

Receptor-coupled signaling

–

Ca2 mobilization, PLC, PLA2, PLD

–

G-protein coupling

Gq/11/4

Gq/11/4/16, Gh

Gq11, Gh

Characteristics *

Mr

§

Exons †

Chromosomal localization

Pharmacological selectivity Subtype-selective agents

Nonselective agents‡

*Apparent molecular weights (Mr) determined by SDS-PAGE are shown (Table adapted from ref. 1). † Refers to the human genome. ‡ These characteristics are the same for all subtypes. § Differences due to species variability and/or to splice variants.

whereas an interaction between the para hydroxyl and another TMV serine (Ser192) contributes only minimally to receptor activation. A further important interaction, for both binding and activation, involves aromatic-aromatic bonding between the catecholamine ring and that of a phenylalanine (Phe310 for the α1B-adrenergic receptor) in TMVI [3]. This interaction is also important in receptor activation, which in addition to the TMIII–TMVII salt bridge disruption, mentioned above, involves movement of TMVI that is likely required to allow interaction between the intracellular third loop and the receptor’s cognate G-protein. Aromatic–aromatic interactions between two phenylalanines (Phe163 and Phe187) in TMIV and TMV, respectively, and the catecholamine ring, have also been suggested to be important for agonist binding [3], but not for activation, although that with Phe163 may be indirect, since this residue is replaced by a leucine in the α1B- and α1D-adrenergic receptors. Residues critical for subtype-selective agonist recognition have been evaluated and, importantly, just two of the approximately 172 residues in the transmembrane domains (Ala204 in TMV and Leu314 in TMVI, and Val185 in TMV and

Met293 in TMVI of the α1B- and α1A-receptors, respectively) have been shown to account entirely for the selective agonist-binding profiles of the α1A- and α1B-subtypes [1]. Interactions between antagonists and α1-ARs are less well-defined, although the selectivity of two α1Aantagonists, phentolamine and WB4101, involves interactions with three consecutive residues (Gly196, Val197 and Thr198) in the second extracellular loop [3].

FUNCTION OF a1-ADRENORECEPTORS The best characterized α1-AR-mediated function is that of smooth muscle contraction. As such, these receptors have a major role in the vascular system in controlling blood pressure, the baroreflex response to changes in blood pressure, and in temperature control. The importance of α1ARs in controlling sympathetically regulated arteriolar tone is evident, for example, from the fact that a component of the blood pressure lowering action of all currently available antihypertensive agents is attributable to an attenuation of α1-AR-responses. This is due either to a direct action at

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

FUNCTION OF α1-ADRENORECEPTORS

VI

VII

V

I

IV III

II

LIGAND

Extracellular

II

VII

IV

LIGAND

V

I Intracellular

III

FIGURE 9.1 Cut away 3D model of the hamster α1B-AR, showing the seven α-helical transmembrane domains indicated by Roman numerals and by dashed circles and backbone ribbons (corkscrews), with the catecholamine agonist, epinephrine (ball and stick model with surrounding dot surface) modeled in its binding pocket. Upper panel: top view (looking down onto the plane of the membrane). Lower panel: side view.

the level of post-junctional, vascular smooth muscle α1-ARactivation or signaling, or to an indirect effect via inhibition of sympathetic outflow and, thus, availability of the endogenous receptor-agonists, the catecholamines, epinephrine and norepinephrine. Therefore α1-ARs agonists or antagonists can be used in the treatment of hypotension or hypertension, respectively, although their use in hypertension may result in heart failure in susceptible patients, due possibly to blockade of important cardiomyocyte pro-survival effects [4,5]. Despite the critical role of α-ARs in mediating arteriolar vasoconstriction, the particular subtype(s) responsible remains unclear. Consistent with findings in rodents, clinical trials of α1A-selective blockers have implicated this subtype as a major regulator of blood pressure [11]. In contrast, studies of a patient with sympathotonic orthostatic hypotension indicate that the α1B-AR may be a major regulator of vascular resistance in humans. Moreover, expression of this subtype increases in the elderly at a time when hypertension is commonly manifested. Thus, there is evidence for both the α1A- and α1B-adrenergic receptors being involved in blood pressure control in humans. Studies in the rat and in mouse knockout-models indicate that the α1D-subtype

53

can also play a role in blood pressure regulation. Indeed, such studies of genetically engineered animal models [5] indicate that sympathetic regulation of vascular tone by the various α1-subtypes is complex and may involve crossregulation of their contractile effects and/or of their expression. For example, inactivation of the α1A-AR results in a small but significant decrease in basal blood pressure but no change in pressor responses to the α1-agonist, phenylephrine. However, inactivation of the α1B-AR results in no effect on basal blood pressure but the pressor responses to phenylephrine were significantly blunted or unchanged. Interestingly, with inactivation of both the α1A-AR and α1B-AR there was no significant fall in blood pressure. Inactivation of the α1D-AR, or both α1D-AR and α1B-AR resulted in a significant drop of the resting blood pressure and impaired vasoconstrictor responses to noradrenaline and phenylephrine. Given that overexpression of the α1Badrenergic receptor in transgenic mice does not increase systemic arterial blood pressure, and that inactivation of this subtype results in attenuation of phenylephrine pressorresponses, with no change in basal blood pressure, it could reasonably be suggested that the α1A- and α1D-AR are the mediators of vasoconstriction and that the α1B-AR has a modulatory rather than a direct contractile role. However, mice lacking both the α1A- and α1B-receptors, or both the α1B- and α1D-receptors, show profound attenuation of phenylephrine pressor-responses. Thus, under appropriate circumstances the α1B-subtype may contribute importantly to sympathetic regulation of arteriolar tone. Further studies are thus required to fully elucidate the mechanisms involved in this apparent latent contribution of the α1B-AR to peripheral resistance, and to determine if these findings in animal models are also germane to the regulation of vascular resistance and blood pressure in humans. Other functions mediated by α1-ARs, include bronchoconstriction, regulation of human lipid metabolism, uptake of glucose into adipocytes, and contractile effects in various tissues, such as the vas deferens and the myocardium. For example, inactivation of the α1A-AR in mice results in a 50% reduction in pregnancy rate, which is markedly enhanced to 10% in animals with inactivation of all three α1A/B/C-AR subtypes – an effect likely due to impaired contraction of the vas deferens and, thus, decreased sperm ejaculation. Cardiac specific overexpression of the wild type α1B-AR in mice results in depressed contractile responses to β-AR stimulation, whereas cardiac-specific overexpression of a constitutively active mutant α1B-AR results in hypertrophic responses to pressure overload, leading to heart failure and premature death. In contrast, α1A-AR overexpression leads to increased cardiac contractility but no hypertrophy and, furthermore, moderate overexpression (66–fold) improves outcomes after pressure overload and after myocardiac infarction. Marked overexpression (170–fold), however, leads to sudden cardiac death that is likely due to calcium overload rather than to arrhythmia development. α1-ARs, particularly, the α1Asubtype, also have important cardiomyocyte pro-survival and developmental growth effects [4,5].

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9. α1-ADRENERGIC RECEPTORS

Other important smooth muscle constrictor effects mediated by α1-ARs, include those relating to the lower urinary tract (bladder and prostate). The role of the α1A-subtype, in particular, is well-documented with α1A-selective antagonists being useful to relax prostate smooth muscle, relieve bladder outlet obstruction and, thus, to enhance urine flow (reviewed in ref. 6). Moreover, urinary incontinence is an unfortunate side effect of therapy with α1-AR blockers in patients with impaired bladder neck function. α1-ARs also regulate prejunctional or pre-synaptic neurotransmitter release, including inhibition of norepinephrine, acetylcholine and vasopressin release, and stimulation of inhibitory GABAergic neurotransmission in neurons and glial cells [7].

SIGNALING OF a1-ADRENORECEPTORS All three α1-ARs subtypes couple to a variety of second messenger proteins via heterotrimeric G-proteins, particularly of the Gαq/11 family, to phospholipase C leading to increased Ca2 entry via voltage-dependent or TRPC3 and or TRPC6 channels, or to release of Ca2 from intracellular stores [8]. Activation of α1-ARs augments arachidonic acid release through stimulation of phospolipase A or D, which can increase cAMP levels [8]. Inotropic effects of α1-ARs in rat hearts involve stimulation of Gαs-protein and reduction of the K current via the cAMP/PKA-mediated pathway. In addition to signaling through hetertriomeric G-proteins, α1ARs also mediate responses through the RhoA/Rho-kinase signaling pathway leading to Ca2 sensitization through phosphorylation of the myosin light chain phosphatase (MLCP) and the α1B and α1D, but not the α1A-subtype may also couple to phospholipase δ1-activation via Gh [9]. There is also evidence that α1-ARs form hetero- and homodimers, which alter their ligand binding and signaling properties, as well as their trafficking [8].

REGULATION OF a1-ADRENORECEPTORS α1-ARs are subject to agonist-induced regulation that results in both short and long-term desensitization of signaling [8,10]. These regulatory responses are mediated by agonist-induced conformational changes that lead to C-terminal tail receptor-phosphorylation, both by

receptor-linked protein kinase C as well as by G-proteincoupled receptor kinases, followed by the binding of arrestins and internalization by the clathrin pathway. These responses have been studied in most detail for the α1Breceptors. With the α1A- and α1D-ARs, the carboxyl terminal tail appears not to be essential for signaling or desensitization, and receptor internalization is not associated with α1A-AR desensitization.

Acknowledgements We are most grateful to Dr J. Novotny for the α1B-AR 3D model. Work from the author’s laboratory is supported in part by a Program Grant from the National Health and Medical Research Council of Australia (#573732), and a Grant-in-Aid from the Heart Foundation of Australia (G09S4342).

References [1] Graham RM, Perez DM, Hwa J, Piascik MT. Alpha(1)-adrenergic receptor subtypes – molecular structure, function, and signaling. Circ Res 1996;78:737–49. [2] Wu B, Chien EYT, Mol CD, Fenalti G, Liu W, Katritch V, Abagyan R, Brooun A, Wells P, Bi FC, Hamel DJ, Kuhn P, Handel TM, Cherezov V, Stevens RC. Structures of the CXCR4 Chemokine GPCR with Small-Molecule and Cyclic Peptide Antagonists. Science 2010;330:1066–71. [3] Perez DM. Structure-function of α1-adrenergic receptors. Biochem Pharmacol 2007;73:1051–62. [4] Huang Y, Wright CD, Merkwan CL, Baye NL, Liang Q, Simpson PC, O’Connell TD. An alpha1A-adrenergic-extracellualr signalregulated kinase signaling pathway in cardiac myocytes. Circulation 2007;115:763–72. [5] Woodcock EA, Du X-J, Reichelt ME, Graham RM. Cardiac α1-adrenergic drive in pathological remodelling. Cardiovasc Res 2008;77:452–62. [6] Schwinn DA. Novel role of α1-adrenerigc receptors in lower urinary tract symptoms. BJU Int 2000;86(Suppl. 2):11–22. [7] Docherty JR. Subtypes of functional alpha1-adrenoceptor. Cell Mol Life Sci 2010;67:405–17. [8] Cotecchia S. The α1-adrenergic receptors: diversity of signaling networks and regulation. J Recept Signal Transduct 2010;30:410–9. [9] Lorand L, Graham RM. Transglutaminases: crosslinking enzymes with pleiotropic functions. Nat Rev Mol Cell Biol 2003;4:140–56. [10] Finch A, Sarramegna V, Graham RM. Ligand binding, activation and agonist trafficking. In: Perez DM, editor. The Adrenergic Receptors in the 21st Century. : Humana Press Inc; 2006. p. 25–85. [11] Plascik MT, Kusiak JW, Barron KW. Alpha 1-adrenoceptor subtypes and the regulation of peripheral hemodynamics in the conscious rat. Eur J Pharmacol 1990;186:273–8.

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C H A P T E R

10 a2-Adrenergic Receptors Qin Wang

PHARMACOLOGY AND FUNCTIONS OF a2-AR SUBTYPES

to be critical for agonist-mediated lowering of blood pressure, sedation, anesthetic sparing, and working memory. The a2A-AR also appears to respond to endogenous catecholamines to suppress epileptogenesis (kindling) and depressive symptoms, the latter measured in mouse behavioral studies. The a2C-AR elicits depressive behaviors, behaving functionally the opposite of the a2A-AR subtype. The a2B-AR, in contrast to the a2A-AR, is involved in the vascular hypertensive effect of a2-AR agonists. Taken together, these findings suggest that subtype-selective agonists may be useful in manipulating one versus another adrenergic response in vivo. An even more refined therapeutic selectivity might be achieved using partial agonists of a2-ARs. It has been observed that agonist-mediated lowering of blood pressure can occur in mice heterozygous for the a2A-AR, whereas agonist-evoked sedation cannot. These data suggest that partial agonists with less than 50% intrinsic activity (efficacy) might be useful in a number of therapeutic settings, such as treatment of attention deficit/hyperactivity disorder (ADHD) and improvement of cognition in the elderly, where sedation as a side effect would undermine the therapeutic value of these agents. Interestingly, imidazoline compounds, for which there is evidence of a role in the central nervous system in regulating blood pressure, nonetheless appear to lower blood pressure via a2-ARs when administered peripherally. Thus, moxonidine and rilmenidine, developed as imidazoline I1-selective agents, are unable to lower blood pressure in D79N a2A-AR or a2A-AR knockout mice. Also of interest is the finding that these agents are partial agonists at the a2A-AR, which may explain their ability to lower blood pressure without evoking sedative side effects.

Alpha2-adrenergic receptors (a2-ARs) bind to their endogenous ligands, epinephrine and norepinephrine, and are blocked by the antagonist yohimbine. There are three subtypes of a2-AR, encoded by three independent, intronless genes. These subtypes are denoted as a2A- (human chromosome 10), a2B- (human chromosome 2), and a2C(human chromosome 4). Ligand selectivity for pharmacological agents does exist for the various a2-AR subtypes, although this selectivity has been observed principally in vitro, as not all of these ligands have been evaluated for their pharmacokinetic properties in vivo (see Table 10.1). The lack of truly specific ligands for each of the a2-AR subtypes, particularly antagonists, has prevented the unequivocal assignment of the differing a2-AR subtypes to various physiological responses. However, tools to genetically manipulate the mouse genome to create mutant (e.g. D79N a2A-AR) or null alleles of each of these subtypes shed light into subtype-specific functions of the different a2-AR subtypes, as outlined in Table 10.2. For example, both the a2A- and a2C-ARs are involved in suppression of catecholamine release from central neurons. However, the a2A-AR serves as the primary presynaptic autoreceptor. The a2C-AR is also critical for suppression of epinephrine release from the adrenal chromaffin cells. Moreover, the a2A-AR appears TABLE 10.1* Relative Selectivity for Ligands at the Three a2Adrenergic Receptor Subtypes Agonists

Shared: norepinephrine, epinephrine, aproclonidine

a2-AR SIGNALING AND TRAFFICKING

Selective: oxynetazoline (A  C  B), clonidine, guanabenz (A,C), UK 14304 (A  C), dexmedetomidine (A,B) Antagonists

All three subtypes of the a2-AR share the same signaling pathways in native cells: decrease in adenylyl cyclase activity, suppression of voltage-gated Ca currents, and activation of receptor-operated K currents and MAP kinase activity. In heterologous cells, the a2A-AR has been shown to activate phospholipase A2 and phospholipase D,

Shared: yohimbine, rauwolscine, phentolamine, idazoxan, RX 821002, atipamezole Selective: ARC 239 (B  C  A), prazosin (B,C  A), BRL 44408 (A  C), mianserin (A,B)

*Modified from Saunders and Limbrid, Pharmacology & Therapeutics 84:193–205, 1999.

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10. a2-ADRENERGIC RECEPTORS

TABLE 10.2*

Subtype-Specific Functions of a2-AR Derived From Gene-Targeted Mouse Models α2-AR Subtypes α2A-AR

Physiological Functions

α2B-AR

α2C-AR

CENTRAL EFFECTS Presynaptic inhibition of norepinephrine, dopamine, serotonin release

Xa

Hypotensive effect

X

Sedative effect

X

Antinociceptive effects of a2-adrenergic receptor agonist

X

Antinociceptive effects of moxonidine

X

Analgesic effect of nitrous oxide

X

X X

Anesthetic-sparing effects of dexmedetomidine

X

Adrenergic-opioid synergy in spinal antinociception

X

Anesthetic-sparing effects of dexmedetomidine

X

Hypothermic effects of dexmedetomidine

X

X

PERIPHERAL EFFECTS Inhibition of noradrenaline release from nerve terminal invaded into tissues

X

X

Inhibition of adrenaline release from adrenal cortex

X

Vasoconstriction, hypertensive effect

X

Salt-induced hypertension

X

Placenta angiogenesis

X

Platelet aggregation

X

X

Prevention of respiratory failure

X

BEHAVIOR EFFECTS Antiepileptogenic effect

X

Special working memory

X

Inhibition of startle responses

X

Antianxiety

X

Inhibition of locomotor stimulation of D-amphetamine

X

Latency to attack after isolation

X

*Above data summarized in Kable et al. J. Pharmacol. Exp. Ther. 2000;293(1):1–7; Brede et al. Biol Cell. 2004;96(5):343–8; and Knaus et al. Neurochem Int 2007;51(5):277–81. Also from Haubold et al. J Biol. Chem. 2010;285(44)34213–19. a Primary autoreceptor.

although these responses have yet to be observed in native target cells. Despite the similarity of the signaling pathways of the a2-AR subtypes, there are interesting differences in the trafficking itineraries of these receptors. Whereas the a2A- and a2B-AR subtypes are enriched on the surface at steady state, the a2C-AR is distributed between the surface and intracellular compartments. Upon agonist stimulation, both the surface a2A-AR and a2B-AR undergo internalization in an arrestin-dependent manner. a2A-AR internalization can be selectively promoted by different agonists such as clonidine and guanfacine, which, at least in part, account for distinct duration of signaling action of these drugs.

REGULATION BY INTERACTING PROTEINS a2-AR trafficking and signaling are tightly regulated by non-G protein interacting partners including GPCRs, kinases and scaffolding proteins such as arrestin and spinophilin (see Table 10.3). Some of these interactions serve to scaffold a2-ARs to particular cellular micro-compartments or to tether them to defined signaling molecules, while other receptor-protein interactions control a2-AR internalization and post-endocytotic sorting as well as the kinetics of a2AR-mediated signaling transduction. For example, whereas arrestin promotes a2A-AR and a2B-AR phosphorylation and

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

a2-AR POLYMORPHISMS

TABLE 10.3*

57

a2-AR-Interacting Proteins α2-AR Subtype(s)

Functional Role(s) Implicated

a2A

Precoupled to G proteins

a2C-AR

a2A

Attenuating agonist-induced a2A-AR to the a2A-AR phosphorylation, reducing arrestin binding

β1-AR

a2A

Altering ligand binding properties of β1-AR, leading to internalization of β1-AR in response to a2-agonist

β2-AR

a2C

Enhancing surface expression and internalization of a2C-AR, enhancing ERK activation by a2C-AR

μOR

a2A

Enhancing morphine-induced GTPγ S binding and ERK activation; however, no transactivation of G proteins, no interdependent internalization

δOR

a2A

Enhancing δOR-mediated neurite outgrowth

GRK2

a2A, a2B

Mediating agonist-induced phosphorylation and homologous desensitization

PKC

a2A

Mediating heterologous desensitization; Modulating constitutive activity of a2A-AR

14-3-3ζ

a2A, a2B, a2C

Function unknown, competed by phosphorylated Raf peptide

APLP1

a2A, a2B, a2C

Increasing a2A-AR-mediated inhibition of adenylyl cyclase activity

arrestin 2

a2B

Receptor affinity for arrestin 2 is lower than arrestin 3

arrestin 3

a2A, a2B

Stabilizing receptor phosphorylation, mediating endocytosis and desensitization, accelerating ERK signaling rate, enhancing sensitivity of in vivo response

arrestin 3

a2C

Mediating endocytosis

eIF-2B

a2A, a2B, a2C

Function unknown

Rab8

a2B

Promoting transport to cell surface

spinophilin

a2A, a2B

Stabilizing receptor at surface, attenuating phosphorylation, decelerating ERK signaling rate, decreasing in vivo response sensitivity

spinophilin

a2C

Function unknown

Uch-L1

a2A

Decreasing a2A-AR-mediated activation of ERK

Interacting Protein(s) GPCR DIMERIZATION HOMO-DIMERIZATION

HETERO-DIMERIZATION

KINASES

OTHER PROTEINS 14-3-3ζ

*Modified from Wang and Limbird, Biochem Pharm 2007;73(8):1135–1145; also from Weber et al. Cell Signal 2009;21(10):1513–21; Dong C et al. J Biol Chem 2010;285(26):20369–80.

internalization and accelerates the initial rate of signaling and desensitization, spinophilin blocks these activities by competing for GRK and arrestin interaction with these two a2-AR subtypes. Importantly, spinophilin antagonism of arrestin functions in regulating a2-ARs has in vivo relevance as manifested by reciprocal modulation of a2A-AR-evoked sedation by these two proteins in vivo.

a2-AR POLYMORPHISMS A number of individual human polymorphisms have been identified for each of the a2-AR subtypes. Some of

these have resulted in alterations in receptor density, G protein coupling, desensitization, or G protein receptor kinasemediated phosphorylation. Genetic association studies have linked a2-AR polymorphisms with a number of disease states and variations in drug responses in human populations. For example, a2A-AR polymorphisms have been linked to increased risk of ADHD, hypertension and type 2 diabetes. Additionally, genetic variants in all three subtypes have been associated with various forms of cardiovascular dysfunction. The study of a2-AR polymorphisms and their association with human diseases provides insights whose functional relevance can, in the future, be assessed by evaluating a variety of a2-AR in genetically engineered mouse

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10. a2-ADRENERGIC RECEPTORS

models that introduce each polymorphism into the genome via homologous recombination. Given the new understanding of the a2-AR subtypes in manipulating different physiological effects, refinement of this understanding with regard to partial agonists at the receptor, and the impact of receptor interactions with other proteins in target cells beyond G protein coupling, it would seem possible that molecular entities might be developed to interfere with particular a2-AR–protein interactions or with a2-AR activation of various pathways and evaluated for efficacy as therapeutic interventions in disease settings still lacking appropriate a2-AR-system regulation.

Acknowledgement I am grateful to Dr. Lee Limbird (Vanderbilt University) for her critical reading of this chapter and invaluable suggestions.

Further Reading Flordellis C, Manolis A, Scheinin M, Paris H. Clinical and pharmacological significance of alpha2-adrenoceptor polymorphisms in cardiovascular diseases. Int J Cardiol 2004;97:367–72.

Kable JW, Murrin LC, Bylund DB. In vivo gene modification elucidates subtype-specific functions of alpha(2)-adrenergic receptors. J Pharmacol Exp Ther 2000;293:1–7. Knaus AE, Muthig V, Schickinger S, Moura E, Beetz N, Gilsbach R, et al. a2-Adrenergic subtypes-unexpected functions for receptors and ligands derived from gene-targeted mouse models. Neurochem Int 2007;51:277–81. Lu R, Li Y, Zhang Y, Chen Y, Shields AD, Winder DG, et al. Epitopetagged receptor knock-in mice reveal that differential desensitization of alpha2-adrenergic responses is due to ligand-selective internalization. J Biol Chem 2009;284:13233–243. Moore CA, Milano SK, Benovic JL. Regulation of receptor trafficking by GRKs and arrestins. Annu Rev Physiol 2007;69:451–82. Rosengren AH, Jokubka R, Tojjar D, Granhall C, et al. Overexpression of alpha2A-adrenergic receptors contributes to type 2 diabetes. Science 2010;327:217–20. Saunders C, Limbird LE. Localization and trafficking of alpha2adrenergic receptor subtypes in cells and tissues. Pharmacol Ther 1999;84:193–205. Small KM, Liggett SB. Identification and functional characterization of alpha(2)-adrenoceptor polymorphisms. Trends Pharmacol Sci 2001;22:471–7. Tan CM, Wilson MH, MacMillan LB, Kobilka BK, Limbird LE. Heterozygous Alpha 2A-adrenergic receptor mice unveil unique therapeutic benefits of partial agonists. Proc Natl Acad Sci USA 2002;99:12471–12476. Wang Q, Zhao J, Brady AE, Feng J, Allen PB, Lefkowitz RJ, et al. Spinophilin Blocks Arrestin Actions in vitro and in vivo at G proteinCoupled Receptors. Science 2004;304:1940–4.

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C H A P T E R

11 β-Adrenergic Receptors C.Michael Stein INTRODUCTION

and norepinephrine, results from having different sub-types of β-ARs that are differentially expressed in particular tissues and that have differences in their affinity for agonists and signal transduction mechanisms, as well as the frequent concomitant presence of α-ARs – additional targets for epinephrine and norepinephrine. The three subtypes of β-ARs share approximately 60% sequence homology and are widely distributed in many cells and organs where they mediate several physiological responses (Figure 11.1; Table 11.1). Stimulation of β1-ARs mediates increased heart rate and cardiac contractility, β2-ARs mediate bronchodilation, vasodilation and presynaptic norepinephrine release, and β3-ARs mediate lipolysis and thermogenesis, relaxation of bladder, uterus and gut smooth muscle, and attenuated cardiac contractility (Table 11.1) [1,2].

There are three subtypes of β-adrenergic receptors (β1ARs, β2-ARs and β3-ARs) that mediate a wide range of physiological responses to the adrenergic agonists epinephrine and norepinephrine, and thus play an important role in regulating cardiovascular responses in health and disease (Table 11.1, Fig. 11.1). These receptors are also the targets of commonly used classes of drugs that block or stimulate signaling. β-Blockers are often used to treat hypertension and heart failure, and to control heart rate in atrial fibrillation, whereas β-agonists are used to treat asthma. There is a wide range of variation in responses to physiological and pharmacological stimulation or blockade of β-ARs; some of this variability is due to genetic variation among individuals in genes encoding β-ARs and their signal transduction proteins.

β-AR SUBTYPES AND THEIR DISTRIBUTION

β-ARs SIGNAL TRANSDUCTION β-ARs are 7-transmembrane spanning G proteincoupled receptors with an extracellular aminoterminus, an intracellular carboxyterminus, and three intracellular and three extracellular loops (Fig. 11.1). The binding of agonist

β-ARs mediate a wide range of responses (Table 11.1). The wide range of physiological responses mediated by just two primary endogenous adrenergic agonists, epinephrine TABLE 11.1 β Adrenergic Receptor Subtypes Subtype

Tissue Distribution

Functions

Gene Localization

Common Coding Variants

β1-AR

Heart Kidney Adipocytes

Positively inotropic and chronotropic Renin release Lipolysis

10q24-q26

Arg389Gly Ser49Gly

β2-AR

Lung and bronchial Vascular smooth muscle Heart Uterus Bladder Adipocytes Eye Liver Skeletal muscle Sympathetic terminal

Bronchodilation Vasodilation Positively inotropic and chronotropic Relaxation Relaxation Lipolysis Increase aqueous humor formation Glycogenolysis Glycogenolysis Norepinephrine release

5q31-q32

Gly16Arg Gln27Glu Thr164Ile

β3-AR

Adipocytes Uterus Bladder Heart

Lipolysis Relaxation Relaxation Negatively inotropic

8p12

Trp64Arg

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11. β-ADRENERGIC RECEPTORS

β1AR Ser or Gly β2AR Arg or Gly

NH2

49

β2AR Gln or Glu

16

27

agonist binding β1AR Gly or Arg 389

β3AR Trp or Arg

64

164

β2AR Thr or IIe

subsequent responses. This occurs by three mechanisms – uncoupling of the receptor from its G-protein, a process mediated by G protein-coupled receptor kinases (GRKs) that phosphorylate the receptor; movement of receptors away from the cell membrane, known as internalization or sequestration; and loss of receptors (downregulation) mediated by several mechanisms including decreased transcription and increased ubiquitination and degradation [4]. β3-ARs are activated at higher concentrations of catecholamines than β1- and β2-ARs; thus they are more likely to activate under conditions of high sympathetic activity and they are also less likely to desensitize [5].

HOOC

THERAPEUTIC IMPORTANCE OF β-ARs

G-protein coupling

desensitization

FIGURE 11.1 Structure of β adrenergic receptors.

to receptor is translated into physiological response by complex signal transduction mechanisms that involves heterotrimeric GTP-binding regulatory proteins (termed G-proteins) and their associated second messenger and effector systems. G-proteins are made up of three subunits, an α subunit that is important for recognition and effector functions, and β and γ subunits that are important for membrane localization but also have effector functions. The α subunit may be stimulatory, for example Gsα stimulates adenylate cyclase and increases cyclic AMP production, or inhibitory, for example Giα decreases cyclic AMP production. All three β-ARs couple with Gsα, but β2- and β3-ARs can also couple to Giα. Receptors exist in a conformational equilibrium between the inactive and active states and this equilibrium shifts with the binding of a ligand. There are three types of ligands: agonists that shift the equilibrium towards the active conformation; inverse agonists that favor the inactive conformation; and neutral antagonists that do not affect the equilibrium. When agonist binds to the receptor there is a conformational change that allows the intracellular part of the receptor to couple with a G-protein. For example, the activated β-AR forms a complex with Gsα and guanosine triphosphate (GTP) and stimulates the production of cyclic AMP, that in turn affects secondary effectors such as protein kinase A (PKA). The signal transduction mechanism is modulated by many other mechanisms, for example desensitization (see later), phosphodiesterases that break down cyclic AMP, and regulators of G-protein signaling (RGS) proteins that promote GTP hydrolysis [3].

RECEPTOR DESENSITIZATION Exposure of β-ARs to agonist and consequent activation triggers a process known as desensitization that acts to limit

From a therapeutic perspective β1-ARs in the heart (the target for β-blockers), and β2-ARs in the lungs (the target for β2 agonists), are most important. β-blockers comprise more than 20 different drugs that vary in their selectivity for β1and β2-ARs, as well as their lipophilicity, distribution, elimination, and intrinsic sympathomimetic effects. The primary target of β-blockers in heart disease is the β1-AR, and drugs that preferentially block this receptor are termed cardioselective β-blockers (e.g. atenolol) whereas non-selective agent such as propranolol block both β1- and β2-ARs. Initially β-blockers were considered to be absolutely contraindicated in patients with heart failure. This was because the marked increase in sympathetic activity that accompanies heart failure was considered to be a beneficial compensatory response; therefore, agents such as β-blockers that antagonized the effects of catecholamines on the heart were considered harmful. However, interventions to increase sympathetic activity increased mortality in heart failure, and in vitro studies showed that β-AR numbers were decreased and the receptors were uncoupled (i.e. desensitization had occurred) [6]. These observations led to the current view that excessive sympathetic activation is harmful in heart failure and clinical studies confirmed that careful treatment with β-blockers could improve cardiac function and decrease mortality [2]. Available β-blockers such as metoprolol, atenolol and propranolol have much lower affinity for the β3-AR than they do for β1-ARs; thus, specific β3-AR antagonists were developed. These agents resulted in weight loss in animal models of obesity but were generally not effective in human clinical trials. Their lack of efficacy has been ascribed to poor selectivity for the β3-AR, and also to the fact that β3ARs appear to play a smaller role in mediating lipolysis in human adipose tissue than they do in rodents [1]. Potential therapeutic clinical applications for future β3-AR antagonists are premature labor, irritable bowel syndrome and overactive bladder [1]. β-agonists, particularly β2-AR selective agonists, are used primarily to treat bronchoconstriction associated with asthma and other diseases of the airways. Hydrophilic drugs such as albuterol have a rapid onset and relatively short

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

GENETIC VARIABILITY IN β-ARS

duration of action (4–6 hours), whereas lipophilic drugs such as formoterol and salmeterol partition into the membrane and have a prolonged action [4]. β2-AR selective agonists are also used to prevent preterm labor. Agonists that are more selective for the β1-AR, for example dobutamine, are used for their ability to increase cardiac contractility.

GENETIC VARIABILITY IN β-ARS Many studies reported ethnic differences in response to adrenergic agonists and antagonists but until recently the potential mechanisms for these observations were unclear. The identification of common genetic variants in all the adrenergic receptors and studies showing that some of these variations affect function in vitro and in vivo led to work to establish the genetic contribution to variability in responses among individuals, and the clinical significance of such variation.

61

changes: Gly16Arg, Gln27Glu, and Thr164Ile. In vitro the Gly16 increased desensitization, Glu 27 decreased desensitization, and the Ile164 receptor had decreased responses to agonist. The clinical pharmacogenetic studies in this area are difficult to interpret because the variants at positions 16 and 27 are in linkage disequilibrium and there are many other non-coding variants that may affect function. Some studies suggest that patients with asthma who carry the Arg16 allele, particularly homozygotes, are less responsive to β-agonist bronchodilators, but there is little consensus regarding the clinical importance of these observations [7,8].

ADRB3 There is a one common coding region variant in the β3AR gene (ADRB3) that results in an amino acid change: Trp64Arg. However, studies performed in vitro and in vivo have been inconsistent in defining functional and clinical consequences of this variant [7,8].

ADRB1

Acknowledgement

There are two common polymorphisms in the β1AR gene (ADRB1) that result in amino acid changes: Arg389Gly and Ser49Gly. The Gly389 variant has impaired coupling to Gs and therefore less response to agonist, and the Gly49 variant has increased desensitization after exposure to agonist in vitro. Thus, the Ser49 and Arg389 alleles code for a β1-AR likely to be associated with greater response. Concordantly, studies in healthy subjects, and patients with hypertension, studied under strictly controlled conditions found that carriers of the Arg389 and the Ser49 alleles were more responsive to β-blockers. Similarly, patients with heart failure who carried the Gly389 allele were less responsive to a β-blocker with a smaller improvement in ejection fraction. In the Beta Blocker Evaluation of Survival Trial (BEST) there was an interaction between Arg389Gly genotype and outcomes of bucindolol treatment. Patients with the Gly389 allele did not benefit, but Arg389 homozygotes receiving bucindolol had improved outcomes. However, other heart failure and hypertension studies found no effect of ADRB1 genotypes on responses – thus, the clinical significance of genotype in β-blocker therapy is uncertain. Genetic variation in other adrenergic receptors that may also affect sympathetic responses (e.g. α2C-AR), and in GRKs and other proteins important in regulating adrenergic responses, may also play a role [2,7].

The previous version of this chapter was written by Dr Stephen B. Liggett and forms the framework for the present chapter.

References [1] Ursino MG, Vasina V, Raschi E, Crema F, De PF. The beta3adrenoceptor as a therapeutic target: current perspectives. Pharmacol Res 2009;59:221–34. [2] Dorn GW. Adrenergic signaling polymorphisms and their impact on cardiovascular disease. Physiol Rev 2010;90:1013–62. [3] McGraw DW, Liggett SB. Molecular mechanisms of beta2-adrenergic receptor function and regulation. Proc Am Thorac Soc 2005;2:292–6. [4] Johnson M. Molecular mechanisms of beta(2)-adrenergic receptor function, response, and regulation. J Allergy Clin Immunol 2006;117:18–24. [5] Dessy C, Balligand JL. Beta3-adrenergic receptors in cardiac and vascular tissues emerging concepts and therapeutic perspectives. Adv Pharmacol 2010;59:135–63. [6] Feldman DS, Carnes CA, Abraham WT, Bristow MR. Mechanisms of disease: beta-adrenergic receptors – alterations in signal transduction and pharmacogenomics in heart failure. Nat Clin Pract Cardiovasc Med 2005;2:475–83. [7] Kirstein SL, Insel PA. Autonomic nervous system pharmacogenomics: a progress report. Pharmacol Rev 2004;56:31–52. [8] Small KM, McGraw DW, Liggett SB. Pharmacology and physiology of human adrenergic receptor polymorphisms. Annu Rev Pharmacol Toxicol 2003;43:381–411.

ADRB2 There are three relatively common coding region variants in the β2-AR gene (ADRB2) that result in amino acid

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12 Dopaminergic Neurotransmission Graeme Eisenhofer, Heinz Reichmann

Dopamine is produced as a neurotransmitter in the central nervous system (CNS) and as an intermediate in the synthesis of norepinephrine in peripheral sympathetic and CNS noradrenergic nerves and of epinephrine in adrenal chromaffin cells. Dopamine is also produced as a catecholamine end product in chromaffin-like glomus cells of the hypoxia-sensing carotid bodies as well as in numerous other peripheral tissues. Synthesis of dopamine within dopaminergic neurons depends on the rate-limiting conversion of tyrosine to L-dihydroxyphenylanaline (L-dopa) by tyrosine hydroxylase, an enzyme confined to catecholamine-producing cells (Fig. 12.1). L-dopa is then converted to dopamine by L-aromatic amino acid decarboxylase, an enzyme with a wide tissue distribution. After this step, dopamine is translocated by vesicular monoamine transporters into storage vesicles from where the amine is available for release by exocytosis. Reuptake back into neurons by the cell membrane dopamine transporter represents the main mechanism for terminating the actions of dopamine at receptor sites, including D2 presynaptic autoreceptors, which act as a break on secretion. Complete inactivation requires metabolism by intraneuronal deamination to dihydroxyphenylacetic acid or extraneuronal O-methylation and deamination to homovanillic acid, the major metabolic end product of dopamine. Dopaminergic neurons within the brain are sparse, representing less than one thousandth of a percent of all CNS neurons. Nevertheless, dopaminergic neurotransmission is crucial for regulating numerous aspects of brain function. Deranged CNS dopaminergic neurotransmission also contributes to several devastating neurological and psychiatric disorders. Consequently the various components of dopamine neuronal systems, such as dopamine transporters, receptors and second messenger systems, represent important therapeutic targets that continue to receive intense scientific attention. CNS dopaminergic neurons can be broadly divided into two major groups: neurons in the mesencephalon project to the striatum and forebrain and subserve diverse functions from the control of movement to modulation of cognition, mood, attention, reward seeking behavior and learning; the other major group located in the arcuate nucleus of the

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hypothalamus has projections that release dopamine in the portal vessels with important neuroendocrine regulatory influences that include inhibition of the release of prolactin and thyroid stimulating hormone. In addition to these two major groups, there are also smaller groups in other CNS

FIGURE 12.1 Schematic representation of a slow synaptic transmission neuromodulatory dopaminergic nerve ending impinging on a dendritic spine of a fast synaptic transmission inhibitory GABAergic neuron. Abbreviations: DOPAC, dihydroxyphenylacetic acid; HVA, homovanillic acid; MTY, 3-methoxytyramine; TH, tyrosine hydroxylase; AADC, aromatic acid decarboxylase; MAO, monoamine oxidase; COMT, catechol-O-methyltransferase; DAT, dopamine transporter; EMT, extracellular monoamine transporter; D1, dopamine D1 receptor; D2 dopamine D2 receptor.

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regions, such as the dopaminergic neurons in the retina and olfactory bulb that have ultra-short projections that transmit signals within these neuronal centers for vision and smell. Mesencephalon dopaminergic neurons localized in the substantia nigra project mainly to the striatum to form the neostriatal pathway, whereas those of the ventral tegmental area project to cortical and limbic regions to form respective mesocortical and mesolimbic pathways. Neostriatal dopaminergic pathways regulate the initiation and maintenance of motor function, while mesocortical and mesolimbic pathways respectively regulate cognitive activity and reward seeking behavior. The importance of neostriatal dopaminergic pathways for control of extrapyrimidal (involuntary) movement is reflected by the loss of motor control when nigro-striatal neurons degenerate in Parkinson’s disease. Rather than acting as a classic excitatory or inhibitory neurotransmitter, dopamine released from midbrain projections acts as a neuromodulator that alters responses of target neurons to other neurons (Fig. 12.1). Such neuromodulatory influences occur by slow modes of synaptic transmission involving complex second messenger systems rather than the ligand-operated ion-channels that characterize fast synaptic transmission. Accordingly, accumulating evidence also indicates that signals from dopamine are generated in a volume mode of transmission in which the amine transmitter does not act rapidly within classical synapses, but instead diffuses from sites of release to act at receptors on more distant neurons. These include glutamatergic and γ-aminobutyric acidergic (GABA) neurons, which respectively release the fast acting excitatory and inhibitory neurotransmitters, glutamate and GABA. Awareness of the above concepts has important implications for understanding the biology and pharmacology of neurological diseases involving dopamine systems, including an appreciation of therapeutic principles underlying the efficacy of L-dopa and dopamine receptor agonists for treatment of the movement disorder characteristic of Parkinson’s disease. In this disorder it is the overall absence of dopamine, not just of the dopaminergic neurons, that disrupts the patterning of firing of intact CNS networks and cell assemblies crucial for control of contextdependent movement. Amelioration of the motor symptoms of Parkinson’s disease, therefore, does not require restoration of the degenerated neural connections, but can be largely achieved at least initially by correcting for the absence of the transmitter itself. Apart from Parkinson’s disease and related syndromes (e.g., multiple system atrophy, progressive supranuclear palsy, diffuse Lewy body disease), derangements in CNS dopamine neuronal systems also appear to be involved in numerous other neuropsychiatric conditions including Huntington's disease, drug addictions, depression, obsessive compulsive behavior, attention deficit/hyperactivity disorder and schizophrenia. The wide-ranging nature of these disorders illustrates the diverse functions subserved by CNS dopamine, with different pathologies and clinical manifestations explained by differences in the nature or

locations of the abnormalities of dopaminergic systems and associated dysfunctions in other neuronal systems. Elucidation of some of the underlying CNS dopaminergic abnormalities associated with the above neuropsychiatric conditions has been achieved through advances in brain imaging technology using positron emission tomographic (PET) or single photon emission computed tomographic (SPECT) imaging with ligands that target components of dopamine neuronal systems (e.g., dopamine receptors and transporters). Whereas the dysfunction of Parkinson’s disease is mainly presynaptic, the lesions associated with Parkinson plus syndromes involve both pre- and postsynaptic deficiencies, explaining why patients with the latter disorders are often unresponsive to therapy with L-dopa and dopamine receptor agonists. PET or SPECT imaging utilizing ligands specific for dopamine D1 and D2 receptors has indicated that in patients with Huntington’s disease there are reductions in striatal D1 and D2 receptor binding indicating postsynaptic dysfunction. In patients with attention-deficit/hyperactivity the disorder appears to involve presynaptic deficits associated with increases in dopamine transporter binding. Patients with chronic drug abuse problems appear to have lowered dopamine release and D2 receptor numbers in dorsal or ventral striatal regions. Furthermore, in these regions and particularly the nucleus accumbens reward centers, many of drugs of abuse – including cocaine, amphetamine, heroin, alcohol and nicotine – stimulate dopamine release, thereby reinforcing drug consumption. A central role of CNS dopaminergic systems in the pathogenesis of schizophrenia followed recognition in the 1950s that antipsychotic drugs interfere with brain dopamine function. Indeed even today, all neuroleptic drugs approved to treat schizophrenia appear to exert their antipsychotic effects through blocking the dopamine D2 receptor. In line with this, PET and SPECT imaging studies have indicated that mid-brain regions in schizophrenia are characterized by increased subcortical dopamine release and sensitivity to D2 receptor activation. Beyond the generally accepted view that psychotic episodes in schizophrenia are associated with a hyperdopaminergic state, the precise neuroanatomical sites and underlying alterations of CNS dopamine systems remain imprecisely elucidated. It is often presumed that production of dopamine within the brain accounts for most of the production of this catecholamine by the body. From measurements of the overflow of dopamine metabolites from the brain into jugular venous blood it is now clear, however, that the contribution of the brain to circulating levels and urinary excretion of dopamine metabolites represents less than 20% of the overall production of dopamine within the body (Fig. 12.2). Furthermore, almost all of the dopamine formed in sympathetic nerves and the adrenal medulla is converted to norepinephrine and epinephrine. Thus, most of the dopamine and dopamine metabolites in the circulation and excreted into urine are derived from other peripheral sources. However, in contrast to norepinephrine, which functions

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DoPAmINERgIC NEuRoTRANsmIssIoN

FIGURE 12.2 Schematic representation of the main sources of dopamine and the principal metabolites of dopamine in plasma and urine. The brain makes a relatively minor contribution, whereas dopamine synthesized in the gastrointestinal tract or derived from the diet contributes substantially to dopamine metabolites in the bloodstream and urine. This contrasts with the free dopamine excreted in urine, which is derived almost entirely from renal extraction of circulating L-dihydroxyphenylalanine and local decarboxylation to dopamine by L-aromatic amino acid decarboxylase. Abbreviations: HVA, homovanillic acid; DOPAC, dihydroxyphenylacetic acid; DA, Dopamine; DA-SO4, dopamine-sulfate; L-Dopa, L-dihydroxyphenylalanine; MTY; 3-methoxytyramine; MTY-SO4, 3-methoxytyramine-sulfate; AADC, aromatic acid decarboxylase.

as an important neurotransmitter in both the CNS and the peripheral sympathetic nervous system, evidence that dopamine acts as a neurotransmitter outside of the CNS is weak. Instead, available evidence indicates that dopamine in the periphery functions not as a neurotransmitter or circulating hormone, but rather as an autocrine or paracrine substance. Findings of increased plasma concentrations of L-dopa, dopamine and dopamine metabolites after consumption of foods indicate that dietary constituents also represent an important source of peripheral dopamine. Such food sources do not, however, account for the substantial amounts of dopamine produced in peripheral tissues outside of the digestive tract. Diet also does not account for findings in fasting individuals in which large arterial to portal venous increases in plasma concentrations of dopamine and its metabolites indicate substantial production of dopamine within mesenteric organs. These findings are consistent with morphological studies demonstrating the presence of cells in the gastrointestinal tract that contain dopamine and express components of dopamine signaling pathways, including catecholamine biosynthetic enzymes and specific dopamine receptors and transporters. In the

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stomach, tyrosine hydroxylase is expressed in epithelial cells, including acid secreting parietal cells. In the small intestine, cells of the lamina propria, including immune cells, also express tyrosine hydroxylase. The enzyme is additionally found in pancreatic exocrine cells. Dopamine and dopamine receptor agonists stimulate bicarbonate secretion and protect against ulcer formation, whereas dopamine antagonists augment secretion of gastric acid and promote ulcer development. Dopamine also appears to influence GI motility, sodium transport, and gastric and intestinal submucosal blood flow. In the pancreas, dopamine may modulate secretion of digestive enzymes and bicarbonate. Thus, dopamine appears to act in mesenteric organs as an enteric neuromodulator or paracrineautocrine substance. In the kidneys, dopamine is an established autocrine and/or paracrine effector substance contributing to the regulation of sodium excretion. Unlike other catecholamine systems, production of dopamine in the kidneys is largely independent of local synthesis of L-dopa by tyrosine hydroxylase. Instead, production of dopamine in the kidneys depends mainly on proximal tubular cell uptake of L-dopa from the circulation. The L-dopa is then converted to dopamine by L-aromatic amino acid decarboxylase, the activity of which is up-regulated by a high-salt diet and down-regulated by a low-salt diet. The presence of a renal dopamine paracrine-autocrine system explains the considerable amounts of free dopamine excreted in the urine. Most derives from renal uptake and decarboxylation of circulating L-dopa and reflects the plasma levels of this amino acid and the function of the renal dopamine paracrine/ autocrine system.

Further Reading Carlsson A, Waters N, Waters S, Carlsson ML. Network interactions in schizophrenia – therapeutic implications. Brain Res Brain Res Rev 2000;31:342–9. Carlsson A. Thirty years of dopamine research. Adv Neurol 1993;60:1–10. Eisenhofer G, Kopin IJ, Goldstein DS. Catecholamine metabolism: a contemporary view with implications for physiology and medicine. Pharmacol Rev 2004;56:331–49. Flagel SB, Clark JJ, Robinson TE, Mayo L, Czuj A, Willuhn I, et al. A selective role for dopamine in stimulus-reward learning. Nature 2011; 469:53–7. Girault JA, Greengard P. The neurobiology of dopamine signaling. Arch Neurol 2004;61:641–4. Nikolaus S, Antke C, Kley K, Poeppel TD, Hautzel H, Schmidt D, et al. Investigating the dopaminergic synapse in vivo. I. Molecular imaging studies in humans. Rev Neurosci 2007;18:439–72. Reichmann H. Long-term treatment with dopamine agonists in idiopathic Parkinson’s disease. J Neurol 2000;247(Suppl 4):IV/17–19. Rice ME, Cragg SJ. Dopamine spillover after quantal release: rethinking dopamine transmission in the nigrostriatal pathway. Brain Res Rev 2008;58:303–13. Sotnikova TD, Beaulieu JM, Gainetdinov RR, Caron MG. Molecular biology, pharmacology and functional role of the plasma membrane dopamine transporter. CNS Neurol Disord Drug Targets 2006;5:45–56. Volkow ND, Fowler JS, Wang GJ, Swanson JM, Telang F. Dopamine in drug abuse and addiction: results of imaging studies and treatment implications. Arch Neurol 2007;64:1575–9.

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13 Dopamine Receptors Sean M. Peterson, Nikhil Urs, Marc G. Caron

INTRODUCTION

receptors, both temporally and spatially, and is cataloged online at GENSAT [1].

Dopamine (DA) is one of the major monoamine neurotransmitters in the mammalian brain. It is generated by hydroxylation and decarboxylation of the amino acid tyrosine, and further metabolized into epinephrine and norepinephrine (collectively referred to as catecholamines). The role of DA in brain function was first proposed by Nobel Laureate, Arvid Carlsson, whereas Oleh Hornykeiwicz and colleagues first demonstrated a DA deficit in the brain of Parkinson’s patients. DA has been shown to regulate a variety of functions such as movement, reward, cognition and emotion. Dysfunction of DA neurotransmission has been implicated in numerous other disease states such as addiction, schizophrenia, ADHD and Tourette’s syndrome. These multiple effects of DA are mediated through its ability to bind to five distinct G protein-coupled receptors (GPCRs) that belong to two subfamilies, the D1-like (comprised of D1R and D5R) and the D2-like (comprised of D2R, D3R, and D4R) receptor families. These families are distinguishable not only by their unique pharmacology but also by their gene structure, expression patterns, protein structure, and signal transduction pathways. A large number of DA receptor (DAR) studies have focused on the receptors in the central nervous system (CNS); however the basic principles apply to both the CNS and autonomic nervous system (ANS).

Autonomic Nervous System DA neurons from the hypothalamus project to the anterior pituitary gland where D1 and D2 family receptors are expressed leading to an inhibition of prolactin release. All DAR subtypes are expressed in the kidney, blood vessels and heart. In the kidney, DARs regulate renin release as well as fluid and sodium reabsorption.

DOPAMINE RECEPTOR STRUCTURE Gene Structure Two distinct receptor families have evolved to bind DA. The D1 family is comprised of D1R, D5R, and the D5R pseudogenes, D5ψ1 and D5ψ2, which are not expressed. The D1 family is intronless in the protein coding region, as are approximately 50% of all GPCRs. The D2 family, which includes D2R, D3R, and D4R, has a more complex gene structure than the D1 family. The D2R gene, DRD2, contains six introns and is alternatively spliced to generate two functional proteins: D2LR (the long isoform) and D2SR (the short isoform lacking a 29 amino acid exon in the third intracellular loop of the receptor). D2SR is expressed predominantly on DA neurons, and functions as an autoreceptor to regulate extracellular levels of DA and modulate neuronal firing, while D2LR is expressed predominantly postsynaptically. The D3R gene, DRD3, contains five introns in the protein coding region and is alternatively spliced, however, DRD3 isoforms are not as well characterized, which may be due to nonsense mediated decay of mRNA products. The D4R gene, DRD4, contains three introns in the protein coding region, and contains a variable number tandem repeat (VNTR) of 48 nucleotides in exon 3 of the gene (which occurs in the intracellular loop 3 of the receptor). The VNTR can be 2–11 repeats, however, the four repeats (4R) is by far the most common, while 7R and 2R occur less frequently.

DISTRIBUTION AND EXPRESSION OF DOPAMINE RECEPTORS Central Nervous System Dopamine receptor distribution in the brain has been extensively studied. In the mammalian brain D1Rs and D2Rs are the most highly expressed receptors of all the five DARs. D1R and D2R expression is highest in areas of DA neuron innervation, whereas D3R, D4R and D5R expression is more specialized. The advent of BAC (bacterial artificial chromosome) recombineering technology has led to advances in clarifying the distribution patterns of these

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The DRD4 7R allele has been proposed to be a contributing factor to the development of ADHD, and has undergone strong positive selection throughout all of modern human history [2].

Receptor Structure Dopamine receptors belong to the superfamily of GPCRs and are all considered to be in the Rhodopsin-like Class A family of 7-transmembrane receptors based on sequence homology and function. For all the five DARs the N-terminus and extracellular loops are glycosylated and cysteine residues between the loops form disulfide bonds. The intracellular loops interact with G proteins, whereas these loops and C-terminal tails are phosphorylated by G protein-coupled receptor kinases (GRKs) and interact with β-arrestins and other kinases and signaling molecules (Table 13.1; Fig. 13.1). The D2 family of DARs have an especially long intracellular loop 3 (IC3) and a short C-terminal tail, while the D1 family has a relatively short IC3 and longer C-terminal tail. Recently, the crystal structure of D3R was solved [3]

TABLE 13.1 Dopamine Receptor signal Transduction Pathways

Receptor

Signaling Molecule

Downstream Effector

Second Messenger System

D1-like

Gαs

AC activation

Increased cAMP

D2-like

Gαi

AC inhibition

Decreased cAMP

D2R

Gαz

AC inhibition

Decreased cAMP

D2R/D1R and D2R/D5R dimers

Gαq

PLC activation

Increased IP3, DAG, and Ca2

D2R

Gβγ

GIRK

K

D1R

β-arrestin

ERK

N/A

D2R

β-arrestin

Akt/GSK3

N/A

Receptors are designated as their family (D1- or D2-like) or their specific receptor subtype. Canonical G protein signaling is compared and contrasted to non-canonical β-arrestin signaling. The details and relationship between these signal transduction pathways are further illustrated in Figure 13.1.

FIGURE 13.1 Relationship of dopamine receptor signal transduction pathways. Dopamine binds to dopamine receptors (DARs) to cause the exchange of GTP for GDP at the Gα subunit and the dissociation of Gβγ, which goes on to interact with effectors like ion channels. Gα hydrolyzes GTP (with the help of RGS proteins in the case of Gαi and Gαq family members) and then re-associates with Gβγ. In addition, binding of dopamine to DARs causes their phosphorylation and the recruitment of β-arrestin, which mediates desensitization of G protein signaling as well as internalization and recycling of competent DARs to the plasma membrane. The DAR/arrestin complex can initiate G protein independent receptor mediated signaling in its own right. Functional selectivity refers to agonists that are able to activate either G protein or β-arrestin pathways to a much greater extent than the other. The intracellular loop 3 and C-terminus of the receptor is represented as a dotted line because of structural differences between D1- and D2-family DARs (see text).

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DoPAmInE RECEPToR FunCTIon: sIgnAl TRAnsDuCTIon PATHwAys

in complex with a selective D3R antagonist. This insight will undoubtedly provide an avenue for the design of highly potent D3R ligands. The importance of DAR pharmacological agents will be discussed in more detail in later sections.

DOPAMINE RECEPTOR FUNCTION: SIGNAL TRANSDUCTION PATHWAYS G protein-coupled receptors have long been known for transducing signals by acting as guanine nucleotide exchange factors for heterotrimeric G proteins, however, recent work has shown that most GPCRs signal through noncanonical pathways both in vitro and in vivo. Dopamine receptors are no exception and the following section summarizes both canonical and noncanonical DAR signal transduction pathways. Figure 13.1 provides a summary of the general relationship between these signaling pathways, while Table 13.1 provides specific details of the important DAR signaling molecules.

G Protein Dependent Signaling The D1 family of DARs activates members of the Gαs family, which are so named because they stimulate adenylate cyclase (AC) leading to the production of adenosine 3,5-cyclic monophosphate (cAMP) as a second messenger. The D2 family of DARs couples to both the pertussis sensitive Gαi and the pertussis insensitive Gαz families resulting in an inhibition of AC and lower cAMP. The D2 family is also known to signal through Gβγ, which activates G protein-coupled inward rectifying potassium channels, phospholipase Cβ, and other proteins.

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molecules and promote signal transduction [6]. Recent studies from our laboratory have shown a role for β-arrestin dependent signaling through both D1R and D2R. D1Rs have been shown to promote β-arrestin dependent ERK signaling upon morphine-stimulated DA release and this signaling pathway regulates morphine/DA-induced psychomotor activation but not its rewarding effects [7]. D2Rs have been shown to promote β-arrestin dependent Akt and GSK3 signaling and regulate several DA sensitive behaviors in mice that can be modeled as endophenotypes of schizophrenia [8].

Oligomerization Higher order GPCR signaling structures have long been hypothesized to exist, however there remains some controversy over the biological significance of such structures. Evidence for oligomers (mostly dimers) between the D1 family and the D2 family has been provided from both in vitro and in vivo approaches. D2R was shown to interact with both D1R or D5R the consequence of which causes the coupling of the receptors to switch from Gαs or Gαi to a different family of G proteins, Gαq, which actives phospholipase C to cause the release of IP3 and Ca2 [9]. Receptor oligomerization adds a layer of complexity to an already complex signaling model, and new investigational tools have and will continue to illuminate these intricacies.

Dopamine Receptor Pharmacology Dopamine’s role in the regulation of complex functions in both the CNS and ANS makes DARs key targets in developing therapeutics for many diseases associated with DA function. Extensive work has been done in characterizing DAR agonists and antagonists.

Signal Regulation

Ligand Specificity

Regulator of G protein signaling (RGS) proteins function to accelerate GTP hydrolysis by the Gα subunit of G proteins, thereby attenuating GPCR signaling. D2Rs are regulated by RGS9, which has been implicated in Parkinson’s disease [4]. Receptor regulation is also accomplished through phosphorylation of the intracellular residues of the receptors by GRKs, which stabilize the receptor in a conformation conducive to β-arrestin binding. All five DARs interact with GRK2, GRK3, GRK5 and GRK6 as well as β-arrestin 1 and 2. Upon binding, β-arrestins desensitize GPCR signaling and induce internalization of receptors by interacting with clathrin-coated pits [5].

Many ligands have been shown to differentiate between the D1 and D2 family of receptors. Classically, the specificity of agonists and antagonists can be used to delineate the contribution of each receptor family in in vitro and in vivo studies. Selective ligands are also used as therapeutic agents in conditions like Parkinsonism and schizophrenia.

β-Arrestin Dependent Signaling In addition to its desensitizing and internalizing functions, recent work has shown a role for β-arrestin dependent signaling through its ability to scaffold signaling

Functional Selectivity The significance of arrestin dependent signaling for GPCRs may be many fold [7]. First, arrestin dependent signaling is usually more persistent than G protein signaling. Second, it has been shown that for certain GPCRs a given compound can act as an agonist at one signaling pathway and an antagonist at the other or vice versa. Moreover, examples are beginning to accumulate demonstrating that these two signaling modes can sub-serve distinct biological functions mediated by the same GPCR.

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Interestingly, all clinically effective antipsychotics for the treatment of schizophrenia interact with D2R. Whereas antipsychotics have a mixed agonist/antagonist profile at D2R/G protein signaling, they appear to behave uniformly as antagonists at the D2R/β-arrestin interactions [10]. This principle is commonly referred to as functional selectivity or biased agonism/antagonism and should become an important element in the development of newer, more selective therapeutic agents.

References [1] The gene expression nervous system atlas (GENSAT) project. New York, NY: The Rockefeller University. [2] Wang E, Ding YC, Flodman P, Kidd JR, Kidd KK, Grady DL, et al. The genetic architecture of selection at the human dopamine receptor D4 (DRD4) gene locus. Am J Hum Genet 2004;74(5):931–44. [3] Chien EY, Liu W, Zhao Q, Katritch V, Han GW, Hanson MA, et al. Structure of the human dopamine d3 receptor in complex with a d2/d3 selective antagonist. Science 2010;330(6007):1091–5. [4] Gold SJ, Hoang CV, Potts BW, Porras G, Pioli E, Kim KW, et al. RGS9-2 negatively modulates L-3,4-dihydroxyphenylalanineinduced dyskinesia in experimental Parkinson’s disease. J Neurosci 2007;27(52):14338–48.

[5] Laporte SA, Oakley RH, Zhang J, Holt JA, Ferguson SS, Caron MG, et al. The beta2-adrenergic receptor/beta-arrestin complex recruits the clathrin adaptor AP-2 during endocytosis. Proc Natl Acad Sci USA 1999;96(7):3712–7. [6] Shenoy SK, Drake MT, Nelson CD, Houtz DA, Xiao K, Madabushi S, et al. beta-arrestin-dependent, G protein-independent ERK1/2 activation by the beta2 adrenergic receptor. J Biol Chem 2006;281(2):1261–73. [7] Urs NM, Daigle TL, Caron MGA. Dopamine D1 Receptordependent beta-arrestin signaling complex potentially regulates morphine-induced psychomotor activation but not reward in mice. Neuropsychopharmacology 2011; 36(3):551–8. [8] Beaulieu JM, Sotnikova TD, Marion S, Lefkowitz RJ, Gainetdinov RR, Caron MG. An Akt/beta-arrestin 2/PP2A signaling complex mediates dopaminergic neurotransmission and behavior. Cell 2005;122(2):261–73. [9] So CH, Verma V, Alijaniaram M, Cheng R, Rashid AJ, O’Dowd BF, et al. Calcium signaling by dopamine D5 receptor and D5-D2 receptor hetero-oligomers occurs by a mechanism distinct from that for dopamine D1-D2 receptor hetero-oligomers. Mol Pharmacol 2009;75(4):843–54. [10] Masri B, Salahpour A, Didriksen M, Ghisi V, Beaulieu JM, Gainetdinov RR, et al. Antagonism of dopamine D2 receptor/betaarrestin 2 interaction is a common property of clinically effective antipsychotics. Proc Natl Acad Sci USA 2008;105(36):13656–61.

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14 Cholinergic Neurotransmission Brett A. English, Carrie K. Jones

ACETYLCHOLINE BIOSYNTHESIS AND METABOLISM

enzymatic inactivation by the enzyme acetylcholinesterase (AChE), which hydrolyzes ACh into acetate and choline. AChE operates at near diffusion-limited rates, thus this enzyme is not rate limiting in overall ACh homeostasis. The byproduct of ACh hydrolysis, choline, is then recycled by the presynaptic cholinergic terminal in a carrier-mediated mechanism of high-affinity choline uptake (HACU) process, mediated by the high-affinity choline transporter (CHT) and which appears to be the critical step in modulating the rate and extent of ACh production. Within cholinergic nerve terminals, choline is required for the biosynthesis of ACh since these neurons cannot synthesize de novo, hence the requirement for a HACU process. Recent studies in transgenic mice exhibiting a genetic reduction in CHT demonstrated age-related reductions in cardiac function and histologic changes consistent with heart failure.

Acetylcholine (ACh), one of the first identified neurotransmitters, was originally termed vagusstoff due to its actions in supporting vagal slowing of heart rate (HR), and has been the focus of intense research for decades. ACh has been shown to regulate many physiologic functions within the central nervous system (CNS) modulating diverse functions including cognition, attention and arousal. Within the autonomic nervous system (ANS), ACh serves as the principal neurotransmitter mediating fast synaptic neurotransmission at the preganglionic junction by both the sympathetic (SNS) and parasympathetic (PNS) nervous systems. The PNS (via the vagus nerve) uses ACh as its primary postganglionic neurotransmitter modulating a number of physiologic processes including heart rate, gastrointestinal motility, exocrine gland secretions and smooth muscle tone (Tables 14.1 and 14.2). In addition to mediating a number of peripheral autonomic effects, recent experimental evidence has demonstrated a role of the PNS (vagus nerve) in mediating cytokine production in response to inflammation. The availability of ACh for cholinergic transmission involves a highly co-ordinated process of ACh synthesis, vesicular packaging, vesicular release, hydrolysis and reuptake into the presynaptic nerve terminal. Within the cholinergic presynaptic terminal, the enzyme choline acetyltransferase (ChAT) synthesizes ACh from the precursors choline and acetyl coenzyme-A (acetyl-CoA). ChAT is not believed to serve as the rate-limited step in the biosynthesis of ACh as the presynaptic concentrations of choline are much lower than the Km (affinity) for ChAT. Upon synthesis, ACh is packaged into synaptic vesicles by the vesicular ACh transporter (VAChT), and released into the synaptic cleft upon depolarization of the neuron by an action potential mediated by voltage-sensitive Ca2channels. Once released into the synaptic cleft, ACh interacts with either nicotinic acetylcholine receptors (nAChRs) or muscarinic acetylcholine receptors (mAChRs). ACh then rapidly dissociates from its receptor and undergoes

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00014-7

ACETYLCHOLINE RECEPTORS The postsynaptic effects of ACh neurotransmission are mediated by two major types of acetylcholine receptors. At the preganglionic junction, both the SNS and PNS use ACh to activate nicotinic acetylcholine receptors (nAChRs) on the postganglionic neuron. The postganglionic effects of ACh by the PNS occur by activating a number of postsynaptic muscarinic acetylcholine receptors (mAChRs) expressed in several target organs (Table 14.1). The classification of the different acetylcholine receptor subtypes is primarily established based upon binding of specific ligands, their respective effector-coupling systems and their primary amino-acid sequence homology. Within the muscarinic acetylcholine receptor family, there are five receptor subtypes that have been identified, termed M1-M5), each with unique tissue distribution and pharmacologic characteristics (Table 14.2). All mAChRs subtypes are monomeric proteins with seven transmembrane spanning domains and mediate their effects by coupling to G-proteins. M1, M3 and M5 receptors couple to Gq/11 and increase phospholipase C (PLC) activity subsequently increasing cytosolic concentrations of

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14. CHolINERgIC NEuRoTRANsmIssIoN

TABLE 14.1 Cholinergic Pharmacology Target

Localization

Effect

Mechanism

Agonist

Antagonist

Nicotinic (N)N

Autonomic ganglia, CNS

Fast depolarization of postganglionic neuron

↑ Na/K channel conductance

Succinylcholine

Trimethaphan

Nicotinic (N)M

Neuromuscular junction (NMJ)

Motor end-plate depolarization, muscle contraction

↑ Na/K channel conductance

Succinylcholine

Tubocurarine

Muscarinic (M1)

CNS, autonomic ganglia

Attention, memory, arousal

Gq/11; ↑ PLC, PKC

Oxotremorine, cevimeline, xanomeline

Scopolamine, pirenzepine

Muscarinic (M3)

Smooth muscle, exocrine glands

Smooth muscle contraction, increased glandular secretions

Gq/11; ↑ PLC, PKC

Cevimeline, pilocarpine

Atropine, solifenacin

Muscarinic (M5)

CNS

?

Gq/11; ↑ PLC, PKC

Sabcomeline

Atropine

Muscarinic (M2)

CNS, heart (SA and AV node)

Reduced HR, AV nodal conduction

Gi; βγ, ↓ AC and ↑ K channel conductance

Carbachol, pilocarpine

Scopolamine, atropine

Muscarinic (M4)

CNS

Cognition

Gi; βγ, ↓ AC and ↑ Kchannel conductance

Xanomeline

Atropine, pirenzepine

Choline transporter (CHT)

Presynaptic; CNS, autonomic, lung

Rate-limiting step for synthesis of ACh

None Na/K-coupled high affinity choline transport

Choline acetyltransferase (ChAT)

Presynaptic terminal

Synthesis of ACh from choline & acetylCoA

AcetylCoA  choline → ACh

None

None

Vesicular acetylcholine transporter (VAChT)

Presynaptic vesicles

Storage of ACh in presynpatic vesicles

H-coupled

None

Vesamicol

Postsynaptic

Hydrolysis of ACh

ACh→choline  acetate

None

Physostigmine, donepezil

RECEPTOR



SYNTHESIS/STORAGE Hemicholinium-3

METABOLISM Acetylcholinesterase (AChE)

inositol-triphosphate (IP3) and diacylglycerol (DAG). Increased IP3 and DAG results in increased release of intracellular Ca2 from endoplasmic reticulum stores and activation of protein kinase C (PKC) respectively. M1 receptors are primarily expressed within the CNS and mediate a number of cognitive functions, while M3 receptors are involved in exocrine gland secretion and smooth muscle function in the PNS. M5 receptors are expressed only within the CNS where they serve to modulate midbrain dopamine release and cerebral vasculature tone. M2 and M4 receptors are coupled to Gi, which results in inhibition of adenylyl cyclase (AC) and activation of inwardly rectifying K-channels (IKACh) and suppression of voltage-gated Ca2-channels. These actions of G-protein coupling to Gi partly explain the cardiovascular effects of M2 activation in the heart via IKACh. Additionally, studies conducted in M2 knockout mice show increased sensitivity to the β-agonist, isoproterenol, developing abnormal ventricular functioning consistent with heart failure. Within the CNS, M2 and M4 receptors provide the major autoreceptor function for control of Ach release. Nicotinic acetylcholine receptors are also divided into nicotinic receptors that mediate neuromuscular junction

activation (NM) or neuronal activation (NN) at the autonomic ganglia and within the CNS. However, unlike muscarinic receptors, nAChRs are ligand-gated ion channels. Upon binding of Ach, these receptors undergo conformational changes that permit the rapid influx of Na with subsequent cellular depolarization. These distinctions are important pharmacologically as nicotinic receptors are an assembly of five heterologous subunits (α1–9 and β1–4) arranged symmetrically around a central ionic selectivity pore, each with different subunit arrangement and pharmacologic binding properties (nicotinic neurotransmission is discussed in detail in Chapter 15). While nAChRs do not directly affect peripheral organ systems, they mediate ganglionic transmission and can facilitate neurotransmission of postganglionic nerve fibers.

CHOLINERGIC PHARMACOLOGIC AGENTS Cholinomimetic drugs can be classified into three categories. Direct-acting cholinomimetics, indirect-acting agents and allosteric modulating compounds. Direct-acting agents bind to and activate either nAChRs or mAChRs; for

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

CHolINERgIC PHARmACologIC AgENTs

TABLE 14.2 Responses of Effectors organs to Cholinergic Transmission of the Autonomic Nervous system Effector Organ

Receptor

Effect

Sinoatrial node

M2

Bradycardia

Atrium

M2, M3

↓ lusitropy

Atrioventricular node

M2

↓ conduction velocity

Arteriole

M2

Vasodilation (via release of nitric oxide, NO)

HEART

GASTROINTESTINAL/UROLOGIC GI Motility

M2, M3

Increased

GI secretion

M3

↑ secretions

Gallbladder

M2

Contraction

Urinary detrusor

M2, M3

Contraction

Urinary sphincter

M2

Relaxation

Penis

M

Erection

M3, M1

Constriction; ↑ secretions

Sweat glands

M3

Diaphoresis

Salivary glands

M3

↑ secretions

Lacrimal glands

M3

↑ secretions

Nasopharyngeal glands

M3

↑ secretions

Iris (pupillae sphincter)

M3

Contraction (miosis)

Ciliary muscle

M3

Contraction (accommodation)

RESPIRATORY Bronchial muscle GLANDULAR

OCULAR

73

(iii) phosphoric acid organic derivatives (e.g. organophosphates). Organophosphates are considered “irreversible” inhibitors, while the other chemical classes of AChEIs are “reversible” acting. Recently, several novel cholinomimetic compounds have been identified that function as positive allosteric modulators (PAMs) or allosteric agonists instead of traditional orthosteric agonists at different mAChRs. Allosteric modulators and agonists of mAChRs do not bind to the orthosteric binding site of Ach but rather interact with topographically distinct binding sites on the receptor that are separate from the orthosteric sites. Because these allosteric modulators and agonists interact at less highly conserved regions of the mAChR, these compounds offer a tremendous advantage in conferring a high degree of subtype selectivity. In addition, since PAMs potentiate the effects of Ach and have no effect in the absence of the endogenous ligand, the physiological signal and transduction is preserved resulting in less receptor desensitization and development of tolerance. A number of mAChR selective PAMs, including the selective M1 PAM BQCA, are being investigated in several preclinical models of cognitive disorders associated with schizophrenia and Alzheimer’s disease. Similarly, cholinergic receptor antagonists are categorized based upon their acetylcholine receptor selectivity. There are few nAChR antagonists used clinically and these consist primarily of the non-depolarizing ganglionic blockers (e.g. hexamethonium) and the depolarizing blocker (e.g. succinylcholine). The prototypical muscarinic antagonist, scopolamine is a non-selective antagonist at all five mAChR subtypes resulting in a reversible blockade that is surmountable by increasing concentrations of ACh. Other muscarinic antagonists display relative selectivity for specific mAChR subtypes, but exhibit dose-dependent effects (e.g. pirenzepine, M1; oxybutynin, M3). Since muscarinic receptors are constitutively active, muscarinic antagonists shift the receptor to an inactive state preventing actions of IP3/DAG and AC.

Further Reading example chemical esters of choline such as carbachol or alkaloid compounds, including pilocarpine. Many of these compounds exhibit preferential selectivity for either nicotinic or muscarinic receptors. However, in the case of the mAChRs, subtype selective compounds within the mAChRs have been difficult to identify and develop due to the high conservation of the orthosteric binding site of ACh across M1–M5. Lack of subtype selectivity of orthosteric compound, especially for the mAChRs, has resulted in a number of adverse side effects due to activation of peripheral mAChRs. Indirect-acting agents increase synaptic concentrations of ACh by inhibiting the hydrolytic metabolism by AChE. While these agents all work by inhibition of AChE, they exhibit significant differences in their chemical properties and pharmacokinetics. There are three chemical groups of AChE inhibitors: (i) quaternary ammonium alcohols (e.g. edrophonium); (ii) carbamic acid esters (e.g. neostigmine);

Dhein S, Van Koppen CJ, Brodde OE. Muscarinic receptors in the mammalian heart. Pharmacol Res 2001;44(3):161–82. Digby GJ, Shirey JK, Conn PJ. Allosteric activators of muscarinic receptors for treatment of CNS disorders. Mol Biosyst 2010;6(8):1345–54. English BA, Appalsamy M, Diedrich A, Ruggiero AM, Lund D, Wright J, Keller NR, Louderback KM, Robertson D, Blakely RD. Tachycardia, reduced vagal capacity and age-dependent ventricular dysfunction arising from diminished expression of the presynaptic choline transporter. Am J Physiol Heart Circ Physiol 2010;299(3):H799–810. Ferguson SM, Blakely RD. The choline transporter resurfaces: new roles for synaptic vesicles. Mol Interv 2004;4:22–7. LaCroix C, Freeling J, Giles A, Wess J, Li YF. Deficiency of M2 muscarinic acetylcholine receptors increases susceptibility of ventricular function to chronic adrenergic stress. Am J Physiol Heart Circ Physiol 2008;294:H810–820. Langmead CJ, Watson J, Reavill C. Muscarinic acetylcholine receptors as CNS drug targets. Pharmacol Therap 2008;117:232–43. Olshanksy B, Sabbah HN, Hauptman PJ, Colucci WS. Parasympathetic nervous system and heart failure: pathophysiology and potential implications for therapy. Circulation 2008;118(8):863–71.

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Rosas-Ballina M, Tracey KJ. Cholinergic control of inflammation. J Intern Med 2009;265:663–79. Shirey JK, Brady AE, Jones PJ, Davis AA, Bridges TM, et al. A selective allosteric potentiator of the M1 muscarinic acetylcholine receptor increases activity of medial prefrontal cortical neurons and restores impairments in reversal learning. J Neurosci 2009;29(45):14271–14286.

Westfall TC, Westfall DP. Neurotransmission: the autonomic and somatic motor nervous system. In: Brunton LL, Lazo JS, Parker KL, editors. Goodman and Gilman’s the pharmacological basis of therapeutics. New York: McGraw-Hill; 2006. p. 137–83.

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C H A P T E R

15 Acetylcholine and Muscarinic Receptors Joan Heller Brown, Nora Laiken The synthesis of acetylcholine (ACh) is catalyzed by choline acetyltransferase, a soluble cytoplasmic enzyme that transfers an acetyl group from acetyl coenzyme A to choline. The activity of choline acetyltransferase is much greater than the maximal rate at which ACh synthesis occurs, and inhibitors of this enzyme have little effect on ACh levels in cholinergic nerve terminals. The rate-limiting step in ACh synthesis is the transport of choline into the nerve terminal by the high-affinity choline transporter (CHT1), the activity of which is regulated in response to neuronal activity. Following its synthesis, ACh is transported into synaptic vesicles by the vesicular ACh transporter (VAChT). The depolarization of the cholinergic nerve terminal by an action potential activates voltage-gated calcium channels; the resulting calcium influx initiates the release of ACh into the junctional space by exocytosis. ACh release can be blocked by botulinum toxin, the etiologic agent in botulism. Botulinum toxin has been used to treat a variety of movement disorders and other conditions, both cosmetic (e.g., glabellar frown lines [facial wrinkles]) and pathological (e.g., strabismus [crossed eyes]); injected locally, the toxin can inhibit ACh release for as long as 3 to 4 months. The ACh released into the synapse associates with postsynaptic cholinergic receptors, triggering various physiological responses. The actions of ACh are terminated by its rapid hydrolysis into choline and acetic acid by acetylcholinesterase (AChE), which is found in the junctional space at all cholinergic junctions; thus, responses mediated by ACh tend to be transient and localized. The choline liberated by AChE can be taken back up into the nerve terminal by CHT1 and resynthesized into ACh. While not important for the hydrolysis of endogenous ACh, a nonspecific cholinesterase (pseudocholinesterase or butyrylcholinesterase) is present in plasma and some organs and important for metabolism of some drugs. Drugs that normally are hydrolyzed by pseudocholinesterase, such as succinylcholine (a neuromuscular blocking agent used in anesthesiology to produce skeletal muscle paralysis), are poorly metabolized by this variant enzyme; succinylcholine can produce a prolonged paralysis in affected patients. Cholinesterases are discussed in detail in Chapter 132 of this Primer. Cholinergic neurotransmission and drugs that affect it are further discussed in Chapter 14 and summarized in Table 15.1.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00015-9

TABLE 15.1 Cholinergic Neurotransmission: Drug Mechanisms Cholinergic neurotransmission can be modified at several sites: (a)

Precursor transport blockade

Hemicholinium (blocks CHT1)

(b)

Choline acetyltransferase inhibition

No clinical example

(c)

Promote transmitter release

Black widow spider venom (latrotoxin)

(d)

Prevent transmitter release

Botulinum toxin

(e)

Storage

Vesamicol (blocks VAChT)

(f)

Cholinesterase inhibition

Physostigmine, neostigmine

(g)

Receptors

mAChR and nAChR agonists/antagonists

CHOLINERGIC NEUROTRANSMISSION: SITES AND RECEPTORS Cholinergic transmission occurs at five important locations: (i) all effector sites innervated by parasympathetic postganglionic neurons; (ii) a small number of effector sites innervated by sympathetic postganglionic neurons (most importantly, most sweat glands); (iii) all autonomic ganglia (including the adrenal medulla), innervated by parasympathetic and sympathetic preganglionic neurons; (iv) all motor end plates on skeletal muscle, innervated by somatic motor neurons; and (v) certain synapses in the CNS. As might be expected for an ancient and ubiquitous neurotransmitter, a variety of ACh (cholinergic) receptor types have emerged in evolution. Based on their responsiveness to the agonists muscarine and nicotine, cholinergic receptors can be divided into two groups, nicotinic cholinergic receptors (nAChRs) and muscarinic cholinergic receptors (mAChRs). nAChRs are found at the following locations: l l l

Autonomic ganglia (including the adrenal medulla). Neuromuscular junction of skeletal muscle. Central nervous system.

There are two nAChR subtypes, NN (neural nicotinic) and NM (muscle nicotinic). NN receptors mediate neurotransmission in autonomic ganglia, while NM receptors

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15. ACETylCHolINE AND MusCARINIC RECEPToRs

TABLE 15.2 Muscarinic Receptor-Mediated Responses at Autonomic Effector sites Iris sphincter muscle

Contraction (miosis)

Ciliary muscle

Contraction (near vision)

Sinoatrial node

Bradycardia

Atrium

Decreased contractility

Atrioventricular node

Decreased conduction velocity

Arteriole

Dilation (through nitric oxide)

Bronchial smooth muscle

Contraction

Gastrointestinal motility

Increased

Gastrointestinal secretion

Increased

Gallbladder

Contraction

Bladder (detrusor)

Contraction

Bladder (trigone, sphincter)

Relaxation

Penis

Erection (but not ejaculation)

Sweat glands

Secretion

Salivary glands

Secretion

Lacrimal glands

Secretion

Nasopharyngeal glands

Secretion

mediate skeletal muscle contraction; thus, NN and NM antagonists are called ganglionic blockers and neuromuscular blockers, respectively. nAChRs in the CNS resemble NN receptors and are primarily presynaptic. nAChRs and the responses that they mediate are discussed in Chapter 16 of this Primer. mAChRs are found at the following locations: l

l

l

l

l

Effector sites innervated by parasympathetic postganglionic neurons. Effector sites innervated by sympathetic cholinergic postganglionic neurons (most sweat glands). Presynaptic sites on noradrenergic and cholinergic nerve terminals. Non-innervated sites in blood vessels (vascular endothelium, smooth muscle). Central nervous system.

Because of their physiological and therapeutic significance, it is important to review the specific responses mediated by mAChRs at autonomic effector sites. All of these sites are innervated by parasympathetic postganglionic neurons except for the sweat glands, which are innervated by sympathetic cholinergic postganglionic neurons (this innervation is sympathetic because of the thoracolumbar origin of the preganglionic neurons and is cholinergic because the postganglionic neurons release ACh). See Table 15.2. There are five mAChR subtypes, designated M1–M5. All mAChRs are G protein-coupled receptors. Stimulation of M1, M3, and M5 receptors generally results in the hydrolysis of phosphoinositides by activating the Gq-phospholipase C

pathway, resulting in mobilization of intracellular calcium; stimulation M2 and M4 receptors generally leads to the inhibition of adenylyl cyclase and the activation of potassium channels by activating Gi and Go. The five mAChR subtypes are widely distributed in both peripheral tissues and the CNS, with most cells expressing at least two subtypes. Due to the paucity of agonists and antagonists that are truly subtype-specific, identifying which subtype(s) mediate a particular mAChR response has been difficult. Studies in knockout mice have helped to elucidate the functions of specific mAChR subtypes. For example, M1 receptors modulate neurotransmitter signaling in the cortex and hippocampus and also have an important role in modulating cholinergic transmission in autonomic ganglia; M2 receptors are the predominant subtype mediating the parasympathetic control of the heart and also mediate mAChR agonist-induced tremor, hypothermia, and presynaptic inhibition of neurotransmitter release; M3 receptors are the predominant subtype mediating the parasympathetic control of smooth muscle contraction and glandular secretion, the sympathetic cholinergic control of sweating, and mAChR agonist-induced increases in food intake and body weight; M4 receptors modulate dopaminergic activity in motor tracts and also mediate the presynaptic inhibition of neurotransmitter release; and M5 receptors modulate central dopaminergic function and the tone of cerebral blood vessels.

MUSCARINIC AGONISTS ACh itself is rarely used as a mAChR agonist because of its rapid hydrolysis after oral or intravenous administration, but several choline esters that are resistant to hydrolysis (methacholine, carbachol, and bethanechol) are available for clinical use. Bethanechol has the additional favorable property of an overwhelmingly high mAChR (vs. nAChR) specificity. In addition, several natural alkaloids are mAChR agonists, including muscarine, pilocarpine, and arecoline; pilocarpine and arecoline (and their synthetic congeners) are used clinically. mAChR agonists are used in the treatment of urinary tract motility disorders and xerostomia (dry mouth due to decreased salivary secretion) and in the diagnosis of bronchial hyperreactivity; they also are used in ophthalmology to produce miosis and to treat glaucoma. Bethanechol, which primarily affects the urinary and GI tracts, is used to facilitate urination in patients with postoperative urinary retention, diabetic autonomic neuropathy, or neurogenic bladder (it formerly was used to increase GI motility in patients with postoperative abdominal distension, gastroparesis, and other motility impairments; more efficacious therapies are now available for these disorders). Pilocarpine is used to stimulate salivary gland secretion in patients with xerostomia resulting from damage to the salivary glands (e.g., due to head and neck radiation treatments) or Sjögren’s syndrome, an autoimmune disorder characterized by decreased secretions

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

MusCARINIC ANTAgoNIsTs

(particularly salivary and lacrimal); antibodies to mAChRs of the M3 subtype have been reported in some patients. Methacholine, administered by inhalation, is used in the diagnosis of bronchial airway hyperreactivity; elicitation of significant bronchoconstriction with inhaled methacholine challenge sometimes leads to the diagnosis of asthma in patients with little baseline abnormality in pulmonary function. Pilocarpine and carbachol are used topically in ophthalmology as miotic agents and to treat glaucoma. Most side effects of mAChR agonists are predictable and include increased salivation, increased sweating, worsening of asthma, diarrhea, nausea, hypotension, and bradycardia; occasionally mAChR agonists cause hiccups. In rare cases, high doses of bethanechol may cause myocardial ischemia in patients with a predisposition to coronary artery spasm; therefore, chest pain in a patient taking bethanechol should be taken seriously.

MUSCARINIC ANTAGONISTS The classical mAChR antagonists are alkaloids derived from plants in the Solanaceae family, including the deadly nightshade (Atropa belladonna), jimson weed (Datura stramonium), and henbane (Hyoscyamus niger). Atropine, scopolamine, and hyoscine are notable examples of the plant-derived mAChR antagonists; these compounds often are termed belladonna alkaloids. The name belladonna (Italian for “beautiful lady”) originates from the historic use of drops prepared from such plants by women who wished to dilate their pupils for beauty. mAChR antagonists cause pupillary dilation (mydriasis) by blocking mAChRs on the iris sphincter muscle; such compounds currently are used in ophthalmology as mydriatics. The hallucinogenic effects of the belladonna alkaloids have been known for centuries; for example, the tiny dark seeds from jimson weed pods were used in sacred ceremonies by aboriginal Americans. In addition to the belladonna alkaloids atropine and scopolamine, the mAChR antagonists used clinically include (i) semisynthetic derivatives of these alkaloids, which differ from their parent compounds primarily in their distribution in the body and/or duration of action; and (ii) synthetic derivatives of the belladonna alkaloids, some of which exhibit some selectivity for specific mAChR subtypes. Notable drugs in these two categories are homatropine and tropicamide, which have a shorter duration of action than atropine; ipratropium, tiotropium, and methscopolamine, which are quaternary amines and, therefore, do not cross the blood–brain barrier or readily cross membranes and epithelial cell barriers; and agents with some mAChR subtype selectivity, such as pirenzepine (selective for M1 receptors), darifenacin (selective for M3 receptors), and solifenacin (selective for M3 receptors). mAChR antagonists are used to inhibit parasympathetic stimulation of the urinary tract, respiratory tract, GI tract, heart, and eye; because of their CNS effects, they also are used to treat Parkinson’s disease, manage extrapyramidal

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(motor) symptoms caused by antipsychotic drugs, and prevent motion sickness. Oxybutynin and tolterodine are examples of drugs used to treat overactive bladder; these agents also can be used to reduce urinary frequency in patients with spastic paraplegia and to treat enuresis in children. Ipratropium and tiotropium, administered by inhalation, are important drugs in the treatment of chronic obstructive pulmonary disease. Ipratropium, administered via a nasal inhaler, is used for the treatment of rhinorrhea associated with the common cold or perennial rhinitis. Pirenzepine (not available in the US) is used for the treatment of acid-peptic disease; however, H2 antihistamines and proton pump inhibitors are more efficacious and more widely used for this purpose. Atropine, hyoscyamine, and other mAChR antagonists are used to treat irritable bowel syndrome and other conditions in which GI motility is increased, although their efficacy is limited in most patients. Atropine also is used to increase heart rate and/ or atrioventricular conduction during situations in which parasympathetic stimulation of the heart is enhanced (e.g., acute myocardial infarction involving the inferior or posterior wall of the left ventricle). Homatropine, tropicamide, and other mAChR antagonists are used in ophthalmology to dilate the pupils and paralyze the accommodation reflex. Benztropine and trihexyphenidyl are used in Parkinson’s disease, especially for the treatment of tremor; these agents also can be used to treat extrapyramidal side effects (such as dystonia and parkinsonian symptoms) caused by antipsychotic drugs. Scopolamine is widely used in a transdermal preparation to prevent motion sickness. Most side effects of mAChR antagonists are predictable and include constipation, xerostomia, anhidrosis (decreased sweating), urinary retention, precipitation of attacks of angle-closure glaucoma in susceptible patients, tachycardia, decreased lacrimation, and decreased respiratory secretions.

Further Reading Abrams P, Andersson K-E, Buccafusco J, et al. Muscarinic receptors: their distribution and function in body systems, and the implications for treating overactive bladder. Br J Pharmacol 2006;148:565–78. Barnes PJ, Hansel TT. Prospects for new drugs for chronic obstructive pulmonary disease. Lancet 2004;364:985–96. Brown JH, Laiken N. Muscarinic receptor agonists and antagonists. In: Brunton LL, editor. Goodman and Gilman’s the pharmacological basis of therapeutics. New York: McGraw-Hill; 2011. p. 219–37. Bymaster FP, McKinzie DL, Felder CC, Wess J. Use of Ml-M5 muscarinic receptor knockout mice as novel tools to delineate the physiological roles of the muscarinic cholinergic system. Neurochem Res 2003;28:437–42. Caulfield MP, Birdsall NJ. International Union of Pharmacology. XVII. Classification of muscarinic acetylcholine receptors. Pharmacol Rev 1998;50:279–90. Conn PJ, Christopoulos A, Lindsley CW. Allosteric modulators of GPCRs: a novel approach for the treatment of CNS disorders. Nature Rev Drug Discov 2009;8:41–54. Conn PJ, Jones CK, Lindsley CW. Subtype-selective allosteric modulators of muscarinic receptors for the treatment of CNS disorders. Trends in Pharmacol Sci 2009;30:148–55. Ferguson SM, Blakely RD. The choline transporter resurfaces; new roles for synaptic vesicles? Mol Interv 2004;4:22–37.

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Higgins CB, Vatner SF, Braunwald E. Parasympathetic control of the heart. Pharmacol Rev 1973;25:119–55. Taylor P, Brown JH. Acetylcholine. In: Siegel GJ, Albers RW, Brady ST, Price DL, editors. Basic neurochemistry: molecular, cellular, and medical aspects. Burlington, Massachusetts: Elsevier Academic Press; 2006. p. 185–209. Waterman SA, Gordon TP, Rischmueller M. Inhibitory effects of muscarinic receptor autoantibodies on parasympathetic neurotransmission in Sjögren’s syndrome. Arthritis Rheum 2000;43:1647–54.

Wellstein A, Pitschner JF. Complex dose-response curves of atropine in man explained by different functions of M1- and M2-cholinoceptors. Naunyn Schmiedebergs Arch Pharmacol 1988;338:19–27. Wess J, Eglen RM, Gautam D. Muscarinic acetylcholine receptors: mutant mice provide new insights for drug development. Nature Rev Drug Discov 2007;6:721–33.

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16 Nicotinic Receptors Palmer Taylor Nicotinic acetylcholine receptors are members of a large pentameric super family of ligand-gated ion channel receptors that include the 5-hydoxytryptamine3 (5-HT3), glycine, γ-aminobutyric acid (GABA), and Zn-activated families of receptors along with several invertebrate and prokaryotic receptors [1–4]. Less closely related are the ligand-gated ion channels that respond to the excitatory amino acids and adenosine. Owing to the abundance of nicotinic receptors in the electric fish and findings showing that several peptide toxins that block motor activity bind to subtypes of nicotinic receptor with high affinity and selectivity, the nicotinic receptor was the first pharmacologic receptor to be purified and the cDNAs encoding its subunits cloned. Appropriately, the nicotinic receptor became the prototype for the ligand-gated ion channel family.

conformational states that lead to activation and desensitization [1–5]. More recently, a soluble protein, termed the acetylcholine binding protein, exported from glial cells of snails, has been shown to bind acetylcholine and many of the classical nicotinic agonists and antagonists [6]. Homologous proteins have been found in other invertebrate species as well. This protein, with identical subunits of slightly over 200 amino acids, is composed of residues with homology to the amino-terminal, extracellular domain of family of nicotinic receptors. Its structural characterization by X-ray crystallography (Fig. 16.3) shows it to be pentameric and to contain an arrangement of amino acid residues consistent with findings of protein modification and site-specific mutagenesis studies conducted with acetylcholine receptors isolated from neuronal and muscle systems [5,6]. The binding protein protein is homomeric, and its five binding sites reside at the five subunit interfaces with binding site determinants coming from both of the proximal subunit surfaces. The ligands bind from an outer radial direction [6,7] and enlodge behind a loop that contains selective binding determinants and proximal cysteines at or near its tip (Fig. 16.3). Hence, consistent with other proteins that exhibit homotropic cooperativity, the binding protein and the nicotinic receptors have their binding sites located at the subunit interfaces.

STRUCTURAL CONSIDERATIONS Nicotinic receptors assemble as pentamers of individual subunits. Assembly occurs in a precise manner and order such that the assembled subunits encircle an internal membrane pore and the extracellular vestibule leading to the pore. The replicated pattern of front to back assembly of subunits insures that identical subunit interfaces are formed from the pentameric assembly of identical subunits. Moreover, the amino acid residues found at homologous positions in the receptors with heteromeric subunit assemblies should occupy the same position in three dimensional space (Fig. 16.1A). Each subunit encodes a protein with four transmembrane spans. The amino-terminal ~210 amino acids lie in the extracellular domain followed by three tightly threaded transmembrane spans. A relatively large cytoplasmic loop is found between transmembrane spans 3 and 4 with a short carboxyl terminus winding up on the extracellular side. The overall structure has been detailed in a series of electron microscopy imaging studies (Fig. 16.2) and reveals a large extracellular domain, a wide diameter vestibule on the extracellular side and the gorge constriction region controlling the gating function to exist within the transmembrane spanning region [3]. Rapid and slow binding of ligands appear to induce shape changes within the receptor structure reflective of the individual

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00016-0

SUBTYPE DIVERSITY OF NICOTINIC RECEPTORS The receptor from skeletal muscle and its homologous forms in the fish electric organ consist of four distinct subunits, where the pentamer has two copies of the α subunit and one copy of β, γ and δ subunits. When muscle becomes innervated, the γ subunit is replaced by an  subunit. Within this pentameric structure, the two acetylcholine binding sites exist at the αδ and αγ() subunit interfaces. Opposing faces of the homologous α and γ, δ or  subunits make up the two binding sites. The subunit compositions of the receptors expressed in the nervous system are far more complex where we find nine different α subtypes and four different β subtypes [8] (Table 16.1). The α subtypes have been defined as those containing vicinal cysteines on a loop on which

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TABLE 16.1 subtypes of Nicotinic Acetylcholine Receptors found in Muscle and Neuronal systems Composition α1, β1, γ, δ,  α6 α7 α9

β2 β3 β4

α2βγδ or α2βδ Various pentameric assemblies of α and β or α alone

α1 (A)

β2

β2

δ

α4

αβγδAChR

β2

α

α αAChR

αβNeuronal AChR

α10

α

α

H2N

ELECTROPHYSIOLOGIC EVENTS ASSOCIATED WITH RECEPTOR ACTIVATION All of the nicotinic receptors that are well characterized, to date, are cationic channels resulting in depolarization after activation. They differ substantially with respect to the permeabilities of the open channel state to Na and Ca. Na permeability is most effective in initiating a rapid depolarization, whereas Ca permeability could

2

COOH

1

reside several binding site determinants. All α subunits, except α5, use the face with the vicinal cysteines to form the binding site. The β subunits which lack this sequence and presumably the α5 subunit use the opposite or complementary face to form the binding site (Fig. 16.1). The α subunits, α2, α3, α4 and α6, combine with certain β subunits, mainly β2 and β4, to form pentameric receptors. α5 has the capacity to substitute for a β subunit in the pentameric assembly. Typically, it is thought that two of the neuronal α subunits assemble with three β subunits. Hence, typical stoichiometries (subscripted) would be α2β3 or α2α51β3 where the generic α subtypes are usually α3 or α4 and the β subtypes β2 or β4. Each of these heteromeric receptors would have two binding sites. Receptors composed of α7 subunits appear as functional homomeric entities where five copies of a single subunit confer function, when assembled. α9 and α10 subunits assemble uniquely in a pentamer and may control nociceptive responses. While the assembly patterns of the neuronal nicotinic receptor subunits are diverse, only certain combinations yield functional receptors, and particular combinations seem to be prevalent within regional areas of the nervous system. For example, pentamers of α3β4 are prevalent in ganglia of the autonomic nervous system, α4β2 is most prevalent in the central nervous system, and α7 appears widely distributed [8]. Central and peripheral nicotinic receptors play a discrete role in nicotine addiction and cardiovascular actions [9]. Mutations in nicotinic receptors give rise to congenital myasthenic syndromes [10] and other manifestations that affect autonomic and central nervous system function.

4

α2 α3 α4 α5

β1

γ

3

Muscle Neuronal

Assembly

α

α4

α1

(B)

FIGURE 16.1 Structure of the nicotinic acetylcholine receptor. (A) Arrangement of receptor subunits as pentamers in muscle and neuronal receptors. The muscle receptor exists as a pentamer with two copies of α1 and one of β, γ and δ, as seen for the receptor found in embryonic muscle. The binding sites as designated are found at the αγ and αδ interfaces. In the case of the innervated receptor in skeletal muscle, an  subunit of different composition replaces γ at the same position. Two types of neuronal receptors are shown: the heteromeric type is thought to have usually two copies of α, where α is α2,α3,α4, or α6, and three copies of β, where β can be β2, β3 or β4. α5 is thought to substitute for a β subunit at one of the non-binding positions. The homomeric neuronal receptors are made up of α7 pentamers. α7 subunits may form heteromeric subtypes with certain β subunits remains open, but not proven. α9 and α10 subunits associate with each other in presumed variable ratios to form pentamers. (B) Threading pattern of the α-carbon chain of the subunits of the nicotinic receptor. The first ~210 amino acids form virtually all of the extracellular domain, contain the residues on the primary and complementary subunit interfaces forming the binding site determinants and govern the assembly process. Transmembrane span 2 from the five subunits forms the inner perimeter of the channel and is involved in the ligand gating. The other transmembrane segments contribute to the structural integrity of the receptor. The extended segment between transmembrane spans 3 and 4 forms the bulk of the cytoplasmic domain.

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DIsTRIbuTIoN of NICoTINIC RECEPToRs

(A)

 

(B)



synapse

synaptic face

M

cytoplasm

N

S

I

S

S

S

L

L

T

G P

F

V

T

L

V

L

L

L

E

V

I

C

S

P

I

cytoplasmic face

(C)

M2



E T

V L K D

FIGURE 16.2 Overall dimensional characteristics of the nicotinic receptor. Image reconstruction of electron micrographs shows the receptor to be some ~140 Angstroms in length and perpendicular to the membrane. Its diameter is 80–90 Angstroms and contains a large central channel on the membrane surface. Rapid and prolonged exposure to acetylcholine followed by rapid freezing has yielded distinctive structures for the agonist bound, unligated and desensitized states of the receptor [3,6].

serve as an intrinsic activating function in cell signaling or in triggering an excitation step. However, depolarization per se through an increase in Na permeability can also mobilize Ca through voltage sensitive Ca channels. Typically postsynaptic nicotinic receptors, such as those found in ganglia and the neuromuscular junction, function through the simultaneous occupation of the receptor by more than a single agonist molecule. Agonist association and the ensuing channel opening reveals positive cooperativity in ligand binding. These rapid binding events cause channel openings to occur in a millisecond time frame. Several openings and closings may occur during the short interval of agonist occupation, and the intrinsic efficacy of an agonist may relate to whether the ligand-receptor complex favors an open or closed state. Upon continuous exposure to agonists, most receptors desensitize. Desensitization confers a receptor state wherein the agonist has a high affinity, but the receptor is locked in a closed channel state. Desensitization provides an additional means by which temporal responses to an agonist can be regulated. Antagonists may be competitive or non-competitive. Competitive antagonists show binding that is mutually exclusive with agonists and exhibit a surmountable block of the receptor in which the dose–response curve to the agonist is shift rightward in a parallel fashion. Occupation by a single antagonist molecule is sufficient to block function. Non-competitive antagonists typically block the channel through which ions pass in the open state of the receptor. In the case of ganglionic nicotinic receptors,

trimethaphan is a competitive agonist, while hexamethonium and mecamylamine are non-competitive, therein blocking channel function. Great interest has developed in allosteric sites where binding of a ligand at a non-agonist site, enhances or diminishes agonist function and are termed allosteric activators or inhibitors. Hence, they are distinguished from orthosteric ligands that occupy the primary agonist or antagonist sites.

DISTRIBUTION OF NICOTINIC RECEPTORS Nicotinic receptors are widely distributed in the central and peripheral nervous systems. In innervated skeletal muscle they are found in very high density localized to the motor end plate. Also, certain motor neurons may contain receptors at the presynaptic nerve ending to control release. In ganglia, the primary nicotinic receptor is found on the postsynaptic dendrite and nerve cell body. Others may exist presynaptically to control release from the presynaptic nerve ending. In the CNS, one finds that the majority of receptors are presynaptic or prejunctional. As such, they control the release of other transmitters or in the case of released acetylcholine, they play an autoreceptor role. Presynaptic nicotinic receptors found in the spinal cord and in higher centers of the brain have a functional role in modulating central control of autonomic function and sensory to autonomic control of reflexes and other functions.

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(A)

(B)

FIGURE 16.3 X-ray crystallographic structure of the acetylcholine binding protein. The structure was produced from crystallographic coordinates of Sixma and colleagues [5]. Surfaces are represented as Connolly surfaces. Left, colors are used to delineate the subunit interfaces in the homomeric pentamer. The vestibule on the extracellular surface that represents the entry to the internal channel in the receptor is shown by the arrow. Right, a single subunit interface Note the residues that have been found to be determinants in the binding of agonists and alkaloid and peptidic antagonists to the receptor. Hence, the numbers delineate the probable binding surface(s) for large and small antagonists. These residues come from seven distinct segments of amino acid sequence in the subunit, as determined from extensive mutagenesis, labeling and crystallographic studies [3,5,6]. The right hand panel shows a superimposition of the acetylcholine binding protein with the presumed structure of the muscle receptor.

References [1] Changeux J-P, Edelstein SJ. Nicotinic acetylcholine receptors. New York: Odie Jacob; 2005. [2] Karlin A. Emerging structures of nicotinic acetylcholine receptors. Nat Rev Neurosci 2002;3:102–14. [3] Changeux J-P. Allosteric receptors: From electric organ to cognition. Annu Rev Pharmacol Toxicol 2010;50:1–38. [4] Thompson AJ, Lester HA, Lummis SCR. The structural basis of function in Cys-loop receptors. Q Rev Biophys 2010;43:449–99. [5] Unwin N. Refined structure of the nicotinic acetylcholine receptor at 4A resolution. J Mol Biol 2005;346:967–89. [6] Brejc K, van Dijk WJ, Klaasen RV, Shuurmans M, van der Oost J, Smit AB, et al. Crystal structure of an acetylcholine binding protein

[7]

[8]

[9] [10]

reveals the ligand binding domain of nicotinic receptors. Nature (London) 2001;411:269–76. Hibbs RE, Sulzenbacher G, Shi J, Talley T, Conrod S, Kem WR, et al. Structural determinants for interaction of partial agonist with the acetylcholine binding protein and the neuronal alpha7 acetylcholine receptor. EMBO J 2009;28:3040–51. Various authors. Neuronal nicotinic receptors. In: Clementi F, Fornasi D, Gotti C, editors. Handbook of experimental pharmacology, vol. 144. Berlin: Springer-Verlag, 2000. 821 pp. Benowitz NL. Nicotine Addiction. N Eng J Med 2010;362:2295–303. Engel AG, Shen X-M, Selcen D, Sine SM. What we have learned from congenital myasthenic syndromes. J Mol Neurosci 2010;40:143–53.

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C H A P T E R

17 Serotonin Receptors and Neurotransmission Elaine Sanders-Bush, Charles D. Nichols Tryptophan hydoxylase, the rate limiting enzyme, is not saturated under normal conditions, rendering 5-HT levels sensitive to changes in blood tryptophan. This translates into the remarkable finding that brain levels of this neurotransmitter can be regulated by the dietary intake of tryptophan. Clinical studies have often utilized a tryptophan-free diet to lower brain 5-HT and, by inference, evaluate its role in a particular behavior or drug effect. The principal metabolism of 5-HT is mediated by the ubiquitous enzyme, monoamine oxidase (MAO), to form an inactive product, 5-hydroxyindole acetic acid (5-HIAA). 5-HIAA is subsequently secreted in the CSF and urine. MAO is a family of mitochondrial enzymes that metabolize all biogenic amines.

Serotonin (5–hydroxytryptamine) is a neurotransmitter as well as a circulating hormone. Serotoninergic neurons in the brain synthesize and store serotonin at axon terminals where it is released and interacts with cell-surface receptors on adjacent neurons. The action of serotonin is terminated by re-uptake into presynaptic terminal mediated by the serotonin transporter or by metabolism by monoamine oxidase. Of the 14 different receptor subtypes, most generate intracellular messengers by coupling to G-proteins and modulate, rather than mediate, fast neurotransmission. This multitude of receptors explains the variety of actions of serotonin in normal and abnormal states and provides ample opportunity for drug development for treatment of nervous system diseases.

NEUROTRANSMISSION

LOCALIZATION

In the central nervous system, the entire pathway of synthesis and metabolism exists at axon terminals. The life cycle of 5-HT at the synapse is illustrated in Figure 17.3. Both synthetic enzymes are present in the presynaptic terminal; MAO is also highly expressed in adjacent cells. Newly synthesized 5-HT is accumulated in synaptic vesicles to protect it from degradation by MAO. Uptake into vesicles is mediated by the vesicular monoamine transporter (VMAT). Stored 5-HT is released from synaptic vesicles by a complex series of phosphorylation-dependent protein-protein interactions initiated by the influx of calcium. Once released, 5-HT is inactivated by MAO in the synaptic cleft or its action is terminated by re-uptake into the presynaptic terminal by the 5-HT transporter (SERT). Once in the presynaptic terminal, 5-HT is either accumulated in synaptic vesicles via VMAT or metabolized by MAO. The two transporters in 5-HT nerve terminals, VMAT and SERT, belong to different gene families and have markedly different properties. SERT is a sodium-dependent carrier that translocates 5-HT into the presynaptic nerve terminal. VMAT is driven by a proton gradient; it is promiscuous, present in 5-HT and catecholamine vesicles. SERT is expressed exclusively in serotoninergic neurons

Serotonin, also referred to as 5-hydroxytrytamine (5-HT), is a simple indoleamine (Fig. 17.1), discovered over five decades ago. Since then, 5-HT has been shown to function as a neurotransmitter in central nervous system and also as circulating hormone [1]. The principal source of circulating 5-HT is the intestinal enterochromaffin cells, where 5-HT is synthesized, stored and released into the bloodstream. 5-HT in blood is concentrated in platelets by an active transport mechanism. In the pineal gland, 5-HT is converted by a two-step process to melatonin (5-methoxy-N-acetyltryptamine), a hormone that regulates ovarian function and has been implicated in the control of biological rhythms. The brain 5-HT containing neurons are localized in raphe nuclei of the brainstem, which project diffusely throughout the brain and spinal cord.

SYNTHESIS AND METABOLISM The pathway of synthesis is common throughout the body. 5-HT is synthesized from tryptophan, an essential amino acid obtained in the diet. 5-HT synthesis requires the action of two synthetic enzymes, as in Figure 17.2.

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17. SERoToNIN RECEPToRS ANd NEuRoTRANSmISSIoN

FIGURE 17.1 The chemical structure of serotonin (5–hydroxytryptamine).

FIGURE 17.2 Biosynthetic and metabolic pathway of serotonin. TABLE 17.1 Serotonin Receptor Subtypes and Pharmacology Receptor Family 5-HT1

Primary Transduction Subtype Pathway

Activation of DOI phospholipase BW723C86 Lorcaserin C (Gaq)

MDL100907 RS127445 RS102221

5-HT3

Ligand gated ion channel

m-CPBG

Ondansetron

5-HT4

Activation of adenylate cyclase (Gas)

Cisapride

GR113808

5-HT6

RECEPTORS 5-HT in the synaptic cleft interacts with cell-surface receptors localized at the postsynaptic membrane or on the

WAY100635 SB224289 BRL15572

5-HT2A 5-HT2B 5-HT2C

5-HT5A 5-HT5B

in the CNS and is also found in the enteric nervous system and in blood platelets. Platelets are devoid of 5-HT synthetic enzymes; transport into platelets by SERT is responsible for the high level of 5-HT found in platelets.

U92016A Anpirtoline PNU-142633

Inhibition of adenylate cyclase (Gai)

5-HT5

5-HT is stored in synaptic vesicles to avoid metabolism to 5-HIAA by mitochondrial MAO. Released 5-HT interacts with receptors in the postsynaptic membrane or on autoreceptors in the presynaptic membrane. The principal mechanism of inactivation is reuptake into the presynaptic terminal by the 5-HT transporter.

Pharmacology: Antagonist

5-HT1A 5-HT1B 5-HT1D 5-HT1E 5-HT1F

5-HT2

FIGURE 17.3 Schematic of 5-HT nerve terminal. Newly synthesized

Pharmacology: Agonist

5-HT7

LY344864

Inhibition of adenylate cyclase (Gai) Activation of adenylate cyclase (Gas) Activation of adenylate cyclase (Gas)

SB699551

EMD386088

SB399885

AS-19

SB258719

presynaptic terminal. The 14 serotonin receptors, reclassified in 1994 [2], segregate into seven families (Table 17.1). All but one are members of the superfamily of G proteincoupled receptors (GPCRs), which are predicted to span the plasma membrane seven times with the N-terminus on the outside of the cell and the C-terminus, intracellular [3]. The intracellular loops and C-terminal tail interact directly with G proteins (Fig. 17.4). As illustrated in Figure 17.3, most of the G protein-coupled 5-HT receptors are localized on postsynaptic membranes and modulate

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RolE IN AuToNomIC PRoCESSES

TABLE 17.2 Clinically Available 5-HT drugs

FIGURE 17.4 5-HT receptors couple to multiple G-proteins. G proteins are classified based on their alpha subunits. 5-HT receptors have been definitively shown to couple to four different families of G-proteins.

neurotransmission via second messenger pathways. In contrast, the 5-HT3 receptor is a multimeric 5-HT gaited cation channel; it is primarily localized on presynaptic terminals of non-serotoninergic neurons where it regulates the release of other neurotransmitters such as acetylcholine. GPCRs generate intracellular second messengers such as cyclic AMP and calcium, which stimulate or inhibit various kinases and phosphatases that, in turn, regulate proteins by changes in phosphorylation state [4]. As such these receptors act as neuromodulators, modulating other receptors and ion channels that mediate fast neurotransmission. The 5-HT3 receptor is the only 5-HT receptor that gates ions, hence directly altering membrane potential. The other 5-HT receptors indirectly modify the membrane potential by regulating voltage-gated ion channels (such as Ca or K channels) or ligand-gated ion channels (such as glutamate receptors). For example, presynaptic 5-HT1B receptors inhibit N-type calcium channels via Go G-protein, thereby decreasing 5-HT release. This receptor, as well as other G protein-coupled 5-HT receptors, are expressed on the presynaptic terminals of other neurotransmitter releasing neurons, where these so-called heteroreceptors inhibit or potentiate neurotransmitter release. Thus, the possibilities of neurotransmitter cross-talk are numerous and widespread.

PHARMACOLOGY AND ROLE IN DISEASE The large number of receptors translates into a complex pharmacology and a myriad of targets for drug development. Specific drugs do exist, but they are rare. The drugs listed in Table 17.1 are at least 50-fold more potent at their primary target, which translates into reasonable specificity in vivo. Readers interested in more information about pharmacological properties should access the extensive database at http://pdsp.med.unc.edu/indexR.html. Serotonin plays a role in a myriad of behaviors [5]. Advances in genetically modified mice have advanced our understanding of the role of specific receptors in behaviors and drug actions [6]. The clinically available drugs that target 5-HT neurotransmission (Table 17.2) have a range of disease targets

Target

Action

Clinical Use

Examples

MAO

Antagonist

Major depressive Tranylcypromine illness

5-HT Transporter Channel blocker

Major depressive Fluoxetine, illness, panic, escitalopram anxiety, OCD

5-HT1A receptor

Agonist

Anxiety

Buspirone

5-HT1D receptor

Agonist

Migraine

Sumatriptan

5-HT2 receptor

Antagonist

Migraine

Methysergide

5-HT2A receptor

Antagonist

Schizophrenia

Clozapine, risperidone

5-HT4 receptor

Agonist

Irritable bowel syndrome

Prucalopride (Europe)

5-HT3 receptor

Channel blocker

Nausea/emesis

Ondansetron

and varying degrees of specificity. Esitalopram, for example, is a highly specific inhibitor of 5-HT transporter with three orders of magnitude lesser affinity for secondary targets (catecholamine transporters). On the other hand, tranylcypromine blocks the degradation of serotonin, dopamine and norepinephrine equally well. One of the most exciting areas of current research deals with genetic variations in 5HT receptor and transporter genes and their association with human diseases, such as schizophrenia and major depressive illness. Research has focused on more common genetic alterations referred to as single nucleotide polymorphisms (SNPs) and, although inconsistent, the early results suggest that SNPs in serotoninrelated genes may be associated with disease symptoms as well as drug response [7].

ROLE IN AUTONOMIC PROCESSES 5-HT serves a key role in the regulation of gut function and motility [8]. Enterochromaffin cells synthesize and store 5-HT in granules, and release 5-HT upon stimulation by norepinephrine released from the myenteric plexus. This 5-HT can then bind to 5-HT4 and 5-HT2 receptors and activate secretion from enterocytes into the intestinal lumen. Furthermore, 5-HT also binds to 5-HT3/4 receptors on the submucosal plexus of the enteric nervous system to modulate smooth muscle contractility and peristalsis. Too much 5-HT in the gut, resulting from a carcinoid tumor or cisplatin chemotherapy for example, induces diarrhea and emesis. Too little 5-HT in the gut results in slow transit times and constipation. Antagonists of 5-HT3 receptors, including ondansetron, are effective anti-emetics to block the nausea associated with chemotherapy. Although agonists of 5-HT4 receptors like cisapride have had some success in the clinic to increase gut motility associated with irritable bowel syndrome and constipation, they have been

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withdrawn from the market due to their ability to produce fatal cardiac arrhythmias in some patients. Other 5-HT4 agonists are available in Europe. Many cardiovascular functions, including heart rate and blood pressure, are modulated by 5-HT acting at a variety of receptors located in the sympathetic ganglia, parasympathetic ganglia, the vagus nerve, and the heart itself [9]. The interactions of 5-HT with its receptors are complex in mediating cardiovascular effects. Increasing circulating 5-HT levels primarily produces tachycardia. Receptor selective drugs, however, can produce either bradycardia or tachycardia, influence heart rate, cardiac outflow, or blood pressure, as well as have adverse developmental effects like cardiac valvulopathies depending on the receptor targeted and site of action.

References [1] Sanders-Bush E, Hazelwood L. 5-Hydroxytryptamine (serotonin) and dopamine. In: Brunton LL, editor. The pharmacological basis of therapeutics. New York: McGraw-Hill; 2011. p. 335–62.

[2] Hoyer D, Clarke DE, Fozard JR, Hartig PR, Martin GR, Mylecharane EJ, et al. International Union of Pharmacology classification of receptors for 5-hydroxytryptamine (Serotonin). Pharmacol Rev 1994;46:157–203. [3] Nichols DE, Nichols CD. Serotonin Receptors. Chem Rev 2008;46:1614–41. [4] Millan MJ, Marin P, Bockaert J, Mannoury la Cour C. Signaling at G-Protein-coupled serotonin receptors: recent advances and future research directions. Trends Pharmacol Sci 2008;29:454–64. [5] Lucki I. The spectrum of behaviors influenced by serotonin. Biol Psychiatry 1998;44:151–62. [6] Murphy DL, Wichems C, Li Q, Heils A. Molecular manipulations as tools for enhancing our understanding of 5-HT neurotransmission. Trends Pharmacol Sci 1999;20:246–52. [7] Hariri AR, Mattay VS, Tessitore A, Kolachana B, Fera F, Goldman D, et al. Serotonin transporter genetic variation and the response of the human amygdala. Science 2002;297:400–3. [8] Spiller R. Serotonin and GI clinical disorders. Neuropharmacology 2008;55:1072–80. [9] Villalon CM, Centurion D. Cardiovascular responses produced by 5-hydroxytriptamine: a pharmacological update on the receptors/mechanisms involved and therapeutic implications. NaunynSchmiedeberg’s Arch Pharmacol 2007;376:45–63.

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18 Purinergic Neurotransmission and Nucleotide Receptors Geoffrey Burnstock

PURINERGIC NEUROTRANSMISSION

Another concept that has had a significant influence on our understanding of purinergic transmission was that of cotransmission. Burnstock wrote a Commentary in Neuroscience in 1976 [4] entitled: “Do some nerves release more than one transmitter?” This position challenged the single-neurotransmitter concept, which became known as “Dale’s Principle”, even though Dale himself never defined it as such. The commentary was based on hints about cotransmission in the early literature describing both vertebrate and invertebrate neurotransmission, and, more specifically, with respect to purinergic cotransmission, on the surprising discovery in 1971 that ATP was released from sympathetic nerves supplying the taenia coli as well as from NANC inhibitory nerves. The excitatory junction potentials (EJPs) recorded in the vas deferens were blocked by α,β-meATP, a selective desensitizer of P2X receptors (Fig. 18.3a and b). This clearly supported the earlier demonstration of sympathetic cotransmission in the vas deferens in the laboratory of Dave Westfall, following an earlier report of sympathetic cotransmission in the cat nictitating membrane. Sympathetic cotransmission was later described in a variety of blood vessels. The proportions of ATP and noradrenaline (NA) vary in different tissues and species, during development and ageing and in different pathophysiological conditions. Acetylcholine (ACh) and ATP are cotransmitters in parasympathetic nerves supplying the urinary bladder. Subpopulations of sensory nerves have been shown to utilize ATP in addition to substance P and calcitonin gene-related peptide; it seems likely that ATP cooperates with these peptides in “axon reflex” activity. ATP, vasoactive intestinal polypeptide and nitric oxide (NO) are cotransmitters in NANC inhibitory nerves. ATP and NA act synergistically to release vasopressin and oxytocin from the hypothalamus, which is consistent with ATP cotransmission in the hypothalamus involved in central nervous system (CNS) control of autonomic functions. Release of ATP from autonomic nerves is by vesicular exocytosis and after release it is broken down by ectonucleotidases. Much is now known about the

The existence of non-adrenergic, non-cholinergic (NANC) neurotransmission in the gut was established in the mid 1960s (Fig. 18.1a). Several years later, after many experiments, a study was published that suggested that the NANC transmitter in the guinea-pig taenia coli and stomach, rabbit ileum, frog stomach and turkey gizzard was adenosine 5-triphosphate (ATP) [1]. The experimental evidence included mimicry of the NANC nervemediated response by ATP (Fig. 18.1b); measurement of release of ATP during stimulation of NANC nerves with luciferin-luciferase luminometry; histochemical labeling of subpopulations of neurons in the gut with quinacrine, a fluorescent dye known to selectively label high levels of ATP bound to peptides; the later demonstration that the slowly-degradable analog of ATP, α,β-methylene ATP (α,β-meATP), which produces selective desensitization of the ATP receptor, blocked the responses to NANC nerve stimulation. Soon after, evidence was presented for ATP as the neurotransmitter for NANC excitatory nerves in the urinary bladder (Fig. 18.1c and d). The term “purinergic” was proposed in a short letter to Nature in 1971 and the evidence for purinergic transmission in a wide variety of systems was presented in Pharmacological Reviews in 1972 [2] (Fig. 18.2). This concept met with considerable resistance for many years. Perhaps understandably, this was probably partly because it was felt that ATP was established as an intracellular energy source involved in various metabolic cycles and that such a ubiquitous molecule was unlikely to be involved in extracellular signaling. However, ATP was one of the biological molecules to first appear and, therefore, it is not really surprising that it should have been utilized for extracellular, as well as intracellular, purposes early in evolution. The fact that potent ectoATPases were described in most tissues in the early literature was also a strong indication for the extracellular actions of ATP. Purinergic neurotransmission is now generally accepted (see [3]).

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FIGURE 18.1 (a) Sucrose gap recording of membrane potential changes in smooth muscle of guinea-pig taenia coli in the presence of atropine (0.3 μM) and guanethidine (4 μM). Transmural field stimulation (0.5 ms, 0.033 Hz, 8 V) evoked transient hyperpolarizations, which were followed by rebound depolarizations. Tetrodotoxin (TTX, 3 μM) added to the superfusing Kreb’s solution (applied at arrow) rapidly abolished the responses to transmural field stimulation, indicating that they were inhibitory junction potentials in response to stimulation of NANC inhibitory nerves. (Reproduced from [19] with permission of Blackwell Publishing.) (b) Mechanical responses of the guinea-pig taenia coli to intramural nerve stimulation (NS, 1 Hz, 0.5 ms pulse duration, for 10 s at supramaximal voltage) and ATP (2  10–6 M). The responses consist of a relaxation followed by a “rebound contraction”. Atropine (1.5  10–7 M) and guanethidine (5  10–6 M) were present. (Reproduced from [20] with permission of the Nature Publishing Group). (c) A comparison of the contractile responses of the guinea-pig bladder strip to intramural nerve stimulation (NS: 5 Hz, 0.2 ms pulse duration and supramaximal voltage) and exogenous ATP (8.5 μM). Atropine (1.4 μM) and guanethidine (3.4 μM) were present throughout. (Reproduced from [21] with permission of the Nature Publishing Group). (d) Effect of changing the calcium ion (Ca2) concentration on the release of ATP from the guinea pig isolated bladder strip during stimulation of intramural nerves. Upper trace: mechanical recording of changes in tension (g) during intramural nerve stimulation (NS: 20 Hz, 0.2 ms pulse duration, supramaximal voltage for 20 seconds). Lower trace: concentration of ATP in consecutive 20 s fractions of the superfusate. The Ca2 concentration in the superfusate varied as follows: (i) 2.5 mM (normal Krebs); (ii) 0.5 mM; (iii) 0.25 mM; (iv) 2.5 mM. The successive contractions were separated by 60 min intervals as indicated by the breaks in the mechanical trace. Atropine (1.4 μM) and guanethidine (3.4 μM) were present throughout. The temperature of the perfusate was between 22°C and 23°C. (Reproduced from [21] with permission).

ectonucleotidases that break down ATP released from neurons and non-neuronal cells. Several enzyme families are involved: ecto-nucleoside triphosphate diphosphohydrolases (E-NTPDases) of which NTPDase 1, 2, 3 and 8 are extracellular; ectonucleotide pyrophosphatase (E-NPP) of three subtypes; alkaline phosphatases; ecto-5-nucleotidase and ecto-nucleoside diphosphokinase (E-NDPK). NTPDase1 hydrolyses ATP directly to AMP and uridine 5-triphosphate (UTP) to uridine diphosphate (UDP), while NTPDase 2 hydrolyses ATP to adenosine 5-diphosphate (ADP) and 5-nucleotidase AMP to adenosine (see [5]). Purinergic receptors have been cloned and characterized in amoeba, Schistosoma and green algae, which resemble P2X receptors found in mammals, suggesting that purinergic signaling was present early in evolution (see [6]). This perhaps explains the wide distribution of purinergic receptors in most non-neuronal as well as neuronal cell types (see [7]).

An important conceptual step was when, in addition to purinergic neuromuscular transmission, purinergic nerve– nerve synaptic transmission was described in coeliac ganglion in 1992 by Silinsky, Surprenant and colleagues. Synaptic transmission has also been demonstrated in the enteric plexuses and in various sensory sympathetic, parasympathetic and pelvic ganglia (see [3]). A hypothesis was proposed that purinergic mechanosensory transduction occurred in visceral tubes and sacs, including ureter, bladder and gut, where ATP, released from epithelial cells during distension, acted on P2X3 homomultimeric and P2X2/3 heteromultimeric receptors on subepithelial sensory nerves initiating impulses in sensory pathways to pain centers in the CNS. Subsequent studies of bladder, ureter, gut, tongue and tooth pulp have produced evidence in support of this hypothesis (see [8]).

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FIGURE 18.2 Purinergic neuromuscular transmission depicting the synthesis, storage, release and inactivation of adenosine 5-triphosphate (ATP). ATP, stored in vesicles in nerve varicosities, is released by exocytosis to act on postjunctional ATP receptors on smooth muscle. ATP is broken down extracellularly by ATPases and 5-nucleotidase to adenosine, which is taken up by varicosities to be re-synthesized and restored in vesicles. If adenosine is broken down further by adenosine deaminase to inosine and hypoxanthine, they are removed by the circulation. (Reproduced from [2] with permission from the American Society for Pharmacology and Experimental Therapeutics.)

While the early emphasis was on short-term purinergic signaling in neurotransmission, neuromodulation and secretion, it was later recognized that ATP, released from autonomic nerves and by paracrine or autocrine release from non-neuronal cells, is involved in long-term (trophic) signaling, involved in cell proliferation, differentiation and death in development and regeneration [9]. Purinergic signaling in the brain stem is involved in control of autonomic functions, including cardiovascular and respiratory control and in the regulation of hormone secretion and body temperature at the hypothalamic level. The nucleus tractus solitarius (NTS) is a major integrative center of the brain stem involved in reflex control of the cardiovascular system; stimulation of P2X receptors in the NTS evokes hypotension. P2X receptors expressed in neurons in the trigeminal mesencephalic nucleus might be involved in the processing of proprioceptive information.

RECEPTORS FOR PURINES AND PYRIMIDINES Implicit in the purinergic neurotransmission hypothesis was the presence of purinoceptors. A basis for distinguishing two types of purinoceptor, identified as P1 and P2 for adenosine and ATP/ADP, respectively, was recognized in

FIGURE 18.3 (a) Excitatory junction potentials in response to repetitive stimulation of adrenergic nerves (white dots) in the guinea pig vas deferens. The upper trace records the tension, the lower trace the electrical activity of the muscle recorded extracellularly by the sucrose gap method. Note both summation and facilitation of successive junction potentials. At a critical depolarization threshold an action potential is initiated which results in contraction. (Reproduced from [22] with permission from Springer). (b) The effect of various concentrations of α,β-methylene ATP (α,β-meATP) on EJPs recorded from guinea pig vas deferens (intracellular recordings). The control responses to stimulation of the motor nerves at 0.5 Hz are shown on the left. After at least 10 min in the continuous presence of the indicated concentration of α,β-meATP, EJPs were recorded using the same stimulation parameters. (Reproduced from [23] with permission from Elsevier.)

1978 [10]. This helped resolve some of the ambiguities in earlier reports, which were complicated by the breakdown of ATP to adenosine by ectoenzymes, so that some of the actions of ATP were directly on P2 receptors, whereas others were due to indirect action via P1 receptors. However, it was not until 1985 that a pharmacological basis for distinguishing two types of P2 receptors (P2X and P2Y) was proposed [11]. A year later, two further P2 receptor subtypes were named, a P2T receptor selective for ADP on platelets and a P2Z receptor on macrophages. Further subtypes followed, perhaps the most important of which being the P2U receptor, which could recognize pyrimidines such as UTP in addition to ATP. However, to provide a more manageable framework for newly identified nucleotide receptors, Abbracchio and Burnstock proposed in 1994 [12] that purinoceptors should belong to two major families: a P2X family of ligand-gated ion channel receptors and a P2Y family of G protein-coupled receptors. This was based on studies of transduction mechanisms and the cloning of nucleotide receptors: P2Y receptors were cloned first in 1993 and a year later P2X receptors were cloned (Fig. 18.4). This nomenclature has been widely adopted and currently seven P2X subtypes and eight P2Y

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FIGURE 18.4 Membrane receptors for extracellular ATP. (a) The P2X family of receptors are ligand-gated ion channels (S-S; disulphide bond; M1 and M2, transmembrane domains), and (b) the P2Y family are G protein-coupled receptors (S-S; disulphide bond; pale blue circles represent amino acid residues that are conserved between P2Y1, P2Y2 and P2Y3 receptors; fawn circles represent residues that are not conserved; and purple circles represent residues that are known to be functionally important in other G protein-coupled receptors). (Part (a) reproduced from [24] with permission from Nature; Part (b) modified from [25] with permission from Elsevier.)

receptor subtypes are recognized, while four subtypes of P1 receptor have been cloned and characterized (see [13]; Table 18.1).

P2X Receptors Members of the family of ionotropic P2X1–7 receptors show a subunit topology of intracellular N- and C- termini possessing consensus binding motifs for protein kinases; two transmembrane-spanning regions (TM1 and TM2), the first involved with channel gating and the second lining the ion pore; a large extracellular loop, with 10 conserved cysteine residues forming a series of disulfide bridges and an ATP-binding site, which may involve regions of the extracellular loop adjacent to TM1 and TM2 (see [14]). The crystal structure of the P2X4 receptor was recently described [15]. Heteromultimers as well as homomultimers are involved in forming the trimer ion pore. P2X2/3, P2X1/2, P2X1/5, P2X2/6, P2X4/6 and P2X1/4 receptor heteromultimers have been identified. P2X7 does not form heteromultimers, and P2X6 will not form a functional homomultimer. Advances have been made by the preparation of knockout mice for P2X1, P2X2, P2X3, P2X4 and P2X7 receptors, and transgenic mice that over-express the P2X1 receptor. Adenoviral expression of a P2X1 receptor-green fluorescent protein construct in vas deferens shows the receptor to be localized in clusters, with larger ones apposing nerve varicosities. The P2X2 receptor is generally described as non-desensitizing, compared with the P2X1 and P2X3 receptors. P2X3 receptors are prominently expressed on nociceptive sensory neurons. P2X2/3 heteromer receptors have been identified in subpopulations of sensory neurons and sympathetic ganglion cells. Homomeric P2X4 receptors

are activated by ATP, but not by α,β-meATP. The most useful distinguishing feature of ATP-evoked currents at P2X4 receptors is their potentiation by ivermectin. The P2X5 receptor cDNA was first isolated from cDNA libraries constructed from rat coeliac ganglion and heart. Cells expressing the heteromeric P2X1/5 receptor are very sensitive to ATP, concentrations as low as 3 or 10 nM evoking measurable currents. The P2X6 subunit is only functionally expressed as a heteromultimer. P2X2/6 heteromeric receptors are prominently expressed by respiratory neurons in the brainstem. The main feature of the P2X7 receptor is that, in addition to the usual rapid opening of the cationselective ion channel, with prolonged exposure to high concentrations of ATP large pores form and this usually leads to cell death. Vesicles are shed after activation of P2X7 receptors, which release inflammatory cytokines.

P2Y Receptors At present, there are eight accepted human P2Y receptors: P2Y1, P2Y2, P2Y4, P2Y6, P2Y11, P2Y12, P2Y13 and P2Y14 (see [16] and Table 18.1). The missing numbers represent either non-mammalian orthologs, or receptors having some sequence homology to P2Y receptors, but for which there is no functional evidence of responsiveness to nucleotides. In contrast to P2X receptors, P2Y receptor genes do not contain introns in the coding sequence, except for the P2Y11 receptor. Site-directed mutagenesis of the P2Y1 and P2Y2 receptors has shown that some positively charged residues in TM3, TM6 and TM7 are crucial for receptor activation by nucleotides. From a phylogenetic and structural (i.e., protein sequence) point of view, two distinct P2Y receptor subgroups characterized by a relatively high level of sequence divergence have been

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TABLE 18.1 Characteristics of Purine-mediated Receptors Transduction Mechanisms

Receptor

Main Distribution

Agonists

Antagonists

P1 (adenosine) A1

Brain, spinal cord, testis, heart, autonomic nerve terminals

CCPA, CPA, S-ENBA

DPCPX, N-0840, MRS1754

Gi/o ↓cAMP

A2A

Brain, heart, lungs, spleen

CGS 21680, HENECA

KF17837, SCH58261, ZM241385

GS ↑cAMP

A2B

Large intestine, bladder

NECA (non-selective)

Enprofylline, MRE2029-F20, MRS17541, MRS 1706

GS ↑cAMP

A3

Lung, liver, brain, testis, heart

IB-MECA, 2-Cl-IB-MECA, DBXRM, VT160

MRS1220, L-268605, MRS1191, MRS1523, VUF8504

Gi/o Gq/11 ↓cAMP ↑IP3

P2X1

Smooth muscle, platelets, cerebellum, dorsal horn spinal neurons

α,β-meATP  ATP  2-MeSATP (rapid desensitization), L-β,γ-meATP

TNP-ATP, IP5I, NF023, NF449

Intrinsic cation channel (Ca2 and Na)

P2X2

Smooth muscle, CNS, retina, chromaffin cells, autonomic and sensory ganglia

ATP  ATPγS  2-MeSATP  α,β-meATP (pH  zinc sensitive)

Suramin, isoPPADS, RB2, NF770

Intrinsic ion channel (particularly Ca2)

P2X3

Sensory neurones, NTS, some sympathetic neurons

2-MeSATP  ATP  α,βmeATP  Ap4A (rapid desensitization)

TNP-ATP, PPADS, A317491, NF110

Intrinsic cation channel

P2X4

CNS, testis, colon

ATP  α,β-meATP, CTP, Ivermectin

TNP-ATP (weak), BBG (weak)

Intrinsic ion channel (especially Ca2)

P2X5

Proliferating cells in skin, gut, bladder, thymus, spinal cord

ATP  α,β-meATP, ATPγS

Suramin, PPADS, BBG

Intrinsic ion channel

P2X6

CNS, motor neurons in spinal cord

– (does not function as homomultimer)

–

Intrinsic ion channel

P2X7

Apoptotic cells in, for example, immune cells, pancreas, skin

BzATP  ATP  2-MeSATP  α,β-meATP

KN62, KN04, MRS2427 Intrinsic cation channel Coomassie brilliant and a large pore with blue G prolonged activation

P2Y1

Epithelial and endothelial cells, platelets, immune cells, osteoclasts

2-MeSADP  ADPβS  2-MeSATP  ADP  ATP, MRS2365

MRS2179, MRS2500, MRS2279, PIT

Gq/G11; PLC-β activation

P2Y2

Immune cells, epithelial and endothelial cells, kidney tubules, osteoblasts

UTP  ATP, UTPγS, INS 37217

Suramin  RB2, AR-C126313

Gq/G11 and possibly Gi; PLC-β activation

P2Y4

Endothelial cells

UTP  ATP, UTPγS

RB2  Suramin

Gq/G11 and possibly Gi; PLC-β activation

P2Y6

Some epithelial cells, placenta, T cells, thymus

UDP  UTP  ATP, UDPβS

MRS2578

Gq/G11; PLC-β activation

P2Y11

Spleen, intestine, granulocytes

AR-C67085MX  BzATP  ATPγS  ATP

Suramin  RB2, NF157, 5-AMPS

Gq/G11 and GS; PLC-β activation

P2Y12

Platelets, glial cells

2-MeSADP  ADP  ATP

CT50547, AR-C69931MX, INS49266, AZD6140, PSB0413, ARL66096, 2-MeSAMP

Gi/o; inhibition of adenylate cyclase

P2Y13

Spleen, brain, lymph nodes, bone marrow Placenta, adipose tissue, stomach, intestine, discrete brain regions

ADP  2-MeSADP  ATP & 2-MeSATP UDP glucose  UDP-galactose

MRS2211, 2-MeSAMP

Gi/o

–

Gq/G11

P2X

P2Y

P2Y14

Updated from [18] with permission from Elsevier. Abbreviations: BBG, Brilliant blue green; BzATP, 2-&3-O-(4-benzoyl-benzoyl)-ATP; cAMP, cyclic AMP; CCPA, chlorocyclopentyl adenosine; CPA, cyclopentyl adenosine; CTP, cytosine triphosphate; IP3, inosine triphosphate; Ip5I, di-inosine pentaphosphate; 2-MeSADP, 2-methylthio ADP; 2-MeSATP, 2-methylthio ATP; NECA, 5-N-ethylcarboxamido adenosine; PLC, phospholipase C; RB2, reactive blue 2.

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identified. The first subgroup includes P2Y1,2,4,6,11 and the second subgroup encompasses the P2Y12,13,14 subtypes. Selective antagonists have been identified for some P2Y receptor subtypes (see Table 18.1). P2Y1, P2Y2, P2Y4 and P2Y6 receptors couple to G proteins to increase inositol triphosphate (IP3) and cytosolic calcium. Activation of the P2Y11 receptor by ATP leads to a rise in both cAMP and in IP3, whereas activation by UTP produces calcium mobilization without IP3 or cAMP increase. The P2Y13 receptor can simultaneously couple to G16, Gi and, at high concentrations of ADP, to Gs. The activation of several P2Y receptors is commonly associated with the stimulation of several mitogen-activated protein kinases, in particular extracellular signal regulated protein kinase 1/2. In most species, ADP is a more potent agonist than ATP at P2Y1 receptors. Site-directed mutagenesis studies on the human P2Y1 receptor have shown that amino acid residues in TM3, TM6 and TM7 are critical determinants in the binding of ATP. P2Y2 receptors are fully activated by ATP and UTP, whereas ADP and UDP are much less effective agonists. Expression of P2Y2 receptor mRNA and protein has been detected in many peripheral tissues. UTP is the most potent activator of the recombinant human P2Y4 receptor. In human and mouse, P2Y4 mRNA and protein was most abundant in the intestine, but was also detected in other organs. The mouse, rat and human P2Y6 receptors are UDP receptors. A wide tissue distribution of P2Y6 mRNA and protein has been demonstrated, with the highest expression in spleen, intestine, liver, brain and pituitary. ATPγS is a more potent agonist at the P2Y11 receptor than ATP. ADP is the natural agonist of the P2Y12 receptor. It is heavily expressed in platelets where it is the molecular target of the active metabolite of the antiplatelet drug clopidogrel. The P2Y13 ADP-sensitive receptor is strongly expressed in the spleen, followed by placenta, liver, heart, bone marrow, monocytes, T-cells, lung and various brain regions. The P2Y14 receptor is activated by UDP, UDPglucose as well as UDP-galactose, UDP-glucuronic acid and UDP-N-acetylglucosamine. The formation of oligomers by P2Y receptors is likely to be widespread and to greatly increase the diversity of purinergic signaling. P2X receptors are often expressed in the same cells as P2Y receptors. Thus, there is the possibility of bi-directional cross-talk between these two families of nucleotide-sensitive receptors. P2X receptors in general mediate fast neurotransmission, but are sometimes located prejunctionally to mediate increase in release of cotransmitters, for example glutamate in terminals of primary afferent neurons in the spinal cord. P2Y receptors are particularly involved in prejunctional inhibitory modulation of transmitter release, as well as cell proliferation. P2Y1,2,4,6 receptors have been described on subpopulations of sympathetic neurons, P2Y2 and P2Y4 receptors in intracardiac ganglia, P2Y1 and P2Y2 receptors on sensory neurons while P2Y1 receptors appear to be the dominant subtype on enteric neurons. P2Y2 (and/or P2Y4) receptors are expressed on enteric glial cells.

CONCLUSIONS Purinergic neurotransmission is now widely established for both autonomic neuromuscular transmission to smooth muscle and synaptic transmission in ganglia and it is a rapidly expanding field. There is particular interest in the physiology and pathophysiology of purinergic signalling in autonomic systems and therapeutic interventions are being explored [17]. The autonomic nervous system shows marked plasticity: that is, the expression of cotransmitters and receptors show dramatic changes during development and ageing, in nerves that remain after trauma or surgery and in disease conditions. There are several examples where the purinergic component of cotransmission is increased in pathological conditions. The parasympathetic purinergic nerve-mediated component of contraction of the human bladder is increased to 40% in pathophysiological conditions such as interstitial cystitis, outflow obstruction, idiopathic instability and also some types of neurogenic bladder. ATP also has a significantly greater cotransmitter role in sympathetic nerves supplying hypertensive compared to normotensive blood vessels. Receptors for purines and pyrimidines have been cloned and characterized. There are 4 subtypes of P1(adenosine) receptors, 7 subtypes of P2X ion channel receptors and 8 subtypes of P2Y G protein-coupled receptors. These are widely distributed in non-neuronal cells as well as neurons.

References [1] Burnstock G, Campbell G, Satchell D, Smythe A. Evidence that adenosine triphosphate or a related nucleotide is the transmitter substance released by non-adrenergic inhibitory nerves in the gut. Br J Pharmacol 1970;40:668–88. [2] Burnstock G. Purinergic nerves. Pharmacol Rev 1972;24:509–81. [3] Burnstock G. Physiology and pathophysiology of purinergic neurotransmission. Physiol Rev 2007;87:659–797. [4] Burnstock G. Do some nerve cells release more than one transmitter? Neuroscience 1976;1:239–48. [5] Zimmermann H. Ectonucleotidases: some recent developments and a note on nomenclature. Drug Dev Res 2001;52:44–56. [6] Burnstock G, Verkhratsky A. Evolutionary origins of the purinergic signalling system. Acta Physiologica 2009;195:415–47. [7] Burnstock G, Knight GE. Cellular distribution and functions of P2 receptor subtypes in different systems. Int Rev Cytol 2004;240:31–304. [8] Burnstock G. Purine-mediated signalling in pain and visceral perception. Trends Pharmacol Sci 2001;22:182–8. [9] Burnstock G, Verkhratsky A. Long-term (trophic) purinergic signalling: purinoceptors control cell proliferation, differentiation and death. Cell Death Dis 2010;1:e9. [10] Burnstock G. A basis for distinguishing two types of purinergic receptor. In: Straub RW, Bolis L, editors. Cell Membrane Receptors for Drugs and Hormones: A Multidisciplinary Approach. New York: Raven Press; 1978. p. 107–18. [11] Burnstock G, Kennedy C. Is there a basis for distinguishing two types of P2-purinoceptor? Gen Pharmacol 1985;16:433–40. [12] Abbracchio MP, Burnstock G. Purinoceptors: are there families of P2X and P2Y purinoceptors? Pharmacol Therap 1994;64:445–75. [13] Burnstock G. Purine and pyrimidine receptors. Cell Mol Life Sci 2007;64:1471–83. [14] North RA. Molecular physiology of P2X receptors. Physiol Rev 2002;82:1013–67.

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CoNClusIoNs

[15] Kawate T, Michel JC, Birdsong WT, Gouaux E. Crystal structure of the ATP-gated P2X4 ion channel in the closed state. Nature 2009;460:592–8. [16] Abbracchio MP, Burnstock G, Boeynaems J-M, Barnard EA, Boyer JL, Kennedy C, et al. International Union of Pharmacology. Update on the P2Y G protein-coupled nucleotide receptors: from molecular mechanisms and pathophysiology to therapy. Pharmacol Rev 2006;58:281–341. [17] Burnstock G. Pathophysiology and therapeutic potential of purinergic signaling. Pharmacol Rev 2006;58:58–86. [18] Burnstock G. Introduction: ATP and its metabolites as potent extracellular agonists. In: Schwiebert EM, editor. Current Topics in Membranes, vol 54. Purinergic Receptors and Signalling. San Diego: Academic Press; 2003. p. 1–27. [19] Burnstock G. The changing face of autonomic neurotransmission. (The First von Euler Lecture in Physiology). Acta Physiol Scand 1986;126:67–91. [20] Burnstock G, Wong H. Comparison of the effects of ultraviolet light and purinergic nerve stimulation on the guinea-pig taenia coli. Br J Pharmacol 1978;62:293–302.

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[21] Burnstock G, Cocks T, Crowe R, Kasakov L. Purinergic innervation of the guinea-pig urinary bladder. Br J Pharmacol 1978;63:125–38. [22] Burnstock G, Costa M. Adrenergic Neurones: Their Organization, Function and Development in the Peripheral Nervous System. London: Chapman and Hall; 1975. pp. 1–225 [23] Sneddon P, Burnstock G. Inhibition of excitatory junction potentials in guinea-pig vas deferens by α,β-methylene-ATP: further evidence for ATP and noradrenaline as cotransmitters. Eur J Pharmac 1984;100:85–90. [24] Brake AJ, Wagenbach MJ, Julius D. New structural motif for ligandgated ion channels defined by an ionotropic ATP receptor. Nature 1994;371:519–23. [25] Barnard EA, Burnstock G, Webb TE. G protein-coupled receptors for ATP and other nucleotides: a new receptor family. Trends Pharmacol Sci 1994;15:67–70.

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C H A P T E R

19 Adenosine Receptors and Autonomic Regulation Italo Biaggioni

Adenosine is an endogenous nucleoside formed by the degradation of adenosine-triphosphate (ATP) during energy-consuming processes. Adenosine modulates many physiological processes through activation of four subtypes of G-protein coupled membrane P1 purinergic receptors, A1, A2A, A2B and A3. Its physiological importance depends on the affinity of these receptors and the extracellular concentrations reached. Extracellular concentrations of adenosine are usually low, but adenosine may have tonic actions even during physiological conditions, mostly through activation of high affinity A2A and A1 receptors. Theoretically, adenosine can be formed within the synapse, from the degradation of ATP that is released as a co-transmitter (see Chapter 18). Significant increases of extracellular adenosine, however, occur mostly in ischemic tissues, when energy demands exceed oxygen supply. Thus, adenosine is considered a “retaliatory” metabolite, whose actions are physiologically relevant during ischemic conditions. Adenosine has perhaps the shortest half-live of all autacoids, particularly in humans. It is rapidly and extensively metabolized to inactive inosine by adenosine deaminase. It is also quickly transported back into cells by an energydependent uptake mechanism, which is part of a purine salvage pathway designed to maintain intracellular levels of ATP. The effectiveness of this adenosine transport system is species-dependent. It is particularly active in humans, and is mainly responsible for the extremely short half-life of adenosine in human blood, which is probably less than one second. Adenosine mechanisms are the target of commonly used drugs. Dipyridamole (Persantin, Aggrenox) acts by blockade of adenosine reuptake, thus potentiating its actions. Conversely, caffeine and theophylline are antagonists of adenosine receptors. Adenosine receptors are ubiquitous and, depending on their localization, may mediate opposite effects. This phenomenon is particularly evident in the interaction of adenosine and the autonomic nervous system; adenosine can produce either inhibition or excitation of autonomic

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00019-6

neurons [1]. I will first outline the effects of adenosine in the efferent, central, and afferent autonomic pathways, emphasizing those with clinical relevance. I will then propose an integrated view that may explain how these seemingly contradictory effects may work together. ATP shares many of the modulatory effects of adenosine on autonomic function and we will discuss these similarities when appropriate. A comprehensive review of the effects of ATP and its P2 purinergic receptors is found in Chapter 18.

POSTSYNAPTIC ANTI-ADRENERGIC EFFECTS OF ADENOSINE Adenosine A1 receptors are found in target organs innervated by the sympathetic nervous system. A1 receptors are coupled to inhibition of adenylate cyclase and their effects are opposite to those of β-adrenoreceptor agonists. For example, adenosine will oppose β-mediated tachycardia and lipolysis. This phenomenon is translated functionally as an “anti-adrenergic” effect. The physiological relevance of this effect is not entirely clear, but some studies suggest that adenosine is more effective in reducing heart rate during isoproterenol-induced tachycardia than in the baseline state or during atropine-induced tachycardia.

PRESYNAPTIC EFFECTS OF ADENOSINE ON EFFERENT NERVES AND GANGLIONIC TRANSMISSION Adenosine inhibits the release of neurotransmitters through putative presynaptic A1 receptors in both the brain and the periphery. Blockade of forearm adenosine receptors with intrabrachial theophylline potentiates sympathetically mediated forearm vasoconstriction, suggesting that endogenous adenosine inhibits noradrenergic neurotransmission in vivo in humans.

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A few studies have investigated the effects of adenosine on ganglionic neurotransmission, and most of them show an inhibitory effect. Adenosine inhibits the release of acetylcholine presynaptically and blocks calcium current postynaptically in ganglia.

ADENOSINE AND CENTRAL AUTONOMIC REGULATION Adenosine acts as a neuromodulator within the central nervous systems, mostly through interaction with A1 and A2A receptors. Of particular relevance to this review are its actions on brainstem nuclei involved in autonomic cardiovascular regulation. In general, the central actions of adenosine result in inhibition of sympathetic tone through complex, and incompletely understood, mechanisms of action. Microinjection of adenosine into the nucleus tractus solitarii (NTS) evokes a dose-related decrease in blood pressure, heart rate, and renal sympathetic nerve activity. These effects appear to be mediated, at least partially, through A2A receptors. The NTS is the site of the first synapse of afferent fibers arising from baroreceptors. The NTS provides excitatory input to the caudal ventrolateral medulla, which in turn provides inhibitory input to the rostro ventrolateral medulla (RVLM), where sympathetic activity is thought to originate. Thus, stimulation of baroreceptor afferents (e.g., by an increase in blood pressure) activates the NTS and this results in inhibition of the RVLM and a reduction in sympathetic tone. The effect of adenosine in the NTS is similar to that of the excitatory neurotransmitter glutamate, implying that adenosine has excitatory neuromodulatory effects on the NTS. The precise mechanisms that explain this phenomenon are not known. It has been proposed that adenosine releases glutamate within the NTS [2,3], or blunts the release of the inhibitory neuromodulator GABA. The effects of adenosine in the NTS are also blunted by microinjection of the nitric oxide synthase inhibitor L-NAME, suggesting an interaction between adenosine and nitric oxide in the NTS. Regardless of the mechanism of action, microinjections of adenosine receptor antagonists into the NTS results in blunting of the baroreflex gain, suggesting a role of endogenous adenosine on central cardiovascular regulation. A1 and A2A receptors are also found in the RVLM and may modulate neuronal activity either directly, or through inhibition of GABA release [4]. ATP and P2 receptors are also important in central autonomic regulation of cardiovascular and respiratory functions [5].

carotid body chemoreceptors, renal afferents, and myocardial and skeletal muscle afferents. The neuroexcitatory actions of adenosine were first recognized in animals in the early 1980s when it was found that adenosine activates arterial chemoreceptors in rats and cats, and renal afferents in rats and dogs. The functional relevance of these findings, however, was not apparent until human studies were done. The most striking effect of intravenous adenosine in humans is a dramatic stimulation of respiration and sympathetic activation. This effect can be explained by carotid body chemoreceptor activation because it is observed when adenosine is injected into the aortic arch at a site proximal to the origin of the carotid arteries, but not if adenosine is injected into the descending aorta. Activation of vagal C-fibers may also contribute to adenosine-induced dyspnea. The effects of intravenous adenosine are dramatically different if autonomic nervous system is absent. For example, adenosine lowers blood pressure and heart rate in autonomic failure patients. Pain resembling angina has been reported with intravenous and intracoronary administration of adenosine presumably because of activation of sensory afferents. Intracoronary adenosine also elicits a pressor reflex in humans, which may be explained by activation of myocardial afferents. The few animal studies that have tested this hypothesis have yielded conflicting results, perhaps because of the confounding effects of anesthesia or to species differences. There is also controversy over whether adenosine activates skeletal muscle chemoreceptors that trigger sympathetic activation in response to ischemic exercise. Studies favoring [6] or opposing [7] the possibility that adenosine is a metabolic trigger of the exercise pressor reflex have been published. ATP has also been implicated as a trigger of skeletal muscle afferent activation [8]. In summary, whereas adenosine inhibits sympathetic efferents, it activates afferent nerves, including arterial chemoreceptors (in animals and humans), renal afferents (animals), and possibly cardiac and muscle afferents (humans). The autonomic excitatory actions of adenosine clearly predominate when adenosine is given intravenously to conscious human subjects [9]. This sympathetic activation may explain the usefulness of intravenous adenosine as a challenge test to diagnose neurogenic syncope during tilt table tests [10]. It remains speculative whether or not endogenous adenosine plays a role in the generation of spontaneous neurogenic syncope. The adenosine receptor antagonist theophylline is used in the treatment of neurogenic syncope, but controlled studies are lacking.

NEUROEXCITATORY ACTIONS OF ADENOSINE ON AFFERENT PATHWAYS

INTEGRATED VIEW OF ADENOSINE AND CARDIOVASCULAR AUTONOMIC REGULATION

In contrast to the “inhibitory” actions of adenosine in efferent pathways, adenosine excites a variety of afferent fibers that evoke systemic sympathetic activation including

These neuroexcitatory actions seem at odds with the postulated protective role of adenosine, which heretofore has been assigned to its “inhibitory” effects. We postulate,

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Sympathetic activation NE NE Systemic

Systemic vasoconstriction blood pressure perfusion pressure

NE

III Local inhibition of NE release and F– vasodilation I Adenosine

Local

F+ II. Activation of afferent fibers

ATP

Ischemic exercise

FIGURE 19.1 Postulated modulation by purines (adenosine and ATP) of autonomic cardiovascular regulation. See text for details.

however, that the neuroexcitatory actions of adenosine work in tandem with its inhibitory effects to provide protection against ischemia. It is also likely that ATP, in addition to or instead of adenosine, participate in these actions. This framework is presented in Figure 19.1. 1. Interstitial levels of adenosine increase under conditions of increased metabolic demand (exercise) and decreased energy supply (ischemia), reaching physiologically relevant concentrations. ATP can also be released as a co-transmitter from noradrenergic neurons. 2. Adenosine (or ATP) then activates sensory afferent fibers that produce pain, and muscle or myocardial afferents (metaboreceptors) that trigger an ischemic pressor reflex. Pain sensation is a primordial defense mechanism that signals the individual to stop exercising. Sympathetic activation leads to systemic vasoconstriction, increase in blood pressure and improved perfusion pressure. 3. This systemic vasoconstriction would be deleterious to the ischemic organ, if it were not for the simultaneous local inhibitory actions of adenosine that produce vasodilation and inhibit norepinephrine (NE) release. These actions are, for the most part, circumscribed to the local ischemic tissue so that it is protected from sympathetically-mediated vasoconstriction while it benefits from the improved perfusion pressure. ATP has been proposed as a mediator of this “functional sympatholysis” [11]. We propose, therefore, that the excitatory actions of adenosine and ATP work in tandem with their inhibitory effects to provide local protection against ischemia, even at the expense of the rest of the organism. Furthermore, we propose that adenosine (and ATP) provides a link between

local mechanisms of blood flow autoregulation and systemic mechanisms of autonomic cardiovascular regulation, heretofore thought to work independent from each other.

References [1] Biaggioni I. Contrasting excitatory and inhibitory effects of adenosine in blood pressure regulation. Hypertension 1992;20:457–65. [2] Phillis JW, Scislo TJ, O’Leary DS. Purines and the nucleus tractus solitarius: effects on cardiovascular and respiratory function. [Review] [29 refs]. Clin Exp Pharmacol Physiol 1997;24:738–42. [3] Biaggioni I, Mosqueda-Garcia R. Adenosine in cardiovascular homeostasis and the pharmacological control of its activity. In: Laragh J, Brenner BM, editors. Hypertension: pathophysiology, managements and diagnosis. New York: Raven Press; 1995. [4] Spyer KM, Thomas T. A role for adenosine in modulating cardiorespiratory responses: a mini-review. Brain Res Bull 2000;53:121–4. [5] Gourine AV, Wood JD, Burnstock G. Purinergic signalling in autonomic control. Trends Neurosci 2009;32:241–8. [6] Costa F, Diedrich A, Johnson B, Sulur P, Farley G, Biaggioni I. Adenosine, a metabolic trigger of the exercise pressor reflex in humans. Hypertension 2001;37:917–22. [7] Cui J, Leuenberger UA, Blaha C, Yoder J, Gao Z, Sinoway LI. Local adenosine receptor blockade accentuates the sympathetic responses to fatiguing exercise. Am J Physiol Heart Circ Physiol 2010;298:H2130–H2137. [8] McCord JL, Tsuchimochi H, Kaufman MP. P2X2/3 and P2X3 receptors contribute to the metaboreceptor component of the exercise pressor reflex. J Appl Physiol 2010;109:1416–23. [9] Biaggioni I, Killian TJ, Mosqueda-Garcia R, Robertson RM, Robertson D. Adenosine increases sympathetic nerve traffic in humans. Circulation 1991;83:1668–75. [10] Shen WK, Hammill SC, Munger TM, Stanton MS, Packer DL, Osborn MJ, et al. PA Adenosine: potential modulator for vasovagal syncope. J Am Coll Cardiol 1996;28:146–54. [11] Rosenmeier JB, Yegutkin GG, Gonzalez-Alonso J. Activation of ATP/UTP-selective receptors increases blood flow and blunts sympathetic vasoconstriction in human skeletal muscle. J Physiol 2008;586:4993–5002.

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20 Nitric Oxide and Autonomic Regulation Alfredo Gamboa GMP causes activation of cyclic GMP-dependent protein kinases, with reduction of intracellular Ca2 and a decrease in the sensitivity of contractile elements to Ca2.

Since the discovery that nitric oxide (NO) was a naturally occurring signal molecule in blood vessels, it has become one of the most widely studied substances in biology. Now it is recognized as an endogenous mediator of numerous physiological processes ranging from regulation of cardiovascular function to memory formation. NO is probably one of the most important metabolic modulator of blood pressure; our previous studies suggest that NO tonically restrains blood pressure by at least 30 mmHg in healthy young adults [1]. The autonomic nervous system is a major regulatory mechanism for both short and long term adjustments of the cardiovascular system in general, and blood pressure in particular. In addition to its control of local vasomotor tone and arterial pressure, NO has been proposed to also modulate autonomic regulation of blood pressure through interactions with autonomic pathways in the central nervous system and at peripheral sites.

CENTRAL NO–AUTONOMIC NERVOUS SYSTEM INTERACTIONS In the central nervous system, nitric oxide functions mainly as a neuromodulator. Nitric oxide can have both central neural sympathoinhibitory and sympathoexcitatory actions. In humans, intravenous infusion of L-NMMA (a competitive stereospecific non-selective inhibitor of NOS) induces an increase of mean arterial pressure of about 10% without the expected compensatory inhibition of sympathetic firing rate mediated by baroreflex loading. In comparison, the same increase in blood pressure induced by phenylephrine produced a decrease of about 50% in sympathetic firing rate. When sodium nitroprusside was given in addition to L-NMMA in order to prevent the increase in mean arterial blood pressure, muscle sympathetic nerve activity increased more than double its baseline values [3]. Furthermore, it has been shown that acute and/or chronic (3 weeks) intracerebroventricular injection of L-NMMA, at a dose ineffective to produce systemic effects; increases mean arterial pressure in normotensive rats [4]. Taken together these findings suggest that NO tonically inhibits central sympathetic tone. Thus, NOS inhibition increases blood pressure in part by increasing sympathetic firing rate but this effect is partially masked by the baroreflex. The excitatory effects of NO on neural sympathetic outflow, and their physiological relevance, are less well understood. In rat brain slices, L-arginine, the substrate to produce NO, stimulates the neuronal activity of some nucleus tractus solitarius neurons and this effect can be blocked by co-infusion of L-NMMA [5]. It should be noted that the NTS provides inhibitory input (through the caudal ventrolateral medulla) to the rostral ventrolateral medulla (RVLM) where sympathetic tone is thought to originate. Therefore, neuroexcitation of NTS neurons by NO may contribute to its tonic inhibition of sympathetic tone. On the other hand, in vivo studies in rabbits have shown that NO has an excitatory effect on renal

NO SYNTHESIS AND ACTIONS NO is produced from L-arginine by three isoforms of the enzyme nitric oxide synthase (NOS). An inducible form is found mostly in macrophages (iNOS). Two isoforms are expressed constitutively, one in epithelial and neural cells (nNOS), and another in endothelial cells, platelets, and myocardial cells (eNOS). Both eNOS and nNOS consists of a C-terminal reductase domain and an N-terminal oxygenase domain, differing only in one aminoacid, namely, Asp597 in nNOS is Asn368 in eNOS [2]. The monomer of NOS is inactive, while the dimer is its active form and the dimerization requires tetrahydrobiopterin (BH4), heme and L-arginine binding. Overall, NOS catalyzes the oxidation of L-arginine to generate citrulline and NO. The synthesis of nitric oxide by these NOS isoforms is inhibited non-selectively by L-arginine analogs, including NG-monomethyl-L-arginine (L-NMMA), NG-nitro-Larginine (L-NA), L-NA methylester (L-NAME), and asymmetric dimethylarginine (ADMA). There are also selective nNOS inhibitors such as 7-nitroindazol (7-NI), S-methylL-thiocitrulline (SMTC) and N[omega]-propyl-L-arginine. Once formed, NO activates soluble guanylyl cyclase and produces cyclic guanosine monophosphate (GMP) from guanosine triphosphate (GTP). Accumulation of cyclic

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FIGURE 20.1 Schematic illustration of the effects of NO and of NOS inhibition on blood pressure regulation both centrally and in the periphery.

sympathetic preganglionic neurons [6]. Additionally, in eNOS/KO mice, selective inhibition of neural NOS produces a drop in blood pressure and it has been hypothesized that NO released in the central nervous system and/or the baroreceptor pathways may increase sympathetic nerve activity and increase blood pressure, while the inhibition of NO may have the opposite effect [7]. In summary, nitric oxide in the central nervous system may have both excitatory and inhibitory effects on sympathetic activity, but the evidence from human studies suggest that tonic inhibition of sympathetic tone by NO predominates and may contribute to its role in lowering blood pressure.

NO–AUTONOMIC NERVOUS SYSTEM INTERACTIONS IN THE PERIPHERY In addition to its role as a central modulator of sympathetic activity, NO may also modulate sympathetic vasoconstrictor tone peripherally. Nitric oxide synthase activity has been localized by immunohistochemistry in preganglionic autonomic fibers and postganglionic parasympathetic nerves innervating vascular smooth muscle. There is

evidence that NO released from nitrergic nerves interferes with the release of norepinephrine from adrenergic nerve terminals and with its vasoconstrictive actions on smooth muscle. In anesthetized paralyzed, baroreflex denervated cats, systemic NOS inhibition by L-NAME produced a greater pressor response compared to intact cats [8]. In humans undergoing thoracic sympathectomy, the vasoconstrictor response to systemic infusion of L-NMMA has been evaluated and it has been found that sympathectomy potentiates the vasoconstrictor effect of NOS inhibition only in the denervated limb. This effect is thought to be NO-dependent since a similar increase in forearm vasoconstriction in both limbs was observed during phenylephrine infusion [9]. NO is tonically produced by endothelial cells and in disease states (e.g. during inflammation) also possibly by vascular smooth muscle cells (iNOS) to induce local vasodilatation and, simply through this mechanism, counteract sympathetically-mediated vasoconstriction. NO can be formed locally by either eNOS and nNOS and their relative contribution to endothelial dependent vasodilatation remains a work in progress driven by the availability of selective nNOS inhibitors. The importance of neuronally derived NO in regulating basal vascular tone independently

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NO–AuTONOmIC NERvOus sysTEm INTERACTIONs IN HEART RATE CONTROl

of eNOS has been recently studied in humans [10] and in isolated mouse aorta [11]. These studies suggest a role for neurally derived NO in regulating basal vascular tone. In summary, NO acting in the CNS places a brake on sympathetic outflow decreasing sympathetic vasoconstriction. In the periphery, NO produced locally from endothelial and vascular smooth muscle cells, or nitrergic nerves innervating them, produces vasodilatation and counteracts sympathetically-mediated vasoconstriction. NOS inhibition has been shown to produce an increase in sympathetic outflow as well as an increase in NE release. These effects however are masked by the baroreflex. Thus the final effect of NOS inhibition on blood pressure will be the resultant of these effects (Figure 20.1).

NO–AUTONOMIC NERVOUS SYSTEM INTERACTIONS IN HEART RATE CONTROL In addition to modulation of vascular tone, NO has been implicated in the control of cardiac function but its exact role is not completely understood. The effects of NO on heart rate modulation appear to be not merely a baroreflex-mediated response to vasodilatation. NO donors have been shown to elicit a positive chronotropic response [12] that seems to be at least in part independent of the autonomic nervous system since this effect is also seen in heart rate transplant recipients with cardiac denervation [13]. Inhibition of NOS with non-isoform specific inhibitors has shown either no effect on baseline heart rate or a modest negative chronotropic effect. Chronic inhibition of NOS in animal models results in sustained bradycardia. Studies on eNOS/ or nNOS/ KO mice show conflicting results and it may be a reflection on the complex interactions between the different NOS isoforms, the regulation of heart rate by the autonomic nervous system and the specific localization of the different NOS isoforms.

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References [1] Gamboa A, Shibao C, Diedrich A, Choi L, Pohar B, Jordan J, et al. Contribution of endothelial nitric oxide to blood pressure in humans. HTN 2007;49:170–7. [2] Flinspach ML, Li H, Jamal J, Yang W, Huang H, Hah JM, et al. Structural basis for dipeptide amide isoform-selective inhibition of neuronal nitric oxide synthase. Nat Struct Mol Biol 2004;11:54–9. [3] Owlya R, Vollenweider L, Trueb L, Sartori C, Lepori M, Nicod P, et al. Cardiovascular and sympathetic effects of nitric oxide inhibition at rest and during static exercise in humans. Circulation 1997;96:3897–903. [4] Sakima A, Teruya H, Yamazato M, Matayoshi R, Muratani H, Fukiyama K. Prolonged NOS inhibition in the brain elevates blood pressure in normotensive rats. Am J Physiol 1998;275:R410–417. [5] Tagawa T, Imaizumi T, Harada S, Endo T, Shiramoto M, Hirooka Y, et al. Nitric oxide influences neuronal activity in the nucleus tractus solitarius of rat brainstem slices. Circ Res 1994;75:70–6. [6] Hakim MA, Hirooka Y, Coleman MJ, Bennett MR, Dampney RA. Evidence for a critical role of nitric oxide in the tonic excitation of rabbit renal sympathetic preganglionic neurones. J Physiol 1995;482:401–7. Pt 2 [7] Kurihara N, Alfie ME, Sigmon DH, Rhaleb NE, Shesely EG, Carretero OA. Role of nNOS in blood pressure regulation in eNOS null mutant mice. HTN 1998;32:856–61. [8] Zanzinger J, Czachurski J, Seller H. Inhibition of sympathetic vasoconstriction is a major principle of vasodilation by nitric oxide in vivo. Circ Res 1994;75:1073–7. [9] Lepori M, Sartori C, Duplain H, Nicod P, Scherrer U. Sympathectomy potentiates the vasoconstrictor response to nitric oxide synthase inhibition in humans. Cardiovasc Res 1999;43:739–43. [10] Melikian N, Seddon MD, Casadei B, Chowienczyk PJ, Shah AM. Neuronal nitric oxide synthase and human vascular regulation. Trends Cardiovasc Med 2009;19:256–62. [11] Capettini LS, Cortes SF, Lemos VS. Relative contribution of eNOS and nNOS to endothelium-dependent vasodilation in the mouse aorta. Eur J Pharmacol 2010;643:260–6. [12] Musialek P, Lei M, Brown HF, Paterson DJ, Casadei B. Nitric oxide can increase heart rate by stimulating the hyperpolarizationactivated inward current, I(f). Circ Res 1997;81:60–8. [13] Chowdhary S, Harrington D, Bonser RS, Coote JH, Townend JN. Chronotropic effects of nitric oxide in the denervated human heart. J Physiol 2002;541:645–51.

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21 Glutamatergic Neurotransmission Deborah Bauer, Michael Robinson Unlike the neuromuscular junction, where acetylcholine mediates rapid excitatory signaling, glutamate mediates essentially all rapid excitatory signaling in the mammalian CNS. This excitatory signaling is counterbalanced by GABA- or glycine-mediated inhibition (see Chapter 22). The levels of glutamate are 1,000 to 10,000fold higher than most of the other “classical” neurotransmitters, including dopamine, serotonin, norepinephrine, and acetylcholine; they approach 10 mmol/Kg. Glutamatedependent signaling is required for essentially all sensory and motor processing. It contributes to neuronal migration and synapse formation during development. In addition, there is substantial evidence to suggest that plasticity of excitatory synapses contributes to memory formation [1]. In considering glutamate as a neurotransmitter, it may be helpful to remember that glutamate has multiple functions that are not necessarily unique to the nervous system. It is an amino acid and is incorporated into proteins. It is one metabolic step from α-ketoglutarate, an intermediate in the tricarboxylic acid cycle. This cycle is responsible for ~90% of the ATP generated from glycolysis. In the brain there is no intact urea cycle; waste nitrogen (ammonia) is incorporated into glutamate with the subsequent export of glutamine to the periphery. Glutamate is the only precursor for the inhibitory neurotransmitter, GABA. In addition, there are other substances in the brain, such as aspartate, that may serve as excitatory neurotransmitters in the CNS using many of the same receptors as glutamate. For the purposes of this review, we will refer to all of this as glutamatergic neurotransmission. From a disease perspective, stroke and head trauma cause an increase in extracellular glutamate, excessive activation of glutamate receptors, and cell death [2]. This form of cell death is referred to as “excitotoxicity”. Remarkably, evidence for an excitotoxic contribution to stroke-induced brain damage first emerged in the late 1980s, and no effective therapies have emerged in spite of significant efforts by pharmaceutical firms. Excitotoxicity may also contribute to the cell loss that accompanies many neurodegenerative diseases, including Alzheimer’s disease, Parkinson’s disease, Huntington’s disease and amyotrophic lateral sclerosis [2]. An imbalance of excitatory and inhibitory signaling contributes to seizure disorders, and glutamate receptor antagonists reduce seizure activity [3]. Finally,

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there is significant evidence that altered glutamatergic function contributes to psychiatric conditions and to developmental disorders. Several drugs targeting specific glutamate receptors have entered clinical trials for psychiatric conditions (anxiety, depression, schizophrenia) and Fragile X syndrome [4,5]. While therapeutics targeting glutamatergic neurotransmission are promising, it may be difficult to target glutamate in specific pathways without altering the myriad signals that are required for normal brain function. In this chapter, we provide an overview of our current understanding of glutamate signaling in the nervous system and describe a few specific roles for glutamate in the autonomic nervous system. A simplified schematic of an excitatory synapse is provided (Fig. 21.1).

SYNTHESIS AND VESICULAR RELEASE Although glutamate is available in the diet, it cannot efficiently cross the intact blood brain barrier and rapid signaling requires a robust mechanism to replenish vesicular pools [6]. Although there are several possible routes for glutamate synthesis, it is generally thought that neurotransmitter pools of glutamate are generated from glutamine by glutaminase [6]. Glutamate can also be synthesized from α-ketoglutarate by glutamate dehydrogenase [6]. There are also other routes for glutamate synthesis from proline or from the dipeptide N-acetyl-alpha-L-aspartyl-L-glutamate (NAAG), but these are not thought to be robust sources of neurotransmitter pools of glutamate [7]. After synthesis, glutamate is actively transported into vesicles by a family of vesicular glutamate transporters (VGLUT1-3) [8]. These transporters utilize a proton electrochemical gradient to actively accumulate glutamate in synaptic vesicles to concentrations estimated to be as high as 100 mM. None of these vesicular transporters can package aspartate, but recently a distant member of the same gene family was shown to transport aspartate [9]. Interestingly, VGLUT1 and 2 have a complementary distribution in the nervous system with VGLUT1 found in the cerebrum and cerebellum and VGLUT2 enriched in brainstem areas. It is not known why there are two VGLUTs, but it has been suggested that the rhythmic activity of

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PRESYNAPTIC NEURON Gln Gln

Glutaminase

Glu

Glutamine Synthetase

VGLUTs

Glutamine Dehydrogenase

ASTROCYTE

Krebs Cycle

GLAST

mGluRII,III

mGluRII,III

Glu

Kainate Glu

Glu

GLT-1

Kainate

Glu

Glu Glu

GLT-1

EAAC1

mGluRI

Glu

AMPA NMDA

mGluRII

Gammaglutamylcysteine + Glutathione Synthetase

EAAT4

Glutathione

POSTSYNAPTIC NEURON

FIGURE 21.1 Schematic of the glutamate synapse. (Adapted from [27].) Glutamate is synthesized from glutamine with the enzyme glutaminase. Glutamate is packaged into presynaptic vesicles by vesicular glutamate transporters (VGLUTs) and released into the synaptic cleft where it can activate NMDA, AMPA, kainate, and metabotropic glutamate receptors (mGluRs). Glutamate is cleared from the synapse by the plasma membrane glutamate transporters EAAC1 in neurons, and GLAST and GLT-1 in astrocytes. Glutamate can then enter the Krebs cycle, be converted into glutathione, or be converted back into glutamine for the cycle to begin again.

brainstem neurons necessitates a specific vesicular transporter. VGLUT3 is mostly found in presynaptic termini of neurons that are not classically considered glutamatergic, including those that utilize serotonin, acetylcholine, and GABA [8]. This provides further evidence that Dale’s hypothesis, postulating that a neuron only uses one neurotransmitter, is no longer supported by experimental findings.

RECEPTORS Once released into the synapse, glutamate activates ionotropic (ligand-gated ion channel) or metabotropic (G-protein-coupled) glutamate receptors, called iGluRs or mGluRs. iGluRs are named for exogenous agonists that selectively activate subtypes of glutamate receptors, including N-methyl-aspartate (NMDA), α-amino-3hydroxyl-5-methyl-4-isoxazole-propionate (AMPA), and kainate [10]. Both groups of receptors are found on

various aspects of the synapse, including the postsynaptic bouton, presynaptic terminus, and nearby astrocytes. AMPA receptors are ligand-gated channels permeable to sodium and potassium, and under some circumstances calcium [11]. Thus, activation of AMPA receptors causes depolarization of the postsynaptic membrane. AMPA receptors are tetramers comprised of the subunits GluR1-4 [10]. These receptors are responsible for the majority of fast glutamate-mediated neurotransmission [2]. Like AMPA receptors, kainate receptors are also ligand-gated channels permeable to sodium and potassium [2]. Kainate receptors are comprised of GluR5-7 and KA1-KA2 [2]. They can exist as homotetramers or heterotetramers [10]. NMDA receptors are ligand-gated channels permeable to Na, K, and Ca2 [11]. Activation of these receptors requires two compounds, glutamate and glycine (or possibly D-serine) [2]. Therefore, in addition to its accepted role as an inhibitory neurotransmitter in brainstem and spinal cord, glycine also contributes to excitatory signaling. NMDA receptors are comprised of four subunits, two

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of which must be NR1 subunits, and the other two can be comprised of either NR2 or NR3 subunits [2]. Thus, it is a dimer of dimers. The binding of both glutamate and glycine is not sufficient to activate NMDA receptors when the membrane is near 70 mV because physiologic concentrations of Mg2 bind to and block channel function [2]. This effect of magnesium is voltage-dependent, such that depolarization removes the Mg2 from the channel and allows current to pass through the channel. Thus, the NMDA receptor is considered a “coincidence detector”, it is activated when another receptor causes sufficient membrane depolarization, as would be expected to occur with high frequency stimulation. This unique property of the NMDA receptor is linked to synaptic plasticity that may underlie memory formation [12]. In addition to the ionotropic receptors, there also exist a series of seven transmembrane domain metabotropic glutamate receptors (mGluR1-8) [2]. These receptors are grouped into three families, based on the sequence similarity and their signaling properties [2]. Type I receptors include mGluR1 and 5. These receptors primarily activate phospholipase C and IP3 production. Type II receptors includes mGluR2 and 3. Type III receptors includes mGluR4, 6, 7, and 8. These two families of receptors primarily signal through inhibition of adenylate cyclase and a reduction in cAMP levels. The mGluRs are situated on all aspects of the synapse and might be considered sensors of glutamate; they regulate neurotransmitter release from the presynaptic nerve terminal, iGluR channel opening, and aspects of astrocytic function [13].

EAAC1/EAAT3 expression is enriched in neurons in the CNS [18]. It is found on postsynaptic excitatory neurons, GABAergic neurons, and on the presynaptic nerve terminal. On GABAergic neurons, it may directly provide glutamate for the synthesis of GABA as a complement to the normal source of carbon backbone, which is thought to be glutamine (see Chapter 22). EAAC1/EAAT3 participates in neuronal import of cysteine, the rate limiting ingredient for the synthesis of glutathione. Interestingly, these transporters also gate Cl– ions to a varying degree. In fact, EAAT4 and EAAT5 are quite slow as transporters, but gate substantial Cl– currents [14,19]. Rather than having significant roles in the clearance of glutamate, these two transporters may serve as receptors in the cerebellum and retina, respectively [19].

CLEARANCE OF GLUTAMATE

GLUTAMATE IN THE AUTONOMIC NERVOUS SYSTEM

Unlike some other neurotransmitters, there is no evidence that glutamate is metabolized by extracellular enzymes [14]; instead, it is cleared by a family of Nadependent transporters. In mammals, there are five related gene products that perform this function, including GLAST, GLT-1, EAAC1 (called EAAT1-3, respectively in human), EAAT4 and EAAT5 [14]. These transporters exist as homotrimers [15] and co-transport 3 Na and 1 H with each molecule of glutamate and counter-transport 1 K ion. With this stoichiometry, these transporters are capable of generating a 1 million-fold concentration gradient of glutamate across the membrane [16]. The vast majority of glutamate uptake is mediated by GLT-1 and GLAST with GLT-1 having a larger role in forebrain and GLAST a larger role in cerebellum [17]. These transporters are found on many cell types in the nervous system, but are highly enriched on glial cells. Therefore, unlike any of the other classical neurotransmitters that are recycled back into the presynaptic nerve terminal (see chapters on dopamine, serotonin, and norepinephrine; 6, 8, 12 and 17), glutamate is not substantially recycled back into the presynaptic nerve terminal.

GLUTAMATE METABOLISM After clearance into astrocytes, most, if not all of the glutamate carbon backbone is conserved, by conversion to glutamine by the astrocyte specific enzyme, glutamine synthetase [6]. Glutamine is then exported by a specific transporter, called System N [20]. Glutamine is thought to be imported back into the presynaptic nerve terminal and used for regeneration of glutamate. This is referred to as the glutamate-glutamine cycle. Glutamate can also be converted to α-ketoglutarate by either glutamate-dehydrogenase or a transaminase (either aspartate aminotransferase or glutamate oxalacetate transaminase), and enter the citric acid cycle [6].

In contrast to the CNS, glutamate is not the predominant excitatory neurotransmitter in the periphery. The sodium dependent glutamate transporters are found throughout the body and thus are not a good marker for glutamatergic signaling. However, glutamate receptors and vesicular glutamate transporters have been detected in a variety of peripheral tissues including the kidney, bone, heart, intestine, pancreas, and platelets [14,21]. Thus, while their role in signaling has not been fully elucidated, glutamate is involved in signaling outside of the CNS. For example, glutamate is critical in regulation of the enteric nervous system. Glutamatergic neurons innervate the gut and influence both motility and secretion [22]. Both ionotropic and metabotropic glutamate receptors have been identified in the gut. Although the sympathetic and parasympathetic nervous systems innervate the organs using norepinephrine and acetylcholine, respectively, glutamate is important for central control of these systems. The nucleus tractus solitarii (NTS) of the medulla, often considered the command center of the autonomic nervous system, controls

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cardiovascular and digestive tract reflexes and contains both glutamatergic afferents and efferents [23] (see Fig. 21.2). The importance of glutamatergic signaling in the NTS is clearly illustrated in the baroreceptor reflex (see Fig. 21.2). This reflex begins with activation of baroreceptors, pressure sensitive neurons, found in the aortic arch and carotid sinus that project to the NTS [24]. At this point, the reflex splits into two pathways, the cardioinhibitory pathway and the sympathoinhibitory pathway [24]. In the cardioinhibitory pathway, second order glutamatergic neurons in the NTS excite cholinergic neurons in the nucleus ambiguus (NA) that project to the cardiac ganglion. The cardiac ganglion projects cholinergically to the heart and controls heart rate. This activation of muscarinic acetylcholine receptors causes a decrease in heart rate [24]. In the sympathoinhibitory pathway, second order glutamatergic neurons in the NTS excite GABAergic interneurons in caudal ventrolateral medulla (CVL) [24]. These CVL interneurons project to and inhibit glutamatergic neurons in the rostral ventrolateral medulla (RVLM), which in turn innervate preganglionic sympathetic neurons in the spinal cord. These spinal cord neurons release acetylcholine at the sympathetic ganglion. Finally, noradrenergic neurons in the sympathetic ganglion activate vasoconstriction. However, because excitation of GABAergic interneurons in the CVL causes downstream inhibition of the pathway, the net result is decreased

-

GABA

CVL

Glutamate

+

Some estimates suggest that glutamate mediates upwards of 80% of all neuron–neuron communication in the mammalian CNS. It is required for virtually every physiologic function in mammals, including most brainstem pathways that contribute to autonomic control. In addition to normal physiologic function, excessive activation of glutamate receptors contributes to cell death observed in both acute and chronic neurodegenerative diseases. As our understanding of this system evolves, it seems likely that therapeutic targeting of this system will become more sophisticated.

NTS

+

Sympathoinhibitory

Cardioinhibitory

Glutamate

SUMMARY

Glutamate

+

Glutamate

Glutamate

RVLM

vasoconstriction. Thus, activation of baroreceptors activates two homeostatic pathways collectively known as the baroreceptor reflex, causing heart rate and blood pressure to decrease. Another major output of the NTS where glutamate plays a role is the hypothalamus. Glutamatergic activity in the hypothalamus also influences the baroreceptor reflex. Microinjection of glutamate in the paraventricular nucleus (PVN) of the hypothalamus results in baroreflex bradycardia through activation of NMDA receptors [25]. In fact, glutamate is the major excitatory neurotransmitter providing input to the PVN and supraoptic nucleus of the hypothalamus [26].

+

Heart Acetylcholine

Acetylcholine

NA

+ Cardiac Ganglion

-

+

Acetylcholine

+ SC

Sympathetic

Norepinephrine

Ganglion

FIGURE 21.2 Simplified baroreflex circuit. (Adapted from [24].) Baroreceptors in the aortic arch and carotid sinus project to the nucleus tractus solitarii (NTS) and release glutamate. In the cardioinhibitory pathway, second order glutamatergic neurons in the NTS excite cholinergic neurons in the nucleus ambiguus (NA) that project to the cardiac ganglion. Cholinergic neurons of the cardiac ganglion project to the heart and decrease heart rate. In the sympathoinhibitory pathway, second order glutamatergic neurons in the NTS excite GABAergic interneurons in caudal ventrolateral medulla (CVL). CVL interneurons project to and inhibit glutamatergic neurons in the rostral ventrolateral medulla (RVLM), which in turn innervate preganglionic sympathetic neurons in the spinal cord. These spinal cord neurons release acetylcholine at the sympathetic ganglion which activate noradrenergic neurons in the sympathetic ganglion causing vasoconstriction. The final result of the baroreceptor reflex is a decrease in heart rate and blood pressure.

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

summARy

References [1] Izquierdo I. Pharmacological evidence for a role of long-term potentiation in memory. FASEB J 1994;8:1139–45. [2] Lau A, Tymianski M. Glutamate receptors, neurotoxicity and neurodegeneration. Pflugers Arch 2010;460:525–42. [3] LaRoche SM, Helmers SL. The new antiepileptic drugs: scientific review. JAMA 2004;291:605–14. [4] Berry-Kravis E, Hessl D, Coffey S, Hervey C, Schneider A, Yuhas J, et al. A pilot open label, single dose trial of fenobam in adults with fragile X syndrome. J Med Genet 2009;46:266–71. [5] Olive MF. Metabotropic glutamate receptor ligands as potential therapeutics for addiction. Curr Drug Abuse Rev 2009;2:83–98. [6] Palmada M, Centelles JJ. Excitatory amino acid neurotransmission. Pathways for metabolism, storage and reuptake of glutamate in brain. Front Biosci 1998;3:d701–18. [7] Robinson MB, Blakely RD, Couto R, Coyle JT. Hydrolysis of the brain dipeptide N-acetyl-L-aspartyl-L-glutamate. Identification and characterization of a novel N-acetylated alpha-linked acidic dipeptidase activity from rat brain. J Biol Chem 1987;262:14498–14506. [8] Takamori S. VGLUTs: “exciting” times for glutamatergic research? Neurosci Res 2006;55:343–51. [9] Miyaji T, Echigo N, Hiasa M, Senoh S, Omote H, Moriyama Y. Identification of a vesicular aspartate transporter. Proc Natl Acad Sci USA 2008;105:11720–11724. [10] Kew JN, Kemp JA. Ionotropic and metabotropic glutamate receptor structure and pharmacology. Psychopharmacology (Berl) 2005;179:4–29. [11] Mayer ML, Armstrong N. Structure and function of glutamate receptor ion channels. Annu Rev Physiol 2004;66:161–81. [12] Tsien JZ. Linking Hebb’s coincidence-detection to memory formation. Curr Opin Neurobiol 2000;10:266–73. [13] Pin JP, Duvoisin R. The metabotropic glutamate receptors: structure and functions. Neuropharmacology 1995;34:1–26. [14] Danbolt NC. Glutamate uptake. Prog Neurobiol 2001;65:1–105. [15] Gendreau S, Voswinkel S, Torres-Salazar D, Lang N, Heidtmann H, Detro-Dassen S, et al. A trimeric quaternary structure is conserved in bacterial and human glutamate transporters. J Biol Chem 2004;279:39505–39512. [16] Zerangue N, Kavanaugh MP. Flux coupling in a neuronal glutamate transporter. Nature 1996;383:634–7.

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[17] Robinson MB. The family of sodium-dependent glutamate transporters: a focus on the GLT-1/EAAT2 subtype. Neurochem Int 1998;33:479–91. [18] Shashidharan P, Huntley GW, Murray JM, Buku A, Moran T, Walsh MJ, et al. Immunohistochemical localization of the neuronspecific glutamate transporter EAAC1 (EAAT3) in rat brain and spinal cord revealed by a novel monoclonal antibody. Brain Res 1997;773:139–48. [19] Dehnes Y, Chaudhry FA, Ullensvang K, Lehre KP, StormMathisen J, Danbolt NC. The glutamate transporter EAAT4 in rat cerebellar Purkinje cells: a glutamate-gated chloride channel concentrated near the synapse in parts of the dendritic membrane facing astroglia. J Neurosci 1998;18:3606–19. [20] Fei YJ, Sugawara M, Nakanishi T, Huang W, Wang H, Prasad PD, et al. Primary structure, genomic organization, and functional and electrogenic characteristics of human system N 1, a Na- and Hcoupled glutamine transporter. J Biol Chem 2000;275:23707–23717. [21] Morrell CN, Sun H, Ikeda M, Beique JC, Swaim AM, Mason E, et al. Glutamate mediates platelet activation through the AMPA receptor. J Exp Med 2008;205:575–84. [22] Kirchgessner AL. Glutamate in the enteric nervous system. Curr Opin Pharmacol 2001;1:591–6. [23] Baude A, Strube C, Tell F, Kessler JP. Glutamatergic neurotransmission in the nucleus tractus solitarii: structural and functional characteristics. J Chem Neuroanat 2009;38:145–53. [24] Benarroch EE. The arterial baroreflex: functional organization and involvement in neurologic disease. Neurology 2008;71:1733–8. [25] Crestani CC, Alves FH, Busnardo C, Resstel LB, Correa FM. N-Methyl-D-aspartate glutamate receptors in the hypothalamic paraventricular nucleus modulate cardiac component of the baroreflex in unanesthetized rats. Neurosci Res 2010;67:317–26. [26] Iremonger KJ, Benediktsson AM, Bains JS. Glutamatergic synaptic transmission in neuroendocrine cells: Basic principles and mechanisms of plasticity. Front Neuroendocrinol 2010;31:296–306. [27] Bauer D, McCullumsmith RE, Meador-Woodruff JH. A role for glutamate receptors, transporters, and interacting proteins in cortical dysfunction in schizophrenia. In: O’Donnell P, editor. In Cortical Deficits in Schizophrenia. New York, NY: Springer; 2008. p. 113–48.

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C H A P T E R

22 GABAergic Neurotransmission Andre H. Lagrange, Mark D. Grier OVERVIEW OF GABA MEDIATED SIGNALING GABA is the primary mediator of neuronal inhibition in the brain, as well as being a very important neurotransmitter in the spine and peripheral nervous system. Major components of the GABAergic neurotransmission system include two enzymes that synthesize GABA, two classes of receptors, two types of transporters, and an enzyme that degrades GABA. Nearly all GABA found in the mammalian brain is synthesized by decarboxlyation of glutamate by two forms of glutamic acid decarboxylase, GAD65 and GAD67. While GAD67 is present as a nearly constitutively active enzyme throughout the cell, GAD65 is localized primarily to the axon terminals and only becomes highly active in periods of increased demand. After GABA is synthesized, it is packaged into synaptic vesicles by the vesicular GABA transporter vGAT (also known as vesicular inhibitory amino acid transporter, VIAAT). Upon stimulation of the neuron, these vesicles fuse to the presynaptic terminal and release GABA into the synapse via calcium dependent exocytosis. GABA is removed from the synapse by another class of GABA transporter, GATs. It is important to note that while the function of vGAT and GAT are similar, their structure and mechanisms of transport are very different. To date, there have been four GAT isoforms identified, each with unique pharmacology and cellular localization. In the CNS, the main transporters are GAT1 and GAT3, expressed primarily on presynaptic terminals and astrocytes, respectively. Once GABA is cleared from the synapse it is degraded by GABA-transaminase. Inhibitors of GABA reuptake and breakdown effectively produce increased levels of extracellular GABA and are used clinically in conditions of CNS hyperexcitability, such as spasticity and epilepsy [1].

GABA RESPONSES ARE MEDIATED BY BOTH IONOTROPIC AND METABOTROPIC RECEPTORS The neuronal response to GABA is mediated through two populations of receptors, GABAA and GABAB receptors. While both populations generally produce neuronal

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00022-6

inhibition, they do so through wholly different mechanisms. GABAA receptors are primarily postsynaptic ligand-gated Cl ion channels, while GABAB receptors are G-protein coupled receptors found on both pre- and postsynaptic terminals. Activation of GABAA receptors produces Cl ion fluxes that directly alter membrane potential. In contrast, the effects GABAB receptor stimulation are mediated by G-proteins that produce a variety of responses, including activation of potassium channels, inhibition of presynaptic voltage-gated calcium channels and modulation of intracellular second messenger systems, including adenylate cyclase and phospholipase C. This review will focus primarily on GABAA receptors; however it is important to note that GABAB receptors also play an important role in inhibition in the central nervous system.

GABAA RECEPTORS MEDIATE MULTIPLE MODES OF NEURONAL SIGNALING GABAA receptors have the important ability to convey two different types of inhibition within the brain [2]. Phasic inhibition involves temporally and spatially precise responses to very brief, high levels of synaptic GABA (≈1 mM, 1 ms). In comparison, synaptic spillover produces prolonged low levels of GABA in the extrasynaptic space that activate highly sensitive GABAA receptor subtypes, thereby producing a tonic inhibitory current (Fig. 22.1). Despite its small amplitude, the prolonged duration of tonic inhibition produces an overall charge transfer that exceeds that of synaptic neurotransmission in certain brain regions. Furthermore, by effectively reducing membrane resistance, tonic inhibition reduces the ability of excitatory currents to depolarize the cell toward the action potential threshold. These dual effects allow tonic inhibition to serve as a critical determinant of overall network excitability in a number of important brain regions. While classically thought of as an inhibitory neurotransmitter, GABAA receptors also have the unusual capacity to convey excitatory responses, depending on the Cl reversal potential across the neuronal membrane [3]. In normal adult neurons, robust activity of the Cl transporter KCC2 results in low intracellular Cl levels

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22. GABAERGIC NEuRoTRANsmIssIoN

Strong Synaptic Input

Weak Synaptic Input

Presynaptic Neurons

Presynaptic Neurons Glia

Glia

Metabolism

Metabolism

GABA-T

GABA-T

VGAT

GABA

GABA

Postsynaptic Neuron GABA A Receptor GAT

GABA B Receptor Synaptic Vesicle

Postsynaptic Neuron GABA A Receptor GAT

GABA B Receptor Synaptic Vesicle

FIGURE 22.1 GABA receptors mediate multiple forms of neurotransmission. During weak synaptic activity, brief surges of GABA activate the ionotropic GABAA receptors clustered within the synapse, producing transient inhibitory postsynaptic currents. However, during more pronounced synaptic activity, GABA can overflow beyond the synapse, activating highly sensitive GABAA receptor subtypes that exist outside the synapse to produce a long-lived tonic current. G-protein coupled GABAB receptors on both the pre- and postsynaptic neurons are also activated by synaptic overflow to help suppress neuronal activity.

compared to the extracellular fluid, thereby producing a hyperpolarizing influx of negative Cl ions when GABAA receptors are activated. In contrast, maturing neurons express very little KCC2, and anion homeostasis is largely determined by a different Cl transporter (NKCC1) that preferentially transports Cl into the cell. This produces high levels of intracellular Cl that allow GABAA receptor activation to produce a depolarizing efflux of anions [3] that can generate action potentials, intracellular calcium waves, and relieve Mg block of NMDA receptors. These excitatory responses regulate a number of important steps in brain development, including the formation of both excitatory and inhibitory synapses. Interestingly, similar depolarizing responses can be found in specific subcellular regions in the normal adult neurons, as well as more widely following brain injury. Furthermore, these excitatory GABA responses may be involved in the pathogenesis of some neurological disorders (e.g. epilepsy) and selective inhibition of NKCC1 with drugs like bumetanide, has been proposed as a potential future treatment for some of these conditions [4].

NEUROTRANSMISSION IS MEDIATED BY A DIVERSE SET OF GABAA RECEPTOR SUBTYPES GABAA receptors are heteropentameric protein complexes, generally composed of a combination of 2α, 2β and either a γ or δ subunit. To date, there are 6α, 3β, 3γ and a single δ isoform subtypes. Despite a vast array of possible subunit combinations, only a limited number of GABAA receptor subunit combinations are actually found in native tissue. The expression of individual subunit combinations is highly region and cell-type specific. While most synaptic responses throughout the brain are thought to be mediated by postsynaptic α1β2γ2 containing GABAA receptors, other

subunit combinations are highly expressed in select brain regions (thalamus, hypothalamus, hippocampus), as well as in specific subcellular domains, including the axon initial segment (α2βxγ) or extrasynaptic space (α4βxδ and α5β3γ2). Since the identity of the constituent subunit isoforms is the primary determinant of GABAA receptors’ sensitivity to GABA and current kinetic properties, specific subunit isoform combinations likely serve very different physiological functions. For example, α4βxδ containing receptors are more than 10–100 times more sensitive to low levels of GABA than some other subunit combinations, allowing them to respond to the very low levels of ambient GABA in the extrasynaptic space. In contrast, the less sensitive, but rapidly activating α1β2γ2 containing GABAA receptors are found clustered within inhibitory synapses, where they are able to convert extremely brief, high levels of GABA to more prolonged postsynaptic currents. However, these receptors also desensitize quickly, and respond poorly during repetitive input. Replacement of the α1 subunit with the α3 subunit, produces receptors that are even less sensitive to low GABA, but are more slowly desensitizing and deactivate extremely slowly, resulting in very prolonged IPSCs that can produce a summating response during repetitive stimulation [5,6]. More recently, it has become clear that these patterns of subunit expression can be profoundly disrupted in a variety of pathological states like ischemia and seizures [7,8]. These changes may be compensatory, but are also thought to contribute to some of the pathological plasticity that occurs after brain injury.

SUBTYPE PREFERRING DRUGS ALLOW FOR SELECTIVE MODULATION OF GABA SIGNALING Given that essentially every neuron in the mammalian brain expresses GABAA receptors, it is not surprising that

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

suBTyPE PREfERRING DRuGs Allow foR sElECTIvE moDulATIoN of GABA sIGNAlING

a number of the most useful medications used in neurology and psychiatry target these receptors. Nonselective GABAA receptor-active drugs, like barbiturates, and inhaled anesthetics, propofol and etomidate, are commonly used for surgery. At low concentrations, these agents enhance the effects of endogenous GABA, but at higher concentrations they directly and indiscriminately activate GABAA receptors throughout the brain. While this allows these agents to serve as general anesthetics, it also introduces the possibility of lethal effects on respiration and blood pressure. Fortunately, there are also a variety of subunit-preferring medications that allow selective modulation of specific GABAA receptor subunits combinations. For example, benzodiazepines increase the GABA affinity of α(1, 2, 3 or 5)βxγx containing GABAA receptors. Even more selective is the benzodiazepine-like hypnotic compound, zolpidem. This drug potently enhances α1βxγ2 mediated inhibition, has much weaker effects on α2,3βxγ2 mediated currents, and is virtually inactive at all other GABAR subunit combinations. Since synaptic GABAA receptors are usually exposed to saturating levels of GABA, the affinityenhancing effects of benzodiazepines and zolpidem produce prolonged IPSCs, without much effect on current amplitude. In contrast, extrasynaptic α5β3γ2 GABAA receptors on hippocampal neurons are exposed to prolonged, low levels of GABA. Consequently, benzodiazepines, but not zolpidem, produce increased tonic current amplitude in these cells. This form of inhibition is thought to be important in learning/memory, and suppression of tonic inhibition with α5βxγ-selective benzodiazepine site inverse agonists is currently being evaluated as a potential mechanism for cognitive enhancement. Similar agents that modulate α2 and α3 containing receptors are in more preliminary stages of development as anxiolytics, muscle relaxants and analgesics [9]. The other predominant mediators of tonic inhibition in the brain are α4βδ containing GABAA receptors. These relatively small GABA-evoked currents can be enhanced several fold by barbiturates and ethanol. While truly α4βδ selective agents are lacking, the drug gaboxadol is a superagonist with particularly high potency and efficacy at this particular subunit combination, and low concentrations of this drug are often used to selectively enhance α4βδ mediated tonic inhibition. Interestingly, these receptors are also especially prone to modulation by endogenous neuroactive steroid hormones produced in the gonads, adrenals and even in the CNS itself. While steroid hormones are generally associated with long-term changes at the genomic level, neuroactive steroids, like allopregnanolone (3α,5α-tetrahydroprogesterone) and THDOC (allotetrahydrocorticosterone) enhance GABAergic signaling within seconds. This enhancement is not mediated by the classic nuclear steroid hormone receptors, nor is it mediated by changes in protein synthesis. Instead, these neuroactive steroids are thought to interact directly with GABAA receptors, possibly involving specific binding site(s) in the transmembrane region. While allopregnanolone and THDOC enhance most GABAA receptor subunit

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combinations to some degree, αxβxδ containing receptors are especially sensitive to even extremely low neurosteroid concentrations. Moreover, αxβxδ containing receptors typically produce small currents due to a lower single channel conductance and mean open times. However, nanomolar concentrations of neurosteroids shift the activation properties of αxβxδ containing receptors, producing a nearly ten-fold increase in the maximal GABA evoked current [10]. In summary, GABA is a multifunctional neurotransmitter capable of producing brief, phasic responses or tonic currents that may be either excitatory or inhibitory, depending on the age, brain region, and cell-type being studied. In addition, the control of GABAergic signaling involves multiple proteins, many of which have multiple isoforms. The regulation of GABAergic signaling is a critical determinant of nervous system function, and disruption of this system is thought to play a crucial role in many neurological disorders. Fortunately, the availability of a number of isoform-selective drugs at these diverse preand postsynaptic targets provides an unparalleled potential for pharmacological manipulation of neuronal activity in the treatment of human illness.

References [1] Bialer M, Johannessen SI, Kupferberg HJ, Levy RH, Perucca E, Tomson T. Progress report on new antiepileptic drugs: a summary of the Seventh Eilat Conference (EILAT VII). Epilepsy Res 2004;61:1–48. [2] Farrant M, Nusser Z. Variations on an inhibitory theme: phasic and tonic activation of GABA(A) receptors. Nat Rev Neurosci 2005;6:215–29. [3] Ben Ari Y, Gaiarsa JL, Khazipov R. GABA: A pioneer transmitter that excites immature neurons and generates primitive oscillations. Phys Rev 2007;87:1215–84. [4] Dzhala VI, Talos DM, Sdrulla DA, Brumback AC, Mathews GC, Benke TA, et al. NKCC1 transporter facilitates seizures in the developing brain. Nat Med 2005;11:1205–13. [5] Cox CL, Huguenard JR, Prince DA. Nucleus reticularis neurons mediate diverse inhibitory effects in thalamus. Proc Natl Acad Sci USA 1997;94:8854–9. [6] Rula EY, Lagrange AH, Jacobs MM, Hu N, Macdonald RL, Emeson RB. Developmental modulation of GABA(A) receptor function by RNA editing. J Neurosci 2008;28:6196–201. [7] Clarkson AN, Huang BS, Macisaac SE, Mody I, Carmichael ST. Reducing excessive GABA-mediated tonic inhibition promotes functional recovery after stroke. Nature 2010;468:305–9. [8] Zhang N, Wei W, Mody I, Houser CR. Altered localization of GABA(A) receptor subunits on dentate granule cell dendrites influences tonic and phasic inhibition in a mouse model of epilepsy. J Neurosci 2007;27:7520–31. [9] Mohler H. GABA(A) receptor diversity and pharmacology. Cell Tissue Res 2006;326:505–16. [10] Wohlfarth KM, Bianchi MT, Macdonald RL. Enhanced neurosteroid potentiation of ternary GABA(A) receptors containing the delta subunit. J Neurosci 2002;22:1541–9.

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C H A P T E R

23 Renin-Angiotensin Amy C. Arnold, Debra I. Diz INTRODUCTION The renin-angiotensin system (RAS) plays an important role in the regulation of blood pressure under normal and pathophysiological conditions. The RAS interacts with the autonomic nervous system for blood pressure regulation, with angiotensin receptors localized to brain regions involved in modulation of both sympathetic and parasympathetic nervous system activity. The following provides a brief overview of the classical and brain RAS, focusing on interactions of this hormonal system with autonomic brain regions integral to cardiovascular control.

THE CLASSICAL RAS

Recent Advances in the Classical RAS

Components and Features of the Classical RAS As recently reviewed [1,2] the classical circulating RAS is a series of enzyme-substrate interactions that generates angiotensin peptides involved in cardiovascular, fluidelectrolyte and neuroendocrine homeostasis. The ratelimiting enzyme, renin, is synthesized in juxtaglomerular cells of the kidney and is secreted into the circulation in response to various stimuli. As illustrated in Figure 23.1, circulating renin acts upon hepatic-derived angiotensinogen to form angiotensin I. Angiotensin converting enzyme (ACE), a lung-derived dipeptidyl carboxypeptidase, subsequently cleaves angiotensin I into angiotensin II, the main effector peptide of the RAS. However, other enzymes including chymase are capable of forming angiotensin II in pathological conditions. Four subtypes of the angiotensin II g-protein coupled receptor (AT1–AT4) have been identified. The majority of angiotensin II effects are mediated by ubiquitously expressed AT1 receptors. Depending on the cell and tissue type, AT1 receptor activation is associated with inhibition of adenylyl cyclase, activation of phospholipase C or phosphoinositide hydrolysis. Angiotensin II stimulation of AT1 receptors induces vasoconstriction, sympathetic activation, suppression of baroreflex function, sodium and water reabsorption, cellular proliferation and hypertrophy, inflammation, oxidative stress and release of aldosterone, vasopressin and noradrenaline (Table 23.1). Angiotensin II also binds AT1 receptors on juxtaglomerular

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00023-8

cells to provide negative feedback suppression of renal renin release. These actions are collectively abrogated by ACE inhibitors or AT1 receptor antagonists, to prevent the formation or binding of angiotensin II, respectively. Angiotensin II also binds AT2 receptors, with expression of these receptors abundant during fetal life, but declining into adulthood. The AT2 receptor opposes AT1 activation with respect to blood pressure and cellular proliferation and is also involved in growth, repair, pain threshold and neuronal cell maturation and differentiation (Table 23.1). Angiotensin II is degraded by aminopeptidases A and N into the shorter fragments angiotensin III and IV, respectively (Fig. 23.2, Table 23.1).

New pathways for the production of angiotensin peptides have been recently revealed, including the formation of the heptapeptide angiotensin-(1-7) (Figs. 23.1 and 23.2). Angiotensin-(1-7) can be formed via cleavage of angiotensin I by various endopeptidases including neprilysin (NEP), thimet oligopeptidase (TOP) and prolyl oligopeptidase (POP) [3,4]. Alternatively, angiotensin-(1-7) is formed through cleavage of angiotensin II by ACE2, a novel homolog of ACE (Fig. 23.1) [3–5]. The actions of angiotensin-(1-7) are mediated by g-protein coupled mas receptors and are blocked by the selective receptor antagonist [D-Ala7]angiotensin-(1-7) [3]. Angiotensin-(1-7) is degraded into the inactive metabolite angiotensin-(1-5) by ACE [3]. The complexity of the RAS is further increased by the recent discovery of angiotensin-(1-12), a C-terminally extended peptide longer than angiotensin I (Figs. 23.1 and 23.2) [6]. Angiotensin-(1-12) is found in plasma and peripheral tissues and appears to serve as a renin-independent precursor for the production of angiotensin peptides [4].

The Classical RAS and Autonomic Regulation Systemic angiotensin II elicits vasoconstriction, sympathetic activation, baroreflex dysfunction and neuroendocrine and fluid-electrolyte changes (Table 23.1), all of which can contribute to blood pressure elevations [1,7]. The sympathetic and baroreflex actions of circulating angiotensin II involve interactions with AT1 receptors in specialized circumventricular organs lacking a functional

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23. REnIn-AngIoTEnsIn

FIGURE 23.1 Schematic diagram of the current view of the reninangiotensin system showing biochemical pathways involved in the formation, degradation and actions of the biologically active peptides angiotensin II and angiotensin-(1-7). Ang, angiotensin; ACE, angiotensin converting enzyme; NEP, neprilysin; TOP, thimet oligopeptidase; POP, prolyl oligopeptidase; AT1-R, angiotensin type 1 receptor; AT2-R, angiotensin type II receptor; mas-R, mas receptor.

TABLE 23.1 Receptors and Actions of Angiotensin Peptides Receptor

Actions

Potential Ligands

AT1

Increase blood pressure, vasoconstriction, sympathetic activation, decrease baroreceptor reflex sensitivity for heart rate control and increase baroreflex set-point, pro-inflammation, pro-oxidative, thirst behaviors, cell growth and hypertrophy, anxiety, stress, neuronal excitation, and stimulate aldosterone, norepinephrine and vasopressin release

Angiotensin II Angiotensin III Angiotensin-(1-12)– likely after conversion to Angiotensin II

AT2

Vasodilation, decrease in blood pressure, apoptosis, antiinflammation, neuronal development and differentiation

Angiotensin II Angiotensin III

AT1-7 [mas]

Vasodilation, decrease in blood pressure, anti-inflammation, anti-oxidant, increase baroreceptor sensitivity for heart rate control, neuronal excitation

Angiotensin-(1-7)

AT4

Memory, learning, Angiotensin IV depression, anxiety, prevents insulin-regulated aminopeptidase (IRAP)mediated metabolism of vasoactive peptides

FIGURE 23.2 Amino acid sequences for peptides of the renin-angiotensin system.

blood-brain barrier [7]. The importance of circulating angiotensin II to hypertension is illustrated by the finding that ACE inhibitors or AT1 receptor blockers lower blood pressure, reset the baroreflex set-point and improve baroreflex sensitivity in hypertensive populations [8]. These therapies also increase levels of angiotensin-(1-7), a peptide that stimulates mas receptor-mediated vasodilation and improvement of baroreflex function (Table 23.1) [1,3]. In rodents, low doses of angiotensin-(1-7) increase vasodilation through interactions with prostaglandinbradykinin-nitric oxide systems [3,4]. However, the systemic effects of angiotensin-(1-7) on blood pressure in humans are controversial. The counter-regulatory actions of angiotensin II and angiotensin-(1-7) suggest that the balance of these two peptides is important to cardiovascular physiology and pathophysiology. Indeed, emerging evidence suggests that cardiovascular diseases are associated with an imbalance in the angiotensin II/ACE and angiotensin-(1-7)/ACE2 axes [3–5]. Possibly shifting the balance of the RAS in the circulation, angiotensin-(1-12) acts as a precursor to angiotensin II for peripheral cardiovascular actions including vasoconstriction and elevations in blood pressure [4,6].

THE BRAIN RAS All components of the RAS are present in distinct, independently-regulated tissue systems including adipose, kidneys, blood vessels, heart and brain [2]. As early as 1961, studies demonstrated that angiotensin II acts centrally to increase blood pressure [7]. At this time, it was hypothesized that angiotensin II interacted with the brain through AT1 receptors in circumventricular organs. A few years later, the existence of an independent brain RAS was postulated by Ganten and colleagues. Indeed, all necessary precursors and enzymes required for the local formation and metabolism of angiotensins are found in brain [2,7]. However, controversy still exists regarding the cellular localization, independence from the circulating system and authenticity of brain RAS peptides [9].

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

THE BRAIn RAs

Components of the Brain RAS Angiotensinogen is widely expressed in the central nervous system with high levels in cerebrospinal fluid, hypothalamic and brainstem regions [2,7]. While angiotensinogen is primarily produced in astrocytes, it is also present in neuronal cells of key cardiovascular nuclei [7]. Renin immunoreactivity is described in discrete glial and neuronal cells of the pituitary, choroid plexus, medulla and hypothalamus. However, renin expression in brain is low suggesting alternate pathways exist for central production of angiotensin peptides [2,7]. All enzymes necessary for processing angiotensin I into bioactive peptides are found in brain (ACE, ACE2, NEP and aminopeptidases) [7]. While the exact mechanisms involved in their formation remains unclear, angiotensin peptides are all reported in brain, with the highest levels demonstrated for angiotensin II and angiotensin-(1-7) [7]. Angiotensin II AT1 receptors are highly localized to brain regions associated with autonomic outflow as well as areas involved in energy homeostasis, respiration, salt appetite, vasopressin release and thirst [2,7]. Low levels of AT2 receptors are also found in brain regions associated with cardiovascular, motor, sensory and limbic activity [7]. The active fragment angiotensin III binds central AT1 receptors to mediate actions attributed to angiotensin II including regulation of blood pressure and vasopressin release (Table 23.1) [1]. Central stimulation of AT4 receptors by angiotensin IV prevents insulin-regulated aminopeptidase-mediated metabolism of vasoactive peptides [7] and is also implicated in depression, learning, memory and anxiety (Table 23.1) [7].

Influence of the Brain RAS on Autonomic Regulation Angiotensin II AT1 and angiotensin-(1-7) mas receptors are abundant at each synaptic relay of the sympathetic and parasympathetic nervous systems, including spinal cord and ganglia [7,8]. AT1 receptors are distributed in sympathetic preganglionic neurons, sympathetic ganglia, sympathetic nerve terminals and sympathetic regulatory brain sites including hypothalamus, dorsal medullary and ventral medullary regions [10]. Central angiotensin II infusion increases arterial pressure and stimulates whole body and regional sympathetic activity in rodents, in part mediated by stimulatory interactions with other neurotransmitters including norepinephrine, dopamine and substance P [10]. Angiotensin II also acts at AT1 receptors within the solitary tract nucleus (NTS) to attenuate baroreflex restraint of sympathetic outflow to the heart, kidney and vasculature [8,10]. The NTS has descending projections to brainstem vasomotor nuclei, including the rostral and caudal ventrolateral medullas, which respond to angiotensin peptides to influence sympathetic activity, blood pressure and baroreflex function [8]. Importantly, a positive feedback loop exists between the RAS and sympathetic nervous system as increased renal sympathetic nerve activity is a major

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stimulus for β-adrenergic receptor-mediated renal renin release [10]. Angiotensin receptors are also distributed in vagal pathways influencing control of the parasympathetic nervous system [7,8,10]. High-affinity AT1 receptor binding sites are located on presynaptic vagal afferent terminals and on NTS cell bodies, with a small population of AT2 receptors in this brain region [8]. Central infusion or NTS microinjection of angiotensin II or angiotensin III impairs the baroreflex sensitivity for control of heart rate for bradycardia to increases in arterial pressure, an established marker of parasympathetic function, independent of changes in resting pressure [8,10]. Furthermore, NTS administration of AT1 receptor antagonists improves baroreflex sensitivity in rats suggesting that angiotensin II endogenous to the brain attenuates baroreflex function [8]. Illustrating the importance of brain angiotensin II to blood pressure regulation, transgenic rodents with increased brain angiotensin II generation are hypertensive and have impaired baroreflex sensitivity while those with reduced brain angiotensin II synthesis are hypotensive and have lower sympathetic and higher parasympathetic tone [2]. Studies in transgenic rodents also suggest that the source of angiotensin II for baroreflex modulation is of glial origin, whereas the source of the peptide for blood pressure regulation is neuronal [11]. Similar to the periphery, central actions of angiotensin-(1-7) oppose angiotensin II. However, both peptides can elicit similar pressor/depressor actions in specific brain nuclei and stimulate hypothalamic vasopressin release [3,4]. Numerous studies document that central administration of angiotensin-(1-7) lowers blood pressure, increases baroreflex sensitivity for control of heart rate and reduces norepinephrine release in hypertensive rodents [5]. Moreover, [D-Ala7]-angiotensin-(1-7) impairs baroreflex sensitivity in normotensive rats suggesting angiotensin-(1-7) endogenous to the brain facilitates baroreflex function [11]. Thus, the prevailing level of baroreflex sensitivity appears to reflect the balance of angiotensin II and angiotensin-(1-7) actions in the brain. This balance may in part depend on local levels of ACE2, the enzyme that converts angiotensin II to angiotensin-(1-7). Indeed, central genetic deletion or NTS pharmacologic inhibition of ACE2 reduces baroreflex sensitivity in rodents [5]. In contrast, central overexpression of ACE2 lowers blood pressure and restores baroreflex function in hypertensive animals demonstrating an emerging role for brain ACE2 in central cardiovascular regulation [5]. The angiotensinogen: renin mismatch is an ongoing topic of controversy for synthesis of angiotensin peptides within brain [9]. Levels of angiotensin-(1-12) are reported similar or higher to angiotensin II in brain [6]. In tissues, angiotensin-(1-12) can form either angiotensin II or angiotensin-(1-7) depending on the local enzyme milieu [4]. However, central immunoneutralization of angiotensin-(1-12) lowers blood pressure and improves baroreflex sensitivity in hypertensive rats [12] and NTS

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microinjection of angiotensin-(1-12) impairs baroreflex sensitivity in normotensive animals [13]. The baroreflex actions of angiotensin-(1-12) within the NTS are blocked by either an AT1 receptor antagonist or ACE inhibitor suggesting that angiotensin-(1-12) serves as a precursor to angiotensin II in brain pathways regulating blood pressure and autonomic function. Since angiotensin-(1-12) metabolism is independent of renin [4], this peptide may provide an alternate pathway for central angiotensin peptide formation.

SUMMARY Due to its key role in blood pressure regulation, the RAS has been an intense subject of research and an important therapeutic target for cardiovascular disease. Independent of blood pressure lowering effects, blockade of angiotensin II activity with ACE inhibitors or AT1 receptor antagonists reduces sympathetic activity and restores baroreflex function, in part mediated by increased levels of angiotensin-(1-7). These observations illustrate the importance of interactions between the RAS and autonomic nervous system in central regulation of blood pressure. A wider influence of the RAS on other facets of autonomic function, including energy metabolism, body weight regulation and glucose homeostasis at peripheral and central nervous system sites is emerging. The discovery of new RAS components, including ACE2, angiotensin-(1-7) and angiotensin-(1-12), will improve our understanding of these interactions that contribute broadly to autonomic regulation and may open new avenues for pharmacologic targeting of this system.

Acknowledgement

References [1] Fyhrquist F, Saijonmaa O. Renin-angiotensin system revisited. J Intern Med 2008;264:224–36. [2] Bader M. Tissue renin-angiotensin-aldosterone systems: Targets for pharmacological therapy. Annu Rev Pharmacol Toxicol 2010;50:439–65. [3] Trask AJ, Ferrario CM. Angiotensin-(1-7): pharmacology and new perspectives in cardiovascular treatments. Cardiovasc Drug Rev 2007;25:162–74. [4] Varagic J, Trask AJ, Jessup JA, Chappell MC, Ferrario CM. New angiotensins. J Mol Med 2008;86:663–71. [5] Xia H, Lazartigues E. Angiotensin-converting enzyme 2: central regulator for cardiovascular function. Curr Hypertens Rep 2010;12:170–5. [6] Nagata S, Kato J, Sasaki K, Minamino N, Eto T, Kitamura K. Isolation and identification of proangiotensin-12, a possible component of the renin-angiotensin system. Biochem Biophys Res Commun 2006;350:1026–31. [7] McKinley MJ, Albiston AL, Allen AM, Mathai ML, May CN, McAllen RM, et al. The brain renin-angiotensin system: location and physiological roles. Int J Biochem Cell Biol 2003;35:901–18. [8] Averill DB, Diz DI. Angiotensin peptides and baroreflex control of sympathetic outflow: pathways and mechanisms of the medulla oblongata. Brain Res Bull 2000;51:119–28. [9] Grobe JL, Xu D, Sigmund CD. An intracellular renin-angiotensin system in neurons: fact, hypothesis, or fantasy. Physiology (Bethesda) 2008;23:187–93. [10] Phillips MI. Functions of angiotensin in the central nervous system. Annu Rev Physiol 1987;49:413–35. [11] Sakima A, Averill DB, Kasper SO, Jackson L, Ganten D, Ferrario CM, et al. Baroreceptor reflex regulation in anesthetized transgenic rats with low glia-derived angiotensinogen. Am J Physiol Heart Circ Physiol 2007;292:H1412–H1419. [12] Isa K, Garcia-Espinosa MA, Arnold AC, Pirro NT, Tommasi EN, Ganten D, et al. Chronic immunoneutralization of brain angiotensin-(1-12) lowers blood pressure in transgenic (mRen2)27 hypertensive rats. Am J Physiol Regul Integr Comp Physiol 2009;297:R111–5. [13] Arnold AC, Isa K, Shaltout HA, Nautiyal M, Ferrario CM, Chappell MC, Diz DI. Angiotensin-(1-12) requires angiotensin converting enzyme and AT1 receptors for cardiovascular actions within the solitary tract nucleus. Am J Physiol Heart Circ Physiol 2010;299:H763–71.

Funding: NIH HL-51952.

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24 Aldosterone and the Mineralocorticoid Receptor J. Howard Pratt Aldosterone, the only true mineralocorticoid, is secreted by the zona glomerulosa, a thin layer of cells at the periphery of the adrenal cortex. Aldosterone’s principal functions are to contain extracellular potassium (K) concentration within narrow limits and to maintain adequate extracellular volume by guarding against sodium (Na) loss. The two stimuli that regulate its secretion are therefore K (only small increments are required to significantly augment the levels of aldosterone) and angiotensin II (which increases in response to the need for volume expansion and retention of additional Na). Although not as well-established, there is compelling evidence that the adrenergic nervous system influences aldosterone secretion. Rays of adrenal medullary cells extending out to the zona glomerulosa have been described in the rat adrenal, and catecholamines have been shown to stimulate aldosterone secretion in vitro. More recently, variants in the β-adrenergic receptor were found to associate with low renin hypertension. Taken together, the findings suggest the possibility of a significant contribution by the adrenergic nervous system to hypertensive states resulting from overproduction of aldosterone [1].

K channel. The amount of K secreted is a principal determinant of extracellular K concentration. Thus, by increasing the number of ENaCs residing on the apical surface, aldosterone achieves a needed level of Na reabsorption, together with maintenance of K homeostasis.

Effects of Aldosterone That Target Distal Nephron

Excess Aldosterone and the Development of Hypertension

The functions of aldosterone begin with its occupancy of the nuclear receptor known as the mineralocorticoid receptor (MR). The ligand coupled to its receptor becomes a functioning transcription factor, and thus there is genomic delay in the onset of its actions. MR is expressed in a variety of tissues including the heart and brain, but the major site for its expression is the kidney’s distal nephron. Here, the principal target is the epithelial Na channel or ENaC located in cortical collecting duct [2]. Aldosterone induces translation for the protein serum glucocorticoid kinase type I (Sgk-1) [3]. The ultimate effect is to render an ubiquitin ligase (Nedd4-2) incapable of removing ENaC from the apical surface of the cell [4]. The Na that reaches the cell interior via ENaC exits the cell at the basolateral surface via the Na,K-ATPase; the exchange of Na for K leads to secretion of K into the tubular lumen via a

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Specificity of MR for Aldosterone; Role of 11-β-Hydroxysteroid Dehydrogenase Type II (11BHSD2) Cortisol is also a ligand for MR and its circulating levels are much higher than those of aldosterone. However, under normal circumstances cortisol is converted to cortisone by 11BHSD2, an enzyme expressed in the vicinity of MR. Cortisone is biologically inactive; it has no binding affinity to MR. A deficiency in 11BHSD2 can result from a rare genetic disorder (apparent mineralocorticoid excess), from eating licorice (glycyrrhizynic acid in licorice inhibits 11BHSD2) or from Cushing’s syndrome (typically due to ectopic ACTH secretion where extremely high cortisol levels overwhelm the enzyme). In each case, there can be hypertension and typically hypokalemia.

Autonomous hypersecretion of aldosterone (primary aldosteronism) not uncommonly results in severe hypertension due to excessive reabsorption of Na in the distal nephron. The ensuing volume expansion suppresses the renin-angiotensin axis. The diagnosis is established when the plasma level of aldosterone is increased while the level of plasma renin activity (PRA) is low. An additional diagnostic approach is to utilize the ratio of plasma aldosterone to PRA (aldosterone/renin ratio). (Fig. 24.1) There are two types of primary aldosteronism (Fig. 24.2): that which is caused by a solitary adrenal adenoma (frequently 1 cm in diameter), also known as Conn’s syndrome [5]; and, the most common form, bilateral adrenal hyperplasia. The prevalence of primary aldosteronism is now estimated to be 10% of patients with hypertension

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Two types of primary aldosteronism

Solitary adenoma MR ENaC (α - β - γ subunits)

Right

Left

Bilateral adrenal hyperplasia

FIGURE 24.2 The great majority of cases of primary aldosteronApical

Basolateral

FIGURE 24.1 Schematic depiction of mechanism for how aldosterone increases Na uptake by ENaC in principal cells of collecting duct. ENaC residing on the apical surface consists of three subunits (alpha, beta, gamma). ENaC is removed from the cell surface by the ubiquitin ligase Nedd4-2. Aldosterone binds to MR in the cell’s interior resulting in a transcription factor that increases expression of Sgk-1. The latter phosphorylates Nedd4-2, and by so doing, inactivates it to where it can no longer displace ENaC from the apical surface. The result is a prolonged residency time with greater Na reabsorption.

[6]. The prevalence increases with age and is exceptionally high in patients with resistant hypertension [7] (patients with uncontrolled hypertension on three or more antihypertensive drugs). Although it was initially thought that hypokalemia was a requisite feature, it is now well accepted that most patients with primary aldosteronism have a normal serum K concentration. Treatment approaches depend on whether the source of excess aldosterone is from a unilateral adrenal adenoma or arises from hyperplasia of both adrenals. Once it is confirmed, often by performing an adrenal venous catheterization, that a single adenoma is causative, it can be removed using laparoscopic surgery. Medical management for bilateral disease is employed most often. There are currently two MR blockers, spironolactone and eplerenone. In addition, direct inhibitors of ENaC can be used such as amiloride. A description of medical treatment options is listed in Table 24.1.

Low-Renin Hypertension but not Primary Aldosteronism Many patients with hypertension who may require multiple antihypertensive drugs and patients who are typically older have suppressed levels of PRA. Their aldosterone levels are in the normal range and thus do not have primary aldosteronism based on established criteria. These patients more often than not respond extremely

ism are caused either by an adrenal adenoma or by bilateral adrenal hyperplasia. The former is less common and is removed with surgery, usually laparoscopically. In the instance where there is bilateral disease, medical management that mitigates the actions of aldosterone is preferred.

TABLE 24.1 Medical Treatment options for Treating AldosteroneInduced Hypertension (Primary Aldosteronism and low-Renin, normal Aldosterone, Hypertension) Drug

Mechanism

Advantages

Disadvantages

Spironolactone

Competive antagonist of MR.

1. Extremely effective in lowering blood pressure. 2. Reduced mortality rate by 30% in RALES clinical trial with CHF patients [9]. 3. Cost is trivial; only pennies a day.

Spironolactone also binds to the androgen receptor which can result in dose-dependent side effects.

Eplerenone

Competive antagonist of MR.

1. Selective for MR (no binding to androgen receptor. 2. Also shown to improve outcomes in CHF [10–11].

Although generic version available, cost is considerably more than cost of spironolactone.

Amiloride

Direct inhibitor of ENaC.

1.Effective in lowering blood pressure. 2. Few side effects. 3. Inexpensive.

A theoretical disadvantage is that it does not block aldosterone’s effects at the tissue level (e.g. the heart).

MR, mineralocorticoid receptor; ENaC, epithelial Na channel; RALES, Randomized Aldactone Evaluation Study.

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AldosTERonE And THE MInERAloCoRTICoId RECEPToR

well to MR blockade. Indeed, recognition of this large body of hypertensive patients who respond to this targeting of antihypertensive therapy has been one of the recent breakthroughs in the area of hypertension.

Aldosterone as a Mediator of Tissue Injury Angiotensin II infused into an experimental animal produces injury to multiple organ systems. It is now known that the injury can be prevented by concurrent treatment with an MR antagonist such as spironolactone indicating that aldosterone and not angiotensin II conveys much of the tissue damage [8]. Thus, aldosterone has more than a single undesirable feature: too much causes not only hypertension but damage to a variety of tissues including kidney, heart, and vasculature. Some of the largest and most noteworthy clinical trials of the effectiveness of MR blockade have been in patients with congestive heart failure (Table 24.1). In summary, aldosterone plays a pivotal role in maintaining K homeostasis and extracellular volume. Its Na reabsorptive actions bolstered by today’s typically rich in Na diet is thought to participate in the pathogenesis of hypertension and in some instances to confer risk of injury to vital tissues such as heart and kidney. Treatment regimens that include MR antagonist are becoming increasingly utilized.

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[2] Canessa CM, Horisberger JD, Rossier BC. Epithelial sodium channel related to proteins involved in neurodegeneration. Nature 1993;361:467–70. [3] Pearce D, Verrey F, Chen SY, Mastroberardino L, Meijer OC, Wang J, et al. Role of SGK in mineralocorticoid-regulated sodium transport. Kidney Int 2000;57:1283–9. [4] Debonneville C, Flores SY, Kamynina E, Plant PJ, Tauxe C, Thomas MA, et al. Phosphorylation of Nedd4-2 by Sgk1 regulates epithelial Na() channel cell surface expression. EMBO J 2001;20:7052–9. [5] Conn JW. Primary aldosteronism, a new clinical sydrome. J Lab Clin Med 1955;45:6–17. [6] Young Jr WF. Minireview: Primary aldosteronism–changing concepts in diagnosis and treatment. Endocrinology 2003;144:2208–13. [7] Calhoun DA, Jones D, Textor S, Goff DC, Murphy TP, Toto RD, et al. Resistant hypertension: Diagnosis, evaluation, and treatment. A scientific statement from the American Heart Association Professional Education Committee of the Council for High Blood Pressure Research. Hypertension 2008;51:1403–19. [8] Rocha R, Chander PN, Khanna K, Zuckerman A, Stier Jr CT. Mineralocorticoid blockade reduces vascular injury in stroke-prone hypertensive rats. Hypertension 1998;31:451–8. [9] Pitt B, Zannad F, Remme WJ, Cody R, Castaigne A, Perez A, et al. The effect of spironolactone on morbidity and mortality in patients with severe heart failure. Randomized Aldactone Evaluation Study investigators. N Engl J Med 1999;341:709–17. [10] Pitt B, Remme W, Zannad F, Neaton J, Martinez F, Roniker B, et al. Eplerenone, a selective aldosterone blocker, in patients with left ventricular dysfunction after myocardial infarction. N Engl J Med 2003;348:1309–21. [11] Zannad F, McMurray JJ, Krum H, van Veldhuisen DJ, Swedberg K, Shi H, et al. Eplerenone in patients with systolic heart failure and mild symptoms. N Engl J Med 2011;364:11–21.

References [1] Pratt JH. The adrenergic nervous system conversing with the adrenal cortex: New implications for salt-sensitive hypertension. Hypertension 2006;48:820–1.

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25 Vasopressin and Disorders of Water Homeostasis Joseph G. Verbalis VASOPRESSIN SECRETION The primary physiologic action of arginine vasopressin (AVP) is its function as a water-retaining hormone. The central sensing system (osmostat) for control of release of AVP is located in the hypothalamus anterior to the third ventricle that also includes the circumventricular organ, the organum vasculosum of the lamina terminalis (OVLT). The osmostat controls release of AVP to cause water retention, and also stimulates thirst to cause water repletion. Osmotic regulation of AVP release and thirst are usually tightly coupled, but experimental lesions and some pathologic situations in humans demonstrate that each can be regulated independently. The primary extracellular osmolyte to which the osmoreceptor responds is sodium. Under normal physiologic conditions, glucose and urea readily traverse neuron cell membranes and do not stimulate release of AVP. Basal osmolality in normal subjects lies between 280 and 295 mOsm/kg H2O, but for each individual osmolality is maintained within narrow ranges. Increases in plasma osmolality as little as 1% will stimulate the osmoreceptors to release AVP. Basal plasma levels of AVP are 0.5 to 2 pg/mL, which are sufficient to maintain urine osmolality above plasma osmolality and urine volume in the range of 2–3 L/day. When AVP levels are suppressed below 0.5 pg/ml, urine osmolality decreases to less than 100 mOsm/kg H2O and a free water diuresis ensues to levels approaching 800–1000 ml/h (18–24 L/d). Increases in plasma osmolality cause a linear increase in plasma AVP and a corresponding linear increase in urine osmolality. At a plasma osmolality of approximately 295 mOsm/kg H2O, urine osmolality is maximally concentrated to 1000–1200 mOsm/kg H2O. Thus, the entire physiologic range of urine concentration is accomplished by relatively small changes in plasma AVP of 0 to 5 pg/ml. AVP secretion is also stimulated by low blood volume and pressure. High-pressure baroreceptors are located in the aorta and carotid sinus, and low-pressure baroreceptors are located in the right and left atria. Stimuli for pressure and volume receptors are carried via the glossopharyngeal (ninth) and vagal (tenth) cranial nerves to the nucleus tractus solitarius in the brainstem. Subsequent

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secondary and tertiary projections converge on the magnocellular neurons, where they provide inhibitory as well as excitatory inputs. Decreases in blood pressure or vascular volume stimulate AVP release, whereas situations that increase blood volume or left atrial pressure (e.g., negative-pressure breathing) decrease secretion of AVP. The release of AVP in response to changes in volume or pressure is less sensitive than the release in response to osmoreceptors, and generally a 10–15% reduction in blood volume or pressure is needed to stimulate release of AVP. However, once arterial pressure falls below this threshold, the stimulated response is exponential, and plasma levels of AVP achieved that are markedly greater than those achieved by osmotic stimulation. Other nonosmotic stimuli, such as nausea and intestinal traction, also act through similar nonosmotic neural pathways to release AVP.

VASOPRESSIN ACTIONS Three known receptor subtypes mediate the actions of AVP. They all are classical G protein-coupled receptors with seven transmembrane domains, and are classified according to the second messenger system to which they are coupled. The AVP V1a (V1aR) and V1b (V1bR) receptors are linked to the phosphoinositol signaling pathway via Gaq/11 GTP binding proteins that activate phospholipase C activity, with intracellular calcium acting as the second messenger. V1aR are present on vascular smooth muscle cells, hepatocytes, and platelets, and mediate the well-known pressor effects of AVP on peripheral resistance and blood pressure. V1bR are found predominately on corticotrophs cells of the anterior pituitary, where they mediate corticotrophin (ACTH) release in concert with the well-known effects of corticotrophin releasing hormone (CRH). V2R, or antidiuretic receptors, are mainly localized in the collecting duct cells of the kidney where they regulate water excretion. V2R are G protein-coupled receptors that activate adenylyl cyclase with subsequent increased intracellular cyclic AMP levels upon ligand activation. The increased cAMP initiates the movement of

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aquaporin-2 (AQP2) water channels from the cytoplasm to the apical (luminal) membrane of the collecting duct cells. Once inserted into the apical membrane, these channels allow facilitated rapid transport of water from the collecting duct lumen into the cell along osmotic gradients. The water then exits the cell through the basolateral membrane and into the kidney medullary circulation via aquaporin-3 and aquaporin-4 water channels, which are constitutively present in the basolateral membrane. This entire process is termed antidiuresis. In the absence of AVP, the AQP2 channels are re-internalized from the apical membrane into subapical vesicles. This prevents active reabsorption of water from the collecting duct lumen, resulting in diuresis. In addition to this rapid “shuttling” of the AQP2 water channels to regulate water reabsorption on a minute-to-minute basis, AVP also acts via V2R to regulate long-term stores of AQP2; i.e., increased AVP stimulates AQP2 synthesis and the absence of AVP results in decreased AQP2 synthesis. The hypertonic medullary interstitium determines the maximum concentration of the final urine, which is isotonic with the inner medulla of the kidney under conditions of maximal antidiuresis.

TABLE 25.1 Pathogenesis of Hypoosmolar Disorders DEPLETION (PRIMARY DECREASES IN TOTAL BODY SOLUTE  SECONDARY WATER RETENTION)a Renal solute loss Diuretic use Solute diuresis (glucose, mannitol) Salt wasting nephropathy Mineralocorticoid deficiency Non-renal solute loss Gastrointestinal (diarrhea, vomiting, pancreatitis, bowel obstruction) Cutaneous (sweating, burns) Blood loss DILUTION (PRIMARY INCREASES IN TOTAL BODY WATER  SECONDARY SOLUTE DEPLETION)b Impaired renal free water excretion Increased proximal reabsorption Hypothyroidism Impaired distal dilution

DISORDERS OF BODY WATER HOMEOSTASIS Disorders of body fluids are among the most commonly encountered problems in clinical medicine. This is because many different disease states can potentially disrupt the finely balanced mechanisms that control the intake and output of water and solutes. Although solute and water homeostasis are closely linked, clinical disorders of body fluids are generally divided into disorders of water homeostasis and disorders of solute homeostasis. Since body water is the primary determinant of the osmolality of the extracellular fluid (ECF), disorders of water homeostasis can be broadly divided into hypoosmolar disorders, in which there is an excess of body water relative to body solute, and hyperosmolar disorders, in which there is a deficiency of body water relative to body solute. Because sodium is the main constituent of plasma osmolality, these disorders are typically characterized by hyponatremia and hypernatremia, respectively.

HYPOOSMOLALITY Hypoosmolality indicates excess water relative to solute in the ECF; because water moves freely between the ECF and the intracellular fluid (ICF), this also indicates an excess of total body water relative to total body solute. Imbalances between body water and solute can be generated either by depletion of body solute more than body water, or by dilution of body solute from increases in body water more than body solute (Table 25.1). This is an oversimplification of complex physiology, and most

Syndrome of Inappropriate antidiuretic hormone secretion (SIADH) Glucocorticoid deficiency Combined increased proximal reabsorption and impaired distal dilution Congestive heart failure Cirrhosis Nephrotic syndrome Decreased urinary solute excretion Beer potomania Excess water intake Primary polydipsia Dilute infant formula a Virtually all disorders of solute depletion are accompanied by some degree of secondary retention of water by the kidneys in response to the resulting intravascular hypovolemia; this mechanism can lead to hypoosmolality even when the solute depletion occurs via hypotonic or isotonic body fluid losses. b Disorders of water retention primarily cause hypoosmolality in the absence of any solute losses, but in some cases of SIADH secondary solute losses occur in response to the resulting intravascular hypervolemia and this can then further aggravate the hypoosmolality (however, this pathophysiology does not likely contribute to the hyponatremia of edema-forming states such as congestive heart failure and cirrhosis, since in these cases multiple factors favoring sodium retention will result in an increased total body sodium).

hypoosmolar states include components of both solute depletion and water retention. Nonetheless, this general concept has proven to be useful because it provides a simple framework for understanding the basic etiologies of hypoosmolar disorders. Definitive identification of the etiology of hypoosmolality is not always possible at the time of presentation, but categorization according to the patient's

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HyPoosmolAlITy

ECF volume status represents the first step in ascertaining the underlying cause of the disorder.

Decreased ECF Volume (Hypovolemia) Clinically detectable hypovolemia indicates some degree of solute depletion. Even isotonic or hypotonic fluid losses can cause hypoosmolality if water or hypotonic fluids are subsequently ingested or infused. A low urine sodium concentration (UNa) suggests a non-renal cause of solute depletion, whereas a high UNa suggests renal causes of solute depletion. Diuretic use is the most common cause of hypovolemic hypoosmolality. Most etiologies of solute losses causing hypovolemic hypoosmolality will be clinically apparent, although some salt-wasting nephropathies and mineralocorticoid deficiency may be difficult to diagnose during early phases of these diseases.

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TABLE 25.2 Criteria for the Diagnosis of sIADH ESSENTIAL 1. Decreased effective osmolality of the extracellular fluid (Posm  275 mOsm/kg H2O). 2. Inappropriate urinary concentration (Uosm  100 mOsm/kg H2O with normal renal function) at some level of hypoosmolality. 3. Clinical euvolemia, as defined by the absence of signs of hypovolemia (orthostasis, tachycardia, decreased skin turgor, dry mucous membranes) or hypervolemia (subcutaneous edema, ascites). 4. Elevated urinary sodium excretion while on a normal salt and water intake. 5. Normal thyroid, adrenal and renal function. SUPPLEMENTAL 6. Abnormal water load test (inability to excrete at least 80% of a 20 ml/ kg water load in 4 hours and/or failure to dilute Uosm to 100 mOsm/ kg H2O). 7. Plasma AVP level inappropriately elevated relative to plasma osmolality. 8. No significant correction of serum [Na] with volume expansion but improvement after fluid restriction.

Normal ECF Volume (Euvolemia) Virtually any disorder causing hypoosmolality can present with a volume status that appears normal by standard methods of clinical evaluation. Because clinical assessment of volume status is not very sensitive, the presence of normal or low blood urea nitrogen and uric acid concentrations are helpful laboratory correlates of relatively normal ECF volume. In these cases, a low UNa (30 mmol/L) suggests depletional hypoosmolality secondary to ECF losses with subsequent volume replacement by water or other hypotonic fluids. Such patients may appear euvolemic by the usual clinical parameters used to assess hydration status. A high UNa (30 mmol/L) generally indicates a dilutional hypoosmolality such as SIADH, the most common cause of euvolemic hypoosmolality. The clinical criteria necessary to diagnose SIADH remain as initially defined by Bartter and Schwartz in 1967 (Table 25.2). Many different disorders are associated with SIADH, which can be divided into four major etiologic groups: tumors, CNS disorders, drug effects, and pulmonary diseases.

Increased ECF Volume (Hypervolemia) Clinically detectable hypervolemia indicates whole body sodium excess, and hypoosmolality in these patients suggests a relatively decreased intravascular volume and/or pressure leading to water retention as a result of elevated plasma AVP levels and decreased distal delivery of glomerular filtrate to the kidneys. Such patients usually have a low UNa because of secondary hyperaldosteronism, but under certain conditions the UNa may be elevated (e.g., diuretic therapy). Hyponatremia generally does not occur until relatively advanced stages of congestive heart failure, cirrhosis, or the nephrotic syndrome, by which time diagnosis is usually not difficult. Renal failure can also cause retention of both sodium and water.

The clinical manifestations of hyponatremia are largely neurological, and primarily reflect brain edema resulting from osmotic water shifts into the brain. These range from nonspecific symptoms such as headache and confusion, to more severe manifestations such as decreased sensorium, coma, seizures, and death. Significant central nervous system (CNS) symptoms generally do not occur until the serum sodium concentration ([Na]) falls below 125 mmol/L, and the severity of symptoms can be roughly correlated with the degree of hypoosmolality. Individual variability is marked, and for any patient the level of serum [Na] at which symptoms will appear cannot be accurately predicted. Several factors other than the severity of the hypoosmolality also affect the degree of neurological dysfunction. The most important is the time course over which hypoosmolality develops. Rapid development of severe hypoosmolality frequently causes marked neurological symptoms, whereas gradual development over several days or weeks is often associated with relatively mild symptomatology despite profound degrees of hypoosmolality. This is because the brain counteracts osmotic swelling by extruding extracellular and intracellular solutes, including potassium and a variety of small organic molecules (amino acids, polyols and methylamines) called organic osmolytes. Since this is a time-dependent process, rapid development of hypoosmolality can result in brain edema before this adaptation occurs, but with slower development of the same degree of hypoosmolality brain cells can lose solute sufficiently rapidly to prevent cell swelling, brain edema, and neurological dysfunction. Underlying neurological disease also affects the level of hypoosmolality at which CNS symptoms appear; moderate hypoosmolality is of little concern in an otherwise healthy patient, but can cause morbidity

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in a patient with an underlying seizure disorder. Nonneurological metabolic disorders (hypoxia, hypercapnia, acidosis, hypercalcemia, etc.) similarly can affect the level of osmolality at which CNS symptoms occur.

HYPEROSMOLALITY Hyperosmolality indicates a deficiency of water relative to solute in the ECF. Because water moves freely between the ICF and ECF, this also indicates a deficiency of total body water relative to total body solute. Although hypernatremia can be caused by an excess of body sodium, the vast majority of cases are due to losses of body water in excess of body solutes, caused by either insufficient water intake or excessive water excretion. Consequently, most of the disorders causing hyperosmolality are those associated with inadequate water intake and/or deficient pituitary AVP secretion (Table 25.3). Although hyperosmolality from inadequate water intake is seen frequently in clinical practice, this is usually not due to an underlying defect in thirst but rather results from a generalized incapacity to obtain and/or ingest fluids, often stemming from a depressed sensorium. Evaluation of the patient’s ECF volume status is important as a guide to fluid replacement therapy, but is not as useful for differential diagnosis since most hyperosmolar patients will manifest some degree of hypovolemia. Rather, assessment of urinary concentrating ability provides the most useful data with regard to the type of disorder present. Using this approach, disorders of hyperosmolality can be categorized as those in which renal water conservation mechanisms are intact but are unable to compensate for inadequately replaced losses of hypotonic fluids from other sources, or those in which renal concentrating defects are a contributing factor to the deficiency of body water.

Diabetes Insipidus Diabetes insipidus (DI) can result from either inadequate AVP secretion (central or neurogenic DI) or inadequate renal response to AVP (nephrogenic DI). Central DI is caused by a variety of acquired or congenital anatomic lesions that disrupt the neurohypophysis, including pituitary surgery, tumors, trauma, hemorrhage, thrombosis, infarction, or granulomatous disease. Severe nephrogenic DI is most commonly congenital due to defects in the gene for the AVP V2R (X-linked recessive pattern of inheritance) or in the gene for the AQP2 water channel (autosomal recessive pattern of inheritance), but relief of chronic urinary obstruction or therapy with drugs such as lithium can cause an acquired form sufficient to warrant treatment. Short-lived nephrogenic DI can result from hypokalemia or hypercalcemia, but the mild concentrating defect generally does not by itself cause hypertonicity and responds to correction of the underlying disorder. Regardless of the

TABLE 25.3 Pathogenesis of Hyperosmolar Disorders WATER DEPLETION (DECREASES IN TOTAL BODY WATER IN EXCESS OF BODY SOLUTE) 1. Insufficient water intake Unavailability of water Hypodipsia (osmoreceptor dysfunction, age) Neurological deficits (cognitive dysfunction, motor impairments) 2. Hypotonic fluid loss* A. Renal: diabetes insipidus Insufficient AVP secretion (central DI, osmoreceptor dysfunction) Insufficient AVP effect (nephrogenic DI) B. Renal: other fluid loss Osmotic diuresis (hyperglycemia, mannitol) Diuretic drugs (furosemide, ethacrynic acid, thiazides) Post-obstructive diuresis Diuretic phase of acute tubular necrosis C. Non-renal fluid loss Gastrointestinal (vomiting, diarrhea, nasogastric suction) Cutaneous (sweating, burns) Pulmonary (hyperventilation) Peritoneal dialysis SOLUTE EXCESS (INCREASES IN TOTAL BODY SOLUTE IN EXCESS OF BODY WATER) 1. Sodium Excess Na administration (NaCl, NaHCO3) Sea water drowning 2. Other Hyperalimentation (intravenous, parenteral) *Most hypotonic fluid losses will not produce hyperosmolality unless insufficient free water is ingested or infused to replace the ongoing losses, so these disorders also usually involve some component of insufficient water intake.

etiology of the DI, the end result is a water diuresis due to an inability to concentrate urine appropriately. Because patients with DI do not have impaired urine Na conservation, the ECF volume is generally not markedly decreased and regulatory mechanisms for maintenance of osmotic homeostasis are primarily activated: stimulation of thirst and pituitary AVP secretion (to whatever degree the neurohypophysis is still able to secrete AVP). In cases where AVP secretion is totally absent (complete DI), patients are dependent entirely on water intake for maintenance of water balance. However, in cases where some residual capacity to secrete AVP remains (partial DI), plasma osmolality can eventually reach levels that allow moderate degrees of urinary concentration. Although untreated DI can lead to both hyperosmolality and volume depletion, until the water losses become severe, volume depletion is minimized by osmotic shifts of water from the ICF into the more osmotically concentrated ECF.

Osmoreceptor Dysfunction The primary osmoreceptors that control AVP secretion and thirst are located in the anterior hypothalamus, and lesions of this region in animals cause hyperosmolality through a combination of impaired thirst and osmotically

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stimulated AVP secretion. Initial reports in humans described this syndrome as “essential hypernatremia,” and subsequent studies used the term “adipsic hypernatremia” in recognition of the profound thirst deficits found in most of the patients. All of these syndromes are now grouped together as disorders of osmoreceptor function. Most of the cases reported to date have represented various degrees of osmoreceptor destruction associated with different brain lesions. In contrast to lesions causing central DI, these lesions usually occur more rostrally in the hypothalamus. For all cases of osmoreceptor dysfunction it is important to remember that afferent pathways from the brainstem to the hypothalamus generally remain intact; therefore, these patients will usually have normal AVP and renal concentrating responses to baroreceptor-mediated stimuli such as hypovolemia and hypotension. The clinical manifestations of hyperosmolality can be divided into the signs and symptoms produced by dehydration, which are largely cardiovascular, those caused by the hyperosmolality itself, which are predominantly neurological and reflect brain dehydration as a result of osmotic water shifts out of the central nervous system, and those which are secondary to excessive renal water losses in patients with DI. Cardiovascular manifestations of hypertonic dehydration include hypotension, kidney failure secondary to decreased renal perfusion, acute tubular necrosis or rhabdomyolysis, and in severe cases hypotensive shock. Neurological manifestations range from nonspecific symptoms such as irritability and decreased sensorium to more severe manifestations such as chorea, seizures, coma, focal neurological deficits, and cerebral infarction. The severity of symptoms can be roughly correlated with the degree of hyperosmolality, but individual variability is marked and for any single patient the level of serum [Na] at which symptoms will appear cannot be predicted. Similar to hypoosmolar syndromes, the length of time over which hyperosmolality develops can markedly affect clinical symptomatology. Rapid development of severe hyperosmolality is frequently associated with marked neurological symptoms, whereas gradual development over several days or weeks generally causes milder symptoms. In this case the brain counteracts osmotic shrinkage by increasing intracellular content of solutes. These include electrolytes such as potassium and organic osmolytes which previously had been called “idiogenic osmoles” (for the most part these are the same organic osmolytes that are lost from the brain during adaptation to hypoosmolality discussed previously). The net effect of this process is to protect the brain against excessive shrinkage during sustained hypertonicity. However, once the brain has adapted by increasing its solute content, rapid correction of the hyperosmolality can cause brain edema since it takes a finite time (24–48 h in animal studies) to dissipate the accumulated solutes, and until this process has been completed the brain will accumulate excess water as the plasma osmolality is normalized. This effect is most often seen in dehydrated pediatric

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patients, who can develop seizures as a result of rapid rehydration.

DISORDERS OF WATER HOMEOSTASIS AND THE AUTONOMIC NERVOUS SYSTEM The major relation between the autonomic nervous system and disorders of water retention such as hyponatremia results from effects on AVP secretion. Osmotic regulation of AVP secretion is not known to be influenced significantly by autonomic inputs to osmoreceptive cells. However, a wide variety of non-osmotic stimuli also stimulate AVP secretion, many of which originate in peripheral baroreceptors or chemoreceptors, and travel to the brainstem via vagal and glossopharyngeal afferent nerves. Prominent among these are hypovolemia, hypotension and nausea. Secretion of AVP from the neurohypophysis represents a balance between excitatory and inhibitory stimuli. In the presence of strong non-osmotic stimuli, osmotic inhibition of AVP secretion is dampened, or prevented, resulting in inappropriate AVP secretion that can lead to water retention with resulting hypoosmolality and hyponatremia. Thus, many cases of SIADH are due to afferent signaling through the autonomic nervous system. Neither hyponatremia nor hypernatremia are known to cause direct effects on autonomic efferent outputs.

Further Reading Anderson RJ, Chung H-M, Kluge R, et al. Hyponatremia: A prospective analysis of its epidemiology and the pathogenetic role of vasopressin. Ann Intern Med 1985;102:164–8. Bartter FC, Schwartz WB. The syndrome of inappropriate secretion of antidiuretic hormone. Am J Med 1967;42:790–806. Ellison DH, Berl T. Clinical practice. The syndrome of inappropriate antidiuresis. N Engl J Med 2007;356:2064–72. Ghirardello S, Malattia C, Scagnelli P, Maghnie M. Current perspective on the pathogenesis of central diabetes insipidus. J Pediatr Endocrinol Metab 2005;18:631–45. Knepper MA. Molecular physiology of urinary concentrating mechanism: regulation of aquaporin water channels by vasopressin. Am J Physiol 1997;272:F3–F12. Schrier RW. Pathogenesis of sodium and water retention in high-output and low-output cardiac failure, nephrotic syndrome, cirrhosis and pregnancy. New Engl J Med 1988;319:1065–72 and 1127–1134. Verbalis JG. The syndrome of inappropriate antidiuretic hormone secretion and other hypoosmolar disorders. In: Schrier RW, editor. Diseases of the Kidney. Philadelphia: Lippincott Williams and Wilkins; 2001. p. 2511–48. Verbalis JG. Brain volume regulation in response to changes in osmolality. Neuroscience 2010;168:862–70. Verbalis JG, Goldsmith SR, Greenberg A, Schrier RW, Sterns RH. Hyponatremia treatment guidelines 2007: Expert panel recommendations. Am J Med 2007;120:S1–S21. Zerbe R, Stropes L, Robertson G. Vasopressin function in the syndrome of inappropriate antidiuresis. Annu Rev Med 1980;31:315–27.

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26 Calcitonin Gene-Related Peptide and Adrenomedullin Donald J. DiPette, Scott C. Supowit INTRODUCTION Calcitonin gene-related peptide (CGRP) and adrenomedullin (AM) belong to a superfamily of closely related genes that also includes calcitonin and amylin. The potent vasodilator activity of CGRP (a sensory neuropeptide) and AM (acting as a circulating endocrine/paracrine factor) and their widespread distribution in peripheral tissues indicate a critical role in protecting tissues from injury, in addition to regulating systemic hemodynamics and regional organ blood flows under normal physiological and pathophysiological conditions. Considerable evidence indicates that both of these peptides possess significant protective activity against hypertension-induced heart and kidney damage, heart failure, and ischemia/reperfusion injury. These protective functions of CGRP and AM are mediated not only by their potent vasodilator activity, but also through the inhibition of oxidative stress, inflammation, and necrosis/apoptosis. The effects of CGRP and AM on these three mechanisms will be the focus of this chapter.

SYNTHESIS AND LOCALIZATION OF CGRP AND AM There are two forms of the 37 amino acid CGRP, α and β, which differ in only two amino acids in rats and three in humans. α-CGRP is derived from the tissue specific splicing of the calcitonin/CGRP gene. Whereas calcitonin is produced mainly in the C cells of the thyroid, CGRP synthesis is limited almost exclusively to specific regions of the central and peripheral nervous systems. The β-CGRP gene does not produce calcitonin and is also synthesized primarily in neuronal tissues. α-CGRP is prevalent in the central nervous system and in the peripheral sensory neural network. β-CGRP is also prevalent in the central nervous system, but peripherally is common in intestinal neurons. However, the biological activities of both peptides are similar in most vascular beds. CGRP and its receptors are widely distributed in the nervous and cardiovascular systems. In the periphery, prominent sites of

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00026-3

CGRP synthesis are the dorsal root ganglion (DRG). These structures contain the cell bodies of sensory nerves that terminate peripherally on blood vessels and all other tissues innervated by sensory nerves and centrally in laminae I/II of the dorsal horn of the spinal cord. A dense perivascular CGRP neural network is seen around the blood vessels in all vascular beds. In these vessels CGRP containing nerves are found at the junction of the adventitia and the media passing into the muscle layer. It is thought that circulating CGRP is largely derived from these perivascular nerve terminals and represents a spillover phenomenon related to the release of these peptides to promote vasodilation or other tissue functions. Receptors for CGRP have been identified in the media and intima of resistance vessels as well as the endothelial layer. Adrenomedullin (AM) is a 52 amino acid peptide that was originally isolated from pheochromocytoma cells. Since that time it has been shown to be produced in a number of different cells and tissues that are relevant to the cardiovascular system including vascular smooth muscle (VSMCs) and endothelial cells. A number of cytokines, growth factors, and hormones have been reported to increase AM expression including TNF-α and -β, IL-1α and -β, dexamethasone, cortisol, retinoic acid, thyroid hormone, and shear stress.

RELEASE OF CGRP FROM SENSORY NERVE TERMINALS CGRP-rich nerve fibers are components of the primary afferent nervous system, comprising principally capsaicin-sensitive C- and Aδ-fiber nerves that respond to chemical, thermal, and mechanical stimuli. Although these nerves have traditionally been thought to “sense” stimuli in the periphery and transmit the information centrally, there was early evidence that they also have an efferent function. It is clear that DRG neuron-derived peptides are released at peripheral sensory nerve terminals in the absence of afferent nerve stimulation. The continuous release of peptides from DRG neurons may reflect a paracrine function implying that these neurons participate in

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the continuous regulation of blood flows and other tissue activities. Sensory nerve terminals can either increase or decrease the release of CGRP in response to factors including protons, nerve growth factor, vascular wall tension, bradykinin/prostaglandins, endothelin, the sympathetic nervous system and the renin-angiotensin-aldosterone system.

CGRP AND AM SIGNALING The CGRP receptor is unique in that CGRP and AM signal through the common receptor CLR (calcitonin-like receptor). Ligand specificity is determined by co-expression of either of two chaperone proteins, receptor activity modifying proteins: RAMP1 for CGRP and RAMP2 for AM. Another RAMP (RAMP3) has also been postulated to confer AM specificity to the CLR. It now appears that a functional CGRP (or AM) receptor must include 3 proteins in a complex: the ligand binding, membrane spanning peptide, CLR, the chaperone peptide, RAMP1 or RAMP2, as well as a third peptide, the receptor component protein, RCP, that couples the receptor to the cellular signal transduction pathway. The CGRP/AM receptor is coupled to G-proteins with the CGRP/AM receptor-mediated stimulation of cAMP, downstream of Gαs, being the primary and best understood signal transduction pathway for CGRP and AM. There are, however, reports that the CGRP/AM receptor can activate other G proteins thereby providing a mechanism for fine-tuning CGRP and/or AM signaling in different cell types and tissues. Other reports indicate that CGRP and AM are capable of activating K-ATP channels of vascular smooth muscle. There is additional evidence that the vasodilator and other responses evoked by CGRP and AM are mediated, in part, by NO release, which in turn stimulates cGMP production, and that various vascular beds differ in their dependence on the endothelium for the dilator response to CGRP. Therefore, the biological activities can be mediated via endothelium-dependent and -independent mechanisms.

ATTENUATION OF OXIDATIVE STRESS BY CGRP AND AM In disease states such as hypertension, heart failure, and ischemia/reperfusion injury, overproduction of reactive oxygen (ROS) species lowers antioxidant defenses and alters signaling pathways resulting in endothelial and vascular dysfunction. Oxidative stress leads to the activation of transcription factors such as NFκ-B resulting in the up-regulation of inflammatory response genes. Multiple studies of vascular inflammation have focused on a critical role for NADPH oxidase. Endothelial, VSCMs, and adventitial cells contain NADPH oxidase. The activation of this enzyme is dependent on multiple stimuli including Ang II, norepinephrine, high glucose, shear stress, and a

number of inflammatory cytokines. Increased NADPH oxidase activity, and subsequent ROS production, plays a central role in a number of cardiovascular disease states. Indirect evidence for an antioxidant role for CGRP is provided by evidence that AM binds and activates the same receptor as CGRP. The mechanisms underlying this activity vary depending on cell types and experimental conditions. In addition, we, and other investigators, have reported that AM can attenuate Ang II-evoked ROS generation in VSMCs and ECs primarily through the inhibition of NADPH oxidase. We have recently demonstrated, for the first time, virtually identical results with CGRP. It has also been demonstrated that AM can inhibit ROS production via thiol redox systems and that CGRP can inhibit mitochondrial generated ROS in cardiac myocytes.

ATTENUATION OF INFLAMMATION BY CGRP AND AM Sensory nerves have long been considered to be involved in the development of inflammation. However, more recent studies indicate that chemical ablation of sensory nerves and genetic deletion of the sensory neuropeptide α-CGRP or the TRPV receptor, a primary activator of sensory nerve fibers, results in a marked increase in the severity of inflammation in hypertension-induced end organ damage and ischemia/reperfusion injury in the heart, liver, and gut. Likewise, AM has been implicated in both the progression and remission of the inflammatory response with the weight of the evidence indicating a protective, anti-inflammatory role for AM in a number of cardiovascular disease states. The enhanced inflammatory response in the aforementioned pathological conditions exhibits increased levels of pro-inflammatory markers and cytokines as well as tissue accumulation of neutrophils. Attenuation of inflammatory responses in the liver and the gut by sensory nerve activation appears to be due, at least in part, to a CGRP-induced increase in the endothelial production of prostacyclin, a potent anti-inflammatory agent. In vitro evidence indicates that CGRP can markedly stimulate prostacyclin production and release in endothelial cells. CGRP can significantly increase intracellular Ca2 in several different cell types including epithelial cells. In one study it was reported that this increase in intracellular Ca2 was mediated by the activation of phospholipase C (PLC). It was demonstrated that CGRP activates adenylate cyclase and releases prostacyclin from human umbilical vein endothelial cells, however, a mechanism for prostacyclin release was not established. As described previously it is well established that CGRP and can significantly induce NO production in vascular endothelial cells. It has also been reported that in endothelial cells endogenous and exogenous NO is a potent activator of cyclooygenase 1 (COX 1), a key enzyme for prostacyclin synthesis. AM treatment of bovine aortic endothelial cells induced a marked increase in cAMP accumulation and intracellular

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Ca2 through independent pathways. The intracellular free Ca2 increase resulted from PLC activation and inositol 1,4,5-triphosphate formation. This increase in free Ca2+ resulted in an activation of endothelial NO synthase (eNOS). Similarly, studies involving the AM-evoked endothelium-dependent vasorelaxation in rat aorta indicated that AM induced the PI3K/Akt pathway that was implicated in the production of NO, which in turn induced endothelium-dependent vasodilation.

INHIBITION OF APOPTOSIS BY CGRP AND AM In cardiovascular disease states such as hypertensioninduced end organ damage, heart failure, and ischemia/ reperfusion injury, there are two types of cell death, necrosis and apoptosis. Activation of pro-survival kinase cascades has been hypothesized to attenuate cell death via anti-apoptosis mechanisms. The PI3K-Akt and Erk 1/2 kinase cascades activate the cardio-protective pathway. Several lines of evidence suggest that both CGRP and AM have significant anti-apoptotic activity in vivo and in vitro (isolated rat vascular smooth muscle cells and isolated rat cardiomyocytes) in the context of cardiovascular disease states and that this is correlated with the activation of PI3K-Akt and Erk 1/2 kinase pathways. The PI3K-AKT and ERK 1/2 kinase cascades are activated by a number of receptors including G-protein coupled receptors. These pathways participate in the regulation of proliferation, differentiation, and survival. The mechanism of the pro-survival activities of these pathways is mediated, in part, by the inhibition, via phosphorylation, of the pro-apoptotic proteins BAD, BAX, BIM, inactivation of capsases, and inhibition of Bcl-2. CGRP has also been implicated in the attenuation of ischemia/reperfusion-evoked cardiac mitochondrial dysfunction through inhibition of GSK 3β via its phosphorylation by the PI3k-Akt and ERK 1/2 kinase cascades. Both CGRP and AM also appear to be protective against necrosis in cardiovascular diseases, however, it is beyond the scope of this chapter to address this issue.

CONCLUSION The CGRP/AM system represents a unique homeostatic mechanism that is relevant to normal cardiovascular function and the pathophysiology of several cardiovascular disease states. There are two related vasodilator peptides where one is a sensory neuropeptide (CGRP) and the other acts as a circulating hormone (AM). Both proteins signal through the same receptor, which has a unique composition compared to other G-protein coupled receptors. While both CGRP and AM contribute to the progression of the inflammatory response and tissue damage in some contexts, in a number of cardiovascular pathological conditions CGRP and/or AM play a compensatory protective role via the attenuation of oxidative stress, inflammation, and necrosis/apoptosis.

Further Reading Bowers MC, Katki KA, Rao A, Koehler M, Spiekerman A, DiPette DJ, et al. Role of calcitonin gene-related peptide in hypertension-induced renal damage. Hypertension 2005;46:51–7. Brain SD, Grant AD. Vascular actions of calcitonin gene-related peptide and adrenomedullin. Physiol Rev 2004;84:903–34. Dickerson I. The CGRP-receptor component protein: a regulator for CLR signaling. In: Hay DL, Dickerson IM, editors. The calcitonin generelated peptide family: form, function, and future perspectives. New York, NY: Springer  Business Media; 2010. p. 59–73. Huang R, Ma H, Karve A, DiPette DJ, Bowers MC, Supowit SC, et al. Deletion of the mouse α-calcitonin gene-related peptide gene increases the vulnerability of the heart to ischemia/reperfusion injury. Am J Physiol. Heart Circ Physiol 2008;294:1291–7. Okajima K, Harada N. Regulation of inflammatory responses by sensory neurons: Molecular mechanisms and possible therapeutic applications. Cur Med Chem 2006;13:2241–51. Prado MA, Evans-Bain B, Dickerson IM. Receptor component protein (RCP): a member of a multi-protein complex required for G-proteincoupled signal transduction. Biochem Soc Trans 2002;30:460–4. Supowit SC, Rao A, Bowers MC, Zhao H, Fink G, Patel P, et al. Calcitonin gene-related peptide protects against hypertension-induced heart and kidney damage. Hypertension 2005;45:109–14. Walker CS, Conner AC, Poyner DR, Hay DL. Regulation of signal transduction by calcitonin gene-related peptide receptors. Trends in Pharm Sci 2010;31:476–83. Yanagawa B, Nagaya N. Adrenomedullin: molecular mechanisms and its role in cardiac disease. Amino Acids 2007;32:157–64.

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27 Leptin Signaling and Energy Homeostasis Kamal Rahmouni INTRODUCTION Historically, the importance of the central nervous system in the regulation of energy homeostasis derived from the clinical observation of excessive subcutaneous fat in patients with pituitary tumors. The role of the hypothalamus in body energy storage was later established using discrete lesions or surgical transection of neural pathways [1]. For years, it was postulated that to control body fat stores, the brain must receive afferent input in proportion to the current level of body fat. The identification in 1994 of leptin [2], the ob gene product, provided powerful support for the concept of a feedback loop between the periphery and the brain for energy homeostasis (Fig. 27.1). Leptin, a 167-amino acid protein secreted by adipocytes, circulates in proportion to the adipose tissue mass. This hormone relays a satiety signal to the hypothalamus after entering the brain by a saturable specific transport mechanism. Leptin gene expression and secretion are increased by overfeeding, high fat diet, insulin and glucocorticoids and decreased by fasting and sympathetic nerve activation. The severe obesity and the hyperphagia caused by the absence of leptin or its receptor in rodents and humans make it clear that this hormone is fundamental for the control of body weight and food intake. Leptin promotes weight loss by reducing appetite and by increasing energy expenditure through stimulation of sympathetic nerve activity (Fig. 27.1). The effect of leptin on the sympathetic nervous system is an important aspect in the regulation of energy homeostasis and several other physiological functions. Leptin is also involved in regulation of glucose metabolism, sexual maturation and reproduction, the hypothalamic-pituitary-adrenal system, thyroid and growth hormone axes, angiogenesis and lipolysis, hematopoiesis, immune or proinflammatory responses, and bone remodeling (Fig. 27.1). Leptin also contributes to the regulation of cardiovascular function and appears to be involved in the pathophysiology of obesity-associated hypertension [3].

LEPTIN RECEPTOR The leptin receptor is a single transmembrane protein belonging to the cytokine-receptor super-family. Due to alternative splicing of the mRNA, at least six leptin

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00027-5

receptor isoforms have been identified (designated Ob-Ra to Ob-Rf). Five isoforms (Ob-Ra to Ob-Rd and Ob-Rf) differ in the length of their intracellular domain, while Ob-Re, which lacks the transmembrane domain, is a soluble form of the receptor [4]. The Ob-Rb form encodes the full receptor, including the long intracellular domain, which contains all the motifs necessary to stimulate the intracellular machinery involved in leptin signaling. The high levels of the short intracellular domain forms in the choroid plexus may act to transport leptin across the blood–brain barrier [4].

INTRACELLULAR MECHANISMS ASSOCIATED WITH THE LEPTIN RECEPTOR The divergent signaling capacities of the leptin receptor, ObRb, mediate the stimulation of various intracellular pathways that are important for leptin control of physiological processes (Fig. 27.2). Leptin binding to ObRb triggers the activation of the receptor-associated Janus kinase (Jak) 2 tyrosine kinase. Once activated, Jak2 phosphorylates other tyrosine residues within the ObRb including tyrosine (Tyr)1138, Tyr1077 and Tyr985, each of which mediates the activation of distinct downstream signaling pathways [5]. ObRb activation also promotes the activation of phosphatidylinositol 3 kinase, although this appears to be cell-type specific and the mechanisms underlying this regulation remain unclear. Whereas phosphorylated Tyr1138 of ObRb recruits and activates signal transducer and activator of transcription STAT3, the phosphorylation of Tyr1077 promotes the tyrosine phosphorylation and activation of STAT5. Activated STAT3 and STAT5 translocate to the nucleus to modulate gene transcriptional with important implications for the regulation of metabolism and body energy balance. For instance, knock-in mice that have a mutation at Tyr1138 of ObRb, which disrupts leptininduced STAT3 signaling, are severely obese and hyperphagic, but in contrast to the mice lacking leptin or ObRb these mice remain fertile and are less diabetic [5]. On the other hand, phosphorylation of Tyr985 creates a binding site for the COOH-terminal SH2 domain of the tyrosine phosphatase, PTPN11 (also known as SHP2), leading to the activation of extracellular signal-related kinase (ERK)

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-Insulin secretion and sensitivity -Kidney function -Endothelium function -Hematopoiesis -Immune system -Angiogenesis -Thermogenesis -Reproduction -Wound healing

↑ SNA

↑ Leptin ↓ Food intake

↑ Energy expenditure -Blood pressure -Neuroendocrine function -Osteogenesis -Reproduction -Insulin secretion -Glucose homeostasis

Fat tissue

FIGURE 27.1 Role of leptin in the negative loop regulating body weight. Leptin is secreted by adipocytes and circulate in the blood in concentration proportional to fat mass content. Action of leptin on its receptor in the brain inhibits food intake and increases energy expenditure through stimulation of sympathetic nerve activity (SNA). This leads to a decrease in adipose tissue mass and body weight. In addition, leptin is involved in the regulation of several physiological processes either directly or through its action in the brain.

ObRb

Tyr

P

IRS IRS

Tyr985

P

Tyr1138

P

Shp2 Shp2

P Tyr1077

STAT5 STAT5

STAT3 STAT3

ERK ERK

PI3K PI3K

FIGURE 27.2 Molecular mechanisms involved in leptin receptor (ObRb) signaling. In the central nervous system, there are four primary intracellular signaling pathways that emanate from ObRb: signal transducer and activator of transcription STAT3 and STAT5 proteins, phosphoinositol-3 kinase (PI3K) and the extracellular signal-related kinase (ERK). Each pathway is activated by phosphorylation of specific tyrosine residues in ObRb.

signaling pathway. While Tyr985 mediates most ERK stimulation during ObRb signaling, tyrosine phosphorylation sites on Jak2 appear to account for a fraction of ERK activation by leptin independently from ObRb phosphorylation. Tyr985 also binds suppressors of cytokine signaling-3 which act as a negative regulator to inhibit STAT3 signaling.

SITES OF LEPTIN ACTION IN THE BRAIN The arcuate nucleus of the hypothalamus is considered a major site for the regulation of physiological processes by leptin [5–7]. Supporting this view, the arcuate nucleus contains the highest concentration of ObRb and

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is the most responsive brain nucleus to leptin in terms of activation of intracellular signaling pathways associated with the leptin receptor (e.g. STAT3). The relevance of leptin signaling in the arcuate nucleus was further supported by lesioning studies which demonstrated a lack of feeding response to leptin after the arcuate nucleus had been destroyed. In addition, restoration of the expression of ObRb in the arcuate nucleus of leptin receptor-deficient Koletsky rats or in mice that have a leptin receptor null allele leads to decreased food intake and body weight. These findings indicate that leptin receptor signaling in the arcuate nucleus is critical to the effects of leptin on energy homeostasis. However, recent evidence demonstrating ObRb expression and leptin actions in several other brain regions has led to the concept of a distributed brain network of leptin action [8].

INTERACTION OF LEPTIN AND NEUROPEPTIDES IN THE HYPOTHALAMUS After activation of leptin receptors in the central nervous system, the signal is transduced by a series of integrated neuronal pathways that regulate endocrine and autonomic function [1,6]. Although the arcuate nucleus contains several neuronal populations, two classes of neurons have been well characterized with regard to leptin action: proopiomelanocortin (POMC) neurons (which also express cocaine- and amphetamine-related transcripts (CART) that are activated by leptin; and neuropeptide Y (NPY) neurons (which also express agouti-related protein (AgRP) that are inhibited by leptin (Fig. 27.3).

Melanocortin System There is strong evidence that many of leptin’s actions are mediated by stimulation of the melanocortin system [6,7,9]. The melanocortins are peptides that are processed from the polypeptide precursor POMC, which is produced by neurons in the arcuate nucleus of the hypothalamus and the nucleus of the tractus solitarius. POMC neurons are known to express the leptin receptor, and leptin binding leads to stimulation of neuronal firing activity and increase the gene expression of POMC and CART genes. This result in higher secretion of alpha-melanocyte stimulating hormone (α-MSH), which in turn binds to a number of a family of melanocortin receptors. Five melanocortin receptors (MC-1R to MC-5R) have been identified. The MC-3R and MC-4R are highly expressed in the central nervous system [9]. The critical role for MC-4R in energy balance was demonstrated by target disruption of the MC-4R gene inducing hyperphagia and obesity in mice [9,10]. Antagonism of a central melanocortin receptor is also important in the regulation of energy homeostasis [7,9]. This concept emerged with the discovery of production within the hypothalamic neuron of a potent and selective

Leptin

⊕

NPY neuron

POMC neuron

α −MSH

NPY

AgRP

-

⊕

MC3/4-R

NPY-R

Energy balance FIGURE 27.3 Schematic illustration of the leptin-sensitive neuronal populations in the hypothalamic arcuate nucleus; those activated (catabolic pathway represented by proopiomelanocortin (POMC) neurons) and those inhibited (anabolic pathway represented by the neuropeptide Y (NPY) neurons which also express agouti related protein, AgRP). In POMC neurons, leptin increases neuronal firing and POMC gene expression promoting the secretion of alpha-melanocyte stimulating hormone (α-MSH), an agonist of the melanocortin 3 and 4 receptors (MC3/4-R) located in the second order neurons. Conversely, in NPY neurons, leptin inhibits the neuronal firing rate and decreases the expression and secretion of NPY and AgRP (antagonist of the MC3/4-R), promoting MC3/4-R activation and NPY-R inhibition, respectively. Leptin suppression of the NPY anabolic pathway and stimulation of the POMC catabolic pathway reduce food intake and promote thermogenesis (resulting in a decrease in body weight).

antagonist of MC-3R and MC-4R. This molecule known as agouti related peptide (AgRP) is expressed only in the arcuate nucleus of the hypothalamus by the same neurons that express NPY. The expression levels of AgRP are upregulated by fasting and by leptin deficiency.

Neuropeptide Y NPY, a 36-amino acid peptide, is the most potent orexigenic (promote increased energy intake) peptide activated by decreases in leptin [9]. In the hypothalamus, NPY is synthesized by neurons of the arcuate nucleus and secreted from their terminals in the paraventricular nucleus and lateral hypothalamus. Injection of NPY into the cerebral ventricles or direct hypothalamic administration increases food intake and promotes obesity. In NPY neurons, leptin inhibits neuronal firing and decreases the expression of NPY and AgRP genes. Accordingly, levels of NPY are dramatically increased in the hypothalamus of leptin-deficient mice. Moreover, knockout of the NPY gene reduces the obesity and other endocrine alterations resulting from chronic leptin deficiency in ob/ob mice by about 50% [9].

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Other Mediators

CONCLUSION

The complicated nature of leptin signaling pathways may be suggested from the essentially normal phenotype of NPY knockout mice [9], despite the potent stimulatory effects of NPY on food intake and body weight. This suggests that there are complementary and/or overlapping effector systems that compensate for the absence of NPY. Subsequently, other candidate molecules that can mediate the effects of leptin have been identified. For example, leptin-dependent sympathetic activation to brown adipose tissue appears to be mediated by corticotrophin releasing factor as the sympathoexcitatory effect of leptin to this tissue was substantially inhibited by pretreatment with the corticotrophin releasing factor receptor antagonist [10]. Several hypothalamic neuropeptides, monoamines, and other transmitter substances have emerged as candidate mediators of leptin action in the central nervous system (Table 27.1). TABLE 27.1 Example of neuropeptide and monoamine Candidate mediators in the Transduction of Leptin Action in the Central nervous System Catabolic Molecules

Anabolic Molecules

Proopiomelanocortin (POMC) and Neuropeptide Y (NPY) derived peptides Corticotrophin releasing factor (CRF)

Agouti related peptide (AgRP)

Cocaine-and amphitamineregulated transcript (CART)

Melanin-concentrating hormone (MCH)

Urocortin

Hypocritin 1 and 2/Orexin A and B

Neurotensin

Galanin

Interleukin 1β

Noradrenaline

Glucagon like peptide 1 Oxytocin Neurotensin

Energy balance is a highly regulated phenomenon, and the importance of the central nervous system in this regulation of energy homeostasis is well-established. The centers of regulation of food intake and body weight are distributed throughout the central nervous system with the hypothalamus playing a major role. The discovery of leptin has illuminated this field of neuroscience. This hormone constitutes the signal from adipose tissue that acts in the brain to complete the feedback loop that regulates appetite and energy expenditure. The identification of the leptin receptor and its sites of action in the brain have resulted in the striking progress in dissecting the brain circuitries that regulate energy homeostasis. Despite the lack of many pieces of this puzzle, the network of the brain pathways that control energy balance is rapidly being defined.

References [1] Elmquist JK, Elias CF, Saper CB. From lesions to leptin: hypothalamic control of food intake and body weight. Neuron 1999;22:221–32. [2] Zhang Y, Proenca R, Maffei M, Barone M, Leopold L, Friedman JM. Positional cloning of the mouse obese gene and its human homologue. Nature 1994;372:425–32. [3] Rahmouni K. Obesity, sympathetic overdrive and hypertension: the leptin connection. Hypertension 2010;55:844–5. [4] Tartaglia LA. The leptin receptor. J Biol Chem 1997;272:6093–6. [5] Myers Jr MG. Deconstructing leptin: from signals to circuits. Diabetes 2010;59:2708–14. [6] Morton GJ, Cummings DE, Baskin DG, Barsh GS, Schwartz MW. Central nervous system control of food intake and body weight. Nature 2006;443:289–95. [7] Flier JS. Obesity wars: molecular progress confronts an expanding epidemic. Cell 2004;116:337–50. [8] Grill HJ. Distributed neural control of energy balance: Contributions from hindbrain and hypothalamus. Obesity 2006;14:216–21. [9] Inui A. Transgenic approach to the study of body weight regulation. Pharmacol Rev 2000;52:35–61. [10] Rahmouni K, Haynes WG. Leptin and the cardiovascular system. Recent Prog Horm Res 2004;59:225–44.

Serotonin Dopamine

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C H A P T E R

28 The Endothelin System Ernesto L. Schiffrin INTRODUCTION: ENDOTHELIN SYSTEM COMPONENTS The 21 amino acid peptide endothelin (ET) was isolated and cloned in 1985 by Yanagisawa et al. [1] ET-1, -2 and -3 are isopeptides with different functions [2], and there are also larger 31 and 32-amino acid peptides. ET-1 is the most abundant ET produced in blood vessels and the kidney, whereas ET-3 is mainly a neuropeptide. In endothelial and other cells, furin and other enzymes act on proETs to generate 38–39 amino acid peptides (big ETs) that are converted into mature 21-amino acid ETs by zinc-dependent endoproteases, the endothelin-converting enzymes (ECE-1 and 2). ECEs cleave big ET-1 at the Val21-Trp22 bond, yielding ET-1. ECE-1, of which there are four differentially spliced isoforms encoded by a single gene resulting from four alternative promoters, is present in endothelial cells. The four isoforms of ECE-1 differ by their N-terminal amino-acid, which is responsible for their cellular localization. ECE-1a, c and d are extracellular, whereas ECE-1b is an intracellular enzyme which heterodimerizes with other ECE-1 isoforms and regulates their activity. ECE-2 in smooth muscle cells converts big ET-1 to ET-1 in the vicinity of ET receptors, thus protecting it from degradation. Other enzymes that generate ETs include matrix metalloproteinase-2 that cleaves the Gly32-Leu33 bond to generate ET-1[1–32], chymase from mast cells that cleaves big ET-1 at the Tyr31-Gly32 peptide bond, yielding ET-1[1–31], and neutral endopeptidase, but their physiological importance is unclear. ET production is modulated by inhibitors such as shear stress (in blood vessels) and nitric oxide (NO), and stimulators (epinephrine, thrombin, angiotensin II (Ang II), vasopressin, cytokines, insulin, growth factors (TGF-β1) and hypoxia). Leptin stimulates ET-1 generation by endothelial cells, which may be a mechanism involved in vascular injury in the metabolic syndrome and in type 2 diabetes mellitus. ET-1 induces potent vasoconstriction, inflammation and cell growth by acting on ETA and ETB receptors present in the vascular wall on smooth muscle cells, whereas endothelial cells only possess ETB receptors which stimulate release of NO and prostacyclin, mediating vasodilation [2]. ETA or ETB receptors appear to predominate in the

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00028-7

adrenal gland depending on the species [3]. ETA receptors predominate in the kidney. ETB receptors are the predominant receptor subtype in the brain and peripheral nervous system.

EFFECTS OF PREPROET-1 OR ET RECEPTOR GENE DELETION Inactivation in mice of the ET-1 gene or the ETA receptor gene results in minor blood pressure elevation [4]. This occurs as a consequence of abnormal craniofacial development that interferes with breathing and raises blood pressure through anoxia. The aorta exhibits developmental abnormalities as well, with the phenotype resembling the Pierre Robin syndrome. ET-3 is the cognate ligand of ETB receptors and acts mainly on neural or neurally derived tissues. It regulates migration of neural crest cells, and mutations or gene inactivation of ETB receptors induce aganglionic megacolon and pigmentary abnormalities. Hereditary and sporadic human aganglionic megacolon (Hirschprung’s disease) result in some cases from mutations of the ETB receptor gene. Heterozygous ETB receptor knockout mice have slightly elevated blood pressure, which supports the hypothesis that the physiological action of ETB receptors is vasodilatory.

MECHANISM OF ACTION OF ET ET receptors stimulate phospholipase C, inositol trisphosphate generation and calcium release, leading to calmodulin activation, diacylglycerol production and protein kinase C stimulation [2]. The ras-raf-mitogen activated kinase (MAPK) cascade and non-receptor tyrosine kinases also participate in the intracellular signaling pathways activated by ET receptor stimulation. Reactive oxygen species (ROS) generation by reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, xanthine oxidase, mitochondria, and uncoupled NO synthase contribute to intracellular signaling via growth factor receptor transactivation and MAPK activation. ETA receptors also induce cell growth and apoptosis through NFκB activation. ETB receptors may also have apoptotic effects.

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28. THE EndoTHElIn SySTEm

PreproET-1 N

C Furinor PC7 N

ET-1[1-32] MMP-2

Big ET-1(38-39 aa)

RENAL EFFECTS OF ET-1

C ECE-1 M t Met

Leu

Ser Ser

Cys

Ser Cys

Asp

N

ET‐1(1‐21)

Lys Glu Cys

Val

Tyr

Phe Cys

ETA receptors

His

Leu Asp

ETB receptors

Ile

mice have enhanced vascular lipid biosynthetic enzyme gene expression, and demonstrate increased atherosclerosis when crossed with apoE knockout mice [11].

Ile

Trp C

ET-3

FIGURE 28.1 Biosynthesis and molecular structure of endothelin-1 (ET-1). PreproET-1 is cleaved by furin or protein convertase (PC)7 to generate the 38 or 39 amino acid (aa) big ET-1. The latter is cleaved by endothelin convertase-1 (ECE-1a, b, c, d) to generate the mature 21-aa ET-1, which can bind with high affinity to both ETA and ETB receptors. ET-2 and ET-3 exhibit 2 and 5 aa differences respectively. ET-3 is the cognate ligand of ETB receptors. ET-1 [1-31] is an additional peptide of the system produced in the vasculature and the airway that functions as a vasoconstrictor of tracheal and vascular smooth muscle and could be involved in allergic inflammation ET-1 [1-32] is generated in the vascular wall by the action of matrix metalloproteinase-2 (MMP-2).

PATHOPHYSIOLOGY OF THE ENDOTHELIN SYSTEM IN EXPERIMENTAL MODELS The endothelin system plays a pathophysiological role in hypertension [5–7], atherosclerosis, coronary artery disease, heart failure, subarachnoid hemorrhage and cerebral vasospasm, diabetes, primary pulmonary hypertension (the only approved indication of ET antagonists), pulmonary fibrosis, scleroderma, diabetic and non-diabetic renal disease, renal failure, hepatorenal syndrome, glaucoma, prostate cancer and its metastasis, and may also be involved in pheochromocytoma. In salt-dependent models of experimental hypertension such as DOCA-salt hypertension or Dahl salt-sensitive rats, and in severe hypertension such as stroke-prone spontaneously hypertensive rats (SHRsp), in particular when salt-loaded or treated with the nitric oxide synthase inhibitor L-NAME, enhanced production of ET-1 [8] induces hypertrophic remodeling of large and small arteries rather than the usual “eutrophic” remodeling without true vascular hypertrophy more often found in essential hypertension and in SHR, which regresses after treatment with ET antagonists [6–9]. In a mouse that overexpresses human preproET-1 in the endothelium by use of the endothelium-specific promoter Tie-2, small artery hypertrophic remodeling, vascular inflammation and endothelial dysfunction occurs despite the fact that blood pressure is not elevated, demonstrating the ability of ET-1 to induce blood pressure-independent vascular remodeling [10]. These

Although salt loading stimulates ET-1 production, activation of renal ETB receptors inhibits sodium reabsorption [12]. In Ang II-infused mice, the dual ETA/ETB receptor blocker bosentan partially prevented activation of the procollagen gene and rats overexpressing human angiotensinogen and human renin which develop malignant hypertension, exhibited reduced renal and myocardial damage after treatment with bosentan. In salt-loaded SHRsp, increased expression of ET-1 was associated with enhanced generation in the kidney of transforming growth factor (TGF)-β1, basic fibroblast growth factor (bFGF), procollagen I expression and matrix metalloproteinase (MMP)-2 activity, and were reduced by a selective ETA antagonist. ETs are implicated in both the development and progression of chronic kidney disease (CKD) [13]. The major pathological effects in CKD are ETA receptor-mediated. In a recent study, selective ETA receptor antagonism slightly reduced blood pressure, and to greater degree proteinuria and arterial stiffness in renal patients suggesting that the reduction in proteinuria and arterial stiffness is partly independent of blood pressure, and that selective ETA receptor antagonism may confer cardiovascular and renal benefits in patients with CKD [14,15].

CARDIAC EFFECTS OF ET-1 TGF-β1 expression and collagen deposition in the heart of DOCA-salt hypertensive rats are increased, and are prevented by ETA blockade. ETA receptor antagonism also blocked the expression of inflammatory mediators (NFκB and adhesion molecules) and the anti-apoptotic molecule X inhibitor of apoptosis peptide (xIAP) [16]. ETA receptor blockade prevented aldosterone-induced cardiac and vascular fibrosis, which suggests that ET-1 mediates in part effects of aldosterone on the heart and blood vessels [17]. Patients with essential hypertension, primary aldosteronism or renovascular hypertension have enhanced ultrasound backscatter signals resulting from tissue heterogeneity in the myocardium, which correlates with plasma aldosterone and immunoreactive ET, suggesting that as in experimental animal models, aldosterone and ET-1 contribute to myocardial fibrosis in human hypertension. However, studies using ETA receptor antagonists in heart failure patients have not demonstrated efficacy in chronic studies, although acute short-term trials suggested beneficial effects.

ET-1 IN ESSENTIAL HYPERTENSION In primary human hypertension plasma concentrations of immunoreactive ET are normal in Caucasians,

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EndoTHElInS And THE nERvouS SySTEm

SFO Hypothalamus AP

PVN VLM PP

Sympathetic Outflow Baroreceptors

Vasopressin

Kidney

Blood vessels

NTS

Sympathetic ganglia

Heart

Adrenal medulla

FIGURE 28.2 Role of endothelins (ETs) in the brain and sympathetic nervous system. ETs produced in rostral areas of the brain including the subfornical organ (SFO) act on the hypothalamus to stimulate secretion of vasopressin and on the brainstem to stimulate cardiovascular regulatory centers. The latter, in the ventrolateral medulla (VLM) area postrema (AP) and nucleus tractus solitariius (NTS) modulated by baroreceptor input, stimulate sympathetic outflow, which via sympathetic ganglia, modulates heart rate, vascular tone, kidney blood flow and renin secretion as well as water and sodium handling, and catecholamine release by the adrenal medulla. ETA receptors are involved at most stages, except some in which ETB receptors play a role.

but elevated in African-Americans, who have a volume expanded low-renin form of hypertension and in whom an increase in vascular smooth muscle vasoconstrictor ETB receptors has been documented. High plasma ET levels may be related to subclinical renal dysfunction and smoking rather than hypertension. However, vascular mRNA levels of preproET are increased in stage 2 hypertension of the JNC 7 classification [7]. ETA receptor antagonists caused greater vasodilatation in the forearm of hypertensive than normotensive subjects, suggesting that ETA receptors play an important role in ET-1-dependent vascular tone in essential hypertension [18,19]. The ETB blocker BQ-788 constricted forearm resistance arteries in normotensive subjects, which suggests that ETB receptors are vasodilator in normotensive subjects, whereas the forearm in hypertensive subjects was vasodilated, suggesting that ETB receptors are vasoconstrictor in hypertensive patients. Whereas in normotensive subjects forearm blood flow response to the ETA receptor blocker BQ-123 was similar in white and black subjects, ETA receptor antagonism was a more potent vasodilator in blacks than in whites among hypertensive individuals although ET-1 induced equipotent vasoconstriction, which suggests that increased ETA-mediated vasoconstriction

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may contribute to blood pressure elevation in hypertensive blacks. ET-1 at concentrations of 1011 mol/L may potentiate other vasoconstrictors (e.g. phenylephrine or serotonin), a mechanism which is under the influence of the EDN1 K198N polymorphism in the coding region of the preproET-1 gene, and could be enhanced in hypertension, contributing to blood pressure elevation [20]. Clinical trials in human hypertension with the ETA/ETB antagonist bosentan [21] and the ETA antagonist darusentan resulted in reduced blood pressure. However, ETA antagonists may induce liver damage and liver enzyme elevation, which has resulted in halting of human hypertension studies with ETA antagonists.

MOLECULAR GENETICS OF THE ENDOTHELIN SYSTEM A polymorphism (EDN1 K198N) in the coding region of the prepro-ET-1 gene has been associated with hypertension in overweight individuals [22], and with increased vasoreactivity [20]. ECE1 C-388A is a polymorphism present in the 5-regulatory region of the ECE-1b gene. It results in enhanced expression of ECE-1b, with increased generation of ET-1. Its presence was reported in untreated hypertensive German women in whom the A allele had effects on daytime and night-time BP, and in women in the French epidemiological study Étude du Vieillissement Artériel (EVA). However, the EDN1 K198N polymorphism of preproET-1 was not associated with BP in either men or women in this study, but interacted with the ECE1 C-338A variant to influence systolic and mean BP levels in women. Stimulation of ET by androgens could explain the absence of effect in males.

ENDOTHELINS AND THE NERVOUS SYSTEM ETs are found in rostral portions of the brain, in the hypothalamus (paraventricular nuclei, colocalized with vasopressin containing neurons) and the dorsal motor nucleus of the vagus and medulla oblongata as well as brainstem areas [23] known to regulate cardiovascular function such as the area postrema, the ventrolateral medulla and the nucleus tractus solitarius. ETs cans stimulate the central and peripheral sympathetic nervous system and act on the carotid bodies and on cervical superior and nodose ganglia, influencing hemodynamic regulation by the baroreflex and by chemoreflexes [24–29]. Also, ETs released by postganglionic sympathetic neurons modulate catecholamine release and vascular tone. They also stimulate catecholamine release from adrenal glands via ETA and possibly ETB receptors [30]. ET release has been implicated in vasospasm in the cerebral circulation, and particularly in the vasospasm

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28. THE EndoTHElIn SySTEm

of subarachnoid hemorrhage (SAH). In the latter condition, high concentrations of immunoreactive ET have been reported in cerebrospinal fluid (CSF). Although in experimental models of SAH endothelin antagonists have demonstrated some efficacy in combatting vasospasm, ET antagonists are not indicated in SAH in humans. Recently, the interactions between the sympathetic nervous system and endogenous endothelin in patients with essential hypertension have been studied by recording muscle sympathetic nerve activity during infusion of an ETA antagonist [31]. Endogenous ET-1 appeared to have a sympatho-excitatory effect both in normotensive and hypertensive subjects through ETA receptors, contributing to basal sympathetic vasomotor tone. Essential hypertensive subjects demonstrated an increased susceptibility to the sympatho-excitatory effect of endogenous ET-1.

CONCLUSION ET-1 is a potent vasoconstrictor that also promotes cardiac, vascular and renal inflammation, hypertrophy and fibrosis. ET receptor antagonists could prevent some of the complications of hypertension, atherosclerosis and diabetes, and it is possible that they could achieve blood pressure-independent cardiovascular protection. However, because of side effects, their potential usefulness in hypertension, heart failure, atherosclerosis, CKD, diabetes and other diseases cannot be currently exploited. The only approved indication to date of ETA receptor blockade is primary pulmonary hypertension. Whether the role of ETs on the central and sympathetic nervous system will allow their therapeutic use in humans in the future is uncertain.

Acknowledgements The work of the author was supported by grant 37917 from the Canadian Institutes of Health Research (CIHR), a Canada Research Chair (CRC) from the CIHR/ Government of Canada CRC Program, and a Canada Fund for Innovation grant.

References [1] Yanagisawa M, Kurihara H, Kimura S, Tomobe Y, Kobayashi M, Mitsui Y, et al. A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature 1988;332:411–5. [2] Schiffrin EL, Touyz RM. Vascular biology of endothelin. J Cardiovasc Pharmacol 1998;32:S2–S13. [3] Davenport AP, Hoskins SL, Kuc RE, Plumpton C. Differential distribution of endothelin peptides and receptors in human adrenal gland. Histochem J 1996;28:779–89. [4] Clouthier DE, Hosoda K, Richardson JA, Williams SC, Yanagisawa H, Kuwaki T, et al. Cranial and cardiac neural crest defects in endothelin-A receptor-deficient mice. Development 1998;125:813–24. [5] Larivière R, Thibault G, Schiffrin EL. Increased endothelin-1 content in blood vessels of deoxycorticosterone acetate-salt hypertensive but not in spontaneously hypertensive rats. Hypertension 1993;21:294–300.

[6] Schiffrin EL. Endothelin: Potential role in hypertension and vascular hypertrophy. Hypertension 1995;25:1135–43. [7] Schiffrin EL, Deng LY, Sventek P, Day R. Enhanced expression of endothelin-1 gene in resistance arteries in severe human essential hypertension. J Hypertens 1997;15:57–63. [8] Goel A, Su B, Flavahan S, Lowenstein CJ, Berkowitz DE, Flavahan NA. Increased endothelial exocytosis and generation of endothelin-1 contributes to constriction of aged arteries. Circ Res 2010;107:242–51. [9] Li JS, Larivière R, Schiffrin EL. Effect of a nonselective endothelin antagonist on vascular remodeling in deoxycorticosterone acetatesalt hypertensive rats. Evidence for a role of endothelin in vascular hypertrophy. Hypertension 1994;24:183–8. [10] Amiri F, Virdis A, Neves MF, Iglarz M, Seidah NG, Touyz RM, et al. Endothelium-restricted overexpression of human endothelin-1 causes vascular remodeling and endothelial dysfunction. Circulation 2004;110:2233–40. [11] Simeone SMC, Li Melissa W, Paradis P, Schiffrin EL. Vascular gene expression in mice overexpressing human endothelin-1 targeted to the endothelium. Physiol Genomics 2011;43:148–60. [12] Kohan DE, Rossi NF, Inscho EW, Pollock DM. Regulation of blood pressure and salt homeostasis by endothelin. Physiol Rev 2011;91:1–77. [13] Saleh MA, Boesen EI, Pollock JS, Savin VJ, Pollock DM. Endothelin-1 increases glomerular permeability and inflammation independent of blood pressure in the rat. Hypertension 2010;56:942–9. [14] Dhaun N, MacIntyre IM, Melville V, Lilitkarntakul P, Johnston NR, Goddard J, et al. Blood pressure-independent reduction in proteinuria and arterial stiffness after acute endothelin-A receptor antagonism in chronic kidney disease. Hypertension 2009;54:113–9. [15] Wenzel RR, Littke T, Kuranoff S, Jurgens C, Bruck H, Ritz E, et al. Avosentan reduces albumin excretion in diabetics with macroalbuminuria. J Amer Soc Nephrol 2009;20:655–64. [16] Ammarguellat FZ, Gannon PO, Amiri F, Schiffrin EL. Fibrosis, matrix metalloproteinases, and inflammation in the heart of DOCAsalt hypertensive rats: role of ET(A) receptors. Hypertension 2002;39:679–84. [17] Pu Q, Neves MF, Virdis A, Touyz RM, Schiffrin EL. Endothelin antagonism on aldosterone-induced oxidative stress and vascular remodeling. Hypertension 2003;42:49–55. [18] Cardillo C, Kilcoyne CM, Waclawiw M, Cannon RO, Panza JA. Role of endothelin in the increased vascular tone of patients with essential hypertension. Hypertension 1999;33:753–8. [19] Haynes WG, Hand MF, Johnstone HA, Padfield PL, Webb DJ. Direct and sympathetically mediated venoconstriction in essential hypertension. Enhanced responses to endothelin-1. J Clin Invest 1994;94:1359–64. [20] Iglarz M, Benessiano J, Philip I, Vuillaumier-Barrot S, Lasocki S, Hvass U, et al. Preproendothelin-1 gene polymorphism is related to a change in vascular reactivity in the human mammary artery in vitro. Hypertension 2002;39:209–13. [21] Krum H, Viskoper RJ, Lacourcière Y, Budde M, Charlon V. The effect of an endothelin-receptor antagonist, bosentan, on blood pressure in patients with essential hypertension. Bosentan Hypertension Investigators. N Engl J Med 1998;338:784–90. [22] Tiret L, Poirier O, Hallet V, McDonagh TA, Morrison C, McMurray JJV, et al. The Lys198Asn polymorphism in the endothelin-1 gene is associated with blood pressure in overweight people. Hypertension 1999;33:1169–74. [23] Giaid A, Gibson SJ, Herrero MT, Gentleman S, Legon S, Yanagisawa M, et al. Topographical localisation of endothelin mRNA and peptide immunoreactivity in neurons of the human brain. Histochemistry 1991;95:303–14. [24] Dai X, Galligan JJ, Watts SW, Fink GD, Kreulen DL. Increased .O2– production and upregulation of ETB receptors by sympathetic neurons in DOCA-salt hypertensive rats. Hypertension 2004;43:1048–54.

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ConCluSIon

[25] Dai SM, Shan ZZ, Miao CY, Yin M, Su DF. Hemodynamic responses to endothelin-1 and endothelin antagonists microinjected into the nucleus tractus solitarius in rats. J Cardiovasc Pharmacol 1997;30:475–80. [26] Itoh S, van den Buuse M. Sensitization of baroreceptor reflex by central endothelin in conscious rats. Amer J Physiol 1991;260:H1106–H1112. [27] Kopp UC, Grisk O, Cicha MZ, Smith LA, Steinbach A, Schluter T, et al. Dietary sodium modulates the interaction between efferent renal sympathetic nerve activity and afferent renal nerve activity: role of endothelin. Amer J Physiol Regul Integr Comp Physiol 2009;297:R337–51.

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[28] Lau YE, Galligan JJ, Kreulen DL, Fink GD. Activation of ETB receptors increases superoxide levels in sympathetic ganglia in vivo. Amer J Physiol Regul Integr Comp Physiol 2006;290:R90–5. [29] Mosqueda-Garcia R, Inagami T, Appalsamy M, Sugiura M, Robertson RM. Endothelin as a neuropeptide: cardiovascular effects in the brainstem of normotensive rats. Circ Res 1993;72:20–35. [30] Yamaguchi N. Role of ET(A) and ET(B) receptors in endothelin-1induced adrenal catecholamine secretion in vivo. Amer J Physiol 1997;272:R1290–R1297. [31] Bruno RM, Sudano I, Ghiadoni L, Masi L, Taddei S. Interactions between sympathetic nervous system and endogenous endothelin in patients with essential hypertension. Hypertension 2011;57:79–84.

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29 Pharmacology of the Nucleous Tractus Solitarii Ching-Jiunn Tseng, Pei-Wen Cheng, Che-Se Tung The nucleus tractus solitarii (NTS) is the primary integrative center for cardiovascular control and other autonomic functions in the central nervous system (CNS). The NTS not only integrates covergent information but itself is the site of substantial modulation. Several neurotransmitters or neuromodulators are involved in cardiovascular regulation in the NTS. This section presents some of the recent findings and their molecular mechanisms in the NTS.

NUCLEUS TRACTUS SOLITARII The NTS is located in the dorsal aspect of the medulla oblongata, and receives visceroceptive information from cardiopulmonary and gastrointestinal sites. It is a bilateral structure, that when extending caudally, comes medial and close to the walls of the fourth ventricle. At the level of the posterior tip of the area postrema, the neuronal groups within the NTS fuse medially and form the commisural nucleus of the NTS. This part of the NTS lies dorsal to the central canal and the dorsal motor nucleus of the vagus. The caudal limit of the NTS is located at the level of the pyramidal decussation [1]. The NTS has long been identified as a site where the first synape of the baroreceptor reflex is located. Therefore the NTS, as well as other key central nuclei in the hypothalamus and other forebrain regions, have important roles in mediating cardiovascular responses to acute stresses. From the NTS, pathways project to cholinergic parasympathetic neurons located in the dorsal vagal motor nucleus and the nucleus ambiguus, as well as to a group of inhibitory neurons in the caudal ventrolateral medulla (CVLM), which, in turn, project to tonically active sympathoexcitatory neurons in the rostral ventrolateral medulla (RVLM). A rise in arterial blood pressure in the periphery is detected by baroreceptors located in the carotid arteries and aortic arch and these activate neurons in the NTS via the branches of the vagus and glossopharyngeal nerves. The activation of NTS neurons stimulates inhibitory neurons in the CVLM, which attenuate the activity of the RVLM neurons, leading to

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00029-9

a reduction in sympathetic activity and a fall in blood pressure [2]. Hypertension can be due to central interruption of baroreflexes in the NTS since electrolytic lesions of blood flow was similar to that produced by sinoaortic denervation [3]. An analogous condition has also been found in the clinic: a very rare case of continuous hypertension and tachycardia after excision of a cerebellar hemangioblastoma at the dorsal medulla oblongata showed a small injury at the dorsocaudal medulla that was located at the caudal site of the NTS by postoperative MR imaging [4]. The NTS is richly innervated by neurons containing a number of potential neurotransmitters and neuromodulators, such as glutamate, noradrenaline, adrenaline, acetylcholine, serotonin, angiotensin II (Ang II), vasopressin, β-endorphin, enkephalins, neuropeptide Y, adenosine, nitric oxide (NO), insulin, etc. Among these neuromodulators related to central cardiovascular regulation, this chapter addresses the cardiovascular effects of NO, insulin, adenosine and Ang II and their molecular signaling pathways in the NTS.

CARDIOVASCULAR EFFECTS OF NO IN THE NTS NO, synthesized from the semiessential amino acid L-arginine by nitric oxide synthase (NOS), is a remarkable regulatory molecule and plays an important role in physiological functions. By using immunohistochemistry, NADPH-diaphorase staining and autoradiography, neuronal NOS (nNOS) was found at a high concentration in regions of the brain stem, especially in the NTS and rostral ventrolateral medulla. Unilateral microinjection of L-arginine into the NTS produced prominent doserelated depressor and bradycardic effects and reduced renal sympathetic nerve activity [5]. In addition, 4 to 6 hours after intravenous injection of bacterial endotoxinlipopolysaccharide, there was a time-related potentiation of the L-arginine-induced depressor and bradycardic effects in the NTS. In the investigation of the mechanisms of action of NO in the NTS, evidence suggests that

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29. PHARmACology of THE NuClEouS TRACTuS SolITARII

the cardiovascular effects produced by L-arginine in the NTS are inhibited by pharmacological interventions that block NO production and the cGMP-PKG signaling pathway within the nucleus [5]. Furthermore, the interactions among NO and other important neurotransmitters have been investigated in the NTS. NO, adenosine and glutamate have reciprocal inhibition in the release of other transmitters [6,7].

CARDIOVASCULAR EFFECTS OF ADENOSINE IN THE NTS The endogenous nucleoside adenosine has been studied for its potential role as a neuromodulator in a number of autonomic functions. The highest density of adenosine uptake sites within the CNS has been observed in the NTS. The natural precursor of adenosine in the CNS is ATP, which is dephosphorylated and rapidly becomes adenosine via the activity of ectonucleotidases to synaptic release. Among four adenosine receptor subtypes, A2A receptor is expressed at high levels in limited regions of the brain that are primarily linked to adenylyl cyclase activation. In the NTS, activation of A2A receptors has been demonstrated to elicit dose-related decreases in blood pressure (BP). These studies suggest that the cardiovascular modulatory effects of adenosine in the NTS are mediated predominantly by the A2A receptor. Based on the finding, adenosine has been shown to play an important role in central cardiovascular control. The depressor and bradycardic effects of adenosine in the NTS are attenuated by a NOS inhibitor [6]. Endothelial NOS (eNOS), originally identified in vascular endothelium, is expressed in several nonendothelial cell types, including neurons of various rat brain regions. In addition, eNOS-generated NO in the NTS plays a role in the control of baroreflex gain and arterial pressure [8]. Suppression of the endogenous eNOS by overexpression of dominant-negative eNOS in bilateral NTS increased spontaneous baroreceptor reflex gain in conscious Wistar rats and decreased BP in mature spontaneously hypertensive rats [8]. Signaling molecules that regulate eNOS activity include phosphatidylinositol 3-kinase (PI3K)/Akt, cyclic nucleotide-dependent kinases (protein kinase A and protein kinase G), protein kinase C, CaMKII and ribosomal protein S6 kinase (RSK). Experimental studies demonstrate that adenosine may phosphorylate eNOS through activation of MEK/ERK cascades in the NTS [9]. This result is further supported by reports that inhibition of ERK1/2 activation attenuated the eNOS phosphorylation that was induced by estrogen and vascular endothelial growth factor [9]. Although it has been reported for years that ERKs could phosphorylate and activate eNOS, the exact signaling mechanisms that couple the activated ERK1/2 to eNOS activation remained uncertain. Evidence indicates that there is a novel adenosine-ERKeNOS signaling pathway related to the production of NO in the regulation of BP and heart rate in the NTS [9].

CARDIOVASCULAR EFFECTS OF INSULIN IN THE NTS Insulin receptors are unevenly distributed throughout the brain, with a particularly high density in the choroid plexus, cerebral cortex, olfactory bulb, hippocampus, cerebellum, and hypothalamus, which indicates the significant role of insulin in the CNS. Recent evidence has demonstrated that the peripheral and central influence of insulin on cardiovascular regulation is due to effects on the sympathetic nervous system (SNS). However, the significance of the insulin-mediated system in the control of blood pressure is not well-understood. It is well-known that the binding of insulin with its receptors results in activation of the insulin receptor (IR) tyrosine kinase. The activated kinase then phosphorylates tyrosine residues of IR substrates (IRSs). IRSs are adaptor proteins that are linked to the activation of two main signaling pathways including PI3K-Akt/protein kinase B (PKB) pathway, which is responsible for most of the metabolic actions of insulin, and the Ras-mitogen-activated protein kinase (MAPK) pathway, which regulates expression of some genes and cooperates with the PI3K pathway to control cell growth and differentiation [10]. In addition, several studies have shown that enhancing insulin sensitivity with insulin sensitizers may improve insulin resistance and limit the development of adverse cardiovascular consequences. These observations suggest that signaling defects at the IR or post-receptor levels can lead to insulin resistance and may be associated with cardiovascular diseases, including hypertension We reported a novel insulin-PI3K-Akt-NOS signaling pathway related to the production of NO in the regulation of BP and HR in the NTS of normotensive rats [11]. Conversely, there is an additional signal transduction pathway involved in insulin-mediated cardiovascular effects in the NTS of insulin resistance rats that is NOSdependent but independent of the protein kinase Akt [12].

CARDIOVASCULAR EFFECTS OF ANGIOTENSIN II IN THE NTS Ang II is a powerful vasoconstrictor. Hyperactivity of Ang II has been shown to play a major role in hypertension. However, the underlying molecular mechanisms are still not fully understood. These pathological and physiological actions of Ang II are mediated through its type 1 receptor (AT1R). It is wellknown that the expression of the AT1R is 2 to 4-fold higher in the neurons from spontaneously hypertensive rat (SHR) compared with those from WKY rat. In normotensive rats, the Ang II-AT1R-PKC-eNOS signaling pathway relates to production of NO in the regulation of BP and HR. Several lines of evidence demonstrate that reactive oxygen species (ROS) play a role in central autonomic networks that are involved in Ang II-mediated signaling [13]. For instance,

II. BIOCHEMICAL AND PHARMACOLOGICAL MECHANISMS

CoNCluSIoN

143

FIGURE 29.1 The proposed adenosine, insulin and Ang II signaling pathway in the regulation of blood pressure in the NTS.

inhibition of Ang II activity by losartan (AT1R antagonist) significantly increased the expression of extracellular signal-regulated kinase (ERK)1/2, RSK, and also increased nNOS phosphorylation, which is involved in Ang II- ROSmediated modulation of BP in the NTS of SHR [14]. Thus the ERK1/2-RSK-nNOS signaling pathway may play a significant role in Ang II-mediated central BP regulation [14].

CONCLUSION The sympathetic nervous system has moved toward center stage in cardiovascular medicine. Recently studies have shown that sympathetic hypereractivity participates in the development, maintenance and progression of elevated blood pressure. The CNS–mean arterial pressure (CNS-MAP) set-point theory has recently been proposed. It has been hypothesized that hypertension occurs as the result of a primary shift of the CNS-MAP set point to a higher operating pressure, which results in increased sympathetic nerve activity. The NTS, located at the dorsal part of the brainstem, is recognized to be an important integral center in the CNS. Ubiquitously distributed neuromodulators such as adenosine, insulin and Ang II, were found to participate in sympathetic activity regulation in the NTS. Several

studies have investigated the signaling mechanism of neuromodulator with regard to cardiovascular modulation in the NTS and found that the adenosine-ERK-eNOS, insulin-PI3K-Akt-NOS and Ang II-ERK1/2-RSK-nNOS signaling pathways participate in central cardiovascular control (Fig. 29.1). Further investigation of the molecular mechanisms involved in sympathetic nervous activity modulation might elucidate the pathogenesis of the CNS-MAP set-point shift and sympathetic overactivity in essential hypertension. These observations should be of help in further understanding of the pathogenesis and optimal treatment of blood pressure disorders.

References [1] Mosqueda-Garcia R. Central autonomic regulation. In: Robertson D, Low PA, Polinsky RJ, editors. Primer on the autonomic neuvous system. U.S.A: Academic Press; 1996. p. 3–12. [2] Guyenet PG. The sympathetic control of blood pressure. Nat Rev Neurosci 2006;7:335–46. [3] Doba N, Reis DJ. Role of central and peripheral adrenergic mechanisms in neurogenic hypertension produced by brainstem lesions in rat. Circ Res 1974;34:293–301. [4] Ideguchi M, Kajiwara K, Yoshikawa K, Kato S, Ishihara H, Fujii M, et al. Continuous hypertension and tachycardia after resection of a hemangioblastoma behind the dorsal medulla oblongata: relationship to sympathetic overactivity at the neurogenic vasomotor center. J Neurosurg 2010;113:369–73.

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[5] Tseng CJ, Liu HY, Lin HC, Ger LP, Tung CS, Yen MH. Cardiovascular effects of nitric oxide in the brain stem nuclei of rats. Hypertension 1996;27:36–42. [6] Lo WC, Jan CR, Wu SN, Tseng CJ. Cardiovascular effects of nitric oxide and adenosine in the nucleus tractus solitarii of rats. Hypertension 1998;32:1034–8. [7] Lin HC, Wan FJ, Kang BH, Wu CC, Tseng CJ. Systemic administration of lipopolysaccharide induces release of nitric oxide and glutamate and c-fos expression in the nucleus tractus solitarii of rats. Hypertension 1999;33:1218–24. [8] Waki H, Murphy D, Yao ST, Kasparov S, Paton JF. Endothelial NO synthase activity in nucleus tractus solitarii contributes to hypertension in spontaneously hypertensive rats. Hypertension 2006;48:644–50. [9] Ho WY, Lu PJ, Hsiao M, Hwang HR, Tseng YC, Yen MH, et al. Adenosine modulates cardiovascular functions through activation of extracellular signal-regulated kinases 1 and 2 and endothelial nitric oxide synthase in the nucleus tractus solitarii of rats. Circulation 2008;117:773–80.

[10] Taniguchi CM, Emanuelli B, Kahn CR. Critical nodes in signalling pathways: insights into insulin action. Nat Rev Mol Cell Biol 2006;7:85–96. [11] Huang HN, Lu PJ, Lo WC, Lin CH, Hsiao M, Tseng CJ. In situ Akt phosphorylation in the nucleus tractus solitarii is involved in central control of blood pressure and heart rate. Circulation 2004;110:2476–83. [12] Hsiao M, Lu PJ, Huang HN, Lo WC, Ho WY, Lai TC, et al. Defective phosphatidylinositol 3-kinase signaling in central control of cardiovascular effects in the nucleus tractus solitarii of spontaneously hypertensive rats. Hypertens Res 2008;31:1209–18. [13] Sun C, Zubcevic J, Polson JW, Potts JT, Diez-Freire C, Zhang Q, et al. Shift to an involvement of phosphatidylinositol 3-kinase in angiotensin II actions on nucleus tractus solitarii neurons of the spontaneously hypertensive rat. Circ Res 2009;105:1248–55. [14] Cheng WH, Lu PJ, Ho WY, Tung CS, Cheng PW, Hsiao M, et al. Angiotensin II inhibits neuronal nitric oxide synthase activation through the ERK1/2-RSK signaling pathway to modulate central control of blood pressure. Circ Res 2010;106:788–95.

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C H A P T E R

30 Entrainment of Sympathetic Rhythms Michael P. Gilbey SYMPATHETIC RHYTHM The term “sympathetic rhythm” describes the mo­ ment to moment waxing and waning in amplitude (sig­ nal strength) of population activity recorded from whole sympathetic nerves that contain thousands of fibers. Such rhythms are commonly observed in the discharges of pre­ and postganglionic nerves regulating the heart and blood vessels. Rhythm is frequently only an emergent property of population activity; i.e., the discharges of single neurons sampled from the population may not necessarily dem­ onstrate rhythmicity, however, the tendency for subpopu­ lations of neurons to discharge action potentials almost coincidentally but intermittently gives rise to rhythmic­ ity in aggregate activity. Although Adrian and colleagues described rhythmic activity in the first published record­ ings of mammalian sympathetic nerves in 1932, underly­ ing mechanism(s) and possible functional significance are still uncertain [1–3].

Cardiac- and Respiratory-related Rhythms The most common sympathetic rhythms are cardiac­ (pulse­) and respiratory­related. Concerning respiratory rhythm, two components can be distinguished; one as­ sociated with central respiratory activity and another dependent upon afferent activity related to pulmonary ventilation (e.g., from arterial baroreceptors and/or pul­ monary stretch receptors) [4,5]. Mechanisms Underlying Rhythms Two major hypotheses have been proposed to account for cardiac­ and respiratory­related rhythms in sympa­ thetic discharges. Phasic Inputs Generate Rhythms The classic view holds that these rhythms are imposed upon sympathetic discharge by “external” inputs. In the case of the cardiac­related rhythm, an increase in barore­ ceptor discharge during systole is considered to inhibit tonic excitatory drive to sympathetic nerves and thereby give rise to waxing and waning of sympathetic discharges [3]. A similar mechanism is proposed for rhythms related

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00030-5

to pulmonary ventilation cycles where activation of lung stretch and/or baroreceptor afferents, for example, may cause periodic inhibition of activity (independent of the central respiratory rhythm generating network) [4,4a,5]. Concerning central respiratory­related rhythms it is sug­ gested that elements within the central respiratory net­ work provide excitatory and/or inhibitory inputs to central networks providing tonic drive to sympathetic nerves [4,5]. Additionally, Richter and Spyer have argued for the existence of a common cardiorespiratory network [6], and see Chapter 31. Entrainment of Rhythms The observation of a non­respiratory­ and non­cardiac­ related sympathetic rhythm (“10 Hz” rhythm: Green and Hoffman, 1967) provided the first indication that sympa­ thetic rhythms might not arise exclusively from phasic inputs to tonic sympathetic tone generating networks. Some 8 years later Taylor and Gebber observed that sym­ pathetic rhythms with a frequency similar to heart rate persisted following baroreceptor denervation [1,3]. As a result the idea developed that cardiac­related rhythms in sympathetic nerve discharge might be a consequence of the entrainment of central oscillator(s) within the brain­ stem (i.e., that central networks driving sympathetic out­ flow may be intrinsically capable of generating their own rhythms). In this scheme, phasic baroreceptor input acts as a forcing input that can entrain a central oscillator. Consequently, in the absence of, or with reduced barore­ ceptor activity, there is a continuous phase shift between cardiac cycle and sympathetic rhythm resulting from lack of entrainment [1–3]. Additionally, there is evidence to support the hypothesis that respiratory­related rhythms in sympathetic discharges arise from oscillator(s) other than those within the central respiratory networks [7]. First, in vagotomized anesthetized animals, a “slow” rhythm in the frequency range of the central respiratory rhythm has been observed during central apnea (indicated by absence of rhythmic phrenic discharge). Second, “slow” rhythms in the discharges of pairs of sympathetic nerves at the fre­ quency of central respiratory drive have been observed to remain correlated after mathematical removal of the com­ ponent of these signals common to phrenic nerve activity (an indicator of central respiratory drive); i.e., “theoretical

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removal of central respiratory drive”. Third, locking ratios other than 1:1 can be observed between rhythmic phrenic and sympathetic discharge (e.g. 2:1, 3:1, 2:3). It is also apparent that afferents activated during pulmonary ventilation can act as an entraining force [2]. However, the observations concerning the entrainment of respira­ tory rhythm must be mindful of findings that indicate great complexity in central respiratory rhythm generating networks and their differential influence on the various somatic motor outflows driving respiratory muscles.

How Many Central Oscillators? Gebber and colleagues have provided evidence that separate oscillators, capable of coupling, may drive activ­ ity to different sympathetic nerves: they noted from paired nerve recordings, that rhythmic discharges of similar fre­ quency could be phase­locked, but this was not obliga­ torily; their variation in amplitude was not necessarily proportionate [7]. In addition, the observations of Gilbey and colleagues have raised the intriguing possibility that activity of sympathetic neurons regulating the same tar­ get may be influenced by a family of weakly coupled or uncoupled oscillators and that their degree of synchro­ nization is influenced by inputs related to lung inflation, central respiratory drive, various afferents (e.g., somatic and baroreceptor) and possibly arousal state [2,8].

The Spinal Cord and Sympathetic Rhythms Traditionally, supraspinal sites have been the focus of interest regarding sympathetic rhythm generation [1,5,6]. However, recently it has been demonstrated, both in situ and in isolated slices, that sympathetic rhythms can be generated within the spinal cord [2,9]. Therefore, it appears that sympathetic rhythms generated within the spinal cord may be entrained by inputs arising both from peripheral afferents and supraspinal networks; e.g., respi­ ratory [2]. Intrathecal application of 5­hydroxytryptamine (5­HT) induced a cutaneous vasomotor sympathetic rhythm in a spinalized rat preparation similar to that seen in an intact preparation [2] and spontaneous rhythmic activity was recorded from the intermediolateral cell columns (IML: the location of the cell bodies of sympathetic pregangli­ onic neurons) of thoracic spinal cord slices taken from neonatal rats [9]. In a spinal cord slice preparation, rhyth­ mic activity could be induced or enhanced by 5­HT recep­ tor agonists and reduced by gap junction blockers. These neurophysiological observations are strengthened by the findings that the IML receives a dense 5­HT­containing innervation and membrane localized Cx36 immunoreac­ tivity, indicative of gap junctions, has been observed in sympathetic preganglionic neurons. These findings point to a potentially important func­ tion of spinal cord circuitry both for the generation and

entrainment of sympathetic rhythms. Furthermore, they demonstrate the potential for the neurochemical modula­ tion of sympathetic rhythmogenesis both at the single cell and neural network level. Consequently, aberrant sympa­ thetic rhythm generation and/or entrainment induced by changes in CNS function may result in peripheral disease (e.g., cardiovascular: see Chapter 31).

Functional Significance Whatever mechanism(s) lie behind the generation of sympathetic rhythms, their characteristic phasic nature in­ dicates co­ordination of neuronal discharges. Whereas it is clear why co­ordinated phasic discharges are required in locomotor and respiratory motor control, the need for pat­ terning and synchronization in sympathetic motor control of heart and blood vessels is not readily apparent. With regard to neuroeffector transmission, many rhythms will be filtered­out (i.e., a sympathetic rhythm above 1 Hz will not lead to a 1 Hz vasomotion) as the time constant for response is ~2 seconds [1–3]. It has been suggested that co­ordination may be par­ ticularly easy to achieve between oscillating neural networks. Gebber and colleagues have suggested, based upon experimental observation, that coupled sympa­ thetic oscillators may be important in the generation of differential patterns of sympathetic activity to blood ves­ sels of muscle, skin and viscera associated with behav­ ioural alerting [1]. Furthermore, the observations on entrainment of sympathetic rhythms indicate that when appropriate sympathetic and respiratory networks may “bind” together to form a highly co­ordinated “super­net­ work” [2]. There are also many indications that pattern and syn­ chrony coding are used in addition to rate coding in vari­ ous nervous system functions [2]. At the level of the single neuron, pattern of firing appears important in determin­ ing probability of transmitter release, synaptic plasticity, types of transmitter released/co­released and receptors activated. In these ways, firing pattern probably influ­ ences ganglionic and neuroeffector transmission [2,3,10]. Synchrony may be important as it favours summation, which raises the efficacy of transmission and can also have longer­term influences on synaptic and neuroeffec­ tor function. Synchrony therefore may enhance ganglionic transmission by increasing the probability of summa­ tion of weak inputs and improve neuroeffector control by coordinate activation of post­junctional receptors [2,3]. Consequently, if as suggested by Gilbey and co­workers [2] a population of neurons regulating a single target is influenced by a family of oscillators their dynamic and graded synchronization through variable entrainment could lead to the modulation of target organ function. In this manner aberrant entrainment of sympathetic rhythms might lead to peripheral pathology consequent to inappro­ priate sympathetic activity.

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SYMPATHETIC RHYTHM

References [1] Barman SM, Gebber GL. “Rapid” rhythmic discharges of sympa­ thetic nerves: Sources, mechanisms of generation, and physiological relevance. J Biol Rhythm 2000;15:365–79. [2] Gilbey MP. Sympathetic rhythms and nervous integration. Clin Exp Pharmacol Physiol 2007;34:356–61. [3] Malpas SC. The rhythmicity of sympathetic nerve activity. Prog Neurobiol 1998;56:65–96. [4] Habler HJ, Janig W, Michaelis M. Respiratory modulation in the activity of sympathetic neurons. Prog Neurobiol 1994;43:567–606. [4a] Huang C, Marina N, Gilbey MP. Impact of lung inflation cycle fre­ quency on rat muscle and skin sympathetic activity recorded using suction electrodes. Auton Neurosci 2009;150:70–5. [5] Koepchen H­P, Klussendorf D, Sommer D. Neurophysiological background of central neural cardiovascular­respiratory coordina­ tion: Basic remarks and experimental approach. J Auton Nerv Syst 1981;3:335–68.

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[6] Richter DW, Spyer KM, Gilbey MP, Lawson EE, Bainton CR, Wilhelm Z. On the existence of a common cardiorespiratory network. In: Koepchen H­P, Huopaniemi T, editors. In cardiorespiratory and motor coordination. Berlin: Springer Verlag; 1991. p. 118–30. [7] Zhong S, Zhou SY, Gebber GL, Barman SM. Coupled oscillators account for the slow rhythms in sympathetic nerve discharge and phrenic nerve activity. Am J Physiol 1997;272:R1314–R1324. [8] Staras K, Change HS, Gilbey MP. Resetting of sympathetic rhythm by somatic afferents causes post­reflex coordination of sympathetic activity in rat. J Physiol London 2001;533:537–45. [9] Pierce ML, Deuchars J, Deuchars SA. Spontaneous rhythmogenic capabilities of sympathetic neuronal assemblies in the rat spinal cord slice. Neuroscience 2010;170:827–38. [10] Karila P, Horn JP. Secondary nicotinic synapses on sympathetic B neurons and their putative role in ganglionic amplification of activ­ ity. J Neurosci 2000;20:908–18.

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C H A P T E R

31 Cross-talk Between Body Systems: Respiratory-Cardiovascular Coupling in Health and Disease Julian F.R. Paton, Anthony E. Pickering

INTRODUCTION Although the common perception of the pulse is one of a stable and constant beat, periodic heart rate variability has been observed in fish, amphibians, reptiles and mammals suggesting that its appearance during evolution was not coincidental and that it confers a survival advantage. Much of this periodic heart rate fluctuation is linked to the phases of respiration. Classically, in man, heart rate increases on inhalation and falls during exhalation (so called respiratory sinus arrhythmia; RSA) and is most prominent in the young and well trained athletes. Why RSA exists is not wholly clear, yet we know that its loss is a powerful predictor of morbidity and mortality from cardiovascular disease [1]. The latter indicates a major protective role for RSA in cardiovascular health, which is a good reason to discuss its potential function. Blood pressure also expresses a respiratory oscillation, often called the high frequency oscillation or HFO. The functional significance of the HFO is if anything even less clear than that of RSA but both oscillations demonstrate the strong cross-talk between the circulatory and respiratory systems. This chapter focuses on the central neural inter-connectivity between the brainstem respiratory pattern generator and neural networks governing sympathetic and parasympathetic activity. The putative functional roles of RSA and HFO in health are discussed; additionally their plasticity will be illustrated with examples of alterations in coupling strength and patterns between respiration and cardiovascular autonomic motor outputs. The importance of these changes is discussed in the context of disease states and how this may provide novel insights into our mechanistic understanding of cardiovascular pathologies.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00031-7

CARDIOVASCULAR AUTONOMIC ACTIVITY: COUPLING TO THE RESPIRATORY PATTERN GENERATOR Anrep et al. [2] described RSA for the first time and provided mechanistic insight into its causes. These included a baroreceptor reflex-mediated modulation of heart rate, triggered by respiratory induced changes in venous return (secondary to oscillations in intrathoracic pressure) and abdominal thoracic pumping, and consequent blood pressure waves in time with the respiratory cycle. When this peripheral change in venous return was negated, RSA still persisted leaving the heart rate modulation as being due either to pulmonary stretch receptor feedback following lung inflation and/or direct coupling to central respiratory drive. Both mechanisms were found to play a role. Since those seminal studies, it has been found that cardiac sympathetic motor activity exhibited prominent bursting during late-inspiration [3] whereas cardiac vagal activity fired preferentially during early expiration (postinspiration) [4]. Recordings from in situ perfused preparations (without either pulmonary stretch receptor activation or respiratory related changes in venous return) show prominent respiratory sinus arrhythmia, confirming the important role of central coupling between the respiratory and cardiac parasympathetic preganglionic neurons (Fig. 31.1) [5]. The burst of cardiac vagal activity seems to originate centrally at the level of the preganglionic neurons in the nucleus ambiguus that are inhibited during inspiration but excited during post-inspiration [4]. The magnitude of RSA is principally a function of this cardiac vagal tone or the excitability state of the preganglionic cardiac vagal motoneurons [4]. Differences in the cardiac vagal tone explain the variable degree of RSA between individuals.

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In terms of vasomotor sympathetic neurons, recordings at either the pre- or post-ganglionic levels show respiratory related busting during late-inspiration and early expiration contributing to the HFO [6–8]. The respiratory-related alterations in venous return/cardiac output described above also contribute to the respiratory modulation of arterial pressure. However, the fluctuations persist in the working heart brainstem preparation indicating again there is an important central neural component to the coupling [8]. These data provided unequivocal evidence that manifestation of both RSA and HFO was dependent, in part, on a central respiratory modulation of both limbs of the autonomic nervous system. An argument supporting the concept that the cardiovascular and respiratory systems are actually a single system (even within the brain) comes

Respiration Phrenic nerve

BRAINSTEM

inspiration - 120

Arterial blood Pressure mmHg

- 80 - 70

Heart rate beats/min

3s

- 60

FIGURE 31.1 Respiratory sinus arrhythmia is in part due to central nervous coupling between the brainstem respiratory rhythm generator and autonomic motor outflows. Schematic depicting respiratory sinus arrhythmia expressed here as a rise in heart rate during central inspiratory drive (i.e. phrenic nerve discharge) and a bradycardia coincident with the start of expiration. These data were obtained in the absence of lung inflation; the latter removes any influence from pulmonary stretch receptor feedback and baroreceptor reflex activation secondary to respiratory induced changes in venous return. This indicates that cross-talk between the central respiratory pattern generator with sympathetic and cardiac vagal neural circuitry within the brainstem contributes significantly to the arrhythmia.

WKY

WHY IS THE CIRCULATORY SYSTEM RESPIRATORY-MODULATED? The exact function/s of RSA and HFO are debated with no unified consensus and require further investigation.

RSA Four suggestions are: 1. The most likely role is to provide close matching of ventilation with pulmonary perfusion. It has been experimentally demonstrated that oxygenation of blood is optimized if increases in cardiac output are timed to occur when fresh air arrives within the alveoli [9], which is the case with RSA as heart rate rises during inspiration. However, no respiratory related alterations in arterial tension of oxygen are detectable but there could be in the pulmonary veins. In contrast, “reverse” RSA increased ventilatory dead space and intrapulmonary shunting [9]. 2. Based on the fact that a red blood cell spends the same amount of time in an alveolar capillary per cardiac cycle (in human), it has been proposed that RSA enhances cardio-respiratory efficiency and prevents “wasted” heart beats by decreasing heart rate/cardiac output on exhalation [9]. 3. The bradycardia of RSA assists in stabilizing blood pressure and counter acts the respiratory related increase in cardiac output that results from increased venous return. 4. It is plausible that slowing of heart rate during expiration enhances coronary blood flow by prolonging diastole. A constant frequency of cardiac contraction (B)

SHR

PNA

PNA

HF(SBP) (mmHg/Hz)

1/2

(A)

from their tight central nervous coupling reflecting the commonality of function.

7

**

6

5

4

Wistar

SHR

FIGURE 31.2 Enhanced respiratory-sympathetic (SNA) coupling in the spontaneously hypertensive rat (SHR). (A) comparison of phrenic nerve activity (PNA) triggered sympathetic activity between normotensive (Wistar Kyoto, WKY) and SHR from the in situ arterially perfused rat preparation. In the SHR there was a greater strength in coupling as central inspiratory drive was comparable between rat strains. Note, the increased coupling produced a larger wave in arterial perfusion pressure (PP). (B) in conscious rats blood pressure was measured using radio-telemetry and showed a high frequency (HF) component in the systolic blood pressure (SBP) that was greater in the SHR than the normotensive control rat. Data from ref 8.

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CARDIovASCulAR DISEASE: oPPoSITE EffECTS on RSA AnD Hfo

(i.e. loss of RSA) could limit coronary blood flow at any given blood pressure as may occur in heart failure and hypertension. The loss of this mechanism may contribute to the increased susceptibility of patients with these diseases to left ventricular tachycardia and sudden cardiac death. Of these four possibilities direct evidence only exists to support the first hypothesis.

CARDIOVASCULAR DISEASE: OPPOSITE EFFECTS ON RSA AND HFO

HFO There is no definitive explanation for the functional role of HFO. This may be related to the experimental difficulty in removing and reconstituting the sympathetic vasomotor drive as has been elegantly done for vagal tone and RSA [9]. Rhythmic contraction and relaxation of vascular smooth muscle of arterioles (i.e., vasomotion) is a physiological process thought to aid blood flow through tissues by reducing resistance [10]. Such a process may become more important when metabolic demand increases such as during exercise. It is likely that the increased central respiratory drive that accompanies exercise may augment HFO to optimise vascular conductance in skeletal muscle to both increase oxygenation and remove end products. There is no reason why such a mechanism cannot also apply to the lungs to optimize ventilation perfusion matching, although this has not been directly shown to our knowledge. HFO may also improve capillary exchange to ensure both low interstitial pressure and constant para-capillary fluid flux [10]. In diabetic patients with neuropathy the amplitude of flow-motion was reduced compounding issues with perfusion. However, in diabetics without neuropathy no such reduction in

(A)

vasomotion was seen indicating a high dependence on sympathetic innervation [10]. In hypertensive animals and humans, vasomotion is increased and this positively correlates with increased blood pressure [10]. Whether hypertension triggers more vasomotion or whether excessive vasomotion drives the hypertension is unclear but discussed below (see HFO, below.)

RSA Loss of vagal tone in cardiovascular diseases can be clearly demonstrated by the diminished change in heart rate on administration a vagolytic drug like atropine and also by the loss of RSA [11]. The cause of this absence of cardiac vagal tone and RSA may have a common origin – depression of the cardiac vagal limb of the baroreceptor reflex. The latter normally provides a major excitatory synaptic drive to cardiac vagal motoneurons [4]. At which point this depression occurs (afferent, brainstem, efferent) is not known. Depressed synaptic transmission at the level of the cardiac vagal ganglion in heart failure dogs was reported [12] and this could be restored by the administration of acetycholinesterase inhibitors. Whether this involves a pre- and/or postsynaptic effect and the precise mechanism(s) involved remain unknown. Interestingly it has been shown that reductions in nitric oxide within the cardiac ganglia dis-faciliate vagal transmission, which can be restored by over expression of neuronal nitric oxide synthase [13]. However, transmission failure at the neuroeffector junction cannot be ruled out. A central component

central apnea

(B) ∗

∆ PP with return of PNA (mmHg)

20 15 10 5 0

1 min

WKY

SHR

Tone developed by respiratorysympathetic coupling

FIGURE 31.3 In hypertension the increased vascular resistance is driven, in part, by the enhanced respiratory-sympathetic coupling. (A) Reinstating breathing after suppressing central respiratory drive (with hypocapnia) allows an assessment of the contribution of respiratorysympathetic coupling to vasomotor tone. As phrenic nerve activity (PNA) returns this couples to sympathetic nerve activity (SNA) driving up arterial perfusion pressure (PP). Notice the immediate saw-toothed increase in PP when PNA returns with summation of the PP oscillations driving pressure up. (B) Graph indicates that the respiratory-sympathetic coupling contributed more to PP (and vascular resistance) in the SHR than the normotensive Wistar Kyoto rat (WKY). All data from the arterially perfused in situ preparation. From ref 8.

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to this vagal dysfunction is suggested from a model of intermittent hypoxia induced cardiovascular pathology exhibiting decreased excitability of cardiac vagal preganglionic neurons [14].

the stimulus coincides with the onset of expiration [20]. This may aid in reducing arterial pressure transiently by limiting venous return.

NEW HORIZONS

HFO In contrast to a decline in RSA, HFO are augmented in hypertension, at least in the spontaneously hypertensive (SH) rat (Fig. 31.3) [8]. The amplitude of respiratory-related sympathetic bursting was augmented in SH compared to normotensive rats and showed a phase shift peaking in late inspiration rather than early expiration [8]; the significance of the latter is unknown but agrees with earlier observations in the SHR [15]. The increased coupling of respiratory-sympathetic modulation was without significant change in central inspiratory drive [8]. This supporting the notion that the coupling strength was augmented. The resultant increase in coupling strength produced greater HFO in arterial pressure in the SH rat. These waves summated raising vascular resistance and arterial pressure was greater in the SH than normotensive rat (i.e. 15 vs. 5 mmHg) (Fig. 31.2) [8]. Alteration in coupling strength and pattern of coupling have also been seen in the angiotensin II/high salt induced hypertensive rat [16], the Goldblatt hypertensive rat (Oliveira-Sales EB, Campos RR and Paton, JFR – unpublished) and rats treated with chronic intermittent hypoxia (CIH) [17]. In the latter case, CIH produced hypertension associated with increased respiratory modulation of sympathetic activity produced by the unique appearance of an extra respiratory-related burst in sympathetic activity occurring during late expiration. This was subsequently found to correlate with abdominal motor discharge, which was not present in control animals, suggesting that CIH treated rats expire actively (i.e. forced expiration) and that this rhythm drives the additional sympathetic burst contributing to the hypertension [17]. The importance of increased blood pressure variability in mediating the end organ damage is increasingly appreciated in animal models such as the SHR [18] and also in man [19]. This respiratory-sympathetic coupling therefore represents an interesting alternative potential target for treatment.

CARDIOVASCULAR MODULATION OF RESPIRATORY ACTIVITY This review would be incomplete without mentioning the emerging data supporting the reciprocity of cardiovascular-respiratory coupling. Brainstem neurons receptive to pulmonary stretch receptor inputs (essential for inspiratory phase termination) receive convergent inputs from baroreceptors. This now appears important for mediating baroreceptor reflex depression of breathing [20]. Prolongation of the expiratory pause can be evoked by baroreceptor stimulation but this is most effective when

There are some key studies that need to be performed. Given its protective role, the site(s) and mechanisms of blockade of cardiac vagal transmission must be identified in cardiovascular disease to allow novel, targeted therapy to be devised. Regarding HFO we need a better comprehension of its functional role from basic experimental investigations and the factors that govern its plasticity and amplification. Similarly we need to know whether such amplified coupling exist in human patients with heart failure and/or hypertension. Our pilot data support that this may be the case (Fisher J, Pickering AE & Paton JFR – unpublished). If so, an understanding of the changes in breathing rate and/ or pattern may provide new insight into methods to control excessive sympathetic activity in cardiovascular diseases. Changes in the sensitivity of central and peripheral chemoreception are potentially most relevant. Breathing rate and pattern may be a sensitive prognostic indicator for cardiovascular disease allowing earlier intervention and prevention treatment. All said, cardiovascular-respiratory coupling is in take-off mode for translation to the clinic.

References [1] La Rovere MT, et al. Baroreflex sensitivity and heart-rate variability in prediction of total cardiac mortality after myocardial infarction. ATRAMI (Autonomic Tone and Reflexes After Myocardial Infarction) Investigators. Lancet 1998;351:478–84. [2] Anrep GV, Pascual W, Rössler R. Respiratory variations of the heart rate. II. The central mechanism of the sinus arrhythmia and the inter-relationship between central and reflex mechanism. Proc Roy Soc Lond 1936;119(Series B):218–30. [3] Paton JF, Boscan P, Pickering AE, Nalivaiko E. The yin and yang of cardiac autonomic control: vago-sympathetic interactions revisited. Brain Res Rev 2005;49:555–65. [4] McAllen RM, Spyer KM. The baroreceptor input to cardiac vagal motoneurons. J Physiol 1978;282:365–74. [5] Paton JFR. A working heart-brainstem preparation. J Neurosci Meth 1996;65:63–8. [6] Adrian ED, Bronk DW, Phillips G. Discharges in mammalian sympathetic nerves. J Physiol 1932;74:115–33. [7] Habler HJ, Janig W, Michaelis M. Respiratory modulation in the activity of sympathetic neurons. Prog Neurobiol 1994;43:567–606. [8] Simms AE, Paton JF, Pickering AE, Allen AM. Amplified respiratory– sympathetic coupling in the spontaneously hypertensive rat: does it contribute to hypertension? J Physiol 2009;587:597–610. [9] Hayano J, Yasuma F, Okada A, Mukai S, Fujinami T. Respiratory sinus arrhythmia. A phenomenon improving pulmonary gas exchange and circulatory efficiency. Circ 1996;94:842–7. [10] Nilsson H, Aalkjaer C. Vasomotion: mechanisms and physiological importance. Mol. Interv. 2003;3:79–89. 51 [11] Polson JW, McCallion N, Waki H, Thorne G, Tooley MA, Paton JFR, et al. Evidence for cardiovascular autonomic dysfunction in neonates with coarctation of the aorta. Circ 2006;113:2844–50. [12] Bibevski S, Dunlap ME. Ganglionic mechanisms contribute to diminished vagal control in heart failure. Circ 1999;99:2958–63.

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[13] Wang L, Li D, Dawson TA, Paterson DJ. Long-term effect of neuronal nitric oxide synthase over-expression on cardiac neurotransmission mediated by a lentiviral vector. J Physiol 2009;587:3629–37. [14] Yan B, Soukhova-O'Hare GK, Li L, Lin Y, Gozal D, Wead WB, et al. Attenuation of heart rate control and neural degeneration in nucleus ambiguus following chronic intermittent hypoxia in young adult Fischer 344 rats. Neuroscience 2008;153:709–20. [15] Czyzyk-Krzeska MF, Trzebski A. Respiratory-related discharge pattern of sympathetic nerve activity in the spontaneously hypertensive rat. J Physiol 1990;426:355–68. [16] Toney GM, Pedrino GR, Fink GD, Osborn JW. Does enhanced respiratory-sympathetic coupling contribute to peripheral neural mechanisms of angiotensin II-salt hypertension? Exp Physiol 2010;95:587–94.

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[17] Zoccal DB, Simms AE, Bonagamba LH, Braga VA, Pickering AE, Machado BH, et al. Increased sympathetic outflow in juvenile rats submitted to chronic intermittent hypoxia correlates with enhanced expiratory activity. J Physiol 2008;586:3253–65. [18] Miao CY, Xie HH, Zhan LS, Su DF. Blood pressure variability is more important than blood pressure level in determination of endorgan damage in rats. J Hypertens 2006;24:1125–35. [19] Tatasciore A, Renda G, Zimarino M, Soccio M, Bilo G, Parati G, et al. Awake systolic blood pressure variability correlates with target-organ damage in hypertensive subjects. Hypertension 2007;50:325–32. [20] Baekey DM, Molkov YI, Paton JFR, Rybak IA, Dick TE. Effect of baroreceptor stimulation on the respiratory pattern: Insights into respiratory-sympathetic interactions. Resp Physiol Neurobiol 2010;174:135–45.

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32 Circadian Rhythms and Autonomic Function Diego A. Golombek Nested deep within the brain, a master clock guides most of our temporal regulation of physiology and behavior. Indeed, mammalian diurnal rhythms are generated by the hypothalamic suprachiasmatic nuclei (SCN) and finely tuned to environmental periodicities, the most important being the light–dark cycle. Light reaches the clock through a direct retinohypothalamic tract (RHT), primarily through glutamatergic innervation, and its action is probably regulated by a variety of other neurotransmitters. The SCN continue to work precisely under constant conditions, originating circadian (i.e., with a period of about 24 h) rhythms in free-running conditions. This simple scheme can be better described by considering diverse entrainment agents (zeitgebers) which include food availability, arousal and temperature. In addition. a variety of so-called peripheral oscillators are able to generate autonomous circadian cycles in vitro (e.g., the liver, fibroblasts, diverse brain regions, lung, retina, etc.), although their influence is probably restricted to their local environment. Photic entrainment is achieved primarily by stimulation of retinal photoreceptors that include the classical rod and cone signal transduction, but also through melanopsin, a photopigment present in retinal ganglion cells involved in non-visual responses. Within the SCN, rhythms are generated in circadian pacemaker cells by a complex of molecular feedback loops that positively and negatively regulate the transcription of core genes (e.g. period, cryptochrome, bmal1) of the circadian clock. The transcription-translation loop that generates molecular oscillations of clock genes is remarkably conserved among species and even distant groups, suggesting a possibly monophyletic origin of such mechanism. Forward genetic approaches have unraveled several genetic components isolated from distinct circadian phenotypes, advancing our knowledge of the molecular circadian clock. In the mid-1990s the first mammalian clock gene (CLOCK) was discovered and led to the description of the loop in which CLOCK and BMAL heretodimerize and promote the transcription of PER (period) and CRY (cryptochrome) genes, whose proteic products in turn heterodimerize, translocate to the nucleus and negatively regulate the activity of CLOCK/BMAL. Deletions or

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mutations in these genes or in their post-transcriptional regulation (such as phosphorylation) result in arrhythmicity or in profound changes in the circadian structure. In addition, genome-wide identification of clock-controlled genes, as well as their protein products and their interactions, provides important information for the understanding of the systemic behavior of circadian cogs and wheels within and between cells. In addition, post-translational modifications of clock proteins are deeply related to circadian regulation, as well as the recent description of epigenetic mechanisms that are controlled by the clock and also regulate the transcription of clock genes.

SCN OUTPUT AND AUTONOMIC CONTROL The output of the SCN includes both humoral (i.e., by vasopressin release) and neural mechanisms, which are directly related to the autonomic nervous system and, in turn, to the circadian regulation of body functions. The best known example of SCN output is probably the regulation of melatonin synthesis by the pineal gland. Pineal melatonin content exhibits a clear diurnal and circadian rhythm which is regulated by light in a daily and seasonal fashion. The SCN regulates pineal function by innervation of paraventricular (PVN) neurons which project to pre-ganglionic neurons in the intermediolateral nucleus of the thoracic spinal cord, which in turn project to the noradrenergic neurons of the superior cervical ganglion (SCG) (Fig. 32.1). The noradreneregic unnervation of the pineal is crucial in the activation and light-response of the enzymes responsible for melatonin synthesis and release. Melatonin (and, as stated below, corticosterone) might be considered the humoral hand of the circadian clock that might help in setting the pace and phase of diverse physiological variables. This SCN control of pineal melatonin can be extrapolated to other autonomic controls of hormonal secretion. For example, tracing experiments revealed a SCNPVN-sympathetic nervous system-adrenal gland axis that controls corticosterone release, by means of a diurnal

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FIGURE 32.1 The circadian clock located in the hypothalamic suprachiasmatic nuclei (SCN) sends temporal information to the body through humoral and neural pathways interacting with the autonomic nervous system. The SCN is entrained by photic stimulation through the retinohypothalamic tract (RHT) which receives information from retinal photoreceptors (PRs) and intrinsically photoreceptive ganglion cells (iPRGCs). Besides targeting neuroendocrine neurons directly (depicted here as “endocrine cells”, such as GnRH- or CRH-releasing cells), the SCN synapses with diverse hypothalamic nuclei such as the medial preoptic nucleus or the dorsomedial nucleus, as well as thalamic projections (all of which are here grouped as “other nuclei”). However, the best known efferent target of the SCN are the paraventricular nuclei (PVN), which integrate sympathetic and parasympathetic signals from the circadian system and connect with relay stations in the brainstem and the spinal cord. Parasympathetic projections to the dorsal motor nucleus of the vagus (DMV) and sympathetic projections to preganglionic neurons of the intermediolateral column of the spinal cord (IML). Autonomic circadian control of peripheral organs follows these pathways. As an example, noradrenergic innervation from the IML, through the sympathetic superior cervical ganglion, to the pineal gland (P) is shown.

change in ACTH responsiveness. In a similar fashion, the SCN regulates glucose metabolism by a circadian modulation of insulin sensitivity and hepatic glucose production. In addition, the SCN relays neural information to the pancreas, the liver and the adipose tissue, the heart and the kidney (among others) by sympathetic/parasympathetic pathways. Metabolism is linked to the circadian system not only through autonomic innervation but also because the core clock genes, expressed in several tissues throughout the body, are directly related to cellular metabolic pathways. Besides the humoral and neural regulation of autonomic pathways, the SCN can be modulated by feedback from the periphery, by means of hormones (e.g., melatonin), immune factors or information conveyed by circumventricular organs such as the arquate nuclei. In addition, the master clock in the SCN might use autonomic signaling in order to keep peripheral clock oscillations synchronized, as suggested by experiments in which manipulations of autonomic connections to peripheral organs such as the liver reset clock gene expression. Disruption of the normal synchrony between central and peripheral circadian oscillators may result in severe pathological outcomes, as suggested by recent reports that indicate that abnormalities in circadian sympathetic homeostasis results in tumor developments in mice.

CIRCADIAN AND SLEEP-CONTROL OF THE AUTONOMIC NERVOUS SYSTEM The light–dark cycle influences autonomic activity, and it has been demonstrated that this regulation depends on the SCN, since lesions of the nuclei eliminate photic modulation. Clever tracing experiments performed by the group of Kalsbeek and Buijs in the Netherlands demonstrated that control of parasympathetic and sympathetic pre-autonomic neurons is completely separated and independent along the circadian pathway from the hypothalamus to the brainstem. In this way, the SCN and the circadian system are able to modulate both subdivisions of the autonomic nervous system independently, thus resulting in the most adequate responses depending on the time of day and other environmental and internal temporal cues. An imbalanced temporal regulation of sympathetic and parasympathetic activity might lead to profound changes in metabolic physiology and, eventually, to metabolic syndrome-like disease. Indeed, in experimental forced desynchronization clear endogenous circadian regulation of autonomic activity has been demonstrated (assessed by, for example, R-R interval variation in the electrocardiogram), by independent control of sympathetic and vagal innervation of the heart. However, besides it endogenous circadian control, autonomic activity also exhibits profound changes throughout

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the sleep–wake cycle. For example, sympathetic activation is reduced significantly in the transition from wakefulness to slow-wave sleep stage 4 (accompanied by a decrease in arterial blood pressure and bradycardia) determining the parasympathetic predominance typical of this stage – although it goes back to awake levels during REM sleep. Indeed, sleep styles affect autonomic regulation of several variables, including blood pressure and cardiac function: among other changes, short sleep duration correlates with the development of hypertension (as well as diabetes). It is also well-known that cardiovascular incidents do not occur at random throughout the day, but tend to be more common in the morning, suggesting that at the rest-activity transition the SCN is crucial in regulating the sympathetic/parasympathetic balance to the heart, and any changes in clock function put the system at risk. Treatments that increase the amplitude of circadian rhythms (such as exercise, diet, chronobiotics, etc.) might be useful to prevent such clock-dependent risks.

CARDIAC EVENTS AND CIRCADIAN RHYTHMS As an example of circadian autonomic regulation, we shall mention that most (if not all) cardiovascular variables exhibit strong diurnal and/or circadian rhythms, including blood pressure, heart rate, heart rate variability and many others. As a consequence, cardiovascular events also tend to occur at specific times of day, with higher frequencies in the morning and lower risks during sleep which relate to rhythmic changes in human susceptibility to such events. Besides the general pattern of higher blood pressure and heart values during the day, there are marked interindividual differences in temporal allocation of physiological variables (i.e., chronotypes), with extreme morning (“larks”) and evening types (“owls”) exhibiting a difference of several hours in the maxima of temperature, blood pressure and heart rate. Forced desynchronization protocols have demonstrated a mixed contribution from both endogenous circadian factors and sleep in the regulation of diurnal autonomic variations such as changes in heart rate and heart rate variability. Although less studied, seasonal variations in cardiac function have also been reported, including pathological consequences. As for pathological situations, it has long been known that there is a significant peak of acute myocardial infarction in the morning hours (and sometimes a secondary peak in the afternoon), as well as for ischemic and hemorrhagic stroke. Moreover, an alteration of circadian cardiac regulation in coronary disease has been demonstrated.

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Circadian desynchronization of autonomic (sympathetic/ parasympathetic) regulation of the heart correlates closely with heart disease. Finally, extreme environments (e.g., space missions, Antarctic expeditions) can also induce an autonomic/circadian disruption with accompanying neurovegetative changes. In summary, the mammalian circadian system is deeply interrelated with the autonomic nervous system. The two main output channels for the circadian clock are through humoral release or sympathetic/parasympathetic relay stations that convey temporal information throughout the body. The SCN sends neural projections to other hypothalamic nuclei and from there to the brainstem or spinal cord autonomic centers that control physiological activity. In addition, neurovegetative mechanisms exhibit a double modulation by the circadian system (and, in turn, are synchronized to environmental synchronizers) and by arousal/sleep-related mechanisms in a homeostaticlike process. In this sense, autonomic regulation should not only be regarded as a spatial/functional control of physiology and behavior but also in terms of a strong temporal modulation that helps the body to adequately devote resources and energy throughout the day.

Further Reading Buijs RM, Kalsbeek A. Hypothalamic integration of central and peripheral clocks. Nat Rev Neurosci 2001;2:521–6. Golombek DA, Rosenstein RE. Physiology of circadian entrainment. Physiol Rev 2010;90:1063–102. Kalsbeek A, Bruinstroop E, Yi CX, Klieverik LP, La Fleur SE, Fliers E. Hypothalamic control of energy metabolism via the autonomic nervous system. Ann NY Acad Sci 2010;1212:114–29. Kalsbeek A, Fliers E, Hofman MA, Swaab DF, Buijs RM. Vasopressin and the output of the hypothalamic biological clock. J Neuroendocrinol 2010;22:362–72. Kalsbeek A, Kreier F, Fliers E, Sauerwein HP, Romijn JA, Buijs RM. Circadian control of metabolism by the suprachiasmatic nuclei. Endocrinology 2007;148:5635–9. Lee S, Donehower LA, Herron AJ, Moore DD, Fu L. Disrupting circadian homeostasis of sympathetic signaling promotes tumor development in mice. PLoS One 2010;5(6):e10995. Maywood ES, O’Neill JS, Reddy AB, Chesham JE, Prosser HM, Kyriacou CP, et al. Genetic and molecular analysis of the central and peripheral circadian clockwork of mice. Cold Spring Harb Symp Quant Biol 2007;72:85–94. Scheer FA, Kalsbeek A, Buijs RM. Cardiovascular control by the suprachiasmatic nucleus: neural and neuroendocrine mechanisms in human and rat. Biol Chem 2003;384:697–709. Schibler U. Circadian time keeping: the daily ups and downs of genes, cells, and organisms. Prog Brain Res 2006;153:271–82. Takahashi JS, Hong HK, Ko CH, McDearmon EL. The genetics of mammalian circadian order and disorder: implications for physiology and disease. Nat Rev Genet 2008;9:764–75.

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33 Baroreceptor Reflexes Mark W. Chapleau Changes in blood pressure (BP) and/or blood volume are “sensed” within specific compartments of the cardiovascular system by “baroreceptors” (Fig. 33.1). Baroreceptors are mechanosensitive nerve endings that are activated by vascular and/or cardiac distension during increases in intraluminal BP. The activity of arterial baroreceptors innervating large arteries (primarily aortic arch and carotid sinuses) is increased when arterial BP rises, and decreased when BP falls. The changes in baroreceptor activity evoke rapid reflex adjustments that buffer or oppose the changes in arterial BP in a negative-feedback manner. Cardiopulmonary baroreceptors innervate the heart, vena cava, and pulmonary vasculature. Since the activity of cardiopulmonary baroreceptors correlates with intrathoracic (central) blood volume, these nerve endings are often referred to as “volume receptors” or “low-pressure” baroreceptors. The reflex adjustments triggered by changes in cardiopulmonary baroreceptor activity regulate blood volume in addition to influencing BP.

NEURAL PATHWAYS AND EFFECTOR MECHANISMS Arterial Baroreflex The neural pathways and effector mechanisms involved in baroreflex control of the circulation are summarized in Figures 33.1 and 33.2. The cell bodies (somata) of carotid sinus and aortic arch baroreceptor neurons are located in petrosal and nodose ganglia, respectively. The corresponding afferent baroreceptor activity is transmitted to the nucleus tractus solitarius (NTS) in the medullary brain stem via carotid sinus and glossopharyngeal nerves, and aortic depressor and vagus nerves, respectively. The baroreceptor inputs are integrated and relayed through a network of central nervous system (CNS) neurons controlling efferent parasympathetic nerve activity (paraSNA), sympathetic nerve activity (SNA), and release of the vasoconstrictor and antidiuretic peptide vasopressin (AVP) from the posterior pituitary gland (Figs 33.1 and 33.2). The multiple effector mechanisms by which these systems buffer increases in BP are depicted in Figure 33.2. The effector mechanisms operate in the opposite direction when arterial BP and baroreceptor activity are reduced.

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BP-sensitive sensory nerves also innervate the juncture of the right carotid and right subclavian arteries (Fig. 33.1) and the coronary arteries. These nerves travel in the aortic depressor and vagus nerves with cell bodies located in the nodose ganglia. Activation of the coronary artery baroreflex modulates peripheral SNA and vascular resistance, but unlike carotid sinus and aortic arch baroreflexes, has little or no affect on HR.

Cardiopulmonary Baroreflex The cardiopulmonary region is innervated by multiple types of mechanosensitive and chemosensitive sensory nerves that affect autonomic and cardiovascular functions in a variety of ways (see Chapter 35). We focus here on vagal afferent neurons with cell bodies in the nodose ganglia and nerve endings in the heart, vena cava, and pulmonary vasculature that are sensitive to changes in central blood volume (Fig. 33.1). The electrophysiological properties of these sensory neurons, the CNS pathways engaged by their activation, and their influence on efferent effectors are similar to that of arterial baroreceptor neurons (see Figs 33.1 and 33.2), but not identical. While changes in cardiopulmonary baroreceptor activity during changes in central blood volume evoke powerful reflex changes in peripheral SNA, vascular resistance, and release of renin and AVP; the reflex has little affect on HR. The changes in SNA and vascular resistance contribute to orthostatic adjustments. Renal actions of SNA, the renin-angiotensinaldosterone system, and AVP leading to changes in Na and water reabsorption play a major role in regulation of blood volume.

DETERMINANTS OF AFFERENT BARORECEPTOR ACTIVITY Rate Sensitivity of Baroreceptors Baroreceptor activity is dependent not only on the mean level of BP, but also on the direction and rate of change in BP. Consequently, baroreceptor activity will increase or decrease to a greater extent when the change in BP occurs more rapidly leading to a more effective reflex compensation. Similarly, baroreceptor activity is higher

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FIGURE 33.1 Location of baroreceptors and neural pathways mediating baroreflex responses. (A) Arterial baroreceptor afferents innervate carotid sinuses, aortic arch and the right carotid artery-right subclavian artery juncture. Cardiopulmonary baroreceptors innervate veno-atrial juncture, atria, ventricles and pulmonary vasculature. The baroreflexes modulate paraSNA and SNA to numerous organ systems and vasopressin (AVP) release. Key targets involved in cardiovascular regulation are illustrated. (B) Major nuclei involved in baroreflex control. Increases in BP and baroreceptor activity activate excitatory neural projections from nucleus tractus solitarius (NTS) to preganglionic parasympathetic neurons in nucleus ambiguus (NA) and dorsal motor nucleus of the vagus (DMNX) resulting in increases in paraSNA and decreases in HR. Activation of excitatory projections from NTS to caudal ventrolateral medulla (CVLM) causes subsequent inhibition of premotor sympathetic neurons in rostral ventrolateral medulla (RVLM) that project to preganglionic sympathetic neurons in the intermediolateral (IML) column of the thoracolumbar spinal cord. Increased baroreceptor activity also inhibits secretion of AVP from magnocellular neurons in paraventricular nucleus (PVN) and supraoptic nucleus (SON) of hypothalamus. Other CNS regions interact with these areas to modulate baroreflexes.

FIGURE 33.2 Effector mechanisms mediating reflex responses to increases in baroreceptor activity. Increases in arterial BP and baroreceptor activity increase paraSNA, decrease SNA, and inhibit release of AVP leading to an array of cardiovascular, hormonal and renal responses. Decreases in arterial BP evoke directionally opposite reflex responses. A-V node, atrio-ventricular node; Ang II, angiotensin II; H2O, water.

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during the systolic phase of the arterial pressure pulse and lower or absent during diastole. The phasic discharge of afferent activity facilitates reflex inhibition of SNA.

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and autocrine/paracrine factors including norepinephrine, prostacyclin, serotonin, nitric oxide, and reactive oxygen species modulate baroreceptor sensitivity through actions on these ion channels and membrane pumps.

Large Artery Compliance Baroreceptors are not directly sensitive to BP, but rather are sensitive to mechanical deformation of the nerve endings during distension of the arterial wall. Therefore, large artery compliance (specifically of the carotid sinuses and aortic arch) is a major determinant of the baroreceptor sensitivity to changes in BP. Decreased arterial compliance contributes to decreased baroreceptor sensitivity in atherosclerosis, hypertension, and aging.

Neuronal Mechanisms Mediating Sensory Transduction The prevailing view is that baroreceptors are activated by the opening of mechanosensitive ion channels in the sensory terminals. The resulting depolarization, if of sufficient magnitude, will trigger action potential discharge upon opening of voltage-gated Na and K channels. The action potentials are propagated towards the CNS at frequencies related to the magnitude of deformation and depolarization of the sensory terminals. Evidence suggests that members of the epithelial sodium channel (ENaC) superfamily including acid-sensing ion channel 2 (ASIC2) are components of the mechanosensitive ion channel complex. Transient receptor potential (TRP) channels have also been implicated in baroreceptor sensory transduction and/ or signaling, perhaps functioning as a mechanosensor. A variety of voltage- and ligand-gated ion channels and membrane pumps modulate the membrane potential and excitability of baroreceptors including Kv1, Kv4, BK and KCNQ (M-type) K channels; tetrodotoxin-insensitive, voltage-gated Na channels; hyperpolarization-activated cyclic nucleotide-gated (HCN) channels; serotonin 5HT3 receptor/ channels; and the Na/K–ATPase. Several neurohumoral

BAROREFLEX ADAPTATION AND RESETTING IN ACUTE HYPERTENSION Baroreceptor activity increases with a rise in arterial BP but declines over time if the acute hypertension is maintained. Furthermore, “post-excitatory depression” (PED) of baroreceptor activity occurs when BP decreases rapidly after a period of increased BP. Different mechanisms have been implicated in these two phenomena with opening of 4-aminopyridine sensitive K channels contributing to adaptation and activation of the Na/K–ATPase causing PED. Baroreceptor adaptation and PED contribute to acute resetting of the baroreceptor pressure-activity relationship to higher mean pressures in hypertension. The baroreceptor function curve is shifted in a parallel manner with little or no change in baroreceptor sensitivity (slope), and is usually accompanied by resetting of the arterial BP-HR relation (Fig. 33.3). Central mechanisms may exacerbate or oppose resetting of the baroreflex function curve. While baroreflex resetting compromises the ability to counter the sustained hypertension, it helps preserve the ability to buffer acute fluctuations in BP at the new higher prevailing level of BP.

DECREASED BAROREFLEX SENSITIVITY IN DISEASE Control of HR vs. SNA and BP, and Underlying Mechanisms Baroreflex sensitivity (BRS) for control of HR is consistently decreased in numerous pathological states including chronic hypertension, coronary artery disease,

FIGURE 33.3 Baroreceptor and baroreflex resetting during acute hypertension. An increase in mean arterial BP increases baroreceptor activity (left) and reflexively decreases HR (right). During a sustained increase in BP, baroreceptor activity decreases or adapts over time (left) and HR increases at the same level of BP (right). The baroreceptor and baroreflex function curves are shifted (reset) to higher BP during acute hypertension with preservation of the slope of the curves (sensitivity) (dashed lines).

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post-myocardial infarction, heart failure, diabetes mellitus, and obesity, and with aging. Afferent, central, and efferent mechanisms contribute to varying degrees to decreased BRS in these diseases. Chronic structural changes such as decreased large artery compliance and cardiac hypertrophy impair the afferent sensitivity of arterial and cardiopulmonary baroreceptors. In addition, neurohumoral activation and oxidative stress impair baroreflex function. For example, increased circulating levels of angiotensin II (Ang II) reset the baroreflex function curve to a higher mean level of arterial BP. This resetting is mediated by actions of Ang II on circumventricular organs that lack a blood–brain barrier (e.g., area postrema) and is independent of the rise in BP. Furthermore, Ang II acts at multiple central and peripheral sites in the nervous system to increase SNA, and decrease paraSNA and BRS. Aldosterone inhibits BRS by reducing afferent baroreceptor activity and by central actions. Factors released from activated platelets and reactive oxygen species decrease baroreceptor afferent sensitivity. Oxidative stress in the CNS contributes to increased SNA and BP in animal models of hypertension. Antioxidant therapies improve BRS in hypertension, heart failure, and in aging. In contrast to control of HR, the effects of cardiovascular disease and aging on arterial baroreflex control of SNA and BP are controversial with reports of both impaired and preserved BRS. Differences in results between studies can be explained in part by differences in: (i) the methods used to quantify changes in SNA and evaluate BRS; (ii) engagement of arterial vs. cardiopulmonary baroreflexes; (iii) control of SNA to different peripheral targets; (iv) severity of disease; and (v) experimental conditions (e.g., use of anesthesia). In most conditions, decreased BRS for parasympathetic control of HR usually precedes and/ or exceeds the decrease in BRS for sympathetic control. Structural damage to baroreceptor afferents, usually resulting from surgery or radiation, can cause “baroreflex failure” with loss of cardiovagal tone and severe, episodic periods of sympathetic-mediated hypertension (see Chapter 72). A similar phenotype is observed in patients with familial dysautonomia, a rare genetic disease with severe developmental sensory nerve defects (see Chapter 103).

Genetic Determinants of BRS Decreased BRS may be secondary to underlying cardiovascular disease or may precede and contribute to disease. BRS for control of HR is impaired in normotensive subjects with a family history of hypertension, and heritability of BRS has been confirmed in twin studies. Polymorphisms in several genes have been reported to be associated with BRS (see Chapter 34). Therefore, BRS screening in high risk patients identified by disease and/ or by the presence of specific polymorphisms may be advisable. The ability to measure BRS noninvasively from spontaneous fluctuations in systolic BP and pulse interval makes this clinical application feasible.

BRS: A DETERMINANT OF CARDIOVASCULAR RISK AND THERAPEUTIC TARGET Decreased BRS and Cardiovascular Risk Increased BP variability causes target organ damage, e.g., endothelial dysfunction, vascular and cardiac hypertrophy, kidney disease and cerebral vascular dysfunction. These insults lead to myocardial infarction, stroke, and heart and kidney failure. By minimizing BP variability and restraining SNA and BP, the arterial and cardiopulmonary baroreflexes reduce target organ damage. In addition to regulating BP, baroreflexes exert a major influence on the electrical properties of the heart through modulation of cardiac SNA and paraSNA. Myocardial infarction, heart failure, and diabetes are associated with decreased BRS for control of HR, cardiac arrhythmias, and sudden cardiac death. The decrease in BRS predicts occurrence of arrhythmias and mortality in patients suffering from these diseases suggesting a causal relationship.

BRS is a Therapeutic Target The strong inverse relationship between BRS and cardiovascular risk encourages targeting therapy to improve BRS. Baroreflexes may contribute to the benefit of standard antihypertensive therapies. For example, lowering of BP of hypertensive patients by pharmacological or dietary interventions rapidly resets the baroreflex function curve to lower mean arterial BPs. The baroreflex resetting helps stabilize BP at the new lower prevailing level of BP. Reversal of vascular and cardiac stiffening and hypertrophy with longer periods of antihypertensive treatment increases BRS. Antagonists of the renin-angiotensinaldosterone system and antioxidants increase BRS independent of BP lowering, thus providing further reductions in cardiovascular risk. Recent findings have rejuvenated the concept of specific therapeutic targeting of baroreflex pathways in cardiovascular disease. Cholinesterase inhibitors promote increases in cardiovagal tone and BRS by increasing the concentration of the neurotransmitter acetylcholine at cholinergic synapses and sinoatrial node, and amplify cholinergic signaling in left ventricle. Administration of cholinesterase inhibitors and chronic electrical stimulation of the vagus nerve each result in novel downstream anti-inflammatory effects and increased survival post-myocardial infarction. A recent study examining effects of chronic vagus nerve stimulation in patients with heart failure has provided promising results. Chronic electrical stimulation of carotid sinus baroreceptors in dog models of hypertension and in patients with drug-resistant hypertension have demonstrated long-term efficacy in lowering BP and reducing target organ damage. Chronic carotid sinus baroreceptor stimulation has also been shown to improve cardiac function, decrease arrhythmias, and prolong survival in

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BRs: A DETERmINANT of CARDIoVAsCulAR RIsk AND THERAPEuTIC TARgET

dog models of heart failure. Clearly, favorable effects of increasing baroreceptor activity extend far beyond shortterm control of BP.

Further Reading Brooks VL, Sved AF. Pressure to change? Re-evaluating the role of baroreceptors in the long-term control of arterial pressure. Am J Physiol Regul Integr Comp Physiol 2005;288:R815–8. Chapleau MW, Li Z, Meyrelles SS, Ma X, Abboud FM. Mechanisms determining sensitivity of baroreceptor afferents in health and disease. Ann NY Acad Sci 2001;940:1–19. Chapleau MW, Lu Y, Abboud FM. Mechanosensitive ion channels in blood pressure-sensing baroreceptor neurons. Hamill OP, editor. Current topics in membranes, Vol 59. : Elsevier Science; 2007. p. 541–67. Chapleau MW, Sabharwal R. Methods of assessing vagus nerve activity and reflexes. Heart Fail Rev 2011;16:109–27. Glazebrook PA, Ramirez AN, Schild JH, Shieh C-C, Doan T, Wible BA, et al. Potassium channels Kv1.1, Kv1.2 and Kv1.6 influence excitability of rat visceral sensory neurons. J Physiol 2002;541.2:467–82. Glazebrook PA, Schilling WP, Kunze DL. TRPC channels as signal transducers. Pflugers Arch 2005;451:125–30. Guyenet PG. The sympathetic control of blood pressure. Nature Rev Neurosci 2006;7:335–46. Hainsworth R. Reflexes from the heart. Physiol Rev 1991;71(3):617–58. Handa T, Katare RG, Kakinuma Y, Arikawa M, Ando M, Sasaguri S, et al. Anti-Alzheimer’s drug, donepezil, markedly improves long-term survival after chronic heart failure in mice. J Cardiac Fail 2009;15:805–11.

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Korner PI. Baroreceptor resetting and other determinants of baroreflex properties in hypertension. Clin Exp Pharmacol Physiol Suppl 1989;15:45–64. La Rovere MT, Bigger Jr. JT, Marcus FI, Mortara A, Schwartz PJ for the ATRAMI investigators. Baroreflex sensitivity and heart-rate variability in prediction of total cardiac mortality after myocardial infarction. The Lancet 1998;351:478–84. Lu Y, Ma X, Sabharwal R, Snitsarev V, Morgan D, Rahmouni K, et al. The ion channel ASIC2 is required for baroreceptor and autonomic control of the circulation. Neuron 2009;64:885–97. Monahan KD, Eskurza I, Seals DR. Ascorbic acid increases cardiovagal baroreflex sensitivity in healthy older men. Am J Physiol Heart Circ Physiol 2004;286:H2113–H2117. Parati G, Di Rienzo M, Mancia G. How to measure baroreflex sensitivity: from the cardiovascular laboratory to daily life. J Hypertens 2000;18:7–19. Schwartz PJ, De Ferrari GM, Sanzo A, Landolina M, Rordorf R, Raineri C, et al. Long term vagal stimulation in patients with advanced heart failure: First experience in man. Eur J Heart Fail 2008;10:884–91. Sun H, Li D-P, Chen S-R, Hittelman WN, Pan H-L. Sensing of blood pressure increase by transient receptor potential vanilloid 1 receptors on baroreceptors. J Pharmacol Exp Ther 2009;331:851–9. Taylor JG, Bisognano JD. Baroreflex stimulation in antihypertensive treatment. Curr Hypertens Rep 2010;12:176–81. Wladyka CL, Feng B, Glazebrook PA, Schild JH, Kunze DL. The KCNQ/ M-current modulates arterial baroreceptor function at the sensory terminal in rats. J Physiol 2008;586.3:795–802. Wright C, Drinkhill MJ, Hainsworth R. Reflex effects of independent stimulation of coronary and left ventricular mechanoreceptors in anaesthetized dogs. J Physiol 2000;528.2:349–58.

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34 Genetic Determinants of Baroreflex Function Italo Biaggioni The baroreflex is the archetypical mechanism of autonomic cardiovascular regulation. Its neural pathways, physiological relevance, and pathological consequences of its failure are described in detail in Chapters 33 and 72 and will not be repeated here. The importance of the baroreflex in a given subject can be assessed by the gain of the system, i.e., the magnitude of the reciprocal change in heart rate or sympathetic activity in response to alterations in blood pressure. The slope of the relationship between blood pressure and heart rate or sympathetic activity is used as a measure of baroreflex sensitivity (BRS). There is substantial inter-individual variability in these parameters. This is not surprising given the complex neural pathways involved and the biological measurements used in its estimation. Furthermore, most of the methods used to assess baroreflex function cannot isolate the predominant contribution of carotid sinus arterial baroreceptors to that of other arterial and venous afferents. In this chapter we will review the evidence suggesting that a significant component of the variability in baroreflex function is genetically determined. Less is known about the determinants of this genetic influence. A few studies have identified discrete polymorphisms associated with BRS. In general these studies have included relatively small number of subjects and results have not always been replicated. In some cases, the predicted functional consequences of these polymorphisms in the gene products provide a plausible biological explanation of their putative effect in BRS, but a direct causal relationship has been difficult to document. Earlier studies showed that baroreflex function was decreased in normotensive offspring of hypertensive parents, and that decreased baroreflex gain was a predictive factor for the subsequent development of hypertension [1]. More definitive proof of genetic influences on baroreflex function came from studies in twins showing significant correlation for BRS in monozygotic but not dizygotic twins [2]. It has been difficult to ascertain the role of individual genes in the modulation of complex phenotypes like the baroreflex. The baroreflex is the result of the interplay between afferent fibers located in vascular structures

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00034-2

responsive to stretch, the integration of these afferent signals in brainstem nuclei, efferent fibers carrying sympathetic and parasympathetic outflow and coupling of these fibers with cardiac and vascular structures (Chapter 33). It is likely that different genes influence each of these individual components of the baroreflex. A case in point is the observation that normotensive children with Williams syndrome have a significant reduction in BRS [3]. This syndrome is caused by the microdeletion of the long arm of chromosome 7 encompassing the locus of the elastin gene. Elastin is the main component of the extracellular matrix of arteries, and affected patients develop an arteriopathy with intimal thickening. It is plausible, therefore, that this pathology affects pressure transduction at the level of baroreceptor afferents resulting in the observed reduction in BRS. However, in humans it is not feasible to test this hypothesis by measuring afferent firing rates from the carotid sinus in response to pressure changes. The same is true for other individual components of the baroreflex arc; a gene polymorphism of deletion can theoretically affect the function of afferent structures, central brainstem pathways, or efferent neurovascular coupling. We are only able, however, to measure integrated baroreflex function and this approach will necessarily “dilute” any genetic influence on the individual components of the baroreflex. An additional level of complexity is introduced by the fact that the “baroreflex” is not a single entity, but has separate afferent inputs arising from high pressure (the carotid sinus, aortic arch and others) and low pressure (large veins) vascular structures. The efferent pathways are also differentiated into those modulating heart rate, which are predominately parasympathetic, and vascular tone, which are mostly sympathetic. It would be impossible to phenotype each of these components in the large number of patients required for genetic studies. The most commonly used method, therefore, is to estimate the relationship between systolic blood pressure and RR interval during spontaneous periods when parallel increases (or decreases) in these parameters occur. There is inherent variability in these measurements, which may also “dilute” a genetic effect.

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Despite all these limitations, a handful of studies have described associations between discrete gene polymorphisms and BRS. Polymorphisms of KCNMB1 gene coding the β1 subunit of the calcium-sensitive potassium channel BK was associated with BRS in 298 normotensive twins [4]. In particular, AA homozygous in the polymorphism of exon 4b had greater BRS than heterozygous or CC homozygous. Potassium channels (BK) are expressed in vascular smooth muscle cells and are important to calcium-mediated relaxation. They are, however, also expressed in neural tissues and the site where this polymorphism may alter BRS could not be determined from these studies. AA homozygous also had higher heart rate variability in the high frequency range, a measure of increased vagal tone. It is possible, therefore, that the increased BRS was due to the putative increase in vagal tone. A common polymorphic variant in the promoter region of the bradykinin B2 receptor gene, T-58C, was also associated with baroreflex function; greater number of T alleles was associated with increased BRS in 129 untreated hypertensive subjects, but not in 95 normotensive controls [5]. This polymorphism could account for 12% of BRS variation even after accounting for blood pressure, BMI, age and gender. In transfected cell models the  58C allele was associated with reduced B2R expression, leading the authors to suggest that decreased bradykinin actions could contribute to the reduction in BRS. In a study of Finnish subjects, a C-344T polymorphism of the aldosterone synthase gene (CYP11B2) was associated with BRS measured from the overshoot phase of the Valsalva maneuver, with the CC allele associated with decreased BRS [6]. No comparable associations were found for BRS with an insertion/deletion (I/D) polymorphism of angiotensin-converting enzyme (ACE), or the M235T variants of angiotensinogen (AGT). The C-344T polymorphism is a common variant in the promoter region of the gene. The C allele has been found to be associated with elevated plasma levels of aldosterone. Ormezzano et al studied 17 polymorphisms of 11 genes relevant to blood pressure regulation in 146 hypertensive individuals and 105 healthy controls. Only the C  1222T polymorphism of endothelin receptor A, EDNRA, correlated with BRS in both populations after adjustment for age, gender, blood pressure and BMI [7]. Importantly, the association remained significant after accounting for the multiple number of genes tested. No other polymorphism was significantly correlated with BRS, including the CYP11B2 previously reported by Ylitalo [6]. Similarly, the C allele of the A1166C polymorphism in the AT1 receptor gene was associated with reduced BRS in 135 normotensive individuals [8]. Of note, this variant occurs in an untranslated region of the gene and is not associated with a functional change in the AT1 receptor but could be in linkage disequilibrium with other sites. Also, this appears to be one of the polymorphisms previously reported not to be associated with BRS [7].

Recently, Xing-Sheng et al evaluated the influence of nine polymorphisms in six genes, in 182 normotensive Chinese men, and found that carriers of the TT genotype of the T-786C polymorphism of the endothelial nitric oxide synthase gene had lower BRS than subjects carrying either the TC or the CC genotype [9]. It is believed that the C allele is associated with reduced expression of eNOS. The finding that subjects carrying the C allele had higher BRS would imply that eNOS-generated NO normally depresses baroreflex function. They also found that subjects with the TT genotype of the C-344T polymorphism in the aldosterone gene had higher BRS as compared to those with CC confirming the results from Ylitalo et al. [6]. Finally, subjects with the CC genotype of the T-58C polymorphism of the B2R gene had lower BRS compared to subjects with TT. These results are similar to those reported by Milan et al. in hypertensive subjects, even though the latter group of investigators found no association in normotensive subjects. Multivariable regression indicated that the three polymorphisms reported by XingSheng could explain 16% of BRS variability [9]. The measurement of BRS in virtually all of the studies described above relies on spontaneous reciprocal changes in blood pressure and heart rate analyzed with spectral analysis techniques from non-invasive measurements. This parameter estimates only the vagal modulation of heart rate by the baroreflex. Cardiovagal BRS has consistently shown to be reduced in hypertensive populations, but it is reassuring that none of the polymorphisms associated with reduced cardiovagal BRS were found to correlate with blood pressure itself, suggesting that differences in BRS reported were not secondary to blood pressure changes. It is known that the hypertensive trait has a significant genetic component, but most attempts to identify relevant genes have been disappointing; in most cases, the genes that have been identified account for only a few mmHg in blood pressure. There is less information about the hereditability of the sympathetic limb of the baroreflex modulating vascular tone. Yamada et al. studied three groups of adolescents: normotensives with a positive family history of hypertension, borderline hypertensives with a positive family history, and normotensives with a negative family history [10]. “Sympathovascular” BRS was estimated by the slope of the percent decrease in muscle sympathetic nerve activity in response to increases in systolic blood pressure induced by phenylephrine. “Sympathovascular” BRS was significantly smaller in the groups with a family history of hypertension (  8  1 %/mm Hg for both the borderline and normotensive offspring) compared to the normotensive controls (  16  1 %/mm Hg). The sympathetic contribution to blood pressure, as determined by baseline MSNA and the fall in blood pressure induced by the ganglionic blocker trimethaphan, was greater in in borderline hypertensive offspring than in normotensives with a positive family history or normotensive controls. These results indicate an impaired baroreflex capacity to dampen

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sympathetic activity resulting in greater “sympathetically driven” blood pressure in adolescents with a family history of hypertension, even before they develop hypertension. We should note that cardiovagal BRS is consistently reduced in hypertensive subjects, but baroreflex control of sympathetic tone in most studies has not been different than normotensive controls. Whereas these studies evaluated arterial high-pressure baroreceptors, Ookuwa et al. studied the function of low pressure baroreceptors located in the venous side of the circulation. Cardiopulmonary baroreflex control of forearm vascular resistance was evaluated in 12 normotensive young subjects with a family history of hypertension and compared with normotensives without a family history of hypertension [11]. Low-pressure cardiopulmonary baroreceptors were preferentially unloaded using low levels (  20 mmHg) of lower body negative pressure (LBNP). It should be noted, however, that even these low levels of LBNP can unload high-pressure baroreceptors. The reflex increase in forearm vascular resistance in response to LBNP was significantly lower in normotensive with a family history of hypertension (38  8%) than in normotensives with a negative family history (86  19%). These results suggest that cardiopulmonary baroreflexes are also impaired in subjects with a genetic predisposition to hypertension, even during the normotensive stage. In summary, there is substantial evidence indicating that a significant component of baroreflex function is genetically determined. Furthermore, reduced BRS is found in offspring of hypertensive parents, and predicts the development of hypertension. It is not clear whether reduced baroreflex buffering contributes to the development of hypertension or is mostly a trait of an underlying abnormality in autonomic regulation of blood pressure. A few polymorphisms have been associated with reduced baroreflex function, but it is not clear where in the baroreflex arc those genes are operative, nor whether there is a

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causal relationship. This, perhaps, is not surprising given the complex nature of baroreflex pathways and inherent limitations in clinical measures of its function.

References [1] Ducher M, Fauvel JP, Cerutti C. Risk profile in hypertension genesis: A five-year follow-up study. Am J Hypertens 2006;19:775–80. [2] Tank J, Jordan J, Diedrich A, Stoffels M, Franke G, Faulhaber HD, et al. Genetic influences on baroreflex function in normal twins. Hypertension 2001;37:907–10. [3] Girard A, Sidi D, Aggoun Y, Laude D, Bonnet D, Elghozi JL. Elastin mutation is associated with a reduced gain of the baroreceptor-heart rate reflex in patients with Williams syndrome. Clin Auton Res 2002;12:72–7. [4] Gollasch M, Tank J, Luft FC, Jordan J, Maass P, Krasko C, et al. The BK channel beta1 subunit gene is associated with human baroreflex and blood pressure regulation. J Hypertens 2002;20:927–33. [5] Milan A, Mulatero P, Williams TA, Carra R, Schiavone D, Martuzzi R, et al. Bradykinin B2 receptor gene (-58T/C) polymorphism influences baroreflex sensitivity in never-treated hypertensive patients. J Hypertens 2005;23:63–9. [6] Ylitalo A, Airaksinen KE, Hautanen A, Kupari M, Carson M, Virolainen J, et al. Baroreflex sensitivity and variants of the renin angiotensin system genes. J Am Coll Cardiol 2000;35:194–200. [7] Ormezzano O, Poirier O, Mallion JM, Nicaud V, Amar J, Chamontin B, et al. A polymorphism in the endothelin-A receptor gene is linked to baroreflex sensitivity. J Hypertens 2005;23:2019–26. [8] Jira M, Zavodna E, Honzikova N, Novakova Z, Vasku A, Izakovicova HL, et al. Association of A1166C polymorphism in AT(1) receptor gene with baroreflex sensitivity. Physiol Res 2010;59:517–28. [9] Xing-Sheng Y, Yong-Zhi L, Jie-Xin L, Yu-Qing G, Zhang-Huang C, Chong-Fa Z, et al. Genetic influence on baroreflex sensitivity in normotensive young men. Am J Hypertens 2010;23:655–9. [10] Yamada Y, Miyajima E, Tochikubo O, Matsukawa T, Shionoiri H, Ishii M, et al. Impaired baroreflex changes in muscle sympathetic nerve activity in adolescents who have a family history of essential hypertension. J Hypertens 1988;6:S525–8. [11] Ookuwa H, Takata S, Ogawa J, Iwase N, Ikeda T, Hattori N. Abnormal cardiopulmonary baroreflexes in normotensive young subjects with a family history of essential hypertension. J Clin Hypertens 1987;3:596–604.

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35 Cardiac and Other Visceral Afferents John C. Longhurst, Liang-Wu Fu INTRODUCTION Visceral afferents convey information to the central nervous system about local changes in chemical and mechanical environments of a number of organ systems. Physiologically, the autonomic sensory nervous system provides information about the function of abdominal and thoracic organ systems, allowing for reflex responses that typically aid in the normal function of these systems. Pathophysiologically, these sensory nerves provide a warning system to alert the organism to the presence of injury or conditions that can lead to injury and cell death. Thus, they assist both in normal function and maintain homeostasis in adverse conditions. The reflex arc includes the afferent or sensory limb, central neural processing and efferent motor system innervating effector organs. This chapter focuses on afferent fibers present in vagus and sympathetic (spinal) pathways that respond to mechanical and chemical alterations in the environment, and the resulting reflex responses, using ischemia as the stimulus paradigm. Ischemia constitutes an important condition associated with cardiovascular disease. Stimulation of visceral sensory nerves leads to important cardiovascular reflex responses mediated by the autonomic nervous and humoral systems.

ANATOMICAL FRAMEWORK Finely myelinated (Aβ, Aδ) and unmyelinated (C fibers) afferent pathways innervating either unspecialized dense, diffuse or bare nerve endings, form the afferent pathway of visceral cardiovascular reflexes, Table 35.1 [1]. The nerve endings typically are located within the interstitial space and respond to mechanical and/or chemical events. Afferents ascend to the central nervous system (CNS) through mixed nerves, including the vagus or sympathetic (spinal, e.g., spinothalamic and spinoreticular) pathways. A number of nuclei in the thalamus, hypothalamus, midbrain, pons and medulla, including the nucleus tractus solitarii, caudal and rostral ventral lateral medulla, parabrachial nucleus, paraventricular nucleus, periaqueductal gray, lateral tegmental field, medullary raphé, and nucleus ambiguus, among others, process input from visceral afferents ultimately regulating sympathetic and

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00035-4

vagal autonomic motor fiber discharge activity directed to cardiovascular effector organs. These reflex arcs form the pathways for visceral reflexes concerned with regulation of autonomic outflow to the cardiovascular system.

AFFERENT STIMULI Visceral afferent fibers responsive to mechanical stimuli are either high or low threshold with the high threshold endings frequently serving as nociceptors [2–4]. Low threshold mechanosensitive receptors respond to changes in stress or strain and provide information relevant to digestion (gut), cardiac filling or function (cardiac venoatrial, atrial and ventricular). For example, cardiac atrial receptors, located mainly in the veno-atrial junctions, that are innervated by myelinated afferents that course through the vagus, respond to changes in balloon distension involving modest changes in volume but high tensions [5]. Many ventricular mechanosensitive C-fibers respond to changes in end-diastolic volume, and hence stretch, rather than systolic pressure, i.e., compression [6]. Sensory nerves that respond to changes in stress or strain in the gastrointestinal tract largely course through the vagus nerves and likely are concerned with transmission of information related to digestion rather than cardiovascular function [2].

ISCHEMIA Many high threshold mechanosensitive endings are triggered by chemical events and hence are bimodal in their sensitivity. Chemical stimuli activating these endings depend on the organ in which they are situated and the condition imposed [4]. Ischemia, for example, leads to the production and release of protons, kinins, serotonin, histamine, cyclooxygenase products like thromboxane, endothelin and reactive oxygen species, including hydroxyl radicals, among others (Fig. 35.1). These mediators stimulate afferent endings and frequently sensitize them to the action of other chemical mediators [7]. Other ischemia-related chemical changes associated with activation of peripheral chemoreceptors, such as hypoxia and hypercapnia, are not important stimuli of chemosensitive

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TABLE 35.1 Classification of Afferent fibers from Abdominal Viscera CrossSectional Diameter (µm)

Conduction Velocity (m/sec)

Aβ (myelinated)

6–12

Aδ (finely myelinated)

C (unmyelinated)

Fiber Type

Terminal Ending

Effective Stimulus

20–84

Pacinian corpuscle

Vibration

2–6

3–30

Unknown, bare nerve endings

Vibration, pulse pressure, contraction, distension, chemicals, noxious stimuli

0.3–1.5

0.3–2.5

Unknown, bare nerve endings

Strong mechanical stimuli, chemicals, noxious stimuli

visceral sensory nerve endings [8,9]. This differential sensitivity has led to the use of the term “chemosensitive” visceral receptors to distinguish them from the arterial chemoreceptors. It is clear that visceral organs, such as the heart, are selectively responsive to chemical events related to altered metabolism associated with deprivation of blood flow and oxygen. The sources of many mediators are parenchymal cells in the organ, for example cardiac myocytes and endothelium [10]. Reactive oxygen species likely are derived from cardiac myocytes, while the endothelium is a major source for endothelin [11,12]. Circulating precursors or enzymes activated by ischemia initiate a cascade of events leading to the production of mediators like kinins, serve as an important source along with activated blood elements, such as platelets, which release several mediators during ischemia and reperfusion. Platelets aggregate at the site of injury of arterial endothelium following rupture of an atherosclerotic plaque or following occlusion of a coronary artery [13]. Activated platelets release serotonin, histamine and

FIGURE 35.1 Diagram of mediators, their receptors and cellular messaging mechanisms involved in activation of cardiac afferents during ischemia and reperfusion. Coronary artery occlusion, for example following plaque rupture induces myocardial ischemia leading to production and release of a number of mediators including 5-hydroxytryptamine (5-HT), histamine, bradykinin (BK), endothelin (ET), reactive oxygen species (ROS), lactic acid, ATP and COX pathway products, including thromboxane A2 (TxA2) and prostaglandins (PG) from various cells. Other abbreviations: adenosine 5'-triphosphate (ATP), purinergic 2X receptor (P2X), purinergic 2Y receptor (P2Y), TxA2/prostaglandin H2 receptor (TP), histamine 1 receptor (H1), 5-HT3 receptor (5-HT3), BK2 receptor (BK2), endothelin A receptor (ETA), transient receptor potential A1 (TRPA1), protons (H), phospholipase A2 (PLA2), phospholipase C (PLC), adenyl cyclase (AC), cyclic adenosine monophosphate (cAMP), phosphatidylinositol-4,5-bisphosphate (PIP2), inositol-1,4,5triphosphate (IP3), 1,2-diacylglycerol (DAG), protein kinase A (PKA), protein kinase C (PKC), arachidonic acid (AA) and cyclooxygenase (COX).

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Nerve Activity (impulses/5 s)

2.5

A

Control Ischemia

*

2.0

Nerve Activity (impulses/s)

Nerve Activity (impulses/s)

AuTOnOmIC REflEx REsPOnsEs TO VIsCERAl AffEREnT ACTIVATIOn

*

1.5 1.0 0.5 0.0

-Rabbit serum

+Rabbit serum

C

225 a 200 175 150 125 100 75 50 25 0 -5 -3 -1

225 200 175 150 125 100 75 50 25 0 1

3

5

7

9

2.5

B

*

2.0

*†

1.5 1.0 0.5 0.0

-Antiplatelet Ab

D

b

Anterior view

•∇•• •• •

∇ -5 -3 -1

Time (min)

1

3

5

7

9

+Antiplatelet Ab

Posterior view

•• ∇ • • ∇

FIGURE 35.2 Responses of cardiac ventricular afferents to repeated ischemia before and after treatment with control rabbit serum (A), exposure to a polyclonal rabbit antiplatelet antibody (B) and composite histogram of responses of eight afferents to five minutes of regional ischemia induced by occlusion of a left coronary artery branch before (a) and after (b) treatment with the antiplatelet antibody (C). Means and standard errors are shown. Panel D shows locations of the receptive fields of ischemically sensitive cardiac afferents on epicardial surface of left ventricle. Receptive fields of afferents included in this study: , Aδ afferents (n  4) and •, C-fiber afferents (n  10). * indicates nerve activity during ischemia was higher than during control (P  0.05) and † indicates response to ischemia after treatment with antibody was less than before (P  0.05). Modified from Fu, L-W and Longhurst, JC (2002) Role of activated platelets in excitation of cardiac afferents during myocardial ischemia in cats. Am J Physiol 282:H100–H109. Reproduced by permission.

thromboxane A2, each of which independently or in combination activate cardiac sympathetic afferent endings (Fig. 35.2). The action of each mediator is receptor mediated, with many, if not all of the receptors located on afferent endings. For example, bradykinin exerts its action through BK2, thromboxane through TP, histamine through H1, serotonin through 5-HT3 and endothelin through ETA receptors. Some mediators, like adenosine, remain controversial with regard to their role in stimulating afferent endings during ischemia [7,10]. Some reports, consisting mainly of reflex studies in the dog suggest that adenosine in the heart mediates autonomic reflexes that influence renal function. Other studies in cats suggest that adenosine does not stimulate ischemically sensitive cardiac afferents during ischemia [14].

AUTONOMIC REFLEX RESPONSES TO VISCERAL AFFERENT ACTIVATION Cardiovascular reflex responses to visceral afferent stimulation are either excitatory or inhibitory (Fig. 35.3). In this respect, stimulation of vagal afferents causes reflex

cardiovascular inhibition, including decreased heart rate, blood pressure, and myocardial contractility, consequent to reduced sympathetic outflow to the heart and blood vessels and increased vagal motor output to the heart [15]. Conversely, stimulation of sympathetic afferents that project centrally through sympathetic nerves and spinal pathways evokes reflex cardiovascular excitation, including increases in heart rate, blood pressure and myocardial performance, through increased sympathetic motor activity and, possibly, withdrawal of parasympathetic tone to the heart. Cardiovascular reflex responses originating from the heart consist of either reflex inhibitory or excitatory responses, or, more often, a combination of the two [16]. Thus, stimulation of the posterior-inferior and inner regions of the wall of the left ventricle leads to reflex bradyarrhythmias and hypotension, while stimulation of the anterior and superficial regions of the wall of the left ventricle leads to reflex tachyarrhythmias and hypertensive responses [5]. More commonly both vagal and sympathetic afferent pathways are stimulated concomitantly with a resulting reflex reflecting a mixed response consisting often of a small increase in blood pressure (Fig. 35.3) due to processing in the CNS, for example in the tractus solitarii and possibly the parabrachial nuclei [17].

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FIGURE 35.3 Dose-dependent responses of mean arterial pressure (MAP), heart rate (HR), and left ventricular dP/dt at 40 mmHg developed pressure (LV dP/dt40) to application of graded doses of hydrogen peroxide (H2O2) to the anterior surface of the ventricle in intact (vagal and sympathetic cardiac afferents intact, n  8), bilateral cervical vagotomized (sympathetic afferents intact, n  6) or bilateral T1–T4 ganglionectomized (vagal afferents intact, n  6) cats. Circles and brackets represent means and standard errors, respectively. * denotes significant difference comparing saline control to H2O2 (P  0.05). Modified with permission from Longhurst, JC. (2011) Regulation of autonomic function by visceral and somatic afferents. In: Central Regulation of Autonomic Functions. I. Llewellyn-Smith and A. Verberne, eds. New York: Oxford University Press.

Although stimulation of cardiac afferents can lead to reflex cardiovascular depression or excitation or a mixture of responses due to simultaneous stimulation of sensory fibers in both vagal and sympathetic pathways, stimulation of abdominal visceral afferents more commonly leads to reflex excitation since activation of afferents ascending through spinal pathways predominates. Thus, stimulation of chemosensitive, mechanosensitive and polymodal sensory endings in a number of abdominal organs activate spinal pathways that increase heart rate, myocardial contractility and arteriolar constriction in several regional circulations, including the coronary system [1,18]. Vagal afferent pathways from the abdominal region typically regulate digestive organ function, although recent studies suggest that during hypercapnic acidosis both spinal and vagal afferent activation by gastric distension leads to reflex cardiovascular depression as a result of sympathetic withdrawal and vagal activation [19]. Hence, the normal sympathoexcitatory responses evoked by abdominal visceral organ stimulation is converted to sympathoinhibition and vagal excitation by the altered baseline arterial blood gas status. These new findings may have important implications for patients undergoing abdominal surgery, when visceral afferents are stimulated either during vascular

compromise or mechanical traction. Either increases or decreases in blood pressure potentially can occur depending on the arterial blood gas and pH status. In addition to cardiovascular regulation, stimulation of visceral afferents can cause a number of other important reflex events. For example, stimulation of cardiac vagal afferents during myocardial ischemia can lead to relaxation of the stomach, the antecedent of nausea and vomiting that frequently accompanies inferior myocardial infarctions [5,10]. Stimulation of sympathetic afferents in the heart or abdominal region frequently leads to pain, e.g., angina pectoris [7]. Hence many high threshold visceral sympathetic afferents function as nociceptors [3].

PATHOLOGICAL ALTERATIONS OF VISCERAL AFFERENTS In addition to ischemia, a number of other conditions activate visceral afferents. For example, inflammation typically is associated with the production of chemical (and sometimes mechanical) changes, including increases in kinins, activation of the cyclooxygenase system and enhanced formation of reactive oxygen species. Interestingly,

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FIGURE 35.4 Bar graph summarizing average discharge activity (A) and total response over time (B) of 15 ischemically sensitive cardiac sympathetic afferents to injection of BK (1 μg), U46619 (5 μg), a thromboxane A2 (TxA2) mimetic or BK (1 μg)  U46619 (5 μg) into the left atrium (LA). Neurohistograms provide summed 2 s impulse activity of 15 cardiac afferents during administration of U46619 (C1), BK (C2) or BK  U46619 (C3). The total responses of afferent to the mediators were calculated by counting all spikes that occurred during the entire period of response when activity exceeded baseline rates by 20%. Columns and error bars represent means  standard errors. *Indicates significant difference comparing stimuli to controls (P  0.05), # significantly different from either BK or U46619 alone (P  0.05) and †denotes significant difference comparing actual BK  U46619 to predicted BK  U46619 (P  0.05). Modified from Fu L-W and Longhurst JC (2009) Bradykinin and thromboxane A2 reciprocally interact to synergistically stimulate cardiac spinal afferents during myocardial ischemia. Am J Physiol 298:H235–H244. Reproduced by permission.

some chemical mediators appear to act as primary stimuli while others may sensitize in either an additive or synergistic fashion (e.g., TxA2, Fig. 35.4) or modulate (e.g., histamine) sensory nerve endings to the action of the primary stimulus (bradykinin) [7]. Other clinical conditions, like hypertension or heart failure modify the responsiveness of nerve endings in the atria or ventricles [5]. Hypertension may do this by altering the parenchymal substrate in which the endings are located. Heart failure may do this by altering the sensitivity of the nerve ending to mechanical or chemical events.

Acknowledgments Research cited was supported by NIH grants HL066217.

References [1] Longhurst JC. Cardiovascular reflexes of gastrointestinal origin. In: Shepherd AP, Granger DN, editors. Physiology of the intestinal circulation raven. New York; 1984. p. 165–78.

[2] Longhurst J. Reflex effects from abdominal visceral afferents. In: Zucker IH, Gillmore JP, editors. Reflex control of the circulation. Caldwell, NJ: Telford Press; 1991. p. 551–77. [3] Pan H-L, Longhurst JC. Ischaemia-sensitive sympathetic afferents innervating the gastrointestinal tract function as nociceptors in cats. J Physiol (Lond) 1996;492:841–50. [4] Longhurst JC. Regulation of autonomic function by visceral and somatic afferents. In: Llewellyn-Smith I, Verberne AJM, editors. Central regulation of autonomic function (Second ed.). New York: Oxford University; 2011. [5] Longhurst JC. Cardiac receptors: Their function in health and disease. Prog Cardiovasc Dis 1984;XXVII:201–22. [6] Thoren PN. Characteristics of left ventricular receptors with nonmedullated vagal afferents. Circ Res 1977;40:415–21. [7] Fu L-W, Longhurst JC. Regulation of cardiac afferent excitability in ischemia. Handb Exp Pharmacol 2009:185–225. [8] Fu L-W, Pan H-L, Pitsillides K, Longhurst J. Hypoxia does not directly stimulate ischemically sensitive abdominal visceral afferents during ischemia. Am J Physiol 1996;271:H261–6. [9] Mark AL, Abboud FM, Heistad DD, Schmid PG, Johannsen UJ. Evidence against the presence of ventricular chemoreceptors activated by hypoxia and hypercapnia. Am J Physiol 1974;227: 273–9.

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[10] Longhurst J, Tjen-A-Looi S, Fu L-W. Cardiac sympathetic afferent activation provoked by myocardial ischemia and reperfusion: mechanisms and reflexes. Ann N Y Acad Sci 2001;940:74–95. [11] Grill HP, Zweier JL, Kuppusamy P, Weisfeldt ML, Flaherty JT. Direct measurement of myocardial free radical generation in an in vivo model: effects of postischemic reperfusion and treatment with human recombinant superoxide dismutase. J Am Coll Cardiol 1992;20:1604–11. [12] Hans G, Schmidt BL, Strichartz G. Nociceptive sensitization by endothelin-1. Brain Res Rev 2009;60:36–42. [13] Flores NA, Sheridan DJ. The pathophysiological role of platelets during myocardial ischemia. Cardiovasc Res 1994;28:295–302. [14] Pan H-L, Longhurst J. Lack of a role of adenosine in activation of ischemically sensitive cardiac sympathetic afferents in cats. Am J Physiol 1995;269:H106–13. [15] Fu L-W, Longhurst JC. Reflex pressor response to arterial phenylbiguanide; role of abdominal sympathetic visceral afferents. Am J Physiol 1998;275:H2025–H2035.

[16] Huang H-S, Stahl G, Longhurst J. Cardiac-cardiovascular reflexes induced by hydrogen peroxide in cats. Am J Physiol 1995;268:H2114–H2124. [17] Tjen-A-Looi S, Bonham A, Longhurst J. Interactions between sympathetic and vagal cardiac afferents in nucleus tractus solitarii. Am J Physiol 1997;272:H2843–H2851. [18] Longhurst JC. Chemosensitive abdominal visceral afferents. In: Gebhart GF, editor. Proceedings: visceral pain symposium. Seattle: IASP Press; 1995. p. 99–132. [19] Tjen-A-Looi SC, Hsiao AF, Longhurst JC. Central and peripheral mechanisms underlying gastric distension inhibitory reflex responses in hypercapnic-acidotic rats. Am J Physiol 2011;300:H1003–12.

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36 Autonomic Control of the Heart Rachel C. Drew, Lawrence I. Sinoway INTRODUCTION The cardiovascular system consists of the heart and a network of blood vessels that circulate blood to tissues and organs within the body. The heart works as a pump by forcing blood into the arterial circulation in order to deliver oxygen and nutrients to tissues, and remove carbon dioxide and waste products from these tissues in the venous circulation that carries blood back to the heart. This circulatory system is essential for normal homeostatic function, as well as regulation of body temperature, fluid maintenance and adjusting to altered physiological states such as exercise, and is dependent on the heart’s ability to operate properly.

THE HEART The heart is a powerful muscle that comprises of four chambers; right atrium, right ventricle, left atrium and

left ventricle (see Fig. 36.1). Blood enters the right atrium via the superior vena cava from the upper body and the inferior vena cava from the lower body. The right atrium contracts to push blood down into the right ventricle. When the right ventricle contracts, blood is forced out of the heart through the right and left pulmonary arteries towards the lungs. Oxygen-depleted blood entering this pulmonary circulation exchanges carbon dioxide for oxygen, and so oxygen-rich blood returns to the heart via the right and left pulmonary veins and into the left atrium. Contraction of the left atrium pushes blood down into the left ventricle, and left ventricular contraction forces blood out of the heart again via the aorta to the systemic circulation. Blood flow through the heart is unidirectional, which is achieved by valves positioned between the chambers and within the vessels. Blood entering the heart will pass the tricuspid valve between the right atrium and right

Aorta (to body) Pulmonary artery

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FIGURE 36.1 Internal structure and blood flow within the human heart. From Diseases and Conditions Index – Heart and Blood Vessel Diseases; How the Heart Works; Anatomy; http://www.nhlbi.nih.gov/health/dci/Diseases/hhw/hhw_anatomy.html; National Heart Lung Blood Institute, National Institutes of Health, US Department of Health and Human Services, 2010. Permission granted.

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ventricle, and then the pulmonary valve when being pumped out of the right ventricle and towards the lungs. When re-entering the heart, blood will pass the mitral valve between the left atrium and left ventricle, and finally the aortic valve when being forced out of the heart through the aorta to the rest of the body. The heart contracts in a repeated, rhythmic manner, yet blood is continuously delivered to tissues and organs due to the innate properties of arterial blood vessels. During ventricular contraction, the aorta and its branches distend and then during ventricular relaxation, the walls of the large arteries recoil elastically, which forces blood through the circulation continually.

the SA and AV nodes and activate them by releasing acetylcholine. This action decreases heart rate (negative chronotropy), force of atrial contraction (negative inotropy), rate of relaxation (negative lusitropy), and conduction velocity of the SA and AV nodes (negative dromotropy). As the parasympathetic nervous system is responsible for regulating functions primarily when the body is at rest, this increased vagal tone results in low heart rates of ~70 beats per minute in humans. As such, the parasympathetic nervous system is said to induce the “rest and digest” state. Sympathetic nervous control of the heart arises from the upper thoracic region of the spinal cord. Short preganglionic efferent nerve fibers, compared to long preganglionic vagal efferent fibers, enter the paravertebral chains of ganglia that are located on either side of the spinal column. These preganglionic fibers synapse with postganglionic sympathetic fibers and release acetylcholine, which binds to nicotinic receptors on the postganglionic fibers. From here, relatively long sympathetic adrenergic efferent fibers extend to the SA and AV nodes in the heart where they release the neurotransmitter norepinephrine at synapses with beta-adrenergic receptors. In contrast to parasympathetic effects, this action increases heart rate (positive chronotropy), force of ventricular contraction (positive inotropy), rate of relaxation (positive lusitropy), and conduction velocity of the SA and AV nodes (positive dromotropy). These responses are indicative of the “fight or flight” state that the sympathetic nervous system is said to elicit, and are capable of increasing heart rate up to ~200 beats per minute in humans.

AUTONOMIC NERVOUS CONTROL The autonomic nervous system can be divided into two sub-divisions, the parasympathetic nervous system and the sympathetic nervous system (see Fig. 36.2). Parasympathetic nervous control of the heart arises from vagal nuclei within the medulla oblongata in the brainstem, and efferent nervous outflow occurs via the tenth cranial nerve, known as the Vagus nerve. These long preganglionic efferent nerve fibers extend down to the heart where they synapse with small ganglia located near the sinoatrial (SA) and atrioventricular (AV) nodes in the heart. At this target organ, the neurotransmitter acetylcholine is released, which binds to nicotinic receptors and activates short postganglionic efferent nerve fibers. These postganglionic fibers synapse with muscarinic receptors in

Parasympathetic (vagus)

Hypothalamus Paravertebral Ganglia

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FIGURE 36.2 Anatomy of the autonomic nervous system and its control of the heart. From Cardiovascular Pharmacology Concepts – Autonomic Ganglia; http://cvpharmacology.com/autonomic_ganglia.htm; Richard E. Klabunde, 2010. Permission granted.

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The actions of the parasympathetic and sympathetic nervous systems are opposed to each other, yet they work reciprocally to bring about the necessary responses to internal and external stimuli. At rest, parasympathetic tone is predominant, which can be illustrated by observing the effects of inhibiting parasympathetic and sympathetic activation [2] (see Fig. 36.3). Atropine is a muscarinic receptor antagonist that inhibits parasympathetic activity and when administered in humans at rest, heart rate increases to ~110 beats per minute. In contrast, when propranolol, which is a beta-adrenergic receptor antagonist that inhibits sympathetic activity, is given at rest, heart rate decreases only slightly to ~50 beats per minute. When both nervous systems are blocked, heart rate remains ~100 beats per minute, which is known as the intrinsic heart rate. When an increase in heart rate from resting levels is necessary, for example during exercise, parasympathetic tone is initially withdrawn and then sympathetic tone is enhanced. Conversely, when heart rate recovers following a bout of exercise or other physiologically stressful event, sympathetic activity is first reduced and then parasympathetic activation is augmented [1].

CONTROL OF THE HEARTBEAT The heart itself is capable of generating its own electrical impulses to cause coordinated and rhythmic contractions of its chambers. The SA node, a specialized group of cardiac myocytes located in the right atrium, generates action potentials that spread throughout the atria. This causes depolarization and consequent contraction of atrial

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muscle. This produces the normal sinus rhythm, thus the SA node is known as the “pacemaker”. These electrical impulses travel from the atria to the ventricles via the AV node, another specialized group of cardiac myocytes located in the posterior inferior region of the interatrial septum. From here, these impulses are transmitted through the bundle of His, specialized cardiac myocytes that conduct electrical activity, at the base of the ventricles along the left and right bundle branches in the interventricular septum down to the apex of the heart. These branches then divide into an extensive network called Purkinje fibers that cover the inner ventricular walls, and these impulses travel throughout this system of fibers causing depolarization and contraction of ventricular muscle. This complex yet highly coordinated sequence results in one cardiac cycle, which lasts for ~0.8 seconds. Within each cycle, there are periods of relaxation, known as diastole, and contraction, known as systole. When the whole heart is in diastole, the atria fill with blood, which then contract during atrial systole and force blood into the ventricles. The ventricles then contract during ventricular systole and eject blood into the aorta and pulmonary arteries while the atria are in diastole. The ventricles then return to diastole and the cycle begins again. In addition to the heart’s intrinsic ability to regulate its electrical activity, autonomic nervous mechanisms can also directly influence the conduction of electrical impulses throughout the heart. This action is primarily mediated via the AV node. Parasympathetic activation releases acetylcholine that binds to muscarinic receptors in the AV node, which causes slower depolarization of cardiac myocytes. This results in negative dromotropy and decreased ventricular contractility. Acetylcholine release from parasympathetic nerve fibers can also inhibit the release of norepinephrine from sympathetic nerve fibers. This neurotransmitter antagonism can also cause reduced ventricular contraction. Sympathetic activation releases norepinephrine that binds to beta-adrenergic receptors in the AV node, which causes faster depolarization of cardiac myocytes. This results in positive dromotropy and increased ventricular contractility, as the time between atrial and ventricular contraction is reduced. It is possible for the conduction velocity of action potentials to be slowed to such an extent that they completely stop, leading to AV block. In the treatment of certain medical conditions, giving drugs to induce AV block can be an effective method of stabilizing heart rate. For example, patients prone to cardiac arrhythmias can be given medication to block beta-adrenergic receptors, known as ‘beta-blockers’ [4]. This reduces the conduction velocity of electrical impulses through the AV node, thereby preventing detrimental changes in heart rhythm.

DRUG # 2

FIGURE 36.3 Effects of atropine (0.04 mg/kg total) and propranolol (0.2 mg/kg total) on heart rate in 10 nonathletes. In half of the trials, atropine was given first (top line) and in the other half, propranolol was given first (bottom line). From Katona et al. (1982) J Appl Physiol 52(6):1652–7. Permission granted.

ALTERED AUTONOMIC CONTROL Autonomic nervous control of the heart can be altered under many different physiological conditions. Exercise

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and stress require acute changes in heart function, which are predominantly brought about by alterations in autonomic nervous control [5]. Reduced parasympathetic and augmented sympathetic nerve activity induce raises in heart rate, contractility and conduction velocity of electrical impulses, which provide the necessary increases in blood delivery to the appropriate tissues and organs that demand a greater supply. Chronic alterations in autonomic nervous control manifest in the process of aging and certain disease states. Aging is associated with heightened cardiac sympathetic and reduced parasympathetic activity, which can potentially augment the detrimental effects of any existing cardiovascular disease [3]. Several cardiovascular disease states, such as heart failure and hypertension, have also been linked to similar increases in sympathetic tone and decreases in parasympathetic activity [6–8]. These alterations can lead to a vicious cycle where the disease state is enhanced due to the autonomic changes that have occurred to compensate for the disease itself. In summary, autonomic control of the heart is integral to the regulation of heart function. It plays a major role in ensuring optimal performance of the heart, both when the body is at rest and when responding to acute or chronic physiological changes.

References [1] Arai Y, Saul JP, Albrecht P, Hartley LH, Lilly LS, Cohen RJ, et al. Modulation of cardiac autonomic activity during and immediately after exercise. Am J Physiol Heart Circ Physiol 1989;25:H132–41. [2] Katona PG, McLean M, Dighton DH, Guz A. Sympathetic and parasympathetic cardiac control in athletes and nonathletes at rest. J Appl Physiol 1982;52(6):1652–7. [3] Kaye DM, Esler MD. Autonomic control of the aging heart. Neuromol Med 2008;10:179–86. [4] Kennedy HL, Brooks MM, Barker AH, Bergstrand R, Huther ML, Beanlands DS, et al. Beta-blocker therapy in the cardiac arrhythmia suppression trial. Am J Cardiol 1994;74:674–80. [5] Robinson BF, Epstein SE, Beiser GD, Braunwald E. Control of heart rate by the autonomic nervous system: studies in man on the interrelation between baroreceptor mechanisms and exercise. Circ Res 1966;19:400–11. [6] Schlaich MP, Lambert E, Kaye DM, Krozowski Z, Campbell DJ, Lambert G, et al. Sympathetic augmentation in hypertension: role of nerve firing, norepinephrine reuptake, and angiotensin neuromodulation. Hypertension 2004;43:169–75. [7] Schwartz PJ, de Ferrari GM. Sympathetic-parasympathetic interaction in health and disease: abnormalities and relevance in heart failure. Heart Fail Rev 2011;16:107–7. [8] Thayer JF, Yamamoto SS, Brosschot JF. The relationship of autonomic imbalance, heart rate variability and cardiovascular disease risk factors. Int J Cardiol 2010;141:122–31.

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37 Cardiac Vagal Ganglia Javier G. Castillo, David H. Adams THE VERTEBRATE NERVOUS SYSTEM

THE CARDIAC CONDUCTION SYSTEM

The vertebrate nervous system is divided into two major components: the central nervous system, consisting of the spinal cord and the brain, and the peripheral nervous system, which can be in turn subdivided into somatic and autonomic components [1]. The autonomic component of the peripheral system regulates the function of the internal organs by way of efferent nerve fibers originating from the central system [2]. In this regard, sympathetic and parasympathetic efferent nerves generate complementary motor outputs in response to sensorial impulses directed to the brain from internal organs via either mechanical or chemical receptive afferent neurons. It is important to highlight that both branches of the autonomic system use pre and postganglionic relays. Sympathetic efferent nerves containing preganglionic neurons arise segmentally from the central portion of the spine, and terminate in sympathetic ganglia that divide into bilateral chains that run parallel to the spinal cord (Fig. 37.1). In this area, they synapse with postganglionic sympathetic neurons that travel to the effector organ. In the case of the heart, these neurons are able to provide chronotropic and inotropic stimulation. The main neurotransmitters of the sympathetic nervous system are norepinephrine and acetylcholine. Parasympathetic preganglionic efferent nerves arise mainly in the brainstem and also from the spinal cord and synapse with postganglionic neurons located in the heart [3]. Acetylcholine is the main neurotransmitter, which contributes to reduction of heart rate and contractility. In addition, sensory afferent pathways with sympathetic and parasympathetic components relay information from the heart to the brain. While cardiac parasympathetic afferent pathways control sympathetic and parasympathetic output, the sympathetic afferent pathways do not have an impact on the autonomic output of the heart. These parasympathetic afferent pathways connect to efferent pathways via interneurons located within the brainstem. Sympathetic afferent nerves travel through the stellate ganglia of the sympathetic chain, and connect with motor neurons in the spinal cord.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00037-8

The cardiac conduction system initiates, conducts, and controls the heart beat. It is integrated by the sinus (SA) node, the atrioventricular (AV) node, penetrating bundle, and ventricular bundle branches [4]. The sinus node is a wedge shaped structure situated at the junction of the superior vena cava with the terminal crest musculature, usually arranged about a central artery. Its cells are fascicular in nature and frequently embedded in a very fibrotic matrix. Acetylcholine positive nerves represent the main cellular subpopulation identified in the SA. The AV node is located within the atrial aspect of the atrioventricular septum, separated from the ventricle by the annulus fibrosus. The node consists of an elongated half oval with a superficial transitional layer arranged circumferentially. This transitional layer crosses through the node and terminates in the base of the tricuspid valve, while additional transitional fibers also enter the node from the floor of the coronary sinus. These fibers are arranged in parallel structures separated by fibrous tissue. The main distinguishing feature between the distal end of the AV node and the proximal penetrating bundle is the penetration of the conduction axis into the annulus fibrosus. When these fibers advance towards the ventricles, the cells reorient in a more parallel fashion and remain small in size. After reaching the central fibrous body and then the left ventricular outflow tract, the bundle subdivides into right and left bundle branches. The left bundle branch ends up descending as a sheet of cells within the septal subendocardium, whereas the right bundle branch descends intramyocardially (Fig. 37.2).

ANATOMY OF THE CARDIAC VAGAL GANGLIA The anatomy of the cardiac vagal ganglia has been consistently studied for over a century [5]. Intrinsic cardiac neurons have been reported to be located in small ganglia organized primarily over the posterior surfaces of the atria, particularly over the atrioventricular groove [6]. Only a few neurons have been associated with ventricular tissues. Recent reports have identified the presence

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Brainstem

Atrial Ganglionated Plexuses There are five major atrial ganglionated plexuses in the human heart, all of them named based on their anatomical location:

Craneal Cervical Ganglia

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1. The superior right atrial ganglionated plexus is located on the posterior surface of the right atrium adjacent to the junction of the superior vena cava and the right atrium. 2. The superior left atrial ganglionated plexus is a collection of ganglia identified on the posterior surface of the left atrium between the pulmonary veins. It is in this area where an important number of mediastinal nerves synapse. 3. The posterior right atrial ganglionated plexus lies over the posterior surface of the right atrium adjacent to the interatrial groove. 4. The posteromedial left atrial ganglionated plexus courses on the posterior medial surface of the left atrium. The subgroup of ganglia formed after the fusion of the two posterior atrial ganglionated plexuses extending anteriorly into the interatrial septum is named interatrial septal ganglionated plexus. The largest concentration of ganglia is associated with the two major ganglionated plexuses on the posterior surface of the two atria. 5. The posterolateral left atrial ganglionated plexus is the smallest among the atrial plexuses and can be identified on the posterior lateral surface of the left atrial base on the atrial side of the atrioventricular groove (Fig. 37.3).

FIGURE 37.1 The heart is innervated by preganglionic sympathetic and parasympathetic efferent nerves that originate from the spinal cord and dorsal motor nucleus of the brainstem respectively. Preganglionic (dashed) to postganglionic (solid) sympathetic transfer occurs in the sympathetic ganglion chain adjacent to the spinal cord. Postganglionic sympathetic neurons travel to the cardiac plexus, where preganglionic parasympathetic neurons synapse to postganglionic elements. From the cardiac plexus, autonomic nerves can synapse with intrinsic cardiac neurons located within the network of cardiac ganglia.

Ventricular Ganglionated Plexuses

of stained (1% solution of methylene blue) ganglia and nerves mostly in fatty areas, and only 10% of them are located adjacent to underlying muscle or between muscle fascicles [7]. Interestingly, no ganglia are usually identified in the bulk of the fat that accompanies the major coronary arteries in the atrioventricular groove. The human heart comprises an average of 450 ganglia in atrial tissues and approximately 90 ganglia in ventricular tissues. The precise anatomical configuration of every ganglionated plexus varies among individuals as does the size of ganglia within a particular region. Nerves (up to 0.2 mm in diameter) course between regional ganglia forming neural networks. In addition, interconnecting nerves in ganglionated plexuses form complete loops, which range from less than 2 mm to 1 cm in diameter.

1. The major ventricular ganglionated plexus is totally surrounded by the aortic root fat and therefore named as aortic root ganglionated plexus. This plexus is intimately related to the nerves coursing along the coronary arteries, with its left component projected towards the origin of the circumflex coronary artery. 2. At the same time, this latter component of the aortic root plexus is connected to a plexus located at the origin of the anterior descending coronary artery. Consequently, this plexus has been named anterior descending ganglionated plexus. 3. One of the smallest plexuses follows the trajectory of the posterior descending coronary artery thus called posterior descending ganglionated plexus. The last two plexuses are located adjacent to the right acute marginal

The five ventricular ganglionated plexuses may be found in the fatty areas of both ventricular bases, particularly on the superior aspects of the ventricles, the interventricular grooves, and the origins of the marginal arteries. Additionally, occasional neurons can be identified outside these areas, mostly in fat associated with branching segments of larger coronary arteries.

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FIGURE 37.2 View of the human heart illustrating the conduction system. The table shows the patterns of innervation by immunofluorescence according to developing stages. DBH: Dopamine β-hydroxylase (sympathetic activity); PGP: Protein gene product (presence of nerve fibers); TH: Tyrosine hydroxylase (sympathetic activity); –No detected nerve fibers; Scattered individual nerve fibers; Moderate number of nerve fibers, Large number of nerve fibers.

coronary artery and the left obtuse marginal coronary artery and their names are: 4. Right acute marginal ganglionated plexus; and 5. Obtuse marginal ganglionated plexus. Finally, scattered ungrouped neurons can be found embedded in fatty areas on the ventricular side of the atrioventricular groove. However, these neurons are not consistently clustered in a determined region (Fig. 37.3).

THE AUTONOMIC NERVOUS SYSTEM The integrated control of the circulatory system results from intrinsic (e.g., endothelium) and extrinsic mechanisms (e.g., autonomic nervous system). In this regard, the

autonomic nervous system mostly affects the vasomotor tone and cardiac function through the sympathetic and parasympathetic divisions. Moreover, it also has an impact on the systemic volume and the peripheral resistance by modulating the release of certain peptide hormones such as angiotensin II (AGII) or nitric oxide (NO) [8]. In fact, this neuronal control involves an accurate acquisition of inputs from the cerebral cortex and individualized sensors, the integration into particular brain regions, and the transmission of efferent activity to the heart over the sympathetic and parasympathetic pathways. Three anatomical areas of the brain have been identified as an integrated functional system that coordinates the vasomotor status of the cardiovascular system. These are: 1. The upper anterolateral medulla (vasoconstrictor).

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37. CARdIAC VAGAl GAnGlIA

2

1

5

3

7

4

6

<100 Neurons

100-200 Neurons

>200 Neurons

Ganglia per Heart

Superior right atrial (1)

+++

+

-

31 ± 5

Superior left atrial (2)

+++

++

+

56 ± 12

Posterior right atrial (3)

+++

++

++

194 ± 22

Posteromedial left atrial (4)

+++

++

+

161 ± 27

Posterolateral left atrial (5)

+++

+

-

16 ± 2

Aortic root

++

-

-

16 ± 2

Anterior descending

++

+

-

11.2 ± 1.1

Posterior descending (6)

+

-

-

5.2 ± 1.9

Right acute marginal

+

-

-

6.2 ± 2.8

Ganglionic Plexus Atrial Ganglionated Plexuses

Ventricular Ganglionated Plexuses

FIGURE 37.3 SuperiorObtuse view of the human of ganglionated plexuses on the surface of the atria and ventricles. The marginal (7) heart illustrating the distribution 5.2 ± 2.0 + table shows the number of neurons and ganglia in these regions. –None; Scattered; Moderate; Abundant.

2. The lower anterolateral medulla (vasodilator). 3. The nucleus tractus solitarii which integrates vasoconstrictor and vasodilator stimuli. The cerebral regions that modify the chronotropic activity of the heart are located in the thalamus, posterior and posterolateral regions of the hypothalamus, and dorsal region of the medulla. Variations of the sympathetic activity exert a very powerful control over the peripheral circulation. These efferent nerve fibers course either in specific sympathetic pathways to innervate the heart or join the paravertebral sympathetic chain and synapse in secondary

ganglia forming spinal nerves that innervate peripheral vessels. These vascular nerves terminate on small arteries, arterioles, venules, and veins and adjust vascular resistance and volume capacity and therefore the heart pacing. Cardiac nerves, which mainly arise from the stellate ganglia, innervate the atria and ventricle. Reflex sympathetic stimulation causes vasoconstriction by releasing norepinephrine (NE) from sympathetic nerve endings and epinephrine from the adrenal medulla, which stimulates α and β receptors. In this case, epinephrine provides a dual pattern of action, produces vasodilation and cardiac stimulation at low concentrations, and

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THE AuTonomIC nERVous sysTEm

vasoconstriction at higher concentrations. This is a very important concept since reflex sympathetic stimulation increases cardiac output during metabolic stress, maintaining systemic pressures and heart perfusion [8]. On the other hand, although the parasympathetic nervous system plays a very insignificant role in arterial pressure regulation, it plays a crucial role in modulating the heart rate. The parasympathetic fibers of the vagus nerve provide innervation to the cardiac conduction system including the SA and AV nodes as well as the atrial myocardium, thus providing chronotropic control. When the vagus nerve is stimulated, acetylcholine reduces heart rate and contractility.

References [1] Van Stee EW. Autonomic innervation of the heart. Environ Health Perspect Oct 1978;26:151–8.

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[2] Armour JA, Murphy DA, Yuan BX, Macdonald S, Hopkins DA. Gross and microscopic anatomy of the human intrinsic cardiac nervous system. Anat Rec Feb 1997;247(2):289–98. [3] Martins J. Parasympathetic regulation of the heart. Heart Rhythm Aug 2010;7(8):1120–1. [4] Chow LT, Chow SS, Anderson RH, Gosling JA. Innervation of the human cardiac conduction system at birth. Br Heart J May 1993;69(5):430–5. [5] Chow LT, Chow SS, Anderson RH, Gosling JA. Autonomic innervation of the human cardiac conduction system: changes from infancy to senility--an immunohistochemical and histochemical analysis. Anat Rec Oct 1 2001;264(2):169–82. [6] Hildreth V, Anderson RH, Henderson DJ. Autonomic innervation of the developing heart: origins and function. Clin Anat Jan 2009;22(1):36–46. [7] Crick SJ, Wharton J, Sheppard MN, et al. Innervation of the human cardiac conduction system. A quantitative immunohistochemical and histochemical study. Circulation Apr 1994;89(4):1697–708. [8] McGrath MF, de Bold ML, de Bold AJ. The endocrine function of the heart. Trends Endocrinol Metab Dec 2005;16(10):469–77.

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38 Neural Control of Blood Vessels Julian H. Lombard, Allen W. Cowley, Jr. AUTONOMIC VASOMOTOR CONTROL AND CARDIOVASCULAR HOMEOSTASIS The sympathetic and parasympathetic components of the autonomic nervous system play a crucial role in maintaining cardiovascular homeostasis and enabling the body to respond to physiological stressors. Neurogenic control, especially on the arterial side of the circulation, is generally superimposed on intrinsic tone of the vessels, although neurogenic vasoconstriction also occurs in vessels possessing little or no intrinsic tone, e.g. large veins. The effects of the autonomic nervous system on the cardiovascular system are coordinated by a combination of centrally driven autonomic nerve activity that is modulated by cardiovascular receptors (mainly the arterial baroreceptors). Neurogenic mechanisms are not only essential to maintain and regulate arterial blood pressure, but also play a crucial role in regulating the distribution of blood flow between and within individual vascular beds. As discussed below, the net effect of adrenergic constrictor mechanisms on vascular resistance and blood flow within specific vascular beds is determined not only by the frequency of sympathetic nerve traffic, but also by the modulating influences of multiple vasoactive stimuli such as hormones, autacoids, and local autoregulatory mechanisms. Although autonomic neurogenic mechanisms predominantly mediate vasoconstriction via noradrenergic nerve terminals, neurogenic vasodilation does occur in some vascular beds. Two of the primary functions of neurogenic vasodilation are in erectile tissue of the genetalia (mediated by parasympathetic nerves) and sympathetic cholinergic vasodilation of skeletal muscle arterioles in some species, although likely not in man (see below). A tonic level of sympathetically-mediated vasoconstriction in larger vascular beds, e.g., skeletal muscle and the splanchnic circulation, is essential to prevent catastrophic drops in arterial blood pressure and to regulate blood flow to match tissue needs. Due to the large volume of blood (approximately 70% of the total blood volume) on the venous side of the circulation, neurogenic regulation of the capacity of the venous side of the circulation by the sympathetic nervous system (SNS) plays a crucial role in cardiovascular homeostasis via regulation of cardiac output and ventricular filling pressure.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00038-X

Sympathetically mediated vasoconstriction can decrease flow in the splanchnic circulation to ~25% of resting values. About half the blood volume in the splanchnic circulation (~15% of total blood volume) can be mobilized by passive collapse of the veins and by active venoconstriction at higher levels of SNS activation, e.g., those occurring during conditions of hemorrhagic stress. Under those conditions, sympathetically mediated constriction of precapillary arterioles can also reduce capillary pressure sufficiently to absorb interstitial fluid into the circulation when capillary pressure falls below plasma colloid osmotic pressure. In contrast to sympathetic nervous effects, parasympathetic nervous activity dilates splanchnic vessels indirectly by stimulating intestinal motility and glandular secretion, which increases intestinal metabolism. The cutaneous vascular bed is another important vascular bed for recruiting blood volume in response to increases in sympathetic nerve activity. Another important role for sympathetic regulation of cutaneous blood vessel function is in thermoregulation. In this case, vasoconstriction to restrict heat loss and vasodilatation to promote heat loss play crucial roles in regulating body temperature. Finally, the extracranial arteries in the head receive both sympathetic innervation via the superior cervical ganglion, with norepinephrine (NE) and neuropeptide Y (NPY) as transmitters or parasympathetic innervation via the sphenopalatine and otic ganglia, with vasoactive intestinal peptide (VIP), acetylcholine and nitric oxide as neurotransmitters. The primary function of the sympathetic innervation of the cerebral arteries is to shift the upper limit of cerebral autoregulation to higher pressures. By contrast, the parasympathetic nerves, which are potent dilators of the cerebral circulation, do not appear to play a significant role in regulating cerebral blood flow, although they may be involved in migraine headaches. In addition, sensory input from the trigeminovascular pathway, with calcitonin gene related peptide (CGRP) as the transmitter, may act as a protective mechanism to restore cerebral vessel tone after contractile stimuli. The latter pathway may also have a role in migraine headache. After entering the brain parenchyma, cerebral arteries lose their sympathetic and parasympathetic innervation. Within the parenchyma itself, changes in cerebral blood flow occurring in response to neuronal activity are effected

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38. NEuRAl CoNTRol of Blood VEssEls

Cardiac

Sympathetic Nervous System

Inotropic Effect (+) Chronotropic Effect (+)

Arteries Veins Microvessels

Targets

Norepinephrine (NE) Vascular

Transmitters

ATP Neuropeptide Y (NPY) Alpha 1 (constriction) Adrenergic (NE) Alpha 2 (constriction/dilation)

Receptors

Beta 1 and beta 2 (vasodilatation) P2 Purinergic (ATP)-constriction Y1 (NPY)-Constriction

FIGURE 38.1 Learning map summarizing cardiovascular control by the sympathetic nervous system.

via multiple mediators coordinated by the neurovascular unit, consisting of neurons, astrocytes, and the target blood vessels.

SYMPATHETIC COMPONENT OF AUTONOMIC VASOMOTOR CONTROL The sympathetic component of the autonomic nervous system plays the predominant role in regulating vascular tone and whole-body hemodynamics (see Fig. 38.1). Activation of the sympathetic nervous system usually elicits a vasoconstriction that is mediated by three main neurotransmitters (norepinephrine, ATP, and neuropeptide Y), and is roughly proportional to the level of neural activity. Stimulation of sympathetic nerve fibers in skeletal muscle can also cause a transient dilation of larger arterioles that is blocked by atropine (sympathetic cholinergic vasodilation) in some species. However, this does not appear to be the case in man, where vasodilation in anticipation of stresses such as exercise appears to be due to withdrawal of sympathetic tone and vascular relaxation mediated by β2 adrenergic receptors.

Neuroeffector Junction Figure 38.2 shows a schematic diagram of the vascular neuroeffector apparatus; and Figure 38.3 shows the dense adrenergic innervation of a small artery and the less dense adrenergic innervation of a small vein in the rat mesentery. The postganglionic autonomic nerves of the sympathetic nervous system that innervate blood vessels ramify into small bundles, which form a primary plexus located in the adventitial layer of the vessel. The terminal effector plexus is located near the medial layer where adrenergic nerve fibers approach the surface of the smooth muscle

cells and establish neuromuscular contact. These nerves end in strings of varicosities that are devoid of Schwann cell sheaths, and which release transmitter in response to action potentials in the nerves. It is now recognized that many varicosities form en passant synapses with their targets. Compared to the narrow synaptic clefts in the central nervous system, the synapses in peripheral neuroeffector units are much wider, often no closer than 100 nm from the vascular smooth muscle cells.

Sympathetic Neurotransmitters In the varicosities at the neuroeffector junction, sympathetic neurotransmitters are stored in either small or large dense-cored granular vesicles, which also contain ATP, enzymes involved in norepinephrine synthesis such as dopamine β-hydroxylase, and proteins such as chromogranin. There are three major sympathetic co-transmitters – norepinephrine, ATP and neuropeptide Y (NPY), all of which contribute to sympathetic vasoconstriction, although the relative contribution of each varies with among vascular beds and with the amount of sympathetic nerve traffic. Norepinephrine (NE) is the classical neurotransmitter released from adrenergic nerve terminals during sympathetic discharge and produces vasoconstriction by activating α1-adrenergic receptors located on vascular smooth muscle cells. Norepinephrine is found in the small dense core vesicles and seems to be primarily responsible for the moderately fast phase of adrenergic vasoconstriction. ATP is released together with norepinephrine from sympathetic nerves and produces a rapid contraction of vascular smooth muscle by activating P2 purinergic receptors, which include ligand-gated ion channels and G-protein coupled receptors. Neuropeptide Y (NPY) is also found in sympathetic nerve fibers, most likely co-localized with norepinephrine

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189

Primary adventitial plexus Schwann nucleus Vasa vasorum Adventitio-medial junction

Adventitia

Terminal effector plexus

Media

Terminal effector plexus in outermost muscle lamellae

FIGURE 38.2 Arrangement of vascular neuroeffector apparatus. Postganglionic autonomic nerves ramify into small bundles forming a primary plexus, which is located in loose adventitia. Bundles give rise to varicosed fibers forming terminal effector plexus, located on surface of medial layer. (Reprinted with permission from Verity MA. Morphologic studies of the vascular neuroeffector apparatus. In: Physiology and Pharmacology of Vascular Neuroeffector Systems, JA Bevan, RF Furchgott, RA Maxwell and AA Somlyo, eds. Basel: Karger, 1971, pp. 2–12.)

compound may play a more important role with circulatory stress, as it appears to be released at the moderate to high levels of sympathetic discharge that occur with circulatory stress. NPY causes a slowly developing and persistent phase of constriction via Y1 receptors, and also potentiates vasoconstriction in response to norepinephrine.

Neurotransmitter Release and Effector Action

FIGURE 38.3 Fluorescence micrograph of a whole mount of rat mesentery showing perivascular sympathetic nerves demonstrated by Falck– Hillarp formaldehyde technique. A very dense plexus of fluorescent noradrenergic fibers supplies a small artery, whereas the corresponding vein has less well developed innervation, although still prominent. Nerve terminals are seen to accompany small vessels of arteriolar caliber. 155. (Reprinted with permission from Falck B. Observations on the possibilities of cellular localization of monoamines by a fluorescence method. Acta Physiol Scand (Suppl.) 1962;197:1–25.)

in the large dense-core vesicles. NPY is released along with norepinephrine and ATP at moderate to intense levels of sympathetic activation. The role of NPY in normal regulation of blood pressure is uncertain, but this

NE released from the adrenergic nerves binds to receptors on the VSM cells acting primarily on α1-adrenergic receptors, which cause contraction of the vascular smooth muscle via membrane depolarization with extracellular Ca2 influx and by liberation of Ca2 ions from intracellular stores via the phospholipase C/IP3 mechanism. Because not all the VSM cells are in close proximity to the adrenergic nerve varicosities, gap junctions between adjacent smooth muscle cells provide low resistance pathways, enabling electrical coupling between the cells. This allows smooth muscle cells that are not adjacent to varicosities to be activated and, in the microcirculation, allows excitation to be propagated longitudinally along the arteriole. Norepinephrine is removed from the junctional cleft primarily by active reuptake by the neurons, enzymatic degradation by catechol-O-methyl transferase, and by spillage into the circulation.

Parasympathetic Component of Autonomic Vasomotor Control The cranial and sacral nerves of the parasympathetic nervous system also regulate vascular tone, although much less is known regarding their functional roles apart

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from their role of increasing blood flow during engorgement of erectile tissue in the genitalia. It is generally accepted that cholinergic nerves contain predominantly small agranular vesicles (35–60 nm), although large granular vesicles with a dense core are often present as well. Cholinergic vasodilatation appears to be mediated via the combined action of acetylcholine, vasoactive intestinal peptide (VIP) and nitric oxide (NO). Despite its high endogenous concentration in some blood vessels, e.g. cerebral arteries, acetylcholine appears to exhibit a negligible direct effect on vascular smooth muscle tone, most likely due to a combination of a wide synaptic distance and a low synaptic concentration as a result of the activity of acetylcholinesterase. However, NO synthase and choline acetyltransferase coexist in parasympathetic ganglia and in the perivascular nerves innervating cerebral blood vessels of several species. Acetylcholine and NO are coreleased from the same nerves, and there is also evidence that acetylcholine acts as a presynaptic transmitter to modulate NO release. Taken together, those findings suggest that nitric oxide (NO) mediates the major component of parasympathetic neurogenic vasodilation in cerebral arteries. In addition to acetylcholine, parasympathetic neurons also contain vasoactive intestinal peptide (VIP) which, together with NO, contributes to vasodilation in some vascular beds, particularly the larger arteries and veins in the head and pelvis. Vasodilator neurons that synthesize VIP also contain structurally unrelated peptides, including NPY, dynorphins, enkephalin, galanin, somatostatin and calcitonin gene-related peptide (CGRP), although little is known regarding their physiological roles. Despite the existence of cholinergic vasodilation in some vascular beds, the overall contribution of the parasympathetic nervous system to the regulation of vascular tone and hemodynamics is small. Rather, the primary regulatory role of the parasympathetic nervous system in the cardiovascular system is mediated via its negative chronotropic and inotropic effects on the heart (see Fig. 38.4).

Differential Vasomotor Control Except for fight or flight responses, the sympathetic nervous system does not usually produce uniform effects on all target organs, Different postganglionic sympathetic neurons have distinct properties, and release other transmitters in addition to NE. This specific distribution of neuroactive chemicals among neurons is called chemical coding. For example, depolarization of postganglionic sympathetic neurons in the lumbar sympathetic ganglia of the guinea pig causes a brief burst of action potentials in nearly all the neurons, leading to release of ATP and NPY to cause arterial vasoconstriction; while depolarizaton of sympathetic preganglionic neurons in the inferior mesenteric ganglion causes sustained firing in about 80% of the neurons, leading to release of both NE and somatostatin, which regulates gut motility and secretion. Sympathetic neurons act in a discrete and organspecific manner, allowing blood flow in individual vascular beds to be regulated independently, depending on physiological conditions. This differential regulation involves discrete neurons in the pre-vertebral ganglia that exhibit specific electrophysiological, neurochemical, and morphological phenotypes. The final neurons in the sympathetic vasomotor pathways are small and receive fewer preganglionic inputs than the non-vasomotor neurons of the autonomic nervous system. Specific functional pools of postganglionic neurons receive convergent input from different pools of preganglionic neurons, many of which contain neuropeptides that affect the excitability of the neurons. Pools of sympathetic neurons project to specific segments of the vasculature, allowing for selective regulation of regional resistance in the proximal and distal portions of the vascular bed. In addition, the functional pools of vasomotor neurons can contain characteristic combinations of co-transmitters. Thus, vasomotor neuron pools appear to be grouped into functional pools and can be recruited as necessary to provide highly graded and specific flow within and between the vascular beds.

Heart (Major)

Parasympathetic Nervous System

Chronotropic Effect (–) Intropic Effect (–)

Vasodilator Action Acetylcholine (ACh) Vascular (Minor)

Transmitters

Nitric Oxide (NO) Vasoactive Intestinal Peptide (VIP) Muscarinic Receptors (ACh)

Receptors

Guanylyl cyclase (NO) VIP (VIP)

FIGURE 38.4 Learning map summarizing cardiovascular control by the parasympathetic nervous system.

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symPATHETIC ComPoNENT of AuToNomIC VAsomoToR CoNTRol

Modulation of Adrenergic Vasoconstriction There are significant regional variations in the responsiveness of blood vessels to sympathetic activity as a result of a variety of factors, including: (i) the density of α-adrenergic innervation; (ii) vascular smooth muscle sensitivity to norepinephrine; (iii) differences in α-adrenergic receptor populations between organs; (iv) differences in neuronal reuptake of norepinephrine; and (v), the structure and size of blood vessels, which affects the access of norepinephrine to the receptors, i.e. in small vessels with small junctional clefts, the actions of released norepinephrine can be more localized. An important factor in determining the ultimate effect sympathetic activity on blood flow in individual vascular beds is the modulating influence of factors that can override the vasoconstrictor effects of adrenergic activation. For example local autoregulatory mechanisms cause vasodilation in response to increases in metabolic activity particularly in the skeletal muscle, cerebral, and coronary circulations. Increases in blood flow velocity in response to shear stress on the endothelium of arterioles and small resistance arteries can also cause vasodilation by releasing nitric oxide and potentially other vasodilator compounds such as epoxygenase metabolites of the cytochrome P450 pathway of arachidonic acid metabolism. In addition, circulating hormones can either potentiate (e.g., angiotensin II) or inhibit (e.g., atrial natriuretic peptide) adrenergic vasoconstriction. Finally, the release of endogenous vasoactive substances (autacoids) such as nitric oxide, eicosanoids, histamine, kinins, adenine nucleotides, and locally produced vasodilator metabolites all can counteract sympathetic vasoconstriction and contribute to regional modulation of vascular sympathetic responses.

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Thus, the influence of the sympathetic nervous system on regional vascular beds may be affected by any of these factors, which must be carefully considered when evaluating the contribution of sympathetic nerves to the regulation of vascular tone and regional blood flow within a specific vascular bed.

Further Reading Boron WF, Boulpaep EL. Medical physiology, 2nd ed. Philadelphia: Saunders/Elsevier; 2009. Falck B. Observations on the possibilities of cellular localization of monoamines by a fluorescence method. Acta Physiol Scand 1962;197(Suppl):1–25. Franchini KG, Cowley AW. Neurogenic control of blood vessels. In: Robertson D, editor. Primer of the autonomic nervous system (2nd ed). San Diego: Elsevier Science USA; 2004. p. 139–43. Gibbins IL, Jobling P, Morris JL. Functional organization of peripheral vasomotor pathways. Acta Physiol Scand 2003;177:237–45. Hamel E. Perivascular nerves and the regulation of cerebrovascular tone. J Appl Physiol 2006;100:1059–64. Hodges GJ, Jackson DN, Mattar L, Johnson JM, Shoemaker JK. Neuropeptide Y and neurovascular control in skeletal muscle and skin. Am J Physiol Regul Integr Comp Physiol 2009;297:R546–55. Lee TJ. Nitric oxide and the cerebral vascular function. J Biomed Sci 2000;7:16–26. Joyner MJ, Halliwill JR. Sympathetic vasodilatation in human limbs. J. Physiol 2000;526(3):471–80. Koehler RC, Gebremedhin D, Harder DR. Role of astrocytes in cerebrovascular regulation. J Appl Physiol 2005;100:307–17. Verity MA. Morphologic studies of the vascular neuroeffector apparatus. In: Bevan JA, Furchgott RF, Maxwell RA, Somlyo AA, editors. Physiology and pharmacology of vascular neuroeffector systems. Basel: Karger; 1971. p. 2–12. Watts SW, Kanagy NL, Lombard JH. Receptor-Mediated Events in the Microcirculation. In: Tuma RL, Duran WN, Ley K, editors. Handbook of physiology-microcirculation. San Diego: Academic Press/Elsevier; 2008. p. 285–348.

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C H A P T E R

39 Physiology of Upright Posture Wouter Wieling, Jan T. Groothuis INTRODUCTION Orthostatic stresses are common daily events for humans. They result in a shift of blood away from the chest to the distensible venous capacitance system below the diaphragm. Venous pooling is the term commonly used to describe this process. Such pooling in dependent parts results in a rapid diminution of central blood volume, i.e. the volume of blood directly available to the cardiac ventricles, which is of paramount importance for the beat-to-beat adjustment of blood pressure. Unless compensatory adjustments are promptly instituted, blood pressure falls and the subject faints within minutes. This chapter discusses these adjustments. It is concerned with a general description of available regulatory mechanisms. For details of the difference between the initial (first 30 s) cardiovascular adjustments between active and passive changes of posture, refer to Wieling et al., 2007.

REGULATORY MECHANISMS INVOLVED IN THE ADJUSTMENTS OF THE HUMAN BODY TO ORTHOSTATIC STRESS Orthostatic pooling of blood begins almost immediately upon the change from the supine to the upright posture and is estimated to total 300–800 mL; the bulk of the total change occurs within the first 5–10 s (Fig. 39.1). In humans

FIGURE 39.1 Influence of gravity on intravascular fluid shift.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00039-1

orthostatic adjustments are provided by an effective set of blood pressure regulatory mechanisms. They maintain blood pressure at an appropriate level for perfusion of the vital organs, even for the brain, which is located above the heart. To achieve this, the regulatory systems increase heart rate, cardiac contractility and vascular tone to stabilize blood pressure at the level of the heart and brain. The initial reflex adjustments to orthostatic stress are mediated exclusively by neural regulatory systems. During prolonged orthostatic stress additional reflex activation of humoral regulatory systems takes place.

Arterial Baroreceptors The main sensory receptors involved in orthostatic cardiovascular reflex adjustment are the arterial baroreceptors located in the carotid sinuses and aortic arch and mechanoreceptors located in the heart and lungs (Fig. 39.2). The latter consist of a variety of stretch receptors located within the heart and the lungs. “Cardiopulmonary receptors” can be considered to function as rapid acting volume receptors, ideally suited to detect changes in the filling of the central venous circulation. Cardiopulmonary receptors act in concert with arterial baroreceptors to effect the necessary adjustments, but are not essential for the orthostatic cardiovascular adjustments. The arterial baroreceptors tonically inhibit the vasomotor centers in the brainstem. A decrease in blood pressure, as occurs on the assumption of the upright posture, removes this tonic inhibition with a resultant decrease in vagal outflow and an increase in sympathetic activity causing an increase in heart rate, cardiac contractility and vasomotor tone (Figs 39.1 and 39.2). Arterial baroreflex adjustments to orthostatic stress are fast. Decreased vagal outflow increases heart rate within one or two heartbeats. Increased sympathetic activity needs 1–3 seconds to increase heart rate, cardiac contractility and vasomotor tone. The sympathetic mediated increase in vasomotor tone is the key factor in the maintenance of blood pressure in the upright posture. Pronounced increases in heart rate are insufficient to maintain cardiac output, since the heart cannot pump blood that it does not receive. To examine the relative role of the carotid and aortic mechanoreceptors, the following points are important. First, the observation that carotid sinus receptors respond

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39. PHysIology of UPRIgHT PosTURE

deafferentation of the carotid baroreceptors blood pressure control in the stabilized upright posture is impaired. Remaining aortic receptors, however, limit the fall in blood pressure to 10–20 mmHg. These data seem to indicate that the carotid baroreceptors by their location are most important for orthostatic reflex adjustments and for defending the constancy of perfusion pressure of the brain during prolonged orthostatic stress. Additional data support this view. Functional abnormalities can be detected in carotid baroreflex function by neck suction in apparently healthy individuals with unexplained syncope. In addition, the decrease in orthostatic tolerance observed after spaceflight or on mobilization after prolonged bed rest is related to depressed efficiency of the carotid sinus baroreflex. Moreover, in the elderly with an excessive fall in blood pressure in the upright posture afferent baroreflex dysfunction is reported and much overlap is present between the carotid sinus syndrome and orthostatic hypotension.

Local Vasoconstrictor Mechanisms

FIGURE 39.2 Schematic drawing of the afferent and efferent pathways of the arterial baroreceptor reflex arc. Nerve fibers from the carotid and aortic join the glossopharyngeal nerve and vagus nerve respectively toward the vasomotor centre (VMC) in the brainstem. Nerve fibers from the lungs and the heart (not shown) join the vagus nerve as cardiopulmonary afferents.

more vigorously to rapid than to slow changes in pressure makes it likely that they play the major role in the initial reflex adjustments. The abnormally large initial fall and a delayed recovery of blood pressure observed in a patient with bilateral denervation of the carotid baroreceptors supports this view. However, the results of the reverse experiment, i.e. selective denervation of aortic receptors, are not available in humans. Second, neural adjustments in the stabilized fully upright posture result in an increase in diastolic blood pressure with little change in systolic blood pressure at heart level resulting in an increase in mean arterial blood pressure of about 5–10 mmHg. The aortic receptors, which are located just above heart level, sense a reduced pulse pressure, but an increased instead of a decreased mean blood pressure in the upright posture. The blood pressure sensed by the carotid baroreceptors, in contrast, drops and remains below the recumbent level since they are located about 20–25 cm above heart level. This hydrostatic effect lowers the effective pressure at the carotid baroreceptor by about 15 mmHg. A permanent state of diminution in the stimulation of the carotid baroreceptors by the fall in both mean arterial blood pressure and pulse pressure must therefore persist as long as the upright posture is maintained. Third, after surgical

Central modulation of vasomotor outflow is reinforced by local vasoconstrictor mechanisms, such as the veno-arteriolar axon reflex and a myogenic response. The veno-arteriolar axon reflex is triggered when venous pressure exceeds 25 mmHg, which results in vasoconstriction of the corresponding arteriole and is reported to elicit up to 30–45% of the total vasoconstriction in the legs in the upright posture. The myogenic response of the smooth muscle of resistance vessels in the dependent parts is triggered by an increase in transmural pressure across an arteriole. Recent studies suggest that the myogenic response can increase leg vascular resistance up to 30%. It seems to be the most important vasoconstrictor mechanism during orthostatic stress in autonomic failure.

Role of Capacitance Vessels to Orthostatic Reflex Adjustments Capacitance vessels also contribute to reducing the gravitational shift of blood. Reflex venoconstriction in the lower limbs appears of little importance. The cutaneous veins are richly innervated, but venoconstriction of these vessels is not a consistent response to the upright posture; if it occurs it is transient. The capacity of cutaneous veins to contain blood seems primarily determined by thermoregulatory and psychological stimuli. Heat markedly increases venous capacity and thus reduces orthostatic tolerance. Cold has the opposite effects. Muscle veins in human limbs have little smooth muscle and little or no sympathetic innervation and therefore, respond little, if at all, to neural stimuli. Their capacity is determined mainly by the properties of the surrounding skeletal muscle (see below). In humans, intact innervation of the splanchnic bed is of paramount importance for orthostatic tolerance. The upright posture is accompanied by constriction of

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REgUlAToRy MECHAnIsMs InvolvEd In THE AdjUsTMEnTs of THE HUMAn Body To oRTHosTATIC sTREss

splanchnic resistance vessels increasing systemic vascular resistance. The increase in splanchnic vascular resistance also causes a passive expulsion of blood out of the large venous reservoir of the splanchnic bed by elastic recoil of venous vessels. Active constriction of the splanchnic capacitance vessels is potentially of great importance in mobilizing additional venous blood to maintain the cardiac filling pressures and, hence, stroke volume during orthostatic stress. The rich innervation and the great sensitivity and rapidity of the reflex responses of these vessels already to very low frequencies of sympathetic discharge appears indicative for their importance in responding to postural changes. However, because of technical and ethical constraints it has not been possible to determine whether active contraction of splanchnic veins does indeed occur.

Role of Skeletal Muscle Pump Mechanical factors play an important adjunctive role in promoting venous return in the upright posture. First, with quiet standing, the body behaves more or less as an inverted pendulum that sways about the ankles. The static increase in skeletal muscle tone involved opposes venous pooling in lower limb veins. Postural sway during quiet standing is thought to be able to compensate for otherwise poor orthostatic tolerance. The importance of static muscle contractions of the lower body in opposing gravitational pooling of venous blood has been clearly demonstrated in patients with severe orthostatic hypotension due to autonomic failure and in otherwise healthy subjects with a tendency to vasovagal fainting. Leg crossing and contraction of leg and abdominal muscles have been shown beneficial to combat orthostatic hypotension in these subjects. These maneuvers translocate venous blood pooled below the diaphragm to the chest and thereby partially restore cardiac filling pressure, stroke volume and thereby cardiac output (see Chapter 127). Second, activation of the musclevenous pump of the legs during tiptoeing or walking, in the presence of competent venous valves, pumps blood back to the heart and partially restores cardiac filling pressure. The leg-muscle pump can be considered as a “second heart”. Third, the thoraco-abdominal pump may also contribute to improve venous return; with inspiration intrathoracic pressure decreases and intra-abdominal pressure increases thereby promoting venous return. A sighing respiration often precedes an actual faint; it has been suggested that this helps to prevent syncope by enhancing the thoraco-abdominal pump and by inducing venoconstriction in the skin. However, continuous deep breathing and

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the consequent hypocapnia cause vasoconstriction in the brain and the skin venoconstriction is only transient.

Humoral Mechanisms The activity of humoral mechanisms (renin-angiotensin system and vasopressin) is altered by postural changes. The contribution of the humoral system to circulatory orthostatic adjustments depends on the adequacy of the effective circulating blood volume, which is the component of blood volume that the volume-regulatory system responds to by initiating renal retention of water and sodium. When the effective blood volume is adequate, the humoral mechanisms are minimally involved in the initial circulatory adjustment. Activation of the humoral mechanisms becomes more important during prolonged orthostasis, particularly in combatting imminent hypotension in the volume-depleted state. During severe orthostatic stress, both activation of the renin-angiotensin system and vasopressin release are necessary for maintaining blood pressure. Under these circumstances, vasopressin may rise sharply to levels that promote reabsorption of water by the kidneys and have profound vasoconstrictor effects.

Further Reading Claydon VE, Hainsworth R. Increased postural sway in control subjects with poor orthostatic tolerance. J Am Coll Cardiol 2005;46:1309–13. Cooper VL, Hainsworth R. Effects of head-up tilting on baroreceptor control in subjects with different tolerances to orthostatic stress. Clin Sci 2002;103:21–226. Crandall CG, Shibasaki M, Wilson TE. Insufficient cutaneous vasoconstriction leading up to and during syncopal symptoms in heat stressed human. Am J Physiol (Heart Circ Physiol) 2010;299:H1168–1173. Fu Q, Witkowski S, Levine BD. Vasoconstrictor reserve and sympathetic neural control of orthostasis. Circulation 2004;110:2931–7. Groothuis JT, Thijssen DH, Lenders JW, Deinum J, Hopman MT. Leg vasoconstriction during head-up tilt in patients with autonomic failure is not abolished. J Appl Physiol 2011;110:416–22. Miller JD, Pegelow DF, Jacques AJ, et al. Skeletal muscle pump versus Respiratory muscle pump: modulation of venous return from the locomotor limb. J Physiol 2005;563:925–43. Smit AAJ, Halliwill JR, Low PA, et al. Topical Review. Pathophysiological basis of orthostatic hypotension in autonomic failure. J Physiol 1999;519:1–10. Timmers HJLM, Wieling W, Karemaker JM, et al. Denervation of carotid baro- and chemoreceptors in humans. J Physiol 2003;553:3–11. Van Heusden K, Gisolf J, Stok WJ. Mathematical modelling of gravitational effects on the circulation: importance of the time course of venous pooling and blood volume changes in the lung. Am J Physiol (Heart Circ Physiol) 2006;291:H2152–H2165. Wieling W, Krediet CT, van Dijk N, Linzer M, Tschakovsky ME. Initial orthostatic hypotension: review of a forgotten condition. Clin Sci 2007;112:157.

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40 Cerebral Circulation Ronald Schondorf The cerebral circulation is tasked with selectively and specifically directing cerebral blood flow (CBF) to metabolically active regions of the brain while simultaneously responding to or in some cases defending the brain from changes in cerebral perfusion pressure (CPP), carbon dioxide (CO2) and oxygen (O2). A myriad of mechanisms are involved in this complex interplay of demands and the precise integration of these is not well understood. This chapter will touch on some of the clinically relevant factors and challenges that pertain to the control of the human cerebral circulation.

dendritic activity, or to local neuronal spiking. Lastly, increased demand from the brain’s microcirculation must be matched by increased vasodilatation of the upstream extracerebral (pial) resistance vessels. Some of this signaling may be achieved via tight gap junctions within vascular smooth muscle or the endothelium. Alternatively, increased endothelial shear in these vessels may release vasodilating substances such as nitric oxide (NO). Other mechanisms to be discovered undoubtedly exist.

REGULATION OF CEREBRAL CIRCULATION

NEUROVASCULAR COUPLING Local increases in brain activity such as occur during cognitive tasks, are reliably accompanied by parallel increases in CBF and glucose metabolism that greatly exceed the rate of oxygen consumption. The resulting decrease in paramagnetic deoxyhemoglobin can be detected as a change in brain oxygen level dependent (BOLD) signal, the basis for functional MRI. The substrate underlying this response mainly resides in the brain microcirculation beyond the Virchow Robin space. Blood flow through the intraparenchymal vessels is regulated through a combination of signals closely linked to local metabolic activity and shear stress, that originate from locally apposed astrocytes and neurons and possibly from the vascular endothelium itself [1–4]. The intrinsic neural input to the intraparenchymal vessels originates form local interneurons as well as from discrete region-specific intrinsic brain pathways that are located in part in the locus coeruleus, raphe, ventral tegmental region and nucleus basalis [3]. There are several implications that stem from this brief description of the cerebral microcirculation. First, the multiplicity of redundant mechanisms that regulate intraparenchymal CBF makes it highly unlikely that a single factor mediates vasoconstriction or vasodilatation even under well-defined conditions. Second, the complex interplay of metabolically active astrocytes and neurons underscores that caution is required when relating BOLD to a particular element of neuronal activity. Indeed BOLD is imperfectly linked either to extracellularly recorded indices of synaptic and

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00040-8

Basic Considerations The complex hardware (MRI or PET) used in brain mapping requires the subject to perform tasks in the supine position, thereby eliminating many other concomitant challenges to cerebral perfusion [5]. Those who treat patients with autonomic failure more often encounter the opposite end of the spectrum, routine activities of daily living that cause transient or maintained cerebral hypoperfusion and syncope. Many patients however adequately maintain cerebral perfusion despite profound orthostatic hypotension for reasons that are far from clear. The effects of the myriad of endothelial derived factors, circulating or locally released neurotransmitters, changing levels of PCO2, O2, pH, lactate and glucose as well as local cerebrovascular myogenic responses to stretch that have been identified in animals preclude any simple understanding of how cerebral perfusion is regulated or maintained in humans [6,7]. Moreover some of the mechanisms identified in animals may be species specific or alternatively not easily demonstrable in humans using techniques or interventions that are deemed permissible and ethical.

Cerebral Autoregulation Currently most of the useful clinical inferences concerning human cerebral circulatory control have been derived from observing responses to stereotyped challenges such as standing or exercise. Our understanding has been

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further enhanced by the availability of non-invasive transcranial Doppler (TCD) ultrasound and near-infrared spectroscopy (NIRS) which provide robust beat-to-beat approximations of CBF and cerebral oxygenation [6,8]. These techniques have redefined our understanding of cerebral autoregulation, one of the more important processes that defend against cerebral hypoperfusion. Cerebral autoregulation is classically defined as the intrinsic ability to maintain CBF constant (plateau portion of the autoregulation curve) over a wide range of stepwise changes in CPP. Maintenance of such a plateau, however, necessitates high feedback gains not usually encountered in biological systems [8]. Indeed the hydrostatic challenge of standing alone causes a reduction in CBF that is independent of changes in CO2 and is not offset by a siphon mechanism [9]. Whether the reduction in cardiac output during stand contributes in some fashion to the reduction in CBF is still an open question [9]. The dynamic nature of cerebral autoregulation is demonstrated by the rapid correction of CBF during relatively reproducible rapidly induced or alternatively spontaneous changes in CPP [6,8]. The presence of dynamic autoregulation can easily be appreciated by eye when CBF returns to baseline before blood pressure (BP) does, an apparent phase lead of CBF. If dynamic cerebral autoregulation is severely impaired the change in CBF essentially mirrors the change in BP. Dynamic cerebral autoregulation is optimal at frequencies 0.1 Hz and negligible at higher frequencies. A variety of time and frequency domain techniques have been employed in an attempt to describe and more importantly, to quantitate the integrity of dynamic

cerebral autoregulation [6,8]. Not surprisingly, autoregulation varies between individuals and within the same individual both minute to minute and day to day [10]. Input magnitude (BP fluctuations), changes in CO2, cerebrovascular tone and local metabolic demands of the perfused brain as well as the intrinsic non linearity of cerebral autoregulation all contribute to this variability and confound quantitation of autoregulation. For example large BP swings generated by repeat squats improve estimations of cerebral autoregulation [11]. Conversely, increased metabolic activity within the visual cortex results in impairment of posterior but not middle cerebral autoregulation [12]. In neither instance would we say that that the inherent properties of autoregulation are now different. Sufficient sample size, uniformity of population characteristics and strict standardization of test conditions and input magnitude are all necessary to detect true changes in autoregulatory efficacy between populations.

Clinical Considerations Do patients with cerebral hypoperfusion have impairment of cerebral autoregulation? Figures 40.1 and 40.2 show examples of raw tracings from one patient with neurally-mediated syncope and two with autonomic failure. In all three patients the percent decline in BP is much greater than the decline in CBF suggesting that cerebral autoregulation is intact. Some patients with autonomic failure appear to have a reduced autoregulatory capacity whereas others have an autoregulatory range that appears to be expanded. The mechanisms that could possibly

FIGURE 40.1 Raw and filtered (0.2 Hz) blood pressure (BP) and transcranial Doppler (TCD) recordings of right middle cerebral artery blood flow velocity response in a patient with neurally-mediated syncope. The small decline in CBF relative to the large decrease in BP suggests preservation of cerebral autoregulation.

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FIGURE 40.2 Raw and filtered (0.2 Hz) blood pressure (BP) and transcranial Doppler (TCD) recordings of right middle cerebral artery blood flow velocity response from two different patients with autonomic failure.

be responsible for the modification of the cerebral autoregulatory range in patients with autonomic failure are unknown. Dynamic cerebral autoregulation does not appear to be impaired in patients with neurally-mediated syncope but may be altered in patients with autonomic failure [6].

sympathetic activity in human circulatory control remains unclear, phasic increases of cerebral sympathetic activity are critical in defending against spikes in BP that may occur during many activities of daily living and during REM sleep.

References Role of Autonomic Innervation If cerebral perfusion is relatively preserved in patients with profound autonomic failure, then what is the role of the rich extrinsic autonomic innervation of the cerebral vasculature? The parasympathetic innervation of cerebral vasculature may be involved in cerebral vasodilatation but its exact role in human cerebral circulatory control remains unclear. A lively debate concerning the role of sympathetic activity on regulation of CBF has been recently published [13]. To summarize the major points ganglionic blockade or sympathodenervation does not affect resting CBF suggesting that tonic baseline sympathetic outflow to cerebral vasculature is negligible. However, changes in sympathetic activity have been implicated in the reduction of CBF to stand [9] or following beta-adrenergic blockade in subjects during high intensity exercise [13] and alphaadrenergic blockade does affect dynamic cerebral autoregulation. Lastly sympathetic stimulation does restrain the CBF increase during hypertension and recently activation of many neurons in the superior cervical ganglion of anesthetized lambs during imposed hypertension has been directly demonstrated [14]. Although the role of tonic

[1] Andresen J, Shafi NI, Bryan Jr. RM. Endothelial influences on cerebrovascular tone. J Appl Physiol 2006;100:318–27. [2] Cauli B, Hamel E. Revisiting the role of neurons in neurovascular coupling. Front Neuroenerg 2010;2:9. [3] Hamel E. Perivascular nerves and the regulation of cerebrovascular tone. J Appl Physiol 2006;100:1059–64. [4] Koehler RC, Gebremedhin D, Harder DR. Role of astrocytes in cerebrovascular regulation. J Appl Physiol 2006;100:307–17. [5] Raz A, Lieber B, Soliman F, Buhle J, Posner J, Peterson BS, et al. Ecological nuances in functional magnetic resonance imaging (FMRI): Psychological stressors, posture, and hydrostatics. NeuroImage 2005;25:1–7. [6] Singer W, Low PA, Schondorf R. Transcranial doppler evaluation in autonomic disorders. In: Low PA, Benarroch EE, editors. Clinical autonomic disorders (3rd ed.). Baltimore, Philadelphia: Lippincott, Williams & Wilkins; 2008. p. 198–218. [7] Ainslie PN, Duffin J. Integration of cerebrovascular CO2 reactivity and chemoreflex control of breathing: Mechanisms of regulation, measurement, and interpretation. Am J Physiol Regul Integr Comp Physiol 2009;296:R1473–1495. [8] Panerai RB. Transcranial doppler for evaluation of cerebral autoregulation. Clin Auton Res 2009;19:197–211. [9] Immink RV, Truijen J, Secher NH, Van Lieshout JJ. Transient influence of end-tidal carbon dioxide tension on the postural restraint in cerebral perfusion. J Appl Physiol 2009;107:816–23. [10] Brodie FG, Atkins ER, Robinson TG, Panerai RB. Reliability of dynamic cerebral autoregulation measurement using spontaneous fluctuations in blood pressure. Clin Sci (Lond) 2009;116:513–20.

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[11] Claassen JAHR, Levine BD, Zhang R. Dynamic cerebral autoregulation during repeated squat-stand maneuvers. J Appl Physiol 2009;106:153–60. [12] Nakagawa K, Serrador JM, LaRose SL, Moslehi F, Lipsitz LA, Sorond FA. Autoregulation in the posterior circulation is altered by the metabolic state of the visual cortex. Stroke 2009;40:2062–7.

[13] van Lieshout JJ, Secher NH. Point/Counterpoint: Sympathetic activity does/does not influence cerebral blood flow. J Appl Physiol 2008;105:1364–6. [14] Cassaglia PA, Griffiths RI, Walker AM. Sympathetic nerve activity in the superior cervical ganglia increases in response to imposed increases in arterial pressure. Am J Physiol Regul Integr Comp Physiol 2008;294:R1255–1261.

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41 Autonomic Control of the Lower Airways Peter J. Barnes Airway nerves regulate the caliber of the airways and control airway smooth muscle tone, airway blood flow and mucus secretion.

receptors (RAR) account for 10–30% of the myelinated nerve endings in the airways. These endings are sensitive to mechanical stimulation and to protons, low chloride solutions, histamine, cigarette smoke, ozone, serotonin and prostaglandin F2α, although it is possible that some responses are secondary to the mechanical distortion produced by bronchoconstriction. C-fibers. There is a high density of unmyelinated (C-fibers) in the airways, which contain neuropeptides, including substance P (SP), neurokinin A (NKA) and calcitonin gene-related peptide (CGRP). They are selectively stimulated by capsaicin and also activated by bradykinin, protons, hyperosmolar solutions and cigarette smoke. Cough. Cough is an important defense reflex that may be triggered from either laryngeal or lower airway afferents [2]. Both RAR and C-fibers mediate the cough reflex, which may be sensitized in inflammatory diseases by the release of mediators, including neurotrophins [3]. Transient receptor potential A1 (TRPA1) channels play a key role in mediating activation of cough receptors in response to many tussive agents and inflammatory mediators [4] Neurogenic inflammation. Activation of C-fibers may result in the antidromal release of neuropeptides, such as SP, NKA and CGRP (Fig. 41.2). This may increase inflammation in the airways in asthma and COPD, although the role of neurogenic inflammation is debated [5].

OVERVIEW OF AIRWAY INNERVATION Three types of airway nerve and several neurotransmitters are recognized (Table 41.1): l

l l

Parasympathetic nerves which release acetylcholine (ACh). Sympathetic nerves which release norepinephrine. Afferent (sensory nerves) whose primary transmitter is glutamate.

In addition to these classical transmitters, multiple neuropeptides have now been localized to airway nerves and may have potent effects on airway function. Several neural mechanisms are involved in the regulation of airway caliber, and abnormalities in neural control may contribute to airway narrowing in diseases, such as asthma and chronic obstructive pulmonary disease (COPD), contributing to the symptoms and possibly exacerbating the inflammatory response. There is a close interrelationship between inflammation and neural responses in the airways, since inflammatory mediators may influence the release of neurotransmitters via activation of sensory nerves leading to reflex effects and via stimulation of pre-junctional receptors that influence the release of neurotransmitters [1]. In turn, neural mechanisms may influence the nature of the inflammatory response, either reducing inflammation or exaggerating the inflammatory response.

Afferent Nerves At least three types of afferent fiber have been identified in the lower airways (Fig. 41.1). Slowly adapting receptors. Myelinated fibers associated with smooth muscle of proximal airways are probably slowly adapting (pulmonary stretch) receptors that are involved in reflex control of breathing and in the cough reflex. Rapidly adapting receptors. Aδ myelinated fibers in the epithelium show rapid adaptation. Rapidly adapting

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00041-X

TABLE 41.1 Neurotransmitters in the Airways

Neurotransmitter Receptors

Airway Smooth Muscle

Mucus Secretion

Airway Vessels

Acetylcholine

M3

Constrict

Increase

Dilate

Norepinephrine

α1

No effect

No effect

Constrict

Nitric oxide

GC

Dilate

Increase

Dilate

VIP

VIP

Dilate

Increase

Dilate

CGRP

CGRP

Constrict?

No effect

Dilate

Substance P

NK1, NK2

Constrict

Increase

Dilate

Neurokinin A

NK2

Constrict

No effect

No effect

VIP, vasoactive intestinal polypeptide; GC, guanylyl cyclase; CGRP, calcitonin gene related peptide; NK, neurokinin.

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Cholinergic Nerves

Muscarinic receptors. Three subtypes of muscarinic receptor are found in human airways [7]. M1-receptors are localized to parasympathetic ganglia and facilitate neurotransmission. M2-receptors serve as feedback inhibitory receptors on postganglionic nerves (and may be dysfunctional in asthma), whereas M3-receptors mediate the bronchoconstrictor and mucus secretory effect of ACh. Cholinergic reflexes. Reflex cholinergic bronchoconstriction may be activated by afferent receptors in the larynx or lower airways. Cholinergic reflexes are exaggerated in asthma and COPD because of increased responsiveness to ACh. Anticholinergics in airway disease. Muscarinic antagonists (anticholinergics) such as ipratropium and tiotropium, cause bronchodilatation in airway disease through the relief of intrinsic cholinergic tone. They are the bronchodilators of choice in COPD, but less effective than β2-adrenergic agonists in asthma in which several other bronchoconstrictor mechanisms are operative.

Cholinergic nerves are the major neural bronchoconstrictor mechanism in human airways, and are the major determinant of airway caliber [6].

Cholinergic Efferents Cholinergic nerve fibers arise in the nucleus ambiguus in the brainstem and travel down the vagus nerve and synapse in parasympathetic ganglia which are located within the airway wall. From these ganglia short post-ganglionic fibers travel to airway smooth muscle and submucosal glands and release acetylcholine (ACh) that acts on muscarinic receptors (Fig. 41.3).

mechanical stimuli capsaicin, bradykinin (bronchoconstriction) cigarette smoke, SO2 water, hyperosmolar solutions water, low Cl–

Airway epithelium

RAR Aδ-fiber

C-fiber

NP release SP, NKA, CGRP

Bronchial vessel

BRONCHODILATOR NERVES Cough Cholinergic reflex

Neural bronchodilator mechanisms exist in airways and there are considerable species differences.

Airway smooth muscle

Sympathetic nerves. Sympathetic innervation of human airways is sparse and there is no functional evidence for direct innervation of airway smooth muscle, although sympathetic nerves regulate bronchial blood flow and to a lesser extent mucus secretion (Fig. 41.4). Adrenergic tone in the airways is primarily regulated by circulating epinephrine.

SAR

FIGURE 41.1 Afferent nerves in airways. Slowly-adapting receptors (SAR) are found in airway smooth muscle, whereas rapidly adapting myelinated (RAR) and unmyelinated C-fibers are present in the airway mucosa. NP, neuropeptide; SP, substance P; NKA, neurokinin A; CGRP, calcitonin gene-related peptide.

Nodoseljugular ganglion

C.N.S.

Vagus nerve

Laryngeal Esophageal afferents

Parasympathetic nerve C-fiber Aδ-fiber

C-fiber receptors Irritant receptors

ACh

Parasympathetic ganglion Inflammatory cell ACh

ACh

Submucosal gland

Mediators

Airway epithelium Irritants (e.g. cigarette smoke)

FIGURE 41.2 Neurogenic inflammation. Possible neurogenic inflammation (axon reflex) in asthmatic airways via retrograde release of peptides from sensory nerves via an axon reflex. Substance P (SP) causes vasodilatation, plasma exudation and mucus secretion, whereas neurokinin A (NKA) causes bronchoconstriction and enhanced cholinergic reflexes and calcitonin gene-related peptide (CGRP) vasodilatation.

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NEuRAL CoNTRoL of AIRwAys IN DIsEAsE

CNS

M3-receptors ß2-receptors α-receptors

Sympathetic ganglion Vagus nerve

Sympathetic nerve Adrenal medulla

E

NE

Parasympathetic nerve

NE Parasympathetic ganglion NE

Bronchial vessel

ACh Airway smooth muscle

FIGURE 41.3 Cholinergic control of airway smooth muscle. Pre-ganglionic and post-ganglionic parasympathetic nerves release acetylcholine (ACh) and can be activated by airway and extra-pulmonary afferent nerves. NE, norepinephrine; E, epinephrine.

Epithelial shedding

Eosinophils Sensory nerve activation

Vasodilatation

Plasma exudation

Mucus secretion

Neuropeptide release SP, NKA, CGRP, …. Bronchoconstriction Cholinergic activation

Cholinergic facilitation

FIGURE 41.4 Adrenergic control of airway smooth muscle. Sympathetic nerves release norepinephrine (NE), which may modulate cholinergic nerves at the level of the parasympathetic ganglion or postganglionic nerves, rather than directly at smooth muscle in human airways. Circulating epinephrine (E) is more likely to be important in adrenergic control of airway smooth muscle. SP, substance P; NKA, neurokinin A; CGRP, calcitonin generelated peptide.

Inhibitory NANC nerves. The bronchodilator nerves in human airways are non-adrenergic non-cholinergic (NANC) and the major neurotransmitter is nitric oxide (NO). Neuronal NO synthase is expressed mainly in cholinergic neurons.

is a bronchoconstrictor and vasoconstrictor. The neuropeptides in sensory nerves (SP, NKA, CGRP) act as bronchoconstrictors and vasodilators, and also increase mucus secretion and inflammation in the airways [9].

NEURAL CONTROL OF AIRWAYS IN DISEASE

NEUROPEPTIDES Multiple neuropeptides have been localized to nerves in the respiratory tract and function as co-transmitters of classical autonomic nerves to fine-tune airway function [8]. Vasoactive intestinal peptide and related peptides act as bronchodilators and vasodilators, whereas neuropeptide Y

Autonomic control of airways may be abnormal and contribute to the pathophysiology in several airway diseases. Asthma. Neural mechanisms contribute to the pathophysiology of asthma is several ways [10]. Several triggers activate reflex cholinergic bronchoconstriction

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and inflammatory mediators, including neurotrophins, may sensitize the cough reflex. The role of neurogenic inflammation and neuropeptides is still uncertain, however. COPD. The structurally narrowing of the airways in COPD means that the normal vagal cholinergic tone has a relatively greater effect on caliber than in normal airways for geometric reasons. Cholinergic mechanisms are the only reversible component of COPD and may contribute to the mucus hypersecretion of chronic bronchitis.

References [1] Barnes PJ. Modulation of neurotransmission in airways. Physiol Rev 1992;72:699–729. [2] Undem BJ, Carr MJ. Targeting primary afferent nerves for novel antitussive therapy. Chest 2010;137:177–84.

[3] Freund-Michel V, Frossard N. The nerve growth factor and its receptors in airway inflammatory diseases. Pharmacol Ther 2008;117:152–76. [4] Geppetti P, Patacchini R, Nassini R, Materazzi S. Cough: the emerging role of the TRPA1 channel. Lung 2010;188(Suppl 1):S63–8. Epub;2009 Nov 30:S63–S68 [5] Barnes PJ. Cytokine modulators as novel therapies for airway disease. Eur Respir J Suppl 2001;34:67s–77s. [6] Racke K, Matthiesen S. The airway cholinergic system: physiology and pharmacology. Pulm Pharmacol Ther 2004;17:181–98. [7] Barnes PJ. Muscarinic receptor subtypes in airways. Life Sci 1993;52:521–8. [8] Barnes PJ, Baraniuk J, Belvisi MG. Neuropeptides in the respiratory tract. Am Rev Respir Dis 1991;144(1187-98):1391–9. [9] Joos GF, Germonpre PR, Pauwels RA. Role of tachykinins in asthma. Allergy 2000;55:321–37. [10] Undem BJ, Carr MJ. The role of nerves in asthma. Curr Allergy Asthma Rep 2002;2:159–65.

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42 Gastrointestinal Function Michael Camilleri Proper function of the gastrointestinal tract is essential for the orderly digestion, absorption, and transport of food and residue. Digestion requires secretion of endogenous fluids from the salivary glands, stomach, pancreas, and small bowel to facilitate intraluminal breakdown of foods. Fluids, electrolytes, and smaller building blocks of the macronutrients are then absorbed, leaving non-digestible residue to be excreted. The motor activity of the gut is one of the integrated functions that is essential for the normal assimilation of food. Gut motility facilitates the transport of nutrients, brings together digestive enzymes and their substrates, temporarily stores content for optimal absorption (particularly in the distal small bowel and right colon) and, finally, excretes non-digestible residue by defecation in a wellcoordinated function under voluntary control. The extrinsic autonomic nervous system is critically important for almost all secretory and motor functions in the digestive tract by modulating the intrinsic or enteric nervous system of the alimentary tract (Fig. 42.1).

Salivary Secretion Presentation of food to the mouth and olfactory stimulation trigger afferent nerves that stimulate secretory centers in the medulla. These reflexly stimulate efferent fibers along parasympathetic and sympathetic pathways: parasympathetic fibers course along the facial nerve to sublingual and submaxillary glands, and the glossopharyngeal nerve to the parotid gland. Synapse with postganglionic fibers occurs in or near the glands. Sympathetic fibers reach the salivary glands through the cervical sympathetic trunk, but the brainstem centers are unclear. Parasympathetic efferents stimulate secretion; sympathetic fibers serve to cause contraction of myoepithelial cells on the duct. The human salivary glands secrete 0.5–1.0 liter saliva per day at a maximal rate of 4 mL/min. Saliva facilitates speech, lubricates food for swallowing and contains the amylase ptyalin which begins the digestion of starch. Bicarbonate in saliva neutralizes noxious acidic ingesta.

Gastric Secretion Gastric secretion is stimulated by the act of eating (cephalic phase) and the arrival of food in the stomach

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00042-1

(gastric phase). Arrival of the food in the small intestine also controls gastric secretion (intestinal phase). The secreted fluid contains hydrochloric acid, pepsinogen, intrinsic factor, bicarbonate and mucus. Gastric secretion of acid and pepsinogen follows stimulation of oral and gastric vagal afferents. Efferent vagal pathways synapse with submucous plexus neurons which innervate secretory cells via several important bioactive molecules including gastrin, histamine, and somatostatin. In the stomach, there is some digestion of carbohydrate and protein, but very little absorption except for some fat-soluble substances. The mucus-bicarbonate layer protects the stomach lining from auto digestion by acid.

Pancreaticobiliary Secretion Pancreatic juice consists of alkaline (chiefly bicarbonate) fluid and enzymes; 200–800 mL is produced each day. The enzymes, trypsin, lipase, and amylase are essential for digestion of most of the protein, fat, and carbohydrate in the meal. The pancreas consists of exocrine and endocrine portions: bicarbonate and fluid are secreted by ductular cells, chiefly under the influence of secretin; enzymes are produced by acinar cells in response to vagal stimulation of intrapancreatic cholinergic neurons. Cholecystokinin (CCK), which is released from the duodenal mucosal enteroendocrine cells after chemical stimulation by food, activates pancreatic enzyme secretion by stimulating vagal afferents.

Bile Bile is continuously secreted by the liver as two fractions: the bile salt-independent fraction, controlled by secretin and CCK, is similar to the pancreatic ductular secretion; the bile salt-dependent fraction contains bile salts. Bile flow into the intestine is controlled by storage in the gallbladder and by the sphincter of Oddi. Postprandially, the gallbladder contracts under vagal and CCK stimulation, and the basal sphincter tone within the ampulla of Vater falls to allow bile to enter the duodenum. There is evidence that pancreaticobiliary secretion cycles during the interdigestive period, and the cycling is synchronous with the main phases of the gut’s cyclical migrating motor complex (vide infra).

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Parasympathetic Cranial

Salivary secretion Appetite Swallowing

Gallbladder contraction

Hypothalamic and thalamic nuclei

Bile secretion

Sphincter of Oddi relaxation

Gastric secretion and motility

Pancreatic secretion Small bowel and colonic motility and absorption

Sympathetic Thoracolumbar

Visceral sensation, nociception Parasympathetic Sacral roots S2,3,4

Continence and defecation

FIGURE 42.1 Gastrointestinal physiology: functions under extrinsic autonomic control.

Intestinal Secretion and Absorption The small bowel produces about 5 liters of fluid per day during the equilibration of osmotic loads that arrive in the small intestine as ingested nutrients and are the result of intraluminal digestion. Yet, most of the 7 liters entering the digestive tract each day (5 liters from small intestine, 1 liter from stomach, 1 liter from liver and pancreas) is reabsorbed (about 80% in small bowel, 20% in colon) during the flow of chyme through the small bowel and colon; thus, stool weight is usually below 200 g/day in health. Water and electrolyte fluxes are generally independent of extrinsic neural control; on the other hand, the submucosal plexus is increasingly recognized as a key factor influencing mucosal blood flow and enterocyte function. Absorption of macro- and micronutrients is generally determined by concentration gradients or active carriermediated, energy-requiring transport processes. These are indirectly influenced by the autonomic nervous system through its effects on the secretion of salivary, gastric, and pancreaticobiliary juices and by the motor processes of mixing and delivery of substrate to sites of preferential absorption, e.g. B12 to the terminal ileum.

Control of Gut Motility The function of the gastrointestinal smooth muscle is intimately controlled by release of peptides and transmitters by the intrinsic (or enteric) nervous system; modulation of the latter input arises in the extrinsic autonomic nerves, the craniospinal (vagus, and S2, 3, 4 nerves) parasympathetic excitatory input, and the thoracolumbar

sympathetic outflow, which is predominantly inhibitory to the gut, but excitatory to the sphincters (Fig. 42.2). Gastrointestinal smooth muscle forms an electrical syncytium whereby the impulse that induces contraction of the first muscle cell results in efficient transmission to a sheet of sequentially linked cells in the transverse and longitudinal axes of the intestine. The pacemaker of the intestinal muscle syncytium is the network of interstitial cells of Cajal which serve to coordinate contraction circumferentially and longitudinally along the gut. In several species, including humans, the enteric nervous system is formed of a series of ganglionated plexuses, such as the submucosal (Meissner’s), myenteric (Auerbach), deep muscular (Cajal), mucosal and submucosal plexuses. Together these enteric nerves number almost 100 million neurons; this number is roughly equivalent to the number of neurons in the spinal cord. At the level of the diaphragm, the vagus nerve consists predominantly of afferent fibers. Thus, the classical concept that preganglionic vagal fibers synapse with a few motor neurons is not tenable, in view of the overwhelmingly larger number of effector cells that would need to be innervated by the smaller number of preganglionic nerves. The current concept (Fig. 42.3) is that each vagal command fiber supplies an integrated circuit that is hard-wired in the intestinal wall and results in a specific motor or secretory response. These hard-wired circuits in the enteric nervous system are also important in many of the automated responses of the gut, such as the peristaltic reflex. These hard-wired circuits persist and, therefore, they retain their functions even in a totally extrinsically denervated intestine. The enteric nerves also control pacemaker activity.

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GAsTRoInTEsTInAl FunCTIon

Parasympathetic Cranial Hypothalamic and thalamic nuclei

cervical

Glossopharyngeal nucleus NA DMV Vagal nuclei NTS

}

thoracic LES

Vagus N.

pylorus CG (T6-9)

Sympathetic Thoracolumbar

SMG (T9-10)

IMG (T11-L1)

ICS IAS

Parasympathetic Sacral roots S2,3,4

Parasympathetic efferents to digestive tract

EAS

Sympathetic efferents to digestive tract Visceral afferents follow main nerve pathways

FIGURE 42.2 Neural pathways with sympathetic and parasympathetic nervous systems to the gastrointestinal tract. ICS, ileocolonic sphincter; IAS, internal anal sphincter; EAS, external anal sphincter; LES, lower esophageal sphincter; CG, celiac ganglion; SMG, superior mesenteric ganglion; IMG, inferior mesenteric ganglion; NA, nucleus ambiguous; DMV, dorsal motor nucleus of vagus; NTS, nucleus tractus solitarius; T, thoracic.

Vagal efferent fibers stimulate programmed patterns of neural activity; Sympathetic post-ganglionic efferent fibers inactivate neural circuits

Vagal command fiber

Intrinsic neuronal circuits

X

Parasympathetic Cranial

Vagal afferents: visceral reflexes, and CNS-mediated responses

S Sympathetic Thoracolumbar

Enteric effector neurons

Parasympathetic Sacral roots S2,3,4

Muscle cell

Epithelial cell

Visceral afferents: viscero-visceral reflexes, nociception Parasympathetic efferents to digestive tract Sympathetic efferents to digestive tract Visceral afferents follow main nerve pathways

FIGURE 42.3 Integration between extrinsic and intrinsic (or enteric) neural control. Hardwired programs controlling such functions of peristalsis are modulated by efferents in the vagus and sympathetic nerves, which also contain afferent fibers mediating visceral sensation, nociception, and reflex responses. CNS, central nervous system.

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42. GAsTRoInTEsTInAl FunCTIon

Pacemakers located on the greater curve of the gastric corpus and the duodenal bulb “drive” the maximum intrinsic frequency of contractions: three per minute in the stomach and 12 per minute in the small intestine. As in the heart, malfunction of the pacemaker with the highest frequency results in “take over” of pacemaker function by the region with the next highest intrinsic contractile frequency. Derangement of the extrinsic neural control of the gastrointestinal smooth muscle forms the basis for disorders of motility encountered in clinical practice, such as diabetic gastroparesis and fecal incontinence following obstetric trauma of the anal sphincters. Other diseases result from disorders of enteric neural function including achalasia or Hirschsprung's disease.

Normal Gastrointestinal Motor Function Swallowing involves chewing of food, transfer from the oral cavity to the hypopharynx, ejection of the bolus into the esophagus and esophageal peristalsis. The lower esophageal sphincter relaxes at the onset of the swallowing reflex and remains open for a period of about 8 seconds until the bolus passes through the entire esophagus. Then the sphincter contracts to prevent gastroesophageal reflux. Reflux is also prevented by the positive intra-abdominal pressure and contraction of the diaphragmatic crura that occlude the short intra-abdominal portion of the esophagus. Extrinsic neural control reaches the esophagus through efferent pathways in the glossopharyngeal (upper esophagus) and vagus nerves. The motor functions of the stomach and small bowel differ greatly between the fasting and postprandial periods.

(A)

During fasting, cyclical motor events sweep through the stomach and small bowel and are associated with similarly cyclical secretion from the biliary tract and pancreas. The cyclical motor activity is called the interdigestive migrating motor complex; this consists of a phase I of quiescence, phase II of intermittent pressure activity, and phase III or the activity front when contractions of the maximal frequency typical of each region (3 per minute in stomach, 12 per minute in the small bowel) sweep through the gut like a housekeeper, transporting non-digestible residue, products of digestion, epithelial debris and large numbers of commensal bacteria that constitute the normal microbionia and contribute to a large proportion of fecal mass towards the colon for subsequent excretion through the anal canal. The pacemaking functions, cyclical motor activity, and peristalsis are essentially controlled by intrinsic neural pathways, but they can be modulated by extrinsic autonomic nerves. Postprandially, this cyclical activity is abolished, and the different regions of the gut subserve specific functions. Tonic contractions in the gastric fundus result in the emptying of liquids; irregular but high amplitude antral contractions triturate solid food and propel particles that are less than 2 mm in size from the stomach. Particles that are digestible but larger than 2 mm in size are retained by a sieving function of the pylorus. Completion of gastric emptying of solids and liquids takes 3 to 4 hours, depending on the total caloric and fat content of the meal. Irregular, frequent contractions in the postprandial period serve to mix food with digestive juices in the duodenum and jejunum and to propel it aborally. The duration of small bowel transit is on average about 3 hours, and the ileum is a site of temporary storage of chyme,

(B)

At Rest

During Straining

Pubis

Puborectalis

External anal Sphincter Internal anal Sphincter

Coccyx Anorectal angle

Anorectal angle Descent of the pelvic floor

FIGURE 42.4 Dynamics of normal defecation: note the straightening of the rectoanal angle by relaxation of the puborectalis to facilitate evacuation.

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GAsTRoInTEsTInAl FunCTIon

allowing salvage of nutrients, fluids and electrolytes, that were not absorbed upstream. Residues are finally discharged from the ileum to the colon in bolus transfers that probably result from prolonged propagated contractions or reestablished interdigestive cyclical motor activity (the “housekeeper”). The vagus nerve is critically important in efferent control of the fed phase; splanchnic and vagal afferents convey signals from the gut to the prevertebral ganglia, spinal cord and brain to evoke reflex responses and coordinate secretory and motor functions. In animal species such as the dog, the colon also demonstrates cyclical activity, but this is less understood than in the small bowel. This cyclical activity is not recognized in human colon. The proximal colon (ascending and transverse regions) stores solid residue. The ascending colon has variable patterns of emptying: relatively linear, or constant; intermittent; or sudden mass movements. The descending colon is mainly a conduit, and the rectosigmoid functions as a terminal reservoir allowing emptying under voluntary control when there is a call to defecate. Defecation results from a well-coordinated series of motor responses (Fig. 42.4). The rectoanal angle is maintained relatively acute by the puborectalis muscle sling or pelvic floor and this is also important to maintain continence. For defecation to occur, this sling relaxes, thereby

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opening the rectoanal angle to allow a straighter rectal conduit; the external anal sphincter is inhibited voluntarily by parasympathetic (S2, 3, 4) input, and the internal sphincter (under sympathetic control) is inhibited reflexly as a result of rectal distension. Propulsion is facilitated by intracolonic pressure increases, predominantly by a rise in intra-abdominal pressure associated with straining and by propagated contractions of the colon. In contrast, continence is maintained by contraction of the puborectalis (parasympathetic S2, 3, 4), contraction of the internal sphincter (sympathetic lumbar colonic nerves) and contraction of the external sphincter (pudendal nerve).

References [1] Davenport HW. Physiology of the digestive tract. 5th ed. 1982 [2] Camilleri M. Autonomic regulation of gastrointestinal motility. In: Low PA, editor. Clinical autonomic disorders: Evaluation and management (2nd ed.). Philadelphia: Lippincott-Raven; 1997. p. 135–45. [3] Camilleri M. Gastrointestinal motor mechanisms and motility: hormonal and neural regulation. In: Singer MV, Ziegler R, Rohr G, editors. Gastrointestinal tract and endocrine system. Dordrecht, The Netherlands: Kluwer Publishers; 1995. p. 237–53. [4] Cooke HJ. Role of “little brain” in the gut in water and electrolyte homeostasis. FASEB J 1989;3:127–38. [5] Lembo T, Camilleri M. Chronic constipation. N Engl J Med 2003;349:1360–8.

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43 The Splanchnic Circulation Gregory D. Fink, John W. Osborn OVERVIEW The splanchnic circulation is composed of blood vessels serving the stomach, spleen, pancreas, small intestine, colon and liver. The urogenital organs, including the kidneys, are not part of the splanchnic system. The three major arteries serving the splanchnic organs are the celiac and superior and inferior mesenteric. These parallel circuits branch into numerous smaller arteries that in some instances may form anastomoses. Blood flow to the liver occurs through two distinct routes. Approximately one third enters via the hepatic artery (a branch of the celiac artery); another two-thirds arrives via the portal vein. The portal vein is formed by the confluence of smaller veins draining the spleen, pancreas, small intestine and colon. All venous blood leaving the splanchnic region passes through the three major hepatic veins.

Local Regulation The splanchnic organs receive ~25–30% of the cardiac output under normal conditions. Basal blood flow to the splanchnic region far exceeds that required by tissue oxygen utilization, thus total flow is not determined primarily by metabolic needs. Flow increases substantially, however, during feeding and digestion. Local (intrinsic) control mechanisms are responsible for most blood flow changes necessary to support specific functions of the splanchnic organs. For example, blood flow in the GI mucosa is regulated by intramural cholinergic vasodilator nerves activated in response to exposure of the mucosa to nutrients. Collectively the splanchnic vascular beds show some capacity for autoregulation of blood flow, but this is much weaker than in the renal circulation. Veins in the splanchnic region represent a major site for blood storage (capacitance). The splanchnic vessels contain 20–25% of total circulating blood volume. The high compliance and elasticity of veins allows passive forces to exert a major influence on their degree of filling. However, venous volume also is affected by the degree of contraction of smooth muscle in the wall of veins (venoconstriction). Myogenic tone and local humoral factors affect venous smooth muscle activity in response to hemodynamic and metabolic perturbations in the splanchnic organs.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00043-3

Effect of Splanchnic Circulation on Overall Circulatory Function The splanchnic circulation powerfully influences systemic arterial pressure via two distinct mechanisms. Widespread contraction of arteries in the splanchnic bed reduces blood flow to the region. The low oxygen consumption of splanchnic organs allows for a very large reduction in blood flow without producing ischemia. Arterial constriction causes dramatic increases in systemic arterial pressure and total peripheral resistance. Cardiac output also may increase due to passive discharge of stored blood from downstream veins into the central circulation. Active constriction of veins in the splanchnic organs reduces regional blood volume. This has relatively little effect on total peripheral resistance but raises cardiac output and arterial pressure by increasing central blood volume and thus cardiac preload. Generalized arterial and venous constriction in the splanchnic circulation occur mainly in response to extrinsic neural and hormonal inputs. Catecholamines, angiotensin II and vasopressin are among the most powerful hormonal vasoconstrictors. They serve this function particularly in response to major challenges to overall circulatory homeostasis, for example acute hypovolemia caused by hemorrhage. Extrinsic neural regulation is achieved almost exclusively through the sympathetic branch of the autonomic nervous system. That topic is the focus of the remainder of this chapter.

SYMPATHETIC CONTROL OF THE SPLANCHNIC CIRCULATION – GENERAL FEATURES A large majority of the sympathetic nerves innervating the splanchnic organs originate from the T4–T9 segments of the spinal cord as cholinergic preganglionic neurons, pass through the greater splanchnic nerves, synapse in the celiac ganglion plexus, then distribute to various target sites as post-ganglionic adrenergic nerve fibers. Smaller numbers originate in the T10–T12 segments and pass through the lesser and least splanchnic nerves to the prevertebral ganglia. The major neurotransmitters released from sympathetic varicosities are norepinephrine (NE),

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43. THE SPlAnCHnIC CIRCulATIon

which binds to adrenoceptors, and ATP (or possibly a related purine), which binds to P2 purinoceptors and NPY, which binds to Y1 and Y2 receptors. Neurotransmission is regulated prejunctionally by feedback actions of the neurotransmitters, but also by a variety of other factors including angiotensin, nitric oxide, adenosine, and prostanoids. Removal of released transmitters is via neuronal and extraneuronal uptake, metabolism and simple diffusion. Both arteries and veins in the splanchnic region are densely innervated. The degree of innervation is not static: it can change in response to altered physiological demands [1]. In humans the density of sympathetic innervation is substantially greater in mesenteric veins than in arteries [2], presumably reflecting the importance of sympathetic control of splanchnic vascular capacitance during upright posture. The potential exists for distinct sympathetic control of splanchnic arteries versus veins, because they receive some separate postganglionic sympathetic inputs [3]. Furthermore, neurotransmission differs substantially in arteries and veins. For example, venoconstriction occurs at lower levels of nerve activity than does arterial constriction and is better maintained during sustained increases in activity. Alpha-2 adrenergic receptors play a larger role in constriction of mesenteric veins versus arteries. And the relative contribution of ATP and NPY to neurotransmission also varies in arteries versus veins. The functional significance of possible separate regulation of venous and arterial contraction in the splanchnic region remains a matter of dispute [ref 4, and related commentaries]. Splanchnic sympathetic nerve activity (splSNA) is controlled by arterial and cardiopulmonary baroreceptors, and by inputs from other somatic and visceral afferents. Some stimuli are able to alter splSNA in a selective fashion. For example, cholecystokinin and leptin decrease splSNA while generally increasing activity in other sympathetic nerves [5]. Selective changes in sympathetic activity appear to be enabled by the existence of neurons in the rostral ventrolateral medulla of the brainstem that specifically target the splanchnic region. It is unclear if central mechanisms can drive differential activation of arterial versus venous sympathetic nerves. As discussed earlier, sympathetic input to the splanchnic vasculature is not critical for normal functioning of the splanchnic organs, but instead mainly participates in overall regulation of the circulation. Systemic circulatory – and splanchnic hemodynamic – responses to splSNA have been well characterized using both nerve activation and inactivation (e.g. denervation or regional anesthesia). Maximal increases in splSNA can reduce splanchnic blood flow by ~80% and splanchnic blood volume by ~70%. Systemic arterial pressure, total peripheral resistance and cardiac output all increase; the relative proportion of resistance and output changes depends on circulating blood volume, with cardiac output changes predominant when blood volume is low. During sustained increases in splSNA, effects on cardiac output are

generally more persistent than on total peripheral resistance. Catecholamines from the adrenal medulla contribute to both regional and systemic circulatory responses to increased splSNA, especially at higher levels of nerve activity. In conscious, supine subjects inhibition of resting splSNA decreases arterial pressure and cardiac output, but not total peripheral resistance. Splanchnic vascular resistance is also generally unaffected; however, splanchnic blood volume is increased. Thus, basal levels of splSNA appear to regulate splanchnic vascular capacitance more strongly than resistance. This is consistent with evidence from animal studies that splSNA is low under basal conditions.

SYMPATHETIC CONTROL OF THE SPLANCHNIC CIRCULATION IN SPECIAL CIRCUMSTANCES Intact splanchnic sympathetic nerves are important for normal cardiovascular adjustments to upright posture in humans, and to exercise, heat stress and blood loss in humans and other animals. The relative contribution of changes in splanchnic blood flow versus blood volume in these conditions varies, but are driven both by increased splSNA to the vasculature and by release of adrenal catecholamines. Responsiveness of splSNA to these physiological perturbations is reduced with aging. Reduced sympathetically mediated splanchnic vasoconstriction also is an important cause of cardiovascular de-conditioning after bed rest or exposure to microgravity.

SYMPATHETIC CONTROL OF THE SPLANCHNIC CIRCULATION IN DISEASE Impaired sympathetic splanchnic vasoconstriction is an important cause of orthostatic hypotension and other orthostatic disorders such as postural tachycardia syndrome. In portal hypertension there is a very marked increase in sympathetic nervous system activity, including to the splanchnic organs. However, the density of sympathetic innervation in the mesenteric vascular bed is dramatically reduced [6]; this may contribute to the splanchnic vasodilation characteristic of portal hypertension. Autonomic nervous system neuropathy and dysfunction, including orthostatic hypotension, are a significant complication of diabetes. Sympathetic neurons innervating the splanchnic region are most likely to be affected, possibly due to their increased susceptibility to glucose-induced oxidative stress. Increased splSNA worsens intestinal ischemia in septic shock [7]. During the development of arterial hypertension in humans, increased vascular resistance is observed first in the splanchnic circulation. And the splanchnic sympathetic nerves clearly contribute to the pathophysiology of essential hypertension since early work in humans showed that

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splanchnicectomy or celiac ganglionectomy often lowered blood pressure. More recent studies in experimental animals also support the idea that sympathetically mediated splanchnic vasoconstriction can cause hypertension, at least in part by affecting vascular capacitance [8]. Interestingly, it appears that very selective increases in splSNA (with decreases or no change in sympathetic activity to muscle or kidney) account for this effect [9]. A relatively new area of interest is the interaction between sympathetic innervation to the intestine and inflammation. Major changes in sympathetic nerve density and neurotransmission have been reported, for example, in inflammatory bowel disease in humans and animal models [10]. Inflammatory factors may play a larger role than heretofore appreciated in dysregulated sympathetic neurotransmission in a variety of disease states.

References [1] Monos E, Lorant M, et al. Long-term adaptation mechanisms in extremity veins supporting orthostatic tolerance. News Physiol Sci 2003;18:210–4.

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[2] Birch DJ, Turmaine M, et al. Sympathetic innervation of human mesenteric artery and vein. J Vasc Res 2008;45(4):323–32. [3] Kreulen DL. Properties of the venous and arterial innervation in the mesentery. J Smooth Muscle Res 2003;39(6):269–79. [4] Rothe CF. Point: active venoconstriction is/is not important in maintaining or raising end-diastolic volume and stroke volume during exercise and orthostasis. J Appl Physiol 2006;101(4):1262–4. discussion 1265-6, 1270. [5] Sartor DM, Verberne AJ. Gastric leptin: a novel role in cardiovascular regulation. Am J Physiol Heart Circ Physiol 2010;298(2):H406–414. [6] Coll M, Martell M, et al. Atrophy of mesenteric sympathetic innervation may contribute to splanchnic vasodilation in rat portal hypertension. Liver Int 2010;30(4):593–602. [7] Daudel F, Freise H, et al. Continuous thoracic epidural anesthesia improves gut mucosal microcirculation in rats with sepsis. Shock 2007;28(5):610–4. [8] King AJ, Osborn JW, et al. Splanchnic circulation is a critical neural target in angiotensin II salt hypertension in rats. Hypertension 2007;50(3):547–56. [9] Osborn JW, Fink GD. Region-specific changes in sympathetic nerve activity in angiotensin II-salt hypertension in the rat. Exp Physiol 2010;95(1):61–8. [10] Lomax AE, Sharkey KA, et al. The participation of the sympathetic innervation of the gastrointestinal tract in disease states. Neurogastroenterol Motil 2010;22(1):7–18.

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44 Autonomic Control of the Kidney Edwin K. Jackson INTRODUCTION The 1970s ushered in four decades of meticulous investigation of the renal autonomic nervous system. Several comprehensive reviews [1–4] describe the voluminous research underpinning our current state of knowledge of the renal autonomic nervous system and provide a detailed listing of citations in this regard.

INNERVATION OF THE KIDNEY Autonomic control of the kidney is predominantly sympathetic, and there is only scant evidence for parasympathetic innervation. The soma of neurons that project directly to renal sympathetic preganglionic neurons reside predominantly in the paraventricular nucleus of the hypothalamus (PVN), the A5 noradrenergic cell group in the caudal ventrolateral pons (A5), the caudal raphe nuclei (CRN) and the rostral ventrolateral medulla (RVLM) (Fig. 44.1). The RVLM is most critical because it provides the primary tonic excitatory input to preganglionic sympathetic neurons. In this regard, the nucleus tractus solitarius (NTS) receives direct inputs from the peripheral arterial and cardiopulmonary baroreceptors and communicates with the RVLM via the caudal ventrolateral medulla (CVLM). Moreover, the somatosensory and visceral systems gather information from chemoreceptors and mechanoreceptors and send direct inputs to the NTS, CVLM and RVLM. Finally, higher cortical centers may influence renal sympathetic tone via inputs to the aforementioned hypothalamic and brainstem nuclei. There is considerable inter-species and intra-species variation in the neuroanatomical arrangement of the renal sympathetic preganglionic and postganglionic neurons. The cell bodies of renal sympathetic preganglionic neurons reside primarily in the intermediolateral column of the thoracic spinal cord, and the axons of these neurons exit the spinal cord at the last few thoracic segments and first few lumbar segments. Sympathetic preganglionic neurons synapse with renal sympathetic postganglionic neurons both in paravertebral ganglia and in outlying prevertebral ganglia (including the celiac, superior mesenteric, aorticorenal and posterior renal ganglia, as well a variable number of smaller renal ganglia). The thoracic

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00044-5

and lumbar splanchnic nerves provide the dominant pathways for renal sympathetic innervation. Postganglionic sympathetic neurons densely innervate the kidneys and have varicosities on renal vascular smooth muscle cells in the interlobar, arcuate and interlobular arteries and afferent and efferent arterioles, on juxtaglomerular cells and on epithelial cells in proximal tubules, thick ascending limbs of Henle’s loops, distal convoluted tubules and collecting ducts. In this regard, the density of sympathetic varicosities is much greater on vascular structures compared with tubular elements.

AUTONOMIC RECEPTORS IN THE KIDNEY The basal discharge rate of renal sympathetic nerves is 0.5 to 2 Hz, and this causes the continuous basal release of the dominant neurotransmitter in the sympathetic varicosity, norepinephrine, along with lesser amounts of the co-transmitter, neuropeptide Y1–36 (Fig. 44.2). Both norepinephrine and neuropeptide Y1–36 cause autoinhibition (prejunctional negative feedback) of neurotransmitter release via prejunctional α2-adrenoceptors and Y2 receptors, respectively. In addition, several other humoral and paracrine factors also prejunctionally modulate renal noradrenergic neurotransmission, for example angiotensin II via AT1 receptors and epinephrine via β2-adrenoceptors, both of which facilitate neurotransmission. Studies involving direct electrical stimulation of renal nerves support the conclusion that different frequency threshold levels of efferent renal sympathetic nerve activity alter renin release (0.5 Hz), tubular transport (1 Hz) and renal hemodynamics (2.5 Hz). With regard to renin release, neuronally-released norepinephrine acts directly on granular juxtaglomerular cells, a type of modified vascular smooth muscle cell in afferent arterioles, to increase the rate of renin secretion. This effect of norepinephrine is mediated exclusively by β1-adrenoceptors and involves in part the following signal transduction process: (1) activation of Gs; (2) stimulation of adenylyl cyclase; (3) increased levels of cAMP; (4) stimulation of protein kinase A; (5) phosphorylation of proteins leading to increased H translocation into renin-containing granules; (6) increased KCl/H exchange; (7) osmotically-driven influx of H2O into the granule; (8) granule swelling; and (9) exocytosis of renin.

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Arterial & Cardiopulmonary Baroreceptors

Somatosensory/VisceralSystems & Higher Cortical Centers

NTS CVLM

RVLM

Projecting Neuron

A5, CRN, PVN Projecting Neuron

Intermediolateral Column of Spinal Cord (Lower Thoracic & Upper Lumbar Segments)

Sympathetic Preganglionic Neuron

Sympathetic Preganglionic Neuron

Paravertebral Ganglia Prevertebral Ganglia (Celiac, Aorticorenal, Superior Mesenteric, Posterior Renal)

Sympathetic Postganglionic Neuron

Innervated Tubular Structures

Sympathetic Postganglionic Neuron

Innervated Vascular Structures

PT TALH DCT CD

Interlobar Arteries Arcuate Arteries Interlobular Arteries Afferent Arterioles Juxtaglomerular Cells Efferent Arterioles

FIGURE 44.1 Diagram of efferent innervation of the kidney. NTS, nucleus tractus solitarius; CVLM, caudal ventrolateral medulla; RVLM, rostral ventrolateral medulla; A5 area, A5 noradrenergic cell group in the caudal ventrolateral pons; CRN, caudal raphe nuclei; PVN, paraventricular nucleus of the hypothalamus; PT, proximal tubule; TALH, thick ascending limb of Henle’s loop; DCT, distal convoluted tubule; CD, collecting duct.

Low hydrostatic pressure in the afferent arteriole stimulates renin release via the “intrarenal baroreceptor mechanism.” Decreased influx of NaCl into columnar epithelial cells located at the end of the thick ascending limb of Henle’s loop augments renin secretion via the “macula densa mechanism.” Importantly, a powerful synergy exists between β1-adrenoceptor-induced and intrarenal baroreceptor- and macula densa-induced renin secretion. See Castrop et al. [5] for a recent, comprehensive review of mechanisms of renin release. Norepinephrine released from renal sympathetic varicosities also acts directly on renal epithelial cells to accelerate

the rate of solute and water reabsorption from the tubular lumen. This effect of noradrenergic neurotransmission occurs in all nephron segments, but is particularly pronounced in the proximal tubule and ascending limb of Henle’s loop. Although norepinephrine directly affects epithelial cell transport, norepinephrine-induced changes in renal blood flow and glomerular filtration rate may also contribute to sympathetically-induced decreases in renal excretion of electrolytes and water. The direct effects of norepinephrine to enhance tubular transport are mediated mostly by α1-adrenoceptors (predominantly α1Badrenoceptors), which appear to engage the following signal

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REflEx REgulATIon of Blood VolumE

Juxtaglomerular Cell Renal Sympathetic Varicosity

Renin

(+)

β1

Renin Release

α2 α1 Basal Frequency = 0.5–2Hz

NE NPY

(–) (–)

Y1

Renal Vascular Smooth Muscle Cell

Other Prejunctional Modulators

(+)

Ang II (AT1) PGs Epi (β2) Histamine Ach ANP NO Bradykinin Adenosine (A1)

(+)

Vascular Contraction

(+)

Y2 (–)

(+) α2

NE NPY

α1 Renal Epithelial Cell (+)

Solute & H20

Sodium Reabsorption

FIGURE 44.2 Summary of autonomic receptors in the kidney. NE, norepinephrine; NPY, neuropeptide Y1–36; α1, α1-adrenoceptor; β1, β1-adrenoceptor; Y1, Y1 receptor for NPY; Y2, Y2 receptor for NPY; PGs, prostaglandins; Ach, acetylcholine; ANP, atrial natriuretic peptide; NO, nitric oxide; Ang II, angiotensin II; Epi, epinephrine; A1, A1 adenosine receptor; AT1, angiotensin type 1 receptor; β2, β2-adrenoceptor. “Threshold” refers to the frequency of renal nerve stimulation necessary to initiate the indicated response.

transduction process throughout the nephron: (1) activation of Gq; (2) stimulation of phospholipase C-β; (3) increased production of inositol trisphosphate; (4) release of intracellular calcium; (5) calcium-mediated activation of the phosphatase calcineurin; and (6) calcineurin-mediated dephosphorylation and activation of basolateral Na-K-ATPase. In addition, α1-adrenoceptors activate Na/H exchangers (NHE1 and NHE3) in the proximal tubule and the Na/K/2Cl cotransporter (NKCC2) in the thick ascending limb of Henle’s loop, and these actions also contribute to solute reabsorption. Stimulation of renal sympathetic nerves decreases renal blood flow and glomerular filtration rate. These effects are mediated mostly by norepinephrine-induced activation of α1-adrenoceptors (predominantly α1Aadrenoceptors) which mediate intense contraction of vascular smooth muscle cells leading to vasoconstriction of the renal microcirculation. The signal transduction pathway is in part: stimulation of Gq → activation of phospholipase C-β → inositol trisphosphate-induced calcium release  diacylglyerol production → activation of protein kinase C. In this regard, preglomerular microvessels appear to be more responsive to norepinphrine than are

postglomerular microvessels, an imbalance that contributes to norepinephrine-induced reductions in glomerular capillary hydrostatic pressure. Norepinephrine reduces single nephron glomerular filtration rate by decreasing both glomerular capillary hydrostatic pressure and single nephron blood flow. Although α1-adrenoceptors mediate most of the renal hemodynamic effects of renal sympathetic nerve stimulation, α2-adrenoceptors and Y1 receptors also participate, with considerable species variation. Both α2-adrenoceptors and Y1 receptors may contribute to renal vasoconstriction by inhibiting adenylyl cyclase via a Gi-mediated mechanism.

REFLEX REGULATION OF BLOOD VOLUME Autonomic control of the kidney enables an important renal reflex that buffers changes in blood volume and contributes to the rapid restoration of normal blood volume following a positive or negative perturbation in volume status due to for instance a large volume load or blood loss (Fig. 44.3). Increases in blood volume stimulate cardiopulmonary baroreceptors, particularly those

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Positive Volume Perturbation (e.g., Large Volume Load)

(+) Blood Volume (–)

NaCl and H20 Excretion

Stimulation of Cardiopulmonary Baroreceptors

ADH Secretion

Integration of Signals in CNS

Negative Volume Perturbation (e.g., Large Blood Loss)

Renin Release

Renin Release

Tubular Transport

Tubular Transport

RBF & GFR

RBF & GFR

Postganglionic Sympathetic Activity

Postganglionic Sympathetic Activity

Preganglionic Sympathetic Activity

Preganglionic Sympathetic Activity

Premotor Neuron Activity

Premotor Neuron Activity

NaCl and H20 Excretion

(–) (+) Blood Volume

Inhibition of Cardiopulmonary Baroreceptors

ADH Secretion

Integration of Signals in CNS

FIGURE 44.3 Reflex regulation of blood volume by the renal autonomic system. ADH, antidiuretic hormone; RBF, renal blood flow; GFR, glomerular filtration rate; CNS, central nervous system.

in the left atrium. Stimulation of cardiopulmonary baroreceptors by an expanding blood volume increases afferent vagal signals to the NTS, which relays these incoming signals to cardiovascular centers in the brain for further integration. The net result is an inhibition of antidiuretic hormone secretion from the posterior pituitary and a reduction in efferent renal sympathetic nerve activity. The decrease in efferent renal sympathetic nerve activity decreases renin release and renal tubular epithelial transport and increases renal blood flow and glomerular filtration rate. These changes, in conjunction with decreases in antidiuretic hormone levels, markedly increase the excretion rate of NaCl and water, which accelerates the restoration of a normal blood volume. Conversely, a reduction in blood volume (for example by severe bleeding) increases antidiuretic hormone levels and efferent renal sympathetic nerve activity, and these changes reduce NaCl and water excretion to prevent further reductions in blood volume. As a general rule, evolution provides multiple homeostatic mechanisms controlling physiological parameters critical to life, and blood volume is no exception. Redundancy exists in the homeostasis of blood volume, and the extent to which autonomic control of the kidney contributes to blood volume regulation depends on multiple factors such as the magnitude of blood volume perturbation and the physiological status of other regulatory mechanisms.

THE RENORENAL REFLEX Since two kidneys regulate blood volume, pressure and composition, it is not surprising that a mechanism exists to balance these critical tasks between the two kidneys. This mechanism is the renorenal reflex (Fig. 44.4). Increased renal blood flow and glomerular filtration rate to one kidney results in ipsilateral increases in renal venous and pelvic pressures due to greater volumes of blood and urine, respectively, in those compartments. Pressure in the renal venous and pelvic structures activates renal mechanoreceptors residing in the major renal veins, the renal pelvis and the corticomedullary connective tissue. Release of substance P and calcitonin generelated peptide from afferent nerve endings, as well as local formation of prostaglandin E2, may augment the discharge of afferent renal sensory nerves. Renal afferent nerves have their cell bodies in the ipsilateral dorsal root ganglia of the lower thoracic and upper lumbar cord segments, and incoming signals pass via the spinal cord to cardiovascular/renal integration centers in the central nervous system. With increased incoming afferent traffic from the ipsilateral kidney, these centers of integration command a decrease in efferent renal sympathetic nerve activity to the contralateral kidney, which results in increases in renal blood flow to and glomerular filtration by the contralateral kidney, thereby increasing the workload on the contralateral kidney. Moreover, diuresis and

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AuTonomIC ConTRol of THE KIdnEy In PATHoPHysIologICAl sTATEs

219

(–) Increased RBF & GFR to Ipsilateral Kidney

Increased Renal Pelvic Pressure

Increased Renal Venous Pressure Activation of Ipsilateral Renal Mechanoreceptors (Involves SP, CGRP, PGE2)

Activation of Ipsilateral Afferent Renal Nerves with Cell Bodies in DRG Spinal Pathways

Integration in CNS Spinal Pathways

Decreased Efferent Nerve Activity to Contralateral Kidney

Decreased Tubular Transport Increased RBF & GFR to Contralateral Kidney Diuresis and Natriuresis by Contralateral Kidney

FIGURE 44.4 How the renorenal reflex distributes workload evenly between the kidneys. RBF, renal blood flow; GFR, glomerular filtration rate; SP, substance P; CGRP, calitonin gene-related peptide; PGE2, prostaglandin E2; CNS, central nervous system; DRG, dorsal root ganglia.

natriuresis by the contralateral kidney gradually decreases arterial blood pressure, which ultimately reduces the renal blood flow and glomerular filtration rate of the ipsilateral kidney. The net result is a near equal renal blood flow and glomerular filtration rate between the kidneys and consequently an equal sharing of the workload of maintaining a constant blood volume, pressure and composition.

AUTONOMIC CONTROL OF THE KIDNEY IN PATHOPHYSIOLOGICAL STATES The relationship between mean arterial blood pressure and the renal excretion rate of sodium, i.e., the renal pressure-natriuresis curve, determines long-term levels

of arterial blood pressure [6]. Increased renal efferent sympathetic nerve activity impairs renal sodium excretion and shifts the renal pressure-natriuresis curve to the right such that higher long-term levels of blood pressure are required to maintain sodium excretion in balance with sodium intake. It is not surprising, therefore, that efferent renal sympathetic nerve activity contributes to the pathophysiology of hypertension. Evidence for this conclusion includes: (1) complete renal denervation delays and/or attenuates the development of hypertension in a wide spectrum of experimental animal models; (2) efferent renal sympathetic nerve activity is usually increased in hypertension; (3) chronic low-level renal nerve stimulation or chronic intrarenal infusions of norepinephrine induce hypertension; and (4) sympatholytic drugs lower blood

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44. AuTonomIC ConTRol of THE KIdnEy

pressure. However, the pathophysiology of hypertension is multifactorial and efferent renal sympathetic nerves are only one among many participating mechanisms. Renal retention of NaCl and water is a prerequisite for edema formation in congestive heart failure, hepatic cirrhosis and nephrotic syndrome. In these pathophysiological states, blood pressure and intravascular extracellular fluid volume are often diminished, even though total extracellular fluid volume is usually expanded. These perturbations inappropriately engage an arterial baroreceptor-mediated and/or cardiopulmonary baroreceptormediated reflex increase in efferent renal sympathetic nerve activity which contributes importantly to the retention of NaCl and water and consequently to the edematous state. Accordingly, maneuvers that attenuate efferent renal sympathetic nerve activity, for example head-out water immersion, bilateral lumbar sympathetic anesthetic block and administration of sympatholytics, increase NaCl and water excretion in edema associated with heart, liver or kidney disease. Renal afferent nerves may also contribute to over-activation of the sympathetic nervous system in renal diseases. Renal ischemia, hypoxia and injury increases renal afferent signaling to the central nervous system, which activates sympathetic tone to the kidneys, heart and other innervated organs and may elevate arterial blood pressure. The important role of renal sympathetic efferent and

afferent nerves in cardiovascular regulation is underscored by recent advances in the use of catheter-based, radiofrequency ablation of renal efferent and afferent nerves for the treatment of drug-resistant hypertension [4, 7, 8, see Chapter 137].

References [1] DiBona GF, Kopp UC. Neural control of renal function. Physiol Rev 1997;77:75–197. [2] DiBona GF. Physiology in perspective: The Wisdom of the Body. Neural control of the kidney. Am J Physiol Regul Integr Comp Physiol 2005;289:R633–41. [3] Johns EJ, Kopp UC. Neural control of renal function. In: “Seldin and Giebisch’s the kidney: Physiology and pathophysiology. 4th ed. vol. 1, Chapter 33, Amsterdam: Elsevier; 2008. p. 925–46. [4] DiBona GF, Esler M. Translational medicine: the antihypertensive effect of renal denervation. Am J Physiol Regul Integr Comp Physiol 2010;298:R245–53. [5] Castrop H, Höcherl K, Kurtz A, Schweda F, Todorov V, Wagner C. Physiology of kidney renin. Physiol Rev 2010;90:607–73. [6] Johnson RJ, Feig DI, Nakagawa T, Sanchez-Lozada LG, RodriguezIturbe B. Pathogenesis of essential hypertension: historical paradigms and modern insights. J Hypertens 2008;26:381–91. [7] Schlaich MP, Sobotka PA, Krum H, Whitbourn R, Walton A, Esler MD. Renal denervation as a therapeutic approach for hypertension. Novel implications for an old concept. Hypertension 2009;54:1195–201. [8] Schlaich MP, Krum H, Sobotka PA. Renal sympathetic nerve ablation: The new frontier in the treatment of hypertension. Curr Hypertens Rep 2010;12:39–46.

III. AUTONOMIC PHYSIOLOGY

C H A P T E R

45 Dopamine Mechanisms in the Kidney Robert M. Carey RENAL DOPAMINE (DA) FORMATION AND EXCRETION Circulating concentrations of DA (picomolar range) are generally insufficient to activate DA receptors, but concentrations within the kidney (high nanomolar to low micromolar range) are able to activate DA receptors. DA biosynthesis in the kidney occurs in proximal tubule cells (PTC) as a result of uptake of filtered L-dihydroxyphenylalanine (L-DOPA) by way of a sodium (Na) transporter in the apical (brush border) membrane. Once inside the PTC, L-DOPA is rapidly decarboxylated to DA via aromatic amino acid decarboxylase (AAAD), the activity of which is dependent on the Na load to the tubule. DA synthesized within the PTC is not stored but is secreted across the apical and basolateral membrane. The basolateral outward transporter also is dependent upon Na and pH. The supply of L-DOPA to the PTC is the major regulator of DA synthesis and secretion. In vivo, DA is preferentially APICAL MEMBRANE

BASOLATERAL MEMBRANE

L-dopa

L-dopa

Na+ Transporter Na+

DOPA DECARBOXYLASE Na+

DA

DA D1R

AC

secreted across the apical membrane into the tubule lumen, as the increase in urinary DA far exceeds its increment in the renal interstitial fluid [1]. In contrast to the usual pathway for DA biosynthesis in neurons, within the kidney DA is synthesized independently of nerve activity. Figure 45.1 depicts the scheme of DA formation and secretion in the PTC.

RENAL DA RECEPTOR EXPRESSION The D1-like and D2-like receptor families are expressed at post-junctional sites within the kidney. D1 receptors are localized in the smooth muscle cells of renal arterioles, juxtaglomerular (JG) cells, PTCs and cortical collecting ducts (CCD) both by immunohistochemistry and by in situ amplification of mRNA [2,3]. The D5 receptor is predominantly localized to the distal nephron. It is currently thought that D1 receptor function is preferentially exerted over that of the D5 receptor in the PTC, whereas the reverse is true in the distal nephron. The D3 receptor, the predominant D2-like receptor in the kidney, is localized in renal arterioles, glomeruli, PTC, medullary thick ascending limb cells and the CCD by immunohistochemistry [4]. The D4 receptor also is localized specifically in the cortical collecting duct. Renal DA receptor distribution is shown in Table 45.1 and the functions of renal DA receptors is shown in Table 45.2. TABLE 45.1 Renal Dopamine Receptor Expression

DA

ATP

PIP2

cAMP

DAG

PLC

Receptor Subtype

D 1R

IP3

PKC P

Na+ Protein

Protein

P

Na+K+ ATPase

NHE-3 H+

3Na+

Protein

2K+

FIGURE 45.1 Schematic depiction of dopamine formation and cell signaling mechanisms activating sodium transport across the proximal tubule cell. DA, dopamine; D1R, dopamine D1 receptor; PLC, phospholipase C; DAG, diacylglycerol; and AC, adenylyl clyclase.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00045-7

221

Tissue Expression

D1

D2

D3

D4

D5

Arterioles











Glomerulus











Proximal tubule











mTal











Distal tubule











Cortical collecting duct











Renal nerves











Juxtaglomerular cells





?



?

© 2012 Elsevier Inc. All rights reserved.

222

45. DoPAMInE MECHAnIsMs In THE KIDnEy

TABLE 45.2 Effects of Renal Dopamine Receptor stimulation D1-LIKE RECEPTORS (D1 and D5) l l l l

Vasodilation of renal arterioles (requires circulating DA) Inhibition of proximal and distal tubule sodium reabsorption Stimulation of renin secretion Inhibition of angiotensin AT1 receptor expression

D2 RECEPTOR l

Inhibition of norepinephrine release from renal sympathetic neurons

D3 RECEPTOR l l l l l

Inhibition of renin secretion Inhibition of angiotensin AT1 receptor expression Possible increase in glomerular filtration Possible vasodilation of renal arterioles Possible inhibition of tubule sodium reabsorption (natriuresis)

D4 RECEPTOR l

Possible inhibition of sodium reabsorption in the cortical collecting duct

DOPAMINERGIC REGULATION OF RENAL Na EXCRETION D1-like Receptors The renal D1-like receptor family plays a major role in the regulation of tubule Na transport. The natriuretic action of DA is due to inhibition of both proximal and distal tubule Na reabsorption. The action of endogenous DA to inhibit tubule Na transport is distinct from the renal vasodilator action of exogenous DA, which depends upon increased plasma concentrations of DA [5]. A major cell signaling process whereby renal DA induces natriuresis is inhibition of Na, K-ATPase along the entire nephron (PTC, thick ascending limb of Henle’s loop, distal tubule and CCD). This action is thought to be mediated by the D1-like receptor family, although synergism between D1-like and D2-like receptors in the inhibition of Na, K-ATPase has been demonstrated. The signaling processes whereby DA inhibits Na/  K -ATPase activity are nephron-specific. Both protein kinase A (PKA) and protein kinase C (PKC) are involved in the PTC, while only PKA is required in the medullary thick ascending limb and CCD. PKA and PKC both phosphorylate the catalytic subunit of the enzyme inhibiting its activity. A primary component of the regulation of Na/  K -ATPase by DA is the action to inhibit the Na/H exchanger (NHE) and the Na/P1 co-transporter in the apical membrane of the tubule cell. This action of DA inhibits transport of Na into the cell, thus rendering intracellular Na too low to stimulate Na/ K-ATPase. DA’s ability to inhibit NHE activity is primarily due to activation of cyclic AMP and PKA, but can also occur via D1-like receptors directly without involving cAMP and

PKA. DA also can inhibit NHE activity by stimulation of P-450 eicosanoids such as 20-HETE. Endogenous intrarenal DA is a major physiological regulator of urinary Na excretion in vivo. About 60% of basal sodium excretion during normal Na balance is controlled by DA. There is clear evidence that renal DA acts as a paracrine substance (cell-to-cell mediator) locally modulating renal Na excretion by an action at the renal tubule independently of renal hemodynamic function [7]. This has been demonstrated both by pharmacologic blockade of the renal D1-like receptor family and also by antisense oligonucleotide-induced inhibition of D1A receptor protein expression [7,8].

D2-like Receptors Recent evidence suggests that the renal D3 receptor may increase glomerular filtration rate (GFR) via post-glomerular (efferent) arteriolar constriction. Also, a D2-like receptor, possibly D3 or D4, in the basolateral membrane of CCD cells is probably responsible for a natriuretic action of DA in this tubule segment. However, relatively little is known about the D2-like receptor family, as compared with D1like receptors, in the control of Na excretion.

PHYSIOLOGIC INTERACTIONS OF THE RENAL DOPAMINERGIC SYSTEM AND THE RENIN-ANGIOTENSIN SYSTEM (RAS) In addition to direct modulation of renal tubule Na transport, DA affects renin release from renal JG cells via the D1-like receptor family. The interaction of DA with the RAS (Fig. 45.2) is an area of continuing investigation. While D1 receptor activation stimulates renin secretion DOPAMINERGIC SYSTEM RENIN-ANGIOTENSIN SYSTEM D3RECEPTOR

ANGIOTENSINOGEN RENIN

DOPAMINE

ANG I ACE

D1-LIKE RECEPTOR

ANG II

AT1RECEPTOR ANTINATRIURESIS

AT2RECEPTOR NATRIURESIS

FIGURE 45.2 Schematic depiction of the interactions between the renal dopaminergic system and the renal renin-angiotensin system. ANG I, angiotensin I; ANG II, angiotensin II, ACE, angiotensin converting enzyme.

III. AUTONOMIC PHYSIOLOGY

REnAl DA AnD HyPERTEnsIon

directly at JG cells, D1 receptors inhibit renin release by inhibiting macula densa cyclooxygenase activity in vivo [10]. DA also has been demonstrated to decrease PTC angiotensin AT1 receptor expression via the D1 receptor [9], so it appears that the net effect of DA is to increase intrarenal angiotensin II and down-regulate its actions. Current evidence suggests that DA may promote natriuresis both by direct action at the renal tubule and indirectly by decreasing the activity of the RAS and angiotensin II-stimulated Na reabsorption. The renal D3 receptor is also thought to inhibit renin secretion and additionally may inhibit AT1 receptor expression. Renal DA serves as one of several paracrine mediators of renal Na excretion. During low Na intake, Na is retained to meet the body’s requirement for Na homeostasis. Under these conditions, the RAS is stimulated and DA biosynthesis is markedly curtailed, both leading to antinatriuresis. During Na loading, on the other hand, the RAS is suppressed and DA biosynthesis is activated, both leading to natriuresis. When DA synthesis is increased, DA may inhibit renin secretion, reducing the activity of the RAS. Thus, the interaction between the RAS and renal dopaminergic systems leads to a series of overlapping counter-regulatory steps, which attempt to normalize each other to bring the control of Na excretion into equilibrium. In addition to the interactions of renal DA with AT1 receptors, recent evidence has demonstrated a major interaction of renal D1 and AT2 receptors. AT2 receptor activation induces vasodilation and natriuresis via a bradykinin - nitric oxide – cyclic GMP signaling cascade. DA-induced natriuresis is dependent on renal AT2 receptor activation in vivo [11].

RENAL DA AND HYPERTENSION Two fundamental defects in the renal DA system have been described in hypertension: [1] deficient DA production due to decreased PTC uptake and/or decarboxylation of DOPA and [2] defective D1-like receptor-G protein coupling so that DA is ineffective in transmitting a signal to inhibit Na excretion. The latter defect is confined to the PTC and current evidence suggests that the defect is due to hyperphosphorylation of the D1 receptor by a mutation in G-protein-coupled receptor kinase-4 (GRK-4). Under these circumstances, the hyperphosphorylated D1 receptor is internalized in the cytoplasm and desensitized. More work will have to be done to determine whether this defect is responsible for states of salt-sensitivity and sodium-dependent hypertension in humans.

223

Clearly, the role of DA receptors in hypertension has been substantiated by studies in animals in which a specific DA receptor has been disrupted. Knockout of the D1 receptor in mice leads to hypertension, but a mutation in the coding region of the receptor has not been found in human essential hypertension or in genetically hypertensive rats. Disruption of the D2 receptor also induces hypertension, but the increase in blood pressure is related to noradrenergic discharge at the whole body level, and there is no Na retention. Absence of the D3 receptor generates a renin-dependent form of hypertension with inability to excrete a Na load. While these studies show potentially interesting interactions, particularly between renin-angiotensin and dopaminergic systems, compensatory mechanisms may alter the resulting phenotype. Additional work needs to be done especially with renal- and nephron-specific knockout of the individual DA receptors and combinations of receptors of the DA and renin-angiotensin systems.

References [1] Wang Z-Q, Siragy HM, Felder RA, Carey RM. Intrarenal dopamine production and distribution in the rat: physiological control of sodium excretion. Hypertension 1997;29:228–34. [2] O’Connell DP, Botkin SJ, Ramos SP, Sibley DR, Ariano MA, Felder RA, et al. Localization of dopamine D1A receptor protein in the rat kidney. Am J Physiol 1995;268:F1185–F1197. [3] O’Connell DP, Aherne AM, Lane E, Felder RA, Carey RM. Detection of dopamine receptor D1A subtype-specific mRNA in rat kidney by in situ amplification. Am J Physiol 1998;43:232–41. [4] O’Connell DP, Vaughan CJ, Aherne AM, Botkin SJ, Wang Z-Q, Felder RA, et al. Expression of the dopamine D3 receptor protein in rat kidney. Hypertension 1998;32:886–95. [5] Hughes J, Ragsdale NV, Felder RA, Chevalier RL, King B, Carey RM. Diuresis and natriuresis during continuous dopamine-1 receptor stimulation. Hypertension 1988;11(Suppl 1):I-169–I-174. [6] Aperia AC. Intrarenal dopamine: a key signal in the interactive regulation of sodium metabolism. Ann Rev Physiol 2000;62:621–47. [7] Siragy HM, Felder RA, Howell NL, Chevalier RL, Peach MJ, Carey RM. Evidence that intrarenal dopamine acts as a paracrine substance at the renal tubule. Am J Physiol 1989;257:F469–77. [8] Wang Z-Q, Felder RA, Carey RM. Selective inhibition of renal dopamine subtype D1A receptor induces antinatriuresis in conscious rats. Hypertension 1999;33:504–10. [9] Cheng H-F, Becker BN, Harris RC. Dopamine decreases expression of type-1 angiotensin II receptors in renal proximal tubule. J Clin Invest 1996;97:2745–52. [10] Zhang MZ, Yao B, Fang X, Wang S, Smith JP, Harris RC. Intrarenal dopaminergic system regulates renin expression. Hypertension 2009;53:564–70. [11] Salomone LJ, Howell NL, McGrath HE, Kemp BA, Keller SR, Gildea JJ, et al. Intrarenal dopamine D1-like receptor stimulation induces natriuresis via an angiotnesin typr-2 receptor mechanism. Hypertension 2007;49:155–61. [12] Felder RA, Sanada H, Xu J, Yu P-Y, Wang Z, Wang W, et al. G protein-coupled receptor kinase 4 gene variants in human essential hypertension. Proc Nat Acad Sci USA 2002;97:3872–7.

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C H A P T E R

46 Autonomic Control of the Lower Urinary Tract Lori Birder, William C. de Groat INTRODUCTION The storage and periodic elimination of urine is controlled by neural circuitry in the brain and spinal cord that regulates the activity of two functional units in the lower urinary tract: (1) a reservoir (the urinary bladder); and (2) an outlet (consisting of bladder neck, urethra, and striated sphincter muscles). Under normal conditions, the urinary bladder and urethral outlet exhibit reciprocal activity. During storage, the bladder neck and proximal urethra are closed and the detrusor muscle is quiescent, allowing intravesical pressure to remain low over a wide range of bladder volumes. During voluntary micturition the initial event is a reduction of intraurethral pressure, which reflects a relaxation of the pelvic floor and the urethral striated muscles followed in a few seconds by a detrusor contraction and a rise in intravesical pressure that is maintained until the bladder empties. These effects are mediated by three sets of peripheral nerves: sacral parasympathetic (pelvic nerves), thoracolumbar sympathetic nerves (hypogastric nerves and sympathetic chain), and sacral somatic nerves (primarily the pudendal nerves) (Fig. 46.1).

INNERVATION Parasympathetic Pathways The sacral parasympathetic outflow, which in humans originates from S2 to S4 segments of the spinal cord, provides the major excitatory input the bladder. Cholinergic preganglionic neurons located in the intermediolateral region of the sacral spinal cord send axons to cholinergic ganglion cells in the pelvic plexus and in the bladder wall. Transmission in bladder ganglia is mediated by a nicotinic cholinergic mechanism, which is sensitive to modulation by various transmitter systems, including muscarinic, adrenergic, purinergic, and peptidergic (Table 46.1). The ganglion cells in turn excite the bladder smooth muscle. A large proportion of the ganglia and nerves supplying the human lower urinary tract contain acetylcholinesterase

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00046-9

(AChE) as well as the vesicular ACh transporter (VAChT), and therefore must be cholinergic. AChE- and VAChTpositive nerves are abundant in all parts of the bladder but are less extensive in the urethra. Parasympathetic neuroeffector transmission in the bladder is mediated by ACh acting on postjunctional muscarinic (M) receptors. Both M2 and M3 muscarinic receptor subtypes are expressed in bladder smooth muscle, however, studies with subtype-selective muscarinic receptor antagonists and muscarinic receptor knockout mice have revealed that the M3 subtype is the principal receptor involved in excitatory transmission. In bladders of various animals, stimulation of parasympathetic nerves also produces a noncholinergic contraction that is resistant to atropine and other muscarinic receptorblocking agents. ATP (Table 46.1) has been identified as the excitatory transmitter mediating the noncholinergic contraction. ATP excites the bladder smooth muscle by acting on P2X receptors that are ligand-gated ion channels. Among the seven types of P2X receptors that have been identified in the bladder, the P2X1 subtype is the major subtype expressed in the rat and human bladder smooth muscle. Although purinergic excitatory transmission is not important in the normal human bladder, it has been identified in bladders from patients with pathological conditions such as chronic urethral outlet obstruction or interstitial cystitis. Smooth muscle contractions are initiated by an increase in intracellular Ca2 concentration that can occur by intracellular release of Ca2 from the sarcoplasmic reticulum or by influx of Ca2 from the extracellular fluid. The former mechanism is an essential step in the cholinergic activation of the detrusor muscle. It has been shown that stimulation of M3 receptors triggers the formation of inositol triphosphate (IP3) and this in turn activates IP3 receptors on the sarcoplasmic reticulum, which then causes the release of Ca2. On the other hand, activation of P2X purinergic receptors causes the influx of extracellular Ca2 as well as depolarization of the cells, leading to an opening of voltage-gated Ca2 channels. This triggers intracellular Ca2-induced Ca2 release by activation of ryanodine-sensitive receptors in the sarcoplasmic reticulum.

225

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226

46. AUTonomIC ConTRoL of THE LowER URInARy TRACT

FIGURE 46.1 Nerve pathways controlling urine storage and voiding. (A) Urine storage. During the storage of urine, distention of the bladder produces a low level of firing in vesical afferent axons, which in turn stimulates (1) the sympathetic outflow to the bladder outlet (base and urethra) and (2) pudendal outflow to the external urethral sphincter (EUS). These responses occur by spinal reflex pathways and represent guarding reflexes, which promote continence. Sympathetic firing also inhibits the detrusor muscle and modulates transmission in bladder ganglia. A region in the rostral pons (the pontine urine storage center) increases activity of the EUS. (B) Voiding reflexes. During elimination of urine, intense firing of bladder afferents activates spinobulbospinal reflex pathways passing through the pontine micturition center, resulting in stimulation of the parasympathetic outflow to the bladder and urethral smooth muscle and inhibition of the sympathetic and pudendal outflows to the urethral outlet. Ascending afferent input from the spinal cord may pass through relay neurons in the periaqueductal gray (PAG) before reaching the pontine micturition center. () or , excitatory mechanisms or synapses; () or , inhibitory mechanisms or synapses. ON, motor neurons in Onuf’s nucleus; PPN, parasympathetic preganglionic neurons; SPN, sympathetic preganglionic neurons.

Intracellular Ca2 combines with calmodulin to activate the contractile proteins. Activation of M2 muscarinic receptors also appears to enhance contractions by suppressing β-adrenergic inhibitory mechanisms by blocking adenylyl cyclase or K channels. Parasympathetic pathways to the urethra induce relaxation during voiding. In various species, the relaxation is not affected by muscarinic receptor antagonists and therefore is not mediated by ACh. However, inhibitors of NOS block the relaxation in vivo during reflex voiding or block the relaxation of urethral smooth muscle strips induced in vitro by electrical stimulation of intramural nerves, indicating that nitric oxide is the inhibitory transmitter involved in relaxation.

Sympathetic Pathways Sympathetic preganglionic pathways that arise from the T11 to L2 spinal segments pass to the sympathetic chain ganglia and then to prevertebral ganglia in the superior hypogastric and pelvic plexuses, and also to short adrenergic neurons in the bladder and urethra. Sympathetic postganglionic nerves that release norepinephrine provide and

excitatory input to smooth muscle of the urethra and bladder base, an inhibitory input to smooth muscle in the body of the bladder, as well as inhibitory and facilitatory inputs to vesical parasympathetic ganglia. Histofluorescence microscopy in animals and humans has shown that adrenergic terminals richly innervate the smooth muscle of the bladder base, but the bladder body has a considerably weaker adrenergic innervation. Radioligand receptor binding studies show that α-adrenergic receptors are concentrated in the bladder base and proximal urethra, whereas β-adrenergic receptors are most prominent in the bladder body. These observations are consistent with pharmacological studies showing that sympathetic nerve stimulation or exogenous catecholamines produce β-adrenergic receptor-mediated inhibition of the bladder body and strong α-adrenergic receptor-mediated contractions of the base and urethra and weak contractions of the bladder body. Molecular and contractility studies have shown that β3-adrenergic receptors elicit inhibition and α1-adrenergic receptors elicit contractions. The α1A-adrenergic receptor subtype is most prominent in normal bladders, but the α1D subtype is upregulated in bladder from patients with outlet obstruction due to benign prostatic hyperplasia. This

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227

THERAPy

TABLE 46.1 Receptors for Putative Transmitters in the Lower Urinary Tract Tissue

Cholinergic

Adrenergic

Other

Bladder body

 (M2)  (M3)

– (β2) – (β3)

Bladder base

 (M2)  (M3)

(α1)

Urothelium

 (M2)  (M3)

α β

Urethra

 (M)

 (α1)  (α2) – (β)

 Purinergic (P2X1) – VIP  Substance P (NK2) – VIP  Substance P (NK2)  Purinergic (P2X)  TRPV1  TRPM8  P2X  P2Y  Substance P  Bradykinin (B2)  Purinergic (P2X) – VIP – Nitric oxide

(α1)

– NPY

(α1)

– NPY

Sphincter striated  (N) muscle Adrenergic nerve – (M2/4) terminals  (M1) Cholinergic nerve – (M2/4) terminals  (M1) Afferent nerve terminals Ganglia

 (N)  (M1)

 Purinergic (P2X2/3) (α1) –(α2) (β)

 TRPV1 – Enkephalinergic (δ) – Purinergic (P1)  Purinergic (P2X)  Substance P

VIP, vasoactive intestinal polypeptide; NPY, neuropeptide Y; TRP, transient receptor potential. Letters in parentheses indicate receptor type (M, muscarinic; N, nicotinic; NK2, neurokinin-2 receptor). Plus and minus signs indicate excitatory and inhibitory effects.

finding raises the possibility that enhanced α1-adrenergic receptor excitatory mechanisms in the bladder body might contribute to irritative lower urinary tract symptoms in patients with prostatic disease. Activation of β-adrenergic receptors in bladder smooth muscle stimulates adenylyl cyclase and increases cyclic adenosine monophosphate (cAMP), which in turn activates protein kinase A. Protein kinase A is thought to act in part by inducing a hyperpolarization of the cells, either by opening of K channels or by stimulating an electrogenic ion pump. Excitatory responses in the urethra and bladder neck mediated by α1-adrenergic receptors are attributed to an increased release of Ca2 from intracellular stores.

CENTRAL NEURAL CONTROL Urine storage is facilitated by sympathetic and pudendal nerve reflex mechanisms organized in the lumbosacral spinal cord; while voiding is dependent on neural circuitry in the brain that inhibits spinal storage reflexes and activates the parasympathetic outflow to the bladder and urethra. Pathways in the forebrain that are responsible for voluntary control of voiding modulate reflex mechanisms in the pontine micturition center in the brain stem that mediate the coordination between the bladder and urethral sphincter.

NEUROPATHOLOGY Injuries to the neuraxis that interrupt connections between pontine micturition center and the lumbosacral spinal cord lead to complete loss of bladder function and in turn urinary retention. In most spinal cord injured patients, bladder reflexes slowly recover as a result of a reorganization of synaptic connections in the spinal cord and the emergence of sacral reflex mechanisms that initiate involuntary bladder contractions. However, micturition in these patients is usually compromised due to a lack of coordination between bladder and sphincter activity (a condition termed detrusor-sphincter dyssynergia). This condition is characterized by simultaneous contractions of the bladder and the striated urethral sphincter causing incomplete emptying and urinary retention. Experimental studies in animals and humans indicate that the emergence of involuntary voiding reflexes following spinal cord injury is due in part to plasticity in bladder afferent pathways and the unmasking of reflexes triggered by capsaicin-sensitive, C-fiber bladder afferent neurons. C-fiber afferents have also been implicated in the bladder hyperactivity associated with other neurological disorders such as multiple sclerosis. Damage to peripheral neural pathways to the lower urinary tract or to the lumbosacral spinal cord (i.e. a lower motoneuron lesion) causes a loss of bladder sensations as well as loss of voluntary and reflex voiding. On the other hand more selective injury to the motor nerves of the urethral sphincter or the pelvic floor or injury to these striated muscles can often occur during pregnancy and/or childbirth resulting in decreased urethral closure mechanisms and involuntary loss of urine (stress urinary incontinence) during straining, sneezing or coughing that increase intraabdominal pressure.

Somatic Motor Pathways to the Urethral Sphincter The innervation of the striated sphincter muscle of the urethra arises from cholinergic motoneurons in the sacral spinal cord and travels in the pudendal nerves. Transmission in the sphincter muscle is mediated by acetylcholine and activation of nicotinic receptors.

THERAPY Neurogenic involuntary voiding occurring as a result of central nervous system lesions or idiopathic urinary frequency, urgency and urgency incontinence (the overactive bladder syndrome) is usually treated with antimuscarinic

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agents that reduce involuntary bladder contractions. These drugs increase bladder capacity and reduce urgency sensations as well as incontinent episodes. Patients resistant to antimuscarinic therapy can be treated with botulinum toxin injected into the bladder wall. This toxin is also injected into the urethral sphincter to suppress detrusorsphincter-dyssynergia in patients with spinal cord injury.

Further Reading Andersson KE. Detrusor myocyte activity and afferent signaling. Neurourol. Urodyn 2010;29:97–106. Andersson KE, Hedlund P. Pharmacologic perspective on the physiology of the lower urinary tract. Urology 2002;60:13–20.

Birder L, de Groat W, Mills I, Morrison J, Thor K, Drake M. Neural control of the lower urinary tract: peripheral and spinal mechanisms. Neurourol Urodyn 2010;29:128–39. de Groat WC. Integrative control of the lower urinary tract: preclinical perspective. Br J Pharmacol 2006;147:S25–40. Fowler CJ, Griffiths D, de Groat WC. The neural control of micturition. Nat Rev Neurosci 2008;9:453–66. Klausner AP, Steers WD. The neurogenic bladder: an update with management strategies for primary care physicians. Med Clin N Amer 2011;95:111–20. Tai C, Roppolo JR, de Groat WC. Spinal reflex control of micturition after spinal cord injury. Restor Neurol Neurosci 2006;24:69–78. Yoshimura N, Kaiho Y, Miyazato M, Yunoki T, Tai C, Chancellor MB, et al. Therapeutic receptor targets for lower urinary tract dysfunction. Naunyn Schmied Arch Pharmcol 2008;377:437–48.

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47 Bladder Function in Health and Disease Marcus J. Drake, Brian A. Parsons The lower urinary tract (LUT) stores and intermittently expels urine; the alternation between these functions is termed the micturition cycle. The control of the micturition cycle depends on several levels of the neuraxis (see Chapter 46), which enables low pressure urine storage, and volitional initiation of voiding. The complexity of control renders it vulnerable to problems, and LUT symptoms are prevalent clinically.

STRUCTURE OF THE LOWER URINARY TRACT The bladder and its outlet comprise a reservoir and a conduit respectively through which urine is periodically expelled. The bladder is a smooth muscle organ which in the human can vary in volume between 0 and 500 mL. The smooth muscle is termed the detrusor, and is primarily controlled by the parasympathetic innervation. Additional functional regulation is derived from interstitial cells within the detrusor and subjacent to the urothelial lining [1]. The urothelium itself releases active substances which can influence the subjacent afferent nerves and maybe also the detrusor [2]. The conduit of the outlet is the urethra; the urethral smooth muscle also contains interstitial cells – the urothelium here is not as active pharmacologically as in the bladder, but is important in maintaining urethral closure by coaptation. The outlet is supported by the pelvic floor; the skeletal muscle external urethral sphincter contributes to maintaining outlet closure by compression and kinking. The bladder neck in men is relatively welldeveloped and receives a sympathetic nervous system supply mediated by α-1-adrenergic receptors; this structure is important in maintaining bladder neck closure during ejaculation. In health, the bladder alternates between storage and voiding functions, the transition between the two phases being mediated by the periaqueductal grey (PAG) and pontine micturition center (PMC), with extensive input from other CNS centers [3]. The PMC is fundamental to healthy bladder function in ensuring that the bladder and its outlet behave synergistically – which is to say that at any one time only one structure is actively contracting; the bladder during voiding and the outlet during storage.

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PROPERTIES OF THE HEALTHY LOWER URINARY TRACT Properties of the LUT are summarized in Table 47.1. Sensory information from the bladder is derived from the suburothelial afferent complex and from in-series stretch receptors in the detrusor [4]. Once the sensory information reaches the cerebral cortex, it gives rise to perceived sensations, comprising awareness of filling, normal desire to void, and strong desire to void [5]. The sensations are perceived intermittently and can be suppressed, except for a strong desire to void. As the bladder fills, adaptive relaxation means that the pressure change between full and empty is very small (“compliance”). The human bladder is capable of holding around about 500 mL, but typical voided volumes are often less, as people tend to make decisions about the timing of voiding based on practical influences (e.g., anticipated activities) rather than leaving it until their bladder reaches capacity. Volitional control is mediated by higher centers of the central nervous system meaning that people can make a conscious decision regarding initiation of voiding. TABLE 47.1 Sensory and Motor Properties of the Bladder and LUT Outlet During the Phases of the Micturition Cycle Storage

Voiding

BLADDER Motor

Sensory

Low amplitude micromotions [6] Generalized contraction Non-micturition contractions of detrusor by No widespread propagation parasympathetic of motor activity innervation In-series stretch receptors and Off-loading of stretch suburothelial afferents [4] receptors Intermittent conscious awareness [5]

OUTLET Motor

Sensory

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Tonic contraction of circular smooth muscle Tonic contraction of skeletal muscle sphincter, with exertional or voluntary enhancement (“guarding”) Minimal afferent activity

Active relaxation of circular smooth muscle and sphincters Shortening of longitudinal muscle of urethra Reporting of urethral flow through pudendal afferents

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Voiding should result in complete emptying of the bladder. Clinically, urine flow is characterised by key parameters of pattern, maximum flow rate (Qmax) and voided volumes. The latter is important to assessing individual patients as poor urinary flow can result if the bladder is suboptimally filled at the time of testing (Fig. 47.1). Animals have additional voiding characteristics; territorial marking is a rapid release of small volumes of urine not resulting in complete emptying, and voiding in some rodents is associated with high frequency oscillations in sphincter activity. The LUT has to coordinate with the genital tract. The bladder neck remains closed at the time of ejaculation in males, whilst the main urethral sphincter is open to allow expulsion.

CLINICAL EVALUATION LUT symptoms [7] comprise those affecting urine storage (incontinence, urgency, frequency and nocturia), voiding (retention, poor stream, hesitancy, dribbling) or post-voiding (dribbling and sensation of incomplete emptying). Other symptoms include: pain, hematuria (blood in the urine), reduced awareness of filling, and dysuria (burning sensation with urine flow). Physical examination includes a general overview of the patient’s health, checks for any neurological impairment, bladder palpation after voiding, and pelvic examination to check for voluntary pelvic floor squeeze and for prostate enlargement in men. Symptom assessment tools such as the International Consultation on Incontinence Questionnaires [8] and the International Prostate Symptom Score are used to quantify symptoms and measure the associated quality of life

impairment. A frequency volume chart (FVC) is used to measure voided volumes over periods of a few days to evaluate maximum and typical voided volumes, day- and night-time urinary frequency, and overall output [9] (Fig. 47.2). Symptom tools and FVC are assessed before and after treatment to gauge response. Flow rate testing (Fig. 47.1) is used to assess outlet function and bladder contractility. A normal flow rate signifies good bladder contraction, satisfactory outlet channel, and good neurological coordination of the components. Ultrasound bladder scanning after flow rate testing can assess completeness of emptying. Filling and voiding cystometry, “urodynamics”, is an invasive test to measure bladder pressures during storage and voiding. A filling line is placed in the bladder so that fluid can be introduced. A separate line is used to measure pressure in the bladder. Since the bladder is an intra-abdominal organ, additional recording to correct for abdominal pressure changes is done, using a rectal or vaginal line. Detrusor contraction can be assumed if bladder pressure goes up in the absence of a rise in rectal pressure, when both lines are measuring accurately (Fig. 47.3). Additional tests may include cystoscopy which is a visual examination of the inside of the bladder and outlet. Radiological imaging can assess whether adjacent organs are influencing the bladder or whether there is any neurological abnormality.

CLINICAL CONDITIONS Incontinence Incontinence is the involuntary loss of urine. It arises when bladder pressure exceeds outlet resistance, which can occur if bladder pressure is abnormally high, outlet

FIGURE 47.1 Flow rate testing. Three successive flow rate tests on a woman with stress urinary incontinence (left) and a man with multiple sclerosis (right); both patients were aged 34. The left hand traces are basically normal, with a rapid rise, a high maximum flow rate and short duration; the importance of voided volumes is illustrated, as maximum flow rate is clearly lower for the small volume void at the top. On the right, maximum flow rate is clearly lower, and the flow is more protracted. The fluctuations in flow are because the patient used abdominal straining to try to improve flow, suggesting impaired bladder contractility – a well-recognized problem in multiple sclerosis.

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CLInICAL COnDITIOnS

resistance is abnormally low, or from a combination of both. Detrusor overactivity (DO) results when spontaneous or provoked bladder contractions arise inappropriately during the storage phase (Fig. 47.3); if DO contractions are very strong, they can overcome outlet resistance, causing DO incontinence. DO (with or without

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incontinence) is a subgroup of the overactive bladder syndrome (OAB), in which patients complain of excessive urinary urgency – the sudden compelling desire to pass urine which is difficult to defer [7]. OAB is defined by the presence of the urgency symptom, but DO can only be diagnosed by performing urodynamic testing.

FIGURE 47.2 Frequency volume charts. Two three-day frequency volume charts in the same patient, recorded before starting treatment and after one month starting treatment, which included an antimuscarinic drug. Time and volume (if measured) of each void is recorded, allowing the physician to evaluate maximum voided volume (in this case “functional bladder capacity” is 450 ml), typical voided volume and total voided volume per 24 hours. Additional parameters recorded in this case are an urgency score (1–5, with 5 being severe) and number of incontinence pads used. After treatment, daytime frequency, urgency score and number of pads used is lower. DNM, did not measure.

FIGURE 47.3 Urodynamic test. A filling and voiding cystometry trace. The patient was a woman with symptoms of urgency urinary incontinence. Illustrated are traces showing the bladder pressure (Pves; top trace) and abdominal pressure recorded from the rectum (Pabd; second trace). The third trace is the detrusor pressure (Pdet), which the urodynamic computer calculates by subtracting Pves from Pabd. The bottom trace shows urine flow, recorded by a flow rate meter placed under the commode on which the woman was seated. Time in minutes and seconds is shown at the very top. Between time 0 and 10:00, the bladder is being filled with saline at a rate of 30 ml/min (not illustrated); this is the filling phase. The first filling phase event is when the patient was asked to cough, at about 10 seconds; this caused a sharp spike in Pves and Pabd – coughs are repeated throughout, and since each event is associated with minimal deflection of Pdet, it serves as a useful check that pressures are being recorded successfully. Other fluctuations simultaneously present in Pabd and Pves (such as at 1:50) may be due to the patient speaking. At 5:00, the Pves rises, but Pabd does not, so Pdet also rises. This is detrusor overactivity (DO); since it also leads to urine flow, the diagnosis is DO incontinence. At 10:00, the patient reported that she had a strong desire to void. Filling was stopped and the patient was given permission to void, which is illustrated at 10:50. Another cough after the void is included to confirm that the process of voiding did not displace the bladder line, which would lead to erroneous pressure recording.

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Stress urinary incontinence (SUI) results from weakening of the bladder outlet. Physical stresses and straining increase abdominal pressure; this is transmitted to the bladder, and will cause leakage if outlet resistance is impaired. The most common etiology is following childbirth in women, as the pelvic floor is no longer able to support the sphincter as effectively. SUI can occur in men who have suffered loss of sphincter function as a complication of prostatectomy. Mixed urinary incontinence means that both DO incontinence and SUI are present. OAB is managed with fluid advice, bladder training and antimuscarinic medication [10]. Stress incontinence is managed by pelvic floor exercises; surgical measures may be needed to support the bladder outlet, such as placement of a tape at the midpoint of the urethra. Artificial urinary sphincter devices have been developed for SUI in men and neurological SUI.

Bladder Outlet Obstruction The prostate encircles the proximal urethra in men. It enlarges with age and encroaches on the urethral lumen, reducing the calibre of the outlet conduit. This gives rise to both voiding and post micturition symptoms, and is diagnosed according to the fact that detrusor pressures are high, and maximum flow rates during voiding reduced compared with normal [11]. Conservative, medical and surgical interventions are available according to degree of quality of life impairment and confounding risk factors for this older patient population.

Neurourology LUT dysfunction is highly prevalent in neurological disease because of the fundamental importance of several CNS centers in controlling LUT activity. As a consequence, various combinations of the following clinical LUT dysfunctions are possible: 1. DO incontinence: DO is common, as the CNS should inhibit generalised detrusor activity during the storage phase. In low spinal lesions, the adaptive detrusor relaxation underlying filling compliance can be impaired, so pressure rises at higher bladder volumes. 2. SUI: the CNS regulates outlet closure, so CNS disease can weaken sphincter and pelvic floor function. 3. Voiding dysfunction is common, including difficulty initiating voiding (hesitancy) and failure to maintain detrusor contraction until the bladder is emptied. An important subgroup is outlet obstruction caused by detrusor sphincter dyssynergia (DSD), which is simultaneous sphincter and detrusor contraction. 4. Reduced awareness of bladder filling or urine flow, as afferent traffic and conscious perception can be impaired at multiple points in the neuraxis. 5. Ejaculatory dysfunction results from impaired coordination of the sympathetic and parasympathetic spinal nuclei.

6. Autonomic hyperreflexia affects some high spinal cord injury patients. It is a life-threatening hypertension resulting from uninhibited activity of the spinal sympathetic nucleus, elicited by noxious stimuli in patients [12]. Neurourological management [13] primarily consists of ensuring renal function is protected and that the patient remains safe in the longer term. Factors that can impair renal function are poor bladder compliance and DSD, as increased bladder pressure hinders emptying of the ureters, raising pressure within the renal collecting systems. In the longer term this leads to renal failure and was the major cause of death in spinal cord injury up until recent years. In controlling symptoms, measures to improve urine storage include antimuscarinic medication and botulinum injections into the bladder to counteract DO, autologous sling placement or artificial urinary sphincter placement where the outlet function is impaired. To manage voiding dysfunction, intermittent self-catheterization or indwelling catheter drainage may be used.

Painful Bladder Syndrome A rare syndrome which can have a very severe impact on quality of life [14]. Bladder pain is reported during filling and can be slightly eased by voiding. As a consequence, affected patients report continuous pain and frequent trips to the toilet to pass urine, day and night. Bladder capacity is markedly impaired and in extreme cases the bladder may hold only 50 mL.

CONCLUSIONS Normal LUT function requires co-ordination of several structures to achieve the two contrasting roles of storage and voiding. Synergic function of the bladder and its outlet, the latter further complicated in males by the genital outflow, is achieved by complex regulatory mechanisms throughout the neuraxis. Consequently, clinical LUT problems are highly prevalent.

References [1] McCloskey KD. Interstitial cells in the urinary bladder – localization and function. Neurourol Urodyn 2010;29(1):82–7. [2] Birder LA. Urothelial signaling. Auton Neurosci 2010;153(1–2):33–40. [3] Drake MJ, Fowler CJ, Griffiths D, Mayer E, Paton JF, Birder L. Neural control of the lower urinary and gastrointestinal tracts: supraspinal CNS mechanisms. Neurourol Urodyn 2010;29(1):119–27. [4] Kanai A, Andersson KE. Bladder afferent signaling: recent findings. J Urol 2010;183(4):1288–95. [5] Wyndaele JJ, De Wachter S. Cystometrical sensory data from a normal population: comparison of two groups of young healthy volunteers examined with 5 years interval. Eur Urol 2002;42(1):34–8. [6] Drake MJ, Harvey IJ, Gillespie JI, Van Duyl WA. Localized contractions in the normal human bladder and in urinary urgency. BJU Int 2005;95(7):1002–5.

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COnCLUSIOnS

[7] Abrams P, Cardozo L, Fall M, Griffiths D, Rosier P, Ulmsten U, et al. The standardisation of terminology of lower urinary tract function: report from the Standardisation Sub-committee of the International Continence Society. Neurourol Urodyn 2002;21(2):167–78. [8] Abrams P, Avery K, Gardener N, Donovan J. The International Consultation on Incontinence Modular Questionnaire: www.iciq. net. J Urol 2006;175(3 Pt 1):1063–6. [discussion 66] [9] Bright E., Drake M.J., Abrams P. Urinary diaries: evidence for the development and validation of diary content, format, and duration. Neurourol Urodyn 2011; In press. [10] Abrams P, Andersson KE, Birder L, Brubaker L, Cardozo L, Chapple C, et al. Fourth International Consultation on Incontinence Recommendations of the International Scientific Committee:

[11]

[12]

[13]

[14]

Evaluation and treatment of urinary incontinence, pelvic organ prolapse, and fecal incontinence. Neurourol Urodyn 2010;29(1):213–40. Rosario DJ, Woo HH, Chapple CR. Definition of normality of pressure-flow parameters based on observations in asymptomatic men. Neurourol Urodyn 2008;27(5):388–94. Khastgir J, Drake MJ, Abrams P. Recognition and effective management of autonomic dysreflexia in spinal cord injuries. Expert Opin Pharmacother 2007;8(7):945–56. Stohrer M, Blok B, Castro-Diaz D, Chartier-Kastler E, Del Popolo G, Kramer G, et al. EAU guidelines on neurogenic lower urinary tract dysfunction. Eur Urol 2009;56(1):81–8. Hanno P, Nordling J, Fall M. Bladder pain syndrome. Med Clin North Am 2011;95(1):55–73.

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48 Physiology and Pathophysiology of Female Sexual Function Max J. Hilz In a 1999 survey, female sexual dysfunction (FSD) was prevalent in 43% of 1749 American women younger than 60 years [1]. In a British study sexual dysfunction was prevalent in 34.8% of men and 53.8% of women for prob­ lems lasting for at least one month, and in 6.2% of men and 15.6% of women for dysfunction lasting for at least 6 months during the previous year [2]. Common classifications are the International Classifi­ cation of Diseases (ICD)­10 [3] and the Diagnostic and Statistical Manual of Mental Disorders (DSM­IV­TR) [4]. Both include four major FSD categories: disorders of desire, arousal, orgasm and pain. The Second Consensus of Sexual Medicine [5] revised FSD definitions as follows: (A) Hypoactive sexual desire/interest disorder: Absent or diminished feelings of sexual interest or desire, sexual thoughts or fantasies, and a lack of responsive desire; scarce or absent motivations for attempting to become sexually aroused, lack of interest is beyond normal decrease experienced with lifecycle and relationship duration. (B) Arousal disorders: Distress is caused by absence of arousal. Subtypes are subjective (diminished feelings of sexual arousal), genital (complaints of absent/ impaired genital sexual arousal, e.g. minimal vulvar swelling, reduced sexual sensations), combined genital and subjective arousal disorder, and persistent genital arousal disorder (with spontaneous, unwanted genital arousal without sexual interest). (C) Women’s orgasmic disorder: Despite sexual arousal no or delayed low intensity orgasm. (D) Dyspareunia: Pain with attempted or complete vaginal entry. (E) Vaginismus: Persistent or recurrent difficulties to allow vaginal entry despite expressed desire. (F) Sexual aversion disorder: Extreme anxiety/disgust at anticipation or attempt of sexual activity. Masters and Johnson described the sexual response cycle as a linear sequence of excitement, plateau, orgasm and resolution phases. Whipple and Brash­McGreer introduced a circular female sexual response pattern that implements positive or negative reinforcement of sexual

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00048-2

function, with pleasant and satisfying experiences during one sexual experience promoting pleasant further sexual experiences, or vice versa with negative experiences [6].

PHYSIOLOGY OF THE FEMALE SEXUAL RESPONSE CYCLE Sexual Arousal Sexual arousal causes increased pelvic blood flow, geni­ tal vasocongestion, vaginal dilatation and lengthening, engorgement of the labia minora and tissue surround­ ing the urethral lumen, clitoris and vestibular bulbs. Sites of genital sensory activation include the clitoris, clitoral sheath, anterior vaginal wall, labial and introital area, ure­ thra, Halban’s fascia between the anterior vaginal wall and bladder, and the controversially discussed G­spot, an anterior vaginal wall area along the urethra [6,7]. Afferent and Central Pathways Afferent arousal pathways include several sensory sys­ tems, e.g., somatosensory, auditory, vestibular and gusta­ tory stimuli reaching the nucleus tractus solitarii, as well as visual and olfactory stimuli. Stimuli from the pudendal, pelvic hypogastric, and genitofemoral nerves contribute to arousal. There is evidence of a direct vagus nerve innerva­ tion of the upper vagina and cervix [8] that bypasses the spinal cord, and thus allows women to experience arousal and orgasm, even after spinal cord injury above the spinal entry level of the genito­spinal nerves [8]. Impulses reach­ ing the spinal cord ascend to the thalamus the somato­ sensory cortex [6,9]. Among the central areas involved in arousal and responses are the ventral medullary reticular formation that facilitates sexual excitation, the nucleus paragiganto­ cellularis in the ventral medulla that mediates descending inhibitory impulses suppressing the so called urethro­ genital reflex [9]. The periaqueductal grey, mesencephalic ventral tegmental area, neurons in the central tegmental mesencephalic region, and the medial amygdala acti­ vate the medial preoptic area (MPOA), that is involved

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48. PHySIology And PATHoPHySIology oF FEmAlE SExuAl FunCTIon

in arousal [6]. The basomedial hypothalamus seems more important for female than male sexual behavior [6,9]. Neurotransmitters Among central neurotransmitters mediating arousal are dopamine (enhances sexual desire, arousal and willingness towards sexual engagement), norepineph­ rine (gates sensory input from genitalia and maintain­ ing sexual arousal), acetylcholine (mediates peripheral lubrication and vaginal engorgement), histamine, and serotonin (decreases hippocampal norepinephrine release and diminishes excitatory effects of norepinephrine and dopamine, [7] promotes arousal and orgasm in periph­ eral tissue, but also interferes with arousal and orgasm, as evidenced by anorgasmia often associated with intake of selective serotononin reuptake inhibitors [9]). Prolactin causes sexual satiety, reduces libido and gonadal func­ tion, offsets central dopamine effects after orgasm and correlates with post­orgasmic sexual satiety and relief. Oxytocin increases during arousal and orgasm, promotes sexual receptivity, social bonding, care for offspring (“cud­ dling hormone”) [6,7].

Innervation of Female Sexual Function Descending motor impulses innervate rhabdosphinc­ ters and striated pelvic floor muscles and thus contrib­ ute to male and female sexual responses. Motor neurons

originate from the Onuf’s nucleus (S2–S4) that receives impulses from the primary motor cortex, ipsilateral para­ ventricular hypothalamus, ipsilateral caudal pontine lateral reticular formation, contralateral caudal nucleus retroambiguus, and other areas. Impulses, e.g., from the ventromedial nucleus of the hypothalamus (VMN) via the mesencephalon to pelvic floor motor neurons modulate motor sexual responses and, e.g., mediate female lordosis [9]. During orgasm, involuntary rhythmic pelvic floor con­ tractions assure male ejaculation while they squeeze the outer third of the vagina and the anal sphincter [6,7]. Central autonomic structures and peripheral autonomic innervation contribute essentially to female sexual func­ tion [6,9]. Sympathetic and parasympathetic pathways are similar in men and women and reach sexual organs via the inferior hypogastric, ovaric and uterovaginal plexus. The inferior hypogastric plexus receives sympathetic fibers from T10–L2, from the sympathetic trunk via sacral splanchnic nerves (from sacral sympathetic ganglion S2– S5), and parasympathetic fibers from sacral spinal nerves via the pelvic splanchnic nerves [6]. Pelvic splanchnic nerve fibers carry parasympathetic fibers from S2–S5 [6]. The inferior hypogastric plexus innervates bladder, rectum, cervix, vagina, urethra, vestibular bulbs and clitoris [6]. The pudendal nerve (S2–S4) innervates the genita­ lia and perineum with somatic sensory and motor fibers, parasympathetic fibers from the sacral spinal cord and sympathetic fibers from the sacral sympathetic trunk [6].

FIGURE 48.1 Autonomic and somatic innervation of female genitalia. (adapted from Rees et al., Lancet, 2007 with permission from the authors and Elsevier). Note: There is evidence that the pudendal nerve also contains sympathetic and parasympathetic fibers supplying the external genitalia and vagina [6].

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There is controversy regarding the functional contribu­ tion of parasympathetic and sympathetic innervation to female sexual responses [6]. There might be a dual auto­ nomic mediation of sexual arousal with cerebral sym­ pathetic activation during arousal being accompanied by spinal parasympathetic activation. Arousal induced tumescence of clitoris and vaginal bulbs is mediated by trabecular smooth muscle relaxation, similar to male erec­ tion [6]. Vaginal smooth muscle contraction very likely occurs only during late excitation, before orgasm, and is mediated via alpha­adrenoreceptors [6]. Sympathetic activation dominates during late arousal and orgasm [6]. Knowledge about peripheral neu­ rotransmitters of female sexual response is still limited. Adrenergic, cholinergic, nonadrenergic­noncholinergic (NANC) neurotransmitters, vasoactive intestinal polypep­ tide (VIP), nitric oxide synthase, neuropeptide Y, calcito­ nin gene­related peptide, substance P, pituitary adenylate cyclase activating polypeptide, helospectine and peptide histidine methionine have been identified in the female genital tract [6,7].

Common Etiologies of Female Sexual Dysfunction

Hormones Influencing Female Sexual Function

Neurogenic Etiologies

Progesterone seems to further receptivity to part­ ner approach. Estrogens are vasoprotective and support female sexual response by enhancing desire, arousal, nerve transmission, sensory thresholds, vaginal, clitoral and urethral arterial blood flow. Menopausal estrogen decrease leads to vaginal atrophy, inadequate vasoconges­ tion and lubrication, often causing painful intercourse and other FSDs with reduced genital sensation, sexual desire, activity and responsiveness, or anorgasmia. Testosterone is also relevant for desire and initiation of sexual activ­ ity. Decreased levels may also cause reduced wellbeing, libido, sexual receptivity, and pleasure, and induce fatigue and dysphoria, loss of pubic hair, and thinning of the vagi­ nal mucosa [6,7]. Correlations between patient complaints, FSD and age­ related androgen decrease are difficult to assess. Multiple factors may decrease sexual arousability and desire. Female testosterone replacement has gained interest but diagnosis of female androgen insufficiency requires care­ ful differential exclusion of psychosocial issues, psycho­ logical/psychiatric disorders, multiple medical conditions, effects of pharmacologic and recreational drugs, and is not proven by a single – difficult – assessment of low andro­ gen levels [6,7,10]. Transdermally applied testosterone improves sexual desire, responsiveness and activity, and bone mineral den­ sity in postmenopausal women. However, there are sig­ nificant concerns regarding increased cardiovascular risk and effects on breast cancer with testosterone supplementa­ tion. In postmenopausal women, it is non­physiological to prescribe testosterone only, without concomitant estrogen therapy. Breast or uterine cancer, cardiovascular or liver dis­ eases are contraindications against testosterone therapy [10].

These include stroke, epilepsy, Parkinson’s disease, movement disorders, spinal cord injuries, multiple sclero­ sis, cauda equina syndrome, peripheral neuropathies, and surgical disruption of the genital autonomic nerve supply. Impairment of pelvic neurovascular and autonomic struc­ tures may compromise sexual function [6,7].

These include psychogenic and psychiatric disorders (e.g., emotional and relational problems, low self­esteem, body image, depression etc.), cardiovascular, atheroscle­ rotic, neurological or muscular problems (e.g., pelvic floor spasticity causing dyspareunia or vaginism), trauma (or chronic perineal pressure), endocrine problems includ­ ing menopause, hypothalamic­pituitary­adrenal axis dys­ function, premature ovarian failure, medical or surgical castration, metabolic syndrome, diabetes, androgen defi­ ciency, hyperprolactinemia, hypo­ and hyperthyroidism, adrenal insufficiency, female hyperandrogenic disorders and estrogen deficiency. Pelvic, genital or general infec­ tious diseases may cause FSDs. Medications causing FSDs include antihypertensives, antidepressants, antacids, anti­ psychotics, anticholinergic substances, and oral contracep­ tives. Selective serotonin reuptake inhibitors (SSRIs) often inhibit libido and orgasm. SSRI­related FSDs may improve with dosage reduction or switching, e.g., to bupropion or mirtazapine [6,7].

Assessment of Female Sexual Dysfunction Assessment includes a detailed history, physical, geni­ tal examination, neurological and neurophysiological examination. Pressure and touch sensitivity of the genita­ lia should be quantified using Semmes–Weinstein mono­ filaments. Mechanical pain thresholds can be measured by weighted pinprick stimulators. Quantitative sensory testing (QST) of vibratory thresholds evaluates afferent, thickly myelinated A beta­fibers, Paccinian corpuscles and Meissner bodies. QST assessment of cold, warm or heat– pain perception evaluates function of thinly­myelinated A delta­fibers and unmyelinated C­fibers, i.e., small nerve fibers that mediate much of the sensation from erogenous zones [6]. Bulbocavernosus reflex testing or pudendal nerve somatosensory evoked potentials and tests such as puden­ dal nerve distal motor latencies or central motor conduc­ tion time to pelvic floor muscles are useful in suspected central, spinal or peripheral motor pathway lesions. Vaginal photoplethysmography assesses vaginal perfu­ sion. Transcutaneous partial oxygen pressure correlates with sexual arousal and orgasm and may be of diagnos­ tic use. Doppler ultrasound measures clitoral, labial, urethral, vaginal and uterine systolic peak blood flow velocity and end­diastolic velocity at rest and during

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sexual stimulation. Vaginal pH measurement determines lubrication as transsudate during arousal increases vaginal pH [6]. l

Laboratory Investigation Laboratory investigation may include microscopic and cultural examination of vaginal discharges and cervical Papanicolaou smear testing, as well as blood cell count, glu­ cose, thyroid­ and thyroid­stimulating hormone levels [6].

COMMON THERAPEUTIC APPROACHES IN FEMALE SEXUAL DYSFUNCTION [6,7] l

l

l

l

l

Psychoeducation and open discussion are a first step towards therapy. Estrogen replacement may help in postmenopausal women with FSD complaints such as dyspareunia due to sore or atrophic vaginal tissue. Treatment of female sexual arousal disorder is difficult. Multiple formulations, mostly without documented safety or efficacy, have been suggested. Phosphodiesterase type 5 inhibitors seem to be ineffective in female arousal disorders. The FDA approved “EROS­Clitoral Therapy Device”, a battery­ operated device producing a gentle vacuum and low­level vibratory genital sensation seems to increase blood flow to clitoris, vagina and pelvis [11]. Treatment of orgasmic disorders is also difficult and requires sex therapy including cognitive behavioral therapy, sensate focus therapy, encouraging self­ stimulation, pelvic muscle exercises or biofeedback. Benefits may occur with the “EROS­Clitoral Therapy Device” [11]. Sexual pain treatment should address the underlying cause, e.g., vaginal entry pain due to vestibulitis, vaginal dryness or atrophy, or deep pain due to endometriosis or levator spasm. Physiotherapy, vaginal dilators, biofeedback have been helpful. Hypoactive sexual desire disorder is the most common FSD. Often androgen replacement therapy is initiated, yet the approach is controversial. So far, non­ pharmacologic treatment, psychoeducation, lifestyle

changes or stress management and psychological therapies remain the mainstay of managing female sexual interest and desire disorders. A novel treatment option was expected with the development of flibanserin, a 5­HT1a receptor agonist, 5­HT2A receptor antagonist and partial dopamin D4­receptor agonist that was supposed to increase dopamine and norepinephrine and decrease serotonin in selective brain regions and thus facilitate sexual excitation [12]. However, the substance was not approved by the Food and Drug Administration.

References [1] Laumann EO, Paik A, Rosen RC. Sexual dysfunction in the United States: Prevalence and predictors. Jama 1999;281:537–44. [2] Mercer CH, Fenton KA, Johnson AM, Copas AJ, Macdowall W, Erens B, et al. Who reports sexual function problems? Empirical evi­ dence from Britain’s 2000 national survey of sexual attitudes and lifestyles. Sex Transm Infect 2005;81:394–9. [3] World Health Organization (WHO). International statistical clas­ sification of diseases and related health problems 10th revision. Geneva: World Health Organization (WHO); 1994. [4] American Psychiatric Association. Diagnostic and statistical man­ ual of mental disorders. Washington, DC: American Psychiatric Association Press; 2000. [5] Basson R, Althof S, Davis S, Fugl­Meyer K, Goldstein I, Leiblum S, et al. Summary of the recommendations on sexual dysfunctions in women. J Sex Med 2004;1:24–34. [6] Hilz MJ. Female and male sexual dysfunction. In: Low PA, Benarroch EE, editors. Clinical autonomic disorders. Philadelphia: Lippincott Williams & Wilkins; 2008. p. 657–711. [7] Clayton AH, Hamilton DV. Female sexual dysfunction. Psychiatr Clin North Am 2010;33:323–38. [8] Komisaruk BR, Whipple B, Crawford A, Liu WC, Kalnin A, Mosier K. Brain activation during vaginocervical self­stimulation and orgasm in women with complete spinal cord injury: FMRI evi­ dence of mediation by the vagus nerves. Brain Res 2004;1024:77–88. [9] Schober JM, Pfaff D. The neurophysiology of sexual arousal. Best Pract Res Clin Endocrinol Metab 2007;21:445–61. [10] North American Menopause Society. The role of testosterone ther­ apy in postmenopausal women: Position statement of the North American Menopause Society. Menopause 2005;12:496–511. [11] Wilson SK, Delk 2nd JR, Billups KL. Treating symptoms of female sexual arousal disorder with the Eros­clitoral therapy device. J Gend Specif Med 2001;4:54–8. [12] Clayton AH, Dennerstein L, Pyke R, Sand M. Flibanserin: A poten­ tial treatment for hypoactive sexual desire disorder in premeno­ pausal women. Women’s Health (Lond Engl) 2010;6:639–53.

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C H A P T E R

49 Control of the Pupil Karen M. Joos, Mark R. Melson The iris is surrounded by aqueous humor. The actions of the sphincter and dilator muscles on the size of the pupil are not impeded by bulky tissue, are visible, and measureable. It is not surprising that 80 to 100 years ago, at the very beginning of autonomic pharmacology, the pupil was frequently used as an indicator of drug action. In those years it was shown that parasympathetic and sympathetic neural impulses to the iris muscles could be modified by drugs at the synapses and at the effector sites because it was at these locations that the transmission of the impulse depended on chemical mediators. In the following paragraphs these well-known, autonomically active drugs are grouped according to the site and mechanism of their action. Precaution should be taken about the interpretation of pupillary responses to topically instilled drugs. There are large interindividual differences in the responsiveness of the iris to typical drugs, and this becomes most evident when weak concentrations are used. For example, 0.25% pilocarpine will produce a minimal constriction in some patients and an intense miosis in others. This means that the most secure clinical judgments stem from comparisons with the action of the drug on the other, normal eye. The general status of the patient will also influence the size of the pupils. If the patient becomes uncomfortable or anxious while waiting for the drug to act, both pupils may dilate. If the patient becomes drowsy, both pupils will constrict. Thus, if a judgment is to be made about the dilation or contraction of the pupil in response to a drug placed in the conjunctival sac, one pupil should be used as a control whenever possible [1,2].

PARASYMPATHOLYTIC (ANTICHOLINERGIC) DRUGS The belladonna alkaloids occur naturally. They can be found in various proportions in deadly nightshade (Atropa belladonna), henbane (Hyoscyamus niger), and jimsonweed (Datura stramonium). Potions made from these plants were the tools of professional poisoners in ancient times. The word “belladonna” (“beautiful lady”) was derived from the cosmetic use of these substances as mydriatics in sixteenth-century Venice. The mischief caused by the ubiquitous jimsonweed is typical of this group of plants.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00049-4

Jimsonweed has been used as a poison, has been taken as a hallucinogen, and has caused accidental illness and death, and it can cause an alarming accidental mydriasis. These solanaceous plants, which are related to the tomato, potato, and eggplant, are still cultivated for medical purposes. Atropine and scopolamine block parasympathetic activity by competing with acetylcholine at the effector cells of the iris sphincter and ciliary muscle, thus preventing depolarization. After conjunctival instillation of atropine (1%), mydriasis begins within about 10 minutes and is fully developed at 35 to 45 minutes; cycloplegia is complete within 1 hour. The pupil may stay dilated for several days, but accommodation usually returns in 48 hours. Scopolamine (0.2%) causes mydriasis that lasts, in an uninflamed eye, for about 2 days; it is less effective cycloplegic than atropine. Tropicamide and cyclopentolate (are synthetic parasympatholytics with a relatively short duration of action. Tropicamide (1%) is an effective, short-acting mydriatic (3 to 6 hours), which results in only a very transient paresis of accommodation. Compared with tropicamide, cyclopentolate (1%) seems to be a more effective cycloplegic and a slightly less effective mydriatic, especially in dark eyes; accommodation takes about half a day to return and the pupil still may not be working perfectly after more than 24 hours. Botulinum toxin blocks the release of acetylcholine, and hemicholinium interferes with the synthesis of acetylcholine both at the preganglionic and at the postganglionic nerve endings, thus interrupting the parasympathetic pathway in two places. The outflow of sympathetic impulses is also interrupted by systemic doses of these drugs, since the chemical mediator in sympathetic ganglia is also acetylcholine.

PARASYMPATHOMIMETIC (CHOLINERGIC) DRUGS Pilocarpine and methacholine are structurally similar to acetylcholine and are capable of depolarizing the effector cell, thus causing miosis and spasm of accommodation. Methacholine is still sometimes used in a weak (2.5%) solution to test for cholinergic supersensitivity of the

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sphincter muscle in autonomic failure. It has been generally replaced by weak pilocarpine (0.1%). Arecoline is a naturally occurring substance with an action similar to that of pilocarpine and methacholine; its chief advantage is that it acts quickly; a 1% solution produces a full miosis in 10 to 15 minutes (compared to 20 to 30 minutes for 1% pilocarpine) [3]. Carbachol acts chiefly at the postganglionic cholinergic nerve ending to release the stores of acetylcholine. There is also some direct action of carbachol on the effector cell. A 1.5% solution causes intense miosis, but the drug does not penetrate the cornea easily and is therefore usually mixed with a wetting agent (1:3500 benzalkonium chloride). Acetylcholine is liberated at the cholinergic nerve endings by the neural action potential and is promptly hydrolyzed and inactivated by cholinesterase. Cholinesterase, in turn, can be inactivated by any one of the many anticholinesterase drugs. These drugs either block the action of cholinesterase or deplete the stores of the enzyme in the tissue. They do not act on the effector cell directly, they just potentiate the action of the chemical mediator by preventing its destruction by cholinesterase. It follows from their mode of action that these drugs will lose their cholinergic activity once the innervation has been completely destroyed. Physostigmine (eserine) is the classic anticholinesterase. Along the Calabar coast of West Africa the native tribes once conducted trials “by ordeal” using a poison prepared from the bean of the plant Physostigma venenosum. The local name for this big bean was the “esere nut”. The organic phosphate esters (echothiophate [phospholine], isoflurophate [diisopropyl fluorophosphates – DFP], tetraethyl pyrophosphate, hexaethyltetraphosphate, parathion), many of which are in widespread use as insecticides, cause a much longer lasting miosis than the other anticholinesterases, but even this potent effect, thought to be due to interference with cholinesterase synthesis, can be reversed by pralidoxime chloride (P-2-AM).

SYMPATHOMIMETIC (ADRENERGIC) DRUGS Epinephrine stimulates the receptor sites of the dilator muscle cells directly. When applied to the conjunctiva, the 1:1000 solution does not penetrate into the normal eye in sufficient quantity to have an obvious mydriatic effect. If, however, the receptors have been made supersensitive by previous denervation, or if the corneal epithelium has been damaged, allowing more of the drug to get into the eye, then this concentration of epinephrine will dilate the pupil. Phenylephrine in the 10% solution has a powerful mydriatic effect. Its action is almost exclusively a direct alpha stimulation of the effector cell. The pupil recovers in 8 hours and shows a “rebound miosis” lasting several days. A 2.5% solution is now commonly used for mydriasis. Ephedrine acts chiefly by releasing endogenous

norepinephrine from the nerve ending, but it also has a definite direct stimulation effect on the dilator cells. Tyramine (5%) and hydroxyamphetamine (1%) have an indirect adrenergic action; they release norepinephrine from the stores in the postganglionic nerve endings; as far as is known this is their only effective mechanism of action. Cocaine (5% to 10%) is applied to the conjunctiva as a topical anesthetic, a mydriatic, and a test for Horner syndrome. Its mydriatic effect is the result of an accumulation of norepinephrine at the receptor sites of the dilator cells. The transmitter substance builds up at the neuroeffector junction because cocaine prevents the reuptake of the norepinephrine back into the cytoplasm of the nerve ending. Cocaine itself has no direct action on the effector cell nor does it serve to release norepinephrine from the nerve ending, and it does not retard the physiologic release of norepinephrine form the stores in the nerve ending. Its action is indirect, it interferes with the mechanism for prompt disposition of the chemical mediator, and in this respect its action is analogous to that of the anticholinesterases at the cholinergic junction. If the nerve action potentials along the sympathetic pathway are interrupted, as in Horner syndrome, the transmitter substance will not accumulate and the pupil will not dilate. The duration of cocaine mydriasis is quite variable; it may last more than 4 hours. It does not show “rebound miosis.” Apraclonidine (0.5%) is a relatively selective alpha2adrenergic agonist which is used topically to lower intraocular pressure. It will also dilate the pupil in Horner syndrome [4] including pupillary sympathetic denervation in diabetes mellitus [5]. This drug is easily obtainable and is a positive test for sympathetic denervation. Less than 0.5 mm dilation occurs in normal eyes.

SYMPATHOLYTIC DRUGS (ADRENERGIC BLOCKERS) Thymoxamine HCl (0.5%) and dapiprazole are alphaadrenergic blockers that will reverse phenylephrine mydriasis by taking over the alpha-receptor sites on the iris dilator muscle.

OTHER AGENTS l

l

l

l

Substance P affects the sphincter fibers directly, and will constrict the pupil of a completely atropinized eye. The chief pupillary action of morphine is to cut off cortical inhibition of the iris sphincter nucleus in the midbrain, with resultant miosis. Topical morphine, however, even in strong solutions (5%), has a minimal miotic effect on the pupil. Nalorphine and levallorphan are antinarcotic drugs that, given parenterally, reverse the miotic action of morphine. Intravenous heroin seems to produce miosis in proportion to its euphoric effect. In a habituated

III. AUTONOMIC PHYSIOLOGY

PuPIl SymPATHETIC DEfECTS

l

l

heroin user, the same dose of the drug seems to produce less pupillary constriction than in a naïve subject. Thus, given the plasma drug concentration and the size of the pupil in darkness it should be possible to come up with a measure of the degree of physical dependence in a given individual. During the induction of anesthesia the patient may be in an excited state and the pupils are often dilated. As the anesthesia deepens, supranuclear inhibition of the sphincter nuclei is cut off and the pupils become small. If the anesthesia becomes dangerously deep and begins to shut down the midbrain, the pupils become dilated and fail to react to light. The concentration of calcium and magnesium ions in the blood may affect the pupil. Calcium facilitates the release of acetylcholine, and when calcium levels are abnormally low, the amount of acetylcholine liberated by each nerve impulse drops below the level needed to produce a postsynaptic potential, thus effectively blocking synaptic transmission and causing dilation. Magnesium has an opposite effect: a high concentration of magnesium can block transmission and this may dilate the pupil.

241

IRIS PIGMENT AND PUPILLARY RESPONSE TO DRUGS In general, the more pigment in the iris, the more slowly the drug takes effect and the longer its action lingers. This is probably due to the drug being bound to iris melanin and then slowly released. It should be noted that there are wide individual differences in pupillary responses to topical drugs. There is probably a greater range of responses among blue eyes than there is between the average response of blue eyes and the average response of dark brown eyes. Some of these individual differences are due to corneal penetration of the drug [6]. Intense exercise will significantly increase pupil diameter [7]. Melanopsincontaining intrinsically photosensitive retinal ganglion cells also influence pupil diameter besides circadian clock synchronization, sleep, and pineal melatonin production [8].

PUPIL SYMPATHETIC DEFECTS A defect in the sympathetic pathway (Fig. 49.1) affects the pupillary dilator muscle and results in a Horner

FIGURE 49.1 The innervation of the iris muscles, showing the pathways and the terminology in general use. Note that an alerting stimulus dilates the pupil in two ways – both of them with a noradrenergic step in the pathway. The alerting stimulus inhibits the iris sphincter nucleus and, at the same time sends a message down to the cervical cord and the out along the cervical sympathetic pathway. This arrives at the iris about half a second after the sphincter-relaxing message and causes the radial dilator muscle to tighten, thus widening the pupil. III. AUTONOMIC PHYSIOLOGY

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syndrome. If unilateral, asymmetry of pupil diameter is more obvious in darkness than in light. Disorders causing a Horner syndrome include: (1) a distal third-order neuron lesion such as internal carotid dissection, surgery around the carotid artery, or tumor extension into the cavernous sinus; (2) a second-order neuron lesion such as apical lung tumor, chest surgery, thoracic aortic aneurysms, pediatric neuroblastoma, or brachial plexus injury; (3) a central firstorder neuron lesion such as vascular occlusion near the lateral medulla, or tumors or disc disease near the upper cervical spinal cord. Bilateral defects have been reported in diabetes mellitus [5], pure autonomic failure [9], and dopamine beta-hydroxylase deficiency [10].

PUPIL PARASYMPATHETIC DEFECTS A defect in the parasympathetic pathway (Fig. 49.1) affects the pupillary sphincter muscle and results in a larger pupil. If unilateral, asymmetry of pupil diameter is more obvious in the light than in darkness. Disorders of postganglionic parasympathetic defects include lesions of the ciliary ganglion or short posterior ciliary nerve causing a tonic Adie pupil [11]. Bilateral defects have been reported in Miller Fisher syndrome [9], acute pandysautonomia [9], paraneoplastic autonomic neuropathy [9], Sjögren syndrome [9,10], and rarely systemic lupus erythematosus [9]. Pupil involvement accompanied by a third cranial nerve palsy requires investigation for an aneurysm at the junction of the internal carotid and posterior communicating arteries.

PUPIL COMBINED SYMPATHETIC AND PARASYMPATHETIC DEFECTS

in Guillain–Barre syndrome [9], Lambert–Eaton myasthenic syndrome [9], diabetes mellitus [9], acute and subacute dysautonomia [10], and amyloidosis [10].

Acknowledgements This chapter was updated with permission by the author in previous editions: H. Stanley Thompson, M.D., Oxford, Iowa. Support: NIH Core Grant 2P30EY008126-22 and an unrestricted departmental grant from Research to Prevent Blindness, Inc., NY.

References [1] Loewenfeld IE. The pupil: anatomy, physiology and clinical applications. Ames, IA: Iowa State University Press; 1993. pp 797-826 and 1255-1558. (Reprinted by Butterworth-Heinemann in 1997) [2] Thompson HS. The Pupil. In: Hart WmM, editor. Alder’s physiology of the eye (9th ed.). St. Louis: Mosby-Year Book; 1992. p. 429. [3] Babikian PV, Thompson HS. Arecoline miosis [Letter]. Am J Ophthalmol 1984;98:514–5. [4] Brown SM, Aouchiche R, Freedman KA. The utility of 0.5% apraclonidine in the diagnosis of Horner syndrome. Arch Ophthalmol 2003;121:1201–3. [5] Koc F, Kansu T, Kavuncu S, Firat E. Topical apraclonidine testing discloses papillary sympathetic denervation in diabetic patients. J Neuro-Ophthalmol 2006;26:25–9. [6] Kardon R. Drop the Alzheimer’s drop test. [Editorial]. Neurology 1998;50:588–91. [7] Hayashi N, Someya N, Fukuba Y. Effect of intensity of dynamic exercise on pupil diameter in humans. J Physiol Anthropol 2010;29:119–22. [8] Bailes HJ, Lucas RJ. Melanopsin and inner retinal photoreception. Cell Mol Life Sci 2010;67:99–111. [9] Toth C, Fletcher WA. Autonomic disorders and the eye. [Editorial]. J Neuro-Ophthalmol 2005;25:1–4. [10] Bremner FD, Smith SE. Pupil abnormalities in selected autonomic neuropathies. J Neuro-Ophthalmol 2006;26:209–19. [11] Bremmer F. Pupil evaluation as a test for autonomic disorders. Clin Auton Res 2009;19:88–101.

Defects in both pathways (Fig. 49.1) affect the pupillary dilator and sphincter muscles. Disorders of the sympathetic and parasympathetic pathways have been reported

III. AUTONOMIC PHYSIOLOGY

C H A P T E R

50 Central Thermoregulation Shaun F. Morrison Central neural circuits orchestrate a homeostatic repertoire to maintain body temperature during environmental temperature challenges (Fig. 50.1) and to alter body temperature during the inflammatory response. Body temperature regulation is effected primarily through dedicated pathways in the brain which function to produce an optimal operating temperature for neurons and for the many tissues on which the brain depends for survival. The principal non-behavioral effector mechanisms for cold defense, recruited in order of increasing energy costs, include heat conservation resulting from cutaneous vasoconstriction (CVC) and piloerection and heat production from thermogenesis, a byproduct of the inefficiency of mitochondrial ATP production and of ATP utilization, in brown adipose tissue (BAT), the heart and skeletal muscle (shivering). Heat loss mechanisms for heat defense include cutaneous vasodilation and evaporative cooling. The activation of these effectors is regulated by parallel but distinct, effector-specific, core efferent pathways within the central nervous system that share a common peripheral thermal sensory input. A wide variety of non-thermal physiological parameters, disease processes, neurochemicals and drugs can influence the central regulation of body temperature and their effects are hypothesized to result from an alteration of the activity within the core neural circuit for thermoregulation. The core central thermoregulatory network (Fig. 50.2) comprises the fundamental pathways through which cutaneous cold and warm sensation and/or reductions or elevations in brain temperature elicit changes in thermoregulatory effector tissues to counter or protect against deviations from a homeostatic temperature of the brain and other critical organ tissues.

CUTANEOUS THERMAL RECEPTOR AFFERENT PATHWAY The central thermoregulatory system receives signals related to changes in environmental temperature through thermoreceptors in primary sensory nerve endings distributed in the skin. Members of the transient receptor potential (TRP) family of cation channels comprise the molecular mechanisms of cutaneous cool and warm thermoreception [1]. Primary thermal somatosensory afferents

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00050-0

synapse on thermoreceptive-specific, lamina I spinal (or trigeminal) dorsal horn cells that respond linearly to graded, innocuous cooling or warming stimuli, but are not activated further in the noxious temperature range. In turn, spinal and trigeminal lamina I neurons collateralize and innervate the thalamus and the pontine lateral parabrachial nucleus (LPB) (Fig. 50.2). Densely clustered neurons that project principally to the midline subregion of the POA, including the median preoptic nucleus (MnPO), from the dorsal (LPBd) and external lateral (LPBel) subnuclei of the LPB are activated following warm or cold exposure, respectively, in parallel with respective skin warming-evoked inhibitions and skin cooling-evoked activations of BAT sympathetic nerve activity (SNA) and BAT thermogenesis [2,3]. Activation of LPBd or LPBel neurons evokes respective decreases or increases in BAT thermogenesis, metabolism and heart rate (HR) that mimic skin warming-evoked or skin cooling-evoked physiological response. Either inhibition of LPBel neurons or blockade of their glutamate receptors eliminates skin cooling-evoked cold-defense responses, including the activation of BAT and shivering thermogenesis and increases in metabolism and in HR [2]. Similar inhibition of LPBd neurons eliminates skin warming-evoked heat-defense responses, including the inhibition of cutaneous vasoconstrictor SNA (mediating cutaneous vasodilation) [3]. Thus, activations of LPBd and LPBel neurons, likely by glutamatergic inputs from lamina I neurons, driven respectively by cutaneous warming and cooling signals, transmit the respective warm and cold cutaneous thermal afferent stimuli that initiate heat defense and cold defense responses to defend body temperature during environmental thermal challenges (Figs 50.1, 50.2). The spinoparabrachiopreoptic thermal afferent pathway that triggers involuntary thermoregulatory responses is distinct from the spinothalamocortical pathway, in which lamina I neurons synapse on neurons in the thalamus that project to the primary somatosensory cortex, which leads to perception and discrimination of cutaneous temperature [2] (Fig. 50.2). The relative contributions of the spinothalamic vs. spinoparabrachial pathways in initiating thermoregulatory behaviors, the stereotypical somatic motor acts directed toward minimizing or optimizing heat transfer from the body to the environment, remain to be elucidated.

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THERMOREGULATORY SENSORIMOTOR INTEGRATION IN THE PREOPTIC AREA

FIGURE 50.1 Modulation of thermal effectors by activation of cutaneous thermal receptors. (A) Reductions in skin temperature (Tskin) elicit increases in brown adipose tissue (BAT) sympathetic nerve activity (SNA), in BAT thermogenesis and BAT temperature (Tbat), in expired CO2, an indicator of increased metabolism, in heart rate (HR), but little change in arterial pressure (AP). (B) With lowered core and brain temperatures (Tbrain), reductions in skin temperature elicit shivering, indicated by marked increases in neck muscle EMG. Note the simultaneous increase in BAT temperature and thermogenesis. (C) Increases in skin temperature inhibit the cutaneous vasoconstrictor (CVC) sympathetic outflow monitored as the action potentials of a single postganglionic fiber (unit) in the sural nerve. Note also the increase in tail skin temperature (Ttail), an indicator of cutaneous vasodilation promoting heat loss.

Glutamatergic stimulation of MnPO neurons, rather than those in medial (MPO) or lateral preoptic areas, evokes thermogenic, metabolic and tachycardic responses similar to those evoked during cold-defense [4]. Colddefense responses triggered either by LPBel stimulation or by skin cooling are blocked by antagonizing glutamate receptors in the MnPO. Similarly, skin warming signaling, mediated via LPBd neurons, is transmitted preferentially to neurons in MnPO and in the rostral dorsomedial portions of MPO and blockade of glutamate receptors in this region of the POA interrupts skin warming-evoked responses [3]. Thus, activation of MnPO neurons mediates cold- or warm-defensive responses to environmental cooling or warming challenges (Figs 50.1, 50.2). Transection of the neural pathways immediately caudal to the POA or reducing the activity of neurons in the MPO produces hyperthermia by stimulating BAT thermogenesis, metabolism, shivering and CVC [1]. Local warming of the POA inhibits CVC and eliminates shivering, whereas cooling of the local environment of POA neurons evokes BAT and shivering thermogenesis. These findings are indicative of a tonically-active, local-warming-mediated mechanism in the POA capable of driving a potent inhibition of cold-defense effectors responsible for heat conservation and thermogenesis (Fig. 50.2). The neuronal substrate for these effects likely resides in GABAergic, warm-responsive neurons [5,6] in the POA whose tonic discharge is reduced by skin cooling. Whether warm-sensitive POA neurons project axons outside of the POA remains to be demonstrated. BAT and shivering thermogenesis, as well as increases in metabolism and HR that are evoked by skin cooling are blocked by antagonizing GABAA receptors in the MPO [7], suggesting that cutaneous cool signals received by MnPO neurons drive a GABAergic inhibition of inhibitory warmsensitive, MPO projection neurons [4] (Fig. 50.2). Thus, warm-sensitive, GABAergic POA projection neurons integrate cutaneous and local thermal information and are tonically active at thermoneutral temperatures to suppress, to varying degrees, shivering and non-shivering thermogenesis and cutaneous vasoconstriction (Fig. 50.2). Different populations of warm-sensitive, GABAergic POA projection neurons, whose firing rates contribute significantly to the balance of thermoregulatory effector activation that determines core body temperature, are expected to control the activation of different thermal effectors, thereby providing the substrate for the graded thermal thresholds for the cold defense activation of different thermal effectors. Further, the binding of prostaglandin E2 (PGE2), an intermediary in the fever cascade, to inhibitory EP3 receptors on POA inhibitory neurons that project to the dorsomedial hypothalamus (DMH) or to the rostral raphe pallidus (rRPa) region (Fig. 50.2) provides a substrate for the disinhibitory activation of cold defense effectors during fever [1].

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THERmoREgulAToRy EffECToR DRIvE fRom THE DoRsomEDIAl HyPoTHAlAmus

245

FIGURE 50.2 Functional neuroanatomical and neurotransmitter model for the core pathways providing the thermoregulatory control and pyrogenic activation of cutaneous vasoconstriction (CVC), brown adipose tissue (BAT) and shivering thermogenesis. Cool cutaneous thermal signals or prostaglandin (PG) E2 stimulate CVC-mediated heat retention and BAT and shivering thermogenesis. Warm cutaneous thermal signals produce inhibitions of BAT and shivering thermogenesis and of CVC sympathetic outflow, the latter allowing cutaneous vasodilation and thereby promoting heat loss. DRG, dorsal root ganglia; DH, dorsal horn; GLU, glutamate; LPBel, external lateral subnucleus of the lateral parabrachial nucleus; LPBd, dorsal subnucleus of the LPB; POA, preoptic area; MnPO, median preoptic; W-S, warm-sensitive; MPO, medial preoptic; rRPa, rostral raphe pallidus; IML, intermediolateral nucleus; 5-HT, serotonin; DMH, dorsomedial hypothalamus; α, alpha motoneuron; γ, gamma motoneuron.

THERMOREGULATORY EFFECTOR DRIVE FROM THE DORSOMEDIAL HYPOTHALAMUS The dorsal portion of the rostral DMH and the dorsal hypothalamic area contain neurons mediating the BAT thermogenic and HR responses to skin cooling and to injection of PGE2 into the POA [1,7,8]. Activation or

disinhibition of neurons in this region of the DMH elicits potent increases in BAT thermogenesis, HR, and metabolism [1]. In contrast, although activation of DMH neurons can increase CVC sympathetic outflow, the activity of DMH neurons is not required for the cutaneous vasoconstriction stimulated by cooling or by injection of PGE2 into the MPO [8]. Thus, skin cooling- and febrile-evoked BAT and cardiac sympathoexcitatory and somatic shivering

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excitatory signals are respectively transmitted to BAT and cardiac sympathetic and somatic shivering premotor neurons in the rRPa from those DMH neurons that are disinhibited following cold cutaneous or pyrogenic stimuli in the POA (Fig. 50.2). In contrast, the parallel activations of CVC sympathetic outflow are mediated by POA projection neurons that bypass the DMH. Although many POA neurons branch to innervate both the DMH and the rRPa regions, almost none of these express the EP3 receptor, suggesting that at least the febrile activations of BAT and shivering thermogenesis and of CVC are activated by relief of a tonic inhibition from separate populations of POA neurons (Fig. 50.2).

ROSTRAL RAPHE PALLIDUS AREA CONTAINS PREMOTOR NEURONS FOR THERMOREGULATORY EFFECTORS The rRPa is a prominent site of neurons that multi-synaptically innervate BAT, the heart, skeletal muscle fibers and cutaneous blood vessels. Activation or disinhibition of neurons in the rRPa elicits pronounced increases in BAT SNA, BAT thermogenesis, shivering, HR and CVC [9]. Blockade of neuronal activity in the rRPa inhibits and prevents skin-cooling and febrile stimulations of BAT and shivering thermogenesis, of HR and of CVC heat retention [7]. Thus, neurons in the rRPa and the immediately surrounding rostral ventromedial medulla play a key role as sympathetic and somatic premotor neurons controlling BAT and shivering thermogenesis and CVC, providing essential excitatory drives to activate spinal motor networks during cold defense and fever (Fig. 50.2).

SPINAL SYMPATHETIC MECHANISMS CONTROLLING THERMAL EFFECTORS The discharges of BAT, cardiac and CVC sympathetic preganglionic neurons and those of alpha and gamma motoneurons that determine the levels and the rhythmic bursting characteristics of BAT, cardiac and CVC SNAs and of skeletal muscle shivering and, in turn, BAT, cardiac and shivering thermogenesis and cutaneous heat loss, are governed primarily by their supraspinal inputs, but also by the excitability of the networks of spinal interneurons (Fig. 50.2) that influence the discharge of the spinal motor neurons for these thermal effectors. A significant fraction of the BAT, cardiac and CVC sympathetic and the somatic shivering premotor neurons in the rRPa are glutamatergic and/or serotonergic neurons, giving rise to at least a portion of the 5-hydroxytryptamine (5-HT)-containing and VGLUT3-containing terminals in the IML. Spinal glutamate and serotonin receptors in the intermediolateral nucleus (IML) play significant roles in mediating the activation of BAT thermogenesis [10] and CVC, including the cold-evoked and rRPa stimulus-evoked increases in BAT

thermogenesis and CVC. Different synaptic mechanisms and postsynaptic targets are engaged in the spinal 5-HT regulation of different thermal effectors (Fig. 50.2).

SUMMARY The activity of thermal effectors is strongly influenced by shared cutaneous thermal afferent signals in a pathway that includes synapses in the spinal dorsal horn leading to glutamatergic activation of neurons in the LPB, where cool and warm afferent signals are processed by anatomically distinct, POA projecting neurons that influence, in turn, the discharge of different, effector-specific populations of warm-sensitive, GABAergic POA projection neurons, that provide for the integration of local temperature and of peripheral thermal sensory signals. The core efferent pathway for thermoregulatory activation of BAT and shivering thermogenesis and HR involves a tonicallyactive inhibitory input from the POA to sympathoexcitatory neurons in the DMH, which project to sympathetic premotor neurons in the rRPa, which, in turn, provide the excitatory drive to spinal sympathetic preganglionic and somatic motor neurons that is transmitted to brown adipocytes, cardiac pacemaker cells and somatic muscle cells. The core efferent pathway for CVC also involves a tonically-active inhibition emanating from the POA. However, these POA projection neurons send axons to the rRPa where they influence the discharge of CVC sympathetic premotor neurons and, consequently, the level of excitation to CVC sympathetic preganglionic neurons to elicit cutaneous vasoconstriction. Identifying key structures and neurochemical mechanisms within each of these core thermoregulatory pathways provides a framework for understanding how body temperature regulation is influenced by a wide variety of neurotransmitters, peptides, cytokines, and genetic, nutritional and perinatal manipulations, as well as how thermoregulation is integrated with other homeostatic systems regulating oxygen and fuel substrate availability, body water, salt appetite and energy balance.

References [1] Morrison SF, Nakamura K, Madden CJ. Central control of thermogenesis in mammals. Exp Physiol 2008;93:773–97. [2] Nakamura K, Morrison SF. A thermosensory pathway that controls body temperature. Nat Neurosci 2008;11:62–71. [3] Nakamura K, Morrison SF. A thermosensory pathway mediating heat-defense responses. Proc Natl Acad Sci USA 2010;107:8848–53. [4] Nakamura K, Morrison SF. Preoptic mechanism for cold-defensive responses to skin cooling. J Physiol 2008;586:2611–20. [5] Lundius EG, Sanchez-Alavez M, Ghochani Y, Klaus J, Tabarean IV. Histamine influences body temperature by acting at H1 and H3 receptors on distinct populations of preoptic neurons. J Neurosci 2010;30:4369–81. [6] Griffin JD, Kaple ML, Chow AR, Boulant JA. Cellular mechanisms for neuronal thermosensitivity in the rat hypothalamus. J Physiol 1996;492(Pt 1):231–42.

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summARy

[7] Nakamura K, Morrison SF. Central efferent pathways mediating skin cooling-evoked sympathetic thermogenesis in brown adipose tissue. Am J Physiol 2007;292:R127–36. [8] Rathner JA, Madden CJ, Morrison SF. Central pathway for spontaneous and prostaglandin E2-evoked cutaneous vasoconstriction. Am J Physiol 2008;295:R343–354.

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[9] Morrison SF, Sved AF, Passerin AM. GABA-mediated inhibition of raphe pallidus neurons regulates sympathetic outflow to brown adipose tissue. Am J Physiol 1999;276:R290–7. [10] Madden CJ, Morrison SF. Serotonin potentiates sympathetic responses evoked by spinal NMDA. J Physiol 2006;577:525–37.

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C H A P T E R

51 Sweating Phillip A. Low INTRODUCTION

Density and Distribution

There are two types of human sweat glands, apocrine and eccrine. The full complement of eccrine sweat glands is present at birth and undergoes a gradual reduction in numbers with increasing age. It is primarily innervated by sympathetic cholinergic nerve fibers. There is much complexity of the sweat gland. It is adrenergic in utero and undergoes an adrenergic to cholinergic innervational switch during development. Apart from cholinergic and adrenergic innervation of M3 receptors on sweat glands, human eccrine sweat glands are also innervated by VIP, CGRP, and SP fibers. The density of human sweat glands varies greatly, being most dense around the palms. The primary role of sweat glands is thermoregulation. With repeated stimulation, sweat glands undergo hypertrophy. Males have the same number of glands as women, but the volume of each gland is several-fold larger. With denervation, preganglionic or postganglionic, sweat gland size and function become greatly reduced and undergo significant atrophy. Tran-synaptic degeneration has also been suggested. Regulation of sweating in humans serves an important role in thermoregulation (Chapter 50) and loss of the ability to sweat can result in thermoregulatory failure, including heat stroke. The focus of this chapter will be on the anatomy, function, and innervation of the sweat gland.

ANATOMY AND FUNCTION OF THE SWEAT GLAND

Eccrine sweat glands are of greater neuroscience interest and the rest of the description will focus on eccrine sweat glands. They each weigh between 30–40 μg [1]. They first appear in the 3½-month-old fetus in the volar surface of the hands and feet. Eccrine glands show area differences with the greatest density in the palms and soles. They vary in density from 400/cm2 on the palm to about 80/cm2 on the thigh and upper arm. The total numbers are approximately 2 to 5 million. Males and females have the same number of sweat glands. However, the size and volume secreted by each gland is about five times greater in males [2]. Surrounding the secretory cells are myoepithelial cells whose contraction is thought to aid the expulsion of sweat. These glands receive a rich supply of blood vessels and sympathetic nerve fibers but are unusual in that sympathetic innervation is largely cholinergic. The full complement of eccrine glands develops in the embryonic state [3]. No new glands develop after birth.

Physiology of Sweat Glands The physiology of human sweat response is known from the detailed in vitro studies of Sato [4]. Acetylcholine secretion results in the production of an ultrafiltrate (isotonic) by the secretory coil. Directly collected sweat

TABLE 51.1 Comparison of Eccrine with Apocrine Sweat Gland

Type

Parameter

Eccrine

Apocrine

There are two types of sweat glands, eccrine and apocrine. The eccrine sweat glands are simple tubular glands that extend down from the epidermis to the lower dermis. The lower portion is a tightly coiled secretory apparatus consisting of two types of cells. The apocrine gland is a dark basophilic cell that secretes mucous material, and the eccrine sweat gland is a light acidophilic cell that is responsible for the passage of water and electrolytes. Differences between the two types of glands are described in Table 51.1. Apocrine sweat glands are found in the axilla, the anogenital zone, the areola of the nipple, and the external auditory meatus.

Size Duct Ductal opening

Relatively small Long and thin Skin surface (near hair)

Secretory coil

Small ext diameter very narrow lumen Secretory (clear); dark myoepithelial Present

Large Short and thick Directly into upper follicular canal Large ext diameter wide lumen Columnar secretory; myoepithelial Absent

Present at birth Chol  β-adr  α-adr Continuous high Serous

Present at birth Chol  β-adr Intermittent variable Milky protein-rich

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Cell type Intercellular cannaliculi Development Pharmacology Sweat secret rate Secretory product

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samples yield Na and K values identical with plasma. Reabsorption of sodium ions by the eccrine sweat duct results in hypotonic sweat, confirmed in directly collected sweat samples from proximal duct (Na 20–80 mM; K 5–25 mM; [1]). Extracellular Ca2 is important since removal of periglandular Ca2 with EGTA completely inhibits sweat secretion, while Ca ionophore A23187 strongly and persistently stimulates sweating [1]. Magnesium ions appear to be unimportant.

INNERVATION OF SWEAT GLAND

Parameter

Comments

Mental stress

Greatest effect on palmar, sole, axillary sites

Exercise

precedes rise in core temperature

Rehydration

Lowers core temperature

Male sex

Greater sweat gland volume

Race

Blacks  whites but difference quite small

Acclimatization

Increased gain (sweating/temperature change); reduced sweat sodium content

Circadian rhythm

Higher in PM

Seasonal variation

Greater response in winter

Innervation is mainly by sympathetic postganglionic cholinergic fibers. In isolated human eccrine sweat gland regulation, the regulation of sweating is cholinergic and muscarinic, being completely inhibited by atropine [1]. Sudomotor function is metabolically active. It is inhibited by cold, involves active transport, and inhibited by metabolic inhibitors. Microtubules may be important since vinblastine strongly but reversibly inhibits sweating. Endogenous cyclic AMP appears to be the second messenger, since theophylline by phosphodiesterase inhibition markedly increases the sweat response. The muscarinic receptor subtype is M3 [6]. The prostaglandin PGE1 has a sudorific effect in vitro comparable to ACh and was thought to act via cAMP. Histochemical labeling studies show prominent innervation with vasoactive intestinal polypeptide and CGRP fibers and presence of sP and tyrosine hydroxylase fibers [7]. There is, however, dual innervation with a loose network of catecholamine-containing nerves around sweat glands. Innervation of human sweat glands show similarities to rat and mouse sweat glands. In rodents, innervation is initially completely adrenergic followed by an adrenergic to cholinergic switch during development [8]. It is assumed that such a switch occurs in humans during development, so that human sweating is predominantly cholinergic. There is some plasticity and there is some indication that a switch-back to adrenergic sweating can occur with neuropathic states [9]. There are a number of observations that suggest adrenergic innervation increases with certain diseases. Human sweat gland responds to intradermal and intra-arterial adrenaline (10% of cholinergic) [10]. There is evidence of both α-adrenergic (blocked by dibenaline) and β-adrenergic mechanism (blocked by propranolol but not phentolamine). Prominent hyperhidrosis and adrenergic sensitivity occurs in certain neuropathies and Chronic Regional Pain Syndrome I (CRPS I) suggesting increased sympathetic innervation. Indeed the postganglionic sympathetic neuron has recently been shown to enhance adrenergic sweating in CRPS I [9]. In vitro studies suggest the following rank order of sudorific effect ACh  epinephrine (α  β)isoproterenol (β)  phenylephrine (α).

Alcohol and drugs

Cutaneous vasodilatation; reduced hypothalamic set-point

Denervation

Function The major function of the sweat gland in humans is thermoregulatory (see Chapter 50). There are a number of factors that affect the sweat response (Table 51.2). With repeated episodes of profuse sweating, the salt content of the sweat progressively declines. In individuals that have acclimatized to a hot climate, the salt content is reduced, probably reflecting an increase of mineralocorticoids in response to thermal stress [3]. Sweat glands are very prone to atrophy and hypertrophy. Repeated stimulation can result in a several-fold increase in the size and function of the gland. Diffuse loss or absence of sweat can occur due to absence of sweat glands or widespread denervation. Heat intolerance can be a major problem, especially in young patients with widespread anhidrosis, as in the condition chronic idiopathic anhidrosis [5]. TABLE 51.2 Factors that Affect the Sweat Responses FACTORS THAT INCREASE THE SWEAT RESPONSE

FACTORS THAT REDUCE THE SWEAT RESPONSE Parameter

Comments

Skin pressure

Mechanoreceptor stimulation; inhibition of local sympathetic efferents

Hydromeiosis

Water on skin surface reduces sweating rate

Dehydration

Reduced skin blood flow

Hyperosmolarity

Reduced skin blood flow

Cold stimulus

Inhibition of sudomotor activity

Denervation of sweat glands occurs in preganglionic lesions (such as spinal cord injury or multiple system atrophy) or postganglionic lesions (as in the autonomic neuropathies). The size and function of the sweat gland under these circumstances undergoes dramatic atrophy (Fig. 51.1 [4]). Another mechanism of injury is that of transynaptic degeneration of postganglionic axons that could occur with chronic preganglionic lesions. There is also evidence that in early or mild neuropathy affecting the feet, there is excessive forearm response suggesting hypertrophy. The

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InnERvATIon oF SwEAT GlAnd

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References

FIGURE 51.1 Human eccrine sweat gland from a normal subject (A) and a patient with chronic idiopathic anhidrosis (B). There is marked sweat gland atrophy secondary to disuse. D, duct; SC, secretory coil. Reprinted with permission from. Sato K. (1997). Normal and abnormal sweat gland function. In: Clinical Autonomic Disorders: Evaluation and Management. P.A. Low, ed. Philadelphia: Lippincott-Raven, pp. 97–108.

postganglionic sweat response progressively fails with increasing age [11]. The sweat loss is associated with a loss of cholinergic unmyelinated fiber stained with the panaxonal marker PGP9.5 and AChE [12]. In summary, the eccrine sweat gland is an important appendage, subserving thermoregulation. Its absence results in heat intolerance. Alterations in its function provide important clues to the status of the autonomic nervous system.

[1] Sato K, Sato F. Individual variations in structure and function of human eccrine sweat gland. Am J Physiol 1983;245:R203–8. [2] Ogawa T, Low PA. Autonomic regulation of temperature and sweating. In: Low PA, editor. Clinical autonomic disorders: evaluation and management. Boston: Little, Brown and Company; 1993. p. 79–91. [3] Kuno Y. Human perspiration. Springfield, IL: Charles C. Thomas; 1956. [4] Sato K. Normal and abnormal sweat gland function. In: Low PA, editor. Clinical autonomic disorders: evaluation and management. Philadelphia: Lippincott-Raven; 1997. p. 97–108. [5] Low PA, McLeod JG. Autonomic neuropathies. In: Low PA, editor. Clinical autonomic disorders: evaluation and management. Philadelphia: Lippincott-Raven; 1997. p. 463–86. [6] Torres NE, Zollman PJ, Low PA. Characterization of muscarinic receptor subtype of rat eccrine sweat gland by autoradiography. Brain Res 1991;550:129–32. [7] Low PA, Kennedy WR. Cutaneous effectors as indicators of abnormal sympathetic function. In: Morris JL, Gibbins IL, editors. Autonomic innervation of the skin. Amsterdam: Harwood Academic Publishers; 1997. p. 165–212. [8] Landis SC. 1988 Neurotransmitter plasticity in sympathetic neurons. In: Handbook of chemical neuroanatomy: the peripheral nervous system. Amsterdam: Elsevier; p. 65–115. [9] Chemali KR, Gorodeski R, Chelimsky TC. Alpha-adrenergic supersensitivity of the sudomotor nerve in complex regional pain syndrome. Ann Neurol 2001;49:453–9. [10] Sato K. Sweat induction from an isolated eccrine sweat gland. Am J Physiol 1973;225:1147–52. [11] Low PA. The effect of aging on the autonomic nervous system. In: Low PA, editor. Clinical autonomic disorders: evaluation and management. Philadelphia: Lippincott-Raven; 1997. p. 161–75. [12] Abdel-Rahman TA, Collins KJ, Cowen T, Rustin M. Immunohistochemical, morphological and functional changes in the peripheral sudomotor neuro-effector system in elderly people. J Auton Nerv Syst 1992;37:187–97.

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52 Regulation of Metabolism Christopher Bell

CONTRIBUTION OF THE SYMPATHOADRENAL SYSTEM TO COMPONENTS OF TOTAL DAILY ENERGY EXPENDITURE

CONTRIBUTION OF SYMPATHETICALLY STIMULATED ORGANS/TISSUES TO RESTING METABOLIC RATE

The sympathoadrenal system, predominantly via stimulation of β-adrenergic receptors (β-ARs), is a significant biological determinant of total daily energy expenditure [1]. The most compelling evidence comes from animal data: compared with wild-type mice, mice genetically modified such that they do not express any of the three β-AR subtypes demonstrate accelerated weight gain, despite similar energy intake [2]. In humans, the largest component of total daily energy expenditure is resting metabolic rate (RMR). RMR accounts for up to 75% of 24-hour caloric utilization (Fig. 52.1 and Reference [3]). During intra-venous administration of the non-selective β-AR antagonist, propranolol, RMR is decreased [4]. Similarly, during transdermal administration of the centrally-acting, pre-junctional alpha-2-adrenergic receptor (α2-AR) agonist, clonidine, sympathoadrenal activity is inhibited and RMR is decreased [5]. The thermogenic effect of feeding (TEF) is the increase in energy expenditure above RMR following consumption of food. TEF accounts for approximately 10% of total daily energy expenditure and, similarly to RMR, also receives support from the sympathoadrenal system. During sympathoadrenal inhibition or β-AR blockade, TEF is decreased. Further, TEF is positively associated with the thermogenic response to intravenous administration of the non-selective β-AR agonist, isoproterenol [6]. Given the overall contribution of RMR and TEF to total daily energy expenditure, and the important contribution of β-AR stimulation to each of these components, then observations of weight gain in patients prescribed β-AR blockers are perhaps not surprising. From a strictly bioenergetic perspective, the remaining component of total daily energy expenditure, physical activity, is generally unaffected by sympathoadrenal inhibition. During β-AR blockade, submaximal oxygen consumption at the same absolute work rate is not altered, although perceived rates of exertion are increased.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00052-4

In order to appreciate the metabolic demands, and hence energy requirements, of adult humans, the quantification of the metabolic rate of specific organs and tissues is of obvious significance. This quantification has been attempted using a variety of techniques, including the combination of indirect calorimetry, magnetic resonance imaging and dual-energy X-ray absorptiometry [7]. These techniques have revealed that, per unit mass of tissue, the most thermogenic organs/tissues (in descending order) are the heart, kidneys, brain, liver, skeletal muscle and adipose tissue, all of which receive considerable input from the sympathoadrenal system (Fig. 52.2).

MOBILIZATION/UTILIZATION OF SPECIFIC MACRONUTRIENTS: CARBOHYDRATE The liver and pancreas are the tissues most important for mobilization of carbohydrate (glucose). Stimulation of α-ARs on the beta cells of the pancreas inhibits the secretion of insulin, while stimulation of β-ARs on the alpha cells promotes the release of glucagon. In the liver, in addition to the effects of glucagon, stimulation of β-ARs will promote gluconeogenesis and glycogenolysis. With respect to utilization, epinephrine impairs insulin mediated glucose uptake, possibly by inhibition of insulin receptor substrate-1 (IRS-1)-associated activation of phosphatidylinositol 3-kinase (PI3-kinase), decreased glucose transporter protein type-4 (GLUT4) translocation, and/or inhibition of hexokinase and glucose phosphorylation [8].

MOBILIZATION/UTILIZATION OF SPECIFIC MACRONUTRIENTS: LIPID Stimulation of β-ARs in adipose tissue activates hormone sensitive lipase, initiating lipolysis, the breaking down of triacyglycerols into free fatty acids and glycerol.

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52. REgulATIon of METAbolIsM

Once mobilized, these free fatty acids bind to albumin in the blood, and are delivered to organs/tissues. Free fatty acids gain entry into skeletal muscle through various active transport proteins, and subsequent entry into the mitochondria by the use of carnitine pathways. In the mitochondria they are oxidized via beta-oxidation. During β1-AR stimulation, energy expenditure and fatty acid oxidation are increased, however these are both attenuated when

lipolysis is inhibited [9]. This suggests that β1-ARs are more important for mobilization than utilization of lipid. A recent and exciting development pertaining to lipid regulation and thermogenesis is the confirmation of functional brown adipose tissue in humans [10]. Using positron emission tomography (PET), in combination with computed tomography (CT) and administration of [18F] fluoro-deoxyglucose (18F-FDG), metabolically active brown adipose tissue has been detected. Brown adipose tissue appears to be predominantly located in the supraclavicular and anterior neck regions, but also in the anterior thorax. Uptake of 18F-FDG by brown adipose tissue is attenuated during β-AR blockade, suggesting that the brown fat is under sympathoadrenal control.

MOBILIZATION/UTILIZATION OF SPECIFIC MACRONUTRIENTS: PROTEIN

FIGURE 52.1 Contribution of the sympathoadrenal system to components of total daily energy expenditure. % Values represent estimated contribution of each component to total daily energy expenditure. Shaded area represents the contribution of the sympathoadrenal system to each component. TEF: Thermic effect of feeding. Data based on references [3,5,14].

In contrast to the sympathetic regulation of the mobilization/utilization of carbohydrate and lipid, relatively little is known about the sympathetic contribution to protein oxidation. Instead, more is understood about the potential role of the sympathoadrenal system in protein turnover; that is, the balance between protein synthesis and breakdown, and not necessarily the use of protein as a substrate for adenosine tri-phosphate (ATP) production. In skeletal muscle, β2-AR stimulation appears to be important for protein synthesis (anabolic effect), possibly via activation of PI-3 kinase and Akt signaling, and inhibition of calpain

FIGURE 52.2 Contribution of sympathetically stimulated organs/tissues to resting metabolic rate. % Values refer to estimated contribution of specific organs/tissues to resting metabolic rate. Unaccounted % attributed to contribution of residual mass. Listed beneath are the major (metabolic) adrenergic receptors specific to that organ/tissue and their influence on substrate mobilization/utilization. ↑ Stimulation. ↓ Inhibition. Data based on reference [7].

III. AUTONOMIC PHYSIOLOGY

RolE of THE syMPATHoAdREnAl sysTEM In THE dysREgulATIon of METAbolIsM

activity [11]. While this has been clearly demonstrated in animals (mostly rodents), it has been more difficult to document in humans.

ROLE OF THE SYMPATHOADRENAL SYSTEM IN THE DYSREGULATION OF METABOLISM Studying physiological systems during sickness and disease can often provide valuable regulatory information. There are many examples of metabolic dysregulation that lead to a variety of common disease states, including obesity, diabetes, hypercholesterolemia and hypertriglyceridemia. Arguably, the most prevalent of these disease states is obesity. Sympathoadrenal activation is typically high in obesity, with a few notable population exceptions that include the Pima Indians. Visceral obesity is thought to be a particularly important determinant of high sympathoadrenal activation [1], although the reasons for this increased activation remain poorly understood. Several theories exist that are not necessarily mutually exclusive: increased adipose tissue leads to greater secretion of various adipocyte-derived and/ or adiposity-associated hormones including leptin, insulin and angiotensin II; all of which are able to stimulate direct central sympathetic outflow. Further, insulin has vasodilatory properties and thus may also increase sympathoadrenal activation via a baroreflex mediated response. Given the role of the sympathoadrenal system in lipid mobilization/utilization and adipocyte proliferation [12], elevated sympathoadrenal activity might intuitively seem like a favorable response to obesity. Unfortunately this persistently high degree of β-AR stimulation eventually leads to β-AR desensitization and down-regulation. Recent evidence [13] suggests obesity-associated sympathoadrenal activation evokes the cyclic-adenosine monophosphate (cAMP) response element binding protein (CREB) coactivator, CRTC3. This, in turn, leads to upregulation of the GTP-ase activating protein, Rgs2, thereby inhibiting adenyl cyclase activity. Consequently, greater than usual sympathoadrenal stimulation is required in order to elicit a given β-AR mediated response, leading to a vicious cycle of further desensitization and increased tonic activation. New

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data suggest that the thermogenic response to β-AR stimulation is augmented following short-term sympathoadrenal inhibition [5]. Clearly the sympathoadrenal system remains an important therapeutic target for obesity-intervention.

References [1] Davy KP, Orr JS. Sympathetic nervous system behavior in human obesity. Neurosci Biobehav Rev 2009;33:116–24. [2] Bachman ES, Dhillon H, Zhang CY, Cinti S, Bianco AC, Kobilka BK, et al. betaAR signaling required for diet-induced thermogenesis and obesity resistance. Science 2002;297:843–5. [3] Ravussin E, Lillioja S, Knowler WC, Christin L, Freymond D, Abbott WG, et al. Reduced rate of energy expenditure as a risk factor for body-weight gain. N Engl J Med 1988;318:467–72. [4] Monroe MB, Seals DR, Shapiro LF, Bell C, Johnson D, Jones PP. Direct evidence for tonic sympathetic support of resting metabolic rate in healthy adult humans. Am J Physiol Endocrinol Metab 2001;280:E740–4. [5] Newsom SA, Richards JC, Johnson TK, Kuzma JN, Lonac MC, Paxton RJ, et al. Short-term sympathoadrenal inhibition augments the thermogenic response to beta-adrenergic receptor stimulation. J Endocrinol 2010;206:307–15. [6] Stob NR, Bell C, van Baak MA, Seals DR. Thermic effect of food and beta-adrenergic thermogenic responsiveness in habitually exercising and sedentary healthy adult humans. J Appl Physiol 2007;103:616–22. [7] Wang Z, Ying Z, Bosy-Westphal A, Zhang J, Schautz B, Later W, et al. Specific metabolic rates of major organs and tissues across adulthood: evaluation by mechanistic model of resting energy expenditure. Am J Clin Nutr 2010;92:1369–77. [8] Hunt DG, Ivy JL. Epinephrine inhibits insulin-stimulated muscle glucose transport. J Appl Physiol 2002;93:1638–43. [9] Schiffelers SL, Brouwer EM, Saris WH, van Baak MA. Inhibition of lipolysis reduces beta1-adrenoceptor-mediated thermogenesis in man. Metabolism 1998;47:1462–7. [10] Lee P, Greenfield JR, Ho KK, Fulham MJ. A critical appraisal of the prevalence and metabolic significance of brown adipose tissue in adult humans. Am J Physiol Endocrinol Metab 2010;299:E601–6. [11] Koopman R, Gehrig SM, Leger B, Trieu J, Walrand S, Murphy KT, et al. Cellular mechanisms underlying temporal changes in skeletal muscle protein synthesis and breakdown during chronic {beta}adrenoceptor stimulation in mice. J Physiol 2010;588:4811–23. [12] Foster MT, Bartness TJ. Sympathetic but not sensory denervation stimulates white adipocyte proliferation. Am J Physiol Regul Integr Comp Physiol 2006;291:R1630–R1637. [13] Song Y, Altarejos J, Goodarzi MO, Inoue H, Guo X, Berdeaux R, et al. CRTC3 links catecholamine signalling to energy balance. Nature 2010;458:933–41. [14] Tappy L. Thermic effect of food and sympathetic nervous system activity in humans. Reprod Nutr Dev 1996;36:391–7.

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53 Autonomic Innervation of the Skeleton Florent Elefteriou, J. Preston Campbell BONE SYMPATHETIC INNERVATION: ONTOGENY AND ANATOMY Autonomic innervation of the skeleton is documented at the gross anatomical and histological levels with both long and flat bones being innervated with myelinated sensory and unmyelinated sympathetic nerve fibers [1–3]. Retrograde tracing and axotomy experiments have demonstrated that these intraosseus nerve fibers are structurally and functionally connected to primary afferent neurons in dorsal root ganglia and paravertebral sympathetic ganglia [4]. Although the terminal nerve branches feeding the bones have been largely unstudied, they are likely derived from the overlying muscle innervations. Postganglionic fibers from cervical sympathetic ganglion and glossopharyngeal nerve innervate external and internal bones of the skull. The long bones of the upper extremities receive nerve supply from the brachial plexus which then branches to the median nerve to innervate the humerus and the ulnar and radian nerves which supply the forearm bones. Osseus innervation of the flat rib bones is achieved via the anterior branches of the 12 pairs of intercostals nerves. Sympathetic innervation of the lower limbs originates in the lumbar plexus which supplies the femoral and deep saphenous nerves to the femur, and the tibial, medial, and popliteal nerves to the tibia and fibula. Basivertebral nerves in the spine supply intraosseus autonomic innervations of the vertebral bodies [5]. Due to the inherent technical difficulties of working with osseous tissue, little research exists concerning the development of bone innervation. In rats, nerves penetrate uncalcified osseous tissues relatively late, after gestational day 17, and continue to develop more fully after birth [4]. These nerve fibers follow alongside blood vessels and grow into the bone presumably in response to bone marrow expression of neurotrophic factors [6] though this has not been demonstrated in vivo. Despite their presence in developing embryonic tissues, it is still unclear whether sympathetic nerves have a functional role in early bone developmental processes. The lack of obvious bone developmental or morphogenesis abnormalities in mutant mouse models lacking Dopamine β-hydroxylase, the β1, 2 and 3-adrenergic receptor, NPY or the Y1, 2 or 4 receptors for instance argues against a major role of sympathetic nerves during development, despite the

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00053-6

demonstrated need for NE for embryonic cardiovascular development [7]. The first detailed description and histology of bone innervation was given by de Castro (1930) who developed a silver staining technique that demonstrated ramification of neurons with mesenchymal cells in bone. Subsequent research has shown that intraosseous nerves express tyrosine hydroxylase [8], indicating catecholamine synthesis, PGP 9.5, vasoactive intestinal peptide, calcitonin related gene product, and substance P, indicating both sensory and autonomic innervation (see [9] for review). The location of these neural markers show that neurosteal fibers can be divided into two major groups: marrow innervation and periosteal innervation (Fig. 53.1A). Most nerve fibers in the periosteum are sensory, but autonomic TH-positive fibers are also present. These sympathetic nerves branch around the periosteum and enter the cortical bone where they are organized parallel to the long axis of the bone in the Haversian canals or perpendicular, alongside Sharpey’s fibers, which anchor the periosteum to the cortical bone (Fig. 53.1B). In the marrow, nerves accompany blood vessels through a nutrient foramen into the bone’s interior where they can be seen to branch and make vascular and cellular contacts (Fig. 53.1C). The majority of these nerves are perivascular, forming spiral patterns around blood vessels, and can be categorized as vasomotor, controlling the vascular hemodynamics in the bone marrow. Schwann cells are also present in the marrow though they are believed to accompany sensory nerves rather than the sympathetic axons. Electronic microscopy studies have shown nerve fibers at the proximity of osteoblasts but the existence of synapses has not been demonstrated (Fig. 53.1D) [10].

EFFECT OF SYMPATHETIC NERVES ON BONE REMODELING The cells forming bone and allowing its constant remodeling during adult life include osteoblasts, the bone-forming cells of mesenchymal origin, osteoclasts, the bone resorbing cells of monocytic origin, and osteocytes, which are fully mature and bone matrix-embedded osteoblasts. Both osteoblasts and osteoclasts express the

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recognized, and the effect of sympathetic nerves on both lineages, hematopoietic and mesenchymal, is likely to be interrelated. The recently identified effect of sympathetic nerves on hematopoietic stem cell egress from the bone marrow, mediated by osteoblasts, supports this hypothesis [15]. NPY is co-released with NE from sympathetic nerves and a role of this neuropeptide in the regulation of bone remodeling is also well established. The Y2 receptor expressed by hypothalamic neurons exerts an inhibitory influence of bone formation, as demonstrated by the high bone mass phenotype of mice lacking the Y2 receptor specifically in these neurons [16]. On the other hand, the Y1 receptors expressed by bone stromal cells appear to regulate the number of mesenchymal stem cells in bone, downstream of NPY signals from bone sympathetic nerves [17]. Surprisingly, NPY is also expressed by osteocytes, which are fully differentiated osteoblasts embedded within bone matrix [18].

RELEVANCE TO BONE PHYSIOLOGY AND DISEASES

FIGURE 53.1 Intraosseal innervation. Branches of the tibial nerve (yellow) supply the bones of the lower limb (A) alongside the post. tibial and peroneal branches of the popliteal artery (red). Blood vessels and nerves enter tibia (arrow) through the nutrient foramen. Neurosteal fibers in the periosteum run parallel with Sharpey’s fibers and throughout the canaliculi of the cortex (B). Most nerve fibers in the central canal are perivascular vasomotor nerves (C) though some branch into the bone. Some sympathetic fibers ramify near mesenchymal cells such as osteoblasts (D), but whether these derive from the periosteal or perivascular nerves is unknown.

β2AR and are thus responsive to catecholamines [10]. Catecholamines and adrenergic receptor agonists indeed potently increase intracellular cAMP levels in osteoblasts and trigger changes in gene expression for osteoclastogenesis (RANKL) and proliferation (G1 cyclin and Ap1) related genes [11,12]. In agreement with these cellular effects, isoproterenol administration in rodents causes severe bone loss caused by increased bone resorption and decreased bone formation [10,13]. On the other hand, pharmacological blockade of the βAR by propranolol increases bone mass in mice and rats by inhibiting bone resorption and promoting bone formation [10,14]. The use of mutant mouse models corroborated these findings, with the demonstration that genetic lack of Dopamine β-hydroxylase (DBH) or of the β2AR in mice causes an increase in bone mass [10,11]. In addition to their structural function, bones are also hematopoietic tissues and host a wide variety of immune cells at different stages of differentiation, most of which express βARs. The fact that immune cells are involved in the complex regulation of bone remodeling is increasingly

Despite the fairly well established nature of these findings in murine models, it is still unclear to what extent sympathetic signaling regulates bone mass or contributes to bone pathologies in humans. Most studies in humans are restricted to the effect of β-blockers on bone mineral density and tend to support a protective effect of this class of drugs on bone mineral density [19]. They did not yet investigate drugs or conditions influencing sympathetic outflow or responsiveness. Sympathetic signaling in bone may be regulated by several physiological and/or pathological signals. Activation of the cannabinoid receptor 1 (CB1) by 2-arachidonoylglycerol (2AG) may represent such a signal since it regulates sympathetic signaling at the presynaptic level in bone nerve endings by reducing NE release and the antiosteogenic effect of the sympathetic nervous system [20]. Glucocorticoids on the other hand may promote sympathetic responsiveness by osteoblasts to induce bone loss, by their action on β2AR expression and signaling [21]. The effect of thyroid hormones on bone may involve a similar mechanism, although this has not been demonstrated yet. Sympathetic outflow is also regulated centrally and particularly by brainstem and hypothalamic centers controlling major bodily homeostatic functions, including body weight and reproduction. The observation that β-blockade by propranolol or β2AR deficiency prevented the bone catabolic effect of leptin hypothalamic administration in mice clearly linked hypothalamic neurons to the regulation of bone remodeling, and functionally positioned the skeleton as a target organ of central neurons, via the sympathetic nervous system [11] (Fig. 53.2). Severe stress and depression are two CNS pathological conditions that may stimulate sympathetic outflow and have effects on bone via this mechanism, as suggested by a study in mice [22] and by

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RElEvAnCE To BonE PHySIology And dISEASES

FIGURE 53.2 The sympathetic nervous system links central neurons to bone cells and regulates the endocrine function of the skeleton. HSC, hematopoietic stem cell; NE, norepinephrine.

the association between clinically diagnosed depression and low BMD [23]. Lastly, the skeleton is increasingly recognized as a true endocrine organ, whose secretion is controlled by sympathetic nerves. Osteocalcin secretion by osteoblasts is indeed controlled by catecholamines and regulates glycemia and testosterone production [24,25].

References [1] Hurrell DJ. The Nerve Supply of Bone. J Anat 1937;72(Pt 1):54–61. PMCID: PMC1252438. [2] Calvo W. The innervation of the bone marrow in laboratory animals. Am J Anat 1968;123(2):315–28. [3] Duncan CP, Shim SS. J. Edouard Samson Address: the autonomic nerve supply of bone. An experimental study of the intraosseous adrenergic nervi vasorum in the rabbit. J Bone Joint Surg Br 1977;59(3):323–30. [4] Gajda M, Litwin JA, Tabarowski Z, Zagolski O, Cichocki T, Timmermans JP, et al. Development of rat tibia innervation: colocalization of autonomic nerve fiber markers with growth-associated protein 43. Cells Tissues Organs 2010;191(6):489–99. [5] Antonacci MD, Mody DR, Heggeness MH. Innervation of the human vertebral body: a histologic study. J Spinal Disord 1998;11(6):526–31. [6] Wang J, Ding F, Gu Y, Liu J, Gu X. Bone marrow mesenchymal stem cells promote cell proliferation and neurotrophic function of Schwann cells in vitro and in vivo. Brain Res 2009;1262:7–15. [7] Thomas SA, Matsumoto AM, Palmiter RD. Noradrenaline is essential for mouse fetal development. Nature 1995;374(6523):643–6. [8] Bjurholm A, Kreicbergs A, Terenius L, Goldstein M, Schultzberg M. Neuropeptide Y-, tyrosine hydroxylase- and vasoactive intestinal polypeptide-immunoreactive nerves in bone and surrounding tissues. J Auton Nerv Syst 1988;25(2–3):119–25.

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[9] Elefteriou F. Neuronal signaling and the regulation of bone remodeling. Cell Mol Life Sci 2005;62(19–20):2339–49. [10] Takeda S, Elefteriou F, Levasseur R, Liu X, Zhao L, Parker KL, et al. Leptin regulates bone formation via the sympathetic nervous system. Cell 2002;111(3):305–17. [11] Elefteriou F, Ahn JD, Takeda S, Starbuck M, Yang X, Liu X, et al. Leptin regulation of bone resorption by the sympathetic nervous system and CART. Nature 2005;434(7032):514–20. [12] Fu L, Patel MS, Bradley A, Wagner EF, Karsenty G. The molecular clock mediates leptin-regulated bone formation. Cell 2005;122(5):803–15. [13] Bonnet N, Benhamou CL, Brunet-Imbault B, Arlettaz A, Horcajada MN, Richard O, et al. Severe bone alterations under beta2 agonist treatments: bone mass, microarchitecture and strength analyses in female rats. Bone 2005;37(5):622–33. [14] Bonnet N, Benhamou CL, Malaval L, Goncalves C, Vico L, Eder V, et al. Low dose beta-blocker prevents ovariectomy-induced bone loss in rats without affecting heart functions. J Cell Physiol 2008;217(3):819–27. [15] Katayama Y, Battista M, Kao WM, Hidalgo A, Peired AJ, Thomas SA, et al. Signals from the sympathetic nervous system regulate hematopoietic stem cell egress from bone marrow. Cell 2006;124(2):407–21. [16] Baldock PA, Allison S, McDonald MM, Sainsbury A, Enriquez RF, Little DG, et al. Hypothalamic regulation of cortical bone mass: opposing activity of y2 receptor and leptin pathways. J Bone Miner Res 2006;21(10):1600–7. [17] Lundberg P, Allison SJ, Lee NJ, Baldock PA, Brouard N, Rost S, et al. Greater bone formation of Y2 knockout mice is associated with increased osteoprogenitor numbers and altered Y1 receptor expression. J Biol Chem 2007;282(26):19082–091. [18] Igwe JC, Jiang X, Paic F, Ma L, Adams DJ, Baldock PA, et al. Neuropeptide Y is expressed by osteocytes and can inhibit osteoblastic activity. J Cell Biochem 2009;108(3):621–30. PMCID: 2754602 [19] de Vries F, Souverein PC, Cooper C, Leufkens HG, van Staa TP. Use of beta-blockers and the risk of hip/femur fracture in the United Kingdom and The Netherlands. Calcif Tissue Int 2007;80(2):69–75. [20] Tam J, Ofek O, Fride E, Ledent C, Gabet Y, Muller R, et al. Involvement of neuronal cannabinoid receptor CB1 in regulation of bone mass and bone remodeling. Mol Pharmacol 2006;70(3):786–92. [21] Ma Y, Nyman JS, Tao H, Moss HH, Yang X, Elefteriou F. β2-Adrenergic receptor signaling in osteoblasts contributes to the catabolic effect of glucocorticoids on bone. Endocrinology 2011; 152(4): 1412-22. [22] Yirmiya R, Goshen I, Bajayo A, Kreisel T, Feldman S, Tam J, et al. Depression induces bone loss through stimulation of the sympathetic nervous system. Proc Natl Acad Sci U S A 2006;103(45):16876–881. [23] Cizza G, Ravn P, Chrousos GP, Gold PW. Depression: a major, unrecognized risk factor for osteoporosis? 2001;12(5):198–203. Trends Endocrinol Metab 2001;12(5):198–203. [24] Hinoi E, Gao N, Jung DY, Yadav V, Yoshizawa T, Myers Jr. MG, et al. The sympathetic tone mediates leptin's inhibition of insulin secretion by modulating osteocalcin bioactivity. J Cell Biol 2008;183(7):1235–42. [25] Oury F, Sumara G, Sumara O, Ferron M, Chang H, Smith CE, et al. Endocrine Regulation of Male Fertility by the Skeleton. Cell 2011.

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54 Sex Differences in Autonomic Function Emma C. Hart, Nisha Charkoudian, Michael J. Joyner INTRODUCTION In humans, both normal and pathophysiological mechanisms of autonomic control of blood pressure are modified by sex. The incidence of hypertension is lower in young women than in young men, and the incidence of “hypotensive” disorders such as orthostatic intolerance is much greater in young women. However, even in healthy normotensive humans, resting autonomic tone and autonomic support of blood pressure tend to be different between men and women. Interestingly, aging increases the risk of developing hypertension and other cardiovascular diseases in both sexes, but this effect is even more marked in women after the menopause than it is in men of a similar age range. In the context of this discussion, there has been some debate regarding whether the term “gender” or “sex” should be used when referring to differences between men and women, particularly with regard to biomedical findings. The word “gender” originates from the old French word “gendre”, referring to a grammatical category which indicates whether a word is masculine or feminine. More recently (over the last century) the word gender is typically used to refer to the general sociological roles of females and males, and a person who is biologically of one sex or the other can identify with either gender. In contrast, sex differences refer to biological differences based on reproductive organs and function. In the present chapter, the word “sex” is used to refer to biological differences between men and women. The cultural and sociological influences of gender (i.e., differences based on how a person defines him or herself by gender) are outside the scope of the present discussion.

SEX DIFFERENCES IN NORMAL AUTONOMIC FUNCTION Tonic sympathetic nerve activity can be measured using microneurography to measure muscle sympathetic nerve activity (MSNA) or norepinephrine spill-over techniques [1,2]. Studies utilizing either approach typically indicate that resting sympathetic nerve activity is lower in young women compared to men [3]. In addition, baseline MSNA

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00054-8

is altered during different phases of the menstrual cycle, where MSNA is higher during the luteal (high hormone) compared to the early follicular (low hormone) phase in women not taking oral contraceptives [4]. These differences in MSNA appear to be partially mediated via the effects of the female sex hormones on central autonomic nuclei [5]. Central estrogen administration also increases resting vagal tone in female mice, suggesting the sex hormones might also influence parasympathetic outflow to the heart [5]. However, whether resting cardiac vagal tone is different in men and women is unclear, with some studies (using heart rate variability analysis) reporting higher vagal tone in women vs. men [6] and others indicating that there are no differences between men and women [7]. Despite the fact that MSNA is tightly linked to arterial pressure via the baroreflex, tonic levels of MSNA do not determine baseline arterial pressure in young normotensive people. In young men and women, there is no relationship between tonic MSNA and resting arterial pressure [1,8,9] (Fig. 54.1). Investigations into inter-individual differences in arterial blood pressure regulation have been essential in explaining this apparent paradox. In young men, MSNA is positively related to TPR and inversely related to cardiac output [8] (Fig. 54.1). Therefore, lower cardiac output values balance high levels of MSNA, (which contribute to high TPR) in young men, thus explaining the lack of direct relationship between MSNA and arterial pressure in young men. Interestingly, in young women, MSNA is not related to TPR or cardiac output (Fig. 54.1). Consequently, MSNA does not appear to determine the overall level of peripheral vasoconstrictor tone in young women. Therefore, in young women other factors must offset the vasoconstrictor influence of sympathetic vasoconstrictor nerves. Other mechanisms may relate to the potent vasodilator effect of estrogen (for review see ref [10]) and differences in β-adrenergic receptor sensitivity between men and women [11]. Because norepinephrine released from adrenergic nerves can stimulate both the α- and β-adrenergic receptors, α-adrenergic vasoconstriction may be offset by increased β-adrenergic receptor mediated vasodilation in young women. In this context, recent data suggest that when the β-adrenergic receptors are blocked, the relationship between MSNA and TPR becomes positive in young women [12].

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FIGURE 54.1 Relationship of muscle sympathetic nerve activity (MSNA) to mean arterial pressure (MAP) cardiac output (CO) and total peripheral resistance among young men and women. There is no relationship between MSNA and MAP among men and women. However, in young men MSNA is positively related to TPR and inversely related to CO. In young women, there is no such relationship. Modified from Hart et al. [9].

ORTHOSTATIC INTOLERANCE AND HYPOTENSION Both orthostatic intolerance and orthostatic hypotension are more common in young women compared to

young men. Normotensive, otherwise healthy women, are less tolerant to changes in central blood volume compared to men [13]. That is, young women tend to develop symptoms (dizziness, lightheadedness, or fainting) earlier during orthostatic stress such as lower body negative

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ovERAll SummARy AnD ConCluSIonS

pressure. This tendency is exacerbated after spaceflight or prolonged bed rest [13]. Moreover, individuals suffering from postural orthostatic tachycardia syndrome (POTS) are most often young women under the age of 35 years. Individuals with POTS tend to have similar MSNA responses to orthostatic stress as age matched controls, suggesting that lower MSNA is not a major contributor to POTS symptoms. Patients with POTS have a smaller stroke volume and blood volume compared to healthy controls [14], which might explain why more women suffer from POTS; i.e., women have smaller stroke volumes and blood volumes vs. men (due to smaller body size). Consequently, a chronic decrease in stroke volume (due to various reasons including physical de-conditioning) may be less well tolerated in women and thus more women express symptoms of POTS. A recent study suggests that exercise training improves left ventricular mass and blood volume in women with POTS and can improve or even cure the symptoms of POTS [14]. It is well accepted that even healthy young women who do not have symptoms of POTS are predisposed to orthostatic intolerance or hypotension at a greater rate compared to men. The exact mechanisms underlying this predisposition are unclear. First, differences in baroreflex function in men and women could explain why more women experience syncope during head-up-tilt or lower body negative pressure. However, this is unlikely since the sympathetic responses and increases in total peripheral resistance to postural stress in women tend to be similar to that in men [15]. Furthermore, studies indicate that sympathetic baroreflex sensitivity is not different between men and women. In women, however, the heart rate response to orthostatic stress appears to be greater than that in men [13]. This may be due to differences in the cardiac physiology between men and women. Along these lines, women have a lower stroke volume and stroke index preceding syncope vs. men [13]. Furthermore, reduced orthostatic tolerance in women appears to be associated with a steeper Frank–Starling curve; that is, for a given decrease in pulmonary capillary wedge pressure, there is a larger decrease in stroke volume in women vs. men [13]. Finally, the withdrawal of sympathetic nerve activity precipitates, and invariably results in, syncope. It is possible that women reach the threshold for sympathetic withdrawal earlier than men due to larger decreases in cardiac filling during orthostatic stress [13,15].

SEX, AGING AND HYPERTENSION The sympathetic nerve system is involved in the pathogenesis of hypertension, with many studies demonstrating that sympathetic nerve activity is elevated among individuals with hypertension. In general young men are more at risk of developing hypertension than women of the same age, which may be related to differential effects that the sympathetic nerves appear to have on the vasculature of

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men and women. Aging is associated with an increased risk of developing hypertension. In this context, aging is associated with a gradual augmentation in sympathetic nerve activity and arterial pressure [1]. Although SNA is not related to blood pressure in young men or women, this relationship becomes significantly positive in older men and women, so that higher sympathetic nerve activity is related to increased arterial pressure [3]. This might explain why arterial pressure is typically higher in older men and women vs. young men and women. The slope of this relationship between arterial pressure and MSNA is steeper in older women, so that for a given increase in MSNA there is a greater increase in arterial pressure in women vs. men of the same age. Consequently, sympathetic nerve activity becomes more important in determining resting arterial pressure in older women. This might be explained by the loss of the protective influence of estrogen after menopause. Along these lines, the risk of developing hypertension is greater in postmenopausal women than in men of the same age.

OVERALL SUMMARY AND CONCLUSIONS In summary, sex and sex hormones have substantial influences on the interaction between the autonomic and cardiovascular systems, such that resting arterial pressure is controlled differently in men and women across the lifespan. In young women, the sex hormones modulate the transduction of sympathetic nerve activity into peripheral vasoconstriction. “Hypotensive” disorders such as orthostatic intolerance and orthostatic hypotension occur more frequently in young women than in young men. Although women tend to have lower resting sympathetic vasoconstrictor nerve activity, this may not be a major contributor to orthostatic intolerance, as recent data suggest that, orthostatic intolerance/hypotension in women appears to be primarily due to less cardiac filling and/or a smaller cardiac size. Interestingly, as humans age, the overall level of arterial pressure becomes related to sympathetic nerve activity; this relationship is especially prominent in women. This might explain why the increase in the rate of hypertension is greater in older women after the menopause vs. men. Overall, understanding sex-based differences in autonomic function is becoming fundamental for the treatment of autonomic dysfunction and related disorders such as hypertension.

References [1] Sundlof G, Wallin BG. Human muscle nerve sympathetic activity at rest. Relationship to blood pressure and age. J Physiol 1978;274:621–37. [2] Wallin BG, Thompson JM, Jennings GL, Esler MD. Renal noradrenaline spillover correlates with muscle sympathetic activity in humans. J Physiol 1996;491(Pt 3):881–7. [3] Narkiewicz K, Phillips BG, Kato M, Hering D, Bieniaszewski L, Somers VK. Gender-selective interaction between aging,

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[4]

[5]

[6]

[7]

[8]

[9]

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blood pressure, and sympathetic nerve activity. Hypertension 2005;45:522–5. Minson CT, Halliwill JR, Young TM, Joyner MJ. Influence of the menstrual cycle on sympathetic activity, baroreflex sensitivity, and vascular transduction in young women. Circulation 2000;101:862–8. Saleh TM, Connell BJ. 17 beta-estradiol modulates baroreflex sensitivity and autonomic tone of female rats. J Auton Nerv Syst 2000;80:148–61. Liao D, Barnes RW, Chambless LE, Simpson Jr. RJ, Sorlie P, Heiss G. Age, race, and sex differences in autonomic cardiac function measured by spectral analysis of heart rate variability – the ARIC study. Atherosclerosis Risk in Communities. Am J Cardiol 1995;76:906–12. Evans JM, Ziegler MG, Patwardhan AR, Ott JB, Kim CS, Leonelli FM, et al. Gender differences in autonomic cardiovascular regulation: spectral, hormonal, and hemodynamic indexes. J Appl Physiol 2001;91:2611–8. Charkoudian N, Joyner MJ, Johnson CP, Eisenach JH, Dietz NM, Wallin BG. Balance between cardiac output and sympathetic nerve activity in resting humans: role in arterial pressure regulation. J Physiol 2005;568:315–21. Hart EC, Charkoudian N, Wallin BG, Curry TB, Eisenach JH, Joyner MJ. Sex differences in sympathetic neural-hemodynamic balance: implications for human blood pressure regulation. Hypertension 2009;53:571–6.

[10] Miller VM, Duckles SP. Vascular actions of estrogens: functional implications. Pharmacol Rev 2008;60:210–41. [11] Kneale BJ, Chowienczyk PJ, Brett SE, Coltart DJ, Ritter JM. Gender differences in sensitivity to adrenergic agonists of forearm resistance vasculature. J Am Coll Cardiol 2000;36:1233–8. [12] Hart EC, Charkoudian N, Wallin BG, Roberts SK, Johnson CP, Joyner MJ. Sex differences in the sympathetic balance of blood pressure: the role of the alpha-adrenergic receptors. FASEB J 2010;24:594–9. [13] Fu Q, Arbab-Zadeh A, Perhonen MA, Zhang R, Zuckerman JH, Levine BD. Hemodynamics of orthostatic intolerance: implications for gender differences. Am J Physiol Heart Circ Physiol 2004;286:H449–57. [14] Fu Q, Vangundy TB, Galbreath MM, Shibata S, Jain M, Hastings JL, et al. Cardiac origins of the postural orthostatic tachycardia syndrome. J Am Coll Cardiol 2010;55:2858–68. [15] Fu Q, Witkowski S, Okazaki K, Levine BD. Effects of gender and hypovolemia on sympathetic neural responses to orthostatic stress. Am J Physiol Regul Integr Comp Physiol 2005;289:R109–16.

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55 Autonomic Control During Pregnancy Virginia L. Brooks, Belinda H. McCully, Priscila A. Cassaglia The physiological adaptations to pregnancy are profoundly complex yet exquisitely tuned to ensure adequate development of the fetus. These changes are largely orchestrated directly or indirectly by the placenta, which is a source of a broad array of hormones, including neuropeptides, steroid hormones, pituitary-like hormones, growth factors, vasoactive hormones, and metabolic hormones. The cardiovascular system and its autonomic innervation are particularly impacted. An early event in gestation is systemic vasodilation (Fig. 55.1), which occurs even before uteroplacental vascular growth and blood flow are significantly increased [1,2]. As a result, arterial pressure falls. However, the decrease is modest because of parallel increases in cardiac output (Fig. 55.1), due in part to cardiac remodeling and enlargement, but also to simultaneous activation of fluid-retaining factors, such as the renin-angiotensin-aldosterone system, and increases in blood volume [2].

PREGNANCY ACTIVATES THE SYMPATHETIC NERVOUS SYSTEM In parallel to increases in fluid-retaining hormones, alterations in the autonomic nervous system accompany normal pregnancy. Using microneurography in humans, several studies have documented that muscle sympathetic nerve activity is increased [3,4]. In experimental animals, elevations of basal renal sympathetic nerve activity have also been observed [3]. In line with these direct measurements of sympathetic nerve firing rates, indirect assessments, including quantification of heart rate and arterial pressure variability in both the frequency and time domains [3,5], also suggest that pregnancy increases basal sympathetic activity. Activation of the sympathetic nervous system would be expected to cause vasoconstriction, which could nullify the primary decreases in systemic vascular resistance. However, this effect is minimized during pregnancy, because the vasculature becomes resistant to the actions of vasoconstrictors, including norepinephrine [3]. While pregnancy induces sympathoexcitation, simultaneously, basal parasympathetic tone decreases [5,6]. One consequence of increased cardiac sympathetic activity, and decreased parasympathetic activity, is increased heart rate, which is also evident early in gestation (Fig. 55.1) [1].

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00055-X

The mechanisms by which pregnancy modifies basal autonomic tone have received limited attention. Central actions of increases in circulating angiotensin II appear to contribute to the sympathoexcitation [3]. In contrast, decreased parasympathetic control of the heart is due in part to reductions in the responsiveness of the heart to acetylcholine [6].

PREGNANCY IMPAIRS THE BARORECEPTOR REFLEX While most physiological changes induced by pregnancy are relatively benign, one deleterious consequence is a marked suppression of the function of the baroreceptor reflex [3–5]. As a result, pregnant women are prone to orthostatic hypotension, and pregnant animals are less able to maintain arterial pressure during hemorrhage [3]. Given that hemorrhage accompanies every delivery and that the baroreflex is largely responsible for arterial pressure maintenance during hemorrhage, it is understandable why peripartum hemorrhage is a major cause of maternal mortality [3]. Baroreflex dysfunction has been documented in several species besides humans, including rabbits, rats, goats, sheep, and dogs [3]. Pregnancy attenuates baroreflex control of multiple efferents, such as renal and muscle sympathetic nerve activity, heart rate, as well as hormones like vasopressin and ACTH [3]. The decrease in baroreflex control of the autonomic nervous system has been detected using several methodological approaches, including noninvasive techniques in women employing the analysis of heart rate and arterial pressure variability, assessments of spontaneous baroreflex sensitivity, and infusion of vasoactive drugs to construct complete sigmoidal baroreflex relationships between arterial pressure and heart rate or sympathetic nerve activity (Fig. 55.2) [3,5]. Three features of these curves are commonly attenuated: the maximum gain or slope of the most linear segment of the curves, the maximal level of sympathetic activity or heart rate achieved during severe hypotension, and the “setpoint” or the arterial pressure level associated with the midpoint of the curve. This latter change is likely mediated by resetting of baroreceptor afferents, which causes the baroreflex function curve to shift toward the lower arterial pressure

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FIGURE 55.1 Hemodynamic changes that accompany normal pregnancy in humans. Data were taken from ref [1]. CO, cardiac output; SV, stroke volume; HR, heart rate; MAP, mean arterial pressure; TPR, total peripheral resistance.

FIGURE 55.2 Pregnancy impairs baroreflex control of heart rate and renal sympathetic nerve activity (RSNA) in conscious rats. From Brooks et al. [3]. P, pregnant; NP, nonpregnant.

level during pregnancy [3]. However, the mechanisms that contribute to the decreases in the baroreflex maximums and in baroreflex gain are complex and may be largely distinct.

MECHANISMS OF PREGNANCY-INDUCED BAROREFLEX IMPAIRMENT While pregnancy could depress the function of any or all anatomical links within the baroreflex pathway, current evidence indicates that brain control is particularly impaired [3]. Within the brain, multiple sites and hormonal mediators are involved. As detailed in Chapter 33, the core brainstem baroreflex pathway begins in the nucleus tractus solitarius (NTS), which receives baroreceptor afferent information. Increases in arterial pressure activate the baroreceptors, which excite NTS second order neurons via a glutamatergic (non-NMDA) synapse. These neurons project to and excite (also via glutamate acting on NMDA receptors) interneurons in the caudal ventrolateral medulla that project to and release GABA to inhibit sympathetic premotor neurons in the rostral ventrolateral medulla (RVLM). Thus, hormonal modulation of baroreflex function could occur at any or all of these sites, although in the case of pregnancy the RVLM appears pivotal. One hormonal mechanism that underlies decreases in the baroreflex maximum is the action of a major neurosteroid metabolite of progesterone, 3α-hydroxydihydroprogesterone (3α-OH-DHP), to enhance GABAergic suppression of RVLM premotor neurons [3] (Fig. 55.3). 3α-OH-DHP levels, as well as the enzymes responsible for the synthesis of 3α-OH-DHP from progesterone, are increased in the brain at end-gestation, the time at which baroreflex function reaches its nadir. In addition, acute systemic or RVLM administration of 3α-OH-DHP in virgin rats reduces baroreflex maximum levels of renal sympathetic activity similarly to pregnancy. The link between 3α-OHDHP and increased GABAergic tone in RVLM is supported by the well-documented ability of this neurosteroid, by binding to the GABAA receptor, to enhance its function. More importantly, evidence indicates that RVLM premotor neurons receive greater tonic GABAergic suppression during pregnancy [3]. Insulin resistance is a normal adaption of pregnancy that, by increasing circulating glucose levels, serves to enhance glucose availability into the fetus. However, several lines of evidence support the hypothesis that insulin resistance also contributes to the decrease in baroreflex gain, by decreasing brain insulin levels [3]. First, decreases in insulin sensitivity and baroreflex gain are temporally correlated in rabbits, rats and humans [3,7,8]. Second, treatment of pregnant rabbits with the insulin sensitizing drug, rosiglitazone, improves baroreflex function [8]. Third, insulin enters the brain via transport from plasma across the blood brain barrier, and insulin resistant states are associated with decreases in insulin transport. Indeed, during pregnancy, brain insulin levels fall [8,9]. Fourth, in conscious pregnant rats, intracerebroventricular infusion of insulin normalizes baroreflex gain, while in virgin rats insulin infusion is ineffective [9].

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The brain sites and circuitry by which insulin supports baroreflex function have been investigated [3]. Unlike 3α-OH-DHP, which acts in the RVLM, initial studies revealed that insulin initiates its effects in the forebrain, since lateral cerebroventricular, but not fourth ventricular, insulin infusion increased baroreflex gain. Further work identified the arcuate nucleus as the site at which insulin acts to activate the sympathetic nervous system and increase baroreflex gain, via a neural pathway that includes the paraventricular nucleus of the hypothalamus (PVN) [10]. Two major sets of neurons in the arcuate nucleus project to the PVN: proopiomelanocortin (POMC) neurons, which release alphamelanocyte-stimulating hormone, and neuropeptide Y neurons. Recently, the sympathoexcitatory response to insulin was shown to be mediated by PVN melanocortin receptors, suggesting that POMC neurons convey the signal from the arcuate nucleus to PVN [11]. From the PVN, the neuronal pathway appears to converge with brainstem baroreflex circuitry in the RVLM, since insulin’s sympathoexcitatory effect is prevented by blockade of RVLM ionotropic glutamate receptors [11]. These data provide a mechanistic explanation for why, in pregnant animals, rosiglitazone treatment to increase insulin sensitivity [8] and intracerebroventricular insulin infusion [9] each improved baroreflex gain, yet failed to improve the attenuated baroreflex maximum levels of heart rate. Given that GABAergic inhibition of RVLM premotor neurons is increased during pregnancy, this suppression would prevent insulin’s normal effect to increase baroreflex maximum levels as well (Fig. 55.3).

PREECLAMPSIA Preeclampsia is a potentially fatal hypertensive disorder of pregnancy that is initiated by reduced placental perfusion. Increased sympathetic tone may contribute to the hypertension, since basal muscle sympathetic nerve activity is clearly increased above the levels observed in normal pregnant women [4]. Moreover, baroreflex sensitivity is further decreased [3,4]. Interestingly, early gestational measurements of impaired baroreflex function, coupled with detection of reduced uterine perfusion, may noninvasively herald the subsequent development of this life-threatening disorder [3]. However, despite the clinical significance of these autonomic changes, the mechanisms are currently unknown.

SUMMARY AND CONCLUSIONS Pregnancy increases sympathetic nerve firing and decreases both basal parasympathetic activity and baroreflex gain; these changes are exaggerated in women with preeclampsia. The changes in basal autonomic tone may counteract to some degree the profound vasodilation that is a hallmark of normal pregnancy. In contrast,

BRAIN

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INSULIN

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+ RVLM

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3α-OH-DHP

Baroreflex Gain FIGURE 55.3 Mechanisms that contribute to impaired baroreflex function during pregnancy. Insulin in blood is transported across the blood–brain barrier (BBB) into brain, where, via actions in the hypothalamic arcuate nucleus, it enhances baroreflex function. The excitation initiated by insulin in the arcuate nucleus is conveyed via the paraventricular nucleus to a brainstem baroreflex relay, the rostral ventrolateral medulla (RVLM). During pregnancy, falls in brain insulin (which supports baroreflex function) decreases baroreflex gain. In addition, GABAergic suppression of RVLM premotor neurons mediated by the neurosteroid 3α-OH-DHP decreases maximum levels of sympathetic activity elicited by severe hypotension.

the impaired baroreflex function is not beneficial for the mother. Little is known about the mechanisms that mediate the changes in basal autonomic activity in normal pregnancy and with preeclampsia. The decrease in baroreflex function induced by normal pregnancy is mediated by at least two hormonal systems (Fig. 55.3): increased levels of 3α-OH-DHP act in the RVLM to enhance GABAergic suppression of RVLM premotor neurons and the baroreflex maximum, while reduced actions of insulin in the hypothalamus contribute to decreases in baroreflex gain.

References [1] Robson SC, Hunter S, Boys RJ, Dunlop W. Serial study of factors influencing changes in cardiac output during human pregnancy. Am J Physiol 1989;256:H1060–H1065. [2] Thornburg KL, Jacobson SL, Giraud GD, Morton MJ. Hemodynamic changes in pregnancy. Semin Perinatol 2000;24:11–14. [3] Brooks VL, Dampney RA, Heesch CM. Pregnancy and the endocrine regulation of the baroreceptor reflex. Am J Physiol Regul Integr Comp Physiol 2010;299:R439–51. [4] Fu Q, Levine BD. Autonomic circulatory control during pregnancy in humans. Semin Reprod Med 2009;27:330–7. [5] Rang S, Wolf H, Montfrans GA, Karemaker JM. Non-invasive assessment of autonomic cardiovascular control in normal human pregnancy and pregnancy-associated hypertensive disorders: a review. J Hypertens 2002;20:2111–9.

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[6] Brooks VL, Kane CM, Van Winkle DM. Altered heart rate baroreflex during pregnancy: role of sympathetic and parasympathetic nervous systems. Am J Physiol 1997;273:R960–R966. [7] Brooks VL, Mulvaney JM, Azar AS, Zhao D, Goldman RK. Pregnancy impairs baroreflex control of heart rate in rats: role of insulin sensitivity. Am J Physiol Regul Integr Comp Physiol 2010;298:R419–R426. [8] Daubert DL, Chung MY, Brooks VL. Insulin resistance and impaired baroreflex gain during pregnancy. Am J Physiol Regul Integr Comp Physiol 2007;292:R2188–R2195.

[9] Azar AS, Brooks VL. Impaired baroreflex gain during pregnancy in conscious rats: role of brain insulin. Hypertension 2011;57:283–8. [10] Cassaglia PA, Hermes SM, Aicher SA, Brooks VL. Insulin acts in the arcuate nucleus to increase lumbar sympathetic nerve activity and baroreflex function in rats. J Physiol 2011;589:1643–62. [11] Ward KR, Bardgett JF, Wolfgang L, Stocker SD. Sympathetic response to insulin is mediated by melanocortin 3/4 receptors in the hypothalamic paraventricular nucleus. Hypertension 2011;57:435–41.

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56 Aging and the Autonomic Nervous System Lewis A. Lipsitz, Vera Novak Healthy human aging is associated with several abnormalities in autonomic nervous system function that can impair an older person’s adaptation to the stresses of everyday life. Aging affects central and peripheral autonomic regulation of heart rate (HR), blood pressure (BP), temperature, and visceral organ function. However, aging, per se should not be considered a state of autonomic failure because many mechanisms compensating for demands of daily living remain intact. Orthostatic and postprandial hypotension are two common manifestations of age-associated autonomic nervous system impairment [1,2]. Both are defined as a 20 mmHg or greater decline in systolic BP on assumption of the upright posture, or within 1 hour of eating a meal, respectively. These are two distinct conditions that may or may not occur in the same patient. Orthostatic hypotension is observed in less than 7% of healthy normotensive elderly people and in as many as 30% of people older than 75 years with multiple pathologic conditions. Postprandial BP falls an average of 11 mmHg in as many as three quarters of asymptomatic community-dwelling elderly people over age 70. Postprandial hypotension is particularly common in the nursing home population where it accounts for up to 8% of cases of syncope. Orthostatic hypotension is an important symptom of autonomic failure, commonly associated with diabetes, malignancy, amyloidosis, Parkinson’s disease, multiple system atrophy, Lewy body dementia, pure autonomic failure, and other syndromes in elderly patients. A unique feature of orthostatic and postprandial hypotension in elderly people without overt autonomic failure is that they are commonly associated with supine or sitting systolic hypertension. This may be due to the adverse effects of hypertension on baroreflex sensitivity, vascular reactivity, and diastolic filling that compound age-related abnormalities in these BP regulatory mechanisms. Both hypotension and hypertension are associated with reduced cerebral perfusion, damage to gray and white matter, and cognitive decline [3]. Although it may seem counterintuitive, the gradual and judicious lowering of BP may improve, rather than worsen hypotension and cerebral perfusion in hypertensive patients. This presentation of orthostatic or postprandial hypotension in the setting of systolic hypertension must be distinguished from the supine hypertension seen in frank autonomic failure, where patients

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00056-1

are very symptomatic and develop hypertension later in the course, often in association with the use of pressor medications. Medications that acutely lower BP may also contribute to hypotension in elderly patients, particularly diuretics, antihypertensives, alpha-blockers used for prostatic obstruction, dopamine, tricyclic antidepressants, and neuroleptics. If clinically indicated, these medications should be given in the lowest possible dose and slowly titrated to the desired effect in order to allow BP regulatory mechanisms to adapt. Recent studies suggest that the control of hypertension in elderly people can not only reduce cardiovascular morbidity and mortality, but also reduce orthostatic hypotension [4], increase cerebral blood flow, and improve carotid distensibility [5]. Pathophysiologic mechanisms predisposing healthy elderly people to orthostatic and postprandial hypotension are summarized in Table 56.1.

CARDIAC BAROREFLEX FUNCTION Normal human aging is associated with a reduction in cardiovagal baroreflex sensitivity, which may lead TABLE 56.1 Age-Related Physiologic Changes Predisposing to Hypotension

1. Decreased baroreflex sensitivity a. Diminished HR response to hypotensive stimuli (orthostasis, meal digestion, and hypotensive medications).

b. Decreased adrenergic vascular responsiveness to orthostatic, postprandial, and medication-related BP reduction.

2. Impaired defense against reduced intravascular volume a. Reduced secretion of renin, angiotensin, and aldosterone. b. Increased atrial natriuretic peptide, supine and upright. c. Decreased plasma vasopressin response to orthostasis. d. Reduced thirst after water deprivation. 3. Reduced early cardiac ventricular filling (diastolic dysfunction) a. Increased dependence on cardiac preload to maintain cardiac output.

b. Increased dependence on atrial contraction to fill the ventricles –

leads to hypotension during atrial fibrillation. Decreased cardiac output during tachycardia when ventricular filling time is reduced. Impaired postprandial vasoconstriction a. Vasodepressor action of insulin [10].

c.

4.

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to hypotension due to decreased cardiac output. This is manifest by a blunted cardioacceleratory response to hypotensive stimuli, such as upright posture, Phase II of the Valsalva maneuver, nitroprusside infusions, and lower body negative pressure; as well as the reduced bradycardic response to hypertensive stimuli, such as the administration of phenylephrine. The baroreflex may be impaired at any one of multiple sites along its arc, including carotid and cardiopulmonary pressure receptors, afferent pathways, brainstem (nucleus tractus solitarii) and higher regulatory centers, efferent sympathetic and parasympathetic neurons, postsynaptic cardiac beta receptors, or defects in intracellular signal transduction pathways within the sinoatrial node. An age-associated increase in BP also contributes to baroreflex impairment. Known agerelated abnormalities in components of the baroreflex arc are summarized below.

SYMPATHETIC ACTIVITY Studies of sympathetic nervous system activity in healthy human subjects demonstrate an age-related increase in resting plasma norepinephrine levels, muscle sympathetic nerve activity, and vascular resistance, as well as the plasma norepinephrine response to upright posture and exercise. The increase in plasma norepinephrine is primarily due to an increase in norepinephrine spillover at sympathetic nerve endings and secondarily due to a decrease in clearance. Despite apparent increases in sympathetic tone with aging, cardiac and vascular responsiveness is diminished. Infusions of beta-adrenergic agonists result in smaller increases in HR, left ventricular ejection fraction, cardiac output, and vasodilation in older compared with younger men.

PARASYMPATHETIC ACTIVITY Previous studies demonstrating age-related reductions in HR variability in response to respiration, cough, and the Valsalva maneuver suggest that aging is associated with impaired parasympathetic control of HR. Elderly patients with unexplained syncope have even greater impairments in the HR response to cough and deep breathing than healthy age-matched subjects without syncope [6]. There is evidence that physical activity may help maintain or increase vagal activity in older age.

INTEGRATION OF AUTONOMIC CONTROL NETWORKS When plotted continuously over time, the beat-to-beat HR or BP signal is highly irregular due to the interactions of multiple autonomic control systems operating over different time scales. The technique of power spectral analysis is commonly used to quantify the relative contributions

of sympathetic and parasympathetic nervous systems to this complex cardiovascular variability. The HR or BP power spectrum can be divided into low- and highfrequency components. Previous studies using betablockade, atropine, or both suggest that the low-frequency oscillations (0.05–0.1 Hz) in BP reflect sympathetic modulation of vasomotor tone and in HR reflect a combination of baroreflex-mediated sympathetic and parasympathetic influences. High-frequency components of the HR and BP power spectra (0.15–0.5 Hz) appear to be under parasympathetic control and represent the effects of respiration. Spectral analysis techniques have confirmed that healthy aging is associated with reductions in baroreflex and parasympathetic modulation of HR, with a relatively greater loss of the high-frequency parasympathetic component. The overall complexity of cardiovascular signals, which represents the integration of autonomic, endocrine, hemodynamic, and other control networks, can be quantified using a variety tools derived from nonlinear dynamics, fractals, and complex systems theory. A decline in the complexity of cardiovascular dynamics is associated with aging and is a marker of cardiovascular disease [7].

NEUROTRANSMITTERS Just as the age-related increase in plasma norepinephrine level is related to increased spillover, and decreased clearance at adrenergic nerve terminals, so must the effect of aging on any neurotransmitter be interpreted with regard to changes in its production and clearance. Other neurotransmitters that influence autonomic nervous system functions have received little attention in aging humans. In the brain, a decline in dopamine and norepinephrine is related to a loss of dopaminergic and noradrenergic neurons in the substantia nigra and locus ceruleus. The clinical implications of these changes are not fully understood, but may lead to the slowing of gait and cognitive dysfunction commonly seen in elderly people. The enzymes choline acetyltransferase and acetylcholinesterase, which are responsible for synthesis and degradation of acetylcholine, respectively, decrease in the cerebral cortex with aging. Furthermore, muscarinic and nicotinic receptors have been reported to decrease in cortical structures. These findings provide indirect evidence for a decrease in central cholinergic neurotransmission with normal aging.

CARDIAC BETA-ADRENERGIC RECEPTORS The age-related decrease in chronotropic response to sympathetic stimulation has been attributed to multiple molecular and biochemical changes in beta-receptor coupling and postsynaptic signaling. The number of beta receptors in cardiac myocytes is unchanged with advancing age, but the affinity of beta receptors for agonists is reduced. Postsynaptic changes with aging include

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a decrease in the activity of G protein and the adenylate cyclase catalytic unit, and a decrease in cAMP-dependent phosphokinase-induced protein phosphorylation. As a result of these changes, G protein-mediated signal transduction is impaired. The decrease in cardiac contractile response to sympathetic stimulation has been studied in rat ventricular myocytes, where it appears to be related to decreased influx of calcium ions through sarcolemmal calcium channels and a reduction in the amplitude of the cytosolic calcium transient after beta-adrenergic stimulation. These changes are similar to those seen in receptor desensitization caused by prolonged exposure of myocardial tissue to betaadrenergic agonists. Thus, age-associated alterations in beta-adrenergic response may be caused by desensitization of the adenylate cyclase system in response to a chronic increase in sympathetic activity.

VASCULAR REACTIVITY Aging is associated with increased vascular stiffness and peripheral resistance, and reduced vasoconstriction in response to sympathetic stimulation [8]. However, the impairment in arterial alpha-adrenergic vasoconstriction is reversible by suppression of sympathetic nervous system activity with guanadrel [8]. This remarkable observation suggests that the abnormality in alpha-adrenergic response also represents receptor desensitization caused by heightened sympathetic nervous system activity. It also indicates that some of the physiologic changes associated with aging may be reversible. Endothelium-dependent vasodilatation is progressively impaired with aging. The vasorelaxation response of both arteries and veins to infusions of the beta-adrenergic agonist isoproterenol is attenuated in elderly people. This may also be due to abnormal beta-adrenergic receptor signaling and cAMP production in vascular smooth muscle cells.

VOLUME REGULATION Aging is associated with a progressive decline in plasma renin, angiotensin II, and aldosterone levels, and increases in atrial natriuretic peptide, all of which promote salt wasting by the kidney. In many healthy elderly individuals there is also a defective plasma vasopressin response to upright posture. These physiologic changes predispose elderly people to volume contraction and hypotension. Furthermore, healthy elderly individuals do not experience the same sense of thirst as younger subjects when they become hyperosmolar during water deprivation or hypertonic saline infusion. Consequently, dehydration may develop rapidly during conditions such as an acute illness, preparation for a medical procedure, diuretic therapy, or exposure to a warm climate when fluid losses are increased and access to oral fluids is limited.

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CEREBRAL AUTOREGULATION The process of cerebral autoregulation maintains a relatively constant cerebral blood flow during changes in perfusion pressure. This is accomplished by active vasodilatation of the cerebral microvasculature during hypotensive stimuli and by vasoconstriction during increases in BP. Cerebral autoregulation is generally effective within a mean arterial pressure range of 80 to 150 mmHg and is well-preserved in healthy aging. However, the lower pressure threshold for cerebral hypoperfusion may be shifted higher in patients with hypertension. There is some evidence that the treatment of hypertension may restore this threshold to a more normal range. Cerebral blood flow is also dependent on the arterial concentration of carbon dioxide, such that CO2 inhalation causes vasodilatation and hyperventilation causes vasoconstriction. This CO2 vasoreactivity is mediated by nitric oxide and is an indicator of endothelial function in the cerebral vasculature. Vasoreactivity is reduced with aging and cardiovascular risk factors, such as hypertension and diabetes. Elderly patients with these vascular risk factors are therefore vulnerable to symptomatic cerebral hypoperfusion if BP falls below the autoregulated range. Recent studies suggest that chronic cerebral hypoperfusion is associated with the development of periventricular white matter hyperintensities and their associated impairments in gait and cognition [3,9].

References [1] Gupta V, Lipsitz LA. Orthostatic hypotension in the elderly: diagnosis and treatment. Am J Med Oct 2007;120(10):841–7. [2] Jansen RW, Lipsitz LA. Postprandial hypotension: epidemiology, pathophysiology, and clinical management. Ann Intern Med Feb 15 1995;122(4):286–95. [3] Novak V, Hajjar I. The relationship between blood pressure and cognitive function. Nat Rev Cardiol 2010;7:686–98. [4] Masuo K, Mikami H, Ogihara T, Tuck ML. Changes in frequency of orthostatic hypotension in elderly hypertensive patients under medications. Am J Hypertens Mar 1996;9(3):263–8. [5] Lipsitz LA, Gagnon M, Vyas M, et al. Antihypertensive therapy increases cerebral blood flow and carotid distensibility in hypertensive elderly subjects. Hypertension Feb 2005;45(2):216–21. [6] Maddens M, Lipsitz LA, Wei JY, Pluchino FC, Mark R. Impaired heart rate responses to cough and deep breathing in elderly patients with unexplained syncope. Am J Cardiol Dec 1 1987;60(16): 1368–72. [7] Goldberger AL, Amaral LA, Hausdorff JM, Ivanov P, Peng CK, Stanley HE. Fractal dynamics in physiology: alterations with disease and aging. Proc Natl Acad Sci USA Feb 19 2002;99(Suppl 1):2466–72. [8] Hogikyan RV, Supiano MA. Arterial alpha-adrenergic responsiveness is decreased and SNS activity is increased in older humans. Am J Physiol May 1994;266(5 Pt 1):E717–724. [9] ten Dam VH, van den Heuvel DM, de Craen AJ, et al. Decline in total cerebral blood flow is linked with increase in periventricular but not deep white matter hyperintensities. Radiology Apr 2007;243(1):198–203. [10] Kearney MT, Cowley AJ, Stubbs TA, Evans A, Macdonald IA. Depressor action of insulin on skeletal muscle vasculature: a novel mechanism for postprandial hypotension in the elderly. J Am Coll Cardiol Jan 1998;31(1):209–16.

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57 Exercise Qi Fu, Benjamin D. Levine INTRODUCTION Physical activity is a key component of a healthy lifestyle. Increased physical activity or exercise training is not only protective against cardiovascular disease, type II diabetes, and obesity, but also effective in improving functional capacity in patients with autonomic disorders. Exercise training also improves mental health, helps to prevent depression, and promotes or maintains positive self-esteem. Endurance or “dynamic” exercise, such as running, jogging, cycling, swimming, rowing, or walking, and strength training, such as weight-lifting are two major forms of exercise though many activities include components of both. In this chapter, we predominately focus on endurance exercise. There is now convincing evidence that the protective and therapeutic effects of exercise training are related, in a substantive fashion, to effects on the autonomic nervous system [1]. In addition, training-induced improvement in vascular endothelial function, blood volume expansion, cardiac remodeling, insulin resistance, and renaladrenal function may also contribute to the protection and treatment of cardiovascular, metabolic, and autonomic disorders.

ACUTE EXERCISE The cardiovascular response during exercise is initiated by a feed-forward mechanism, termed “central command”, which involves higher brain centers such as the motor cortex, hypothalamic and mesencephalic locomotor regions that activate parallel circuits controlling locomotor, cardiovascular, and ventilatory functions [2]. As exercise continues, both mechanical and metabolic signals from active skeletal muscle provide feedback to cardiovascular centers in the brain through group III and IV muscle afferents, the so-called “exercise pressor reflex”, to precisely match systemic oxygen delivery with metabolic demand [2,3]. Vascular resistance decreases (via local metabolic factors) to facilitate increases in muscle perfusion and cardiac output increases proportionate with oxygen uptake, thus allowing the maintenance of or even increase in mean arterial pressure. An overview of the neural regulation of the cardiovascular system during exercise is shown in Figure 57.1.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00057-3

During exercise, the uptake and transport of oxygen is required for oxidative phosphorylation and the efficient production of adenosine triphosphate (ATP) to support the metabolic demands of the body [4]. One of the most inviolate relationships in all of exercise physiology is that between oxygen uptake and cardiac output. Regardless of age, sex, or the presence of various disease states, in general, about 56 liters of cardiac output are required for every liter of oxygen uptake above rest (Fig. 57.2) [4,5]. When this relationship is depressed, it may be a sign of severe underlying disease with impending decompensation. Conversely, when it is exaggerated, like in patients with metabolic myopathies, it gives strong clues to the processes regulating cardiac output. Oxygen uptake is a function of the triple-product of heart rate and stroke volume (i.e., cardiac output) and arterial-mixed venous oxygen difference (the Fick principle, Fig. 57.2) [5]. The degree to which each of the variables can increase determines the upper limit for whole-body oxygen consumption, and this limit is called the maximal oxygen uptake (VO2max) [6]. Maximal heart rate and maximal arteriovenous difference are usually relatively similar among individuals of similar ages despite large differences in physical fitness; therefore, the factor most commonly accounting for the different values of VO2max in different individuals is stroke volume [5]. At rest arteriovenous oxygen difference is normally 4.5 ml/100 ml/min (approximately 23% extraction), and at VO2max this difference is commonly close to 16 ml/100 ml/min (about 80 to 85% extraction) [5]. Cardiovascular responses, such as heart rate and blood pressure, are more closely related to the relative metabolic demands than to the absolute demands. At low levels of exercise, heart rate increases almost exclusively via vagal withdrawal, with little evidence for systematic increases in sympathetic activity until the intensity of exercise is at or above the maximal steady state (ventilatory or lactate thresholds) [5]. The key determinant of the magnitude of the heart rate response to exercise is the relative intensity as well as the absolute amount of muscle mass engaged, while central command plays an essential role in the increase in heart rate during exercise [4]. Gravity plays a critical role in determining the distribution of blood within the cardiovascular system, and body posture markedly affects the relative importance of changes in stroke volume [4]. In the upright position,

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FIGURE 57.1 Autonomic neural control of the cardiovascular response during exercise. Central command initiates the exercise pressor response, which is maintained and augmented via feedback from baroreceptors, as well as by stimulation of skeletal muscle mechanically and metabolically sensitive receptors. After integration in the brain, efferent responses via the parasympathetic (vagal) and sympathetic nervous systems result in increased heart rate and contractility, vasoconstriction in non-exercising (inactive) muscle, and vasodilation in exercising (active) muscle beds mediated by release of local vasodilating substances (“functional sympatholysis”). Adapted with permission from Levine [4], originally developed by J. Mitchell.

stroke volume is only about one-half its value in the supine position due to blood pooling in the legs and a reduction in left ventricular end-diastolic volume. At the onset of exercise, the pumping action of skeletal muscle acts to augment venous return substantially, and stroke volume normally increases 50% via the Starling mechanism [4]. Maximal stroke volume in non-athletic individuals is achieved at relatively low levels of exercise intensity (approximately 50% of maximal oxygen uptake), as pericardial constraint serves to limit left ventricular end-diastolic volume. In general, patients with autonomic disorders have low levels of VO2max and blunted cardiovascular response during exercise. It has been found that patients with pure autonomic failure and multiple system atrophy have an abnormal fall in blood pressure during exercise [7,8], which is presumably attributable to reduced sympathetic nerve activity and blunted α-adrenergic vasoconstriction in nonexercising skeletal muscle in the context of exercise induced vasodilation [9]. Conversely, patients with baroreflex failure have an excessive increase in blood pressure during exercise, which is probably due to impaired baroreflex buffering [10].

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FIGURE 57.2 Relationship between the increase in oxygen uptake (VO2) and the corresponding increase in cardiac output (Qc) during exercise in humans, which in most cases is 6/1. VO2 is a function of the triple-product of heart rate (HR), stroke volume (SV), and arterial-mixed venous oxygen difference (A-V O2 Difference). Adapted with permission from Levine [4].

Both central command and the exercise pressor reflex are important in determining the cardiovascular response during exercise, while dynamic interactions between these

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Sympathetic tone Baroreflex sensitivity

Vagal tone

Heart rate Cardiac size/mass Cardiac function

Endothelial function Hemoglobin mass Insulin resistance

Exercise

Blood/plasma volume

Renal-adrenal function

Obesity

Blood pressure

FIGURE 57.3 Effects of exercise training on the autonomic nervous system, cardiovascular system, and renal-adrenal system in humans.

feed-forward and feed-back circuits are associated with beneficial adjustments in the sympathetic and parasympathetic nervous systems. The beneficial effects can be observed soon after the initiation of exercise training and are sustained as long as activity is continued. Since the sympathetic nervous system is activated during each bout of exercise, repeated activation of this system may result in an attenuation of sympathetic nerve activity. Numerous studies have shown that regular exercise can improve cardiac autonomic balance (i.e., increasing parasympathetic while decreasing sympathetic regulation of the heart), as well as increase the sensitivity of the baroreflex [11]. Additionally, exercise training can result in cardiac remodeling (i.e., increased cardiac size and mass, and improved cardiac function). It has been demonstrated that training increases the release of nitric oxide through shear stress during exercise, and chronic increases in nitric oxide lead to functional and histological alterations of vascular endothelium, causing enhanced vascular structure and function. Greater glucose delivery occurs with exercise training due to increased muscle blood flow and capillary density. An improvement in insulin resistance has been found after exercise training in humans. Training-induced muscle adaptations appear to be important in attenuating insulin mediated sympathetic activation, and may be especially enhanced by strength training which increases overall muscle mass [12]. Exercise training has also been shown to improve aerobic capacity and vascular conductance, and

lower body fat, each of which could contribute to a reduction in blood pressure. Many human studies have found that exercise training increases total hemoglobin mass, red blood cell volume, plasma volume, and blood volume. Recent research has reported that training may also improve renal-adrenal function and decrease circulating levels of angiotensin II [13]. Figure 57.3 depicts the beneficial effects of exercise training in humans.

EXERCISE AS A NON-DRUG THERAPY There is abundant evidence showing that regular exercise decreases the risk of cardiovascular disease, hypertension, colon and breast cancer, type II diabetes, and obesity. Numerous clinical studies have proven that exercise training can be used therapeutically to restore the autonomic function towards normal, and thus, contribute to an improvement in outcome. For example, it has been reported that exercise training is effective in treating patients with chronic heart failure, myocardial infarction, or after coronary artery bypass surgery by increasing vagal modulation and decreasing sympathetic tone. Additionally, exercise training seems to be effective in the prevention of sudden cardiac death by augmenting baroreflex sensitivity and heart rate variability [14]. Many clinical investigations have shown that exercise training improves functional capacity in patients with

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autonomic disorders, such as Parkinson’s disease, stroke, multiple sclerosis, spinal cord injury, Guillain–Barré syndrome, muscular dystrophy, or metabolic myopathies. Although the effects of exercise training on orthostatic tolerance in healthy individuals are controversial, increased orthostatic tolerance after mild to moderate training has been found in patients with unexplained syncope or orthostatic hypotension [15]. Increased baroreflex sensitivity and decreased frequency of syncopal episodes after exercise training were observed in patients with neurally mediated syncope [16]. In addition, it was reported that exercise training had a beneficial effect on physiological and subjective parameters in patients with chronic fatigue syndrome [17]. Fu et al. [18] have found that short-term (i.e., 3 months) exercise training improves or even cures the postural orthostatic tachycardia syndrome (POTS, also called chronic orthostatic intolerance) in most patients; more importantly, patient quality of life assessed by the 36-Item Short-Form Health Survey is significantly improved in all the patients after training. Recent research has demonstrated that exercise training improves older adults’ cognitive function. Exercise has also been found to be neuroprotective in many neurodegenerative and neuromuscular diseases in humans. Given the beneficial effects of exercise training, the American Heart Association, the US Surgeon General, the Centers for Disease Control and Prevention, and the American College of Sports Medicine recommend at least 30 minutes per day of at least moderate-intensity exercise, including brisk walking, jogging, cycling, swimming, or running on most, and preferably all, days of the week [19]. Physicians’ advice to increase physical activity can be a strong motivator to patients, and advice conveyed as a written prescription may enhance success. Supervised exercise training is preferable to maximize functional capacity. Heart rate can be used as an easily measured estimate of relative exercise intensity, and the target training heart rate is usually set at approximately 75% of the maximal heart rate [(take 220–age) 5 beats/min]. However, it is important to emphasize that these are only guidelines that in some patients, such as those who are taking beta-blockers or other medications or with underlying autonomic disorders, may affect the heart rate response to exercise and may not accurately reflect exercise intensity. For patients with autonomic disorders, heat and body temperature during exercise may compromise blood pressure further (by further increases in vascular conductance); cooling the skin or semi-recumbent exercise is recommended. For patients with POTS or orthostatic intolerance, exercise training should be initiated by using a recumbent bike, rowing, or swimming [18]. The use of only semi-recumbent exercise at the beginning is a critical strategy, allowing patients to exercise while avoiding the upright posture that elicits their symptoms. As the patients become relatively fit, the duration and intensity of exercise should be progressively increased, and upright exercise (e.g., upright bike, walking on the treadmill, or jogging) can be gradually added as tolerated [18].

CONCLUSION A sedentary lifestyle is considered to be one of the most important modifiable risk factors for morbidity and mortality in humans. Physical activity or exercise training is necessary to maintain overall health and functional capacity, and it plays a crucial role in the prevention of cardiovascular disease, sudden cardiac death, hypertension, type II diabetes, colon and breast cancer, and obesity. Exercise training can be therapeutic for patients with orthostatic intolerance, syncope, or POTS. In addition, exercise training improves mental health, helps to prevent depression, and promotes or maintains positive self-esteem. Adaptations involving the autonomic nervous system play a large role in the protective and therapeutic effects of exercise training. Moderate-intensity exercise at least 30 minutes per day and at least 5 days per week is recommended for the vast majority of people. Supervised exercise training is preferable to maximize functional capacity, and may be particularly important in patients with autonomic disorders.

References [1] Joyner MJ, Green DJ. Exercise protects the cardiovascular system: effects beyond traditional risk factors. J Physiol 2009;587:5551–8. [2] Mitchell JH. Wolffe JB memorial lecture. Neural control of the circulation during exercise. Med Sci Sports Exerc 1990;22:141–54. [3] Michelini LC, Stern JE. Exercise-induced neuronal plasticity in central autonomic networks: role in cardiovascular control. Exp Physiol 2009;94:947–60. [4] Levine BD. 2001 Exercise physiology for the clinician. In: Exercsie and sports cardiology. Medical Publishing Division: McGraw-Hill. p. 3–29. [5] Rowell LB. Central circulatory adjustments to dynamic exercise. Human Cardiovascular Control 1993:162–203. [6] Levine BD. VO2max: what do we know, and what do we still need to know? J Physiol 2008;586:25–34. [7] Humm AM, Mason LM, Mathias CJ. Effects of water drinking on cardiovascular responses to supine exercise and on orthostatic hypotension after exercise in pure autonomic failure. J Neurol Neurosurg Psychiatry 2008;79:1160–4. [8] Smith GD, Mathias CJ. Differences in cardiovascular responses to supine exercise and to standing after exercise in two clinical subgroups of Shy-Drager syndrome (multiple system atrophy). J Neurol Neurosurg Psychiatry 1996;61:297–303. [9] Schrage WG, Eisenach JH, Dinenno FA, Roberts SK, Johnson CP, Sandroni P, et al. Effects of midodrine on exercise-induced hypotension and blood pressure recovery in autonomic failure. J Appl Physiol 2004;97:1978–84. [10] Ziegler MG, Ruiz-Ramon P, Shapiro MH. Abnormal stress responses in patients with diseases affecting the sympathetic nervous system. Psychosom Med 1993;55:339–46. [11] Okazaki K, Iwasaki K, Prasad A, Palmer MD, Martini ER, Fu Q, et al. Dose-response relationship of endurance training for autonomic circulatory control in healthy seniors. J Appl Physiol 2005;99:1041–9. [12] Church TS, Blair SN, Cocreham S, Johannsen N, Johnson W, Kramer K, et al. Effects of aerobic and resistance training on hemoglobin A1c levels in patients with type 2 diabetes: a randomized controlled trial. JAMA 2010;304:2253–62. [13] Rush JW, Aultman CD. Vascular biology of angiotensin and the impact of physical activity. Appl Physiol Nutr Metab 2008;33:162–72.

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[14] La Rovere MT, Bersano C, Gnemmi M, Specchia G, Schwartz PJ. Exercise-induced increase in baroreflex sensitivity predicts improved prognosis after myocardial infarction. Circulation 2002;106:945–9. [15] Mtinangi BL, Hainsworth R. Increased orthostatic tolerance following moderate exercise training in patients with unexplained syncope. Heart 1998;80:596–600. [16] Gardenghi G, Rondon MU, Braga AM, Scanavacca MI, Negrao CE, Sosa E, et al. The effects of exercise training on arterial baroreflex sensitivity in neurally mediated syncope patients. Eur Heart J 2007;28:2749–55. [17] Joosen M, Sluiter J, Joling C, Frings-Dresen M. Evaluation of the effects of a training programme for patients with prolonged fatigue

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on physiological parameters and fatigue complaints. Int J Occup Med Environ Health 2008;21:237–46. [18] Fu Q, Vangundy TB, Galbreath MM, Shibata S, Jain M, Hastings JL, et al. Cardiac origins of the postural orthostatic tachycardia syndrome. J Am Coll Cardiol 2010;55:2858–68. [19] Marcus BH, Williams DM, Dubbert PM, Sallis JF, King AC, Yancey AK, et al. Physical activity intervention studies: what we know and what we need to know: a scientific statement from the american heart association council on nutrition, physical activity, and metabolism (subcommittee on physical activity); council on cardiovascular disease in the young; and the interdisciplinary working group on quality of care and outcomes research. Circulation 2006;114:2739–52.

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58 Effects of High Altitude Luciano Bernardi INTRODUCTION Ascent to high altitude reduces the inspired partial pressure of O2, leading to hypobaric hypoxia. This requires a complex adaptive process (acclimatization) which, in its early phases is largely influenced by the autonomic nervous system. This process is complex and the integrated response depends upon a number of factors, including the extent, rate of ascent, and duration of hypoxic exposure. Acute responses are modified by chronic adaptations that restore circulatory function towards normoxic levels over time periods that may range from a few days or weeks for the sea level sojourner, to years for the high altitude native. Tolerance to hypoxia varies greatly amongst individuals, and subjects with a particular susceptibility may develop inappropriate responses leading to acute mountain sickness and even to life threatening conditions, such as high altitude cerebral and pulmonary edema (HAPE). Sympathetic over activity seems to play an important role in HAPE.

EFFECTS OF ACUTE HYPOXIA The main oxygen sensors involving the autonomic nervous system response are the peripheral chemoreceptors, located in the carotid body and in the arch of the aorta. The carotid sensors respond mainly to a lowering in PaO2 (aortic sensors respond mainly to CaO2). Peripheral chemoreceptor afferents synapse in a primary cardiovascular control center located within nucleus tractus solirarii (NTS) of the dorsomedial medulla. The NTS relays this input via projections terminating in the rostral ventrolateral medulla. Stimulation of neurons in the ventrolateral medulla by hypoxic stimulation of peripheral chemoreceptors leads to hyperventilation, and excitation of both sympathetic and parasympathetic neurons. However, this response is modified by numerous secondary influences such as hyperventilation and hypocapnia [1]. The hyperventilation, which is proportional to the decreases in PaO2, stimulates lung stretch receptors (during inspiration), whose final response is an inhibition of cardiac vagal tone. In addition, hyperventilation induces hypocapnia, which attenuates the sympathetic activation associated with the peripheral chemoreflex, and also reduces the stimulus from the central chemoreflex – so called hypocapnic

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braking effect [2]. Due to the increase in ventilation, the respiratory changes in stroke volume are also increased, and this contributes to increase baroreceptor loading and unloading during expiration and inspiration, respectively. The baroreflexes remain able to maintain adequate cardiovascular modulation during chemoreflex activation [3]. Thus arterial baroreceptors, central/peripheral chemoreceptors and pulmonary stretch receptors all may modulate the sympathetic activation associated with peripheral chemoreflex excitation [1]. Although complete inability to increase ventilation in response to hypoxia (causing dyspnea) is generally admitted as a contraindication to travel to high altitude, the extent of these autonomic/ventilatory responses are not directly proportional to performance at high altitude. Contrary to general belief, climbers able to reach extreme altitudes (Everest and K2 summits) without oxygen supplementation were characterized by a high respiratory efficiency, but only moderate increase in ventilatory responses, and only moderate sympathetic activation during acclimatization at 5200 m. Other climbers unable to reach extreme altitudes were characterized by higher ventilatory responses and higher sympathetic activation. This suggested that better respiratory efficiency was associated, in part, with a reduction in the cardiorespiratory autonomic reactions at intermediate altitudes thereby maintaining a higher reserve for the more demanding tasks of extreme altitudes [4]. The purpose of this combined cardiovascular and autonomic response is to maintain systemic and regional oxygen despite the fall in PaO2 [5]. This is achieved by an increase in cardiac output (proportional to the altitude), mainly due to an increase in heart rate: tachycardia results from increased sympathetic activity due to chemoreceptor stimulation, and probably also from vagal withdrawal, as a consequence of hyperventilation (central pathways, feedback from pulmonary stretch receptors, and hypocapnia) [2]. Acute hypoxia provokes vasodilation in all vascular beds, except the lung (where hypoxic vasoconstriction in susceptible subjects is often exaggerated and may be a crucial factor leading to HAPE). Peripheral vasodilation, in addition to increased in heart rate and cardiac output causes a remarkably effective redistribution of flow to vascular beds with the greatest metabolic demand, similar to the response seen during physical exercise [6]. The increment in blood flow associated with hypoxia, at rest and during exercise,

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precisely matches the decrease in arterial oxygen content, keeping O2 delivery to tissues constant. Both endothelial and neurally derived NO may be involved in this integrated autonomic and cardiovascular response to hypoxia [1,7]. Initially, the level of blood pressure depends on the balance between hypoxic-induced vasodilation and the vasoconstrictor effect of peripheral chemoreceptor reflex sympathetic activation; in this acute phase it shows little change. When peripheral vasodilation is not effectively compensated by sympathetic activity syncope may occur; this is more frequent after very rapid ascent to high altitude. Plasma catecholamines are also involved in the regulation of blood pressure at altitude, but their changes are still not fully understood. At the moment it remains accepted that adrenaline increases predominantly in the early phases of exposure to high altitude, whereas noradrenaline predominates after several days of exposure. These modifications are compatible with the delayed increase in blood pressure, observed in the majority of subjects during sojourn to high altitude [8]. Hypoxia also causes cerebral vasodilation, which is partially counteracted by hypocapnia, but can lead to a pulsating headache, which is a common symptom upon arrival at high altitude.

EFFECTS OF CHRONIC HYPOXIA Over a period of days to weeks, reduction in plasma volume and later increase in red cell mass increase hemoglobin concentration and CaO2 (also with a contribution of hyperventilation). The increased CaO2 reduces the hyperdynamic circulation and both cardiac output and peripheral blood flow returns towards normal. However, the still low PaO2 maintains peripheral chemostimulation and hence sympathetic activation. This results in a progressive increase in vascular resistance and blood pressure, as the increase in CaO2 reduces the vasodilatory effect of hypoxia. The arterial baroreflexes, although diminished, continue to function at high altitude despite the marked increase in chemoreflexes [3], suggesting a central resetting of the baroreflex with exposure to hypoxia [1]. Cardiac output is consistently depressed after acclimatization to high altitude, often below values measured at sea level; this is due to a decrease in stroke volume (with preserved contractile function) due, in turn, to a reduction in left ventricular end-diastolic volume, which results from a 20–30% reduction in plasma volume. Sympathetic activation and slower acting neurohormones, such as angiotensin, aldosterone or vasopressin may be important in sustaining the reduction in plasma volume in chronic hypoxia. This relative dehydration unloads low-pressure cardiopulmonary receptors as well as arterial baroreceptors (by reducing aortic dimensions, and by decreasing pulsatile flow through arterial baroreceptors [1]). Chronic sympathetic hyperactivity leads to downregulation of peripheral β-adrenoceptors, with a gradual diminution of the heart rate response to sympathetic

activation [9]. After a prolonged sojourn to high altitude sympathetic activity diminishes and blood pressure normalizes. After years of acclimatization, high altitude natives appear similar to sea level natives. While this response is typically observed at altitudes varying from 3000 to 5000 m, at extreme altitudes the acclimatization process cannot normalize arterial oxygen content; as a result, peripheral vasodilation and reduction in peripheral resistance persist as in acute hypoxia.

AUTONOMIC NERVOUS SYSTEM AND ALTITUDE ILLNESS HAPE is associated with severe pulmonary hypertension probably with uneven distribution, thus allowing pulmonary areas of hyperperfusion and hypertension; this leads to pulmonary capillary leak, endothelial dysfunction, possibly a late inflammation and alveolar edema. Although the origin of HAPE is still debated, recent studies in the Italian/Swiss Alps, on patients with HAPE susceptibility, have confirmed the essential role of pulmonary hypertension. Such patients have a marked increase in sympathetic activity during acute exposure to hypoxia, even before the development of HAPE. Sympathetic activation may thus play at least a facilitatory role in HAPE, presumably by contributing to the development of pulmonary hypertension in susceptible individuals [10].

References [1] Levine BD. Mountain Medicine and the autonomic nervous system. In: Appenzeller O, editor. Handbook of clinical neurology Vol 75 (31): the autonomic nervous system. Amsterdam: Elsevier; 2000. p. 259–80. Part II [2] Somers VK, Mark AL, Zavala DC, Abboud FM. Influence of ventilation and hypocapnia on sympathetic nerve responses to hypoxia in normal humans. J Appl Physiol 1989;67:2095–100. [3] Bernardi L, Passino C, Spadacini G, Calciati A, Robergs R, Greene ER, et al. Cardiovascular autonomic modulation and activity of carotid baroreceptors at altitude. Clin Sci 1998;95:565–73. [4] Bernardi L, Schneider A, Pomidori L, Paolucci E, Cogo A. Hypoxic ventilatory response in successful extreme altitude climbers. Eur Respir J 2006;27:165–71. [5] Wolfel EE. Sympatho-adrenal and cardiovascular adaptation to hypoxia. In: Sutton JR, Houston CS, Coates G, editors. Hypoxia and molecular medicine. : Queen City Burlington; 1993. p. 62–80. [6] Cerretelli P, Marconi C, Deriaz O, Giezendanner D. After effects of chronic hypoxia on cardiac output and muscle blood flow at rest and exercise. Eur J Appl Physiol 1984;53:92–6. [7] Thomas GD, Victor RG. Nitric oxide mediates contraction-induced attenuation of sympathetic vasoconstriction in rat skeletal muscle. J Physiol (Lond) 1998;506:817–26. [8] Hainsworth R, Drinkhill MJ, Rivera-Chira M. The autonomic nervous system at high altitude. Clin Auton Res 2007;17:13–19. [9] Voelkel NF, Hegstrand L, Reeves JT, McMurtry IF, Molinoff PB. Effects of hypoxia on density of beta-adrenergic receptors. J Appl Physiol 1981;50:363–6. [10] Scherrer U, Allemann Y, Jayet PY, Rexhaj E, Sartori C. High altitude, a natural research laboratory for the study of cardiovascular physiology and pathophysiology. Progr Cardiovasc Dis 2010;52:451–5.

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59 Space Physiology Gilles Clément, Scott Wood INTRODUCTION

SPACE MOTION SICKNESS

Space medicine and space physiology are often viewed as two aspects of space life sciences, with the former being more operational, and the latter being more investigational. Space medicine tries to solve medical problems encountered during space missions. These problems include some adaptive changes to the space environment, including weightlessness, radiation, the absence of the 24-hour day/night cycle; as well as some non-pathologic changes that become maladaptive on return to Earth, such as muscle atrophy and bone demineralization. Space physiology tries to characterize body responses to space, especially weightlessness, reduced activity, and stress. It provides the necessary knowledge required for an efficient space medicine [1]. Space physiology is as old as the first flight of humans in a hot air balloon, when the symptoms of hypoxia were first discovered (at the expenses of the life of the pilot). The interest in this field of research kept growing along with the space program and the opportunities it provided for flying more and more humans in space on board capsules, shuttles, space stations, and soon suborbital space planes. The future of human space flight will inevitably lead to human missions to Mars. These missions will be of long duration (30 months) in isolated and somewhat confined habitats, with the crew experiencing several transitions in levels of gravity, dangerous radiation, and the challenges of landing and living on their own on another planet. Many research questions must be addressed before safely sending humans to explore Mars, when our current knowledge on humans in space does not exceed 14 months in only one individual. A human research roadmap for tackling these research questions has been recently detailed by NASA [http://humanresearchroadmap.nasa .gov/] [2]. The effects of space flight conditions on the autonomic nervous system could be at the origin of two medical issues experienced by a significant number of astronauts. These issues are space motion sickness immediately after entering weightlessness or after returning to Earth’s gravity, and post-flight orthostatic intolerance. As discussed below, due to shared neural pathways, clinical treatment of one condition often interacts with the other condition.

Space motion sickness is a special form of motion sickness, which include such symptoms as depressed appetite, a nonspecific malaise, gastrointestinal discomfort, nausea, and vomiting. Symptoms have their onset from minutes to hours after orbital insertion. Excessive head movement early on-orbit generally increases these symptoms. Symptom resolution usually occurs between 30 and 48 hours, with a reported range of 12 to 72 hours, and recovery is rapid (Fig. 59.1). Even if someone doesn’t literally get sick to their stomach, they may feel a less dramatic motion-sickness effect known as “sopite syndrome”, characterized by lethargy, mental dullness, and disorientation. Many astronauts have noticed this effect, which they call “mental viscosity” or “space fog”. About two-thirds of the Space Shuttle astronauts and Soyuz cosmonauts experience some symptoms of space motion sickness. There are no statistically significant differences in symptom occurrence between pilots versus non-pilots, males versus females, different age groups, or first time flyers versus veterans repeat flyers. An astronaut’s susceptibility to space motion sickness on his/her first flight correctly predicted susceptibility on the second flight in 77% of the cases. In other words, astronauts who have been sick during their first flight are likely to be sick again during their subsequent flights [3]. Many astronauts returning to Earth after long-duration stays on board the International Space Station now experience symptom recurrence at landing (Fig. 59.1). The severity of the symptoms and the functional recovery after the flight seem to be directly proportional to the time on orbit [4]. Head or full body movements made upon transitioning from microgravity to a gravitational field less than that on Earth, and vice versa, may not be as provocative. It is interesting to note that of the twelve Apollo astronauts who walked on the Moon, only three reported mild symptoms, such as stomach awareness or loss of appetite prior to extravehicular activities. None reported symptoms while in the one-sixth gravity of the lunar surface, and no symptoms were noted upon return to weightlessness after leaving the Moon surface. There are considerable individual differences in susceptibility to space motion sickness, and currently it is not

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FIGURE 59.1 Space motion sickness incidence (dashed line) and drug treatment (continuous line) during early in-flight and post-flight periods. The incidence (y-axis on the left) is measured on a scale from 0 (no symptoms) to 10 (vomiting). The drug treatment (y-axis on the right) is measured as the percentage of motion sickness medication taken by the crew compared to the other medication used during the same period. Note that flight rules restrict docking of spacecraft before flight day two and extravehicular activities before flight day three to allow for space motion sickness symptoms resolution.

possible to predict with any accuracy those who will have some difficulty with sickness while in space. Although anti-motion sickness drugs offer some protection, they may interfere with the adaptation process, and symptoms controlled by these drugs are experienced again once treatment ceases. This was observed for scopolamine, which resulted in a shift towards the use of promethazine. There have been anecdotal reports of medication usage prior to extravehicular activities, with concerns about cognitive and performance side effects associated with this usage. Other issues related to the adaptation of the central nervous system through the vestibular pathways include: (a) the perceptual effects and illusions of free-falling, visual reorientation illusions, and acrophobia episodes during extravehicular activity; (b) decreased sensorimotor performance and oscillopsia during re-entry; (c) disequilibrium and ataxia when standing and walking after landing; and (d) g-state flashbacks during unusual stimulation of the vestibular system during the re-adaptation period following landing [5]. A sensory conflict theory of motion sickness postulates that motion sickness occurs when patterns of sensory inputs to the brain are markedly re-arranged, at variance with each other, or differ substantially from expectations of the stimulus relationships in a given environment. In orbit and during re-entry, sensory conflict can occur in several ways. First, there can be conflicting information regarding tilt transmitted by the otoliths and the semicircular canals. Sensory conflict may also exist between the visual and vestibular systems during motion; the eyes transmit information to the brain indicating body movement, but no corroborating impulses are received from the otoliths, such as during car sickness. A third type of conflict may exist in space because of differences in perceptual habits and expectations. On Earth, we develop a neural

store of information regarding the appearance of the environment and certain expectations about functional relationships, e.g., the concepts of “up” and “down”. In space, these perceptual expectations are at variance, especially during the illusions described above.

ORTHOSTATIC INTOLERANCE It is conceivable that in the same way the central nervous system elaborates an internal representation of gravity for spatial orientation using several sensory inputs, the sympathetic activation and maintenance of blood pressure during orthostatism would be based on an internal representation of the intravascular pressure, previously built based on the same information as used for the internal representation of gravity. On return to Earth, post-flight orthostatic intolerance is characterized by a variety of symptoms that follow standing: lightheadedness, increase in heart rate, decrease in blood pressure, and pre-syncope or syncope. Both orthostatic intolerance and diminished exercise capacity become more severe with longer exposure to microgravity and require more lengthy recovery times after returning to Earth. Orthostatic intolerance affects approximately 30% of short-duration and 80% of long-duration crewmembers. This may pose a problem for “space tourists” during suborbital flight. They will be exposed to re-entry forces even higher that those experienced in the Space Shuttle or Soyuz capsules by well cardiovascular fit professional astronauts. Another threat is whether a debilitated crew can respond to an emergency upon landing [6]. The extent of orthostatic intolerance post-flight is variable and depends on the duration of the flight, individual differences in cardiovascular function among the

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astronauts, and the elapsed time after landing and method of post-flight testing. Recovery to the preflight level of orthostatic tolerance occurs within a day or so following flights of less than one-month's duration, but longer recovery is associated with longer duration flights. In contrast to space motion sickness, there is a well-known gender difference in orthostatic tolerance (see Chapters 106–108). Post-flight orthostatic intolerance is probably induced by multiple factors, including changes in hemodynamics, alterations in baroreceptor reflex gain, decreases in exercise tolerance and aerobic fitness, hypovolemia, and altered sensitivity of β-adrenergic receptors in the periphery [7]. Astronauts in orbit experience a headward shift of fluid at the onset of weightlessness resulting in facial puffiness and distended neck veins. This fluid shift causes distension of heart chambers, which in turn activates mechanisms associated with rapid blood volume reduction. The effects of a reduced blood volume are probably amplified by changes in the low-pressure venous system of the lower limbs and by impaired baroreflex function. Changes in the otolith signals, or their reinterpretation, may also be involved. Indeed, the removal of the normal head-to-foot gravity vector acts not only on fluid, because of the loss of hydrostatic pressure gradient, but also on the otolithic system. A central reinterpretation of otolith signals is presumably taking place during re-adaptation to Earth gravity. However, no data are available yet on the possible role of non-cardiovascular inputs controlling the activity of the sympathetic nervous system during or after space flight. Because the otolithic control of the cardiovascular system is supposed to compensate for head tilt coupled with the observation that otolith tilt reflexes generally vanish during adaptation to microgravity, it can be hypothesized that the otolithic control of the cardiovascular system will be altered after space flight. This alteration would then participate in cardiovascular deconditioning. If this hypothesis is confirmed, it could have potential consequences for the design of countermeasures preventing cardiovascular deconditioning. In this context, providing artificial gravity by using centrifugation in supine subjects with the head off-center might be an effective means for maintaining otolith sensitivity and preserving vestibulosympathetic reflexes [8].

CLINICAL INTERACTIONS The pharmacological intervention for motion sickness and orthostatic intolerance has been an issue after returning from space flights. As stated above, promethazine has been the preferred medication by space medicine for treatment of motion sickness. However, promethazine significantly increases the incidence of orthostatic hypotension, presumably via inhibition of sympathetic responses [9]. Midrodrine has been successful in preventing orthostatic hypotension post-flight. However, its effects are negated by promethazine and this combined usage can also lead to akathisia. Other countermeasures, such as fluid loading and compression garments, have provided successful nonpharmaceutical approaches for orthostatic intolerance. The unique challenges provided by adaptation to the conditions of space flight highlight the interactions in the autonomic nervous system, particularly for those physiological mechanisms that contribute to cardiorespiratory regulation and spatial awareness during changes in posture.

References [1] Clément G. Fundamentals of space medicine, 2nd ed. New York: Springer; 2011. [2] Clément G, Reschke MF. Neuroscience in space. New York: Springer; 2008. [3] Bacal K, Bilica R, Bishop S. Neurovestibular symptoms following space flight. J Vestib Res 2003;13:93–102. [4] Ortega HJ, Harm DL. Space and entry motion sickness. In: Barratt M, Pool S, editors. Principles of clinical medicine for space flight. New York: Springer; 2008. p. 211–22. [Chapter 10] [5] Paloski WP, Oman CM, Bloomberg JJ, Reschke MF, Wood SJ, Harm DL, et al. Risk of sensory-motor performance failures affecting vehicle control during space missions: A review of the evidence. J Gravit Physiol 2008;15:1–29. [6] Hamilton D. Cardiovascular disorders. In: Barratt M, Pool S, editors. Principles of clinical medicine for space flight. New York: Springer; 2008. p. 317–60. [Chapter 16] [7] Yates BJ, Kerman IA. Post-spaceflight orthostatic intolerance: Possible relationship to microgravity-induced plasticity in the vestibular system. Brain Res Rev 1998;28:73–82. [8] Antonutto G, Clément G, Ferretti G, Linnarsson D, Pavy-Le Traon A, Di Prampero P. Physiological targets of artificial gravity: The cardiovascular system. In: Clément G, Bukley A, editors. Artificial gravity. Hawthorne and Springer: New York: Microcosm Press; 2007. p. 137–62. [Chapter 5] [9] Shi SJ, Platts SH, Ziegler MG, Meck JV. Effects of promethazine and midodrine on orthostatic tolerance. Aviat Space Environ Med 2011;82:9–12.

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60 Hypothermia and Hyperthermia Nisha Charkoudian INTRODUCTION The human body is remarkably capable of maintaining core temperature within a few tenths of a degree of 37°C over a very wide range of environmental exposures and activity levels. Sympathetic neural mechanisms controlling skin blood flow and sweating are central to these abilities, in addition to shivering which increases heat production during cold exposure.

CENTRAL NEURAL CONTROL OF THERMOREGULATION The main area which controls body temperature within the central nervous system is the preoptic area of the anterior hypothalamus (PO/AH). This region contains temperature-sensitive and temperature-insensitive neurons which interact to regulate the systemic thermoregulatory response to a given environment or situation (for review, see Boulant [1]). For example, when body temperature increases, “warm-sensitive” neurons in the PO/AH region are activated. This then initiates a series of neural events which activates heat dissipation via cutaneous vasodilation and sweating. Changes in the activity of neurons in the PO/AH region are linked to appropriate increases or decreases in heat dissipation, as discussed below, as well as to increases in heat generation (shivering) during body cooling [1]. Thermoregulatory control at the PO/AH occurs via both directly sensed temperature and via integration with input from peripheral thermosensory afferents which give information about peripheral body temperature, in particular surface temperature [1,8]. Thus, skin temperature can have a significant influence on the rate of a thermoregulatory response (e.g., sweating) for a given level of core temperature. A practical example is that exercising in a cool environment might require a lower sweating rate compared to the same exercise in a warmer environment, even if core temperature is similar in the two conditions. Thus, the skin temperature influence can contribute to the optimization of the integrated sweating response. The relative contributions of core and surface temperature can vary, but in general the relative influence of core to mean skin temperature is about 9:1 [1,8].

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The neural network involving the PO/AH and other inputs results in a set-point for thermoregulation, and changes in the activity of the network when body temperature goes above or below that set-point result in activation of efferent neuronal activation of appropriate thermoregulatory pathways [1]. The set-point concept is related to the concept of a threshold for activation of a certain thermoregulatory effector. The temperature (most often core temperature) at which a certain thermoregulatory response (e.g., sweating) is initiated is referred to as the threshold for the activation of that response. The gain or sensitivity of the response refers to the incremental changes (slope) of the response relative to body temperature with further changes in body temperature beyond the threshold [1,2,8]. The integrated control of body temperature acts as a classical negative feedback loop, as summarized in Figure 60.1. Changes in body temperature are sensed peripherally and centrally, and appropriate changes in heat dissipation and/or heat generation act to minimize or reverse those changes. New information subsequently feeds back to the PO/AH which then appropriately modulates the integrated thermoregulatory response.

REGULATION OF BODY TEMPERATURE IN THERMONEUTRAL ENVIRONMENTS During most daily activities in healthy people, only minor changes in heat dissipation are required to maintain normothermia during changes in activity or when one moves to a slightly warmer or cooler environment. This range of environment/activity combinations has been referred to as the “neutral zone” of thermoregulation [7]. The minor changes in thermoregulation required for this “zone” are brought about primarily by small changes in skin blood flow mediated by modulations in the activity of sympathetic noradrenergic vasoconstrictor nerves innervating the cutaneous circulation [2,3,7]. These cutaneous vasoconstrictor nerves exhibit tonic activity in normothermia, and are responsible for keeping normothermic skin blood flow relatively low (whole body skin blood flow during normothermia is ~250 mL/min). In a slightly warmer environment, small decreases in vasoconstrictor activity cause small “passive” vasodilation (and increased heat loss), and conversely in a slightly cooler environment,

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increases in vasoconstrictor activity decrease skin blood flow (and conserve heat).

HYPOTHERMIA During exposure to cold environments, the goal of the thermoregulatory response is to maintain core body temperature, or to minimize any drops in body temperature that may occur (Fig. 60.1, right side). The two physiological “strategies” for the maintenance of core temperature are (1) to decrease dissipation of heat from the body to the environment; and (2) when necessary, to increase metabolic generation of body heat to offset loss of heat to the environment. The initial response to exposure to a cold environment is sympathetic neurally-mediated vasoconstriction in the skin, which decreases convective heat transfer from the core to the surface of the skin and thus decreases heat loss from the body [2,3,9]. During mild body cooling, cutaneous vasoconstriction may be sufficient as a thermoregulatory response. During more severe or prolonged cooling, however, the physical gradient for heat loss (difference in temperature between the body and the environment) is so great that cutaneous vasoconstriction alone is insufficient to maintain body temperature. In this case, an increase in metabolic heat production is required to offset the heat lost, and maintain internal temperature. Shivering involves involuntary, rhythmic muscle contractions which occur solely for the purpose of generating heat; i.e., not for locomotion or to accomplish some other activity. The increase in metabolic rate that occurs during shivering is substantial, in the order of the increase in metabolic rate that occurs during mild to moderate exercise [9]. Because heat is a byproduct of increased

PO/AH

increased body temperature (–) cutaneous vasodilation sweating

decreased body temperature (–) cutaneous vasoconstriction shivering

FIGURE 60.1 Schematic summary of the negative feedback loops involved in human physiological thermoregulation. Responses to hyperthermia are shown on the left side of the figure, where increased body temperature elicits increased heat dissipation via cutaneous vasodilation and sweating. Responses to hypothermia are shown on the right side of the figure, where decreased body temperature elicits cutaneous vasoconstriction (to decrease heat loss) and shivering (to increase metabolic heat production). These responses then minimize or reverse the initial change in body temperature.

skeletal muscle metabolism (i.e., muscle contraction), shivering results in an increase in heat generation. This increased heat production protects against hypothermia by helping to maintain core temperature. In neonatal humans, and in adults of other species such as rats, a phenomenon exists called “non-shivering thermogenesis”. Non-shivering thermogenesis occurs via sympathetic neurogenic activation of brown adipose tissue, in which uncoupling protein-1 (UCP1) uncouples metabolic activity from the generation of ATP. Therefore, activation of brown adipose tissue results in the generation of heat in the absence of shivering. It is generally accepted that nonshivering thermogenesis does not exist in adult humans [9]. Thermoregulatory responses to environment which involve specific activity or behaviors are referred to as “behavioral thermoregulation”. This is in contrast to the physiological thermoregulatory responses outlined above, and include huddling, putting on warmer clothing or going indoors when one is uncomfortable outdoors. In general, humans tend to rely to a greater extent on behavioral thermoregulation in the cold, since human physiological responses to a cold environment are limited compared to other species [9].

HYPERTHERMIA Human exposure to hyperthermia is usually due to some combination of environmental heat exposure and/or exercise. It is relevant in this context that metabolic heat production (from exercising skeletal muscle) can increase as much as 10 to 20-fold with intense exercise. Thus even mild to moderate exercise represents a substantial, “endogenous” heat stress, and would result in dangerous increases in body temperature were it not for appropriate physiological heat loss responses. Even resting heat exposure can increase body temperature substantially, so when the two stresses are combined, the increased thermal load requires substantial heat dissipation. Physiological thermoregulatory responses to heat in humans are much more efficient than responses to cold; therefore, physiological thermoregulation is primary during hyperthermia (although behavioral responses contribute as well, such as seeking out cooler environments or wearing lighter clothing). Thermoregulation during hyperthermia involves increasing the dissipation of heat from the core to the environment. This is done in two ways: (1) by increasing the convective transfer of heat from the core to the skin via increased skin blood flow; and (2) by cooling the skin with evaporative heat loss by sweating.

Skin Blood Flow The sympathetic innervation of the human skin circulation involves two distinct branches. Sympathetic noradrenergic vasoconstrictor nerves, mentioned above, are tonically active and release norepinephrine

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ovERAll SummARy

and cotransmitters which cause vasoconstriction. The sympathetic active vasodilator system is less well understood, but appears to function via cholinergic nerve co-transmission [5]. A specific neurotransmitter(s) for the active vasodilator system has not been identified, although several substances appear to contribute to the reflex vasodilator response (for review, see [2]). During increases in body temperature due to environmental heat exposure or exercise, the initial vasodilator response in the skin is caused by withdrawal of vasoconstrictor neural activity, and is therefore referred to as “passive” vasodilation. Further increases in body temperature result in activation of the active vasodilator system, which is responsible for 80–90% of the very large vasodilation that can occur during whole body heating [4]. During severe hyperthermia, skin blood flow can increase from its low baseline values to as much as 6–8 L/min or 60% of cardiac output [6].

Sweating Sweating works in concert with cutaneous vasodilation to optimize heat dissipation during hyperthermia. The cooler skin accomplished by the sweating response allows the heat transfer from skin blood flow to be more efficient: when the warmer blood arrives at the skin surface, a cool skin results in a larger thermal gradient and therefore more efficient heat loss from the body. The process of sweating involves the secretion of a hypotonic saline solution onto the surface of the skin, which then evaporates, cooling the skin. Particularly when environmental temperature is warmer than body temperature, evaporative heat loss is necessary to cool the skin surface and facilitate heat transfer away from the body core. Sweating is caused by activation of sympathetic sudomotor nerves innervating eccrine sweat glands, which cover most of the body surface (for review, see [8]). These nerves release acetylcholine, which interacts with muscarinic cholinergic receptors at the sweat gland and triggers the onset of sweating. Sweating can therefore be blocked by local or systemic administration of atropine. Because sweat evaporation is essential to the process of cooling the skin, sweating itself will not cause heat dissipation if evaporation does not occur. Therefore, in dry environments, where evaporation of sweat can occur easily, evaporative skin cooling (and therefore overall heat

dissipation) is much more efficient than in humid environments of similar absolute temperature. Sweat which does not evaporate but simply drips off the skin is “wasted” from the perspective of heat dissipation and overall thermoregulation.

OVERALL SUMMARY As summarized in Figure 60.1, human thermoregulation is a complex, integrative process which relies on multiple organ systems working together in a coordinated fashion to optimize body temperature for a variety of environmental and activity conditions. Responses to changes in body temperature are coordinated centrally at the preoptic area of the anterior hypothalamus. Major physiological responses during hypothermia in humans are cutaneous vasoconstriction and shivering. During hyperthermia, heat is dissipated from the body via cutaneous vasodilation and sweating.

References [1] Boulant JA. Neuronal basis of Hammel's model for set-point thermoregulation. J Appl Physiol 2006;100:1347–54. [2] Charkoudian N. Mechanisms and modifiers of reflex induced cutaneous vasodilation and vasoconstriction in humans. J Appl Physiol 2010;109:1221–8. [3] Hodges GJ, Johnson JM. Adrenergic control of the human cutaneous circulation. Appl Physiol Nutr Metab 2009;34:829–39. [4] Johnson JM, Proppe DW Cardiovascular adjustments to heat stress. In: Handbook of physiology environmental physiology. New York: Oxford University Press; 1996. p. 215–44. [5] Kellogg Jr DL, Pergola PE, Piest KL, Kosiba WA, Crandall CG, Grossmann M, et al. Cutaneous active vasodilation in humans is mediated by cholinergic nerve cotransmission. Circ Res 1995;77:1222–8. [6] Rowell L.B. Cardiovascular adjustments to thermal stress. In: Handbook of physiology the cardiovascular system: peripheral circulation and organ blood flow. Bethesda: American Physiological Society; 1983. p. 967–1023. [7] Savage MV, Brengelmann GL. Control of skin blood flow in the neutral zone of human body temperature regulation. J Appl Physiol 1996;80:1249–57. [8] Shibasaki M, Wilson TE, Crandall CG. Neural control and mechanisms of eccrine sweating during heat stress and exercise. J Appl Physiol 2006;100:1692–701. [9] Young AJ, Castellani JW. Exertion-induced fatigue and thermoregulation in the cold. Comp Biochem Physiol A Mol Integr Physiol 2001;128:769–76.

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61 Psychological Stress and the Autonomic Nervous System Michael G. Ziegler INTRODUCTION

Patterns of Autonomic Response to Stress

Autonomic responses to psychological stress prepare the body for fight or flight. Cannon and Selye described stereotypic responses to stress that involved activation of sympathetic nerves and adrenocortical hormone release. Corticotropin-releasing factor (CRF) in the central nervous system activates autonomic and adrenocortical responses to stressors. CRF injected into the brain increases arousal and responsiveness to stressful stimuli. On the other hand, CRF antagonists can reverse behavioral responses to many stressors. CRF alters the discharge of locus coeruleus neurons in the brainstem and these effects are mimicked by some stressors. Locus coeruleus noradrenergic neurons project into the cerebral cortex. In the paraventricular nucleus of the hypothalamus, norepinephrine stimulates further CRF release. Other central nervous system pathways also mediate stress-induced activation of the sympathetic nervous system but they have generally not been studied as well as the CRF pathways [1].

Normal Psychological Stresses and Autonomic Activity There is a 24-hour rhythm in the plasma levels of norepinephrine and epinephrine which tend to be lowest at 3 a.m., and rise rapidly until a peak at 9 a.m. similar to the cortisol rhythm. Sleep decreases sympathetic nerve activity. Arousals during sleep cause a rapid increase in muscle sympathetic nerve electrical activity which falls back to baseline when sleep resumes. Muscle sympathetic nerve activity nearly doubles when going from sleep to wake and doubles yet again when going from recumbent to standing posture. Exercise, pain, and cold lead to even more dramatic increases in sympathetic nerve activity which is detailed in Chapters 80 and 81 of this book. These are normal responses to the stresses of day to day life which may help explain why there is a preponderance of myocardial infarction and sudden death between 6:00 a.m. and noon. The autonomic responses to stress might also help explain early mortality during both unemployment and bereavement.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00061-5

The general response to both physical and psychological stress is activation of the sympathetic nervous system with inhibition of the parasympathetic nervous system. When stress becomes severe or uncontrolled then adrenomedullary release of epinephrine ensues. As stress increases even further then CRF not only activates the sympathetic nervous system but leads to the release of adrenocorticotropic hormone (ACTH) and adrenocortical steroids. Sympathetic nervous stimulation constricts muscle vasculature and increases peripheral vascular resistance. Sympathetic nerves that supply the skin both vasoconstrict and supply sweat glands through sympathetic cholinergic innervation. Activation of this skin sympathetic pathway can precipitate a “cold sweat” or perspiration and flushing of the skin. During sleep, muscle sympathetic nervous activity and skin sympathetic nervous activity have similar firing intensities and frequencies. However, during stress, muscle and skin sympathetic nerve activities diverge. When a doctor took patients’ blood pressure, skin sympathetic nerve activity increased by 38% while muscle sympathetic nerve activity decreased by 25%. This was accompanied by an apparent increase in sympathetic nerve activity to the heart as heart rate and blood pressure increased [2]. Some stimuli such as hypoglycemia elicit a fairly specific activation of adrenomedullary epinephrine release without a marked increase in sympathetic nerve activity. When medical residents climbed stairs, their plasma norepinephrine increased; however, when they presented a public speech, epinephrine levels showed an even greater increase [3]. Psychological stress not only tends to increase epinephrine disproportionately, it also tends to increase sympathetic nerve activity to the heart, leading to increased cardiac output. As we age, norepinephrine release in response to the cold pressor test increases. Less epinephrine is released from the adrenal medullae in the elderly; however, epinephrine blood levels do not decrease because of diminished clearance of epinephrine from the circulation with advancing age.

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61. PSyCHologICAl STRESS ANd THE AuToNomIC NERvouS SySTEm

CARDIAC DISEASE Psychological stress affects the heart in many ways. Stress increases heart rate by withdrawal of parasympathetic input and increased sympathetic stimulation. This is commonly accompanied by increased blood pressure leading to an increased rate–pressure product and myocardial oxygen consumption. Although the increased oxygen demand requires increased blood flow through the coronary arteries, studies of patients during mental stress showed increased norepinephrine spillover from the heart and a reduction of blood flow through regions of high stenosis by an average of 27% [4]. The combination of hypoxia and sympathetic stimulation can cause arrhythmias. Pigs with an experimentally induced coronary occlusion developed ventricular fibrillation when restrained unless they were adapted to the restraint [5]. Implanted cardioverters discharge in response to dangerous ventricular arrhythmias. Patients experiencing intense anger have an estimated seven times greater risk of cardioverter discharge during both mental and physical stress. There is epidemiological evidence that psychological stress can cause arrhythmias and fatal human cardiac events. After the Northridge earthquake, coroners’ records for Los Angeles County found that sudden deaths from cardiac causes increased from an average of 4.6 per day in the week before to 24 on the day of the earthquake [6]. Studies of the 1995 Hanshin-Awaji earthquake in Japan showed similar results [7]. Psychological stress can precipitate angina pectoris, indicating that it causes myocardial ischemia in patients with coronary artery disease. It can also cause cardiac arrhythmias, triggering cardioverter discharge. Emotional stress can also increase blood levels of fibrinogen, von Willebrand factor, factors VII and VIIII and fibrin D-dimer and activate platelets, all of which can predispose to coronary occlusion [7]. These acute cardiac consequences of stress may be exacerbated if chronic stress is associated with high blood pressure (see Chapter 71). Extreme sympathetic nervous system activation in stress probably plays a role in stress (takotsubo) cardiomyopathy as well as in panic attacks accompanied by coronary artery spasm. The high level of sympathetic firing in these disorders is accompanied by release of neuropeptide Y (NPY), which can lead to prolonged vasoconstriction [8].

GASTROINTESTINAL (GI) CONTROL The thought of food can elicit salivation, gastric motility and acid secretion. Stress inhibits GI motility when it activates the sympathetic nervous system, primarily through release of norepinephrine at its synaptic interface with the enteric nervous system. Postganglionic projections from sympathetic nerves terminate in myenteric submucous ganglia of the enteric nervous system where they suppress motility and secretion. Alpha adrenergic stimulation inhibits both GI secretion and GI blood flow. In animal

models, cold water immersion is associated with an inhibition of gastric emptying. While stress decreases gastric emptying and intestinal motility it does not decrease colonic motility. Sacral parasympathetic stimulation of the large bowel during stress is accompanied by degranulation of enteric mast cells triggered by postganglionic sympathetic nerve release of corticotrophin releasing hormone (CRH) [5]. This releases inflammatory mediators and is postulated to underlie the secretory diarrhea and abdominal discomfort associated with stress.

PSYCHOSOMATIC DISORDERS AND THE AUTONOMIC NERVOUS SYSTEM Autonomic responses to stress frequently lead to medical care. Feelings of warmth and cold, palpitations, tachycardia, nausea, abdominal pain, diarrhea, and constipation can all be the consequence of autonomic stress responses. Twenty per cent of patients with borderline hypertension in the doctor’s office have entirely normal home blood pressures. Sympathetic nervous stimulation of the heart increases heart rate, cardiac output and blood pressure in novel or stressful environments. This cardiovascular stress response increases myocardial oxygen consumption and can precipitate angina pectoris in patients with coronary artery disease. A vasovagal response can be triggered by a stressful situation that makes a person want to run away even though social constraints prevent the person from leaving. When this happens, hypothalamic activation of medullary cardiovascular responses triggered by emotional stress can lead to increased inotropic stimulation to the heart. This can stimulate ventricular mechano-receptors and promote vasodilation, bradycardia, and fainting.

POST-TRAUMATIC STRESS DISORDER, PANIC, AND ANXIETY Post-traumatic stress disorder (PTSD) occurs when intrusive thoughts elicit memories of an unusually stressful event. Nineteenth century descriptions of “soldier’s heart” noted abnormal excitability of the cardiac nervous system in the absence of serious cardiac disease. PTSD is often accompanied by tachycardia, palpitations, and high blood pressure [4,10]. In panic disorder, a psychological stimulus elicits an autonomic response characterized by flushing, tachycardia, palpitations, hypertension, and gastrointestinal symptoms. The autonomic response can sometimes be extinguished by repeated exposure to the stimulus under reassuring circumstances. In anxiety disorder, similar autonomic symptoms occur with no inciting stimulus. In all three of these psychological disorders baseline norepinephrine and epinephrine are normal but plasma and urinary catecholamines increase when symptomatic reactions

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Autonomic responses to psychological stress serve the useful function of preparing us for action by increasing muscle blood supply and slowing vegetative functions. However, inappropriate stress responses are the basis for many psychosomatic disorders. Familiarity with the patterns of autonomic response to psychological stress is essential to understanding human disease.

TABLE 61.1 Sympathetic Nerve Responses to Stress Organ

Response

Receptor

Cardiac atrium

Heart rate

β1

Cardiac ventricle

Inotropism

β1

Eye

Pupil dilation

α1

Skin blood vessel

Constriction

α1

Hand sweat glands

Sweat

Cholinergic

Blood vessels

Constriction

α1

Salivary glands

Constriction, dry mouth

α1, α2

Gut

Decrease motility

α1, α2, β2

Gut sphincters

Contraction

α1

Kidney

Renin release

β1

Bladder

Relaxation

β2

Bladder sphincter

Contraction

α1

Hair

Piloerection

α1

Muscle cells

Glycogenolysis

β2

Muscle cells

K uptake

β2

Muscle blood vessels

Dilation

β2

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References

are triggered [5,11]. Beta adrenergic blocking drugs tend to diminish subjective symptoms of palpitations and tremor and they often eliminate episodes of tachycardia.

[1] Koob GF. Corticotropin-releasing factor, norepinephrine, and stress. Biol Psychiatry 1999;46:1167–80. [2] Grassi G, Turri C, Vailati S, Dell'Oro R, Mancia G. Muscle and skin sympathetic nerve traffic during the “white-coat" effect. Circulation 1999;100:222–5. [3] Dimsdale JE. Psychological stress and cardiovascular disease. J Am Coll Cardiol 2008;51:1237–46. [4] Steptoe A, Brydon L. Emotional triggering of cardiac events. Neurosci Biobehav Rev 2009;33:63–70. [5] Chrousos GP. Stress and disorders of the stress system. Nat Rev Endocrinol 2009;5:374–81. [6] Appels CW, Bolk JH. Sudden death after emotional stress: a case history and literature review. Eur J Intern Med 2009;20:359–61. [7] Steptoe A, Brydon L. Emotional triggering of cardiac events. Neurosci Biobehav Rev 2009;33:63–70. [8] Esler M. Pathophysiology of the human sympathetic nervous system in cardiovascular diseases: the transition from mechanisms to medical management. J Appl Physiol 2010;108:227–37. [9] Plourde V. Stress-induced changes in the gastrointestinal motor system. Can J Gastroenterol 1999;13(Suppl A):26A–31A. [10] Lamprecht F, Sack M. Posttraumatic stress disorder revisited. Psychosom Med 2002;64:222–37. [11] Hoehn-Saric R, McLeod DR. Anxiety and arousal: physiological changes and their perception. J Affect Disord 2000;61:217–24.

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62 Mind–Body Interactions Calvin Carter, Daniel Tranel INTRODUCTION Mind–body interactions are a two-way street. For example, perceptions and emotions (mind) influence many bodily functions, such as sweating, heart rate, blood pressure, and gastrointestinal smooth muscle contractions (body). Conversely, different physiological states (e.g., as in health and disease) influence an individual’s mental state and mood. The study of these mind–body interactions has been greatly enhanced by three techniques which quantify arousal of the autonomic nervous system following stimulation: the electrodermal skin conductance response (SCR), the electrogastrogram (EGG), and cardiovascular parameters (heart rate and blood pressure; HR and BP). The SCR is a remarkably powerful and informative psychophysiological index. Because SCRs are relatively easy to measure, and provide reliable indices of a wide variety of psychological states and processes, SCRs have been arguably the most popular aspect of the autonomic nervous system (ANS) activity used to study human cognition and emotion [1]. The EGG records the stomach’s smooth muscle contractions, called peristalsis, measuring the activity of the digestive and gastrointestinal (GI) systems. The primary function of the GI system is to process ingested foods and liquids, digesting and absorbing nutrients and water and excreting wastes. These functions are highly regulated by the enteric nervous system, the only branch of the ANS that can function independently of the sympathetic (SNS) and parasympathetic nervous system (PNS) [2]. Finally, cardiovascular (CV) responses, including changes in heart rate (HR) and blood pressure (BP), are regulated by the SNS and PNS. Cardiovascular responses are particularly useful in ethnic studies due to robust differences in CV arousal between ethnic groups [3]. Our group and others have used these techniques to study mind–body interactions in patient, meditation and ethnic based studies.

SKIN CONDUCTANCE RESPONSE (SCR)

The Somatic Marker Hypothesis

Decision-Making Making advantageous decisions in everyday situations requires conscious reasoning; however, non-conscious

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00062-7

biases can also come into play, and can influence many aspects of decision-making. Using SCRs, we found that healthy individuals generate non-conscious autonomic responses (SCRs) while pondering a decision that later turned out to be risky. These processes are mediated by the ventromedial (VM) prefrontal sector of the brain. In our experiment, participants were asked to win the most and lose the least amount of money in a card game, using a loan of $2000. Study participants chose to play from two different sets of card decks, one set of decks was advantageous in the long run (the “good deck” because participants won more money than they lost) and the other set was disadvantageous (the “bad deck” because participants lost more money than they won). The participants had no way to predict which of the decks was advantageous or disadvantageous. Healthy participants began to generate SCRs before choosing a card from the “bad deck” and eventually avoided these decks whereas VM patients could do neither (Fig. 62.1) [4]. Furthermore, these SCRs preceded conscious awareness of these decks being disadvantageous (“pre-hunch” period) as determined by questioning the participants throughout the course of the experiment. After playing further, healthy participants became aware of the “good and bad” decks (“hunch period”). Later in the game, healthy participants figured out the nature of the game and expressed “why” certain decks were more advantageous than others (“conceptual period”). During all three periods healthy participants continued to generate SCRs whenever they considered choosing a card from a “bad” deck and subsequently chose cards from the good decks (Fig. 62.1). By contrast, none of the VM patients generated anticipatory SCRs nor expressed a hunch and only few became aware of why the decks were good or bad. Yet, VM patients continued to choose from the “bad deck” despite their overt knowledge of what was going on in the game (Fig. 62.1) [4], suggesting that the VM plays a role in pre-monitory decision-making.

The findings reviewed here, and other related observations, have led to the development of a framework that is termed the somatic marker hypothesis. In a nutshell, the theory posits that feelings and emotions give rise to “somatic

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62. MInd–Body InTERACTIons

FIGURE 62.1 Aberrant decision-making in patients with VM damage. The magnitude of anticipatory SCRs is significantly lower in VM patients compared to controls during all experimental periods (pre-hunch, hunch and conceptual period). Healthy controls chose more and more cards from the good decks as they gained more insight into the experiment; in contrast, VM patients continued to choose cards from bad decks despite their increasing knowledge of what was going on in the task. Experimental periods: Pre-hunch  no knowledge of the task; Hunch  Insight into task; Conceptual  Verbalized why the task worked the way it did (adopted with permission from [4]).

markers,” which serve as guideposts that help steer behavior in an advantageous direction. Deprived of these somatic markers, VM patients lose the ability to experience appropriate emotional responses to various stimuli and events. We have proposed that the absence of these emotional responses – evidenced, for example, by the missing SCRs in the experiments described above – leads to defective planning and decision-making; this, in turn, leads to socially inept and inappropriate behavior that is characteristic of VM patients [5].

ELECTROGASTROGRAM (EGG) Emotion One great avenue for studying mind–body interactions is the brain–gut axis. Have you ever felt butterflies in your stomach before a big interview? The brain–gut connection is the likely culprit as both organs are intimately linked via the vagus nerve. The process begins with an external stimulus (e.g. a traumatic life event) causing anxiety which in turn activates hypothalamic outputs to the pituitary and pontomedullary nuclei. Both structures mediate the neuroendocrine and autonomic outputs to the body and the gut via the vagus nerve [5]. The study of this mind–body interaction has been greatly improved with the introduction of the electrogastrogram in recent years [6]. It is wellestablished that psychological factors influence the activity of the gastrointestinal system, including the stomach and intestine, however, the role of the GI system on a person’s

psychological and emotional state has not been welldocumented [6]. Using EGGs, our lab investigated the role of the gastrointestinal system in one’s subjective emotional experiences. We studied patients with Crohn’s disease, an inflammatory disease of the gastrointestinal system resulting in increased gut sensitivity, in the active phase (Crohn’s-active, CA) or silent/remissive phase (Crohn’ssilent, CS) and healthy participants (HP). We hypothesized that the gut influences emotions and feelings and consequently, participants with an actively abnormal GI (CA participants) would have aberrant feelings of emotion compared with participants with near-normal or normal GIs (CS and NC participants). To test this, we measured participant’s EGG activity while watching emotionally charged clips (happy, disgust, fear and sad), baseline measurements were obtained prior to viewing film clips. Following each presentation of a film clip, each participant completed a self-report questionnaire that assessed the intensity of their gastrointestinal sensations and their subjective emotional experience in response to each film clip. We found that patients with CA demonstrated increased gastrointestinal activity before stimuli were introduced (Fig. 62.2A) and self-reported greater feelings of negative emotions (disgust, fear and sad) after viewing emotionladen film clips compared to CS patients and healthy participants (Fig. 62.2B). Additionally, none of the participants from any group stated a change in gastrointestinal feelings after viewing film clips [7]. These results suggest a cause and effect relationship between the body (GI activity) and one’s subjective emotional experience (self-report) in which abnormal GI activity pre-disposed CA patients to

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297

(C)

(D)

Self-reported emotion ratings

(B) 7

*

6 5 4

Disgust

3

Fear

2

Sad

1 0 HP

CA

0.5 1

2

3

4 5 Arousal

6

7

1

2

3

4 5 Arousal

6

7

CS

CA

1

NC

–1.5 –1.0 –0.5 0.0

z-score peak EGG

10

*

z-score peak EGG –1.5 –1.0 –0.5 0.0 0.5 1.0

EGG Activity

(A)

0e.00 1e.07 2e.07 3e.07 4e.07 5e.07 6e.07

CARdIovAsCulAR ARousAl (HR And BP)

CS

FIGURE 62.2 Mind-gut axis. (A) Active Crohn’s patients (CA) demonstrated significant increases in gastrointestinal activity before emotional stimuli were introduced and (B) self-reported greater feelings of negative emotions after viewing emotion-laden film clips compared to healthy participants (HP) and silent Crohn’s patients (CS) suggesting that abnormal GI activity pre-disposed CA patients to a greater emotional experience (* represents a significant difference between groups (p<0.05)). Panels C and D: Correlation of EGG peak amplitude and intensity of self-reported emotional experience in CA(C) and CS(D) patients. An arousal rating of 1 corresponds to “I did not feel any emotion at all” and 7 corresponds to “I felt an extremely intense emotion”. CA patients show significant correlation between their EGG activity and subjective ratings of emotional experience; whereas, CS patients show no significant correlation (CA patients: r  0.61; CS patients: r  20.28; adopted with permission [7]).

a greater emotional experience. Furthermore, there was a correlation between EGG and the intensity of self-reported emotional experience (i.e., arousal) in CA patients but not in CS patients or healthy participants (an arousal rating of 1 corresponded to “I did not feel any emotion at all” and 7 corresponded to “I felt on extremely intense emotion, Fig. 62.2C and D). This result was attributed to greater inflammation in CA patients causing hyperalgesia resulting in CA patients self-reporting a more intense emotional experience while watching emotionally charged film clips [7]. In summary, actively experiencing gastrointestinal abnormalities increased negative emotional states suggesting that physiology can influence the mind.

CARDIOVASCULAR AROUSAL (HR AND BP) Emotion Imagine that you witness a young teen being mugged outside your favorite restaurant at night; or your favorite team has just won the championship. In each

circumstance, you are left with an emotional imprint that can influence your future decisions and life experiences, i.e., being more alert at night or feeling more joyful throughout the day (cf., the Somatic Marker Hypothesis [5]). People vary in their responses to and ability to cope with emotional events, and individuals have different emotional thresholds which depend on both genetic and environmental factors. However, recent work has demonstrated that despite these contributing factors, we can still volitionally alter our physiology through meditation [8,9]. In these blinded, randomized controlled trials, healthy participants were selected and grouped into two groups, high and low risk for hypertension based on their blood pressure and familial history. Healthy participants were randomly assigned to either the experimental or control groups. Participants in the experimental group were taught transcendental meditation (TM) and practiced this technique twice a day, for 20 minutes each session for three months. Blood pressure, psychological distress and coping abilities were assessed pre- and post-TM. A metaanalysis of these studies indicates that TM significantly decreased low and high risk participants’ systolic and diastolic blood pressures (SBP and DBP) (Fig. 62.3A) [8]. The

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62. MInd–Body InTERACTIons

FIGURE 62.3 Transcendental meditation (TM) induced physiological changes. (A) Meta-analysis indicates that TM significantly decreased low and high risk participants’ systolic and diastolic blood pressures (SBP and DBP) [8]. (B) Psychological distress and coping abilities were significantly improved compared to control TM groups in both low and high risk groups (adopted with permission [9]).

decreases were of sufficient magnitude that the authors expected significant reductions in the risk for atherosclerotic cardiovascular disease. In addition, psychological distress and coping abilities were significantly improved compared to control TM groups in both low and high risk groups (Fig. 62.3B) [9]. Taken together, these results indicate that TM is an effective means to alter human physiology and psyche.

white participants’ autonomic responses during imagined emotional situations. The investigators showed that just imagining emotionally charged events alters autonomic responses. Also, blacks demonstrated more positive and less negative emotional facial expressions, as measured by facial EMG activity, and showed larger increase in blood pressure in response to the stimuli. These two results indicate that (1) the mind influences physiology and (2) expressivity of emotion is driven by an external factor, ethnicity.

Imagery-Induced Emotion Studying emotions in the context of cultural and racial experiences has helped to identify environmental influences contributing to individual differences in emotional experience and expression across different cultures and races. For example, cultural values, including the degree to which a culture values emotional expression, can significantly shape how its members express themselves [3]. In these studies, measuring cardiovascular arousal is the most popular psychophysiological technique employed simply because it yields robust and reliable differences between ethnic groups. For example, it has long been speculated that African-Americans are more emotionally expressive than European-Americans and Asian-Americans because African-American culture places the greatest value on emotional expression, followed by European-American and lastly Asian-American cultures [3]. The following study employs imagery to induce emotion and illustrates the use of ethnic studies in mind–body research. Imagery is a potent method of inducing a particular state of mind associated with a specific physiological state. It has been used in emotion studies to induce a target emotion, e.g., by asking participants to “imagine actively participating in a particular emotion-laden scene.” Using such a paradigm, David Rollock’s group [3] recorded black and

CONCLUSIONS The studies above indicate that mind–body interactions are reciprocal; physiology (e.g., brain damage and Crohn’s disease) can influence an individual’s mental and emotional state and psychological factors (e.g., imagining an event and societal influences) contribute to physiological processes (e.g., sweating, cardiovascular and gastrointestinal arousal). Mind–body interactions are being utilized to reduce psychological and physiological distress and improve human health [10]. By monitoring the autonomic nervous system through SCRs, EGGs and cardiovascular arousal, it is possible to study the mechanisms underlying physiological changes in response to stimuli. The advent of novel technologies targeting the ANS in the years to come will usher in a new age of mind–body research.

References [1] Dawson ME, Schell AM, Filion DL. The electrodermal system. In: Cacioppo JT, Tassinary LG, Berntson GG, editors. Handbook of psychophysiology. Cambridge, MA: Cambridge University Press; 2000. p. 200–23. [2] Vianna EPM, Tranel D. Gastric myoelectrical activity as an index of emotional arousal. Int J Psychophysiol 2006;61(1):70–6.

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ConClusIons

[3] Vrana S, Rollock D. The role of ethnicity, gender, emotional content, and contextual differences in physiological, expressive, and self-reported emotional responses to imagery. Cog Emot 2002; 16(1):165–92. [4] Bechara A, Damasio H, Tranel D, Damasio A. Deciding Advantageously Before Knowing the Advantageous Strategy. Science 1997;275:1293–5. [5] Damasio AR. The Somatic Marker Hypothesis and the Possible Functions of the Prefrontal Cortex [and Discussion]. Philos Trans R Soc Lond B Biol Sci 1996;351(1346):1413–20. [6] Mayer EA, Naliboff BD, Chang L, Coutinho SV. Stress and the gastrointestinal tract. Am J Physiol Gastrointest Liver Physiol 2001;280:519–24.

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[7] Vianna EPM, Weinstock J, Elliot D, Summers R, Tranel D. Increased feelings with increased body signals. Soc Cogn Affect Neurosci 2006;1(1):37–48. [8] Anderson JW, Liu C, Kryscio J. Blood pressure response to transcendental meditation: a meta-analysis. Am J Hypertens 2008;21(3):310–6. [9] Nidich SI, Rainforth MV, et al. A randomized controlled trial on effects of the transcendental meditation program on blood pressure, psychological distress, and coping in young adults. Am J Hypertens 2009;22(12):1326–31. [10] Carter C. Healthcare performance and the effects of the binaural beats on human blood pressure and heart rate. J Hosp Mark Public Relations 2008;18(2):213–9.

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63 Alpha-Synuclein and Neurodegeneration Kiren Ubhi, Leslie Crews, Eliezer Masliah Alpha-synuclein (α-syn) entered the neurodegeneration field in the mid-1990s when it was discovered to be a constituent of the amyloid plaques characteristic of Alzheimer’s disease (AD). Since its initial description as the “non-amyloid” component of plaques, α-syn has been identified as a key neuropathological component in Parkinson’s disease (PD) and in a wide range of other neurodegenerative disorders with autonomic features, collectively termed the “α-synucleinopathies”. This chapter will discuss the role of α-syn in neurodegenerative disorders such as AD and PD. Animal models of α-synucleinopathy will be discussed, as will emerging strategies aimed at the therapeutic intervention in disorders characterized by the abnormal accumulation of α-syn.

ALPHA-SYN IN DISEASE Alzheimer’s Disease Alzheimer’s disease (AD) is among the most common neurodegenerative disorders in Europe and the US and is the leading cause of dementia in the aging population [1]. Alpha-syn was originally identified in AD plaques as the precursor protein of the non-Aβ component (NAC) and was thus called non-amyloid component of plaques (NACP) [2] (Fig. 63.1).

tremor, postural instability and autonomic features; at later stages of the disease patients develop cognitive impairment and depression [3]. Neuropathologically, PD is characterized by the loss of dopaminergic neurons in the substantia nigra, which accounts for the motor symptom in PD patients, and the presence of neuronal α-syn-rich aggregates called Lewy bodies [4] (Fig. 63.1). The majority of PD cases are sporadic in nature; however, diseasecausing mutations in the α-syn gene have been reported in familial cases of PD providing direct evidence for the fundamental role of α-syn in the pathogenesis of PD.

Multiple System Atrophy Multiple system atrophy (MSA) is a sporadic neurodegenerative disorder that typically affects patient 50 years of age or above [5]. In contrast to the neuronal aggregation of α-syn found in PD and LBD, MSA is characterized by the oligodendrocytic accumulation of α-syn, however this oligodendrocytic accretion of α-syn in MSA is still accompanied by significant neural loss in the striatum, substantia nigra, pons, inferior olive, cerebellum and cortex. Although glial cytoplasmic inclusions are the primary neuropathological hallmark of MSA, neuronal cytoplasmic and nuclear inclusions of α-syn have also been reported (Fig. 63.1).

TOXIC SPECIES OF ALPHA-SYN

Parkinson’s Disease Parkinson’s disease (PD) is a progressive neurodegenerative disorder characterized by bradykinesia, resting

Many recent studies investigating the toxicity of α-syn support the view that the oligomers of α-syn, rather than

FIGURE 63.1 Alpha-synuclein aggregation in neurodegenerative diseases. Alpha synuclein aggregation is a feature in a number of disorders. In Alzheimer’s disease, α-syn is a component of the amyloid plaques, whilst in Parkinson’s disease and dementia with Lewy bodies (LB) it forms neuronal aggregates. In contract to the neuronal aggregation in Parkinson’s disease and dementia with Lewy bodies, α-syn aggregation in multiple system atrophy (MSA) is predominantly oligodendrocytic in structures called glial cellular inclusions (GCIs).

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FIGURE 63.2 Toxic species of alpha-synuclein and synaptopathology. Emerging research is beginning to increasingly support the hypothesis that, rather than fibrillar α-syn, oligomeric α-syn may be the toxic species. It hypothesized that these toxic α-syn oligomers lead to synaptic dysfunction and consequent neurodegeneration.

the fibrils, may be the toxic species of α-syn [6]. While oligomers might accumulate at synapses, fibrils are present in LBs, which may reflect a compensatory mechanism by the cell to sequester the more toxic α-syn oligomeric species (Fig. 63.2). Modifications such as phosphorylation and oxidative/nitrative modifications have been shown to increase toxicity of α-syn [7], although the precise role and importance of each modification in this toxicity are still unclear. Other posttranslational modifications of α-syn include C-terminal cleavage by calpain [8]. In addition to the post-translational modification of the α-syn present in the neurons themselves, recent studies with tissue grafted into human PD patients has highlighted the possibility that α-syn may travel from one cell to another [9].

TRANSGENIC ANIMAL MODELS OF α-SYNUCLEINOPATHY Overexpression of wild-type α-syn under the regulatory control of the neuronal PDGF-β promoter has been shown to result in motor deficits, dopaminergic loss and formation of inclusion bodies, whilst under the mThy-1 promoter, expression of α-syn results in extensive α-syn accumulation throughout the CNS including, in some cases, in the SN or motor neurons [10] (Fig. 63.3). In addition to models mimicking neuronal α-syn accumulation as observed in PD, many groups have developed models to recapitulate the progressive α-syn accumulation in oligodendrocytes in MSA [11]. These models have reported accumulation of α-syn in oligodendrocytes with demyelination and degeneration, especially in the spinal cord and also have been reported to mimic the autonomic features characteristic of MSA [12]. However since the effects of α-syn accumulation in neurodegeneration in other cortical and subcortical brain regions deserve further consideration we generated new lines of tg mice overexpressing hα-syn under the control of the myelin basic protein (MBP) promoter [13]. By 6 months of age, these mice developed abundant hα-syn immunoreactive inclusions in

FIGURE 63.3 Transgenic mouse models of alpha-synucleinopathies. A number of transgenic mouse models have been developed to recapitulate the behavioral and neuropathological features of diseases characterized by the accumulation of α-syn. Many of our murine models express human α-syn under the control of neuronal promoters such as PDGF or mThy1, resulting in α-syn expression reminiscent of PD or DLB. In contrast, our mouse model of MSA expresses human α-syn under the control of the oligodendrocyte-specific MBP (myelin basic protein) promoter leading to the accumulation of α-syn in oligodendrocytic inclusions reminiscent of the glial cellular inclusions found in human cases of MSA.

oligodendrocytes in the neocortex, basal ganglia, cerebellum and brainstem, accompanied by myelin and neuronal damage and motor deficits, supporting a more general role of α-syn accumulation in the pathogenesis of MSA [13].

THERAPEUTIC APPROACHES TO α-SYN TOXICITY Since accumulation of α-syn oligomers is viewed as being key to the neurodegenerative process, methods aimed at reducing α-syn oligomers, either by reducing α-syn synthesis or aggregation or by increasing the clearance of α-syn, have been proposed as viable therapeutic approaches. An immense amount of work aimed at ameliorating α-syn toxicity has focused on modulating the phosphorylative and oxidative/nitrative modifications of α-syn. Alternative experimental approaches have been focused on the mechanisms involved in the degradation and clearance of α-syn. Neurosin, a trypsin-like serine protease, has been reported to degrade extracellular α-syn in vitro [14] and to reduce α-syn levels in α-syn tg mice (B. Spencer, unpublished observations). Recent work in our lab has shown that active immunization with recombinant α-syn ameliorates α-syn related synaptic pathology in a tg mouse model of PD [15] highlighting the effectiveness of antibody-based approaches in α-synucleinopathies.

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CONCLUSION The last few decades have seen an exponential growth in our knowledge and understanding of the importance of α-syn in the mechanisms leading to neurodegeneration and large strides have been made towards therapies aimed at ameliorating the toxic effects of abnormal accumulations of α-syn. The future holds exciting promise for the fruition of these treatments and the next 20 years may see many of these therapies transition from the bench to the bedside.

References [1] Maslow K. Alzheimer's disease facts and figures. Alzheimers Dement 2010;6:158–94. [2] Iwai A, Masliah E, Yoshimoto M, Ge N, Flanagan L, de Silva HA, et al. The precursor protein of non-A beta component of Alzheimer's disease amyloid is a presynaptic protein of the central nervous system. Neuron 1995;14:467–75. [3] Auluck PK, Caraveo G, Lindquist S. Alpha-Synuclein: membrane interactions and toxicity in Parkinson's disease. Annu Rev Cell Dev Biol 2010;26:211–33. [4] Spillantini MG, Schmidt ML, Lee VM, Trojanowski JQ, Jakes R, Goedert M. Alpha-synuclein in Lewy bodies. Nature 1997;388:839–40. [5] Wakabayashi K, Takahashi H. Cellular pathology in multiple system atrophy. Neuropathology 2006;26:338–45. [6] Cookson MR, van der Brug M. Cell systems and the toxic mechanism(s) of alpha-synuclein. Exp Neurol 2008;209:5–11.

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[7] Norris EH, Giasson BI, Ischiropoulos H, Lee VM. Effects of oxidative and nitrative challenges on alpha-synuclein fibrillogenesis involve distinct mechanisms of protein modifications. J Biol Chem 2003;278:27230–27240. [8] Mishizen-Eberz AJ, Guttmann RP, Giasson BI, Day 3rd GA, Hodara R, Ischiropoulos H, et al. Distinct cleavage patterns of normal and pathologic forms of alpha-synuclein by calpain I in vitro. J Neurochem 2003;86:836–47. [9] Kordower JH, Chu Y, Hauser RA, Freeman TB, Olanow CW. Lewy body-like pathology in long-term embryonic nigral transplants in Parkinson's disease. Nat Med 2008;14:504–6. [10] Rockenstein E, Mallory M, Hashimoto M, Song D, Shults CW, Lang I, et al. Differential neuropathological alterations in transgenic mice expressing alpha-synuclein from the platelet-derived growth factor and Thy-1 promoters. J Neurosci Res 2002;68:568–78. [11] Stefanova N, Bucke P, Duerr S, Wenning GK. Multiple system atrophy: an update. Lancet Neurol 2009;8:1172–8. [12] Stemberger S, Poewe W, Wenning GK, Stefanova N. Targeted overexpression of human alpha-synuclein in oligodendroglia induces lesions linked to MSA-like progressive autonomic failure. Exp Neurol 2010;224:459–64. [13] Shults CW, Rockenstein E, Crews L, Adame A, Mante M, Larrea G, et al. Neurological and neurodegenerative alterations in a transgenic mouse model expressing human alpha-synuclein under oligodendrocyte promoter: implications for multiple system atrophy. J Neurosci 2005;25:10689–10699. [14] Tatebe H, Watanabe Y, Kasai T, Mizuno T, Nakagawa M, Tanaka M, et al. Extracellular neurosin degrades alpha-synuclein in cultured cells. Neurosci Res 2010;67:341–6. [15] Masliah E, Rockenstein E, Adame A, Alford M, Crews L, Hashimoto M, et al. Effects of alpha-synuclein immunization in a mouse model of Parkinson's disease. Neuron 2005;46:857–68.

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64 Insulin Resistance and the Autonomic Nervous System Megan S. Johnson, Vincent G. DeMarco, Adam Whaley-Connell, James R. Sowers INTRODUCTION Obesity and insulin resistance (IR) are increasing in prevalence worldwide across gender and ethnicity, especially in children. Medical complications related to obesity and the development of IR include the progression to diabetes, hypertension, a high risk for cardiovascular (CVD) and chronic kidney disease (CKD), and recently recognized alterations in cognition, dementia and Alzheimer’s disease. In this context, the association between derangements in autonomic nervous system (ANS) function and complications related to IR such as CVD and CKD emphasizes the importance of the ANS as a contributor to cardiovascular target organ injury as well as a potential target for therapeutic intervention. In particular, preclinical, clinical, and epidemiological evidence supports an association between heightened sympathetic nerve activity (SNA) and metabolic dysfunctions such as impaired glucose metabolism, IR, and hyperinsulinemia. However, the exact nature of this relationship remains enigmatic, and whether activation of the SNS is a cause or consequence of these metabolic abnormalities is the subject of many recent and current studies.

parasympathetic dampening and are highly correlated with end organ damage and negative outcomes. IR and autonomic dysfunction are both features of the metabolic syndrome, and evidence for an association between the two comes from numerous studies. In humans, both coexist in many conditions such as obesity, hypertension, and diabetes, and both are related to fat distribution patterns, being correlated with central rather than peripheral fat. Compared to those without the metabolic syndrome, those with have higher heart rates, increased sympathetic nerve norepinephrine (NE) secretion, increased number of sympathetic bursts to skeletal muscle, and baroreflex impairment, all suggesting a degree of autonomic dysfunction. Furthermore, correlations have been made between SNA (measured by muscle SNA (MSNA)) and percent body fat, fasting insulin levels, and HOMA indices. Additionally, clinical strategies that improve IR, such as lifestyle modification and pharmaceutical treatment, tend to also have beneficial effects on SNA. For example, weight loss in obese normotensive subjects reduces MSNA, lowers plasma NE spillover, and improves whole body glucose disposal [2]. Collectively, these close associations suggest coinciding mechanisms between IR and the degree of sympathetic activation.

SYMPATHETIC ACTIVITY AND INSULIN RESISTANCE IR occurs when peripheral tissues, particularly adipose, liver and skeletal muscle, are resistant to the metabolic actions of insulin, resulting in reduced insulin-induced intracellular glucose transport [1]. The feedback mechanisms that are triggered in the IR state include an increase in pancreatic insulin secretion, and thus hyperinsulinemia. Impaired insulin metabolic signaling is associated with cardiovascular and renal abnormalities such as endothelial dysfunction, cardiac diastolic dysfunction and proteinuria. Hypertension-related autonomic dysfunction includes alterations in baroreflex sensitivity and heart rate variability. The net disruptions in sympatho-vagal balance are characterized by sympathetic predominance and

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00064-0

INSULIN RESISTANCE INDUCES SYMPATHETIC ACTIVATION Insulin enhances central nervous system activity and plays an important role in dietary intake-induced sympatho-excitation which, ironically, likely subserves weight maintenance by stimulating thermogenesis at the expense of hypertension. In human subjects, acute physiological insulin can specifically stimulate SNA. For example, experimental hyperinsulinemia during a euglycemic glucose clamp increases MSNA and sympatho-vagal tone. Insulin’s central pathways are well characterized. Insulin passes through the blood–brain barrier and acts on receptors in sites involved in central pathways regulating autonomic activity, resulting in increased sympathetic drive

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and reduced baroreflex sensitivity. In the hypothalamus sites of insulin action include the paraventricular, supraoptic, and arcuate nucleii and the dorsomedial and ventromedial hypothalamus. Brainstem sites include the solitary tract nucleus and nucleus ambiguous [3]. Thus, insulin is integral in neuroregulatory pathways, and the insulin resistant state and resulting hyperinsulinemia contribute to chronic sympathetic activation, hypertension, and ultimately cardiovascular target organ injury. The notion that hyperinsulinemia contributes to sympathetic nerve activation is supported by preclinical studies in which intracerebroventricular (ICV) injection of insulin increases MSNA, and the mechanism appears to be mediated by hypothalamic PI3K and MAPK [4]. Such studies support a potential role for insulin as a direct mediator of sympathetic overdrive in the metabolic syndrome. However, while hyperinsulinemia contributes to obesity-induced sympathetic activation it does not explain the heterogeneity related to hypertension in the metabolic syndrome. For example, in a study of young and aged adult male human subjects MSNA was more closely related to plasma leptin, which also stimulates sympathetic nerve activity (SNA), suggesting factors other than insulin can be responsible for activation of the SNS in metabolic syndrome [5].

HEIGHTENED SNA INDUCES INSULIN RESISTANCE The contribution of autonomic dysfunction, specifically heightened SNA, to hypertension is well-defined and includes influences on the vasculature (arteriole remodeling and an increase in vascular resistance), kidney (increase in renal tubular sodium reabsorption and fluid retention) and heart (increase in cardiac output). A role for the SNS in IR is less understood, although the idea that increased SNA precedes and drives the development of IR is suggested by many observations. In one of the earliest studies demonstrating this phenomenon, acute reflex sympathetic activation in the human forearm (at physiological levels similar to that experienced post-prandially) impaired skeletal muscle glucose uptake in the same location [6]. Subsequent studies have demonstrated that peripheral insulin sensitivity can be modulated by central manipulations (i.e., ICV injection of various substances such as leptin, glucocorticoids, anti-psychotic drugs, specific neuropeptides). Several important clinical studies have demonstrated the emergence of heightened SNA prior to the development of IR. In one 10-year longitudinal study, investigators showed that sympathetic nerve hyperactivity precedes and predicts the appearance of hyperinsulinemia, impaired glucose metabolism, weight gain, and elevated blood pressure [7]. More recently, it was observed that increased sympathetic reactivity to cold pressor test was an independent predictor of subsequent IR [8]. Finally,

the favorable impact of sympatho-inhibitory agents on metabolic parameters gives further credence to the primacy of SNS activation in the pathogenesis of metabolic syndrome. Several mechanisms by which heightened sympathetic drive may cause IR are plausible (Fig. 64.1). First, the SNS could act via a hemodynamic mechanism in which reflex SNA results in vasoconstriction and thus reduced muscle perfusion and reduced glucose delivery and uptake. Sympathetic vasoconstriction is likely occurring at a time in which, due to the insensitivity of peripheral tissues to insulin, insulin’s vasodilatory actions are impaired, and elevated blood pressure results. However, multiple forearm perfusion studies testing the effects of reflex sympathetic activity on glucose uptake report conflicting results with regard to whether blood flow is actually decreased. A more likely mechanism of increased SNA is that of a direct metabolic NE effect at the cellular level, possibly involving beta-adrenergic receptors. The link between beta-adrenergic receptors and insulin signaling is still being established, but a recent report demonstrated that the G-protein coupled receptor kinase (GRK2) that desensitizes (via phosphorylation) beta-adrenergic receptors can also directly inhibit IRS1 signaling [9]. The exact mechanism of beta-adrenergic receptor-induced IR is likely dependent upon the particular tissue and receptor subtype. Interestingly, polymorphisms of the beta-2 adrenergic receptor have been linked to insulin secretion (Arg16Gly) and IR (Gly16) in various human populations. Increased SNA to specific organs can have disparate consequences on IR. In adipose tissue, heightened SNA can cause lipolysis leading to increased circulation of fatty acids, which can in turn directly inhibit cellular glucose transport. In the kidney, high renal sympathetic tone and excess NE release from renal nerve terminals causes increased renin release from juxtaglomerular cells of the kidney, thereby stimulating the renin-angiotensin-aldosterone system (RAAS) with inappropriate elevations in angiotensin (Ang) II and aldosterone (Aldo) that impact blood volume and sodium retention. Indeed, RAAS activation is likely a major mechanism by which sympathetic activation contributes to the development of IR (Fig. 64.2). Enhanced Ang II signaling via the Ang II type 1 receptor (AT1R) and Aldo non-genomic actions on the mineralcorticoid receptor promote NADPH oxidase subunit upregulation and translocation and, subsequently, production of reactive oxygen species (ROS) that can contribute to impaired insulin metabolic signaling. Furthermore, both Ang II and Aldo effects on sympathetic drive have been well-established. Excess RAAS activation can further potentiate sympatho-activation by mechanisms both peripheral (increased NE release from sympathetic terminals) and central (by acting on hypothalamic and brainstem sites to increase firing in regions important in originating sympathetic drive). Thus, RAAS and sympathetic activation function in a positive-feedback loop

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FIGURE 64.1 Systemic effects of heightened SNA on insulin sensitivity and hypertension. High salt intake, obesity, inactivity, and other environmental factors interact to activate the SNS, with subsequent inflammation and oxidative stress that drive maladaptive tissue responses. ACE, angiotensin-converting enzyme; ACTH, adrenocorticotropic hormone; Aldo, aldosterone; Ang I, angiotensin I; Ang II, angiotensin II; ASCVD, atherosclerotic cardiovascular disease; AT1, angiotensin type 1 receptor; AT2, angiotensin type 2 receptor; CAD, coronary artery disease; CHF, congestive heart failure; CRH, corticotropin-releasing hormone; LVH, left ventricular hypertrophy; MR, mineralocorticoid receptor; SNS, sympathetic nervous system. (Reprinted with permission from Sowers et al., 2009. Narrative review: the emerging clinical implications of the role of aldosterone in the metabolic syndrome and resistant hypertension. Ann Intern Med 150(11): 776–83.)

that contributes to the development of hypertension, IR, and organ damage in metabolic syndrome (Fig. 64.2). Clearly, heightened sympathetic drive can influence insulin sensitivity; however, it is not a requirement for the development of IR in all scenarios. For example, Pima Native Americans display higher rates of obesity and hyperinsulinemia compared to Caucasians but have decreased MSNA, which may underlie the low prevalence of hypertension in this population [10]. Certainly other mechanisms can contribute to IR in peripheral tissues.

AN INTEGRATED VIEW Although evidence exists for primary roles by both IR and sympathetic nerve overactivation, neither has been definitively proven as the inciting event in the metabolic syndrome cascade. Indeed it is likely that in some instances a primary defect in one can be exacerbated by the other, and primacy can likely vary among individuals, ethnic groups, medical conditions. Furthermore, it appears

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Sympathetic Activation

Over-stimulation of central cardio-regulatory pathways

HYPERTENSION

Vasoconstriction Beta-adrenergic receptor stimulation ROS generation

RAAS

Insulin Resistance

FIGURE 64.2 The interaction of SNA and IR in the development of hypertension. IR impacts SNA by stimulating cardio-regulatory sites in the brain. Heightened SNA contributes to the development of IR through various mechanisms, with a large role played by the RAAS, which has reciprocal stimulatory effects on SNA. In addition to their compounding effects, IR and heightened SNA also both independently contribute to hypertension.

IR and heightened sympatho-excitation are linked in a positive-feedback relationship in which hyperinsulinemia can act centrally to increase sympathetic output which in turn increases IR and further promotes hyperinsulinemia (Fig. 64.2). In a forearm perfusion study [11], reflex NE release was enhanced by intravenous insulin infusion at physiological, post-prandial levels. The resulting increased SNA had an antagonistic effect on forearm skeletal muscle glucose uptake that was not hemodynamically mediated, as blood flow to the forearm was unchanged. Importantly, both IR and SNS over-activation may also contribute independently to hypertension and end-organ damage (specifically hypertrophy of cardiac and connective tissue, arterial stiffening, and renal damage) via separate pathways involving mitogenic properties (insulin) and reactive oxygen species (ROS) generation in the SNS (Fig. 64.2). Nevertheless, their combinatorial role is multiplicative. Regardless of which precedes the other, IR and enhanced SNA interact to adversely affect insulin metabolic signaling and contribute to metabolic syndrome, hypertension, CVD and CKD.

THERAPEUTIC CONSIDERATIONS Thus, heightened SNA may be a modifiable therapeutic target in the treatment of IR and metabolic syndrome. In fact, many current treatments for metabolic syndrome concomitantly correct aspects of both heightened sympathetic activation and IR. Lifestyle modifications, including physical training programs and energy-restricted diets, can improve insulin sensitivity and metabolic function and reduce sympathetic nerve traffic. Likewise, RAAS-directed compounds (direct renin inhibition, angiotensin-converting enzyme inhibition, and angiotensin II antagonism) not only prevent the Ang II-induced sympatho-excitation but can also improve insulin sensitivity and other indices of

metabolic syndrome [12]. Finally, improvements in plasma triglyceride levels, glucose metabolism, and insulin sensitivity have been observed with direct sympatho-inhibition. The benefits of pharmacological autonomic blockade are likely multifaceted and include pressor effects, normalization of lipid profiles, direct reduction of end organ damage, stabilization of metabolic parameters, and the favorable modification of insulin sensitivity. Early intervention to reduce sympathetic activity should be an effective therapeutic strategy for IR and metabolic syndrome, and, in light of the relationships discussed herein, efforts should be directed toward sympatho-inhibitory compounds with no negative effect on body weight and glucose metabolism.

References [1] Whaley-Connell A, Sowers JR. Hypertension and insulin resistance. Hypertension 2009;54:462–4. [2] Sowers JR, Whitfield LA, Catania RA, Stern N, Tuck ML, Dornfield L, et al. Role of the sympathetic nervous system in blood pressure maintenance in obesity. Hypertension 1982;54:1181–6. [3] Brooks VL, Dampney RA, Heesch CM. Pregnancy and the endocrine regulation of the baroreceptor reflex. Am J Physiol Regul Integr Comp Physiol 2010;299:R439–51. [4] Morgan DA, Rahmouni K. Differential effects of insulin on sympathetic nerve activity in agouti obese mice. J Hypertens 2001;28:1913–19. [5] Monroe MB, Van Pelt RE, Schiller BC, Seals DR, Jones PP. Relation of leptin and insulin to adiposity-associated elevations in sympathetic activity with age in humans. Int J Obes Relat Metab Disord 2000;24:1183–7. [6] Jamerson KA, Julius S, Gudbrandsson T, Andersson O, Brant DO. Reflex sympathetic activation induces acute insulin resistance in the human forearm. Hypertension 1993;21:618–23. [7] Masuo K, Mikami H, Ogihara T, Tuck ML. Sympathetic nerve hyperactivity precedes hyperinsulinemia and blood pressure elevation in a young, nonobese Japanese population. Am J Hypertension 1997;10:77–83. [8] Flaa A, Aksnes TA, Kjeldsen SE, Eide I, Rostrup M. Increased sympathetic reactivity may predict insulin resistance: an 18 year follow-up study. Metabolism 2008;57:1422–7.

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[9] Cipolletta E, Campanile A, Santulli G, Sanzari E, Leosco D, Campiglia P, et al. The G protein coupled receptor kinase 2 plays an essential role in beta-adrenergic receptor-induced insulin resistance. Cardiovasc Res 2009;84:407–15. [10] Weyer C, Pratley RE, Snitker S, Spraul M, Ravussin E, Tataranni PA. Ethnic differences in insulinemia and sympathetic tone as links between obesity and blood pressure. Hypertension 2000;36:531–7.

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[11] Lembo G, Capaldo B, Rendina V, Iaccarino G, Napoli R, Guida R, et al. Acute noradrenergic activation induces insulin resistance in human skeletal muscle. J Am Physiol 1994;266:E242–7. [12] Lastra G, Habibi J, Whaley-Connell A, Manrique C, Hayden MR, Rehmer J, et al. Direct renin inhibition improves systemic insulin resistance and skeletal muscle glucose transport in a transgenic rodent model of tissue renin overexpression. Endocrinology 2009;150:2561–8.

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65 Salt Sensitivity of Blood Pressure Cheryl L. Laffer, Fernando Elijovich Salt-sensitivity of blood pressure (SSBP) is a phenotype of humans and experimental animals in which blood pressure (BP) changes in parallel with a positive or negative salt balance. Guyton proposed that interactions between natriuretic and antinatriuretic systems maintain salt balance in the whole animal. In contrast, the isolated kidney depends on increased perfusion pressure for restoration of salt balance after a salt load (“pressure-natriuresis”). Hence, SSBP must be due to a defect in salt-stimulation of natriuretic systems or salt-inhibition of antinatriuretic ones, making natriuresis in the whole animal dependent on increased arterial pressure (Fig. 65.1). Transplanting the kidney of a salt-sensitive (SS) to a salt-resistant (SR) rodent confers SSBP to the latter and vice versa, indicating that the defect must be renal in origin. SSBP is an abnormal phenotype because it is uncommon in normotensive subjects (25–30%), associates with human and experimental hypertension, and most importantly, confers risk for cardiovascular morbidity and mortality, independent of BP [1].

reproduced in diverse populations), knockout mice strains implicating genes in exaggeration or attenuation of the BP response to salt, and congenic rat strains, in which chromosomal segments different from those that confer

GENETICS Inbred substrains of rodents with pure SS and SR phenotypes (e.g., Dahl-SS and SR rats) are unequivocal proof for genetic determination of SSBP. In contrast, BP responses to salt in humans are not dichotomous but continuous, suggesting polygenic etiology and environmental interactions; heritability of SSBP is 74% in Blacks and 50% in Chinese, both higher than that for hypertension. Therefore, arbitrary cutoffs in BP responses to salt loading or deprivation have been used to classify subjects into SS and SR. Experimental protocols include long-term dietary salt manipulation and short-term intravenous salt loading followed by diuretics. Inheritance of the SSBP phenotype is supported by its reproducibility over time or when assessed with different protocols, and by its concordance in twin and non-twin siblings. Despite different cutoffs to classify subjects into SS and SR groups, certain clinical characteristics and biochemical markers cluster in SS subjects (Table 65.1). Some are shared with those of the metabolic syndrome, which confers a 3–4 fold increase in risk for SS hypertension [2]. Table 65.2 shows positive studies of gene polymorphisms associated with SSBP in humans (not all

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00065-2

FIGURE 65.1 Schematic representation of maintenance of salt-balance after a salt load. In the top panel, after the salt load (arrow), a whole animal exhibits stimulation of natriuretic systems (N) and inhibition of antinatriuretic systems (AN), leading to natriuresis (UNaV) and conservation of salt balance (Na balance) without a change in blood pressure (BP). In contrast, the bottom panel illustrates the situation in salt sensitive animals, in which either N or AN (or both) are unresponsive to the salt load (i.e., “clamped” at an inappropriate level). UNaV and Na balance are still preserved but this occurs at the expense of BP elevation.

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TABLE 65.1 Clinical and Biochemical Characteristics of SS Hypertensive Subjects DEMOGRAPHIC AND CLINICAL GROUPS WITH INCREASED PREVALENCE African-Americans

Elderly

Postmenopausal women

Obese

Diabetic subjects

Metabolic syndrome

Chronic renal failure

Non-modulators of renal blood flow

Post-pregnancy-related HTN

Subjects born with low birth weight

Subjects with nutritional K or Ca deficiencies

OVER-REPRESENTED CLINICAL FEATURES AND TARGET ORGAN DAMAGE Lack of nocturnal dip of blood pressure

Left ventricular diastolic dysfunction

Left ventricular hypertrophy

CHF and CVA rather than ischemic heart disease

Microalbuminuria

Hypertensive nephrosclerosis

OVER-REPRESENTED PHYSIOLOGICAL AND BIOCHEMICAL FEATURES Increased half-life for urine sodium excretion

Suppressed kallikrein-kinin system

Insulin resistance

Low plasma renin activity

Blunted renin and aldosterone responses to salt loading

Blunted norepinephrine responses to salt loading

Increased plasma endothelin

Overactive arginine vasopressin pressor system

SS, salt sensitive; HTN, hypertension; CHF, congestive heart failure; CVA, cerebrovascular accident.

normotensive and hypertensive phenotypes are specifically able to confer SSBP or SRBP to recipient strains. Most genes in these studies affect salt excretion or its regulatory systems.

GENE–ENVIRONMENT INTERACTIONS Notwithstanding its genetics, SSBP is also influenced by environmental factors. Its prevalence is higher in: (a) older subjects, perhaps linked to age-dependent decrease of endogenous ouabain-like Na pump inhibitors [3]; (b) postmenopausal women, much before hypertension prevalence increases, in whom estrogen deficiency increases ADMA and inhibits NO; (c) normotensive women after a hypertensive pregnancy [4]; (d) populations with low calcium and potassium intake; and (e) subjects with chronic renal disease. Fetal programming of SSBP occurs in children and adults with low birth weight, either as a direct effect or secondary to reduced kidney mass and GFR [5,6]. A link between incretin deficiency and SSBP in diabetes is suggested by the correction of impaired natriuresis and SSBP of obese db/db mice by the GLP-1 analog exendin, which inhibits angiotensin II-induced ERK stimulation [7]. Salt consumption itself leads to SSBP increasing conduit artery stiffness, via plasma volume expansion, increased endothelial sheer stress, opening of K channels, activation of cytoplasmic proline-reach tyrosine kinase-2, cytoplasmic Src and MAPKs, generation of endothelial TGFbeta1, and vascular fibrosis. These phenomena may be more prominent in certain subjects, e.g., excess BP responses to salt in SS normotensive blacks are not due to differences in plasma volume, cardiac output or Na balance (compared to SR) but to an increase in systemic vascular resistance.

Humans with a defined psychological profile and some animals respond to stress with non-autonomic hypoventilation, increasing PCO2 and decreasing plasma pH. This enhances renal Na/H exchange, leading to SSBP. Concomitant increase in digitalis-like Na pump inhibitors suggests volume expansion. Consistent with greater SSBP in blacks, end tidal volume CO2 (a good correlate of PCO2) is higher in elderly blacks compared to whites.

VASOREGULATORY AND NATRIURETIC SYSTEMS Many physiologic abnormalities have been described in different vasoactive and renal regulatory mechanisms of SS animals and humans. However, the ultimate cause of SSBP remains elusive because these regulatory systems interact, making it very difficult to pinpoint a causal pathway.

Renin-Angiotensin-Aldosterone System (RAAS) Salt loading inhibits and salt depletion stimulates the RAAS, preventing changes in BP during changes in salt intake. Hall’s group showed that SSBP (BP reduction during salt depletion and BP increase by salt loading) can be produced in dogs by chronic angiotensin-converting enzyme (ACE) inhibition or chronic angiotensin II infusion (i.e., “clamping” the RAAS at low or high levels of angiotensin II). Others made analogous observations in rats. Williams and Hollenberg showed that subjects who fail to increase plasma aldosterone in response to salt deprivation or angiotensin infusion and do not normally increase renal blood flow in response to salt (“nonmodulators”) generally exhibit SSBP. Finally, sensitivity to exogenous angiotensin II during salt-deprivation, which should

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TABLE 65.2 genetics of Salt Sensitivity of Blood Pressure Gene

Polymorphism

Mechanism

Source

ACE 1

Insertion (II) genotype

Unknown, interacts with Na-intake and obesity

Hiraga et al. Hypertension 27:569;1996 Poch et al. Hypertension 38:1204;2001 Zhang et al. Hypertens Res Clin Exp. 29:751;2006

AT1-R

A allele of A-rs4524238-G

Unknown

Gu et al J Hypertens. 28:1210;2010

CYP11B2

TT genotype of promoter T-344C W of CYP11B2 IC polymorphism

Blunts inhibition of aldosterone during salt loading Unknown

Iwai et al. Hypertension. 49:825;2007

A allele of promoter G209A

Blunts gene upregulation by GCR with transcription regulators NF1 or Sp1 Enzyme overexpression? Unknown

Alikhani-Koupaei et al. FASEB J. 21:3618;2007

A allele of G1065A

Increased renal receptor expression?

Caprioli et al. Can J Physiol Pharmacol. 86:505;2008

β2-AR

A allele of G46A

Blunted aldosterone and PRA responses to low salt (DASH) diet

Sun et al. Am J Clin Nutr. 92:444;2010

GRK4

L of R65L with V of A142V, and Activating variants, impair cyclic Sanada et al. Clin Chem. 52:352;2006 V of A486V AMP response to D1-R; 94.4% predictive of SS hypertension

RAAS

11βHSD2

A allele of C-rs5479-A GG genotype of G534A

Pamies-Andreu et al. J Human Hypertens. 17:187;2003

Gu et al J Hypertens. 28:1210;2010 Poch et al. Hypertension. 38:1204;2001

ET SYSTEM ETb-R SNS

NAD(P)H OXIDASE p22phox

T allele of C242T

Decreased renal NO excretion

Castejon et al. J Human Hypertens. 20:772;2006

Lower PRA and heart rate suggest excess Na reabsorption

Barlassina et al. Human Mol Genet. 16:1630;2007

TRANSPORTERS AND REGULATORY PROTEINS CLCNKA

Intronic G of A-rs1010069-G and Thr of Ala447Thr

GNB3

CC genotype of A-rs1129649-C Enhances Na/H exchanger activity

Kelly et al. Am J Hypertens. 22:985;2009

ADD1

Trp of Gly460Trp

Grant et al. Hypertension. 39:191;2002

G allele of G-rs17833172-A

NEDD4L

Increased Na reabsorption via effects on tubular actin cytoskeleton Increased Na reabsorption via effects on tubular actin cytoskeleton

Kelly et al. Am J Hypertens. 22:985;2009

GG genotype of G-rs4149601-A Impaired bindng to P-Y motifs with CC genotype of of ENaC, decreased channel C-rs2288774-T membrane removal for degradation; low PRA due to increased Na reabsorption

Dahlberg et al. PLoS ONE 2:e432;2007

CYP4A11

C allele of T8590C

Reduced catalytic activity, reduced 20-HETE synthesis and natriuresis

Laffer et al. Hypertension. 51:767;2008

CYP3A5

Expressor *1 of A6986G

Increases renal conversion of cortisol to 6βOH-cortisol with increased Na reabsorption

Zhang et al. J Human Hypertens. 24:345;2010 Bochud et al. J Hypertens. 24:923;2006

CYP450 ENZYMES

(Continued)

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TABLE 65.2 (Continued) Gene

Polymorphism

Mechanism

Source

Knockouts

Mechanism Implied

Source

D5-R(/)

↑Expression NKCC2, NaCl, and α/γ ENaC; not ↓↓ by Na

Wang et al. Hypertension. 55:1431;2010

ANP(/)

Impaired blunting of PRA by Na (via AT1-R and SNS)

Melo et al. Am J Physiol. 277:R624;1999

COX1(/)

Suppressed renal PGE2 and natriuretic response to Na

Ye et al. Am J Physiol. 290:F542;2006

BB2-R(/)

Impaired natriuretic response to high Na intake

Alfie et al. Biochem Bioph Res Co. 224:625;1996

eNOS(/)

↑NADPH, ↑ISO and ↑ROS; SS abrogated by antioxidants

Kopkan et al. Am J Physiol. 299:F656;2010

SOD1(/)

↑ISO in renal HTN; SS ↓↓ by antioxidants or SODtg

Carlstrom et al. Am J Physiol 297:R82;2009

NKCC1(/)

↓Vasoconstriction to low Na and ↓PRA suppression by Na

Kim et al. Am J Physiol. 295:F1230;2008

α2B-R(/)

↓SSBP via ↓SNS, ↓vasoconstriction and ↓Na reabsorption

Makaritsis et al. Hypertension. 33:14;1999

Congenics Donor

Recipient

Effect

Region

Source

TRANSFER OF SALT RESISTANCE FROM NORMOTENSIVE TO HYPERTENSIVE STRAIN BN

SHR

QTLs 2a and 16

↓SS of SHR

BN

SHR

D18Rat113/Rat99

↓SS of SHR

BN

DahlSS

Chromosome 13

↓SS of DahlSS

WKY

SHRSP

D2Mgh12/Rat157

↓SS of SHRSP

SBN

SBH

QTLs SS1a-1b

↓SS of SBH

Aneas et al. Physiol Genomics. 37:52;2009 Johnson et al. Hypertension. 54:639;2009 Cowley et al. Hypertension. 37:456;2001 Graham et al. Hypertension. 50:1134;2007 Yagil et al. Physiol Genomics. 12:85;2003

TRANSFER OF SALT SENSITIVITY FROM HYPERTENSIVE TO NORMOTENSIVE STRAIN SHR

WKY

D1Mit3/Rat57

↑SS of WKY

SHR

Lewis

QTL 20RT1 (MHC)

↑SS of Lewis

Iwai et al. Hypertension. 32:636;1998 Kunes et al. Hypertension. 24:645;1994

TRANSFER OF SALT RESISTANCE FROM HYPERTENSIVE TO HYPERTENSIVE STRAIN SHR

DahlSS

Chromosome 19

↓SS of DahlSS

Wendt et al. J Hypertens. 25:95;2007

Polymorphisms associated with salt-sensitivity (SS) of blood pressure (BP) in humans; mouse knockout models implicating regulatory systems in SS of BP, and congenic rat models demonstrating transfer of salt resistance or SS of BP by specific chromosome regions. RAAS, renin-angiotensin-aldosterone system; ACE 1, angiotensin converting enzyme type 1; AT1-R, type 1 angiotensin II receptor; CYP11B2, aldosterone synthase; 11βHSD2, 11-beta-hydroxysteroid-dehydrogenase type 2; ET, endothelin; ETb-R, endothelin receptor b; SNS, sympathetic nervous system; β2-AR, beta-2 adrenergic receptor; GRK4, G-protein-coupled receptor kinase type 4; CLCNKA, chloride channel type A; GNB3, G-protein beta polypeptide 3; ADD1, adducin; NEDD4L, neuronal precursor cell expressed developmentally down-regulated 4-like; D5-R, dopamine receptor type 5; ANP, atrial natriuretic peptide; COX1, cyclooxigenase 1; BB2-R, bradikinin B2 receptor; eNOS, endothelial nitric oxide synthase; SOD1, superoxide dismutase 1; NKCC1, isoform 1 of the Na,K,2Cl cotransporter, α2B-R, alpha-2b adrenergic receptor; BN, Brown Norway; SHR, spontaneously hypertensive rat; WKY, Wistar Kyoto; SHRSP, stroke prone SHR; SBN, Sabra normotensive rat; SBH, Sabra hypertensive rat; GCR, glucocorticoid receptor; PRA, plasma renin activity; D1-R, dopamine receptor type 1; NO, nitric oxide; ENaC, epithelial sodium channel; 20-HETE, 20-hydroxyeicosatetraenoic acid; NKCC2, isoform 2 of the Na,K,2Cl cotransporter; PGE2, prostaglandin E2; ISO, isoprostanes; ROS, reactive oxygen species; SODtg, mouse transgenic for SOD1.

decrease owing to increased endogenous angiotensin II, is maintained or paradoxically enhanced in SS normotensive and hypertensive subjects, another example of “clamping” of the RAAS at an inappropriate level.

Endothelin Endothelins (ET), potent vasoconstrictors and growth factors, also play a role in natriuresis via renal ETb receptors that inhibit Na-K-ATPase. ET-1 knockout mice (collecting duct-specific) and rats deficient in ETb receptor develop SS hypertension. In SS hypertensive subjects urinary endothelin is diminished, which is important because it normally exhibits circadian rhythm parallel to and a positive correlation with Na excretion during a salt

load. Also, urinary endothelin correlates negatively with blood pressure in normotensive and hypertensive subjects. Therefore, reduced basal and salt-stimulated urinary endothelin may contribute to impaired natriuresis in human SS hypertension. Whether pure ETa blockade (with ETb preservation) is useful in patients with SS hypertension remains to be tested.

Nitric Oxide (NO) and Oxidative Stress nNOS-derived NO is important for the regulation of renal blood flow and natriuretic responses to salt in rodents; its inhibition leads to SS hypertension. Salt diminishes the activity of NO synthase; when given jointly with oxidants or methylglyoxal (advanced

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ClInICAl SIgnIfICAnCE

glycation protein endproducts), it elicits SSBP in Sprague– Dawley rats. SOD1 knockout mice have SSBP in response to experimental hydronephrosis, whereas a transgenic strain with excess SOD1 expression does not. The eNOS knockout mouse develops SSBP that can be inhibited by tempol and apocynin. The antioxidant tempol abolishes SSBP in SHR and NADPH inhibitors do so in Dahl-SS rats (which have increased NADPH activity in the renal medulla). These observations suggest that an imbalance between renal NO and oxygen radical generation plays a role in SSBP. In humans with SS hypertension, a salt load paradoxically decreases excretion of NO metabolites. We have shown acute salt-induced increases in free isoprostanes in SS hypertensive subjects, suggesting that NO is diverted to the scavenging of salt-induced oxygen-free radicals. Alternatively, others have suggested that salt-stimulated production of an endogenous inhibitor of NO (asymmetric dimethylarginine) may play a role in SSBP.

Sympathetic Nervous System (SNS) DiBona’s group showed that cardiac atria (and ventricles) sense volume-distension, transmitting information to brain stem integrator centers via cardiopulmonary afferent nerves. Efferent renal sympathetic nerves mediate ensuing neurogenic vasodilation, increased renal blood flow and natriuresis. Disruption of this mechanism by experimental sinoaortic deafferentation or cardiac transplantation in humans leads to SS hypertension. These models differ from SS essential hypertension because impaired natriuresis leads to volume expansion. In contrast, in human SS hypertension salt balance is restored by pressure natriuresis, such that SS and SR subjects differ only in the level of BP required to achieve natriuresis, not in their salt balance. Salt-induced decreases in norepinephrine and concomitant increases in dopamine are blunted in SS hypertension, contributing to abnormal renal and systemic hemodynamics, volume homeostasis, and BP. Renal vasodepressor effects, mediated by transient receptor potential vanilloid type 4 channels located in sensory neurons and renal parenchyma are impaired in Dahl-SS, which have abnormal depression of protein content of this channel in response to salt, opposite to normal enhancement in Dahl-SR rats [8]. Dopamine D5 receptor knockout mice are SS; their impaired natriuresis is due to: (a) baseline over expression of Na transporters (NaK2Cl, NaCl, and α/δ chains of ENaC); (b) blunted inhibition of transporter expression during a salt load; (c) paradoxical over expression of NHE3 and NaPi2 in response to salt; and (d) increased oxidative stress. Finally, SS hypertension is observed in normal subjects with a phenotype characterized by low anxiety scores but increased self-deception (a defensiveness mechanism) and increased autonomic (heart rate and electrodermal) responses to a standardized mental stress [9].

Arachidonic Acid (AA) Metabolites Products of cyclooxygenation, epoxygenation, and ω-hydroxylation of AA play a major role in SSBP. COX-1 knockout mouse and animals given COX-1 and COX-2 blockers develop SSBP because of blunting of the normal increase in PGE2 produced by salt, which is required for removal of ENaC from the cell membrane for proteasome recycling. The COX-1 knockout exhibits impaired sleeprelated fall in BP, analogous to SS humans. Clinically, saltdependent hypertension occurs during administration of COX inhibitors but there is no evidence for PGE2 deficiency in SS essential hypertension. EETs, products of epoxygenation of AA, inhibit distal Na reabsorption by altering the gating properties of ENaC. Their participation in physiologic natriuresis is supported by increased synthesis during salt loading. Renal epoxygenase (CYP2c44) activity and urine EET excretion are diminished in Dahl-SS rats and CYP4a10 knockout mice. Pharmacologic inhibition of CYP2c44 diminishes synthesis of EETs leading to SSBP. Amiloride corrects the CYP4a10 knockout hypertension, supporting a role for ENaC. The major product of AA ω-hydroxylation, 20-HETE, has pro- (vasoconstriction) and anti-hypertensive (natriuresis) actions. Natriuresis is exerted via blockade of potassium channels, (impairing the action of the NaK2Cl cotransporter of the mTAL) and inhibition of Na-KATPase. Diminished 20-HETE content and CYP4A2 expression in the renal medulla of Dahl-SS rats relates to diminished chloride transport, blunted pressure-natriuresis and SSBP. Congenic Dahl-SS strains harboring the region of chromosome 5 containing the normal CYP4A genes exhibit attenuated SSBP. Stimulation of renal tubular CYP4a protein by fenofibrate in renal tubules of C57BL/6 mice (a strain with reduced 20-HETE) antagonizes angiotensin II– induced hypertension. Also, impairment of the transport effects of 20-HETE (despite normal content) may produce SSBP; e.g., young Sprague–Dawley rats have high levels of 20-HETE but are SS owing to poor correlation of 20-HETE with natriuresis, a phenotype that disappears with age. We have shown reduced urinary excretion of 20-HETE and loss of the normal relationship between 20-HETE and Na excretion in SS hypertensive subjects. Impaired 20-HETE responses to salt were related to a gene– environment interaction between the C allele of the T8590C polymorphism of CYP4A11 and insulin-inhibition of 20-HETE [10]. The T8590C polymorphism produces an enzyme with half the catalytic activity of the wild type protein, which has been linked to hypertension in population studies.

CLINICAL SIGNIFICANCE Owing to the prognostic significance of the SSBP phenotype it is important to identify biochemical or genetic markers that could diagnose it in the clinic, without

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having to resort to the cumbersome protocols currently used in research. Such markers could be used for risk stratification of hypertensive and pre-hypertensive subjects, to determine therapeutic BP goals and intensity of antihypertensive therapy for SS subjects. Understanding of the mechanisms of SSBP may lead to development of specific therapy for this phenotype, regardless of the magnitude of BP elevation, and so contribute to decreased cardiovascular risk in pre-hypertensive and hypertensive SS subjects.

References [1] Weinberger MH, Fineberg NS, Fineberg SE, Weinberger M. Salt sensitivity, pulse pressure, and death in normal and hypertensive humans. Hypertension 2001;37:429–32. [2] Chen J, Gu D, Huang J, Rao DC, Jaquish CE, Hixson JE, et al. GenSalt Collaborative Research Group. Metabolic syndrome and salt sensitivity of blood pressure in non-diabetic people in China: a dietary intervention study. Lancet 2009;373:829–35. [3] Anderson DE, Fedorova OV, Morrell CH, Longo DL, Kashkin VA, Metzler JD, et al. Endogenous sodium pump inhibitors and ageassociated increases in salt sensitivity of blood pressure in normotensives. Am J Physiol 2008;294:R1248–54.

[4] Saxena AR, Karumanchi SA, Brown NJ, Royle CM, McElrath TF, Seely EW. Increased sensitivity to angiotensin II is present postpartum in women with a history of hypertensive pregnancy. Hypertension 2010;55:1239–45. [5] Simonetti GD, Raio L, Surbek D, Nelle M, Frey FJ, Mohaupt MG. Salt sensitivity of children with low birth weight. Hypertension 2008;52:625–30. [6] de Boer MP, Ijzerman RG, de Jongh RT, Eringa EC, Stehouwer CDA, Smulders YM, et al. Birth weight relates to salt sensitivity of blood pressure in healthy adults. Hypertension 2008;51:928–32. [7] Hirata K, Kume S, Araki S, Sakaguchi M, Chin-Kanasaki M, Isshiki K, et al. Exendin-4 has an anti-hypertensive effect in salt-sensitive mice model. Biochem Bioph Res Co 2009;380:44–9. [8] Gao F, Wang DH. Impairment in function and expression of transient receptor potential vanilloid type 4 in Dahl salt-sensitive rats: significance and mechanism. Hypertension 2010;55:1018–25. [9] Zimmermann-Viehoff F, Weber CS, Merswolken M, Rudat M, Deter HC. Low anxiety males display higher degree of salt sensitivity, increased autonomic reactivity, and higher defensiveness. Am J Hypertens 2008;21:1292–7. [10] Laffer CL, Gainer JV, Waterman MR, Capdevila JH, LaniadoSchwartzman M, Nasjletti A, et al. The T8590C polymorphism of CYP4A11 and 20-hydroxyeicosatetraenoic acid in essential hypertension. Hypertension 2008;51:767–72.

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C H A P T E R

66 Endothelial Dysfunction Julian P.J. Halcox INTRODUCTION

ENDOTHELIAL CELL DYSFUNCTION

The vascular endothelium comprises the thin inner layer of cells lining arteries and veins. Previously thought to serve as a simple inert “barrier”, it is now well-recognized that vascular endothelial cells have an essential role in the regulation of blood vessel tone and cellular activity. Endothelial cells produce several important vasoactive substances including nitric oxide (NO), prostacyclin, endothelium-derived hyperpolarizing factor (EDHF), carbon monoxide, endothelin, vasoactive prostanoids, and superoxide. These factors, in addition to other substances produced by the endothelium, also modulate local thrombotic and inflammatory pathways influencing the development and progression of atherosclerosis and its complications.

Traditional cardiovascular risk factors such as diabetes, hypertension, dyslipidemia, and tobacco toxins are associated with endothelial dysfunction. This is typically characterized by reduced local bioavailability of NO in the context of increased oxidative stress; the latter in large part due to increased NADH/NADPH and xanthine-oxidase activity. Superoxide and other pro-oxidant radicals metabolize NO to nitrite and nitrate, as well as pro-oxidant peroxynitrite that promotes further generation of free radicals. Indeed, in these circumstances eNOS may be unable to generate NO, but instead generates superoxide, a situation known as “uncoupling” of the enzyme. Concurrently, transcriptional factors such as NF-κB and SREBP-1 become activated and translocate to the nucleus where they promote inflammatory gene transcription. This results in generation of a range of cytokines and chemokines, expression of selectins and adhesion molecules on the endothelial cell surface and also their release into the circulation. These pathways serve to attract and then encourage the arrest, firm adhesion and subsequent transendothelial migration of leukocytes into the sub-intimal space. Ongoing endothelial activation and dysfunction is a well recognized feature within the arterial wall at all stages of the atherosclerotic disease continuum and it is now widely accepted that the endothelium has an important role not only in the initiation, but also an important influence on the development and destabilization of atherosclerotic lesions. Regional mechanical influences on endothelial cell biology also play an important part in the location and evolution of atherosclerotic plaque. These help explain why lesions tend to develop at sites of bifurcation and curvature in the coronary and carotid arteries, which lead to marked shear-stress gradients in these high flow systems. Exposure of endothelial cells in vitro to low and oscillatory shear stress activates inflammatory transcription factors, promotes apoptosis and enhances endothelial cell turnover all contributing to the creation of an atherogenic milieu. Large animal and invasive clinical studies have confirmed the importance of these processes by demonstrating that that shear gradients predict the development

NORMAL ENDOTHELIAL CELL FUNCTION A healthy arterial endothelium is composed of a confluent layer of spindle-shaped endothelial cells that are bound together by tight junctions and communicate directly with each other and the underlying smooth muscle cells via gap-junctions. This forms a protective and biologically responsive cell layer that is relatively impermeable to low-density lipoprotein (the core-component of atherosclerotic lesions), able to sense molecular cues and interact with cellular components of the circulating blood. A functionally intact endothelium is characterized by its ability to secrete NO, a diatomic molecule that is produced from L-arginine by the enzyme endothelial NO synthase (eNOS). NO is constitutively generated and modulates underlying cellular functions by diffusion into vascular smooth muscle cells where it activates G-protein bound guanylyl cyclase. This enzyme generates cyclic GMP which promotes smooth muscle relaxation leading to vasodilatation. NO also has antiplatelet effects and can downregulate inflammatory pathways and generation of endothelin-1, a potent vasoconstrictor polypeptide, which also possesses pro-inflammatory pro-oxidant and pro-proliferative activity.

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of fibroatheromas with low endothelial shear stress in particular associated with mural inflammation, remodeling and lesion progression.

CLINICAL ASSESSMENT OF ENDOTHELIAL VASOMOTOR FUNCTION Clinical measurement of the ability of the endothelium to influence vascular tone is the usual strategy employed to assess endothelial function. The most commonly employed techniques include (a) administration of pharmacologic agents that act on the endothelium (including acetylcholine, bradykinin and substance-P); and (b) measurement of vasodilator response of the brachial artery in response to physiologically increased shear stress. These provoke the endothelium to release vasoactive substances including a significant amount of NO which is a principal determinant of the magnitude of the vasodilator response to these stimuli (see Fig. 66.1).

Basal NO Activity in the Healthy Human Circulation Inhibition of eNOS by administration of L NG monomethyl arginine (L-NMMA), elevates systemic blood pressure, reduces coronary blood flow and constricts epicardial coronary and peripheral arteries in healthy subjects. This suggests that tonic basal NO release contributes significantly to the normal maintenance of resting vasodilator tone. However, subjects with atherosclerosis or risk factors have a diminished vasoconstrictor response to L-NMMA consistent with reduced endothelial NO bioavailability.

Pharmacologic Endothelial Function Testing Conduit vessel and microvascular responses to endothelium-dependent pharmacological agents can be measured in the coronary circulation during cardiac catheterization using quantitative coronary angiography and Doppler flow wire techniques and in the peripheral circulation by a combination of high-resolution ultrasound and venous occlusion plethysmography. Most of the conduit vessel dilator response to standard pharmacologic stimuli is driven by NO. This factor is, however, only partially responsible for endotheliumdependent microvascular dilatation, which is predominantly due to endothelium-dependent hyperpolarization. The nature of EDHF is still a matter of debate, although activation of calcium-sensitive potassium channels and intercellular electrical communication via gap junctions appears to be a consistent requirement. Several putative EDHFs have been described, including hydrogen peroxide, potassium ions and epoxides generated by cytochrome P450 from arachidonic acid. The role of these

individual factors appears to differ between species and vascular beds. Subjects with atherosclerosis or its risk factors typically manifest with impaired dilator or constrictor responses to endothelium-dependent pharmacologic probes. For example a constrictor response to acetylcholine, the most commonly used agent, is observed when direct smooth muscle muscarinic receptor stimulation predominates over an attenuated dilator effect when endothelium-derived NO is reduced or absent.

Physiologic Endothelial Function Testing A major limitation of pharmacologic endothelial function testing is the need for invasive administration of the probes restricting the wider use of these techniques. Conduit vessel endothelium-dependent vasodilator function can also be assessed safely and reliably, and therefore repeatedly using 2-D ultrasound to measure flow-mediated dilatation (FMD) of a conduit vessel. The brachial artery is the most commonly used vessel, but the radial artery and leg arteries have also been studied in this way. This technique harnesses the ability of a brief ischemic stimulus (5-minutes) induced by supra-systolic inflation of a blood-pressure cuff to generate reactive hyperemia on cuff release due to resistance vessel dilatation. This results in an increase in shear stress, which stimulates endothelial release of vasoactive substances. As in the coronary circulation, peripheral endothelium-dependent FMD is predominantly NO-dependent and is depressed in the presence of conventional risk factors for atherosclerosis. This technique is highly reproducible but has to be conducted under careful experimental conditions due to the inherent physiological influence of numerous stimuli on vascular tone. These include ambient environmental temperature, caffeine, fried and fatty food, recent intensive exercise or infection, smoking and psychological stress. A number of studies have shown that FMD is a strong independent predictor of progression of atherosclerotic disease, as is coronary endothelial function, both in terms of lesion progression and development of clinical events. Furthermore, a significant, but imperfect correlation exists between brachial and coronary endothelial dilator responses. Considered together, these studies consistently suggest that FMD can be used as an informative marker of systemic endothelial function with important relevance to the global atherogenic environment. However, FMD is unable to perfectly characterize regional coronary endothelial biology, which is also under the important and variable influence of local shear forces in addition to the systemic risk factors that influence brachial endothelial function. Several other non-invasive techniques have been developed recently to assess endothelial function. These include pulsewave and pulse contour analysis (measurement of the effect of salbutamol, which releases NO from

V. PATHOPHYSIOLOGICAL MECHANISMS

ClInICAl ConsEquEnCEs of EnDoTHElIAl DysfunCTIon

the endothelium, on the arterial waveform), cutaneous Doppler flowmetry (measuring responses to iontophoretic application of pharmacologic probes) and pulsewave velocity (measurement of NO-mediated slowing of brachio-radial pulsewave transit in response to reactive hyperemia). One of the most promising new techniques for assessment of peripheral endothelial function is pulse amplitude tonometry using the EndoPAT device. This technique uses digital tonometry to measure fingertip pulse amplitude at rest and in response to reactive hyperemia in the study finger using the contralateral side as a control. The autonomic nervous system also has an important influence on digital pulse amplitude, which may confound responses to hyperemia but opens up other opportunities for the application of this tool. These novel techniques have several advantages over ultrasound, in that they are cheaper and simpler to perform and are thus potentially more practical for larger studies. However, they require more comprehensive validation and assessment of FMD using ultrasound remains the gold standard non-invasive test for systemic endothelial function.

CIRCULATING BIOMARKERS OF ENDOTHELIAL DYSFUNCTION Markers of Endothelial Activation and Dysfunction Endothelial activation and dysfunction is characterized by a change in the balance of vasomotor factors released by the endothelium, expression of inflammatory cytokines and chemokines, expression and secretion of selectins and adhesion molecules and modulation of local thrombotic pathways (Table 66.1). Measurement of circulating levels of relevant markers/ mediators of these pathways has been shown to provide important pathophysiological insights into the influence of the endothelium on atherosclerotic disease processes, albeit limited by the fact that systemic levels of these TABLE 66.1 Examples of Circulating Biomarkers of Endothelial function Biological Process

Biomarker

Vasomotor function

NO, NO2, Endothelin-1, Big Endothelin Interleukins Chemokines Adhesion molecules (VCAM-1, ICAM-1) Selectins (eSelectin) PAI-1 tPA vWF Endothelial microparticles Circulating endothelial cells Colony forming unit EPC (characterized by flow cytometry)

Inflammation

Thrombosis Endothelial injury Endothelial repair

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biomarkers may not truly represent their local levels and/ or activity in the vascular wall. Endothelial cell injury typically accompanies vascular inflammation. Thus, fragments of activated endothelium, endothelial microparticles, and even entire endothelial cells are shed into the circulation. These can be measured in the blood and their circulating levels are increased in association with coronary endothelial dysfunction, unstable coronary syndromes and vasculitides.

Markers of Endothelial Repair and Regeneration Damage to the endothelium can be repaired by two putative mechanisms: (a) by local endothelial cell division and ingrowth; and (b) via attachment and incorporation of circulating endothelial progenitor cells. The latter cell population can be identified in the circulating blood as a population of cells expressing both stem/progenitor (CD133/CD34) and endothelial surface markers (e.g., VEGF-R2/CD44/CD31 etc) and minimal levels of the leukocyte common antigen, CD45. Culturing peripheral blood mononuclear cells in a specific medium that favors growth of endothelial lineage populations can also identify these cells. These so called “endothelial progenitor cells” (EPCs) are reduced in the presence of risk factors, correlate inversely with the risk factor profile and presence of endothelial vasomotor dysfunction. Furthermore, they appear to be predictive of vascular prognosis and can be increased by known beneficial interventions such as an exercise program or statin use. Although a direct role for circulating EPCs in clinical endothelial repair remains to be proven, there is considerable interest in the therapeutic potential of enhancing their endogenous number and activity in the preclinical situation. However, the exact mechanism by which these “EPCs” influence vascular repair remains controversial. Small animal models have recently provided convincing evidence that although these cells exert a favorable influence on endothelial repair and integrity, they do this by localizing to the area of injury and secreting paracrine factors that promote local endothelial regeneration but do not themselves become functionally incorporated into the endothelium. Human data are largely consistent with this model, but this remains to be confirmed in definitive clinical studies.

CLINICAL CONSEQUENCES OF ENDOTHELIAL DYSFUNCTION Endothelial NO, Physiologic Coronary Vasomotion, and Myocardial Ischemia Physiologic vasodilatation in response to stimuli such as exercise, pacing and cold-pressor testing, is depressed in the circulation of subjects with atherosclerosis and its risk factors, largely as a consequence of reduced NO bioavailability. This limitation may result in myocardial

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ischemia even in the presence of unobstructed, normal epicardial coronary arteries. Constriction of epicardial coronary arteries during physical or psychologic stress can increase the degree of lesional obstruction in these circumstances facilitating the development of myocardial ischemia. Although this phenomenon has been recognized for many years, it is only relatively recently that the critical role of the vascular endothelium in vasoregulation and the ability of shear stress to mediate NO release from the endothelium have been appreciated. Thus, endothelial dysfunction contributes directly and significantly to the ischemic burden in atherosclerosis.

Endothelial Function and Cardiovascular Prognosis The vast majority of acute coronary events occur as a result of sudden plaque rupture or erosion exposing the lipid and inflammatory cell rich core of the atherosclerotic plaque to the circulating blood which triggers thrombosis. Systemic factors such as blood pressure and generalized inflammation have a clear role, but local pathways that affect plaque biology, in particular matrix turnover and the integrity of the fibrous cap of the plaque can influence the ease and location of plaque rupture. The “shoulder region” of the plaque is particularly susceptible to damage due to increased mechanical stress and constrictor tone, as inflammatory cell aggregation and fibrinolytic enzyme activity is typically greater in this region. Emerging data implicate plaque neovascularization as an important determinant of plaque progression. Furthermore, intra-plaque hemorrhage from this dysfunctional neovasculature has been proposed as another common cause of plaque destabilization and acute events. Elegant experiments in large animal models have shown that proliferation of the vasa-vasorum is an early feature of risk factor exposure that is intimately related to endothelial dysfunction and atherosclerotic lesion progression, suggesting another essential link between dysfunctional endothelium and the manifestation of clinical vascular disease.

STRATEGIES TO IMPROVE ENDOTHELIAL FUNCTION Many different behavioral and pharmacological strategies have been shown to improve endothelial dysfunction over the short as well as intermediate to long term. Many pharmacological agents including ACEinhibitors, angiotensin receptor blockers, phosphodiesterase type-5 antagonists, endothelin antagonists and statins can improve endothelial function. With improved standardization of clinical assessment techniques, this may become an increasingly important means of assessing potential beneficial and harmful effects of new drugs on the vascular wall during their clinical development.

Lifestyle intervention with physical exercise can improve coronary and peripheral endothelial dysfunction. Enriching the diet with omega-3 polyunsaturated fatty acids and adoption of a “Mediterranean-style” diet can also improve vascular dysfunction, whereas a highfat meal is able to temporarily induce inflammation and endothelial dysfunction. In a similar vein, acute psychophysiological stress can induce transient endothelial dysfunction whereas viewing a pleasant, humorous movie can improve function. An interaction between the autonomic nervous system and vascular function in acute stress is highly likely and endothelin type A receptor activation is a putative link. A 20-minute period of arm ischemia leads to transient endothelial dysfunction, which can be measured by FMD and has proven to be a reliable clinical model of ischemiareperfusion (IR) injury. This can be ameliorated by direct ischemic preconditioning and pre-treatment with certain pharmaceutical agents but can also remote ischemic preconditioning through 3  5-minute cycles of ischemiareperfusion of the contralateral arm. Interestingly, this protective effect of remote ischemic preconditioning appears to be autonomically mediated as it can be prevented by ganglion blockade with trimetaphan.

ENDOTHELIAL DYSFUNCTION AND CARDIOVASCULAR PATHOPHYSIOLOGY: INTERACTION WITH THE AUTONOMIC NERVOUS SYSTEM It is clear that the autonomic nervous system has many important influences on the vascular endothelium as outlined in this and other chapters. Activation of the sympathetic nervous system is associated with a large number of effects that promote endothelial dysfunction (Table 66.2). Many of these pathways can be treated and result in improved cardiovascular prognosis, however much is incompletely understood and considerable further clinical study will be required to dissect the complex interactions TABLE 66.2 selected Effects of sympathetic Activation on Endothelial function Systemic Effect of Sympathetic Activation Increased BP Increased heart rate and cardiac output Activation of HPA axis

Activation of RAS and endothelin generation Platelet activation

Effect on EC Biology Increased mechanical stress on EC Increased shear gradients at vessel curvatures and bifurcations promoting local EC activation Increased BP, insulin resistance and visceral fat deposition promoting inflammation Increased BP and vascular inflammation and oxidative stress Direct EC activation, enhanced interaction with leukocytes

EC, endothelial cell; BP, blood pressure, HPA, hypothalamo-pituitary-adrenal; RAS, reninangiotensin system.

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FIGURE 66.1 Endothelium-derived vasodilators. Acetylcholine, bradykinin and substance P stimulate endothelial cells to release vasodilator substances via activation of specific receptors on the endothelial cell surface. Acetylcholine binds to M1 or M3 muscarinic receptors, bradykinin binds to B2 kinin receptors, and substance P binds to NK1 neurokinin receptors. Shear stress activates NOS, but does not release prostacyclin. Nitric oxide and prostacyclin also have inhibitory effects on platelet aggregation and adhesion. Endogenous vasoconstrictors. Angiotensin generated in circulating blood and by tissue-bound angiotensin converting enzyme causes vasoconstriction via activation of smooth muscle AT1 receptors. Endothelin vasoconstricts by activating ETA and ETB receptors on vascular smooth muscle. Plateletderived serotonin and thromboxane A2, and norepinephrine from sympathetic nerve terminals stimulate smooth muscles via 5-HT, TxA2 and α1 adrenergic receptors, respectively. BK, bradykinin; SP, substance P; ACH, acetylcholine; NOS, endothelial nitric oxide synthase; EDHF, endothelium-derived hyperpolarizing factor; PGI2, prostacyclin; R, receptor; ACE, angiotensin converting enzyme; AT I, angiotensin I; AT II, angiotensin II; ET-1, endothelin-1; 5-HT, 5-hydroxytryptamine (serotonin); PGF-2α, prostaglandin F-2α; TxA2, thromboxane-A2; O2, superoxide ion.

and identify new therapeutic opportunities. Ideally, prevention of the common causes of autonomic imbalance with predominant sympathetic activation such as obesity and physical deconditioning and minimizing psychophysiological stresses or improving coping strategies are likely to prove most effective at a population level, but will require major sociopolitical leverage to implement in modern societies. (Fig. 66.1)

Further Reading Chatzizisis YS, Coskun AU, Jonas M, Edelman ER, Feldman CL, Stone PH. Role of endothelial shear stress in the natural history of coronary atherosclerosis and vascular remodeling: molecular, cellular, and vascular behavior. J Am Coll Cardiol 2007;49:2379–93. Deanfield JE, Donald AE, Ferri C, Halcox JPJ, Halligan S, Lerman A, et al. Endothelial function and dysfunction. Part I: Methodological issues for assessment in the different vascular beds. A statement by

the Working Group on Endothelin and Endothelial Factors of the European Society of Hypertension. J Hypertens 2005;23:7–17. Deanfield JE, Halcox JPJ, Rabelink T. Endothelial function and dysfunction: Testing and clinical relevance. Circulation 2007;115:1285–95. Donald AE, Halcox JPJ, Charakida M, Storry C, Wallace SM, Cole TJ, et al. Methodological approaches to optimize reproducibility and power in clinical studies of flow-mediated dilation. J Am Coll Cardiol 2008;51:1959–64. Halcox JP, Schenke WH, Zalos G, Mincemoyer R, Prasad A, Waclawiw MA, et al. Prognostic value of coronary vascular endothelial dysfunction. Circulation 2002;106:653–8. Hansson GK. Inflammation, atherosclerosis, and coronary artery disease. N Engl J Med 2005;352:1685–95. Hirschi KK, Ingram DA, Yoder MC. Assessing identity, phenotype, and fate of endothelial progenitor cells. Arterioscler Thromb Vasc Biol 2008;28:1584–95. Quyyumi AA, Dakak N, Andrews NP, Husain S, Arora S, Gilligan DM, et al. Nitric oxide activity in the human coronary circulation. Impact of risk factors for coronary atherosclerosis. J Clin Invest 1995;95:1747–55.

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Rautou PE, Leroyer AS, Ramkhelawon B, Devue C, Duflaut D, Vion AC, et al. Microparticles from human atherosclerotic plaques promote endothelial ICAM-1-dependent monocyte adhesion and transendothelial migration. Circ Res 2011;108:335–43.

Yeboah J, Folsom AR, Burke GL, Johnson C, Polak JF, Post W, et al. Predictive value of brachial flow-mediated dilation for incident cardiovascular events in a population-based study: the multi-ethnic study of atherosclerosis. Circulation 2009;120:502–9.

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C H A P T E R

67 Inflammation, Immunity and the Autonomic Nervous System Paul J. Marvar, David G. Harrison INTRODUCTION Inflammation is the vascular response to foreign pathogens and irritants, which is designed to protect against injurious stimuli and to initiate healing. This response includes vasodilatation, increased blood flow to the affected tissue, increased vascular permeability, upregulation of vascular cell adhesion molecule expression and the production of chemokines. Inflammation enhances rolling and adhesion of inflammatory cells to the endothelium and their translocation into the interstitium. Over the last decade it has become clear that in addition to its protective role in eliminating foreign pathogens, inflammation also contributes to the pathogenesis of many diseases that are traditionally not considered infectious in etiology, including cardiovascular diseases such as atherosclerosis and hypertension [1]. It has also become evident that the brain, by regulating endocrine and autonomic nervous system functions, plays a pivotal role in detecting and modulating inflammation [2–4]. In this chapter we will discuss the neuroimmune system, and in particular the autonomic nervous system and its influence on immunity and inflammation.

GENERAL CONCEPTS REGARDING INNATE AND ADAPTIVE IMMUNITY The innate immune system provides a global, non-specific defense against pathogens. Major contributors to the innate immune system include epithelial cells, which prevent pathogen entry, phagocytes such as neutrophils and macrophages, the complement system and pattern recognition receptors. Among the pattern recognition receptors are the Toll-like receptors (TLRs) that sense “danger signals” such as double-stranded RNA, bacterial coat proteins, bacterial heat-shock proteins and other toxins. Cells such as neutrophils, macrophages, and dendritic cells express pattern recognition receptors that respond to these danger signals by secreting cytokines, producing nitric oxide and generating reactive oxygen species. These secreted molecules either kill or prevent growth of bacteria

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00067-6

and viruses, and condition other cells to participate in the immune response. In contrast to the innate immune system, the adaptive immune system provides highly specific responses to the invading pathogen. The predominant cells involved in adaptive immunity are B and T lymphocytes and antigen presenting cells (APCs), which provide a link between innate and adaptive immunity. During maturation in the thymus, T cells acquire the surface markers CD4 or CD8 and develop very specialized functions that correspond to these markers. CD4 cells are also referred to as helper T cells, and produce cytokines that modulate local immune responses and direct B cells to produce antibodies. CD8 cells also produce cytokines, but in addition promote killing of adjacent cells by release of cytotoxic molecules including granzyme B and perforin. The traditional concept regarding adaptive immunity is that APCs take up foreign proteins, such as those of bacteria and viruses and process them into short peptides that are presented in the context of major histocompatibility complexes (MHC). Dendritic cells, which present antigenic peptides within MHC class II, predominantly activate CD4 lymphocytes. After antigen processing, dendritic cells migrate to secondary lymphoid organs, including the spleen and lymph nodes, where they seek a T cell that has a T cell receptor (TCR) that recognizes the antigenic peptide. The interaction of the MHC with the TCR occurs at a region termed the immunological synapse, where the interaction of many ligands and receptors on the APC and T cells occurs (Fig. 67.1). Coordination of signals transmitted from the APC to the T cell is essential for full T cell activation. Upon activation, T cells proliferate, produce cytokines and express homing markers that promote egress from secondary lymphoid organs and homing to sites of peripheral inflammation. As a result of the above-described activation process, T cells exist in several states. The newly produced T cell that has not undergone antigen challenge is referred to as a naïve T cell. Upon activation, these cells proliferate and the resultant activated cells are referred to as effector cells. Upon resolution of the inflammatory response, most effector T cells undergo apoptosis, however, a minority persist as resting

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cells and inhibit proliferation and cytokine production of other T cells. For an in-depth review of the principles described above, the reader is referred to informative textbooks edited by Abbas and Lichtman [9].

THE AUTONOMIC NERVOUS SYSTEM, INFLAMMATION AND HYPERTENSION

FIGURE 67.1 Mechanisms of T cell activation. Antigen presenting cells (APCs) provide a direct link between innate and adaptive immunity. APCs take up foreign proteins and process them into short peptides that are presented in the context of major histocompatibility complexes (MHC). After antigen processing, dendritic cells migrate to secondary lymphoid organs where they interact with a T cell that has a T cell receptor (TCR) that recognizes the antigenic peptide. The interaction of the MHC with the TCR occurs at the immunological synapse, where the interaction of many ligands and receptors occurs between the APC and T cells, including the co-stimulatory signals between CD28 and the B7 ligands. Coordination of signals transmitted from the APC to the T cell is essential for full T-cell activation and can result in the production of either TH1 (pro-inflammatory) or TH2 (anti-inflammatory) cytokines.

memory T cells. Some T cells return to secondary lymphoid organs, where they are referred to as central memory cells, while others remain in the circulation, and are referred to as peripheral memory cells. Central memory T cells are capable of self-renewal, and exhibit a rapid response to a second challenge with the initial immunizing agent. In addition to the above considerations, CD4 cells further differentiate during activation to assume specialized roles. The cytokine milieu, the type of antigen presenting cell and the types of danger signals promoting the inflammatory response modulate this differentiation process. Classically, CD4 cells are polarized to either a TH1 or TH2 subtype (Fig. 67.1). TH1 cells produce the signature cytokines interferon gamma (INFγ), IL-2 and TNFα and are involved in inflammatory processes such as response to infections, autoimmune encephalomyelitis and arthritis [5,6]. TH2 cells produce cytokines such as IL-4 and IL-13 and are involved in allergic reactions and responses to parasites [7]. In addition, in the last decade, TH17 cells have been identified [8]. These develop independently of the TH1/TH2 pathways, and secrete the cytokines IL-17, IL-21 and IL-22. Finally, 5 to 10% of T cells have immunosuppressive properties. These T regulatory cells (Tregs), characterized by expression of CD25 and the transcription factor FoxP3, modulate function of antigen presenting

The autonomic nervous system is activated in response to infection or injury and its products have major effects on immune function and inflammation. In addition, cytokines released in the periphery, such as IL-1, can traverse the blood–brain barrier and activate the autonomic nervous system and the hypothalamic pituitary axis (HPA), leading to release of neurotransmitters and glucocorticoids, which in turn modulate innate and adaptive immunity [10]. This bi-directional communication impacts virtually all inflammatory diseases. Many primary and secondary immune organs such as the thymus, bone marrow, spleen and lymph nodes receive substantial sympathetic innervation and almost all immune cells express receptors for neurohormones and neurotransmitters [11]. For example, innate and adaptive immune cells express α and β-adrenergic receptors and of these, the β2-subtype predominates on T and B cells, whereas macrophages express both receptor subtypes [12]. The adrenergic receptors of immune cells modulate immune functions such as antigen presentation by dendritic cells, clonal expansion of lymphocytes, migration, cell trafficking and can either suppress or enhance immune responses [13–15]. With regard to adaptive immunity, Swanson et al. demonstrated that norepinephrine can either increase or decrease TH1 polarization of naïve CD4 T lymphocytes depending on the presence of other cytokines and that β2 adrenergic receptor blockade prevents these effects [16]. Others have reported that the duration of adrenergic stimulation can differentially affect the TH1 response. For example, in vitro studies of peripheral blood mononuclear cells have suggested that short-term adrenergic stimulation decreases TH1 responses [17] while prolonged pre-exposure with epinephrine enhances TH1 polarization [18]. In addition to these in vitro effects, norepinephrine seems essential for normal immune function in vivo. Alaniz et al. showed that mice deficient in dopamine β hydroxylase, which is required for norepinephrine synthesis, have impaired TH1-polarization of T cells in response to infection with listeria monocytogenes [19]. Prolonged sympathetic activation and increased circulating catecholamine levels are often present in patients with cardiovascular disease, hypertension and individuals experiencing chronic stress [20–22]. Relatively few studies have examined the in vivo immunoregulatory effects of chronic catecholamine infusion. Harris et al. showed that a 4-week infusion of catecholamines causes splenic atrophy,

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decreases total splenic T cells and reduces suppressor/ cytotoxic T cell subsets in rats [23]. In a more recent study from our laboratory, a two-week infusion of norepinephrine increased peripheral activation of CD4 cells and infiltration of CD3 cells in the aorta [24]. Studies in humans have also shown that catecholamines can modulate T cell phenotypes and adaptive immune responses. For example, in patients with chronic heart failure, the ratio of TH1 to TH2 cytokines was reduced in patients treated with either beta-blockers or angiotensin-converting enzyme (ACE) inhibitors, suggesting that sympathetic tone and activation of the renin/angiotensin system can modulate T cell polarization and cytokine production [25]. In other human diseases, such as rheumatoid arthritis, the ability of catecholamines to modulate T cell function is impaired, and in some cases this promotes a pro-inflammatory TH1 state [26,27]. Based on these studies it appears that catecholamines have immunoregulatory influences in various chronic inflammatory diseases. In summary, the existing data indicate that catecholamines have diverse effects on adaptive immunity, in particular the TH1 response, and this is dependent on differences in target cell type (naïve vs. effector T cells), duration of adrenergic stimulation and type of inflammation [28]. In addition to adaptive immune cells, innate immune cells also express various adrenergic receptor subtypes and antigen-presenting cells such as dendritic cells are affected by the sympathetic nervous system. For example, β2-adrenergic stimulation with norepinephrine enhances dendritic cell emigration from the skin following exposure to TLR agonists [29]. Beta-adrenergic stimulation reduces dendritic cell IL-12 production and increases production of the anti-inflammatory cytokine IL-10 [30]. Based on these and other data it has been suggested that adrenergic receptors on dendritic cells limit their TH1 priming ability and antigen presenting capacity and ultimately dampen inflammation [31–32]. Likewise norepinephrine has been shown to reduce macrophage production of TNFα and IL1β in response to LPS [33]. The parasympathetic nervous system also modulates inflammation by acting as an anti-inflammatory neural circuit [34–35]. The vagus nerve senses peripheral inflammation and transmits action potentials to the brain stem, in particular the area postrema and the nucleus tractus solitarius. This in turn increases vagal stimulation of lymphoid organs, and inhibits pro-inflammatory cytokine production [36]. This pathway is mediated by acetylcholine, the principal neurotransmitter of the parasympathetic nervous system and therefore commonly referred to as the cholinergic anti-inflammatory pathway. The α7 subunit of nicotinic acetylcholine receptor is expressed on monocytes and macrophages and when exposed to acetylcholine, signals a reduction of proinflammatory cytokines such as TNFα and IL6 [34]. Efferent vagal stimulation inhibits synthesis of TNFα in liver, spleen, heart and attenuates serum TNFα levels in inflammatory diseases such as ischemia reperfusion injury

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and peritonitis [37–39]. Besides macrophages, the α7 subunit of nicotinic acetylcholine receptor is also expressed on CD4 T cells, however much less is known regarding its function on these cells [40]. Recently though, Karimi et al. demonstrated that subdiaphragmatic vagotomy increases TNFα and INFγ secretion from CD4 splenic cells [41]. In addition, several clinical studies have shown inverse correlations between markers of inflammation and vagus nerve activity. For example, multiple studies in humans have found that levels of the inflammatory markers C-reactive protein and IL-6 correlate inversely with vagal activity, as estimated by heart rate variability [42–45]. Thus, as suggested by Tracy et al., dysfunction of the cholinergic anti-inflammatory pathway can exacerbate inflammatory responses in various pathological conditions including cardiovascular disease [34]. There is also evidence that inflammation in the CNS leads to autonomic dysfunction [46–48]. Pro-inflammatory cytokines produced in the periphery can diffuse into circumventricular organs (CVO), including the subfornical organ, the organum vasculosum lamina terminalis (OVLT) and the median preoptic eminence (Fig. 67.2) [49]. Paton and colleagues demonstrated that inflammatory cells and cytokines are increased in the brain in experimental hypertension, and that these may impair central autonomic control of blood pressure regulation [46,50]. Zhang et al. recently reported that angiotensin II increases permeability of the blood–brain barrier and causes cerebral microvasculature inflammation [51]. Moreover, recent studies have shown that microglial cells, a type of specialized macrophage in the central nervous system, are activated in experimental hypertension, and that inhibition of these cells lowers blood [47]. These data indicate that the blood– brain barrier is altered during hypertension, allowing increased passage of circulating pro-inflammatory molecules to the brain and ultimately may promote the dysfunctional autonomic regulation of blood pressure. To further understand these signals in the central control of blood pressure and how the neuroimmune interface may impact peripheral vascular inflammation, we recently investigated the role of the circumventricular organs (CVOs) in angiotensin II dependent hypertension. In these studies we inhibited the central actions of angiotensin II through electrolytic ablation of the anteroventral third ventricle (AV3V) region, which encompasses various CVOs in the hypothalamus and has previously been found to be critical to the central control of blood pressure [24]. AV3V lesioning attenuated the rise in blood pressure in response to a slow pressor dose of angiotensin II and completely prevented the activation and vascular infiltration of T cells. This demonstrated that central rather than peripheral effects of angiotensin II, were responsible for systemic inflammation caused by this octapeptide. In other studies we deleted the locally expressed extracellular superoxide dismutase in the CVO of the brain, and found that this increased sympathetic outflow, elevated blood pressure and increased T cell mediated peripheral

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vascular inflammation [52]. These data support previous work highlighting the importance of the CNS-immune interface and demonstrate a new role for the CNS in modulating peripheral T cell activation and vascular inflammation during hypertension.

CONCLUSION AND PERSPECTIVES The existing literature suggests that the autonomic nervous system has diverse effects on inflammation and immunity. While these interactions can vary, the

predominant pathways are summarized in Figure 67.2. The sympathetic nervous system, via catecholamine release and beta-adrenergic stimulation, promotes a proinflammatory response of T lymphocytes while suppressing macrophage and dendritic cell function. In addition, parasympathetic activation has anti-inflammatory effects via muscarinic signaling in macrophages, monocytes and CD4 T cells. Vagal afferents sense peripheral inflammatory states, and feedback to central sites to enhance antiinflammatory efferent cholinergic signals. There are also important direct effects of inflammatory mediators such as cytokines and angiotensin II in the brain. These effects occur at sites such as the circumventricular organs where the blood–brain barrier is poorly developed, and are enhanced because inflammation reduces blood–brain barrier function. Inflammatory signals in the brain, mediated in part by microglial activation, enhance sympathetic outflow and likely contribute to a variety of cardiovascular and metabolic disorders. These roles of the central nervous system almost certainly contribute to the pathology of conditions such as chronic stress, hypertension and heart failure and should be considered as future therapeutic targets.

References

FIGURE 67.2 Effects of the autonomic nervous system on innate and adaptive immune cells. Sympathetic innervation of lymphoid tissue and adrenergic (AR) receptor expression on immune cells contributes to the inflammatory process. Adrenergic stimulation exerts both stimulatory () and inhibitory effects (–) and are primarily inhibitory on innate immune cells such as macrophages and dendritic cells. Under TH1 -dependent conditions catecholamines enhance the pro-inflammatory response leading to increased release of cytokines such as INFγ and TNFα from T cells. The parasympathetic influences of the vagus nerve and its neurotransmitter acetycholine (ACh) are inhibitory. Vagal afferents in peripheral target tissue sense peripheral inflammatory states and feedback to the brainstem to contribute to the neuromodulation of autonomic activity. Cytokines can also penetrate the blood–brain barrier at circumventricular organs and contribute to autonomic dysfunction in chronic diseases. Abbreviations: median preoptic nuclei (MPO), paraventricular nuclei (PVN), subfornical organ (SFO), rostral ventrolateral medulla (RVLM), nucleus of the solitary tract (NTS), organum vasculosum of the lamina terminalis (OVLT).

[1] Harrison DG, Guzik TJ, Lob HE, Madhur MS, Marvar PJ, Thabet SR, et al. Inflammation, immunity, and hypertension. Hypertension 2011;57:132–40. [2] Sternberg EM. Neural regulation of innate immunity: A coordinated nonspecific host response to pathogens. Nat Rev Immunol 2006;6:318–28. [3] Webster JI, Tonelli L, Sternberg EM. Neuroendocrine regulation of immunity. Annu Rev Immunol 2002;20:125–63. [4] Elenkov IJ. Neurohormonal-cytokine interactions: Implications for inflammation, common human diseases and well-being. Neurochem Int 2008;52:40–51. [5] Notley CA, Inglis JJ, Alzabin S, McCann FE, McNamee KE, Williams RO. Blockade of tumor necrosis factor in collagen-induced arthritis reveals a novel immunoregulatory pathway for TH1 and TH17 cells. J Exp Med 2008;205:2491–7. [6] Fletcher JM, Lalor SJ, Sweeney CM, Tubridy N, Mills KH. T cells in multiple sclerosis and experimental autoimmune encephalomyelitis. Clin Exp Immunol 2010;162:1–11. [7] Lloyd CM, Hessel EM. Functions of T cells in asthma: More than just T(H)2 cells. Nat Rev Immunol 2010;10:838–48. [8] Park H, Li Z, Yang XO, Chang SH, Nurieva R, Wang YH, et al. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17. Nat Immunol 2005;6:1133–41. [9] Abbas A.K., Lichtman A.H. Cellular and molecular immunology. 2005:564. [10] Watkins LR, Maier SF. Immune regulation of central nervous system functions: From sickness responses to pathological pain. J Intern Med 2005;257:139–55. [11] Nance DM, Sanders VM. Autonomic innervation and regulation of the immune system (1987–2007). Brain Behav Immun 2007;21:736–45. [12] Sanders V.M., Kasprowicz D.J., Kohm A.P., Swanson M.A. Neurotransmitter receptors on lymphocytes and other lymphoid cells. 2001;2:161–196. [13] Steinman L. Elaborate interactions between the immune and nervous systems. Nat Immunol 2004;5:575–81. [14] Bellinger DL, Millar BA, Perez S, Carter J, Wood C, Thyaga Rajan S, et al. Sympathetic modulation of immunity: Relevance to disease. Cell Immunol 2008;252:27–56.

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[15] Sanders VM, Straub RH. Norepinephrine, the beta-adrenergic receptor, and immunity. Brain Behav Immun 2002;16:290–332. [16] Swanson MA, Lee WT, Sanders VM. IFN-gamma production by TH1 cells generated from naive CD4  T cells exposed to norepinephrine. J Immunol 2001;166:232–40. [17] van der Poll T, Coyle SM, Barbosa K, Braxton CC, Lowry SF. Epinephrine inhibits tumor necrosis factor-alpha and potentiates interleukin 10 production during human endotoxemia. J Clin Invest 1996;97:713–9. [18] Severn A, Rapson NT, Hunter CA, Liew FY. Regulation of tumor necrosis factor production by adrenaline and beta-adrenergic agonists. J Immunol 1992;148:3441–5. [19] Alaniz RC, Thomas SA, Perez-Melgosa M, Mueller K, Farr AG, Palmiter RD, et al. Dopamine beta-hydroxylase deficiency impairs cellular immunity. Proc Natl Acad Sci USA 1999;96:2274–8. [20] Malpas SC. Sympathetic nervous system overactivity and its role in the development of cardiovascular disease. Physiol Rev 2010;90:513–57. [21] Pickering TG. Stress, inflammation, and hypertension. J Clin Hypertens (Greenwich) 2007;9:567–71. [22] Schlaich MP, Lambert E, Kaye DM, Krozowski Z, Campbell DJ, Lambert G, et al. Sympathetic augmentation in hypertension: Role of nerve firing, norepinephrine reuptake, and angiotensin neuromodulation. Hypertension 2004;43:169–75. [23] Harris TJ, Waltman TJ, Carter SM, Maisel AS. Effect of prolonged catecholamine infusion on immunoregulatory function: Implications in congestive heart failure. J Am Coll Cardiol 1995;26:102–9. [24] Marvar PJ, Thabet SR, Guzik TJ, Lob HE, McCann LA, Weyand C, et al. Central and peripheral mechanisms of t-lymphocyte activation and vascular inflammation produced by angiotensin II-induced hypertension. Circ Res 2010;107:263–70. [25] Gage JR, Fonarow G, Hamilton M, Widawski M, Martinez-Maza O, Vredevoe DL. Beta blocker and angiotensin-converting enzyme inhibitor therapy is associated with decreased TH1/TH2 cytokine ratios and inflammatory cytokine production in patients with chronic heart failure. Neuroimmunomodulation 2004;11:173–80. [26] Wahle M, Hanefeld G, Brunn S, Straub RH, Wagner U, Krause A, et al. Failure of catecholamines to shift T-cell cytokine responses toward a TH2 profile in patients with rheumatoid arthritis. Arthritis Res Ther 2006;8:R138. [27] Firestein GS. Evolving concepts of rheumatoid arthritis. Nature 2003;423:356–61. [28] Kohm AP, Sanders VM. Norepinephrine and beta 2-adrenergic receptor stimulation regulate CD4  T and B lymphocyte function in vitro and in vivo. Pharmacol Rev 2001;53:487–525. [29] Maestroni GJ. Dendritic cell migration controlled by alpha 1b-adrenergic receptors. J Immunol 2000;165:6743–7. [30] Maestroni GJ. Sympathetic nervous system influence on the innate immune response. Ann N Y Acad Sci 2006;1069:195–207. [31] Panina-Bordignon P, Mazzeo D, Lucia PD, D’Ambrosio D, Lang R, Fabbri L, et al. Beta2-agonists prevent TH1 development by selective inhibition of interleukin 12. J Clin Invest 1997;100:1513–9. [32] Goyarts E, Matsui M, Mammone T, Bender AM, Wagner JA, Maes D, et al. Norepinephrine modulates human dendritic cell activation by altering cytokine release. Exp Dermatol 2008;17:188–96. [33] Ignatowski TA, Gallant S, Spengler RN. Temporal regulation by adrenergic receptor stimulation of macrophage (M phi)-derived tumor necrosis factor (TNF) production post-LPS challenge. J Neuroimmunol 1996;65:107–17.

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[34] Tracey KJ. The inflammatory reflex. Nature 2002;420:853–9. [35] Czura CJ, Tracey KJ. Autonomic neural regulation of immunity. J Intern Med 2005;257:156–66. [36] Huston JM, Ochani M, Rosas-Ballina M, Liao H, Ochani K, Pavlov VA, et al. Splenectomy inactivates the cholinergic antiinflammatory pathway during lethal endotoxemia and polymicrobial sepsis. J Exp Med 2006;203:1623–8. [37] Bernik TR, Friedman SG, Ochani M, DiRaimo R, Susarla S, Czura CJ, et al. Cholinergic anti-inflammatory pathway inhibition of tumor necrosis factor during ischemia reperfusion. J Vasc Surg 2002;36:1231–6. [38] Borovikova LV, Ivanova S, Zhang M, Yang H, Botchkina GI, Watkins LR, et al. Vagus nerve stimulation attenuates the systemic inflammatory response to endotoxin. Nature 2000;405:458–62. [39] Wang H, Yu M, Ochani M, Amella CA, Tanovic M, Susarla S, et al. Nicotinic acetylcholine receptor alpha7 subunit is an essential regulator of inflammation. Nature 2003;421:384–8. [40] Zhang S, Petro TM. The effect of nicotine on murine CD4 T cell responses. Int J Immunopharmacol 1996;18:467–78. [41] Karimi K, Bienenstock J, Wang L, Forsythe P. The vagus nerve modulates CD4  T cell activity. Brain Behav Immun 2010;24:316–23. [42] Carney RM, Freedland KE, Stein PK, Miller GE, Steinmeyer B, Rich MW, et al. Heart rate variability and markers of inflammation and coagulation in depressed patients with coronary heart disease. J Psychosom Res 2007;62:463–7. [43] Haensel A, Mills PJ, Nelesen RA, Ziegler MG, Dimsdale JE. The relationship between heart rate variability and inflammatory markers in cardiovascular diseases. Psychoneuroendocrinology 2008;33:1305–12. [44] Lampert R, Bremner JD, Su S, Miller A, Lee F, Cheema F, et al. Decreased heart rate variability is associated with higher levels of inflammation in middle-aged men. Am Heart J 2008;156(759):e751–7. [45] Frasure-Smith N, Lesperance F, Irwin MR, Talajic M, Pollock BG. The relationships among heart rate variability, inflammatory markers and depression in coronary heart disease patients. Brain Behav Immun 2009;23:1140–7. [46] Paton JF, Waki H. Is neurogenic hypertension related to vascular inflammation of the brainstem?. Neurosci Biobehav Rev 2009;33:89–94. [47] Shi P, Raizada MK, Sumners C. Brain cytokines as neuromodulators in cardiovascular control. Clin Exp Pharmacol Physiol 2010;37:e52–7. [48] Yu Y, Zhang ZH, Wei SG, Serrats J, Weiss RM, Felder RB. Brain perivascular macrophages and the sympathetic response to inflammation in rats after myocardial infarction. Hypertension 2010;55:652–9. [49] Banks WA, Kastin AJ, Broadwell RD. Passage of cytokines across the blood–brain barrier. Neuroimmunomodulation 1995;2:241–8. [50] Waki H, Gouraud SS, Maeda M, Paton JF. Evidence of specific inflammatory condition in nucleus tractus solitarii of spontaneously hypertensive rats. Exp Physiol 2010;95:595–600. [51] Zhang M., Mao Y., Ramirez S.H., Tuma R.F., Chabrashvili T. Angiotensin ii induced cerebral microvascular inflammation and increased blood–brain barrier permeability via oxidative stress. Neuroscience. 2010;171:852–8. [52] Lob HE, Marvar PJ, Guzik TJ, Sharma S, McCann LA, Weyand C, et al. Induction of hypertension and peripheral inflammation by reduction of extracellular superoxide dismutase in the central nervous system. Hypertension 2010;55:277–83.

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68 Oxygen Sensing Nanduri R. Prabhakar O2 availability is essential for survival of mammalian cells, as it is essential for generating ATP, a major source of energy for cellular functions. Consequently, it is not surprising that all mammalian cells respond to decreased availability of O2 or hypoxia, albeit to different degrees, by readjusting their metabolism. However, this response to hypoxia requires minutes to hours. On other hand, when O2 levels decrease even modestly in the arterial blood (i.e., hypoxemia), within seconds it stimulates breathing and elevates blood pressure. Such an immediate response to systemic hypoxia is mediated by specialized organs called “arterial chemoreceptors”, which sense arterial blood O2 levels and respond with increased sensory nerve activity to hypoxia [1,9]. The afferent nerve activity is transmitted to the central nervous system and triggers reflex activation of autonomic nervous system manifested as increased breathing and sympathetic activity which are critical for ensuring adequate supply of oxygen to tissues. Thus, arterial chemoreceptors are the sensory organs for monitoring arterial blood O2 levels similar to olfactory sensory neurons that detect odors. In the absence of O2 sensing by the chemoreceptors, autonomic responses to systemic hypoxia are either attenuated or absent, leading to tissue/cellular hypoxia which can be potentially deleterious to tissues or cells. The best studied example of arterial chemoreceptors are the carotid bodies, which are situated bilaterally at the bifurcation of the common carotid artery. The purpose of this chapter is to provide a brief up-date on the recent advances on the mechanisms as well as physiological and pathological consequences of O2 sensing by the carotid body on autonomic functions.

MORPHOLOGY OF THE CAROTID BODY AND MEASURES OF O2 SENSING Carotid body, a highly vascular tissue receives afferent innervation from the carotid sinus nerve, which is a branch of the glossopharyngeal nerve. The chemoreceptor tissue is primarily composed of type I (also called glomus cells) and type II cells (also called sustentacular cells) along with vascular endothelial cells and few ganglion cells. Type I cells are of neural crest in origin and express a variety of neurotransmitters, whereas the type II cells resemble glial cells of the nervous system. Type I cells

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form synaptic contacts with the afferent nerve endings whose cell bodies lie in the petrosal ganglion. Much of the available evidence suggests that the type I cells are the initial sites of O2 sensing and they work in concert with the opposing afferent nerve ending functioning as a “sensory unit” [1,9]. Sensory nerve response to hypoxia is often regarded as a “standard” measure of O2 sensing by the carotid body, because it is essential for triggering reflex activation of the autonomic nervous system [8]. Although hypoxia facilitate dopamine secretion from glomus cells and inhibit K conductances in these cells [1], they often do not correlate with the afferent nerve activation by low O2 [8]. Basal sensory activity is low and increases progressively in response to graded severity of hypoxia in a curvilinear fashion.

UNIQUENESS OF CAROTID BODY O2 SENSING The following features distinguish O2 sensing by the carotid body from other tissues. First, a modest level of hypoxia (PO2 of ~80 mmHg) is adequate to stimulate the carotid body sensory activity, whereas much severe hypoxia (PO2 of ~40 mmHg) is required to elicit a response in other tissues, e.g., erythropoietin production from kidney. Second, increase in afferent nerve activity of the carotid body occurs within seconds after the onset of hypoxia, whereas response of other tissues/or cells to hypoxia develop more slowly reaching maximum over minutes to hours. Third, the increased sensory activity of the carotid body is maintained during the entire period of hypoxia with little adaptation. Thus, the exquisite sensitivity and the rapid response to a wide range of hypoxic intensities with little or no adaptation make the carotid body a unique O2 sensing organ in comparison to other tissues.

O2 SENSORS AND TRANSDUCTION OF THE HYPOXIC STIMULUS Several hypotheses have been proposed to explain the mechanisms underlying O2 sensing by the carotid body. Central to all these hypotheses is the premise

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that glomus cells are the primary site of O2 sensing and that hypoxia releases neurotransmitter(s) from glomus cells, which by way of depolarizing the nearby afferent nerve endings increase the sensory nerve discharge [1,9]. It has been proposed that hypoxia inhibits certain K channel/s (e.g., Ca2 activated K channel, TASK) in glomus cells leading to depolarization and the resulting voltage-gated Ca2 influx triggers the neurotransmitter release (membrane hypothesis). Consistent with this possibility, a variety of K channels were shown to be inhibited by hypoxia in glomus cells. However, these effects of hypoxia on K channels are species and developmental age dependent, making the identity of the K channel(s) responsible for initiating depolarization in Type I cells difficult [4]. Inhibitors of mitochondrial electron transport chain (ETC), like hypoxia, stimulate the carotid body activity. Consequently, it was thought that mitochondrial enzyme(s) associated with oxidative metabolism is critical for hypoxic sensing by the carotid body (mitochondrial/metabolic hypothesis). Supporting the mitochondrial hypothesis, hypoxia depolarizes mitochondrial membrane potential in a stimulus- dependent manner. However, it is unclear how depolarization of mitochondrial membrane potential is linked to K channel inhibition and triggers transmitter release. K channels have been identified as downstream targets for AMP-activated protein kinase (AMPK), a serine/threonine kinase that is activated by increases in the cellular AMP/ATP ratio. Consequently, it has been proposed that AMPK activation by hypoxia is necessary for O2 sensing by the carotid body [4]. Whether hypoxia affects AMPK activity, however, has not yet been demonstrated. Thus, there is considerable evidence supporting as well as questioning both the membrane and metabolic hypotheses.

ROLE OF GAS MESSENGERS IN HYPOXIC SENSING BY THE CAROTID BODY Hydrogen sulfide (H2S) is a gas molecule generated by mammalian cells via a variety of enzymes including cystathionine γ-lyase (CSE) using cysteine as substrate. A recent study showed that CSE is expressed in the glomus cells of the rat and mouse carotid bodies [6]. Mice with genetic absence of CSE [6] as well as pharmacological blockade of H2S generation [3] exhibit selective impairment of carotid body and ventilatory responses to hypoxia. Furthermore, H2S generation in the carotid body, like the sensory activity, was low under normoxia and increased during hypoxia in an O2-dependent manner [6]. H2S inhibited O2 sensitive calcium-activated K channels in heterologous expression system [10]. Furthermore, H2S, like hypoxia, leads to reduced red-ox cellular environment. Based on these findings it was proposed that increased H2S generation during hypoxia mediates carotid body sensory excitation [6].

NORMOXIA

HYPOXIA

Hemeoxygenase-2 (+)

Hemeoxygenase-2 (–)

Carbon monoxide (+)

Carbon monoxide (–)

Cystathionine gama-lyase(–)

H2S (–)

Low Sensory activity

Cystathionine gama-lyase (+)

H2S (+)

High Sensory activity

FIGURE 68.1 Schematic illustration of the interaction between heme oxygenase-2 and carbon monoxide and cytathionine gamma-lyase and H2S generation in the carotid body under normoxia and hypoxia and its effects on carotid body sensory activity.

How does hypoxia increase H2S generation in the carotid body? Glomus cells express heme oxygenase-2 (HO-2), an enzyme that catalyzes the formation of endogenous carbon monoxide (CO) from heme, and requires molecular O2 for its enzyme activity. CO functions as an inhibitory gas messenger to the carotid body activity [9]. Pharmacological inhibitor of HO-2 increased H2S generation under normoxia and stimulated carotid body sensory activity in wild type mice but not in CSE knockout mice [6] suggesting a regulatory role for CO in H2S production from CSE. Since molecular O2 is required for HO-2 activity, it is conceivable that CO is continuously produced during normoxia which in turn inhibits H2S generation resulting in low levels of sensory activity. During hypoxia, HO-2 activity is inhibited because of decreased availability of molecular O2, resulting in reduced CO generation, thereby removing its inhibitory influence on H2S generation from CSE and increase sensory activity. Figure 68.1 illustrates the switching interaction between HO-2-CO and CSE- H2S systems in the carotid body.

MOLECULAR DETERMINANTS OF O2 SENSING BY THE CAROTID BODY Hypoxia-inducible factors 1 and 2 (HIF-1 and HIF-2) are a well studied family of transcriptional activators that mediate genomic responses to hypoxia. Both HIF-1 and HIF-2 are expressed in glomus cells. Recent studies showed that mice with partial deficiency of HIF-1α, the O2 sensitive regulatory subunit of the HIF-1complex exhibit selective impairment of the carotid body response to acute hypoxia [2]. In striking contrast, mice with HIF-2α deficiency display augmented hypoxic sensing by the carotid body [5]. These findings suggest that functional

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COnSEQUEnCES OF CAROTID BODy O2 SEnSIng On AUTOnOMIC nERVOUS SySTEM

antagonism between HIF-1 and HIF-2 plays a fundamental role in maintaining the “normal” hypoxic sensitivity of the carotid body. Hyposensitivity of the carotid body to O2 such as that seen with HIF-1α deficiency may otherwise adversely impact physiological adaptations to hypoxia, whereas hypersensitivity to hypoxia seen with HIF-2α deficiency may result in unwarranted sympathetic excitation such as that associated with many pathological situations (see below).

CONSEQUENCES OF CAROTID BODY O2 SENSING ON AUTONOMIC NERVOUS SYSTEM Physiological Situations There is considerable evidence suggesting that reflexes arising from the carotid bodies are critical for maintaining autonomic responses to exercise, diving reflex, adaptations to high altitude as well as during pregnancy [7].

Pathological Situations Sleep disordered breathing with recurrent apneas (transient, repetitive cessations of breathing) cause periodic decreases in arterial blood O2 levels or intermittent hypoxia (IH). Recurrent apnea patients and rodents exposed to IH exhibit elevated sympathetic nerve activity, blood pressure and circulating catecholamines, which were attributed to exaggerated carotid body reflex. Consistent with this possibility are the findings that (a) IH treated rodents exhibit augmented hypoxic sensory response of the carotid body; (b) denervation of the carotid bodies prevent sympathetic activation by IH; and (c) glomectomized human subjects with recurrent apneas do not develop hypertension [7]. Sympatho-humoral activation is a hall mark of congestive heart failure (CHF), which contributes to progression and ultimate mortality of the disease, which is a major health care problem. Recent studies provided convincing evidence that chemo-reflex arising from the carotid body mediates sympathetic activation in CHF [7]. Patients with essential hypertension exhibit enhanced sympathetic nerve activity and ventilatory

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response to hypoxia, which were attributed to exaggerated carotid body response to low O2. Indeed, spontaneous hypertensive rats (SHR) exhibit exaggerated carotid body response to hypoxia [7]. Thus, altered hypoxic sensing by the carotid body mediates autonomic morbidities associated with most commonly prevalent diseases such as recurrent apneas, CHF and hypertension.

Acknowledgements The research in author’s laboratory is supported by National Institutes of Health Grants HL-76537, HL-90554 and HL-86493. N.R.P. is a Harold H. Hines Jr. Professor at the University of Chicago.

References [1] Gonzalez CL, Almaraz L, Obeso A, Rigual R. Carotid body chemoreceptors: from natural stimuli to sensory discharges. Physiol Rev 1994;74:829–98. [2] Kline DD, Peng YJ, Manalo DJ, Semenza GL, Prabhakar NR. Defective carotid body function and impaired ventilatory responses to chronic hypoxia in mice partially deficient for hypoxia-inducible factor 1 alpha. Proc Natl Acad Sci USA 2002;99:821–6. [3] Li Q, Sun B, Wang X, Jin Z, Zhou Y, Dong L, et al. Crucial role for hydrogen sulfide in oxygen sensing via modulating large conductance calcium-activated potassium channels. Antioxid Redox Signal 2010;12:1179–89. [4] Peers C, Wyatt CN, Evans AM. Mechanisms for acute oxygen sensing in the carotid body. Respir Physiol Neurobiol 2010;174:292–8. [5] Peng YJ, Nanduri J, Khan SA, Yuan G, Wang N, Kinsman B, et al. Hypoxia-inducible factor 2α (HIF-2 α) heterozygous-null mice exhibit exaggerated carotid body sensitivity to hypoxia, breathing instability, and hypertension. Proc Natl Acad Sci USA 2011; 108:3065–70. [6] Peng YJ, Nanduri J, Raghuraman G, Souvannakitti D, Gadalla MM, Kumar GK, et al. H2S mediates O2 sensing in the carotid body. Proc Natl Acad Sci USA 2010;107:10719–724. [7] Prabhakar NR, Peng YJ. Peripheral chemoreceptors in health and disease. J Appl Physiol 2004;96:359–66. [8] Prabhakar NR. O2 sensing at the mammalian carotid body: why multiple O2 sensors and multiple transmitters? Exp Physiol 2006;91:17–23. [9] Prabhakar NR. Oxygen sensing by the carotid body chemoreceptors. J Appl Physiol 2000;88:2287–95. [10] Telezhkin V, Brazier SP, Cayzac SH, Wilkinson WJ, Riccardi D, Kemp PJ. Mechanism of inhibition by hydrogen sulfide of native and recombinant BKCa channels. Respir Physiol Neurobiol 2010;172:169–78.

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69 Reactive Oxygen Species and Oxidative Stress Rhian M. Touyz INTRODUCTION

CNS Stress response

ROS, originally considered to induce negative and injurious cellular effects, such as cell death, are now recognized to have important positive actions including induction of host defense genes, activation of kinases/ phosphatases, regulation of transcription factors and mobilization of ion transporters. Molecular processes whereby ROS influence function of cells involve activation of redox-sensitive signaling pathways [1]. Superoxide anion (O2) and hydrogen peroxide (H2O2) stimulate mitogen-activated protein kinases (ERK1/2, p38MAPK, JNK, ERK5), tyrosine kinases (Src, JAK) and transcription factors (NFκB, AP-1, and HIF-1) and inactivate protein tyrosine phosphatizes (SHP1/2, MAPKP). ROS also increase [Ca2]i and upregulate protooncogene and proinflammatory gene expression and activity. These phenomena occur through oxidative modification of proteins by altering key amino acid residues (cysteine and methionine), by inducing protein dimerization, and by interacting with metal complexes such as Fe-S moieties [2]. Changes in the intracellular redox state through glutathione and thioredoxin systems also influence intracellular signaling events. Emerging evidence suggests that ROS play an important role in modulating autonomic balance. For example, whereas nitric oxide (NO) exerts a tonic inhibition of central sympathetic nervous system activity (SNS), increased production of O2– and ONOO (peroxynitrite) enhance inactivation of NO with resultant activation of the SNS (Fig. 69.1). Significantly, neuronal activity affecting cardiac function by NO is site-specific, since NO is inhibitory for the nucleus tractus solitarii and excitatory for the nucleus ambiguus and sinoatrial node SA node [3]. Reactive oxygen species contribute to cardiac sympathovagal imbalance in the brainstem, peripheral neurons and in cardiovascular cells [4]. A major source for neural ROS is a family of non-phagocytic NAD(P)H oxidases, including the prototypic Nox2 homolog-based NAD(P)H oxidase, as well as other NAD(P)H oxidases, such as Nox1 and Nox4 [5]. Other possible sources include mitochondrial electron transport enzymes, xanthine oxidase, cyclooxygenase, lipoxygenase and uncoupled nitric oxide synthase (NOS; eNOS, iNOS, nNOS).

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00069-X

nNOS activity

Nox activity

nNOS uncoupling

Mitochondria

NO/ O–2 production

Others sources

↑ ROS production

OONO– +

channel activity K Ca2+ channel activity

Parasympathetic nerve activity

Sympathetic nerve activity

FIGURE 69.1 Schematic demonstrating the interaction between the central nervous system and the autonomic nervous system with respect to reactive oxygen species biology. Stress responses in the neurons result in activation of ROS-generating enzymes and inhibition of neuronal NOS (nNOS), leading to reduced NO production and increased ROS generation. Uncoupling of nNOS and activation of neural Nox promotes increased O2 formation, which quenches NO to generate peroxynitrite (ONOO). Increased oxidative stress inhibits K channel activity and stimulates Ca2 channel activity resulting in increased activation of sympathetic nerves and depressed parasympathetic nerve activity.

BIOLOGY OF ROS Reactive oxygen species are produced as intermediates in reduction-oxidation (redox) reactions leading from O2 to H2O. The sequential univalent reduction of O2 is: e− e− e− e− O 2 →⋅ O− Of the ROS 2 → H 2 O 2 → OH ⋅ → H 2 O + O 2 . generated by cells of the nervous system, O2, and H2O2 appear to be particularly important [4,5]. In biological systems, O2 is short-lived owing to its rapid reduction to H2O2 by superoxide dismutase (SOD), of which there are three mammalian isoforms, copper/zinc SOD (SOD1),

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mitochondrial SOD (Mn SOD, SOD2), and extracellular SOD (EC-SOD, SOD3). The main source of H2O2 is the dismutation of O2: 2O2  2H → H2O2  O2. This reaction can be spontaneous or it can be catalyzed by SOD. O2 and H2O2 have distinct chemical properties [1,2].  O2 is electrically charged, highly reactive, cell membrane impermeable, rapidly dismutated into H2O2, its concentrations are in the picomolar–nanomolar range and as such O2 acts as an intracellular messenger. In contrast, H2O2 is less reactive, more stable, its concentrations are in the high nanomolar–micromolar range, it diffuses across hydrophobic membranes and hence H2O2 can act both as an intra- and inter-cellular messenger. As such different species of ROS may activate diverse signaling pathways, which lead to divergent, and potentially opposing, biological responses. For example, in the vascular system O2 levels inactivate the vasodilator NO leading to endothelial dysfunction and vasoconstriction, whereas H2O2, acts as a direct vasodilator in some vascular beds, including cerebral, coronary and mesenteric arteries.

PRODUCTION AND METABOLISM OF ROS ROS are produced by many cell types, including those of the nervous system, and can be formed by many enzymes. Enzymatic sources of ROS are xanthine oxidoreductase, uncoupled NO synthase (NOS, including eNOS, iNOS and nNOS), mitochondrial respiratory enzymes, and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase [6–8].

Xanthine Oxidase Xanthine oxidase (XO) and xanthine dehydrogenase (XDH) are interconvertible forms of the same enzyme, known as xanthine oxidoreductase. Physiologically, XO and XDH participate in many biochemical reactions, with the primary role being degradation of purines and the conversion of hypoxanthine to xanthine and xanthine to uric acid. As a byproduct in the purine degradation pathway, XO oxidizes NADH to form O2 and H2O2.

Uncoupled Nitric Oxide Synthase Under physiological conditions, nitric oxide synthase (NOS), in the presence of cofactors L-arginine and tetrahydrobiopterin (BH4), produces NO. In the absence of these cofactors, because of oxidative destruction or down regulation of GTP cyclohydrolase-1, which is the rate-limiting enzyme in BH4 production, uncoupled NOS produces O2 rather than NO. All three NOS isoforms are capable of “uncoupling” that leads to the preferential formation of O2. Whether effects of uncoupled NOS are due to increased O2 generation or to decreased NO bioavailability still remain unclear. Nevertheless BH4 has been suggested as a treatment modality to “recouple” NOS. While

previously difficult to use clinically because of chemical instability and cost, newer methods to synthesize stable BH4 suggest its novel potential as a therapeutic agent to ameliorate oxidative stress [8].

Mitochondrial Respiratory Enzymes Mitochondrial biogenesis is involved in the control of cell metabolism, signal transduction, and regulation of mitochondrial ROS production. More than 95% of O2 consumed by cells is reduced by four electrons to yield two molecules of H2O via mitochondrial electron transport chain complexes (I–IV), with 1–2% of the electron flow leaking onto O2 to form O2 under normoxic conditions. Mitochondrial ROS production is modulated by many factors including mitochondrial electron transport chain efficiency, mitochondrial antioxidant content, local oxygen, NO concentrations, availability of metabolic electron donors, uncoupling protein (UCP) activity, cytokines and vasoactive agonists. Alterations in mitochondrial biogenesis are associated with mitochondrial dysfunction and mitochondrial oxidative stress. Impaired activity and/or decreased expression of mitochondrial electron transport chain complexes I, III and IV have been implicated in vascular aging, cardiovascular disease and neurological disorders [9]. Mitochondrial dysfunction and mitochondrial–localized ROS production in the nervous system are important in cardiovascular function, as evidenced in rabbits with chronic heart failure, elevated mitochondrial superoxide levels in the carotid body contributes to enhanced chemoreceptor activity and peripheral chemoreflex function [10,11].

ROS-Generating Nox-Family NAD(P)H Oxidases Until recently mitochondria were considered to be the major source of ROS in the nervous system. However increasing evidence indicates that NADPH oxidases (Nox) may be important [5]. Neural Noxes have been implicated in cerebrovascular disease, microglial inflammation and neuronal death. NADPH oxidases were originally considered as enzymes expressed only in phagocytic cells involved in host defense and innate immunity. The mammalian Nox family comprises seven members, characterized primarily by the catalytic subunit that they utilize. These include Nox1, Nox2 (formerly gp91phox), Nox3, Nox4, Nox5, Duox1 and Duox2 [12]. The prototypical NAD(P)H oxidase is a multimeric enzyme found in phagocytes and comprises five subunits: p47phox (“phox” stands for phagocyte oxidase), p67phox, p40phox, p22phox, and the catalytic subunit gp91phox (also termed Nox2) [13]. Nox-family NAD(P)H oxidases are expressed in many tissues and mediate diverse biological functions. All Noxes are transmembrane proteins that transport electrons across biological membranes to reduce O2 to O2. Nox1 is found primarily in colon epithelial cells as well as in other cell types such as endothelial cells and vascular

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smooth muscle cells and is involved in host defense and cell growth. Nox2 is the catalytic subunit of the respiratory burst oxidase in phagocytes, but is also expressed in vascular, cardiac, renal and neural cells (neurons and glia). Nox3 is found in fetal tissue and the adult inner ear and is involved in vestibular function. Nox4, originally termed Renox (renal oxidase) because of its extensive abundance in the kidney, is also found in vascular cells, fibroblasts and osteoclasts. Nox4 produces mainly H2O2, while Nox1 generates mostly O2 that is subsequently converted to H2O2. The difference in the products generated by Nox1 and Nox4 may contribute to distinct roles of these Noxes in cell signaling. Nox5 is a Ca2-dependent homologue, found in testes and lymphoid tissue, but also in vascular cells. While all Nox proteins are present in rodents and man, the mouse and rat genome does not contain the nox5 gene. Unlike other Noxes, Nox5 possesses an amino-terminal calmodulin-like domain with four binding sites for Ca2 (EF hands) and does not require p22phox or other subunits for its activation. Nox5 is directly regulated by intracellular Ca2 ([Ca2]i), the binding of which induces a conformational change leading to enhanced ROS generation. Of the Noxes, Nox1–4 have been identified to be functionally active in the nervous system. To date nothing is known about Nox5 in the nervous system.

Regulation of Noxes All Noxes, except Nox5, appear to have an obligatory need for p22phox [11–13]. Whereas Nox2 requires p47phox and p67phox for its activity, Nox1 and Nox3 may interact with homologs of p47phox (NAD(P)H oxidase organizer 1 (NOXO1)) and p67phox (NAD(P)H oxidase activator 1 (NOXA1)). Oxidase activation involves Rac translocation, phosphorylation of p47phox and its translocation, possibly with p67phox, and p47phox association with cytochrome b558. Nox4 is constitutively active. However, induction of Nox mRNA expression is observed in response to physical stimuli, (shear stress, pressure), growth factors (platelet-derived growth factor, epidermal growth factor, and transforming growth factor β), cytokines (tumor necrosis factor-α, interleukin-1, and platelet aggregation factor), metabolic factors (hyperglycemia, hyperinsulinemia, free fatty acids, advanced glycation end products) and G protein-coupled receptor agonists (serotonin, thrombin, bradykinin, endothelin, and Ang II).

Protecting against Oxidative Stress – Antioxidant Defenses Enzymatic and nonenzymatic systems have evolved to protect against injurious oxidative stress. Major enzymatic antioxidants are SOD, catalase, glutathione peroxidases, thioredoxin and peroxiredoxin. Non-enzymatic antioxidants include ascorbate, tocopherols, glutathione, bilirubin and uric acid and scavenge OH·and other free radicals. SOD catalyzes the dismutation of O2 into H2O2

and O2. Glutathione peroxidase (GPX) reduces H2O2 and lipid peroxides to water and lipid alcohols, respectively, and in turn oxidizes glutathione to glutathione disulfide. Oxidized glutathione (GSSG) can be recycled by glutathione reductase to reduced GSH utilizing NADPH as a substrate or it can be exported from the cell via active transport by the multidrug resistance protein 1 (MRP1). Catalase is an intracellular antioxidant enzyme that is mainly located in cellular peroxisomes and catalyzes the reaction of H2O2 to water and O2. Catalase is very effective in high-level oxidative stress and protects cells from H2O2 produced within the cell. Thioredoxin reductase participates in thiol-dependent cellular reductive processes. Low antioxidant bioavailability promotes cellular oxidative stress and has been implicated in many diseases.

ROS AND AUTONOMIC OUTFLOW In physiological conditions, autonomic outflow is regulated by the balance between sympathoinhibitory effects of NO and the sympathostimulatory effects of O2. In pathological conditions, increased ROS generation, due in part to hyperactivation of NADPH oxidase and mitochondrial oxidases, stimulates central sympathetic outflow promoting sympathetic hyperactivity, an effect that is normalized by antioxidant therapy in experimental hypertension and cardiac failure [14,15]. Centrally produced ROS by NAD(P)H oxidase in the hypothalamic and circumventricular organs are implicated in central control of hypertension, in part through sympathetic outflow [16]. Injection of SOD into the rostral ventrolateral medulla (RVLM) decreased sympathetic nerve activity in swine and in rat experimental models of hypertension, RVLM ROS levels are increased [14,16]. Regulation of sympathetic nerve activity by central ROS involves Ang II/AT1R [16]. Rabbits with cardiac failure exhibit increased renal sympathetic nerve activity and arterial baroreflex function, effects associated with enhanced NADPH oxidasederived O2 production in the RVLM and upregulation of Ang II/AT1R. These effects were restored to near normal by tempol, the SOD mimetic, or by apocynin, NADPH oxidase inhibitor [14,16]. Mechanisms whereby O2 mediates an increase in sympathetic outflow by central neurons are unclear but activation of specific populations of neurons through alterations in potassium and calcium channels may be important. In addition, reductions in the sympathoinhibitory influence of NO because of increased scavenging of NO to form ONOO and a reduction in nNOS activity may contribute to sympathoexcitation [17].

CONCLUSIONS Reactive oxygen species play an important role in regulating autonomic balance. Oxidative stress is implicated in sympathetic hyperactivity, neuronal apoptosis and death.

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These findings have evoked considerable interest because of the possibilities that oxidative stress in the nervous system may contribute to disorders associated with autonomic dysfunction and to neurological and psychiatric diseases. Hence therapies targeted to decrease ROS generation and/or strategies to increase NO availability in the central and peripheral nervous system may be useful in minimizing neural injury and thereby prevent oxidative stress-related neurological diseases.

Acknowledgements Work from the author’s laboratory was supported by grants 44018 and 57886, both from the Canadian Institutes of Health Research.

References [1] Droge W. Free radicals in the physiological control of cell function. Physiol Rev 2002;82(1):47–95. [2] Kemp M, Go YM, Jones DP. Nonequilibrium thermodynamics of thiol/disulfide redox systems: a perspective on redox systems biology. Free Radic Biol Med 2008;44(6):921–37. [3] Paton JF, Kasparov S, Paterson DJ. Nitric oxide and autonomic control of heart rate: a question of specificity. Trends Neurosci 2002;25(12):626–31. [4] Campese VM, Ye S, Zhong H, Yanamadala V, Ye Z, Chiu J. Reactive oxygen species stimulate central and peripheral sympathetic nervous system activity. Am J Physiol Heart Circ Physiol 2004;287(2):H695–703. [5] Sorce S, Krause KH. NOX enzymes in the central nervous system: from signaling to disease. Antioxid Redox Signal 2009;11(10):2481–504. [6] Förstermann U. Oxidative stress in vascular disease: causes, defense mechanisms and potential therapies. Nat Clin Pract Cardiovasc Med 2008;5(6):338–49.

[7] Montezano AC, Burger D, Ceravolo GS, Yusuf H, Montero M, Touyz RM. Novel Nox homologues in the vasculature: focusing on Nox4 and Nox5. Clin Sci (Lond) 2011;120(4):131–41. [8] Porkert M, Sher S, Reddy U, Cheema F, Niessner C, Kolm P, et al. Tetrahydrobiopterin: a novel antihypertensive therapy. J Hum Hypertens. 2008;22(6):401–7. [9] Ungvari Z, Labinskyy N, Gupte S, Chander PN, Edwards JG, Csiszar A. Dysregulation of mitochondrial biogenesis in vascular endothelial and smooth muscle cells of aged rats. Am J Physiol Heart Circ Physiol 2008;294(5):H2121–H2128. [10] Nozoe M, Hirooka Y, Koga Y, Araki S, Konno S, Kishi T, et al. Mitochondria-derived reactive oxygen species mediate sympathoexcitation induced by angiotensin II in the rostral ventrolateral medulla. J Hypertens 2002;6(11):2176–84. [11] Ding Y, Li YL, Zimmerman MC, Schultz HD. Elevated mitochondrial superoxide contributes to enhanced chemoreflex in heart failure rabbits. Am J Physiol Regul Integr Comp Physiol 2010;298(2):R303–11. [12] Miller AA, Drummond GR, Sobey CG. Novel isoforms of NADPH-oxidase in cerebral vascular control. Pharmacol Ther 2006;111(3):928–48. [13] Griendling KK. NAD(P)H oxidases: new regulators of old functions. Antioxid Redox Signal 2006;8(9-10):1443–5. [14] Zucker IH. Novel mechanisms of sympathetic regulation in chronic heart failure. Hypertension 2006;48(6):1005–11. [15] Oliveira-Sales EB, Colombari DSA, Davisson RL, Kasparov S, Hirata EA, Campos RR, et al. Kidney-induced hypertension depends on superoxide signaling in the rostral ventrolateral medulla. Hypertension 2010;56:290–6. [16] Hirooka Y, Sagara Y, Kishi T, Sunagawa K. Oxidative stress and central cardiovascular regulation. Pathogenesis of hypertension and therapeutic aspects. Circ J 2010;74(5):827–35. [17] Danson EJ, Li D, Wang L, Dawson TA, Paterson DJ. Targeting cardiac sympatho-vagal imbalance using gene transfer of nitric oxide synthase. J Mol Cell Cardiol 2009;46(4):482–9.

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70 Neurally Mediated Syncope Satish R. Raj INTRODUCTION Syncope is a sudden, transient loss of consciousness with spontaneous recovery that is associated with a loss of postural tone. It is very common, with conservative estimates that 3% of the general population has experienced at least one syncopal spell, with a greater proportion of groups such as the elderly affected [1]. Syncope is responsible for over 1% of hospital admissions [1]. Syncope can have many possible causes, ranging from benign to life-threatening conditions. The common underlying mechanism of syncope is a transient decrease in cerebral perfusion. An excellent evidenced-based review and approach to the management of syncope can be found in the European Society of Cardiology Guideline on Syncope [2]. Neurally mediated syncope (or reflex fainting) is the most common cause of syncope, especially in those patients without evidence of structural heart disease. Neurally mediated syncope most commonly occurs following prolonged sitting or standing, although it can also occur with exercise (initiation or peak exercise) or with emotional/psychological triggers (e.g., phlebotomy).

PATHOPHYSIOLOGY OF NEURALLY MEDIATED SYNCOPE (NMS) The pathophysiology of neurally mediated syncope is not completely understood [3,4]. The most common explanation for NMS is known as the “Ventricular Hypothesis” (Fig. 70.1). This hypothesis argues that the initiating event is a pooling of blood in the legs (from prolonged sitting or standing) with a resultant reduction in venous return (preload) to the heart. The resultant decrease in blood pressure leads to a baroreceptor mediated increase in sympathetic tone. This increased sympathetic tone leads to both an increased chronotropic and inotropic effect. The vigorous contraction, in the setting of an underfilled ventricle, is thought to stimulate unmyelinated nerve fibers (“ventricular afferents”) in the left ventricle. This is then thought to trigger a reflex loss of sympathetic tone and an associated vagotonia (with resultant hypotension and/or bradycardia). There is also release of epinephrine from the adrenal gland that may also potentiate the hypotension. Recent physiological studies suggest that in some patients stroke

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00070-6

volume decreases acutely before fainting. These data suggest that acute venodilation may be as important (or more important) than acute vasodilation [5]. This putative pathophysiology provides a plausible explanation for the “postural prodrome” in many of the episodes of NMS. It also provides a rationale for the use of the tilt table test, which is now commonly employed to aid in the diagnosis of NMS. However, this hypothesis fails to explain the mode of NMS in those patients with an emotional or psychological trigger, and it fails to account for episodes of NMS among denervated patients post cardiac transplantation. Even among patients with postural NMS, there are some experimental observations that do not fit with this hypothesis.

DIAGNOSIS OF NMS The history and physical examination are at the heart of the diagnosis. A clinical diagnosis can be made with these alone in most cases. Much of the effort is focused on excluding more malignant causes of syncope. The history should focus on the circumstances surrounding the syncope, the associated symptoms before and after the event, and any collateral history from witnesses. The past medical history may contain evidence of structural heart disease

BRAIN SNS Withdrawal

↑ SNS Activation

HEART

↑ Vagal Tone

Vasodilation Venodilation

↑ EPI

Hypotension Bradycardia

ADRENAL Central Hypovolemia

SYNCOPE

FIGURE 70.1 The Ventricular Hypothesis of Neurally Mediated Syncope. SNS, sympathetic nervous system.

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70. NEURALLy MEDIATED SyNCOPE

and coexisting medical conditions, which both point away from NMS. Medications may provoke syncope, and a family history of sudden death may point to an arrhythmic cause. Historical features of NMS have been found to include a female gender, younger age, associated diaphoresis, nausea or palpitation, and post syncopal fatigue [1]. A long history of spells also suggests NMS. Recently, some historical features have been incorporated into a point score that can be used to distinguish between NMS and seizures [6], and between tilt positive and tilt test negative syncope in patients with structurally normal hearts [7]. The physical exam should focus on ruling out structural heart disease and focal neurologic lesions. The most useful exam maneuver is the carotid sinus massage. The current technique involves performing up to 10 seconds of massage to the carotid sinus (per side) in both the supine and upright posture, with a positive result requiring a drop in blood pressure or heart rate with an associated reproduction in presenting symptoms [1]. This procedure is associated with a low rate of neurologic complications.

TILT TABLE TESTING Tilt table testing has been widely used since the late 1980s. These tests subject patients to head-up tilt at angles of 60 to 80 degrees, and all aim to induce either syncope or intense presyncope, with a reproduction of presenting symptoms. Passive tilt tests simply use upright tilt for up to 45 minutes to induce vasovagal syncope (sensitivity ~40%, specificity ~90%) [1]. Provocative tilt tests use a simultaneous combination of orthostatic stress and drugs such as isoproterenol, nitroglycerin, or adenosine to provoke syncope with a slightly higher sensitivity, but reduced specificity. There is little agreement about the best protocol. Many physicians are more comfortable treating patients if a diagnosis can be established with a tilt table test. Recent studies with implantable loop recorders have called the value of tilt testing into question. The International Study on Syncope of Uncertain Etiology (ISSUE) investigators have recently reported that in the absence of significant structural heart disease, patients with tilt positive syncope and tilt negative syncope have similar patterns of recurrence (34% in each group over a follow-up of 3–15 months), with electrocardiographic recordings consistent with NMS [8]. Despite these recent data, tilt table testing can still be useful for the evaluation of recurrent NMS [9]. Tilt tests are contraindicated in patients with severe aortic or mitral stenosis, or critical coronary or cerebral artery stenosis.

NATURAL HISTORY OF NMS Most people with neurally mediated syncope faint only once, but for some patients it can be a recurring and troublesome disorder. Most patients do very well following

assessment, with only a 25–30% likelihood of syncope recurrence after tilt testing in patients who receive neither drugs nor a device [10]. The cause for this apparently great reduction in syncope frequency may be due to spontaneous remission, reassurance, or advice about the pathophysiology of syncope, and postural maneuvers to prevent syncope. However, past performance does predict future results; patients with a greater number of historical syncopal spells are more likely to faint in follow-up [7], and patients with recent syncope are more likely to faint again than those with only remote historical syncope [11]. The time to the first recurrence of syncope after tilt testing is a simple and individualized measure of eventual syncope frequency, as those patients who faint early after a tilt test tend to continue to faint often [12].

NMS TREATMENT Most patients should simply be reassured about the usual benign course of NMS and instructed to avoid those situations that precipitate fainting. The use of support stockings or increased salt intake may be helpful. They should be taught to recognize an impending faint and urged to lie down (or sit down if that is not possible) quickly. This will not be enough for some patients, and other treatment options may be necessary. Some drugs may be helpful [1]. Although salt replenishment has not been rigorously studied, it is commonly used because of its low side effect profile and probable efficacy. Fludrocortisone, a mineralocorticoid agonist, has previously only been studied in small trials with mixed results. A large multicenter trial placebo controlled trial of fludrocortisone for recurrent syncope has recently finished enrolling subjects, with results expected by the late 2011 [13]. Although a previously common therapy, betablockers were found to be ineffective in the Prevention of Syncope Trial (POST), but metoprolol did provide benefits to a prespecified older group of patients (age 42 years) [14]. The α1-agonist midodrine has been found to decrease the rate of recurrent syncope among frequent fainters [15], whereas another α1 agonist, etilefrine, was not found to be useful [16]. Further definitive trials of midodrine for neurally mediated syncope may be necessary. The selective serotonin reuptake inhibitor (SSRI) paroxetine has also been shown to be useful in one well-designed study [17]. Anxiety and reflex fainting can have a “feed-forward” relationship. Acute anxiety can certainly trigger a fainting reaction. This is most famous in the context of bloodinjury phobia, but it can also occur in response to other anxiety triggers. Even in the absence of a known anxiety disorder, frequent recurrent fainting can provoke a degree of anxiety related to the unpredictability of the syncope. Anxiolytics, including SSRIs, can be an important part of the management of these patients. Orthostatic (tilt) training has been suggested as an effective non-pharmacological therapy for patients with

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NMS TREATMENT

recurrent NMS [18]. This treatment involves having patients with neurally mediated syncope lean upright against a wall for 30 to 40 minutes, 1 to 2 times per day. While the initial results from this therapy are quite promising, this therapy has not been found to be effective in blinded randomized evaluations [19]. Permanent dual chamber pacemakers have been shown to be associated with a significant reduction in recurrent syncope among highly symptomatic patients with NMS in several unblinded randomized trials [1]. However, this therapy is both expensive and invasive (requiring a small surgical procedure). A more recent double blind placebo controlled trial found that the benefits of dual chamber pacing are much more modest than was originally thought [20]. It is possible that the problem is in the optimal selection of candidates for permanent pacing. Using an implanted loop recorder guided strategy, patients with spontaneous sinus pauses did better with permanent pacing than those patients without a spontaneous pause [21]. The ISSUE 3 investigators are currently testing this hypothesis in a randomized controlled trial [22]. At this time, permanent pacemaker implantation cannot be recommended as a “first line” therapy for NMS.

References [1] Raj SR, Sheldon RS. Syncope: investigation and treatment. Curr Cardiol Rep 2002;4:363–70. [2] Task Force for the Diagnosis and Management of Syncope, European Society of Cardiology (ESC), European Heart Rhythm Association (Heart Failure Association). (Heart Rhythm Society) Moya A, Sutton R, Ammirati F, Blanc JJ, Brignole M, Dahm JB, et al. Guidelines for the diagnosis and management of syncope (version 2009). Eur Heart J 2009;30:2631–71. [3] Mosqueda-Garcia R, Furlan R, Tank J, Fernandez-Violante R. The elusive pathophysiology of neurally mediated syncope. Circulation 2000;102:2898–906. [4] Raj SR. Is cardiac output the key to vasovagal syncope? A reevaluation of putative pathophysiology. Heart Rhythm 2008;5:1702–3. [5] Verheyden B, Liu J, van DN, Westerhof BE, Reybrouck T, Aubert AE, et al. Steep fall in cardiac output is main determinant of hypotension during drug-free and nitroglycerine-induced orthostatic vasovagal syncope. Heart Rhythm 2008;5:1695–701. [6] Sheldon R, Rose S, Ritchie D, Connolly SJ, Koshman ML, Lee MA, et al. Historical criteria that distinguish syncope from seizures. J Am Coll Cardiol 2002;40:142–8. [7] Sheldon R, Rose S, Flanagan P, Koshman ML, Killam S. Risk factors for syncope recurrence after a positive tilt-table test in patients with syncope. Circulation 1996;93:973–81. [8] Moya A, Brignole M, Menozzi C, Garcia-Civera R, Tognarini S, Mont L, International Study on Syncope of Uncertain Etiology Investigators Mechanism of syncope in patients with isolated syncope and in patients with tilt-positive syncope. Circulation 2001;104:1261–7.

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[9] Benditt DG, Ferguson DW, Grubb BP, Kapoor WN, Kugler J, Lerman BB, et al. Tilt table testing for assessing syncope. American College of Cardiology. J Am Coll Cardiol 1996;28:263–75. [10] Sheldon RS. Outcome of patients with neurally mediated syncope following tilt table testing. Cardiologia 1997;42:795–802. [11] Sumner GL, Rose MS, Koshman ML, Ritchie D, Sheldon RS, Prevention of Syncope Trial Investigators Recent history of vasovagal syncope in a young, referral-based population is a stronger predictor of recurrent syncope than lifetime syncope burden. J Cardiovasc Electrophysiol 2010;21:1375–80. [12] Malik P, Koshman ML, Sheldon R. Timing of first recurrence of syncope predicts syncopal frequency after a positive tilt table test result. J Am Coll Cardiol 1997;29:1284–9. [13] Raj SR, Rose S, Ritchie D, Sheldon RS, POST II, I. The Second Prevention of Syncope Trial (POST II) – a randomized clinical trial of fludrocortisone for the prevention of neurally mediated syncope: rationale and study design. Am Heart J 2006;151:1186–7. [14] Sheldon R, Connolly S, Rose S, Klingenheben T, Krahn A, Morillo C, Investigators POST Prevention of Syncope Trial (POST): a randomized, placebo-controlled study of metoprolol in the prevention of vasovagal syncope. Circulation 2006;113:1164–70. [15] Perez-Lugones A, Schweikert R, Pavia S, Sra J, Akhtar M, Jaeger F, et al. Usefulness of midodrine in patients with severely symptomatic neurocardiogenic syncope: a randomized control study. J Cardiovasc Electrophysiol 2001;12:935–8. [16] Raviele A, Brignole M, Sutton R, Alboni P, Giani P, Menozzi C, et al. Effect of etilefrine in preventing syncopal recurrence in patients with vasovagal syncope: a double-blind, randomized, placebocontrolled trial. The Vasovagal Syncope International Study. Circulation 1999;99:1452–7. [17] Di GE, Di IC, Sabatini P, Leonzio L, Barbone C, Barsotti A. Effects of paroxetine hydrochloride, a selective serotonin reuptake inhibitor, on refractory vasovagal syncope: a randomized, double-blind, placebo-controlled study. J Am Coll Cardiol 1999;33:1227–30. [18] Ector H, Reybrouck T, Heidbuchel H, Gewillig M, Van de Werf F. Tilt training: a new treatment for recurrent neurocardiogenic syncope and severe orthostatic intolerance. Pacing Clin Electrophysiol 1998;21:193–6. [19] Duygu H, Zoghi M, Turk U, Akyuz S, Ozerkan F, Akilli A, et al. The role of tilt training in preventing recurrent syncope in patients with vasovagal syncope: a prospective and randomized study. Pacing Clin Electrophysiol 2008;31:592–6. [20] Connolly SJ, Sheldon R, Thorpe KE, Roberts RS, Ellenbogen KA, Wilkoff BL, VPS II, I Pacemaker therapy for prevention of syncope in patients with recurrent severe vasovagal syncope: Second Vasovagal Pacemaker Study VPS II.: a randomized trial. JAMA 2003;289:2224–9. [21] Brignole M, Sutton R, Menozzi C, Garcia-Civera R, Moya A, Wieling W, International Study on Syncope of Uncertain Etiology Early application of an implantable loop recorder allows effective specific therapy in patients with recurrent suspected neurally mediated syncope. Eur Heart J 2006;27:1085–92. [22] Brignole M. International study on syncope of uncertain aetiology 3 (ISSUE 3): pacemaker therapy for patients with asystolic neurally-mediated syncope: rationale and study design. Europace 2007;9:25–30.

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71 Sympatho-Vagal Imbalance in Hypertension Guido Grassi, Gino Seravalle Autonomic control of the cardiovascular system undergoes profound changes in hypertension with resultant sympathetic activation and parasympathetic inhibition. Evidence has been provided that these autonomic alterations (i) are related to the severity of the hypertensive state, also participating in the development of organ damage; (ii) are potentiated when hypertension is complicated by cardiac, metabolic or renal disease; and (iii) can be favorably affected by non-pharmacological as well as pharmacological interventions (Fig. 71.1). This chapter will discuss these three above mentioned issues based on the results collected in the past few years by our group and others.

Evidence for Autonomic Dysfunction in Hypertension and its Role in Hypertension-Related Organ Damage Evidence collected throughout the years have shown that essential hypertension is characterized by a reduction in the inhibitory influence exerted by the vagus on the heart, with a resulting increase in resting heart rate. The sympathetic nervous system also participates in this heart rate increase, due to the tachycardic effects adrenergic neurotransmitters have on sinus node activity. Both vagal and sympathetic cardiovascular influences appear to be already altered in the pre-hypertensive stage and in borderline hypertension, i.e., in conditions in which blood pressure may be still in the normal or in the high–normal range [1,2]. Indeed, evidence exists that in both these two conditions sympathetic nerve traffic, as assessed via the microneurographic technique, appears to be already elevated. Conversely, in these pre-hypertensive stages baroreflex control of cardiac vagal drive, when assessed by evaluating the tachycardic and the bradycardic responses to arterial baroreceptor stimulation and deactivation (vasoactive drug infusion technique) shows a clear-cut reduction. The two above mentioned autonomic alterations appear to be a hallmark of established hypertension [2,3]. However, while the parasympathetic dysfunction remains stable in magnitude in conditions characterized

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00071-8

by more severe increases in blood pressure values, the sympathetic activation undergoes a progressive potentiation from the mild to the more severe hypertensive state [1,2] (Fig. 71.2, left panel). Interestingly, the main features of the adrenergic activation in hypertension are paralleled by a similar behavior of blood pressure variability, i.e., the spontaneous blood pressure oscillations which characterize the 24-hour blood pressure profile [4,5]. This has been taken as a demonstration, although of indirect nature, that sympathetic neural influences are responsible not only for the increase in absolute blood pressure values but also for their beat-to-beat changes during the day-time and the night-time period. Two further issues related to the autonomic alterations in hypertension deserve to be mentioned. First, a state of sympathetic hyperactivity is detectable not only in young and middle-aged hypertensives, but also in the elderly, even when the blood pressure increase selectively affects systolic blood pressure values [2]. Second, the hypertension-related increase in adrenergic drive appears to be (i) specific for some cardiovascular districts, such as the heart, the kidneys and the skeletal muscle vasculature [6]; and (ii) peculiar to the hypertensive state of essential nature [1]. This latter feature is in sharp contrast with the parasympathetic control of heart rate, which appears to be markedly deranged both in essential and in secondary hypertension, such as in primary hyperaldosteronism and in renovascular hypertension [3]. As mentioned above, growing evidence supports the notion that at least one of the components of the autonomic dysfunction described in hypertension, i.e., sympathetic overactivity, is involved in the pathophysiology of several metabolic and cardiovascular alterations frequently accompanying blood pressure elevation [2,6]. This is the case for insulin resistance, given the evidence that an increase in adrenergic drive may trigger the development of a hyperinsulinemic state [2]. This is also the case for the structural and functional alterations of the cardiovascular system known under the term of “target organ damage” [1,2]. Indeed evidence exists that an adrenergic overdrive, of greater magnitude than that seen in uncomplicated hypertension, can be detected in hypertensive patients

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71. SymPATHo-VAgAl ImbAlAnCE In HyPERTEnSIon

Autonomic dysfunction in hypertension

Vagal impairment

Sympathetic activation

• Unrelated to clinical severity

• Related to clinical severity

• Dependent on a baroreflex dysfunction

• Not dependent on a baroreflex dysfunction

• Co-responsible for the increase in heart rate

• Co-responsible for the increase in heart rate

• Potentially co-responsible for an increase in cardiovascular risk

• Probably responsible for TOD and hypertension development or progression

• Largely reversible by some anti-HT drugs

• Potentially co-responsible for an increase in cardiovascular risk • Partially reversible by some anti-HT drugs

FIGURE 71.1 Scheme illustrating the main features of the parasympathetic/sympathetic alterations in hypertension (HT). TOD: target organ damage. bs/100 hb 80

**

bs/100 hb 80 *

**

*

60

bs/100 hb 80 ** *

60

60

*

*

** 40

40

20

NT

MHT

SHT

20

40

NT

HT

O-HT

20

NT

HT

CHF -HT

FIGURE 71.2 Behavior of muscle sympathetic nerve traffic in (left) normotensive subjects (NT), mild (MHT) and severe (SHT) hypertensives; (middle) lean normotensive subjects, lean hypertensive (HT) and obese hypertensive (O-HT) patients and (right) normotensives (NT) and hypertensive patients with a normal (HT) or an altered (CHF-HT) cardiac function. Data are shown as means  SEM. Asterisks (* p 0.05 ** p 0.01) refer to the statistical significance between groups. Figure modified from data reviewed in Refs 1 and 2.

with left ventricular hypertrophy. Evidence also exists that another functional complication of the hypertensive state, i.e., the left ventricular diastolic dysfunction, is also characterized by a pronounced sympathetic overactivity [7]. Finally, data collected in hypertensive patients with an end-stage renal disease indicate that in this condition also sympathetic activation seen in the high blood pressure state undergoes a further potentiation [8]. Recently the same phenomenon has been described in hypertensive patients with very mild renal failure, thereby suggesting that also in the case of the hypertension-related renal damage, the adrenergic abnormalities probably participate not only at the progression but also at the initial development of the disease [9]. All together these data support the concept that cardiac and renal organ damage depends not only on hemodynamic factors, i.e., the blood pressure overload, but also on the neuroadrenergic abnormalities.

Autonomic Dysfunction in Hypertension Complicated by Other Disease A number of studies have recently addressed the issue related to the assessment of the autonomic profile in cardiovascular and metabolic disease that frequently coexist with the hypertensive state. The evidence available so far can be summarized as follows. First, when obesity (particularly of the visceral type) complicates hypertension, sympathetic activity undergoes a much greater increase compared to that seen in lean individuals (Fig. 71.2, central panel) [1]. Second, hypertensive heart failure patients also display a potentiated adrenergic overdrive as compared to individuals with a similar blood pressure elevation but a preserved cardiac function (Fig. 71.2, right panel) [1]. Finally, diabetic patients with hypertension show a greater adrenergic activity as compared to

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SymPATHo-VAgAl ImbAlAnCE In HyPERTEnSIon

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BOX 71.1

P R I O R I T I E S F O R F U T U R E I N V E S T I G AT I O N S I N AU T O N O M I C C O N T R O L O F C I R C U L AT I O N I N H Y P E RT E N S I O N l

l l l

Implementation of new techniques for assessing sympathetic function (i.e., microdialysis, imaging, etc.) Dynamic assessment of autonomic function during sleep Evaluation of genetic/autonomic interactions Assessment of the link between autonomic dysfunction and hypertension prognosis

age-matched diabetic normotensives [10]. Taken together these findings provide evidence in favor of the existence of a positive feedback relationship between sympathetic neural mechanisms, cardiovascular risk factors and disease. They also provide an important background for the finding that in several of the above mentioned clinical conditions, mortality rate is related to the development of life-threatening cardiac arrhythmias, at which occurrence the autonomic dysfunction described above certainly participates [1].

Effects of Therapeutic Interventions on Hypertension-Related Autonomic Dysfunction Given the relevance of autonomic dysfunction (and more specifically of adrenergic overdrive) to the development/progression of the hypertensive state as well as to the hypertension-related end-organ damage, sympathetic deactivation represents an important goal of the nonpharmacological as well as pharmacological interventions aimed at lowering elevated blood pressure values. As far as non-pharmacological interventions are concerned, there is overwhelming evidence demonstrating the sympathomodulatory effects of low-calorie dietary interventions and regular physical exercise programs. Since both the two procedures trigger clear-cut blood pressure lowering effects of magnitude often related to the degree of the sympathoinhibition, the hypothesis has been advanced that the antihypertensive effects of the two interventions are related to their sympathoinhibitory effects [2]. Conversely, an enhancement of the already elevated adrenergic drive has been reported during long-term and marked low sodium diet. This is presumably related to the fact that dietary sodium restriction elicits hyperinsulinemia and renin-angiotensin stimulation, i.e., two effects which promote sympathoexcitation and impair baroreflex control of both vagal and sympathetic drive [2]. Recently, two invasive procedures, i.e., implantation of a device capable of stimulating the carotid baroreceptor (and thus

l l

Greater focus on combo-drug treatment Investigation of new therapeutic procedures (catheterbased renal sympathetic denervation, implantable carotid sinus baroreflex activating system)

inhibiting sympathetic activity and enhancing baroreflex control of cardiac vagal drive) and renal sympathetic denervation throughout a catheter positioned in a renal artery and connected to a radiofrequency generator, have been successfully developed. Initial promising and exciting results have been reported in resistant hypertension, with documented evidence of a marked sympathoinhibitory effect which presumably plays a major role in the blood pressure lowering effects of the two interventions [1,2]. As far as the effects of antihypertensive drug treatment on autonomic cardiovascular function is concerned, there is evidence that some pharmacologic classes of antihypertensive drugs (such as beta-blockers, ace-inhibitors and angiotensin II receptor blockers) may elicit profound sympathoinhibitory effects, while other classes may leave unchanged (long-acting calcium antagonists), or even further increase (diuretics, short-acting calcium antagonists) the adrenergic cardiovascular drive [1,2]. Information on the effects of different antihypertensive drug combinations on autonomic cardiovascular function is scarce at present. This will certainly represent one of the major areas of research in the next few years in the field of the autonomic dysfunction in hypertension (Box 71.1).

References [1] Grassi G. Sympathetic neural activity in hypertension and related diseases. Am J Hypertens 2010;23:1052–60. [2] Grassi G. Assessment of sympathetic cardiovascular drive in human hypertension: achievements and perspectives. Hypertension 2009;54:690–7. [3] Palatini P, Julius S. The role of cardiac autonomic function in hypertension and cardiovascular disease. Curr Hypertens Rep 2009;11:199–205. [4] Grassi G, Seravalle G, Quarti-Trevano F, Dell'Oro R, Bombelli M, Cuspidi C, et al. Adrenergic, metabolic, and reflex abnormalities in reverse and extreme dipper hypertensives. Hypertension 2008;52:925–31. [5] Joyner MJ, Charkoudian N, Wallin BG. A sympathetic view of the sympathetic nervous system and human blood pressure regulation. Exp Physiol 2008;93:715–24.

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[6] Esler M. The 2009 Carl Ludwig Lecture: Pathophysiology of the human sympathetic nervous system in cardiovascular diseases: the transition from mechanisms to medical management. J Appl Physiol 2010;108:227–37. [7] Grassi G, Seravalle G, Quarti-Trevano F, Dell'Oro R, Arenare F, Spaziani D, et al. Sympathetic and baroreflex cardiovascular control in hypertension-related left ventricular dysfunction. Hypertension 2009;53:205–9. [8] Schlaich MP, Socratous F, Hennebry S, Eikelis N, Lambert EA, Straznicky N, et al. Sympathetic activation in chronic renal failure. J Am Soc Nephrol 2008;20:933–9.

[9] Grassi G, Trevano FQ, Seravalle G, Arenare F, Volpe M, Furiani S, et al. Early sympathetic activation in the initial clinical stages of chronic renal failure. Hypertension 2011;57:846–51. [10] Huggett RJ, Burns J, Mackintosh AF, Mary DA. Sympathetic neural activation in nondiabetic metabolic syndrome and its further augmentation by hypertension. Hypertension 2004;44:847–52.

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72 Baroreflex Failure Jens Jordan INTRODUCTION Baroreflexes have an important role in short term blood pressure regulation. Baroreflex mechanisms may also be involved in chronic blood pressure regulation. Blood pressure changes distend arteries, thus, eliciting carotid and aortic baroreceptor stretch. The electrical signal generated in carotid and aortic baroreceptors is conveyed to medullary brainstem nuclei via the glossopharyngeal and vagus nerves where it is integrated with input from other afferents and cortical input. Efferent parasympathetic and sympathetic activities are adjusted to compensate for the change in systemic blood pressure. Thus, baroreflexes attenuate excessive swings in blood pressure and thereby maintain blood flow, especially to the brain. Moreover, the vasculature is protected from large, potentially deleterious, blood pressure fluctuations. Bilateral damage to afferent baroreflex structures results in baroreflex failure. Any afferent arc structure including baroreceptors, afferent neurons transmitting information from baroreceptors, or afferent brainstem nuclei may be involved. In contrast, damage to the efferent part of the baroreflex causes autonomic failure (Table 72.1). Whether or not the afferent baroreflex input must be completely lost to develop baroreflex failure is not known. In most baroreflex failure patients, the lesion of the afferent arc of the baroreflex seems to be associated with damage to efferent neurons in the vagus nerve. The damage results in partial or complete parasympathetic denervation of the heart (“nonselective baroreflex failure”) (Fig. 72.1). In a minority of patients, efferent parasympathetic neurons are intact (“selective baroreflex failure”) (Fig. 72.1). There is a large body of literature on baroreflex function, both, in different animal species and in humans. However, the number of baroreflex failure patients reported in the literature is relatively small. The small number of reported cases may suggest that baroreflex failure is a rare condition. Perhaps, the probability to experience bilateral damage to afferent baroreflex structures is low. An alternative explanation is that many cases of baroreflex failure go undetected.

CAUSES OF BAROREFLEX FAILURE In most patients, the mechanism that led to bilateral afferent baroreflex interruption is suggested by the history.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00072-X

A common cause of baroreflex failure is extensive neck surgery and radiation therapy for cancers of the neck, which may damage baroreceptors and/or afferent baroreflex neurons. In some patients, the bilateral loss results from repeated trauma to the neck. For example, surgical resection of the glossopharyngeal nerve resulted in baroreflex failure in a patient who had previously sustained injury to the contralateral glossopharyngeal and vagus nerves. Another patient reported in the literature developed baroreflex failure after repeated surgery to the cervical spine and an auto accident. Baroreflex failure has also been described in patients with the familial paraganglioma syndrome. Central nervous tumors impinging on afferent baroreflex structures, such as cranial nerve neuromas, can also induce baroreflex failure. Bilateral damage to the nuclei of the solitary tract (NTS), the most important relay station for afferent autonomic input, is a rare cause of baroreflex failure. In a number of patients with typical signs and symptoms of baroreflex failure, no etiology could be documented.

CLINICAL PRESENTATION Most patients who are ultimately diagnosed with baroreflex failure are sent to tertiary care centers for the evaluation of volatile arterial hypertension. The hypertension can be sustained or episodic. Even with sustained hypertension, blood pressure is highly variable (Fig. 72.2). During hypertensive episodes, blood pressure recordings may be in the range of 170–280/110–135 mmHg. Hypertensive episodes are usually accompanied by tachycardia, a so called “tracking” of blood pressure and heart rate. Patients may experience sensations of warmth or flushing, palpitations, headache, and diaphoresis. The hypertensive episodes are triggered by factors such as psychological stress, physical exercise, and pain. A minority of patients presents with episodes of hypotension and bradycardia. Spontaneous hypotension and bradycardia is a feature of selective baroreflex failure. Hypotensive episodes can be observed when patients are resting and cortical input is diminished. With profound hypotension, patients may experience presyncopal symptoms. Frank syncope seems to be uncommon. Severe orthostatic hypotension is not a typical baroreflex

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TABLE 72.1 Distinction of Baroreflex Failure and Autonomic Failure Baroreflex Failure

Autonomic Failure

Labile hypertension



/

Orthostatic hypotension

/



Orthostatic hypertension





Supine hypertension

/–



Postprandial hypotension

/–



Episodic tachycardia





Bradycardic episodes

*

/–

Hypersensitivity to vasoactive drugs





*Bradycardia associated with hypotension is a typical feature of malignant vagotonia due to selective baroreflex failure.

FIGURE 72.1 Selective baroreflex failure (top) contrasted to nonselective baroreflex failure (bottom). Baroreflex afferents (BA) are damaged in selective and nonselective baroreflex failure patients. Efferent sympathetic (SNS) and parasympathetic nerves (PNS) are intact in selective baroreflex failure. In nonselective baroreflex failure, efferent parasympathetic nerves are at least in part damaged. From Jordan et al. [9].

FIGURE 72.2 Continuous blood pressure (BP) and heart rate (HR) recordings at rest in a baroreflex failure patient. There are large spontaneous oscillations of heart rate and blood pressure that parallel each other. The decreases in BP are very brisk. From Jordan et al. [9].

failure symptom. Indeed, some patients feature a marked increase in blood pressure with standing namely, orthostatic hypertension. Orthostatic hypotension may be observed in baroreflex failure patients who are volume depleted or treated with sympatholytic drugs. One possible explanation for the absence of severe orthostatic hypotension in most patients is sparing of cardiopulmonary stretch receptors. An alternative explanation is that other signals, such as visual and vestibular cues, are sufficient to increase sympathetic activity with assumption of the upright posture. The baroreflex buffers the effect of vasoactive medications. Therefore, baroreflex failure patients may experience severe hypotension after ingestion of standard doses of antihypertensive medications (e.g., vasodilators, diuretics, sympatholytic drugs). Medications increasing vascular tone can lead to dramatic blood pressure surges. Clinical observations suggest that baroreflex failure patients also feature emotional lability, particular during hypertensive episodes. However, this issue has not been systematically addressed. The onset of baroreflex failure can be very abrupt or more gradual. An abrupt onset of symptoms typically occurs in patients with baroreflex failure due to neck surgery. A more gradual onset of baroreflex failure has been observed in some patients who had undergone radiation therapy of the neck. The degree of hypertension seems to be different during the acute and the chronic phase of the disease. After acute interruption of afferent baroreflex input, blood pressure is particularly high (“Entzügelungshochdruck”). Apneic spells can be seen during the first 24 hours when carotid body input to the CNS is lost. In the more chronic phase, the average blood pressure tends to decrease. Yet, blood pressure remains highly variable. A similar time-effect has been observed in animal models of baroreflex failure. Hypertension with increased blood pressure variability is associated with increased cardiovascular risk. However,

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the long-term consequences of baroreflex failure have not been investigated. Case reports suggest that sympathetically mediated hypertension in the acute phase of the disorder can lead to cerebral hemorrhage, ischemic strokes, and stress-induced (Tako-Tsubo) cardiomyopathy.

DIAGNOSING BAROREFLEX FAILURE Baroreflex failure should be suspected in patients with volatile arterial hypertension. However, in the majority of patients, volatile arterial hypertension is not caused by baroreflex failure. Alternative causes of volatile hypertension, such as renovascular hypertension, should be considered first. In patients in whom labile hypertension develops immediately after neck surgery, arriving at the correct diagnosis may be straightforward. Patients in whom the loss of afferent baroreflex function develops more gradually over time as a consequence of radiation therapy or a neuropathy may be difficult to diagnose. Pheochromocytoma can sometimes mimic baroreflex failure. Hypertensive episodes, tachycardia, flushing, and impaired baroreflex function have been described in both conditions. Therefore, the differential diagnosis pheochromocytoma should be considered and ruled out by appropriate testing. Other entities from which baroreflex failure needs to be distinguished are panic attack, generalized anxiety disorder, hyperthyroidism, alcohol withdrawal, and drug abuse (e.g., amphetamines, cocaine). Hyperadrenergic orthostatic intolerance may also present with volatility of blood pressure but severe hypertension is uncommon. One patient reported in the literature who featured typical signs of baroreflex failure was later shown to have Munchhausen`s syndrome. Baroreflex failure is a rare cause of hypotension and bradycardia. Baroreflex failure patients exhibit a normal or exaggerated pressor response to psychological (e.g., mental arithmetic test) and physiological stimuli (e.g., cold pressor or handgrip testing). The pressor effect can be markedly prolonged in these patients. Baroreflex testing should be considered in patients with typical signs and symptoms of baroreflex failure. In patients with an atypical clinical presentation or a gradual onset of symptoms, more common entities should be ruled out before baroreflex testing is considered. The diagnostic abnormality in baroreflex failure patients is the absence of a bradycardic response to pressor agents or a tachycardic response to vasodilators (Figs 72.3 and 72.4). Normal subjects will decrease the heart rate 7–21 bpm in response to a phenylephrine dose raising systolic blood pressure 20 mmHg and will increase the heart rate 9–28 bpm in response to a nitroprusside dose that lowers blood pressure by 20 mmHg. In contrast, baroreflex failure patients did not alter their heart rate by more than 4 bpm with either maneuver. The loss of baroreflex blood pressure buffering in these patients is associated with an about 10–20 fold hypersensitivity to vasoactive medications (Fig. 72.3). Therefore, baroreflex testing

FIGURE 72.3 Original recordings of finger blood pressure (BP), heart rate (HR), and muscle sympathetic nerve activity (MSNA) in a baroreflex failure patient (case 1, see also Fig. 72.4). MSNA was substantially increased. A sympathetic discharge was observed with almost every heart beat. Intravenous bolus application of 50 µg phenylephrine at 0 seconds caused a rapid pressor response, reaching a maximum of 25 mmHg above baseline after 30 to 40 seconds. Normally, the phenylephrine pressor response is associated with a baroreflex-mediated decrease in heart rate and MSNA. The compensatory response was absent in this patient. From Heusser et al. [8].

FIGURE 72.4 Changes in the RR interval were plotted over changes in systolic blood pressure (SBP) during phenylephrine and nitroprusside application to obtain baroreflex heart rate curves in baroreflex failure patients and in a group of younger healthy subjects. The physiological baroreflex response is virtually abolished in both baroreflex failure patients. From Heusser et al. [8].

should be conducted starting with low doses of phenylephrine (e.g., 12.5 µg) and nitroprusside (e.g., 0.1 µg/ kg). The doses should be cautiously increased to obtain a change in systolic blood pressure of at least 20–25 mmHg. Baroreflex heart rate control can be assessed noninvasively using cross spectral analysis or the so called sequence method. These methods have not been evaluated in baroreflex failure patients and cannot be recommended

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TABLE 72.2 Treatment of Baroreflex Failure Blood Pressure Reduction

Blood pressure elevation Prevention of bradycardia/asystole

Clonidine Alpha-methyl-DOPA Guanethidine* Guanadrel* Diazepam Fludrocortisone Dietary salt Cardiac pacemaker

*These medications have been applied in the treatment of baroreflex failure but have been withdrawn in many countries.

as a diagnostic test in this setting. Whenever possible, we also assess baroreflex regulation of sympathetic nerve traffic, which is also impaired in baroreflex failure, using the microneurography technique (Fig. 72.3). However, recording of sympathetic activity is not widely available. Abnormal baroreflex tests alone are not sufficient to diagnose baroreflex failure. Absence of heart rate changes during baroreflex testing can also be observed in autonomic failure patients. Biochemical assessment of patients with baroreflex failure demonstrates surges of sympathetic activity associated with hypertensive episodes. Venous plasma norepinephrine concentrations as high as 2660 pg/ml have been reported. Conversely, levels drawn during normotensive periods may be within the normal range. Clonidine profoundly reduces blood pressure and plasma norepinephrine concentrations in baroreflex failure patients. Determination of the norepinephrine response to clonidine can be useful to differentiate baroreflex failure from pheochromocytoma.

TREATMENT Treating baroreflex failure patients is a challenge. The first step in the treatment of baroreflex failure is education of patients, family members, and referring physicians. It is particularly important to convey the information that many medications that do not elicit changes in blood pressure may have a dramatic effect in baroreflex failure patients. Medications that may change sympathetic activity or vascular tone, including a variety of over-the-counter drugs, have to be used with great caution. The main goals in treating baroreflex failure patients is to prevent extreme hypertension (Table 72.2). The pharmacological treatment of choice for the hypertension is clonidine. Clonidine can be given p.o. or applied as a skin patch. Clonidine decreases sympathetic activity, both, in the central nervous system and in the periphery. Moreover, clonidine causes mild sedation. These effects attenuate pressure surges. Alpha-methyl-DOPA can also be used in these patients but there are some concerns regarding possible hepatotoxicity. Newer centrally acting sympatholytic agents, such as moxonidine or rilmenidine, are reasonable therapeutic alternatives. Some baroreflex

failure patients do not tolerate centrally acting sympatholytic agents (e.g., exacerbation of depression). In these patients, peripherally acting sympatholytic agents, such as guanethidine and guanadrel, have been successfully prescribed. Unfortunately, these peripherally acting sympatholytic drugs have been taken off the market in many countries. Because the hypertension in baroreflex failure patients is often driven by cortical input, which is unopposed by the baroreflex, benzodiazepines elicit a reduction in blood pressure. Chronic treatment with relatively large doses of benzodiazepines can be used in selected patients. All the antihypertensive medications have to be taken very regularly even when blood pressure is relatively low. Discontinuation of the medication elicits a particularly severe rebound phenomenon in these patients. Some patients, in particular patients with selective baroreflex failure, experience hypotensive episodes. Sometimes, the hypotension is acutely exacerbated by the antihypertensive treatment. However, in the long term, prevention of hypertension may attenuate pressure induced volume loss through the kidney. Thus, effective control of the hypertension may improve hypotension. We encourage patients who experience hypotension on chronic antihypertensive treatment to increase their dietary salt intake. In selected patients, pharmacological treatment of the hypotension is required. Because of its long duration of action, fludrocortisone is a good choice for the treatment of hypotension in these patients. Other pressor agents should be used with great caution. Pacemakers are generally not indicated in persons with syncope. However, in a few patients with malignant vagotonia, hypotensive episodes may be accompanied by lifethreatening bradycardia and asystole. Implantation of a cardiac pacemaker may be required in selected patients.

Further Reading Aksamit TR, Floras JS, Victor RG, et al. Paroxysmal hypertension due to sinoaortic baroreceptor denervation in humans. Hypertension 1987;9:309–14. Berganzo K, Ciordia R, Gomez-Esteban JC, et al. Tako-tsubo cardiomyopathy in a patient with bilateral lesions in the dorsal medulla. Clin Auton Res 2010 October 21. Biaggioni I, Whetsell WO, Jobe J, et al. Baroreflex failure in a patient with central nervous system lesions involving the nucleus tractus solitarii. Hypertension 1994;23:491–5. Chan WS, Wei WI, Tse HF. “Malignant” baroreflex failure after surgical resection of carotid body tumor. Int J Cardiol 2007;118(3):e81–2. June 12. Fagius J, Wallin BG, Sundlof G, et al. Sympathetic outflow in man after anaesthesia of the glossopharyngeal and vagus nerves. Brain 1985;108:423–38. Ford FR. Fatal hypertensive crisis following denervation of the carotid sinus for the relief of repeated attacks of syncope. Johns Hopkins Med J 1956;100:14–16. Guasti L, Simoni C, Scamoni C, et al. Mixed cranial nerve neuroma revealing itself as baroreflex failure. Auton Neurosci 2006;130(1–2): 57–60. December 30. Heusser K, Tank J, Luft FC, Jordan J. Baroreflex failure. Hypertension 2005;45(5):834–9. May.

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TREATmEnT

Jordan J, Shannon JR, Black B, et al. Malignant vagotonia due to selective baroreflex failure. Hypertension 1997;30:1072–7. Kuchel O, Cusson JR, Larochelle P, et al. Posture- and emotion-induced severe hypertensive paroxysms with baroreceptor dysfunction. J Hypertens 1987;5:277–83. Lampen H, Kezdi P, Koppermann E, et al. Experimenteller Entzügelungshochdruck bei arterieller Hypertonie. Zeitschrift für Kreislaufforschung 1949;38:577–92.

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Phillips AM, Jardine DL, Parkin PJ, et al. Brain stem stroke causing baroreflex failure and paroxysmal hypertension. Stroke 2000;31:1997–2001. Robertson D, Hollister AS, Biaggioni I, et al. The diagnosis and treatment of baroreflex failure. N Engl J Med 1993;329:1449–55. Tellioglu T, Oates JA, Biaggioni I. Munchausen’s syndrome presenting as baroreflex failure. N Engl J Med 2000;343:581.

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73 Blood Pressure Variability Stanley Fernandez, Sirisha Srikakarlapudi, Joseph L. Izzo BP and blood flow patterns in humans are quite variable, allowing energy-efficient responses to diverse stimuli from outside (environmental) and inside (diurnal, postural, metabolic, emotional, etc.) the individual. Pressure– flow regulation is a major component of virtually all integrated physiologic responses and can be systemic or organ-selective. Usually, the most important factor in BP regulation is the level of outflow of the sympathetic nervous system (SNS), which affects immediate (seconds) and long-term (weeks to months) cardiovascular and BP responses. BP variation is the result of normal (and abnormal) discharges from CNS centers (e.g., posterior hypothalamus) but abnormalities of feedback mechanisms (e.g., baroreflexes) can also lead to clinical abnormalities.

PHYSIOLOGIC CONTROL OF SNS OUTFLOW

Given the wide range of responses observed in normal human physiology, it is difficult to identify discrete boundaries of abnormal BP variation. From a clinical perspective, such variation can confound the diagnosis and management of hypertension and may contribute directly to cardiovascular disease (CVD). Isolating the contribution of BP variability to CVD risk is also difficult, in major part to the fact the confounding caused by co-morbidities associated with abnormal BP variability. Well-established CVD risk factors such as advancing age, diabetes mellitus, dyslipidemia and hypertension are common in high risk patients, many of whom also have increased BP variability.

Respiratory Variation

Control of SNS outflow is generated from complex bi-directional signals among interconnected centers within the central nervous system (cortex, hypothalamus, hippocampus, basal ganglia, circumventricular organs, medulla, etc.) and peripheral tissues (heart, lungs, arteries, kidney, skeletal muscle, etc.). The rostral ventrolateral medulla (RVLM) is the final common efferent pathway for acute BP and blood flow increases, which derive from cardiac inotropic and chronotropic responses and peripheral arterial and venous constriction. There are CNS inputs from vestibular nuclei (postural adjustments), cortical centers (conscious behaviors), and from the amygdala, hippocampus, and posterior hypothalamus (behavior and stress). The RVLM receives signals from the nucleus tractus solitarius (NTS) that are both inhibitory (aortocarotid and cardiopulmonary baroreflexes) and stimulatory (renal and skeletal muscle metaboreceptors). Circulating hormones can influence circumventricular regions with no blood-brain barrier; an example is the stimulatory effect of angiotensin II area postrema, which disinhibits the NTS. Powerful central inhibition of SNS responses occurs via the parasympathetic nervous system, which is responsible for “vegetative” functions. With regard to clinical syndromes involving excessive BP variability, direct investigation about primary dysfunction of specific CNS centers is usually not available.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00073-1

CLINICAL PATTERNS OF BP VARIATION

During normal respiration, breathing is not usually coupled tightly with systemic BP, in part because variations in stroke volume are buffered by baroreflex adjustments as well as vascular damping mechanisms. When changes in intrapleural pressure are marked, as in acute asthma, respiratory variation of BP can exceed 10 mmHg (pulsus paradoxus). Slow deep breathing acutely enhances respiratory BP variation but training in “yoga breathing” or device-guided slow breathing tends to decrease systemic BP chronically. When breathing is grossly disordered, as in obstructive sleep apnea syndrome (OSA), episodes of hypoxia cause marked SNS stimulation, corresponding acute BP elevations (20 mmHg), and possibly chronic hypertension.

Diurnal Rhythms and Nocturnal change Clinically, BP variability is determined using 24-hour ambulatory BP monitoring (ABP). BP decreases by about 10 mmHg in most people with normal sleep patterns, a phenomenon called “nocturnal dipping”. Individuals with exaggerated nocturnal BP dipping have been reported to have asymptomatic cerebral injury in the form of increased white matter lesions in the basal ganglia region as assessed by magnetic resonance imaging. Toward the end of the sleep period (early morning hours), blood volume is highest of the day and there is marked activation

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of the sympathoadrenal and renin-angiotensin systems. The result is a “morning BP surge”, which corresponds to the diurnal peak of stroke and myocardial infarction; therapy with adrenergic inhibitors blunts or eliminates the diurnal pattern of CVD events. Loss of the night-time BP drop (“non-dipping pattern”) occurs in hyperaldosteronism, pheochromocytoma, various sodium overload states, and adolescents exposed to violence and is also associated with increased CVD risk.

Postural Adaptation Maintenance of adequate cerebral perfusion during upright posture requires an instantaneous integrated response of the heart and vasculature that is mediated by the SNS. When gravity causes pooling of blood in the legs and lower abdomen, venous return and cardiac preload are reduced, thereby diminishing cardiac stroke volume and arterial pulse pressure. Reduced cardiac and arterial loads activate cardiopulmonary and aortocarotid baroreflexes, which disinhibit the negative feedback of the nucleus tractus solitarius (NTS) on RVLM outflow, thereby increasing venous and arterial tone and heart rate. Orthostatic hypotension (OH) can be caused by disruption of afferent, central processing, and efferent mechanisms within this reflex arc. OH is a major disease risk factor, not only for falls and injury but for CVD mortality. Orthostatic hypertension is an uncommon condition in which there is an excessive SNS response to posture.

Food Intake and Postprandial Hypotension In normal individuals, meal consumption is associated with increased cardiac output to allow increased splanchnic blood flow, usually manifested by a small increase in heart rate and pulse pressure. In elderly individuals, postprandial hypotension may occur 1–2 hours after eating. This phenomenon is probably multi-factorial in most cases and may include relative volume depletion, baroreflex dysregulation, and diminished cardiac responsiveness. Post-prandial hypotension has been associated with the cerebral leukoaryiosis lesions similar to those observed in individuals with exaggerated nocturnal BP dipping.

Body Temperature Reducing body temperature can stimulate the SNS and increase systemic resistance and BP. Thermostatic reflexes are also triggered when body temperature is increased, allowing cutaneous vasodilation and sweating, a cholinergic sympathetic function.

Emotional States There are many CNS control centers involved in emotional responses but the posterior hypothalamus,

hippocampus, and amygdala are prominent; in general, cortical inputs are inhibitory on these centers. Emotions are highly subjective, variable, and extremely difficult to assess clinically. As a result, the impact of emotions on hypertension and CVD have probably been grossly underestimated. Stress is not always a negative emotion; how the person perceives the stimulus is extremely important in determining its effect on the cardiovascular system. If the individual is receptive and has adequate coping mechanisms, the stimulus is often pleasant, with increased blood flow (increased heart rate and pulse pressure) and decreased systemic vascular resistance; mean BP generally does not change. If the individual cannot cope with the stressor, there is net vasoconstriction as well as increased flow, and BP tends to increase. Stress management is also influenced by a person’s social support system. Anger, frustration, and cynicism almost always tend to increase BP, both acutely and chronically. It is also possible that endothelial dysfunction in such conditions as insulin resistance and dyslipidemia may contribute to excessive vasoconstrictive responses to physiologic stressors.

“White Coat” Syndromes Some individuals have substantial BP elevations uniquely in healthcare settings, whether mean BP at home is normal (white coat hypertension, WCH) or elevated (white coat effect); clinic measurements in some individuals can be 80 mmHg higher than home values. For the most part, the pattern of target organ damage in WCH is closer to normotensive individuals, especially if the BP is relatively low in both settings. When the home–office difference is more substantial, there may be greater morbidity or mortality later in life. WCH creates a diagnostic and therapeutic dilemma, including potential over-diagnosis and over-treatment. It is recognized as a cause of resistant hypertension but many WCH patients also report increased side effects of medications, perhaps due to episodic drug-induced hypotension. Most standard BP medications do little to blunt the home–office BP difference. Although poorly studied and not formally indicated for this purpose, anti-anxiety medications may reduce the magnitude of WCH.

Exercise Physical exercise requires a series of complex neurophysiologic and hemodynamic responses. Centrally regulated increased SNS outflow increases cardiac output and muscle blood flow; stimulatory metaboreceptors in skeletal muscle sustain the response. In patients who have exercise-induced hypertension (SBP 220 mmHg with modified Bruce protocol), there is an increased incidence of future hypertension and possibly CVD risk, even if resting BP is normal. Physical conditioning blunts

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ClInICAl PATTERns of BP VARIATIon

exercise-mediated increases in BP and also lowers resting BP and heart rate.

Smoking Each cigarette smoked raises BP by several mmHg, but the effect lasts only for a few minutes and does not cause hypertension per se. The effect is best demonstrated on ABP, where daytime but not night-time BP is elevated by a few mmHg in smokers compared to non-smokers.

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general anesthesia and using labetalol and other supplemental medications as needed. Excessive intraoperative BP variation (% of readings outside standard statistical confidence bands) has been associated with increased morbidity.

Baroreflex Disorders

Not all hypertensives (probably about 50%) experience BP increases when they consume a high-salt diet. In general, such individuals have low activity of the renin-angiotensin system but high resting SNS activity and diminished suppression of SNS activity during salt loading.

Carotid sinus hypersensitivity, which can cause dramatic BP variation, is discussed in detail elsewhere in this book and remains a substantial management challenge. Increased BP variability is more commonly related to arterial baroreflex blunting, especially in elderly individuals with longstanding hypertension and OH. Stiffening of the walls of the aorta and carotid artery is thought to reduce the function of the arterial mechanoreceptors; cardiopulmonary baroreflex blunting may play a role in excessive BP variability post-myocardial infarction.

Drug Effects

Further Reading

Drugs most likely to contribute to a postural BP drop are those that tend to reduce venous return to the heart: venodilators (alpha-beta blockers, alpha-blockers, nitrates, PDE-5 inhibitors) and loop diuretics. On ABP studies, antihypertensive drugs do not tend to have much effect on 24-hour BP variability, especially if the variation is expressed as coefficient of variation (standard deviation/ mean) but statin therapy has a weak effect on day–night BP difference. Increased 24-hour ABP variability (especially the morning BP surge) and within-visit BP variation are related to stroke risk and increased post-stroke diastolic BP variation predicts hemorrhagic transformation and increased 90-day mortality, perhaps due to impaired cerebral autoregulation. Retrospective analyses of the UK-TIA aspirin trial and the ASCOT-BPLA study found that increased visit-to-visit systolic BP variation was associated with higher risk of subsequent stroke and increased stroke mortality. The mechanism for this effect was postulated to be hypotensive episodes but the data could not fully prove this hypothesis. Calcium channel blockers (CCBs) may reduce visit-to-visit BP variability more than other therapeutic classes (diuretics, beta blockers, ACE inhibitors and angiotensin receptor blockers) and are most likely to prevent systolic BP values from exceeding 200 mmHg (diastolic BP 105 mmHg).

Chrousos GP. Stress and disorders of the stress system. Nature Rev Endocrinology 2009;5:374–81. Eguchi K, Ishikawa J, Hoshide S, Pickering TG, Schwartz JE, Shimada K, et al. Night time blood pressure variability is a strong predictor for cardiovascular events in patients with type 2 diabetes. Am J Hypertension 2009;22:46–51. Goldstein DS, Eisenhofer G. Sympathetic nervous system physiology and pathophysiology in coping with the environment Comprehensive Physiology, 2010;1:21–43. John Wiley & Sons, Inc. Izzo Jr. JL, Taylor AA. The sympathetic nervous system and baroreflexes in hypertension and hypotension. Curr Hypertension Reports 1999;1(3):254–63. Izzo Jr. JL. Hemodynamics of Hypertension. In: Hall JE, Lip GYH, editors. Comprehensive Hypertension. Philadelphia: Mosby-Elsevier; 2007. p. 123–34. Chapter 10. Izzo Jr. JL. Blood Pressure Variability and Reactivity. Chapter 56 in Hypertension Primer American Heart Association, Fourth Edition Philadelphia: Wolters-Kluwer/Lippincott, Williams & Wilkins; 2007. pp. 177–181. Parati G, Gavish B, Izzo Jr. JL. Respiration and Blood Pressure. Chapter 43 in Hypertension Primer American Heart Association, Fourth Edition Philadelphia: Wolters-Kluwer/Lippincott, Williams & Wilkins; 2007. pp. 136–139 Pierdomenico SD, Cuccurullo F. Prognostic value of white-coat and masked hypertension diagnosed by ambulatory monitoring in initially untreated subjects: an updated meta analysis. Am J Hypertension 2011;241:52–8. Rothwell PM, Howard SC, Dolan E, O'Brien E, Dobson JE, Dahlof B, et al. Prognostic significance of visit-to-visit variability, maximum systolic blood pressure, and episodic hypertension. Lancet 2010;375:895–905. Webb AJS, Fischer U, Mehta Z, Rothwell PM. Effects of antihypertensive-drug class on interindividual variation in blood pressure and risk of stroke: a systematic review and meta-analysis. Lancet 2010;375:906–15.

Salt-sensitivity

Intraoperative BP Variation Under most circumstances, BP is easily controlled in the operating room by regulating the type and depth of

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74 Obesity-Associated Hypertension Cyndya Shibao EPIDEMIOLOGY Obesity is a condition affecting more than 30% of the US population and is associated with increased mortality, mostly related to cardiovascular events. Approximately 300,000 deaths per year are attributable to this condition with a substantial reduction in life expectancy, by an estimated 5 to 20 years [1]. Epidemiological studies have documented a linear association between blood pressure and adiposity as measured by body mass index (BMI) in lean and obese populations. Indeed, blood pressure increases over time with weight gain. The prevalence of obesity and hypertension grow in parallel across different population cohorts. Obese individuals have 3.5-fold increased likelihood of having hypertension, and on the other hand, an estimated 60% to 70% of hypertension could be attributed to obesity. Obesity-associated hypertension, therefore, is an increasing medical problem, contributing to health care cost and reversing the gains achieved in the treatment of hypertension. In this context, it is important to understand the mechanisms underlying obesity-associated hypertension to better target treatment strategies. The focus of this chapter is to provide a brief overview of the role of the autonomic nervous system in the pathophysiology of this condition.

INCREASED SYMPATHETIC ACTIVATION IN OBESITY The sympathetic nervous system (SNS) plays a major role in the preservation of body homeostasis. Body weight is maintained by a tight balance between energy intake and energy expenditure. The SNS exerts control not only on the cardiovascular system but also in a number of metabolic processes including energy homeostasis; alterations in the activity of this system have been documented associated with the development of obesity. Two hypotheses concerning autonomic mechanisms and their role in obesity have been proposed. The first of these suggests that a reduction in sympathetic activity is the initial event that predisposes to obesity by reducing energy expenditure (MONA LISA, Most Obesities kNown Are

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Low In Sympathetic Activity). The second hypothesis, proposed by Landsberg, postulated that obesity produces a compensatory sympathetic activation, which contributes to the cardiovascular morbidity, i.e., hypertension associated with this condition [2]. These seemingly contradictory hypotheses are not necessarily mutually exclusive because obesity is likely to be a heterogeneous disorder. Nonetheless, there is growing evidence that sympathetic activity is indeed increased in most forms of obesity. Furthermore, sympathetic activation is not uniformly distributed throughout the body, but selectively increased in those vascular beds relevant to blood pressure regulation. Using regional norepinephrine spillover, e.g., Rumantir et al. [3] found that renal norepinephrine spillover was elevated (twice normal) in normotensive obese subjects, but cardiac norepinephrine spillover was reduced by approximately 50%, and whole body norepinephrine spillover was not significantly increased compared to lean controls. Notably, the presence of hypertension modified this profile, in that in obesity-associated hypertension there was elevation of both renal and cardiac norepinephrine spillover compared with normotensive obese individuals. Microneurography, which directly measures sympathetic traffic to skeletal muscle (MSNA), is an alternative approach to estimate sympathetic activation. MSNA may be particularly useful in studying the role of the sympathetic nervous system in obesity not only because skeletal muscle is an organ with important metabolic functions, but also because sympathetic activity to skeletal muscle adequately reflects vasoconstrictive sympathetic tone that is modulated by the baroreflex. This baroreflex-modulated sympathetic tone is central in the regulation of blood pressure. In this regard, MSNA is positively and linearly correlated with quantitative indices of obesity, be it BMI or fat mass [4] (Fig. 74.1). The cause of sympathetic activation in obesity is not fully known and may be multi-factorial. Potential culprits include the increase in insulin, leptin, non-esterified fatty acids and angiotensin II; the decrease of adiponectin; and the sleep apnea associated with obesity. All these potential mechanism can act in the central nervous system and stimulate sympathetic outflow which is selectively increased in organs relevant to blood pressure control as discussed previously.

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SYMPATHETIC ACTIVATION AND OBESITY IN MINORITY POPULATIONS The preponderance of the evidence strongly supports the presence of increased sympathetic activity in obesity and, therefore, would seemingly refute the MONA LISA hypothesis. It should be noted, however, that virtually all of these studies discussed previously were done in Caucasian populations, and may not be applicable to minorities. Indeed, studies in Pima Indians have shown that, even though sympathetic nerve activity is positively correlated with BMI, it is lower compared to Caucasians when matched for obesity. Pima Indians have a very high incidence of obesity, type 2 diabetes and increased morbidity associated with these conditions but, of interest, they do not have an increased incidence of hypertension. It is postulated that the lower sympathetic activation associated with obesity protects them from hypertension [5,6]. Therefore, the contribution of sympathetic activity to the relation between obesity and hypertension can differ between different racial groups.

FIGURE 74.1 Relation between muscle sympathetic nerve activity (MSNA) and fat mass (kg). MSNA recordings were obtained from normal volunteers with wide range of body weight.

The relationship between SNS activity and adiposity was previously studied in African-Americans, the racial group with the highest prevalence of obesity and hypertension. Abate et al. [7] showed a gender difference in the association between SNS activity and body weight. Black men have increased SNS activity independently of body weight. On the contrary in Black women, SNS activity increases with body weight, although this positive association is less strong compared with Caucasians women. There are important differences in body composition in African-American women that might contribute to these differences. Several studies have reported that for the same BMI, they have decreased levels of fat mass, particularly visceral fat mass. Of note, visceral fat mass correlates better with muscle sympathetic nerve activity than any other index of obesity [8]. Further studies are needed to clarify the mechanisms underlying this gender dimorphism and the role of body composition in the association between the SNS and body weight.

SYMPATHETIC ACTIVATION AND OBESITY-ASSOCIATED HYPERTENSION The causal-relationship between the increased sympathetic activation and obesity-associated hypertension was initially examined in animal models. Combined α- and βadrenergic blockade reduced arterial blood pressure to a much greater extend in obese than normal dogs. Likewise, treatment with the central-acting α2 agonist clonidine that reduces sympathetic outflow centrally prevents the development of obesity-associated hypertension in dogs fed with high-fat diet [9]. In humans, Wofford et al. [10] used combined α- and β-adrenergic blockade with doxazosin and atenolol and showed a greater decrease in blood pressure in obese compare with lean hypertensive subjects. Similarly, complete but transient autonomic withdrawal with the ganglionic blocker, trimethaphan, eliminated any association between blood pressure and BMI in 56 unselected subjects with wide range of body fatness (Fig. 74.2). In this study, systolic blood pressure decreased more during autonomic blockade in the obese-hypertensive

FIGURE 74.2 Relation between systolic blood pressure (SBP) and body mass index (BMI) in 56 subjects with wide range of body weight. (A) Left panel at baseline. (B) Right panel during autonomic withdrawal with the ganglionic blocker trimethaphan.

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group than in obese or lean normotensive subjects. These findings support the hypothesis that most of the increase in blood pressure observed in obese subjects was mediated by the autonomic nervous system [11].

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Resting energy expenditure, in contrast to blood pressure, remained significantly elevated during autonomic blockade (Fig. 74.3A). Further analyses showed that the increase in resting energy expenditure was likely attributable to the increase in fat free mass, i.e., muscle mass and organs, that usually accompanies obesity. In this study a 30-kg increase in fat mass was associated by a 12-kg increase in fat free mass. Fat free mass accounted for 83% of the variability in resting energy expenditure in the absence of autonomic influences (Fig. 74.3B). In summary, obesity-associated hypertension is an important problem in western societies where obesity is achieving epidemic proportions. The preponderance evidence supports the hypothesis that obesity is associated with increased SNS activity, at least in Caucasians. This increased SNS activation contributes to hypertension and does not provide any beneficial metabolic effect as previously thought.

References

FIGURE 74.3 Effect of autonomic withdrawal induced by trimethaphan on blood pressure, resting energy expenditure, and the relationship between REE and FFM. (A) Top shows SBP values, left side bars and REE, adjusted for FFM, right side bars at baseline. Bottom shows the changes in these parameters induced by trimethaphan. Both SBP and REE were significantly elevated in obese individuals. Trimethaphan “normalized” the elevated SBP but not REE in the obese group. (B) Linear regression analysis between RRE and FFM, before () and during (•) trimethaphan administration. At baseline, the linear correlation between FFM and REE was defined by the following equation: REE  324  23.65  FFM; r2  0.69. This correlation improved after ganglionic blockade so that FFM explained 83% of the variability in REE in this group, and the intercept was no longer different from 0 (REE  –3.7  28.53  FFM; r2  0.83). (Reprinted with permission from Wolters Kluwer Health/Lippincott, Williams & Wilkins.)

[1] Fontaine KR, Redden DT, Wang C, Westfall AO, Allison DB. Years of life lost due to obesity. JAMA January 8, 2003;289(2):187–93. [2] Landsberg L. Insulin-mediated sympathetic stimulation: role in the pathogenesis of obesity-related hypertension (or, how insulin affects blood pressure, and why). J Hypertens March 2001;19(3 Pt 2):523–8. [3] Rumantir MS, Vaz M, Jennings GL, Collier G, Kaye DM, Seals DR, et al. Neural mechanisms in human obesity-related hypertension. J Hypertens August 1999;17(8):1125–33. [4] Grassi G, Seravalle G, Cattaneo BM, Bolla GB, Lanfranchi CM, Giannattasio C, et al. Sympathetic activation in obese normotensive subjects. Hypertension April 1995;25(4 Pt 1):560–3. [5] Weyer C, Pratley RE, Snitker S, Spraul M, Ravussin E, Tataranni PA. Ethnic differences in insulinemia and sympathetic tone as links between obesity and blood pressure. Hypertension October 2000;36(4):531–7. [6] Vozarova B, Weyer C, Snitker S, Gautier JF, Cizza G, Chrousos G, et al. Effect of cortisol on muscle sympathetic nerve activity in Pima Indians and Caucasians. J Clin Endocrinol Metab July 2003;88(7):3218–26. [7] Abate NI, Mansour YH, Tuncel M, Arbique D, Chavoshan B, Kizilbash A, et al. Overweight and sympathetic overactivity in black Americans. Hypertension 2001;38:379–83. [8] Alvarez GE, Beske SD, Ballard TP, Davy KP. Sympathetic neural activation in visceral obesity. Circulation November 12 2002;106(20):2533–6. [9] Rocchini AP, Mao HZ, Babu K, Marker P, Rocchini AJ. Clonidine prevents insulin resistance and hypertension in obese dogs. Hypertension January 1999;33(1 Pt 2):548–53. [10] Wofford MR, Anderson Jr. DC, Brown CA, Jones DW, Miller ME, Hall JE. Antihypertensive effect of alpha- and beta-adrenergic blockade in obese and lean hypertensive subjects. Am J Hypertens July 2001;14(7 Pt 1):694–8. [11] Shibao C, Gamboa A, Diedrich A, Ertl AC, Chen KY, Byrne DW, et al. Autonomic contribution to blood pressure and metabolism in obesity. Hypertension January 2007;49(1):27–33.

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75 Orthostatic Hypertension David Robertson Orthostatic hypertension (OHT) is an increase in blood pressure on assumption of upright posture of 20/10 mmHg. It is often beneath the radar of physicians and nurses, because it is generally both unexpected and counterintuitive. On recheck it may sometimes regress toward the mean. But at other times the OHT can be dramatic and persistent. Nevertheless our knowledge about its causes and significance remains circumscribed. It is in many ways the last hemodynamic frontier. Recently, there has been increasing interest in OHT and its possible consequences on health. The spectrum of context and magnitude of OHT is very broad. In some cases it can be a dramatic reproducible 50 mmHg or more increase in blood pressure every time a patient stands. In other situations it may be more modest and only be an incidental physical finding. OHT is usually defined as an increase in blood pressure with upright posture or tilt, but precise criteria have not been established. Furthermore few studies have entailed direct measurement of blood pressure in people with OHT. Such measurements would more faithfully reflect intra-arterial pressure and would avoid the introduction of potential artifacts. Sphygmomanometers can underestimate blood pressure when the vasculature is perturbed by pressor reflexes, such as those engaged by upright posture, or if it is increased by pressor agents. Therefore, the actual magnitude of the blood pressure increase upon standing might be even larger than is generally reported in patients with OHT. OHT has been long recognized. Some of the most illuminating early studies were conducted by David H. P. Streeten. He noted that individuals with OHT had a greater decrease in cardiac output when upright, a greater venous pooling in the lower extremities, and a higher plasma norepinephrine level upon standing. He hypothesized that excessive venous pooling led to a decrease in cardiac output, and that the response to this was increased sympathetic activity and increased DBP. This hypothesis of excessive venous pooling upon standing and decreased cardiac output initially seems paradoxical. Why isn’t there reduced rather than increased blood pressure in this circumstance? However, support garments did in fact prevent the OHT during upright posture in Streeten’s subjects. Perhaps in the OHT patients,

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central sympathoexcitation is pathologically excessive. This process could occur in a setting of partial dysautonomia involving capacitance vessels or in individuals with pathological dysregulation in brainstem or higher centers engaged in autonomic control. Why some patients experience orthostatic hypotension and others OHT in this circumstance remains unclear. OHT occurs in some forms of autonomic dysfunction (Box 75.1). It occurs in more than 20% of patients with postural tachycardia syndrome (POTS) in our center. Shibao has reported that 38% of patients who met criteria for both POTS and disordered mast-cell activation had blood pressure elevation on standing. Interestingly, in this “crossover” group of patients, the OHT manifested as either a persistent hypertensive response to upright posture or sometimes as a hypertensive crisis with upright BP as high as 240/140. Patients presenting with acute baroreflex failure experience some of the highest blood pressures encountered in contemporary practice, sometimes with surges above 300 mmHg. In subsequent days and weeks after the acute event, the surges moderate, but substantial OHT may continue, though it usually declines somewhat with continued upright posture. In chronic baroreflex failure, labile blood pressure and heart rate track together in response to physical or psychological stress. In a final phase of baroreflex failure, usually months to years after onset, the orthostatic hypertension may be replaced by orthostatic hypotension as the dominant hemodynamic expression. In the rare syndrome of norepinephrine transporter deficiency, an increase in blood pressure and tachycardia with upright posture is seen, a finding which may also occur during therapy with norepinephrine transport antagonists or combined serotonin/norepinephrine reuptake inhibitors. But it is not only the dramatic and unusual case of OHT that may be significant. OHT and its potential clinical importance are being recognized in two groups of patients with hypertension. The first is elderly patients with essential hypertension. In one study, OHT occurred in 11% of 241 elderly Japanese patients with essential hypertension (defined as those whose SBP increased by 20 mmHg upon standing). In this study, the incidence of silent cerebrovascular infarction was higher in patients with OHT

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BOX 75.1

C O N D I T I O N S A S S O C I AT E D W I T H O RT H O S TAT I C H Y P E RT E N S I O N

Chronic primary conditions l l l l l

Potentially surgically-correctable conditions

Hypertension in elderly Essential hypertension (“extreme dippers”) Hypertension with orthostatic blood pooling Type 2 diabetes mellitus Peripheral neuropathy

l l l

Pharmacologically induced l

Dysautonomias l l l l l

Pheochromocytoma Renovascular hypertension Medullary vascular compression

l

Atomoxetine and other norepinephrine uptake blockers Yohimbine

Postural tachycardia syndrome Mast cell activation disorder Norepinephrine transporter deficiency Baroreflex failure (early) Central autonomic dysregulation

than in hypertensives without OHT. Notably, about the same proportion of subjects had orthostatic hypotension (23 of 241 patients). These patients were also at increased risk of silent cerebrovascular infarction. Some patients with essential hypertension and abnormal diurnal variation in blood pressure – the “extreme dipper” phenotype – exhibit OHT. These patients show a greater than normal decrease in pressure while sleeping. In one study examining the relationship between orthostatic hypertension and diurnal SBP variation, 72% of extreme dippers had OHT, compared with only 11% and 9% of dippers and nondippers, respectively. Extreme dippers have a higher prevalence (53%) of silent cerebrovascular infarction detected by MRI compared with dippers (29%). Moreover, extreme dippers are at increased risk for overt stroke and tend to have a poorer prognosis in the event of a stroke. OHT might have a role in the overall increased risk for stroke in these patients, as two-thirds of strokes in extreme dippers occur in the morning, a time when these patients experience a surge in blood pressure. Matsubayashi and colleagues reported similar findings relating to orthostatic blood pressure changes and central nervous system (CNS) changes in a study of 334 elderly Japanese subjects. In this study, 8.7% (29/334) of subjects exhibited orthostatic hypertension using the same definition as the aforementioned study, and 6% (20/334) of subjects exhibited orthostatic hypotension. Both orthostatic hypertensive (n  15) and orthostatic hypotensive (n  15) subjects had an increased prevalence of CNS lesions detectable by MRI compared with orthostatic normotensives (n  30).

In addition, scores on a number of cognitive and neurobehavioral metrics were lower in orthostatic hypertensives (n  29) and orthostatic hypotensives (n  20) than in orthostatic normotensives (n  285). An important distinction of this study is that the study population was a general sample of elderly Japanese subjects, of whom only approximately 50% were taking antihypertensive medications. Thus, orthostatic hypertension (and orthostatic hypotension) may be associated with cerebrovascular infarction and with measurable neurocognitive deficits independent of the presence of essential hypertension. Fan et al. in China have also suggested that orthostatic hypertension (OHT) is independently associated with target organ damage and stroke in hypertensive subjects, though this study was limited by its cross-sectional design. Yatsuya et al. in the Atherosclerosis Risk in Communities (ARIC) Study which had its baseline in 1987–89 found analogous results. The ARIC study assessed orthostatic BP change within 2 minutes after supine to standing, and the incidence of subsequent stroke through 2007 was examined. The investigators focused on 680 ischemic strokes, classified as lacunar (153), nonlacunar thrombotic (383), and cardioembolic (144) during a mean follow-up of 18.7 years. There was a greater incidence of lacunar stroke in both orthostatic hypotension and orthostatic hypertension. These meticulously collected observations demonstrate that OHT is associated with a greater effect on cardiovascular health than has previously been recognized. Perhaps the time has come to bring emerging investigative strategies to bear on this neglected phenomenon.

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Further Reading Eguchi K, Kario K, Hoshide S, Hoshide Y, Ishikawa J, Morinari M, et al. Greater change of orthostatic blood pressure is related to silent cerebral infarct and cardiac overload in hypertensive subjects. Hypertension Res 2004;27:235–41. Fan XH, Wang Y, Sun K, Zhang W, Wang H, Wu H, et al. Disorders of orthostatic blood pressure response are associated with cardiovascular disease and target organ damage in hypertensive patients. Am J Hypertens 2010;8:829–37. Fessel J, Robertson D. Orthostatic hypertension: When pressor reflexes overcompensate. Nature Clin Pract Nephrol 2006;2:424–9. Kario K, Eguchi K, Nakagawa Y, Motai K, Shimada K. Relationship between extreme dippers and orthostatic hypertension in elderly hypertensive patients. Hypertension 1998;31:77–82. Ketch T, Biaggioni I, Robertson R, Robertson D. Four faces of baroreflex failure: hypertensive crisis, volatile hypertension, orthostatic tachycardia, and malignant vagotonia. Circulation 2002;105:2518–23. Matsubayashi K, Okumiya K, Wada T, Osaki Y, Fujisawa M, Doi Y, et al. Postural dysregulation in systolic blood pressure is associated with worsened scoring on neurobehavioral function tests and leukoariosis in the older elderly living in a community. Stroke 1997;28:2169–73.

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Robertson D, DesJardin JA, Lichtenstein MJ. Distribution and observed associations of orthostatic blood pressure changes in elderly general medicine outpatients. Am J Med Sci 1998;315:287–95. Shannon JR, Flattem NL, Jordan J, Jacob G, Black BK, Biaggioni I, et al. Orthostatic intolerance and tachycardia associated with norepinephrine-transporter deficiency. N Engl J Med 2000;342:541–9. Shibao C, Arzubiaga C, Roberts 2nd LJ, Raj S, Black B, Harris P, et al. Hyperadrenergic postural tachycardia syndrome in mast cell activation disorders. Hypertension 2005;45:385–90. Streeten DH, Anderson Jr GH, Richardson R, Thomas FD. Abnormal orthostatic changes in blood pressure and heart rate in subjects with intact sympathetic nervous function: evidence for excessive venous pooling. J Lab Clin Med 1988;111:326–35. Streeten DH, Auchincloss Jr JH, Anderson Jr GH, Richardson RL, Thomas FD, Miller JW. Orthostatic hypertension: pathogenetic studies. Hypertension 1985;7:196–203. Yatsuya H, Folsom AR, Alonso A, Gottesman RF, Rose KM. Postural changes in blood pressure and incidence of ischemic stroke subtypes: the ARIC study. Hypertension 2011;57:167–73.

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76 Heart Failure John S. Floras Heart failure, a clinical syndrome in which the left ventricle is unable to meet the body’s metabolic requirements without increasing end-diastolic pressure, volume, or both, is accompanied by alterations in cardiovascular regulation by both the parasympathetic and sympathetic limbs of the autonomic nervous system. These disturbances, which intensify as heart failure progresses, play a major role in determining the advance of disease, the severity of symptoms, and the mode of death [1]. Of the methods available to categorize heart failure patients clinically and in research studies, the left ventricular ejection fraction (the ratio between stroke volume and end-diastolic volume, normal 0.55) has proven the most durable. Most studies of the autonomic nervous system have focused on heart failure patients with reduced systolic function. By contrast, the condition of heart failure with preserved ejection fraction, often a consequence of longstanding ventricular hypertrophy due to hypertension, has received less attention. In general, abnormalities of vagal and sympathetic tone exhibited by such patients are of less magnitude [2]. Until recently, the prevailing view was that heart failure caused by left ventricular systolic dysfunction is characterized by vagal withdrawal and generalized sympathetic activation, representing an integrated reflex response to alterations in cardiac and peripheral hemodynamics that is initially compensatory but subsequently pathological. At the bedside, the autonomic phenotype of most patients with chronic heart failure is that of an increased and relatively static heart rate, tachypnea, a narrow pulse pressure, venoconstriction, coolness of the extremities, and sodium and water retention. Plasma norepinephrine concentrations relate inversely to prognosis in cohort studies but their routine measurement is uncommon in practice, due to specificity concerns. To obtain a more detailed and precise understanding of the time course, magnitude, and organ-specific targets of these autonomic disturbances, and for mechanisms responsible for their occurrence in human heart failure, investigators in specialized research centers have applied radiotracer, microneurographic and spectral analytic methods [1,3]. Time and frequency domain estimates of tonic and reflex vagal heart rate modulation have demonstrated consistently loss of parasympathetic tone at the onset of

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heart failure. This diminution has been attributed to several mechanisms which intensify and interact as heart failure progress: attenuated arterial baroreceptor afferent input, angiotensin II-mediated central alterations in the generation of vagal motor tone, defective parasympathetic ganglionic neurotransmission, inhibition of acetylcholine release by norepinephrine stimulating pre-junctional α1-adrenoceptors on vagal nerve endings, and sinus node stretch by elevated atrial pressure. As heart failure advances, loss of sino-atrial responsiveness to neurallyreleased and circulating catecholamines blunt further heart rate variation, which becomes a marker of reduced life expectancy [3]. Studies in human heart failure reveal considerable inter-individual variability in the magnitude and organ targets of sympathetic activation that is independent of left ventricular ejection fraction [1]. Indeed the initial presumption that impaired systolic function, leading to a fall in cardiac output and systemic blood pressure triggers a generalized arterial baroreceptor-mediated elevation in central sympathetic outflow has not been supported by experimental evidence. Rather, in mild heart failure only cardiac norepinephrine spillover (CNES) is elevated. Increases in total body (TNES) or renal norepinephrine spillover (RNES), or muscle sympathetic nerve activity (MSNA) emerge as heart failure advances, but of these several targets, it is the ventricle that is exposed to the greatest relative increase in neurally released norepinephrine. In many patients treated with contemporary medical therapy, muscle sympathetic firing rate remains within the normal range, despite significant compromise of ventricular systolic function, unless co-morbidities associated independently with sympathetic activation are also present [1]. Thus, as illustrated in Figure 76.1, the processes responsible for the autonomic phenotype of heart failure are much more nuanced and personalized, i.e., patientspecific, than initially envisaged. Because heart failure was conceptualized initially as a primarily hemodynamic disorder, attention was directed first at potential baroreceptor reflex-mediated contributions to sympathetic activation and vagal withdrawal. Arterial baroreflex-mediated heart rate responses to druginduced blood pressure changes are clearly diminished in human heart failure, and because of this a parallel

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reduction in the reflex regulation of sympathetic outflow had been assumed. However, a series of human investigations involving norepinephrine spillover methods, sympathetic nerve recordings and blood pressure-MSNA transfer functions have demonstrated the arterial baroreflex regulation of efferent sympathetic outflow responds rapidly and appropriately to acute changes in blood pressure [1,3]. MSNA responses to perturbations in cardiac filling pressures (i.e. stimulation or unloading of “cardiopulmonary baroreceptor reflexes”) are attenuated. A cardiac specific sympathoexcitatory reflex, related directly to left atrial (and presumably left ventricular end-diastolic) pressure and without influence on TNES likely underlies the isolated increase in CSNE of mild heart failure [1]. It is now appreciated that a range of non-baroreceptor mediated mechanisms contribute importantly to the variation between patients in the magnitude and complexity of their sympathetic activation (Fig. 76.1). Few of these are engaged by or relate specifically to the hemodynamic derangements of heart failure.

Augmented peripheral chemosensitivity to hypoxia, present in perhaps half of patients with advanced congestive failure, is associated with higher plasma norepinephrine concentrations, loss of tonic and reflex heart rate modulation, oscillatory patterns in breathing, and ventricular arrhythmias; each of these abnormalities is exacerbated by exercise and anticipates premature death [1]. If chemosensitivity to hypercapnia is increased in addition, these autonomic disturbances are intensified and 4-year survival becomes half that of patients with normal chemosensitivity [4]. Myocardial ischemia can increase sympathetic outflow by stimulating coronary chemoreflexes, and myocardial infarction by destroying afferent nerves exerting inhibitory restraint on efferent nerve traffic; in a comparison involving heart failure patients with a mean ejection fraction of 22%, MSNA was significantly higher in those with ischemic than those with dilated cardiomyopathy [1]. Stimulation of pulmonary stretch receptors by lung inflation inhibits sympathetic outflow, whereas voluntary

FIGURE 76.1 Summary of mechanisms involved in the autonomic disturbances of heart failure. Input from arterial and cardiac baroreceptor, arterial chemoreceptor, pulmonary stretch receptor, muscle metabo- and mechano-receptor, and renal afferent nerves converge to modulate sympathetic outflow about a centrally-mediated set-point increase, involving an angiotensin II-AT1-NADPH-superoxide pathway. With systolic dysfunction, input effecting sympathoinhibition () arising from primarily ventricular mechanoreceptor nerve afferents diminishes (thin line) whereas efferent sympathetic nerve modulation by arterial baroreceptor and lung stretch reflexes is preserved. Inputs eliciting sympathoexcitation () arising from an atrial reflex activated by increases in cardiac filling pressure, from chemically sensitive ventricular afferents, triggered by ischemia, and from renal afferent nerves add to augmented sympathoexcitatory input from arterial chemoreceptors and from exercising skeletal muscle. The central set point for sympathetic outflow (downward arrow) is raised further by central chemoreceptor sensitization, by sleep apnea, and possibly by obesity. Efferent mechanisms for increased norepinephrine (NE) spillover include pre-junctional facilitation of its release, and impaired NE re-uptake. Mild heart failure is characterized by a selective increase in cardiac NE release and a reduction in tonic and reflex vagal heart rate modulation (thin line), whereas in advanced heart failure there is a generalized increase in sympathetic nerve traffic to the heart, adrenal, kidney, skeletal muscle, and other vascular beds (thick lines). Ach, acetylcholine; CNS, central nervous system; E, epinephrine. Adapted from Reference 1. Reproduced with the author’s permission.

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and involuntary apneas elicit intense clustered sympathetic nerve firing. Congested patients with increased lung water, who must generate higher tidal volume than healthy subjects to elicit inspiratory neural silence, manifest low tidal volumes and high respiratory frequency, each of which correlates with higher MSNA [1]. Reflexes arising from exercising muscle assume an important role in determining exercise capacity, which in drug-treated heart failure patients cannot be predicted by ejection fraction, even if the latter is profoundly compromised. In heart failure, unlike healthy individuals, there is an inverse relationship between predicted peak oxygen uptake during exercise and resting MSNA. Patients with preserved exercise capacity exhibit similar values for MSNA at rest as age-matched controls, but a clear difference emerges with exercise. The muscle metabo-reflex is activated by both isometric and intense isotonic handgrip and at lower workloads. Furthermore, this efferent sympathetic response to local accumulation of exercise metabolites is greater in those whose predicted exercise VO2 peak is impaired, compared with those in whom exercise capacity is preserved [1]. Other neural mechanisms arising from skeletal muscle, including a muscle mechanoreflex elicited by passive exercise, have the potential to increase sympathetic outflow [1]. As a consequence, plasma NE or MSNA values obtained in heart failure patients at rest will underestimate the adrenergic burden borne by ambulatory heart failure patients. Stimulation of renal afferent nerves elicits a potent reflex symaptho-excitatory response in chronic kidney failure evident also in patients with heart failure and reduced glomerular filtration; in a recent study, in which MSNA was significantly greater in patients with heart failure plus chronic renal failure as compared to those with preserved renal function, administration of 100% oxygen reduced selectively MSNA in the renal failure cohort, indicating an important interaction with the arterial chemoreflex [5]. Patient characteristics, including anemia [6], obesity with metabolic syndrome [7] and sleep apnea [8] also potentiate independently the sympathetic activation of heart failure. The higher MSNA of heart failure patients with anemia (but normal creatinine) can be suppressed by acute inhalation of 100% oxygen, indicating participation of heightened chemoreflex sensitivity in this neural disturbance [6]. Because of its high prevalence (50% of patients will have either obstructive or central sleep apnea) and its propensity to induce malignant ventricular arrhythmias [9], sleep apnea is a particularly important central cause of sympathetic excitation in heart failure, independent of and additive to the prevailing level of MSNA [8]. By silencing pulmonary stretch receptors and by stimulating peripheral and central chemoreceptors by hypoxia and hypercapnia, both forms of apnea elicit profound clustered increases in MSNA. The abrupt generation of negative intra-thoracic pressure by obstructive apnea increases left ventricular afterload, resulting in a fall in stroke volume and pulse

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pressure, and further baroreflex mediated reflex sympathetic activation. Termination of apnea by brief arousal from sleep causes a final surge in MSNA before breathing resumes. The after effects of sleep apnea in heart failure include a carry-over effect persisting after awakening. MSNA burst incidence is significantly greater in heart failure patients with obstructive or central apnea than without. Indeed, many drug treated heart failure patients without sleep apnea exhibit MSNA no higher than that of control subjects. Reduction of daytime MSNA, following abolition of obstructive apnea by chronic use of nocturnal positive airway pressure, to levels present in heart failure patients without sleep apnea argue for an additive effect of these two stimuli on sympathetic outflow [1,8]. Daytime sympathetic activation in central apnea is more complex, as it is driven to a considerable extent by greater disease severity [8]. The central sympathoexcitation induced by sleep apnea may be both causal and contributory to heart failure and its progression. However, this is likely only one of several mechanisms that act centrally to increase the set-point for the magnitude of efferent sympathetic nerve traffic, and to augment baroreceptor reflex gain. Increased central angiotensin II and aldosterone, implicated in these processes, generate reactive oxygen species that alter neuronal excitability by modulating ion channel function and inducing inflammation. In experimental models of heart failure angiotensin AT1 receptor and NAD(P)H oxidase subunit gene expression within key cardiovascular centers such as the rostral ventral lateral medulla and the paraventricular nucleus are increased [1]. In human heart failure, internal jugular spillover of catecholamines and their metabolites is increased, with brain norepinephrine turnover correlating positively with CNES [1,3]. Abnormalities in efferent cardiac sympathetic nerve function assume greater importance as heart failure advances. Reduced norepinephrine uptake-1 carrier density causes transcardiac myocardial extraction to fall and CNES to increase disproportionally to nerve firing rate and transmitter release, thus contributing to the selective augmentation of CNES of early heart failure. Recently, loss of normal cardiac sympathetic innervation as assessed by 123-iodine metaiodobenzylguanidine (MIBG) imaging (a marker of norepinephrine uptake) has been shown to predict malignant ventricular arrhythmias and cardiac death [10]. In addition to their relevance to the development and grounding of contemporary heart failure therapy, insights gained from the study of autonomic abnormalities in heart failure underscore the importance of more comprehensive characterization of the individual patient phenotype as a guide to personalized or tailored intervention, and suggest new therapeutic approaches for this all too common condition, which carries still persistently high rates of death, hospitalization and other morbidities. As examples, studies in deconditioned heart failure patients of the effects of training on sympathetic responses to muscle exercise, and large randomized clinical trials evaluating the impact of

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treating sleep apnea on survival and hospitalization are in progress. Demonstration of preserved arterial baroreflex modulation of sympathetic outflow provides a compelling physiological rationale to proceed now with heart failure trials involving electrical stimulation of carotid nerves. Percutaneous renal nerve ablation may benefit heart failure patients by attenuating or abolishing efferent renal nerve mediated renin release, sodium retention and vasoconstriction, and afferent renal nerve stimulated generalized sympathoexcitation. Targeting central processes involved in the generation of sympathetic outflow and in the compromise of efferent sympathetic and vagal neurotransmission, as well as the identification and validation of a robust and scalable biomarker for sympathetic activation in heart failure represent important future research topics.

References [1] Floras JS. Sympathetic nervous system activation in human heart failure. JACC 2009;54:375–85. [2] Hogg K, McMurray J. Neurohumoral pathways in heart failure with preserved systolic function. Prog Cardiovasc Dis 2005;47:357–66. [3] Floras JS. Alterations in the sympathetic and parasympathetic nervous system in heart failure. In: Mann D, editor. Heart failure: A companion to Braunwald’s heart disease (2nd Edition). Elsevier Saunder: St Louis 2011. p. 254–78. Chapter 16.

[4] Giannoni A, Emdin M, Bramanti F, Iudice G, Franci DP, Barsotti A, et al. Combined increased chemosensitivity to hypoxia and hypercapnia as a prognosticator in heart failure. J Am Coll Cardiol 2009;53:1975–80. [5] Despas F, Detis N, Dumonteil N, Labrunee M, Bellon B, Franchitto N, et al. Excessive sympathetic activation in heart failure with chronic renal failure: role of chemoreflex activation. J Hypertens 2009;27:1849–54. [6] Franchitto N, Despas F, Labrunée M, Roncalli J, Boveda S, Galinier M, et al. Tonic chemoreflex activation contributes to increased sympathetic nerve activity in heart failure-related anemia. Hypertens 2010;55:1012–7. [7] Grassi G, Seravalle G, Quarti-Trevano F, Scopelliti F, Dell’Oro R, Bolla G, et al. Excessive sympathetic activation in heart failure with obesity and metabolic syndrome. Hypertens 2007;49:535–41. [8] Floras JS. Should sleep apnoea be a specific target of therapy in chronic heart failure? Heart 2009;95:1041. [9] Bitter T, Westehelde N, Prinz C, Hossain MS, Vogt J, Langer C, et al. Cheynes-Stokes respiration and obstructive sleep apnoea are independent risk factors for malignant ventricular arrhythmias requiring appropriate cardioverter-defibillator therapies in patients with congestive heart failure. Eur Heart J 2011;32:61–74. [10] Boogers MJ, Borleffs CJ, Henneman MM, van Bommel RJ, van Ramshorst J, Boersma E, et al. Cardiac sympathetic denervation assessed with 123-iodine metaiodobenzylguanidine imaging predicts ventricular arrhythmias in implantable cardioverter-defibrillator patients. J Am Coll Cardiol 2010;55:2769–77.

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77 Stress Cardiomyopathy and Takotsubo Syndrome David Robertson, Kyoko Sato A link between stress and cardiovascular disorders has long been recognized. The fixed coronary artery atherosclerotic lesions which significantly obstruct blood flow are often a substrate for angina pectoris, with episodes classically brought on by acute emotion or physical exertion. Subsequently a second kind of angina pectoris emerged in which coronary artery spasm (Prinzmetal angina) was responsible for the myocardial ischemia. The vasotonic lesion tended to be brought on by stimuli that often were unrelated to emotion, but in some cases appeared to be due to sensitivity of the coronary vessels to cholinergic and possibly histaminergic stimulation, rather than sympathetic activation. Patients with coronary artery spasm often had frequent episodes of short-duration (minutes) ECG changes, which sometimes were asymptomatic for the first 2 minutes in spite of large ST segment elevation. Figure 77.1 is a tracing from a patient with 10–20 episodes of coronary artery spasm daily. Figure 77.2 is a tracing from a 51-year-old woman with frequent angina but no fixed lesions in the coronary arteries. A third phenomenon, championed for many years by clinicians like Martin Samuels, appeared to arise when a powerful stressor like subarachnoid hemorrhage or stroke or life-threatening fright acted on a susceptible heart, and caused pathological features of myocardial bridging, in which sudden death might occur. This was difficult to study, and was largely neglected as a research direction, and only began to be elucidated in recent years. A fourth entity, likely related in some cases to the third phenomenon, began with studies in Japan around 1990 when patients were observed with chest discomfort and evidence of associated cardiomyopathy which developed in a setting of emotional or physical stress (“stress” or “Takotsubo” cardiomyopathy). Takotsubo cardiomyopathy was named because of the resemblance of the pathologic myocardial wall image to an octopus trap (tako tsubo in Japanese). Stress cardiomyopathy (SCM) is often associated with left ventricular ballooning, and presents with features that initially can be mistaken for acute myocardial infarction, but fixed epicardial coronary disease is absent. The

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00077-9

incidence of SCM is estimated to be 0.3–6.3% (mean 2.3%) of emergency coronary angiograms of patients with suspected acute myocardial infarction. Most patients with SCM are postmenopausal women averaging 65 years of age. Several reports raise the possibility of a genetic predisposition. SCM is often reported in the setting of emotional and/or physiological stress like the perioperative period or natural disasters. Prediction and prevention of SCM in the vulnerable population are important because accessing intensive medical care is limited during the post-disaster recovery period. For instance, SCM occurred 24 times more frequently on the day of Mid Niigata earthquake. However, in spite of the dramatic stressors that are typically reported, many patients with an otherwise typical presentation do not exhibit a prominent history of emotional or physical stress. Moreover, while the great majority of patients presenting with SCM gradually recover over days to weeks, a significant minority, perhaps 10% do have an additional episode after an interval of months to many years. Criteria for diagnosis of SCM include (i) transient hypokinesis, akinesis, or dyskinesis in the left ventricular mid segment with or without apical involvement; regional wall abnormalities that extend beyond a single epicardial vascular distribution; and frequently, but not always, a stressful trigger; (ii) the absence of obstructive coronary disease or angiographic evidence of acute plaque rupture; (iii) new ECG abnormalities (ST-segment elevation and/or T-wave inversion) or modest elevation in cardiac troponin; and (iv) the absence of pheochromocytoma and myocarditis. The ECG abnormalities in SCM are often dramatic, as shown in Figure 77.3. There is typically ST-segment elevation early on, and evolutionary symmetrical T-wave inversions in the precordial leads hours to days later. Left ventricular function is usually normal in 1–3 months. Complications once patients are hospitalized are uncommon but include atrial and ventricular arrhythmias, blood pressure instability, and rarely heart failure. When LV outflow is compromised by the constellation of apical and mid-ventricular wall motion abnormalities, intravenous fluid and beta blockade may reduce contractility and attenuate these symptoms. Rarely, death from rupture

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BOX 77.1

C AU S E S O F S T R E S S C A R D I O M Y O P A T H Y l l l l l

Emotional stress Intracranial bleed Stroke Head trauma Acute medical illness

l l l l

Sepsis Surgery Pheochromocytoma Hyperadrenergic postural tachycardia syndrome

BOX 77.2

P R E C I P I TA N T S O F S T R E S S C A R D I O M Y O PAT H Y l l l l

Acute severe emotional stress Death of a family member or pet Public speaking Altercations

l l l l

Job loss Physical injury Earthquakes Exhausting work

FIGURE 77.1 Coronary artery spasm. A 61-year-old man with normal coronary anatomy on catheterization. Over three weeks of monitoring he had more than 500 episodes of coronary spasm. Note that in a period of 20 minutes of quiet rest, he experienced two episodes of ST elevation, only the second of which he sensed as discomfort. The upper register shows the compressed ECG lead II over a 20-minute period. The three insets below show the ST segment elevation in single complexes during the interval above. The middle register shows the arterial pressure which changes little in the tracing. Likewise the lower register shows only modest changes in heart rate during this period.

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FIGURE 77.2 Coronary artery spasm. A 51-year-old woman with frequent angina but no fixed lesions in the coronary arteries. The upper register shows the compressed ECG lead II over a 28-minute period. The three insets show the ST segment elevation in single complexes during the associated interval. The middle register shows the arterial pressure. This episode was associated with chest pain and following 0.4 mg nitroglycerin, given at the arrows in the right panel, the patient experienced rapid resolution of angina and had what appeared to be a Bezold–Jarisch-like response, with hypotension (register 2), bradycardia (register 3), and at the right side of the ECG tracing in register 1, nausea and vomiting with ECG artifact in the extreme right as she sat up to reach for an emesis basin. She ultimately had resolution of her episodes.

of the left ventricular wall has been reported. The risk of mortality in SCM is in the 1–3% range, with supportive care generally sufficient since left ventricular function recovers fairly early. Several potential causes of SCM have been proposed such as coronary artery spasm, endothelial dysfunction, and catecholamine cardiotoxicity. Usually the pattern of the cardiomyopathy does not fit the distribution of a single coronary artery however, and the time course (compare Figs 77.1 and 77.2) is quite distinct. It could also be that disturbance in the cardiac microcirculation rather than large coronary arteries might occur. Occasional patients with SCM have had myocardial biopsy, which fairly consistently showed interstitial infiltrates of mononuclear lymphocytes, leukocytes and macrophages;

myocardial fibrosis; and contraction bands with or without myocardial necrosis. Might sympathetic activation underlie this process? That has been supported by some reports that plasma catecholamines are raised during SCM. In patients with the distinct disease coronary artery spasm, we have not found plasma catecholamines to be raised either before or during anginal attacks, but did not sample from the coronary sinus. Nevertheless altered autonomic nervous regulation, and in consequence, overexcretion of catecholamines may play an important role in triggering SCM. The careful report by Akashi et al. (2008) provides a thoughtful range of hypotheses, including the possibility that the preponderance of SCM cases in postmenopausal women might reflect a susceptibility engendered

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adenyl cyclase and activate protein kinase A (PKA), that in turn phosphorylates several downstream intracellular targets, resulting in an increased contractile response. Epinephrine (Epi) also binds β1ARs and activates this response, but it has a higher affinity for the β2 adrenoceptor (β2AR). At epinephrine concentrations in the normal physiological range, Epi binding to β2ARs activates the Gs protein–adenyl cyclase–PKA pathway, resulting in a positive inotropic response. At high concentrations Epi, but not NE, might induce coupling of the β2ARs to switch from Gs protein signaling to Gi protein signaling which might elicit a negative inotropic effect. The density of sympathetic nerve endings in human heart is approximately 40% higher in the basal myocardium than in the apical areas. This sympathetic innervation distribution might cause a stronger sympathetic inotropic effect in the basal myocardium and elevate the left ventricle outflow pressure gradient resulting in hemodynamic instability. As there is significant circulating Epi, the apical area might be more sensitive to excess Epi which could contribute to the apical ballooning and diffuse akinesis. Taken together, β2 ARs and their genetic diversity could have a substantial role in the pathogenesis of SCM. However, it seems likely that other, not yet discovered susceptibility mechanisms, are more likely to underlie the propensity of SCM to occur in some but not most individuals.

Further Reading

FIGURE 77.3 ECG in Takotsubo stress cardiomyopathy. (A) Initial ECG in a 65-year-old woman who developed Takotsubo stress cardiomyopathy immediately after an uncomplicated laparoscopic cholecystectomy. Chest pain and hypotension was noted in the recovery room together with ST segment elevation in leads V2–V6, I, aVL, and II. (B) ECG 24 hours later showing typical symmetrical T-wave inversions in the precordial leads. (C) Emergently performed coronary arteriogram documented normal vessels. (D) End-systolic phase left ventriculogram of the patient demonstrating the typical apical and mid-ventricular wall motion abnormalities of Takotsubo syndrome. The end-diastolic phase left ventriculogram is also shown. Echo (not shown) of the patient 1 month later demonstrated normalization of LV wall motion. Figure courtesy of Bybee and Prasad, Circulation 2008:118;397–409.

by altered estrogen levels. At physiological and elevated concentrations, norepinephrine (NE) released from the sympathetic nerves acts predominantly via the β1 adrenoceptors (β1ARs) on ventricular cardiomyocytes, exerting positive inotropic and lusitropic responses. These effects are the result of β1AR coupling to the Gs protein family, which increases intracellular cyclic AMP levels through

Akashi YJ, Goldstein DS, Barbaro G, Ueyama T. Takotsubo cardiomyopathy: a new form of acute, reversible heart failure. Circulation 2008;118(25):2754–62. Akashi YJ, Nakazawa K, Sakakibara M, Miyake F, Koike H, Sasaka K. The clinical features of takotsubo cardiomyopathy. QJM – An Int J Med 2003;96(8):563–73. Bybee KA, Prasad A. Stress-related cardiomyopathy syndromes. Circulation 2008;118(4):397–409. Christoph M, Ebner B, Stolte D, Ibrahim K, Kolschmann S, Strasser RH, et al. Broken heart syndrome: Tako Tsubo cardiomyopathy associated with an overdose of the serotonin-norepinephrine reuptake inhibitor Venlafaxine. Eur Neuropsychopharmacol 2010;20(8):594–7. Galiuto L, De Caterina AR, Porfidia A, Paraggio L, Barchetta S, Locorotondo G, et al. Reversible coronary microvascular dysfunction: a common pathogenetic mechanism in Apical Ballooning or Tako-Tsubo Syndrome. Eur Heart J 2010;31(11):1319–27. Gianni M, Dentali F, Grandi AM, Sumner G, Hiralal R, Lonn E. Apical ballooning syndrome or takotsubo cardiomyopathy: a systematic review. Eur Heart J 2006;27(13):1523–9. Izumi K, Tada S, Yamada T. A case of takotsubo cardiomyopathy complicated by ventricular septal perforation. Circ J 2008;72(9):1540–3. Lopez PR, Peiris AN. Kounis syndrome. South Med J 2010;103(11):1148–55. Madhavan M, Prasad A. Proposed Mayo Clinic criteria for the diagnosis of Tako-Tsubo cardiomyopathy and long-term prognosis. Herz 2010;35(4):240–3. Prasad A, Madhavan M, Chareonthaitawee P. Cardiac sympathetic activity in stress-induced (Takotsubo) cardiomyopathy. Nat Rev Cardiol 2009;6(6):430–4. Robertson D, Hollister AS, Forman MB, Robertson RM. Reflexes unique to myocardial ischemia and infarction. J Am Coll Cardiol 1985;5(6):B99–B104.

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Robertson D, Robertson RM, Nies AS, Oates JA, Friesinger GC. Variant angina-pectoris – investigation of indexes of sympathetic nervoussystem function. Am J Cardiol 1979;43(6):1080–5. Saito Y. Hypoglycemic attack: A rare triggering factor for takotsubo cardiomyopathy. Intern Med 2005;44(3):171–2. Samuels MA. ‘Voodoo’ death revisited: The modern lessons of neurocardiology. Cleve Clin J Med 2007;74:suppl:S1–16. Sato M, Fujita S, Saito A, Ikeda Y, Kitazawa H, Takahashi M, et al. Increased incidence of transient left ventricular apical ballooning (socalled “takotsubo” cardiomyopathy) after the mid-Niigata prefecture earthquake. Circ J 2006;70:947–53. Schneider B, Athanasiadis A, Schwab J, Pistner W, von Scheidt W, Gottwald U, et al. Clinical spectrum of tako-tsubo cardiomyopathy in

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Germany: results of the tako-tsubo registry of the Arbeitsgemeinschaft Leitende Kardiologische Krankenhausarzte (ALKK). Dtsch Med Wochenschr 2010;135(39):1908–13. Smyth BA, Clayton EW, Robertson D. Experimental arrest of cerebral blood flow in human subjects: The Red Wing studies revisited. Perspect Biol Med 2011;54:121–31. Wittstein IS, Kapur NK, Mudd JO, Champion HC. Neurohormonal features of stress-induced (Takotsubo) cardiomyopathy versus acute myocardial infarction. J Am Coll Cardiol 2007;49(9):96A–7A.

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78 Clinical Evaluation of Autonomic Disorders Paola Sandroni What makes autonomic disorders fascinating, also makes them challenging. The spectrum of symptoms and signs is probably unparalleled in any other medical specialty. In fact, autonomic disorders span across many medical specialties and to properly characterize and manage them one often has to involve multiple specialists working as a team. As always, the first step consists of obtaining a detailed history, which must be systematic and cover each system (i.e., cardiovascular, gastrointestinal etc.) [1]. Obviously, not every “autonomic” symptom necessarily means an autonomic disorder or dysfunction is present. As it is intrinsic in its own function, the autonomic system “gets involved” any time there is a perturbation in bodily function, due to internal (i.e., emotional state, other organ pathology) or external (i.e., medications, environmental changes) factors. Patients oftentimes use terms such as weak, dizzy, numb in an interchangeable fashion and clearly more than one condition (neurologic or otherwise) can cause such symptoms (Fig. 78.1). Incorrectly labeling as “autonomic dysfunction” a primary non-autonomic problem is at least as frequent as the lack of recognition of an autonomic dysfunction. In either scenario, patients end up in not receiving the proper treatment while often receiving wrong ones with potentially serious consequences. In recent years, the autonomic disorders field has grown significantly. Tests that used to be available only in specialized centers are now offered in many medical offices, more in depth investigations have been used almost routinely (such as MIBG or PET scans) [2], and autonomic disorders awareness has grown as well. Patients have also become more savvy, thanks to the plethora of information streaming through various media like the internet. Increased awareness and recognition has created a pseudo-epidemic of conditions such as postural tachycardia syndrome. Unfortunately with this we have also witnessed a rise in misdiagnosis. The focus of this section will be on providing key points in history taking, bedside assessment and how to incorporate the most informative available tests in a cost-effective manner, with the goal of aiding the reader in identifying

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00078-0

the more serious autonomic failure, differentiating it from more benign dysfunctional syndromes and from their most common imitators.

APPROACH IN HISTORY TAKING Systematic symptom review by autonomic domain/ system: l l l l l

cardiovascular gastrointestinal genitourinary sudomotor secretomotor. By temporal profile:

l l

acute, subacute, chronic paroxysmal. By associated symptoms to determine level of pathology:

l

brain, spinal cord, peripheral nerves etc.

The goal is to differentiate more serious conditions from benign ones, recognize specific disorders’ pattern, and exclude reversible/treatable conditions.

ASSESSMENT: BEDSIDE AND LABORATORY TESTING A preliminary evaluation can be easily done at the bedside, at least for the most disabling symptom of autonomic dysfunction, i.e., orthostatic intolerance. Determining the presence of heart rate variability with respiration and HR response to standing together with BP measurements can provide valuable information regarding the nature of the problem, specifically in regard to its being a more or less benign one. Significant decrement in BP without compensatory tachycardia is needless to say much worse prognostically than marked tachycardia without significant BP changes.

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78. ClInICAl EvAluATIon of AuTonomIC DIsoRDERs

BOX 78.1

S Y M P T O M S O F AU T O N O M I C D Y S F U N C T I O N

Reduced orthostatic tolerance: Both orthostatic hypotension and orthostatic intolerance (i.e., POTS) l l l l l l l l l l l l l

Lightheadedness Dizziness Visual obscuration Hot sensation Clammy/cold feeling Weakness Disorientation/cognitive difficulties Imbalance Slurred speech “Coat-hanger” headache and shoulder discomfort Presyncopal symptoms Fatigue Exercise intolerance.

l l l l l l l l

Nausea Bloating Vomiting Anorexia, altered taste. Constipation Diarrhea Urgency Involuntary weight loss.

Genitourinary symptoms l l l l l

Urgency Difficulty voiding/emptying Nocturia Incontinence Erectile dysfunction.

Sweating abnormalities More likely in POTS and related disorders: l l l l l l

Palpitations Tremulousness Anxious feeling Shortness of breath Sleep disturbances Other vasomotor manifestations such as migraine, Raynaud’s phenomenon. Caveat: Some patients, particularly elderly, may have OH without “classic” symptoms, but more vague descriptors such as fatigue, lack of energy, “lack of ambition”.

l l l l l l l

Secretomotor symptoms l l l

Factors that often worsen the above symptoms: l l l l l l l l l l l

Heat exposure (including hot shower, etc.) Intercurrent illness/fever Large meals Prolonged rest/immobilization Exertion Stress/anxiety Dehydration Rapid weight loss Alcoholic beverages Caffeine (POTS, not OH) Menstrual cycle/hormonal changes.

Hypo/anhidrosis – heat intolerance Compensatory hyperhidrosis Essential hyperhidrosis Focal hyperhidrosis Acral hyperhidrosis Paroxysmal hyperhidrosis Gustatory sweating.

l

Dry eyes Dry mouth – altered taste Nose bleeds – nasal congestion Painful urination.

Other symptoms l

l l l

Blurry vision (altered pupillary accommodation) – light sensitivity Abnormal body temperature Skin color changes Trophic changes. Exclude medications’ effect: anticholinergics, adrenolytics, diuretics, antidepressants, opioids, recreational drugs etc. Exclude systemic conditions such as CHF, kidney failure, anemia etc.

Gastrointestinal symptoms l

Early satiety

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AssEssmEnT: BEDsIDE AnD lABoRAToRy TEsTIng

BOX 78.2

P H Y S I C A L E X A M I N AT I O N I N AU T O N O M I C D Y S F U N C T I O N

Examination l l l l l l l l

l

Pupillary responses Dryness of mucosae Dryness of skin Skin color, temperature Skin wrinkling with water immersion Dystrophic changes BP supine/standing HR supine/standing

l l l

HR variability Dependent edema Rectal exam for anal tone Bowel sounds.

Neurologic signs suggesting specific disorders: l

Extrapyramidal, cerebellar signs – MSA and related disorder etc.

Lightheadedness spells

Orthostatic Hypotension (OH)

Isolated ? no

Orthostatic intolerance

POTS and related Dysfunctional syndromes (IBS, CFS)

Not caused by autonomic dysfunction

Seizures; cardiac or Other systemic disease Sensory disturbance Motor impairment

yes Idiopathic OH OH due to systemic conditions

Diffuse/generalized Autonomic disorder

Autonomic neuro-, Ganglionopathies, PAF

Central autonomic disorders

FIGURE 78.1 Example of algorithmic approach to most common complaint in autonomic clinics. Abbreviations: postural tachycardia s. (POTS), chronic fatigue s. (CFS), irritable bowel s. (IBS), pure autonomic failure (PAF).

The severity of a patient’s symptom may not, however, correlate with the magnitude of hemodynamic abnormalities, as, for instance, a slow, neurodegenerative process may allow for compensatory changes (such as supine

hypertension, expanded cerebral autoregulation etc.), while acute onset syndromes can be extremely disabling. Lack of the normal sinus arrhythmia is also prognostically relevant, as it has been associated with increased

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mortality, and can be seen relatively early in conditions such as diabetic and amyloidotic autonomic neuropathies [3]. Labile BP, with large fluctuations in hypertensive and hypotensive range, supersensitivity to medications as well as physiologic states (such as a full bladder) or maneuvers (such as hand grip) is an indicator of baroreflex failure and is one of the most difficult situations to manage [4,5]. Secretomotor and sudomotor functions may be assessed simply by observation of the mucosae (mouth, eyes) and by appreciating presence of moisture on the skin by palpation or by dragging a metal object on the skin (the object will glide on dry skin, and encounter resistance on a normally sweating one). Hyperhidrosis may be appreciated quite easily, as sweat droplets over the skin, visibly wet garments or as wet stains in shoes. Certain pain syndromes can be associated with marked skin vasomotor and sudomotor changes, such as complex regional pain syndrome I and II and erythromelalgia. In erythromelalgia the affected body parts are typically red and hot, with or without appreciable sweat loss, and cold exposure markedly reduces the symptoms. On the other hand, in CRPS the affected limb can be warm or cold, red or bluish, dry or with excessive sweat and cold typically worsens pain. Dystrophic changes may occur long term if these conditions do not resolve. In CRPS dystonic posturing can also occur.

Laboratory Testing A formal passive tilt study with non-invasive beat to beat hemodynamic monitoring can provide useful additional details to the bedside assessment [6]. Passive tilt physiology is somewhat different from active standing, as muscle activity is less and does not elicit reflexes that could cause early BP drop. On the other hand, the lower leg muscle contraction reduces preload by generating a lower pump effect, thus delayed BP drop is much more likely on passive tilt than active standing. A tilt study is best done after 30 of supine baseline rest. Upon tilt up, the overall hemodynamic profile can elucidate the nature of the underlying symptomatology: such as the presence of excessive oscillations (common in the more benign syndromes), early BP drop with subsequent recovery, delayed BP drop, magnitude of HR response to BP changes (i.e., excessive or inappropriate, blunted or even absent). Correlation with a patient’s reported symptoms during the study is always important. A BP decrease of 20 mmHg systolic or 10 mmHg diastolic within 3 minutes is considered significant for the diagnosis of OH. In chronic cases, supine hypertension is often seen and should be noted. In syndromes of reduced orthostatic tolerance such as postural tachycardia syndrome (POTS), a HR rise of 30 bpm in adults without significant BP changes for 50% of tilt is usually accepted for diagnosis if accompanied by symptoms. In the young the increment should be 40.

In patients with history suggestive of syncope, the study can demonstrate which parameter drops first (BP in vasodepressor, HR in vagal and both simultaneously in neurocardiogenic syncope). Increased tilt duration and pharmacologic interventions may increase the study sensitivity, but at the price of lower specificity. We have adopted 10´ tilt duration whenever a disorder of reduced orthostatic tolerance is suspected as a reasonable balance between sensitivity, specificity and practicality. Occasionally, when the nature of the spells is unclear despite many investigations, simultaneous EEG monitoring may identify an ictal bradycardia or asystole and on the other hand provide confirmation that convulsive manifestations not uncommonly seen in syncope are secondary to the hemodynamic changes. Holter BP and HR monitoring can be invaluable to identify spells not captured during routine tilt study, may identify lack of the normal nocturnal blood pressure dipping, marked changes at specific times (such as after meals), paroxysmal changes (like those seen in baroreflex failure) and so forth. When disorders such as postural tachycardia syndrome and related conditions are suspected, an exercise test measuring also peak VO2 can be helpful to demonstrate conditioning level and guide retraining program. A very sensitive and informative test is the Valsalva maneuver. It consists of a forced expiratory effort at a standard expiratory pressure (40 mmHg) for 15 seconds. The maneuver generates a cascade of events involving baroreflexes, heart and arterial vasoconstriction. This maneuver results in stoppage of venous return, resulting in a drop in BP, activation of baroreflex, reflex tachycardia and peripheral vasoconstriction. At the end of the maneuver, the events reverse. Analysis of BP and HR changes during the maneuver provides information about each cardiovascular component: vagal and cardio sympathetic, alpha-adrenergic vasoconstricting capacity and integrity of baroreflex arc. The BP waveform during the Valsalva maneuver consists of four well-defined phases. Phase I and III are generated by the change in intrathoracic pressure (and consequent effect on large vessels) at the onset and at the release of the maneuver. The most interesting components to gather information on the integrity of autonomic system are phase II and IV. Phase II is further subdivided in an early phase II (when the BP declines) and a late phase II (where there is an inversion in BP curve with recovery towards baseline and even above baseline). The BP decline activates baroreflexes inducing tachycardia through both vagal withdrawal and cardiosympathetic activation. During phase IV there is a BP overshoot, paralleled by vagal activation leading to bradycardia before both HR and BP return to baseline levels. The factor with greatest impact on phase II profile is alpha-adrenergic function (i.e., arterial vasoconstriction) while phase IV overshoot is mainly beta-adrenergic dependent. Obviously, the overall maneuver is affected by volume status, and integrity

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of the baroreflex arc. The ratio between the highest HR reached during late phase II and the lowest one reached during phase IV represents the Valsalva ratio. Age, sex, expiratory pressure, position during the maneuver (i.e., supine vs. inclined) and of course medications can affect the results. The maneuver as stated above is very sensitive and can identify preclinical dysfunctions. Sinus arrhythmia is measured more precisely with metronymic breathing, paced at 6/min rate which generates the highest HR changes. This is a purely vagal test, with afferents from lung (sensing stretch), heart chambers and large vessels (sensing filling and pressure changes) and efferent to the sinus node modulating the heart rate. Age, body mass index, depth of breaths, and medications can affect the results. Also, the magnitude of the arrhythmia declines as CO2 is washed out. As for the Valsalva maneuver, one should not forget pulmonary and/or cardiac pathology may also cause abnormal results. The number of maneuvers and other available tests to study cardiovascular and cardiovagal functions is quite large, but some are rarely used to poor reproducibility, some are obsolete while others are used exclusively in research setting. Sympathovagal quantification through frequency analysis has remained mainly a research tool as is microneurography. New technology using nuclear medicine has shown to be valuable to assess integrity of cardiac sympathetic innervation. There are two available sweat tests: one assesses postganglionic function and one assesses the entire thermoregulatory pathway. Combining the two can determine the site (i.e., preganglionic, ganglionic and postganglionic) of pathology. There are other techniques currently under study to assess other sweat parameters such as glands recruitment [7]. Skin and mucosal biopsies can also be used to quantify autonomic fibers around vessels, sweat glands and in the gut, but these techniques are still being perfected [8]. In terms of biological specimens, basic blood and urine tests should be obtained more to exclude systemic conditions such as amyloidosis although anemia is often seen in autonomic failure. Supine and standing catecholamine measurements are useful in discriminating central versus peripheral disorders, benign from less so conditions and genetic disorders [9]. Plasma volume measurement (or, as a surrogate, 24 hours urine sodium) may be helpful to establish pathogenesis and guide treatment. Acetylcholine receptor antibody should be checked whenever an autoimmune disorder is suspected [10]. When conditions like chronic regional pain syndrome (CRPS) and erythromelalgia are considered, vasomotor and sudomotor functions are the most sensitive. Vasomotor assessment is possible using laser Doppler probes, but the extreme sensitivity of these techniques makes them of difficult use in clinical setting as they

require significant time, expertise and patient cooperation. Some vasomotor studies are performed in peripheral vascular laboratories. In the autonomic area temperature measurements are used as surrogate to estimate blood flow. Telethermography and infrared thermometry are commonly utilized to detect asymmetry between affected and unaffected areas. There is no specific pattern or test that can diagnose CRPS: the diagnosis is a clinical one and the studies are only supporting it. Other conditions, painful and not, can have asymmetric vasomotor and sudomotor functions. Saliva and tear production are rarely measured, as clinical assessment often is sufficient, but Schirmer’s test, Rose-Bengal stain and lip biopsy (to assess salivary gland status) can be easily done (usually to diagnose Sjögren’s syndrome). Pupillography can provide, together with pharmacologic studies, a wealth of information on pupillary function and dysfunction. The value of additional studies depends upon the suspected underlying condition. For example, brain imaging, sleep study, cardiac nuclear imaging are important in central disorders such as MSA. Gastrointestinal transit studies, anorectal manometry and urodynamic studies are critical to evaluate respectively gastrointestinal dysmotility syndromes and neurogenic bladder. Erectile dysfunction is also best evaluated with specific tests as opposed to a simple “stamp test” to detect presence of nocturnal tumescence (which is only one aspect of normal function).

FINAL THOUGHTS l

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The goal of a thorough evaluation is to ascertain the presence of an autonomic dysfunction, determine its cause, extent and severity, guide the treatment plan and judge its efficacy on follow up studies. A thoughtful approach should balance costeffectiveness with a comprehensive evaluation. Some of these conditions are common but others are quite rare and if they are not carefully looked for, they will never be found. Pseudo autonomic manifestations can be the most difficult ones to be properly diagnosed. If unsure, better not to attach any label to a patient: think autonomic but not only autonomic.

References [1] Cheshire WP, Kuntz NL. Clinical evaluation of the patient with an autonomic disorder. In: Low PA, Benarroch EE, editors. Clinical Autonomic Disorder (3rd ed.). Baltimore-Philadelphia: Lippincott Williams & Wilkins; 2008. p. 112–29. [2] Goldstein DS. Neuroscience and heart-brain medicine: the year in review. Cleve Clin J Med 2010;77(Suppl. 3):S34–9. [3] Astrup AS, Tarnow L, Rossing P, Hansen BV, Hilsted J, Parving HH. Cardiac autonomic neuropathy predicts cardiovascular morbidity

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[4] [5]

[6]

[7]

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and mortality in type 1 diabetic patients with diabetic nephropathy. Diabetes Care 2006;29:334–9. Benarroch EE. The arterial baroreflex; functional organization and involvement in neurological diseases. Neurology 2008;21:1733–8. Robertson D, Hollister AS, Biaggioni I, Netterville JL, MosquedaGarcia R, Robertson RM. The diagnosis and treatment of baroreflex failure. N Engl J Med 1993;329:1449–55. Low PA, Sletten DM. Laboratory evaluation of autonomic failure. In: Low PA, Benarroch EE, editors. Clinical Autonomic Disorder (3rd ed.). Baltimore-Philadelphia: Lippincott Williams & Wilkins; 2008. p. 130–63. Gibbons CH, Illigens BM, Wang N, Freeman R. Quantification of sudomotor innervation: a comparison of three methods. Muscle and Nerve 2010;42:112–9.

[8] Hilz MJ, Axelrod FB, Bickel A, Stemper B, Brys M, WendelschaferCrabb G, et al. Assessing function and pathology in familial dysautonomia: assessment of temperature perception, sweating and cutaneous innervation. Brain 2004;127:2090–8. [9] Goldstein DS. Noradrenergic transmission. In: Roberston D, Low PA, Polinsky RJ, editors. Primer on the autonomic nervous system. : Academic Press; 1996. p. 91–8. [10] Vernino S. Antibody testing as a diagnostic tool in autonomic disorders. Clin Auton Res 2009;19:13–19.

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C H A P T E R

79 Tilt Table Studies Satish R. Raj 10 QUESTIONS ABOUT HEAD-UP TILT TABLE TESTING 1. What is a Head-Up Tilt Table Test (HUT)? HUT is a method of simulating a prolonged “passive stand”, without the active contraction of calf/leg muscles. It is meant to reproduce the blood pooling in the lower extremities that can occur when someone is standing or sitting still for a prolonged period of time. A HUT involves having a patient lie flat on a table with either footboard support or a seat for support. The patient is strapped to the table securely for safety purposes. While monitoring at least the patient’s heart rate (HR), via continuous electrocardiographic monitoring, and blood pressure (BP), the table is tilted up rapidly to 60–80 degrees head-up so that the patient is almost upright. Most commonly, HUT is used in the diagnosis of neurally mediated syncope. The HUT is usually continued until the patient faints or experiences severe presyncope AND develops significant hypotension, or until the HUT protocol is completed. The blood pressure during HUT is usually stable for some time, before a sudden and precipitous fall (Fig. 79.1A).

2. What are the Different Types of Hut that Exist? The early HUT involved tilting subjects to 60–80° head-up tilt for up to 45–60 minutes duration [1]. These are sometime termed “passive HUT” to reflect that there are no “active drugs” given as a provocative agent. While these protocols can be time-consuming, the primary advantage of these protocols is that since only passive orthostatic stress is involved, there are not possible confounding effects of a non-physiological response to a drug. The current common approach to HUT currently involves a shorter drug-free tilt (20–45 min) followed by the administration of a “provocative agent” to further stress the body and potentially bring out a “physiological” neurally mediated reaction that had previously been hidden. Unfortunately, there is always a tradeoff of reduced specificity of the test for the increased sensitivity that the provocative agents provide. The most commonly used

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00079-2

HUT provocative agents are intravenous isoproterenol [2,3] (a beta-agonist that should increase myocardial contractility and promote vasodilation), intravenous [4] or sublingual [5] nitroglycerine (a potent venodilator that should accentuate preload depletion) and intravenous clomipramine [6] (which might acutely alter central serotonin levels).

3. What Types of Monitoring Should there be During HUT? At a minimum, HR and BP need to be monitored during the HUT. The HR is usually monitored from a continuous electrocardiographic monitor. This will provide not only the HR, but also alert the medical staff to occasional heart rhythm problems that can arise during HUT. The bare minimum BP monitoring required is an automated brachial BP cuff. A manual BP cuff should also be available since automated BP machines are often unable to measure a BP during a rapid change or fall in blood pressure (as can be seen during a HUT study). The problem with brachial BP assessments is that they are intermittent and may miss oscillations in blood pressure or rapid drops in blood pressure. A better approach would be to continuously monitor BP. Some laboratories still use arterial lines for BP monitoring, but the more common current approach is to use a non-invasive continuous blood pressure system that either uses a finger BP cuff or sits on top of the radial artery.

4. What is the Physiological Basis Underlying HUT? With assumption of the upright posture, there is a rapid descent of ~500 ml of blood from the thorax to the lower abdomen, buttocks, and legs. There can also be a 10–25% shift of plasma volume out of the vasculature and into the interstitial tissue [7,8] most of which occurs rapidly (within 10 minutes). This decreases cardiac venous return and may reduce the pressure on the baroreceptors, triggering a compensatory sympathetic activation with tachycardia and vasoconstriction. The net effect of assuming upright posture is a 10–20 beat per minute increase in HR,

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a negligible change in systolic BP, and a ~5 mmHg increase in diastolic BP. The most common pathophysiological explanation for neurally mediated syncope is known as the “Ventricular Hypothesis” [9]. This hypothesis argues that the increased sympathetic tone (above) can lead to increased inotropy, which in the setting of an underfilled ventricle, is thought to trigger a reflex loss of sympathetic tone and increase vagal tone (with resultant hypotension and/or bradycardia). HUT acts as a stimulus for the fluid shift and decreased venous return that initiates the neurally mediated reaction.

On the other hand, HUT are not particularly useful for predicting prognosis or eventual clinical outcome, nor the response to therapy.

7. What are the Indications for HUT? According to the European Society of Cardiology (2009) [11] guidelines, HUT was felt to be appropriate in the following clinical circumstances (after each item is the Recommendation Class): l

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5. How Accurate are HUT for Neurally Mediated Syncope? One challenge is that there is no “gold standard” diagnostic test to diagnose neurally mediated syncope. In most studies, the history of recurrent fainting is used as the gold standard. A positive response to HUT was seen in 49% of drug-free tests [10] and 61–69% of drug-provoked tests in patients with prior syncope [11]. This is the rough correlate of test sensitivity. The specificity of HUT can be even more difficult to interpret. The first lifetime syncopal spell can occur at any age up to about 45, and the lifetime prevalence is around 37% [12]. Thus a “false-positive” tilt may be a mistake (truly false) or it may reflect a physiological propensity to a neurally mediated reaction that has yet to manifest clinically. Although data from individual studies are quite variable, overall HUT has both test positivity and specificity around 70–75%. Another challenge is that HUT has very imperfect test reproducibility. HUT reproducibility is 70–85% over days to months when both presyncope and syncope are deemed to be positive test outcomes [11]. This may reflect a “training effect” with physiological adaptation within the patient to the HUT.

6. How Useful are HUT in the Clinical Management of Patients? HUT in isolation does not provide a clinical diagnosis. Similar to the use of exercise stress testing for coronary artery disease, HUT is not very useful if you have either a very high or very low clinical probability for a diagnosis of neurally mediated syncope. Rather, it is most useful when there is an intermediate clinical probability for neurally mediated syncope with some diagnostic uncertainty. HUT can have many clinical uses. First, it can give useful information to aid in a clinical diagnosis when the history has left diagnostic uncertainty. HUT can also be a very useful patient education tool so that they recognize their prodromal presyncopal symptoms as related to the neurally mediated reaction. It can also provide reassurance, especially in the setting of “convulsive syncope”.

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RECURRENT syncope in the absence of structural heart disease (Class I). RECURRENT syncope in the presence of structural heart disease AFTER exclusion of other potential cardiac causes of syncope (Class I). SINGLE syncopal episode if it was associated with physical injury OR if the patient has a high risk occupation (e.g., pilot) (Class I). When there is clinical value in demonstrating neurocardiogenic syncope to the patient (education/ reassurance) (Class I). Differentiating syncope with myoclonic activity from epilepsy (Class IIb). Evaluation of patients with recurrent unexplained falls (Class IIb). Assessment of patients with frequent syncope and psychiatric disease (Class IIb).

8. When is Extra Monitoring Like Transcranial Doppler (TCD) or Electroencephalogram (EEG) Monitoring Useful? In addition to good monitoring of HR and BP during HUT, ancillary monitoring can sometimes be useful. Some HUT laboratories will routinely also monitor cerebral blood flow velocity (via TCD) and/or EEG activity during the HUT. At the time of typical syncope with hypotension on tilt, the cerebral blood flow velocity pattern changes significantly with decreases in diastolic pressure and diminished diastolic cerebral flow, and the EEG activity can slow or even become asystolic. We do not use these ancillary tests routinely, only in selected cases. Specifically, in those cases when a patient has an apparent loss of consciousness in the absence of hypotension, we will then repeat the HUT with additional monitoring. This can help to differentiate rare physiological disorders such as “cerebral syncope” [13] (due to isolated cerebral vasospasm) from non-hemodynamic (or psychogenic) spells.

9. Is HUT Useful in the Diagnosis of Postural Tachycardia Syndrome (POTS)? POTS is discussed in more length in other chapters. Briefly, it is a clinical syndrome that has a cardinal criterion of an excessive increase in HR on assuming an

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FIGURE 79.1 (A) Neurally mediated syncope. Heart rate (HR) and blood pressure (BP) are shown for the duration of the head-up tilt table test. BP was stable and HR increased slightly and was stable over the first 30 minutes. After 35 minutes, there was an abrupt decrease in BP over ~20 seconds with syncope. (B) Postural tachycardia syndrome. There is an excessive increase in heart rate (HR) with upright posture in patients with Postural Tachycardia Syndrome (POTS). This test was terminated early due to severe presyncope with tachycardia in the absence of classic neurally mediated hypotension. (C) Neurogenic orthostatic hypotension. Patients with neurogenic orthostatic hypotension have a rapid and severe drop in blood pressure (BP) almost immediately on upright tilt, with only a modest increase in heart rate (HR). (D) Delayed orthostatic hypotension. Patients with delayed orthostatic hypotension have a slow and gradual decrease in blood pressure (BP). The drop in BP is not 20/10 mmHg within 3 minutes. The pattern of BP fall is akin to gradually “rolling down a hill”. All parts reproduced with permission from Electrophysiological Disorders of the Heart [17].

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79. TIlT TAblE STudIES

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upright posture (HR increase 30 bpm increase within 10 minutes) in the absence of orthostatic hypotension (BP drop 20/10 mmHg). Many autonomic labs use a short 10-minute HUT for the assessment of this orthostatic tachycardia (Fig. 79.1B). POTS patients can also have neurally mediated syncope, but only a minority of patients with POTS (10%) report frank syncope [14].

10. Is HUT Useful in the Diagnosis of Orthostatic Hypotension? Patients with neurogenic orthostatic hypotension present with a decrease in BP 20/10 mmHg within 3 minutes of

upright posture [15] and only a modest reflex increase in HR (Fig. 79.1C). Some patients can maintain their lower BP with ongoing orthostatic stress, while the BP progressively decreases over time in other patients [16]. HUT can provide a safe method for an orthostatic challenge for these patients. A more recently described variant is delayed orthostatic hypotension [15]. This is characterized by a slow and gradual decrease in BP that does not occur immediately (Fig. 79.1D), as is typically seen with neurogenic orthostatic hypotension. The gradual decrease in BP (“rolling down a ramp”) in delayed orthostatic hypotension contrasts to NMS, which has an initially stable BP followed by a

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10 QuESTIonS AbouT HEAd-uP TIlT TAblE TESTIng

sudden drop (“falling off of a cliff”) (Fig. 79.1A). The exact underlying causes are not known.

References [1] Fitzpatrick AP, Theodorakis G, Vardas P, Sutton R. Methodology of head-up tilt testing in patients with unexplained syncope. J Am Coll Cardiol 1991;17:125–30. [2] Morillo CA, Klein GJ, Zandri S, Yee R. Diagnostic accuracy of a lowdose isoproterenol head-up tilt protocol. Am Heart J 1995;129:901–6. [3] Natale A, Akhtar M, Jazayeri M, Dhala A, Blanck Z, Deshpande S, et al. Provocation of hypotension during head-up tilt testing in subjects with no history of syncope or presyncope. Circulation 1995;92:54–8. [4] Raviele A, Gasparini G, Di Pede F, Menozzi C, Brignole M, Dinelli M, et al. Nitroglycerin infusion during upright tilt: a new test for the diagnosis of vasovagal syncope. Am Heart J 1994;127:103–11. [5] Raviele A, Menozzi C, Brignole M, Gasparini G, Alboni P, Musso G, et al. Value of head-up tilt testing potentiated with sublingual nitroglycerin to assess the origin of unexplained syncope. Am J Cardiol 1995;76:267–72. [6] Theodorakis GN, Markianos M, Zarvalis E, Livanis EG, Flevari P, Kremastinos DT. Provocation of neurocardiogenic syncope by clomipramine administration during the head-up tilt test in vasovagal syndrome. J Am Coll Cardiol 2000;36:174–8. [7] Raj SR, Biaggioni I, Yamhure PC, Black BK, Paranjape SY, Byrne DW, et al. Renin-aldosterone paradox and perturbed blood volume regulation underlying postural tachycardia syndrome. Circulation 2005;111:1574–82. [8] Jacob G, Ertl AC, Shannon JR, Furlan R, Robertson RM, Robertson D. Effect of standing on neurohumoral responses and plasma volume in healthy subjects. J Appl Physiol 1998;84:914–21.

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[9] Mosqueda-Garcia R, Furlan R, Tank J, Fernandez-Violante R. The elusive pathophysiology of neurally mediated syncope. Circulation 2000;102:2898–906. [10] Kapoor WN, Smith MA, Miller NL. Upright tilt testing in evaluating syncope: a comprehensive literature review. Am J Med 1994;97:78–88. [11] Moya A, Sutton R, Ammirati F, et al. Guidelines for the diagnosis and management of syncope version 2009.: The Task Force for the Diagnosis and Management of Syncope of the European Society of Cardiology (ESC). Eur Heart J 2009. [12] Ganzeboom KS, Colman N, Reitsma JB, Shen WK, Wieling W. Prevalence and triggers of syncope in medical students. Am J Cardiol 2003;91(1006-8):A8. [13] Grubb BP. Cerebral syncope: new insights into an emerging entity. J Pediatr 2000;136:431–2. [14] Kanjwal K, Sheikh M, Karabin B, Kanjwal Y, Grubb BP. Neurocardiogenic syncope coexisting with postural orthostatic tachycardia syndrome in patients suffering from orthostatic intolerance: a combined form of autonomic dysfunction. Pacing Clin Electrophysiol 2011. [15] The Consensus Committee of the American Autonomic Society and the American Academy of Neurology. Consensus statement on the definition of orthostatic hypotension, pure autonomic failure, and multiple system atrophy. Neurology 1996;46:1470. [16] Gehrking JA, Hines SM, Benrud-Larson LM, Opher-Gehrking TL, Low PA. What is the minimum duration of head-up tilt necessary to detect orthostatic hypotension? Clin Auton Res 2005;15:71–5. [17] Raj SR, Sheldon RS. Head-up tilt-table test. In: Saksena S, Camm AJ, editors. Electrophysiological Disorders of the Heart (2/e ed).: Elsevier Inc; 2011. p. 73-1–73-11.

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C H A P T E R

80 Sympathetic Microneurography B. Gunnar Wallin Microneurography was developed in the mid-1960s for percutaneous recordings of action potentials in human peripheral nerves. The technique was intended for myelinated sensory fibers, but it was found to be useful also for recordings from unmyelinated postganglionic sympathetic axons [1]. Since then, many studies have been published on human sympathetic nerve traffic to skin and muscle, mostly in groups of sympathetic fibers (multiunit activity), but also in single axons. Large extremity nerves are commonly used, but recordings have been made from small nerves in the face and mouth and extremities. The methodology has provided valuable new knowledge on autonomic physiology and pathophysiology, but it is not suitable for diagnostic assessment in individual patients. This chapter summarizes methodologic aspects of microneurography from biological and technical viewpoints. Detailed accounts have been published previously [2–4].

METHODOLOGY Equipment Commonly used equipment includes insulated monopolar tungsten microelectrodes with tip diameters of a few micrometers, but a concentric needle electrode has been described [5]. Electrodes are available commercially or may be produced in the laboratory. Impedances range from about 50 k to several M; single fiber recordings require electrodes with smaller uninsulated tips and higher impedance. Usually, the neurogram is amplified (gain 50–100 k) in two steps: first in a preamplifier/impedance converter positioned close to the recording site, and then in a main amplifier. To quantify multiunit activity, the raw neurogram is full wave rectified and passed through a leaky integrator with a time constant of 0.1 second. Audio monitoring, which facilitates the actual recording, is made after noise reduction. This is achieved by filters (e.g., bandwidth 500–2000 Hz) and a discriminator, which cuts out the central portion of the neurogram, transmitting only the positive and negative peaks of the signals to the audio amplifier [4].

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00080-9

Recently, the recording equipment has been adapted to a magnetic environment, thereby enabling recordings of sympathetic activity during fMRI procedures [6].

PROCEDURE The nerve is located by the paresthesia/muscle twitches evoked first by percutaneous electrical stimulation, and then by stimulation through the microelectrode after it has been inserted through the skin. Peripheral nerves contain many fascicles, and, in the distal part of the extremities, fascicles are connected either to a defined skin area or to a muscle. The fascicle and its innervation zone are identified by the type of peripheral stimuli that evoke afferent mechanoreceptive impulses: muscle stretch and light touch stimuli for muscle and skin fascicles, respectively. Sympathetic nerve fibers are not evenly distributed within the fascicle; they lie together with afferent C fibers in bundles inside Schwann cells (Fig. 80.1); therefore, repeated small needle adjustments may be necessary before a sympathetic recording site is found. Sympathetic fibers discharge spontaneously in synchronized bursts, which occur irregularly in characteristic temporal patterns, which differ between skin and muscle nerve fascicles (Fig. 80.1). Furthermore, the activity increases in a predictable way with certain maneuvers (Fig. 80.2). In muscle nerve fascicles, multiunit bursts presumably contain only vasoconstrictor impulses, which occur in the cardiac rhythm, preferentially during reductions of arterial blood pressure; their number increases with apneas or Valsalva maneuvers. In skin fascicles, sympathetic bursts may contain vasoconstrictor, vasodilator, and/or sudomotor impulses; cardiac rhythmicity is usually absent but any surprising sensory stimulus regularly evokes a single discharge. Recordings from single sympathetic fibers are achieved by repeated minute electrode adjustments in a region of a fascicle in which multiunit sympathetic bursts are present [7]. To facilitate fiber identification in skin nerves, experiments are made while subjects are cooled or heated; this biases sympathetic traffic toward vasoconstriction and sweating, respectively. Sites with adequate

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FIGURE 80.1 Schematic figure of microneurographic recording from the peroneal nerve at the fibular head (upper left). The nerve contains skin and muscle fascicle, and, in both types of fascicles sympathetic fibers are located in bundles inside Schwann cells and surrounded by myelinated axons (upper right). Lower left panel shows spontaneous variations of muscle sympathetic activity (integrated neurogram) and blood pressure. Note cardiac rhythmicity and the inverse relationship between neurogram and blood pressure levels. Lower right panel shows spontaneous skin sympathetic activity in original and integrated neurograms. Note that the single unit can be identified only in the original neurogram (bandpass 0.3–5 kHz) (A)

sound

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expiratory apnea

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FIGURE 80.2 Simple maneuvers aiding the identification of skin and muscle sympathetic activity. In skin nerve fascicles (A), a single sympathetic discharge is regularly evoked by any type of arousal stimulus, e.g., a sudden sound (left), or a deep breath (right). Note that the sympathetic burst is followed by transient signs of vasoconstriction (reduced pulse amplitudes in the plethysmogram) and sweating (reduction of skin resistance) indicating that vasoconstrictor and sudomotor fibers are activated in parallel. In muscle nerve fascicles (B), the activity increases markedly towards the end of an expiratory apnea.

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AnAlySIS

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FIGURE 80.3 Recording from a single sympathetic vasoconstrictor fiber in a muscle nerve fascicle. (A) In association with six multiunit bursts (see integrated neurogram) the fiber discharges a single impulse (*). Rastered (B) and superimposed (C) action potentials suggest that the spikes originated from one axon. Variations in spike amplitude in raw neurogram are a consequence of low signal-to-noise ratio. Figure reproduced from ref. [10].

signal-to-noise ratio for multiunit activity are obtained in ~90% of attempts; the corresponding figure for single-unit activity recordings is much less.

ANALYSIS Multiunit Activity In a given electrode site, the strength of multiunit activity in the mean voltage neurogram can be quantified as the number of bursts multiplied by mean burst area (  total activity). In muscle nerves, burst duration is relatively constant; therefore, burst amplitude can be substituted for burst area. Measurement of total activity is suitable for quantifying

changes of activity induced by maneuvers. The strength of resting sympathetic activity, conversely, can only be quantified in terms of the number of bursts; burst amplitude cannot be compared among electrode sites because it depends on the proximity of the electrode tip to the active fibers, which varies among sites. Because muscle sympathetic activity always displays cardiac rhythmicity, the strength of activity is expressed both as number of bursts/100 heartbeats (burst incidence) and bursts/min (burst frequency). Burst detection often is made visually, but computer-assisted analysis is valuable because it is faster, reduces observer bias, and gives accurate measures of burst area [8]. Recently, a technique has been developed that enables determination of the number of sympathetic action potentials contributing to the multiunit sympathetic bursts [9].

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Single-Unit Activity Single-unit analysis requires (i) fiber identification and (ii) evidence that all impulses derive from the same fiber. Fiber identification is difficult in skin nerves, which contain several different types of sympathetic fibers. Therefore, to aid the identification, a thermally induced bias of the activity during recordings and spike-triggered averaging of effector responses should be used whenever possible. To ensure that all impulses derive from one fiber, spike amplitude is important. However, because signal-to-noise ratio is often low, considerable spike amplitude variability may be induced by the noise. Therefore, only triggering on spikes exceeding a certain amplitude level is inadequate; spikes may be missed or, alternatively, spikes from other axons may be included. For this reason, computer assisted inspection of wave form and amplitude, combined with spike superimposition, is necessary before a population of spikes can be attributed to a single axon (Fig. 80.3). Firing in single muscle sympathetic fibers displays cardiac rhythmicity; therefore, it is useful to quantitate not only firing frequency, but also the number of cardiac intervals with unit activity and the number of spikes/cardiac interval.

POTENTIAL DIFFICULTIES Mixed Sites Usually, the character of the sympathetic activity agrees with the classification of a fascicle on the basis of afferent mechanoreceptor responses to sensory stimuli. However, mixed sympathetic activity may be encountered in a fascicle that appeared “pure” based on afferent testing. To avoid such mistakes, careful testing of the sympathetic responsiveness is needed before accepting a recording site as “pure.”

A slow change of site without obvious artefacts may, however, lead to problems. If signs of such changes are present (e.g., a drifting baseline level or slowly increasing burst amplitudes in the mean voltage neurogram), burst amplitude/area cannot be used in the quantitative analysis.

Acknowledgement The author thanks Göran Pegenius for excellent technical assistance. Work was supported by Swedish Medical Research Council Grant no. 12170.

References [1] Hagbarth K-E, Vallbo Å. Pulse and respiratory grouping of sympathetic impulses in human muscle nerves. Acta. Physiol. Scand. 1968;74:96–108. [2] Vallbo ÅB, Hagbarth K-E, Torebjörk HE, Wallin BG. Somatosensory, proprioceptive and sympathetic activity in human peripheral nerves. Physiol. Rev. 1979;59:919–57. [3] Eckberg DL, Sleight P. Human baroreflexes in health and disease. New York: Oxford University Press; 1992. [4] Gandevia SC, Hales JP. The methodology and scope of human microneurography. J. Neurosci. Methods 1997;74:123–36. [5] Hallin RG, Wu G. Protocol for microneurography with concentric needle electrodes. Brain Res. Protocols 1998;2:120–32. [6] Macefield VG, Henderson LA. Real-time imaging of the medullary circuitry involved in the generation of spontaneous muscle sympathetic nerve activity in awake subjects. Hum Brain Mapp 2010;31:539–49. [7] Macefield VG, Elam M, Wallin BG. Firing properties of single postganglionic sympathetic neurones. Auton. Neurosci. 2002;95:146–59. [8] Hamner JW, Taylor JA. Automated quantification of sympathetic beat-by-beat activity, independent of signal quality. J. Appl. Physiol. 2001;91:1199–206. [9] Steinback CD, Salmanpour A, Breskovic T, Dujic Z, Shoemaker KJ. Sympathetic neural activation: an ordered affair. J. Physiol. (Lond.) 2010;588:4825–36. [10] Macefield GV, Wallin BG, Vallbo ÅB. The discharge behaviour of single vasoconstrictor motoneurones in human muscle nerves. J. Physiol. (Lond.) 1994;481:799–809.

Changes of Electrode Site Movements of the electrode tip during a recording (e.g., in association with a maneuver) are usually easy to detect.

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81 Clinical Applications of Microneurography Tadaaki Mano Microneurography is a unique method of recording postganglionic sympathetic neural traffic directly from human peripheral nerves leading to muscle and skin, called muscle sympathetic nerve activity (MSNA) and skin sympathetic nerve activity (SSNA) respectively. Recordings and quantitative analysis of not only multi-unit burst discharges, but also single-unit discharges are available, allowing the study of how muscle and skin sympathetic neural traffic operate in humans. The recording technique, as well as analytical procedures of sympathetic nerve activities are dealt with in chapter 80. MSNA, composed mainly of muscle vasoconstrictor activity, plays an essential role in regulating peripheral vascular resistance in muscle, and controls systemic blood pressure; SSNA, mainly composed of skin vasoconstrictor and sudomotor activities, regulates skin blood flow and sweating, thus controlling body temperature regulation. Microneurography has been used as a potent tool to understand functional properties and pathophysiological mechanisms of various autonomic disorders [1]. It has been applied even in space to analyze the influence of microgravity on the human sympathetic nervous system. In this chapter, findings concerning clinical applications of microneurography in neurological, cardiovascular, renal, metabolic, and bone disorders are reviewed based on literature referring to microneurographic recordings of MSNA and SSNA.

NEUROLOGICAL DISEASES MSNA and SSNA in Radiculoneuropathies Microneurographic recordings have been carried out to elucidate autonomic disorders and their mechanisms in peripheral and central neurological diseases. In peripheral autonomic neuropathies such as diabetic neuropathy with involvement of postganglionic unmyelinated C-efferent fibers, MSNA and SSNA are often hardly recordable, but when recorded their conduction velocities are in the normal range, indicating all or no damage of the unmyelinated sympathetic fibers. The poor response to head-up tilt associated with orthostatic hypotension is observed in peripheral autonomic failure and diabetic autonomic neuropathy. On the other hand, MSNA and SSNA are

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00081-0

increased in demyelinating rediculoneuropathies such as Guillain–Barré syndrome (GBS) and chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). SSNA is increased during an acute phase of GBS and CIDP, while normalized in the recovery phase of GBS, as well as in CIDP after treatment by plasmapheresis. These findings may indicate that sympathetic neural traffic is enhanced by some unknown disinhibitory mechanism related to the radiculoneural demyelinating process.

MSNA in Multiple System Atrophy In multiple system atrophy (MSA), MSNA recordings at rest and its response to head-up tilt are very poor. Orthostatic hypotension occurs with a poor or absent MSNA response to standing. Hypotensive episodes can also occur in some cases of MSA after taking food or oral glucose administration (postprandial hypotension). An oral glucose administration boosts MSNA in normal subjects; however, not in cases of MSA patients suffering from postprandial hypotension. This result suggests that a lack of MSNA response to oral glucose intake is an important neural mechanism of postprandial hypotension.

MSNA Withdrawal in Hypotensive Attacks In vasovagal syncope, hypotensive attacks occur when MSNA responds normally or even highly to orthostatic stress, accompanied by tachycardia. In these cases, normal or highly activated MSNA during orthostatic stress becomes suddenly withdrawn. The abrupt withdrawal during normal or high MSNA response to standing may be induced by the Bezold–Jarisch reflex, which triggers orthostatic hypotension followed by a syncopal attack. In rare cases with non-orthostatic paroxysmal hypotensive attacks, normal or even highly discharged resting MSNA is periodically withdrawn in concomitance with bradycardia and marked hypotension. The period of MSNA withdrawal varies from several minutes to 2 hours. With the reappearance of MSNA, heart rate and blood pressure recover completely. The mechanism underlying these very rare hypotensive attacks is unknown, but may depend on some central mechanisms to induce MSNA withdrawal.

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FIGURE 81.1 SSNA response to increased ambient temperature in a normal subject and an anhidrotic patient. From the top to the bottom, room temperature (R.T.), sudomotor and vasoconstrictor components of SSNA (bursts/min) recorded in an artificial climatic chamber from the median nerve in an age-matched normal control subject (left) and a patient suffering from acquired idiopathic generalized anhidrosis (right). With elevated room temperature, sudomotor bursts increase, while vasoconstrictor bursts decrease both in normal and anhidrotic subjects. However the increase in sudomotor bursts is much higher in the anhidrotic patient than in the normal subjects. Abscissa indicates time course in minutes. Adapted with permission from Murakami K, Sobue G, Iwase S, Mitsuma T, Mano T. Neurology 1993;43:1137–40.

SSNA in Sweat Disturbances In cases with hypo- or anhidrosis in MSA and Parkinson’s disease as well as autonomic neuropathies, sudomotor SSNA and its responses to physical and mental stimuli are much lower than in normal controls. In contrast to the low sudomotor SSNA in the abovementioned diseases, in hypo- or anhidrosis, sudomotor SSNA in cases of acquired idiopathic generalized anhidrosis with lesions confirmed by skin biopsy in eccrine sweat glands and idiopathic pure sudomotor failure (due presumably to lesions in the muscarinic cholinergic receptors of sweat glands), is normal or even higher than in age-matched healthy controls (Fig. 81.1). These results suggest that sudomotor SSNA is reduced when the preand/or postganglionic sudomotor nerve is impaired, but is normal or rather enhanced when the sweat glands or their receptors are impaired. Enhanced sudomotor SSNA in these cases may depend on some feedback mechanisms due; for example, to heat retention. Thus microneurography can be an effective tool to differentiate mechanisms underlying various types of hypo- or anhidrosis. In cases of palmoplantar hyperhidrosis, spontaneous sudomotor SSNA and its responses to mental stimuli, recorded from the tibial nerve which innervates the glabrous skin, are markedly higher than those in normal subjects, while the same activity in these cases recorded

from the peroneal nerve which innervates hairy skin is almost the same as in normal controls. As mentioned above, sudomotor SSNA is enhanced in an acute phase of demyelinating polyradiculoneuropathies associated with hyperhidrosis. Resting SSNA recorded from the peroneal nerve is higher in patients of amyotrophic lateral sclerosis (ALS) than in normal controls; similarly, increased MSNA occurs in this intractable disease. The latency of SSNA response to electrical stimulation is however prolonged in ALS patients compared with that in normal controls. The mechanism underlying these changes in sympathetic neural traffic is still unknown, but may depend on impaired central pathways in ALS [2].

MSNA in Sleep Apnea It has been well-established that MSNA is highly enhanced by sleep apnea (SA), associated with blood pressure elevation. Elevated MSNA and blood pressure in response to SA are linked to a chemoreflex which responds to intermittent arterial hypoxemia, since these responses are blunted by O2 administration. Impaired baroreflex sensitivity may also contribute to enhanced MSNA in SA patients. Obesity which often accompanies obstructive SA (OSA) is not an important factor in the mechanisms of SA, because sympathoexcitation is observed in OSA patients with obesity but not in subjects with only obesity without SA.

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Single-unit recordings of MSNA in OSA patients reveal an increase in firing rate of individual vasomotor neurons within a cardiac interval [3]. This may signify that the mechanism of sympathoexcitation in OSA is different from that of sympathoexcitation in normal as well as in chronic heart failure subjects in which single vasoconstrictor neurons primarily fire only once per cardiac interval. MSNA excitation in OSA patients appears to depend on both recruitment and an increased firing frequency of muscle sympathetic nerve. A shift towards multiple firing within a sympathetic burst may constitute a risk factor of high release of neurotransmitters induced by neural volleys with high instantaneous firing frequencies. In SA patients, MSNA and blood pressure are higher than in normal subjects even during the daytime. Daytime sympathetic hyperactivity in SA may be related to excessive daytime sleepiness. The sympathoexcitation induced by OSA is ameliorated by continuous positive airway pressure (CPAP), normalizing sympathetic chemoreflex responses in OSA. An application of CPAP can reduce enhanced MSNA and hypertension not only during sleep, but also on daytime in OSAS patients. Thus, polysomnography with sympathetic microneurography provides valuable information related to neural mechanisms underlying cardiovascular dysfunction associated with SA.

CARDIOVASCULAR DISEASES MSNA in Hypertension MSNA which controls systemic blood pressure has been most preferably applied to analyze the mechanisms underlying hypertension. A large number of studies are focused on MSNA changes in essential hypertension (EH) in association with or without heart failure. Higher MSNA is reported in patients suffering from EH, associated with or without obesity and/or congestive heart failure than in age-matched normotensive subjects. Comparison of MSNA in the hypertensive and age-matched normotensive controls indicates that MSNA is enhanced in hypertensive elderly, for whom obesity and hypertension are enhancing factors. A progressive MSNA enhancement is related to advancing hypertension. MSNA is significantly higher in patients with accelerated EH patients (diastolic pressure 130 mmHg) than those with benign EH (diastolic pressure 95 and 105 mmHg). Increased MSNA in EH is likely related to impaired baroreflex functions and also to activated renin-angiotensin axis as reported in accelerated EH. The presence of left ventricular hypertrophy is associated with higher MSNA in moderate or severe EH patients. The sympathetic overactivity in hypertension may account for the increased cardiovascular risk characterizing left ventricular diastolic dysfunction [4]. MSNA is also increased in masked hypertension, but single unit MSNA is more increased in EH than in

masked hypertension. MSNA is increased in normotensive offspring of malignant hypertensive parents compared with offspring of normotensive parents while the MSNA response to mental stress is larger in offspring of hypertensive parents. An increase in single-unit as well as multi-unit MSNA in female patients with EH is lower than in male patients with essential hypertension. These findings suggest that MSNA is elevated in EH on the basis of genetic and gender factors.

MSNA in Heart Failure Sympathoexcitation in heart failure (HF) has been reported by many investigators. MSNA is increased both in hypertension and heart failure, however more strongly in heart failure with ventricular arrhythmia. Increased MSNA associated with HF is estimated to depend on rapid shallow breathing with decreased tidal volume and increased breathing frequency. Single unit recordings of MSNA show that increased firing frequency as well as firing probability rises in HF patients, while the discharge of more than one impulse per sympathetic burst does not [5]. Muscle metaboreflex, in which metabolites in working muscles stimulate group III or IV afferent fibers to increase MSNA, might be enhanced in HF patients to elevate MSNA. Since peripheral and central chemoreceptor stimulation increases MSNA and blunts muscle vasodilatation in heart failure patients, attenuated chemoreflex-mediated sympathosuppression may also explain sympathoexcitation in HF patients. On the other hand, attenuated baroreflex as well as cardiopulmonary reflex is also estimated as the cause of sympthoexitation in HF patient [6]. Impaired baroreflex-dependent sympathoexcitation may induce left ventricular dysfunction with ventricular arrythmia and cardiovascular risk such as sudden death. In HF, arterial baro-, chemo-, and metaboreflexes should be enhanced, thereby contributing to the sympathoexcitation in HF. Higher MSNA in HF is associated with higher probability of death; thus, the intensity of the MSNA enhancement may be a prognostic marker in advanced HF.

KIDNEY DISEASES MSNA increases inappropriately in patients with chronic kidney diseases and patients undergoing hemodialysis. Age-dependent MSNA increase is steeper in chronic kidney disease than in normal controls [7] (Fig. 81.2). Coexistence of renal insufficiency with HF induces higher MSNA than in patients with renal insufficiency alone or HF alone. MSNA is inappropriately high in hypertensive patients with renal parenchymal disease. Since polycystic kidney patients with normal renal function exhibit higher MSNA with hypertension, sympathetic overactivity is considered to depend on renal ischemia due to narrowed renal arteries, which enhance renin-angiotensin

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FIGURE 81.2 Relation between age and MSNA in normal controls and chronic kidney disease patients. MSNA recorded from the peroneal nerve correlates with age both in normal controls () and chronic kidney disease patients (), while the regression line is steeper in the patients than in normal controls. Reprinted with permission from Neumann J, Ligtenberg G, Kleinm IL, Koomans HA, Blankestijn PJ. Kidney Int 2004;65:1568–76.

vasopressor system. Increased MSNA in renovascular hypertension is normalized several days after successful percutaneous renal angioplasty. Since enhanced MSNA is normalized by nephrectomy, this enhancement is apparently mediated by afferent signals arising in the kidneys with the narrowed renal arteries. Tonic chemoreflex activation may also contribute to the elevated MSNA in patients with chronic renal failure. Sympathoexcitation in hypertensive chronic kidney diseases contributes to the pathogenesis of renal hypertension and produces poor cardiovascular outcome. This condition is improved but not normalized by a standard antihypertensive treatment with angiotensin-converting enzyme inhibitor or angiotensin II antagonist. Despite the normalized blood pressure by antihypertensive treatment, MSNA is related to increased left ventricular mass measured by MRI in chronic kidney disease patients. In poorly effective cases with standard antihypertensive treatment, an addition of central sympatholytic drug, moxonidine (MOX), to angiotensin II antagonist improves elevated MSNA in initial states of chronic renal failure. Low doses of MOX in addition to pre-existing antihypertensive treatment produce sustained and substantial reductions of MSNA in patients suffering from end-stage renal disease. MSNA recording is beneficial in revealing the sympathoexcitation which may contribute to both cardiovascular morbidity and the progression of chronic kidney disease, and thus in estimating the prognosis of chronic kidney diseases.

METABOLIC SYNDROME Metabolic syndrome (MS) is characterized by central adiposity, diabetes mellitus, dyslipidemia, and/or hypertension. Sympathetic microneurography in MS has revealed that MSNA is significantly higher in subjects with hypertension and in those with obesity than in controls.

MSNA is higher in subjects with MS even without hypertension, correlating directly with waist circumference. Sympathetic activation is more related to central obesity than peripheral one and metabolic factors such as insulin resistance. Weight loss by hypocaloric diet decreases MSNA and increases insulin sensitivity. Regarding the gender difference of sympathetic activation in MS, MSNA is reported to relate mainly to blood pressure in women, while mainly to body mass index in men. Insulin-resistant subjects with MS which is related to central adiposity have a blunted MSNA response to oral glucose ingestion compared with insulin-sensitive subjects. Recent work indicates that chronic mental stress in patients with MS modulates the pattern of sympathetic activity which may be related to cardiovascular risk [8]. Multi-unit MSNA of women who present higher depressive symptom scores is similar to that of men, but the women display a disturbed firing pattern of single-unit MSNA as indicated by a higher incidence of multiple spikes per burst. The single-unit MSNA firing pattern does not correlate with any aspect of the metabolic profile but significantly associated with an anxiety state. In particular, a higher incidence of multiple firing (more than two spikes) during a sympathetic neural burst is associated with higher anxiety score and higher affective depression symptom. Somatic symptoms bear no association with the sympathetic firing pattern. These findings indicate that patients with MS have in general sympthoexitation, of which discharge pattern may be influenced by mental condition of patients such as anxiety associated with this syndrome.

BONE LOSS Recent studies have indicated that bone metabolism is regulated by sympathetic nervous system. It has been reported that sympathetic neural traffic to bone inhibits the function of osteoblast and enhances that of osteroclast, through leptin, hypothalamus, sympathetic nerve, noradrenaline and β2-adrenoreceptors in bone, and thus induces bone loss [9]. Our recent preliminary study shows that changes in MSNA recorded from the tibial nerve of healthy human subjects who are exposed to head-down bed rest for 20 days to simulate microgravity have a significant correlation with changes in urinary secretion level of deoxypyridinoline, which is used as a specific marker for measuring bone resorption [10] (Fig. 81.3). This finding suggests that increased sympathetic neural traffic in the tibial nerve which also includes bone nerve branches is related to bone resorption. Further investigations are necessary to clarify this issue.

CONCLUSION Clinical applications of microneurography provide a direct approach to the sympathetic nervous system in humans and

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FIGURE 81.3 Correlation between MSNA and deoxypyridinoline after exposure to simulated microgravity. Percentage changes of MSNA recorded from the tibial nerve is significantly correlated with the changes in urinary secretion of deoxypyridinoline, a marker of bone resorption, in 11 young healthy male subjects after an exposure to head-down bed rest to simulate microgravity for 20 days. Reprinted with permission from Mano T, Nishimura N, Iwase S. Acta Physiol Hung 2010;97:354–61.

valuable information on disease characteristics and pathophysiological mechanisms. Microneurographic recordings of MSNA are particularly indispensable to differentiate mechanisms underlying various types of hypotensive attacks. Those of SSNA to reveal pathophysiology of different kinds of hypo- and anhidrosis, cannot be replaced by other methods. In these cases, microneurography can play an essential role like EEG in diagnosis of epilepsy. Besides microneurographic applications on neurological, cardiovascular, renal, metabolic, and bone diseases as exampled in the limited pages of this chapter, this method has been much more widely applied to elucidate functional abnormalities in many other disorders and pathological conditions. The method is

[1] Mano T, Iwase S, Toma S. Microneurography as a tool in clinical neurophysiology to investigate peripheral neural traffic in human. Clin. Neurophysiol. 2006;117:2357–84. [2] Shindo K, Watanabe H, Ohta E, Nagasaka K, Shiozawa Z, Takiyama Y. Sympathetic sudomotor neural function in amyotrophic lateral sclerosis. Amyotrph. Lateral Scler 2011;12(1):39–44. [3] Elam M, McKenzie D, Macefield VG. Mechanisms of sympathoexitation: single-unit analysis of muscle vasoconstrictor neurons in awake obstructive sleep apnea syndrome subjects. J. Appl. Physiol. 2002;93:297–303. [4] Grassi G, Seravalle G, Quarti-Trevano F, Dell'Oro R, Arenare F, Spaziani D, Mancia G. Sympathetic and baroreflex cardiovascular control in hypertension-related left ventricular dysfunction. Hypertension 2009;53:205–9. [5] Macefield VG, Rundqvist B, Sverrisdottir YH, Wallin BG, Elam M. Firing properties of single muscle vasoconstrictor neurons in the sympathoexitation associated with congestive heart failure. Circulation 1999;100:1708–13. [6] Floras JS. Arterial baroreceptor and cardiopulmonary reflex control of sympathetic outflow in human heart failure. Ann. NY Acad. Sci. 2001;940:500–13. [7] Neumann J, Ligtenberg G, Kleinm IL, Koomans HA, Blankestijn PJ. Sympathetic hyperactivity in chronic kidney disease: Pathogenesis, clinical relevance, and treatment. 2004. Kidney Int. 2004;65:1568–76. [8] Lambert E, Dawood T, Straznicky N, Sari C, Schlaich M, Esler M, Lambert G.et Association between the sympathetic firing pattern and anxiety level in patients with the metabolic syndrome and elevated blood pressure. J. Hypertens. 2010;28:543–50. [9] Elefteriou F, Ahn JD, Takeda S, Starbuck M, Yang X, Kiu X, et. al. Leptin regulation of bone resorption by the sympathetic nervous system and CART. Nature 2005;434(7032):514–20. [10] Mano T, Nishimura N, Iwase S. Sympathetic neural influence on bone metabolism in microgravity. Acta Physiol. Hung. 2010;97:354–61.

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82 Clinical Sympathetic Imaging David S. Goldstein Until recently, clinical laboratory means to test patients with known or suspected abnormalities of the autonomic nervous system consisted of physiological independent manipulations (e.g., heat exposure, the Valsalva maneuver, tilt table testing) and physiological dependent measures (e.g., sweating, blood pressure, heart rate and heart rate variability, skin and core temperature, forearm vascular resistance, skin electrical conductance, cutaneous humidity, microcirculatory blood flow, skeletal muscle sympathetic microneurography). Since the late 1970s clinical research on autonomic disorders has also included neurochemical measures (e.g., plasma norepinephrine during supine rest and orthostasis, norepinephrine spillover, dihydroxyphenylglycol, neuronal nicotinic receptor antibodies) and physiological or neurochemical effects of neuropharmacological agents (e.g., tyramine, clonidine, yohimbine, desipramine, trimethaphan, isoproterenol, glucagon, reboxetine, and acetylcholine in the quantitative sudomotor axon reflex test). Sympathetic imaging provides an important supplement to physiological, neurochemical, and neuropharmacological approaches in the evaluation of patients with clinical autonomic disorders. Sympathetic imaging using planar scintigraphy became available in the 1980s [1], followed by imaging using single photon emission computed tomography (SPECT) and positron emission tomography (PET). This chapter teaches concepts underlying sympathetic imaging, summarizes methods and agents used, highlights some applications of clinical sympathetic imaging in dysautonomias, and emphasizes the utility of sympathetic imaging to detect cardiac sympathetic denervation in Lewy body diseases. Neuroimaging assessments can be categorized in terms of anatomic, functional non-specific, and functional specific. Examples of anatomic neuroimaging are computed tomography and magnetic resonance imaging of the brain. Functional non-specific approaches include functional magnetic resonance imaging and assessments of regional perfusion or metabolism by PET after administration of 18 F-fluorodeoxyglucose, 13N-ammonia, or 15O-water. None of these specifically visualizes sympathetic innervation or function. Functional specific methods examine transporters, receptors, or uptake and storage of neurotransmitters. For instance, measurement of putamen 6-[18F]

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00082-2

fluorodopa-derived radioactivity enables assessment of dopaminergic innervation.

SYMPATHETIC IMAGING METHODS AND AGENTS Almost all sympathetic imaging involves visualization of noradrenergic innervation in the left ventricular myocardium. This is because the heart is a relatively large, solid, homogeneous organ with a high concentration of sympathetic nerve fibers that surround the myocardial cells. Cardiac sympathetic images therefore look like myocardial perfusion scans. Other thoracic organs and tissues possess much lower concentrations of sympathetic nerves. By far the most commonly used means world-wide to assess cardiac sympathetic innervation by imaging is SPECT scanning after injection of the sympathomimetic amine 123I-metaiodobenzylguanidine (123I-MIBG, Fig. 82.1). The principle underlying 123I-MIBG scanning, as well as sympathetic imaging by PET imaging agents, is not binding of the tracer-labeled compound to receptors on target cells but physical translocation of the compound into the sympathetic nerves via the cell membrane norepinephrine transporter and then subsequent active transport of the cytosolic compound into vesicles via the vesicular monoamine transporter (Fig. 82.2). Thus, sympathetic imaging depends on radiolabeling of vesicles in sympathetic nerves. Cardiac sympathetic imaging by 123I-MIBG scanning has been used extensively in Japan, to a lesser extent in Europe, and hardly at all in the United States. The dearth of American experience seems to be the result of a cycle involving lack of insurance coverage because of lack of clinical trials because of lack of availability because of lack of insurance coverage. 123I-MIBG scanning is available at many centers in the United States for diagnostic evaluation of pheochromocytoma, since third party payers generally cover this application. 123 I-MIBG is an analog of the sympathomimetic amine, guanethidine, which differs structurally from norepinephrine, the neurotransmitter that mediates sympathetic regulation of the cardiovascular system (Fig. 82.1). Injected MIBG undergoes quite a different fate from catecholamines

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FIGURE 82.1 Chemical structures of compounds related to clinical sympathetic imaging. The compounds can be classified in terms of sympathomimetic amines and catecholamines.

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FIGURE 82.2 Contrasting fates of sympathomimetic amines and catecholamines used for clinical sympathetic imaging. Abbreviations: COMT, catechol-O-methyltransferase; Exo., exocytosis; MAO, monoamine oxidase; NET, cell membrane norepinephrine transporter; U-2, Uptake-2; VMAT, vesicular monoamine transporter.

such as norepinephrine (Fig. 82.2). These differences must be considered when interpreting changes in 123I-MIBGderived radioactivity in terms of sympathetic neuronal function. First, MIBG is not as avidly or selectively removed by sympathetic nerves as are catecholamines. Second, MIBG that is taken up into the sympathoneural axoplasm is not a substrate for monoamine oxidase, the main enzyme metabolizing cytosolic catecholamines in

sympathetic nerves. Third, MIBG is not a substrate for catechol-O-methyltransferase, a major enzyme metabolizing catecholamines in non-neuronal cells. The metabolic fate of MIBG is poorly understood. Since 123I is not a positron-generating isotope, the usual method for detecting 123I-MIBG-derived radioactivity is SPECT, not PET scanning. After intravenous injection of 123I-MIBG, a relatively long time is required

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FIGURE

82.3 Diagram of “uptake” and “washout” of I-metaiodobenzylguanidine-derived radioactivity. Abbreviation: H:M, heart-mediastinum ratio. 123

for obtaining interpretable images. 123I-MIBG “uptake” is measured about 15–30 minutes after injection of the tracer (Fig. 82.3). This is long after myocardial uptake of tracer-labeled catecholamines is complete. 123I-MIBG “washout” is measured 3–4 hours after tracer injection. Radioactivity concentrations in SPECT scans are difficult to calibrate, and so myocardial concentrations of 123 I-MIBG-derived radioactivity are usually expressed in terms of heart:mediastinum (H:M) ratios. In patients with coronary arterial or microvascular narrowing, as in atherosclerotic cardiovascular disease and diabetes mellitus, low H:M ratios of 123I-MIBG-derived radioactivity might reflect decreased delivery of the imaging agent by blood perfusion; 123I-MIBG SPECT studies rarely take this into account. PET scanning offers potential advantages over SPECT scanning. Spatial resolution is better, the amount of injected radioactivity is smaller, radioactivity concentrations in tissues can be measured in absolute terms, and analyses of curves relating tissue radioactivity detected by PET scanning over time (time–activity curves) can yield information about not only innervation but also function. Several agents have been developed for sympathetic imaging by PET scanning. These include 11C-epinephrine, 11 6-[18F]fluorodopamine, C-hydroxyephedrine, and 18 11 F-metaraminol. C-Epinephrine and 6-[18F]fluorodopamine are catecholamines, whereas 11C-hydroxyephedrine and 18F-metaraminol are non-catecholamine sympathomimetic amines (Fig. 82.1). All are investigational agents, and none has been approved yet for clinical diagnostic purposes. Although 6-[18F]fluorodopa, which as noted above is used to assess striatal dopaminergic innervation in neurodegenerative disorders such as Parkinson’s disease, is converted to 6-[18F]fluorodopamine in sympathetic nerves, 6-[18F]fluorodopa is not a sympathetic imaging agent. Structurally, 6-[18F]fluorodopa is a neutral amino acid, not a catecholamine, and so 6-[18F]fluorodopa is taken up by all cells via the neutral amino acid transporter. 6-[18F]Fluorodopa in non-neuronal cells may be converted to 6-[18F]fluorodopamine by L-aromaticamino-acid decarboxylase, but this is probably a minor

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fate compared to incorporation into peptides or proteins and enzymatic breakdown independent of catecholamine formation. After intravenous injection of 6-[18F]fluorodopa, plasma concentrations of 6-[18F]fluorodopamine are too low to enable visualization of cardiac sympathetic innervation.

SYMPATHETIC IMAGING IN DYSAUTONOMIAS The term, “dysautonomias”, has been used to refer to conditions in which altered activity of one or more components of the autonomic nervous system adversely affects health. The following discussion focuses on dysautonomias that involve functional abnormalities or loss of sympathetic noradrenergic nerves. One of the first forms of dysautonomia to be evaluated by clinical sympathetic imaging was diabetic autonomic neuropathy. In general, patients with diabetic autonomic neuropathy have decreased H:M ratios of 123I-MIBGderived radioactivity, especially in delayed images. A major factor complicating interpretation of the results in terms of sympathetic denervation is that patients with diabetic autonomic neuropathy have a high frequency of coronary arterial and arteriolar narrowing, so that low H:M ratios 123I-MIBG-derived radioactivity may reflect a combination of denervation and decreased delivery of the tracer to sympathetic nerves via coronary hypoperfusion. Analyses of 123I-MIBG SPECT scans rarely if ever adjust 123I-MIBG-derived radioactivity for perfusion in the same regions of interest. 11C-Hydroxyephedrine PET scanning, with perfusion assessed by 13NH3 scanning in the same subjects, has revealed imaging evidence for heterogeneously decreased sympathetic innervation in relatively distal (apical, inferior, lateral) cardiac segments [2]. A study of painful diabetic neuropathy of the feet reported decreased local concentrations of perfusion-adjusted 6-[18F]fluorodopamine-derived radioactivity, along with decreased entry of norepinephrine into the venous drainage, indicating local sympathetic denervation [3]. Low H:M ratios of 123I-MIBG-derived radioactivity or increased rates of “washout” of cardiac 123I-MIBGderived radioactivity have been reported in many disorders, including cardiovascular conditions (congestive heart failure, myocardial infarction, ventricular arrhythmias, dilated cardiomyopathy, Chagas disease, essential hypertension with left ventricular hypertrophy, Brugada syndrome), emotional distress, liver or kidney failure, hypothyroidism, and neurodegeneration (Parkinson’s disease, pure autonomic failure, Lewy body dementia). In general, decreased “uptake” and increased “washout” of cardiac 123I-MIBG-derived radioactivity are associated with relatively poor prognosis. This pattern may reflect increased cardiac sympathetic nerve traffic, decreased vesicular sequestration of cytoplasmic amines, or decreased activity of the cell membrane norepinephrine

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82. ClInICAl SymPATHETIC ImAgIng

transporter, abnormalities that may coexist and interact to produce multiple harmful positive feedback loops. Normal

Sporadic

PARK1

PARK4

ASSOCIATION OF NORADRENERGIC DENERVATION WITH SYNUCLEINOPATHY IN LEWY BODY DISEASES Lewy body diseases such as Parkinson’s disease, dementia with Lewy bodies, and pure autonomic failure are characterized by intra-neuronal precipitates of the protein alpha-synuclein. Such diseases are now subsumed under the heading of synucleinopathies. Multiple system atrophy is also considered to be a form of synucleinopathy; however, in multiple system atrophy alphasynuclein deposits are found in glial cells. Lewy body synucleinopathies are associated with imaging evidence of substantial cardiac sympathetic denervation, both by 123I-MIBG SPECT and by 11C-hydroxyephedrine and 6-[18F]fluorodopamine PET. In these diseases cardiac noradrenergic denervation has been confirmed by profoundly decreased tyrosine hydroxylase immunoreactivity in epicardial nerves [4,5]. Moreover, non-motor findings such as dementia, loss of sense of smell (anosmia), REM behavior disorder, baroreflex failure, and orthostatic hypotension seem to be associated with cardiac noradrenergic denervation at least as closely as with striatal dopaminergic denervation [6]. In contrast, most (but not all) patients with multiple system atrophy have normal cardiac sympathetic innervation. Although the finding of imaging evidence for cardiac sympathetic denervation does not exclude multiple system atrophy, the finding of normal cardiac sympathetic innervation probably does exclude Parkinson’s disease with orthostatic hypotension. In the course of Parkinson’s disease, when does cardiac denervation occur? According to the theory proposed by Braak for the pathogenetic sequence [7], an agent is inhaled and ingested, resulting in early deposition of alpha-synuclein in the olfactory bulb and autonomic nerves, with subsequent ascending pathology in the autonomic ganglia, dorsal motor nucleus of the vagus in the caudal medulla, then rostral ventrolateral medulla, then pontine locus ceruleus, then the midbrain substantia nigra, and finally diffuse lesions in the cortex. This concept predicts that imaging evidence of cardiac sympathetic denervation may be a premotor biomarker of Parkinson’s disease. Although imaging evidence of cardiac sympathetic denervation can precede the movement disorder by several years [8], the frequency and consistency of this abnormality have not yet been determined. Cardiac sympathetic imaging and post-mortem neuropathologic findings have linked alpha-synucleinopathy with noradrenergic denervation in Lewy body diseases [9]. Thus, patients with familial Parkinson’s disease from abnormalities of the gene encoding alpha-synuclein have

FIGURE 82.4 PET images of the left ventricular myocardium after i.v. injection of 6-[18F]fluorodopamine in a normal control subject, a patient with sporadic Parkinson’s disease, a patient with familial Parkinson’s disease from mutation of the gene encoding alpha-synuclein (PARK1), and a patient with familial Parkinson’s disease from triplication of the gene encoding alpha-synuclein (PARK4). Note association between inherited alpha-synucleinopathy and neuroimaging evidence of cardiac sympathetic denervation in Parkinson’s disease.

cardiac sympathetic denervation (Fig. 82.4). The pathologic changes seem to progress in a retrograde, centripetal manner. Bases of the association of alpha-synucleinopathy with catecholaminergic denervation remain obscure but are being actively researched. According to the “catecholaldehyde hypothesis”, catecholaldehydes produced from enzymatic deamination of cytosolic catecholamines exert cytotoxic effects because of oxidative stress and oligomerization of alpha-synuclein, resulting in deleterious positive feedback loops. Especially because of the utility of cardiac sympathetic imaging in distinguishing Parkinson’s disease from multiple system atrophy in patients with clinical evidence of central neurodegeneration and orthostatic hypotension, sympathetic imaging seems a valuable addition to physiological, neuropharmacologic, and neurochemical approaches in the diagnosis of autonomic disorders.

References [1] Wieland DM, Brown LE, Rogers WL, Worthington KC, Wu JL, Clinthorne NH, et al. Myocardial imaging with a radioiodinated norepinephrine storage analog. J Nucl Med 1981;22:22–31. [2] Allman KC, Stevens MJ, Wieland DM, Hutchins GD, Wolfe Jr ER, Greene DA, et al. Noninvasive assessment of cardiac diabetic neuropathy by carbon-11 hydroxyephedrine and positron emission tomography. J Am Coll Cardiol 1993;22:1425–32. [3] Tack CJ, van Gurp PJ, Holmes C, Goldstein DS. Local sympathetic denervation in painful diabetic neuropathy. Diabetes 2002;51:3545–53. [4] Orimo S, Oka T, Miura H, Tsuchiya K, Mori F, Wakabayashi K, et al. Sympathetic cardiac denervation in Parkinson's disease and pure autonomic failure but not in multiple system atrophy. J Neurol Neurosurg Psychiatry 2002;73:776–7.

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ASSoCIATIon of noRADREnERgIC DEnERvATIon wITH SynuClEInoPATHy In lEwy BoDy DISEASES

[5] Orimo S, Amino T, Takahashi A, Kojo T, Uchihara T, Mori F, et al. Cardiac sympathetic denervation in Lewy body disease. Parkinsonism Relat Disord 2006;12(Suppl. 2):S99–S105. [6] Goldstein DS, Sewell L, Holmes C. Association of anosmia with autonomic failure in Parkinson disease. Neurology 2009;74:245–51. [7] Braak H, Ghebremedhin E, Rub U, Bratzke H, Del Tredici K. Stages in the development of Parkinson's disease-related pathology. Cell Tissue Res 2004;318:121–34.

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[8] Goldstein DS, Sharabi Y, Karp BI, Bentho O, Saleem A, Pacak K, et al. Cardiac sympathetic denervation preceding motor signs in Parkinson disease. Clin Auton Res 2007;17:118–21. [9] Orimo S, Uchihara T, Nakamura A, Mori F, Kakita A, Wakabayashi K, et al. Axonal alpha-synuclein aggregates herald centripetal degeneration of cardiac sympathetic nerve in Parkinson's disease. Brain 2008;131:642–50.

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C H A P T E R

83 Assessment of the Autonomic Control of the Cardiovascular System by a Frequency Domain Approach Raffaello Furlan, Franca Barbic INTRODUCTION The neural control of heart rate is mainly obtained at the level of the sino-atrial node by the interaction of sympathetic and vagal efferent discharge. In most conditions an increase of cardiac sympathetic activity is accompanied by a simultaneous inhibition of the vagal modulation to the heart and vice versa: hence the concept of sympathovagal balance. It is accepted that the instantaneous sympatho-vagal balance can be broadly assessed by quantifying RR interval variability. Similarly, arterial pressure spontaneously oscillates in part as a consequence of the sympathetic vasomotor control. The various types of blood pressure oscillations are represented in Figure 83.1. The second order oscillations are produced mechanically by the respiratory activity. The third order fluctuations, with a period of about 10 seconds, are related to vasomotion and are modulated by sympathetic activity. Thus, they increase during conditions associated with a sympathetic excitation such as on standing (tilt). Therefore, the study of circulatory rhythms by spectrum analysis techniques may furnish a valuable insight into the autonomic regulation of the cardiovascular system in health and disease.

METHODOLOGY Power spectrum analysis based on Fast Fourier Transform (FFT) or autoregressive modeling (AR) [2] provides the center frequency of rhythmic fluctuations of the different cardiovascular variables (i.e., heart rate, blood pressure, central venous pressure etc.), their time relationship (phase) and amplitude both in absolute and in normalized values [2]. Absolute values are computed as the integral of each oscillatory component (hatched area of Fig. 83.1). Normalization procedure is obtained by dividing the absolute power of each oscillatory component by total variance (minus the power of the frequency

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00083-4

components below 0.03 Hz) and then multiplying by 100 [2,3]. Normalization overcomes the problems due to marked changes in RR variance when comparing different subjects and experimental conditions [3]. The LFRR/HFRR ratio, which is independent of units of measure, assesses the sympathovagal instantaneous relationship (balance) [3].

CARDIOVASCULAR RHYTHMS AND AUTONOMIC NEURAL CONTROL Two major oscillatory components are observed in healthy humans in recumbent position [3]. One is the high frequency (HF ≈0.25 Hz) component. If extracted from RR variability (HFRR), it is an accepted index of the vagal modulation of the sino-atrial node activity [2]. Indeed, HFRR is reduced after muscarinic blockade [2] and is enhanced during the reflex increase of cardiac vagal activity obtained by phenylephrine administration. The HF component of systolic arterial pressure variability reflects the mechanical influence of respiratory activity. The other oscillatory component is indicated as LF (low frequency, ≈0.10 Hz). In the case of systolic arterial pressure variability, LFSAP is a marker of the sympathetic modulation of vasomotor activity [3]. Indeed, it is increased in animals during baroreflex unloading induced by nitroglycerine [4] and in humans during tilt test [5], moderate physical exercise and mental stress. The LF component of RR variability (LFRR) when expressed in normalized units (n.u.) reflects primarily the sympathetic efferent modulation of the sino-atrial node [2,3]. In conscious dogs, the reflex increase of sympathetic activity by nitroglycerin administration enhanced LFRR n.u. whereas the same stimulus after chronic bilateral stellectomy, that selectively abolishes cardiac sympathetic innervation, could not elicit any LFRR enhancement [4]. In humans, conditions characterized by an increased sympathetic activity such as the gravitational stimulus [5], mental and mild physical stresses, baroreflex unloading induced

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FIGURE 83.1 Examples of arterial pressure spontaneous fluctuations. Notice differences in both the amplitude and the period of the rhythmic oscillations. First order fluctuations, generated by cardiac activity, are characterized by the inter-beat period and their amplitude is represented by the differential pressure. A period of 24 hours is the feature of day-night oscillations of systemic pressure. Second and third order fluctuations define the so-called short-term variability mostly related to neural influences.

by nitroprusside administration and lower body negative pressure [6] were associated with a remarkable increase of the normalized power of LFRR. Both acute [7] and chronic [3] beta-blocker assumption reduced LFRR in n.u. The same oscillatory component was undetectable in subjects with pure autonomic failure, a condition characterized by the degeneration of sympathetic efferent neurons. Rhythmic discharge activity is a general property of the nervous system [1]. A 0.1 Hz rhythmicity linked with low frequency fluctuations of RR interval and arterial pressure variability was found to characterize sympathetic neurons located in areas within the central nervous system involved in cardiovascular regulation. Interestingly, these fluctuations remain after baroreflex afferent inputs were removed by a stabilizer device connected to the arterial system of the animal suggesting that baroreceptor activity is not necessary to the genesis of such fluctuations that, instead, may result from a central oscillator. Rhythmic LF and HF periodicities characterize the discharge activity of postganglionic sympathetic fibers of humans (muscle sympathetic nerve activity, MSNA) [5]. In man, the sympathetic activation induced by a gravitational stimulus was accompanied by a marked increase of the 0.1 Hz oscillatory component of MSNA, resembling the changes observed in the same oscillatory component of RR and systolic arterial pressure variability [5]. In addition, the

linear coupling between LF fluctuations of MSNA and heart rate, and MSNA and systolic arterial pressure was increased during the tilt maneuver suggesting the onset of a common oscillatory pattern based on prevailing 0.1 Hz fluctuations which involve the different variables [5].

DIFFERENTIAL NEURAL CONTROL OF CARDIOVASCULAR VARIABLES AND OF THEIR SPONTANEOUS VARIABILITY Frequency domain analysis of heart rate and arterial pressure variability enabled to disclose subtle changes in the neural cardiovascular control that would be hidden when considering the simple variations of heart rate and blood pressure mean values, i.e., when approaching the problem by a conventional time domain analysis. There are several physiological and pathophysiological conditions supporting this concept. In a study performed on ambulant individuals based on spectral analysis of heart rate and arterial pressure variability over 24 hours [8], LFRR in n.u. and LFSAP were reduced during sleeping hours and were high during day-time. Interestingly, awakening in the early morning, while subjects were still lying in the bed, was associated with a remarkable increase in LFSAP compared to night-time, in absence of significant

VI. CARDIOVASCULAR DISORDERS

DIFFEREnTIAl nEuRAl ConTRol oF CARDIovASCulAR vARIAblES AnD oF THEIR SPonTAnEouS vARIAbIlITy

FIGURE 83.2 Example of beat by beat changes of RR interval and spectral markers of cardiac autonomic modulation LFRR and HFRR as observed during tilt in a subject with neurally-mediated syncope. Latency (L, 77 seconds) is defined as the time lag between maximum reached by LFRR during tilt and the onset of bradycardia. Notice that a slow decay of LFRR7, suggesting progressive sympathetic inhibition, and the increase of the vagal related index HFRR preceded bradycardia. Latency was clinically characterized by pre-syncope signs and symptoms. (Modified from ref. 10.)

changes in the values of systolic arterial pressure. This suggests that an increase of the sympathetic activity to the vessels may be primarily mirrored by a change in blood pressure variability and not necessarily result in an enhancement of systolic arterial pressure values. Similarly, patients with Parkinson’s disease without orthostatic hypotension [9] were found to be indistinguishable from healthy controls during the upright posture on the basis of the simple hemodynamic pattern. The frequency domain analysis of systolic arterial pressure variability disclosed lower values of LFSAP compared to healthy individuals during a 75° head-up tilt. This finding is consistent with the presence of early and subtle abnormalities in the sympathetic vasomotor control in patients with Parkinson’s disease even if they were not suffering from symptoms due to orthostatic intolerance nor from orthostatic hypotension. Notably, about 50% of patients with long lasting Parkinson’s disease suffer from clinical signs of dysautonomia including orthostatic hypotension. It is likely that the progressive neural degeneration characterizing Parkinson’s disease may result in early changes in the blood pressure variability. Subsequently, orthostatic

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hypotension appears when the loss of autonomic neurons becomes critical. There is agreement that lower body negative pressure (LBNP) values  –20 mmHg, by markedly decreasing central venous pressure, induce a reflex increase of heart rate mediated by both cardiopulmonary and arterial baroreceptor areas to maintain systemic arterial pressure. From this observation there might appear to be a lack of reflex sympathetic modulation to the heart in response to mild decrease of central venous pressure (i.e., in response to LBNP  –20 mmHg). In healthy volunteers we found that low intensity (–15 mmHg) LBNP elicited a reflex increase of cardiac sympathetic modulation, as evaluated by an enhancement of LFRR, in the absence of concomitant changes in heart rate, systolic arterial pressure and in the indexes of arterial baroreflex control [6]. Thus, the autonomic control of heart beat seemed to result in earlier effects on heart period variability and retarded influences on the values of heart rate. These latter could be observed only for LBNP higher than –20 mmHg [6]. Finally, in a study based on time variant spectrum analysis of RR variability, the two spectral components LFRR and HFRR were stable during a 15-minute lasting tilt maneuver in healthy subjects [10]. However, in a subgroup of patients suffering from a neurally-mediated syncope, the cardiac autonomic modulation preceding the loss of consciousness was characterized by a progressive reduction of the index of sympathetic modulation (LFRR) of the seno-atrial node activity and by a concomitant increase of the index of cardiac vagal modulation HFRR (see Fig. 83.2). Also, there was a time lag of about 1 minute duration between the starts of both decrease in cardiac sympathetic modulation and increase in cardiac vagal activity and the onset of bradycardia. Interestingly, this pattern in the indexes of cardiac autonomic function was associated with unchanged values of heart rate compared with asymptomatic phase of tilt and with a concomitant presence of pre-syncope signs and symptoms such as pallor, sweating, yawning, nausea, blurred vision and dizziness [10] suggestive of a remarkable underlying autonomic modification.

References [1] Malliani A. The sympathovagal balance explored in the frequency domain. In: Principles of Cardiovascular Neural Regulation in Health and Disease. Boston/Dordrecht/London: Kluwer Academic Publishers. 2000. pp. 65-107. [2] Task Force of the European Society of Cardiology and the North American Society of Pacing and Electrophysiology. Heart rate variability: standards of measurements, physiological interpretation, and clinical use. Circulation 1996;93:1043–65. [3] Pagani M, Lombardi F, Guzzetti S, Rimoldi O, Furlan R, Pizzinelli P, et al. Power spectral analysis of heart rate and arterial pressure variabilities as a marker of sympatho-vagal interaction in man and conscious dog. Circ Res 1986;59:178–93. [4] Rimoldi O, Pierini S, Ferrari A, Cerutti S, Pagani M, Malliani A. Analysis of short term oscillations of R-R and arterial pressure in conscious dogs. Am J Physiol 1990;258:H967–76.

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[5] Furlan R, Porta A, Costa F, Tank J, Baker L, Schiavi R, et al. Oscillatory Patterns in Sympathetic Neural Discharge and Cardiovascular Variables During Orthostatic Stimulus. Circulation 2000;101:886–92. [6] Furlan R, Jacob G, Palazzolo L, Rimoldi A, Diedrich A, Harris P, et al. Sequential Modulation of Cardiac Autonomic Control Induced by Cardiopulmonary and Arterial Baroreflex Mechanisms. Circulation 2001;104:2932–7. [7] Cogliati C, Cogliati C, Colombo S, Gnecchi Ruscone T, Gruosso D, Porta A, et al. Acute β-blockade increases muscle sympathetic activity and modifies its frequency distribution. Circulation 2004;110:2786–91.

[8] Furlan R, Guzzetti S, Crivellaro W, Dassi S, Tinelli M, Baselli G, et al. Continuous 24-hour assessment of the neural regulation of systemic arterial pressure and RR variabilities in ambulant subjects. Circulation 1990;81:537–47. [9] Barbic F, Perego F, Canesi M, Gianni M, Biagiotti S, Pezzoli G, et al. Early abnormalities of vascular and cardiac autonomic control in Parkinson's disease without orthostatic hypotension. Hypertension 2007;49:120–6. [10] Furlan R, Piazza S, Dell'Orto S, Barbic F, Bianchi A, Mainardi L, et al. Cardiac autonomic patterns preceding occasional vasovagal reactions in healthy humans. Circulation 1998;98:1756–61.

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C H A P T E R

84 Assessment of Sudomotor Function Ronald Schondorf TESTS OF SUDOMOTOR FUNCTION Laboratory tests of sudomotor function are of value not only as descriptors and quantifiers of sudomotor and thermoregulatory capacity but also as one of the few methods by which the site of autonomic dysfunction can be localized [1,2]. Several measurement options are available. Changes in humidity are measured quantitatively with a hygrometer or qualitatively with indicator dyes (alizarin red or cornstarch iodine). Sweat gland number and activity are measured with indicator dyes [3] or silicone impressions [4]. Image analysis techniques can be applied to quantify these latter measurements and to correlate the temporal profile of changes in humidity with changes in sweat gland number and output. Changes in the electrical activity of the skin can be used to indicate the presence or absence of sudomotor activity and are most useful as a psychophysiological index of arousal [5]. Lastly, sophisticated morphological analysis of stained punch biopsies of the skin is beginning to provide clinical pathological correlates to the physiological measurements outlined above [6]. This methodology is not yet suitable for routine laboratory evaluation. The means through which sweat glands are activated help to localize the site of dysfunction. Direct stimulation with acetylcholine (Ach) or cholinergic agonists such as pilocarpine provides first order information concerning

functional integrity of the sweat gland and its muscarinic innervation. Additional information concerning sweat gland innervation can be obtained by studying the sudomotor axon reflex described in greater detail below. Placing the patient in a heated environment to provoke diffuse thermoregulatory sweating provides maximal information regarding the topographic distribution of sudomotor dysfunction but does not discriminate between preganglionic and postganglionic sites of dysfunction [1].

AXON REFLEX TESTING The axon reflex (schematically shown in Fig. 84.1) occurs when ACh stimulates nicotinic receptors on the postganglionic sympathetic sudomotor axon. The evoked impulse travels antidromically, reaches a branch-point, and then travels orthodromically to mediate sweating by binding to M3 muscarinic receptors on the gland itself. Cessation of the sweat response occurs when acetylcholinesterase cleaves ACh to acetate and choline. Despite the rapid destruction of Ach there is evidence that axon reflex activation of sweat glands propagates for a distance of several cm and for some minutes after local sweating has terminated [7]. The degree of propagation depends on the initial site of stimulation [8]. These and other data suggest that sweat glands are locally interconnected and activated

FIGURE 84.1 QSART. Stimulus of nerve terminals by acetylcholine iontophoresis results in activation of postganglionic sympathetic sudomotor fibers, acetylcholine release and activation of a new population of sweat glands.

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by a dense network of interwoven sudomotor axons that allow propagation of local sweating until local concentrations of Ach drop below a critical level [7]. One of the commonly accepted measures of axon reflex sweating is the quantitative sudomotor axon reflex test or QSART. The standard Ach stimulus that evokes this response is 5 minutes of constant anodal current iontopheresis of 10% Ach in the outer compartment of a multicompartmental sweat cell. The sweat response is recorded for 10–15 minutes with a hygrometer from a second inner compartment isolated from the site of iontopheresis (see Fig. 84.2). Four skin sites are tested, three on the lower extremity and one on the upper extremity. The morphology of the response, latency to onset and integrated sweat response are measured. An extensive normative database from the Mayo Clinic Autonomic Reflex Laboratory for the QSART response has been published [9]. These data have been recently compared with responses simultaneously recorded from the contralateral limbs of normal subjects with a commercially available device [10]. The volumes from the commercial device were approximately half those of the previously published norms although in all other respects responses were similar. The reasons for this difference have not been elucidated [10]. Clearly caution is required with any change in technique and active laboratories should obtain their own normative data. Patterns of abnormality (assuming adequate delivery of Ach) include absence or reduction of integrated sweat response with prolongation of latency to onset. Sometimes single sites are abnormal. Alternatively there is a pattern of dysfunction in the lower limb with amplified sweat response in the upper limb suggestive of a length dependent dying back neuropathy. Milder forms

of postganglionic sudomotor dysfunction may be associated with a sustained “hung up” response to stimulation suggesting hyperexcitability or reverberation within the axon reflex [7,11]. Factors that influence the magnitude of the QSART response include: number of sudomotor fibers activated by Ach, number of responding sweat glands and individual glandular sweat production. The QSART declines with age and is consistently smaller in women [12]. In most preganglionic or central disorders, the QSART is unimpaired although with increasing duration of the preganglionic lesion, the QSART may be reduced. The QSART provides high temporal resolution information regarding sweat gland output but limited information about sweat gland number and activity. Silicone impressions can be made from the skin after Ach iontopheresis is complete and the skin dried [13]. Sweat gland activity from the skin overlying the iontopheresis electrode is a consequence of direct cholinergic stimulation of sweat glands and of activation of the sudomotor axon reflex whereas more remote sweating is a consequence of the axon reflex alone. Image analysis of these impressions provides important information about the final numbers of sweat glands activated but gives no insight as to the temporal profile of sweat gland recruitment [1]. A recent attempt to circumvent this problem has resulted in a new testing paradigm – the quantitative direct and indirect test of sudomotor function or QDIRT [3]. Following the 5-minute iontopheresis, the skin is dried and dusted with indicator. Digital photographs of the sweat response are taken every 15 seconds and analyzed with image processing software. This technique provides better temporal resolution than silicon impressions but the number of sweat glands detected with this technique

FIGURE 84.2 Multicompartmental sweat cell. Stimulus (acetylcholine) is iontophoresed through C and recorded in A. A and C are separated by air gap (B). Acetylcholine is administered through cannula E and current in applied via anode F. A stream of desiccated air is run in via D to evaporate sweat. The capsule is held in place with straps applied to posts G.

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ConCluSIon

is substantially less than the silicon method. This may be due to lack of detection of minimal sweating with indicator dyes and to saturation of large sweat responses once detected. This technique may be of interest to clinicians who need a qualitative index of peripheral sweating and cannot afford the more expensive hygrometers needed for the QSART.

disorders. As an example, patients with multiple system atrophy are more likely to exhibit substantial anhydrosis when compared to patients with Parkinson’s disease who have significant autonomic failure. Anhydrosis may be distal, global or regional reflecting the site of the lesion.

SKIN POTENTIALS

The ideal test of sudomotor function would combine the topographical information provided by the TST with the exquisite localization provided by axon reflex sweating and the morphological and neurochemical information from skin punch biopsies. Absent this, evaluation of sudomotor function must rely on judicious use of available tests complemented by clinical acumen.

The sympathetic skin response (SSR) is a polysynaptic reflex that requires integrity of hypothalamic, brainstem and spinal circuits, and postganglionic sympathetic sudomotor axons. The SSR is generated in deep layers of the skin by sympathetically-mediated activation of sweat glands. The morphology of the SSR potential is determined by the interaction between these sweat glands and the surrounding epidermal tissue. At normal ambient temperatures SSR is recorded from the glabrous skin of the palms and soles. This activity is referenced to the adjacent hairy skin whose sweat glands are not typically active under these conditions. In routine clinical practice, the SSR is recorded following stimulation of median or posterior tibial nerve afferents at an intensity at three least times sensory threshold. In cases of peripheral neuropathy where afferent input may be insufficient to evoke an SSR, the response to acoustic stimuli or to an inspiratory gasp should also be recorded. Although the SSR is easy to perform using standard EMG equipment, its clinical utility in the evaluation of sympathetic sudomotor function is questionable. The criteria for what constitutes an abnormal SSR are controversial. Some consider absence of an SSR as an abnormal result, but SSRs may not be obtained from the lower extremities in 50% of normal subjects above 60 years. There is often very little correlation between severity of autonomic dysfunction and presence or absence of SSR. The SSR specifically tests sudomotor fibers that do not participate in thermoregulation and hence the correlation between presence or absence of SSR and other modalities of measurement of sudomotor function is poor [1,14].

THERMOREGULATORY SWEATING The thermoregulatory sweat test (TST) is a sensitive qualitative test of sudomotor function that provides important information on the pattern and degree of sweat loss [1]. A color indicator powder is applied onto dry skin. The environmental temperature is raised until an adequate core temperature rise is attained and the presence of sweating causes a change in the indicator color. The TST may be partially quantitated by estimating the percent of anterior surface anhydrosis. This percentage has proven to be a useful parameter in differentiating autonomic

CONCLUSION

References [1] Illigens BM, Gibbons CH. Sweat testing to evaluate autonomic function. Clin Auton Res 2009;19:79–87. [2] Kimpinski K, Iodice V, Sandroni P, Fealey RD, Vernino S, Low PA. Sudomotor dysfunction in autoimmune autonomic ganglionopathy. Neurology 2009;73:1501–6. [3] Gibbons CH, Illigens BM, Centi J, Freeman R. QDIRT: Quantitative direct and indirect test of sudomotor function. Neurology 2008;70:2299–304. [4] Kennedy WR, Sakuta M, Sutherland D, Goetz FC. Quantitation of the sweating deficiency in diabetes mellitus. Ann Neurol 1984;15:482–8. [5] Critchley HD, Melmed RN, Featherstone E, Mathias CJ, Dolan RJ. Volitional control of autonomic arousal: A functional magnetic resonance study. Neuroimage 2002;16:909–19. [6] Gibbons CH, Illigens BM, Wang N, Freeman R. Quantification of sudomotor innervation: A comparison of three methods. Muscle & Nerve 2010;42:112–9. [7] Schlereth T, Brosda N, Birklein F. Spreading of sudomotor axon reflexes in human skin. Neurology 2005;64:1417–21. [8] Schlereth T, Brosda N, Birklein F. Somatotopic arrangement of sudomotor axon reflex sweating in humans. Auton Neurosci 2005;123:76–81. [9] Low PA, Deng JC, Opfer-Gehrking TL, Dyck PJ, O’Brien PC, Slezak JM. Effect of age and gender on sudomotor and cardiovagal function and blood pressure response to tilt in normal subjects. Muscle & Nerve 1997;20:1561–8. [10] Sletten, D.M., Weigand, S.D., Low, P.A. Relationship of Q-sweat to quantitative sudomotor axon reflex test (QSART) volumes 2009. Muscle & Nerve 2010;41:240–6. [11] Schlereth T, Dittmar JO, Seewald B, Birklein F. Peripheral amplification of sweating – a role for calcitonin gene-related peptide. J Physiol 2006;576:823–32. [12] Low PA, Opfer-Gehrking TL, Proper CJ, Zimmerman I. The effect of aging on cardiac autonomic and postganglionic sudomotor function. Muscle & Nerve 1990;13:152–7. [13] Kihara M, Opfer-Gehrking TL, Low PA. Comparison of directly stimulated with axon-reflex-mediated sudomotor responses in human subjects and in patients with diabetes. Muscle & Nerve 1993;16:655–60. [14] Schondorf R. The role of sympathetic skin responses in the assessment of autonomic function. In: Low PA, editor. Clinical autonomic disorders: Evaluation and management. Philadelphia: LippincottRaven; 1993. p. 221–31.

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C H A P T E R

85 Cutaneous Autonomic Innervation: Assessment by Skin Biopsy Christopher H. Gibbons, Roy Freeman INTRODUCTION The availability of punch skin biopsies stained with specific neuronal markers has ushered in a new era in the investigation of autonomic and peripheral nerve fibers. Although neurocutaneous investigation of skin biopsies was originally developed to study unmyelinated sensory nerve fibers in the assessment of small fiber neuropathy, recent investigations have focused on the large number of autonomic nerve fibers that are present within dermal tissue. Peripheral adrenergic and cholinergic fibers are present and provide innervation to a number of organelles in the skin including hair follicles, blood vessels, sweat glands and arrector pili muscles. The simplicity with which skin biopsies may be obtained and the opportunity for repeated testing provides an unparalleled opportunity to study autonomic nerve fibers in health and disease states.

CUTANEOUS NEUROANATOMY Punch skin biopsies provide tissue from the epidermal and dermal layers, which are separated by a basement membrane. Unmyelinated nociceptive C fibers pierce the basement membrane to innervate the epidermis from a subdermal plexus that runs just below the basement membrane. Within deeper dermal tissue bundles of nerve fibers, both myelinated and unmyelinated, course through the tissue. Autonomic and sensory fibers innervate a number of structures with the dermal tissue (Fig. 85.1).

Sweat Glands Sweat glands are tubular structures located within the deeper dermal tissue that contain a rich network of capillaries and nerve fibers. The investigation of sweat gland innervation began simultaneously to the initial studies of cutaneous epidermal nociceptive C-fibers using skin biopsies [1]. Recently, the widespread availability of skin biopsies for evaluation of intra-epidermal sensory nerve fiber density combined with scientific advances in quantitative immunohistochemistry staining of specific neuronal subtypes have extended the utility of studies of sudomotor

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00085-8

innervation [2]. Clinical skin biopsies can be stained with the pan-axonal marker protein gene product (PGP) 9.5 and imaged by light microscopy to highlight the nerve fibers that surround the sweat gland tubules [2,3]. The nerve fibers that surround sweat gland tubules are primarily sympathetic cholinergic (Fig. 85.2A,C), although some sympathetic adrenergic and sensory fibers are present (Fig. 85.2D). In addition to a dense network of nerve fibers, there are also a large number of capillaries within sweat glands that have separate innervation (Fig. 85.2B). The complexity of sweat gland innervation limits the utility of descriptive and semi-quantitative methods for determining sudomotor density [3,4]. More recent studies have described both an unbiased stereologic and a rapid automated method for quantitation of sudomotor density [2,3]. Both techniques can differentiate groups of patients with diabetic neuropathy from healthy control subjects. The automated approach is much faster, but has larger confidence intervals, thereby reducing the utility in individual patients, but proving of value in larger group studies where time constraints may play a factor. Thus far, quantitation of sudomotor nerve fibers has been limited to healthy control subjects and patients with diabetes. Sudomotor fiber density correlates with both neurophysiologic sweat testing and questionnaire responses about sweat output.

Hair Follicles Hair follicles extend from the deeper dermal tissue, through the basement membrane and epithelial layer and extend beyond the border of the skin. The base of the hair follicle is deep within the dermal tissue, the hair follicle itself is anchored to the skin by arrector pili muscles and sebaceous glands. Hair follicles have large numbers of sensory fibers that circumferentially wrap around the base of the follicle and extend up the shaft in order to provide sensory feedback. The majority of the innervation to the hair follicle is sensory. Autonomic nerve fibers also innervate the base of the hair follicle. The innervation is primarily sympathetic cholinergic but there are some sympathetic adrenergic fibers noted as well. To date, there have been no efforts to quantify the density of sensory or autonomic nerve fibers around hair follicles.

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85. CuTAnEouS AuTonomIC InnERvATIon: ASSESSmEnT By SkIn BIoPSy

Subdermal plexus Epidermal layer Dermal layer

HF HF

SG

Intra-epidermal nerve fibers

FIGURE 85.1 Cutaneous neuroanatomy. A sample skin biopsy stained with the panaxonal marker protein gene product 9.5 (PGP 9.5 – the green immunofluorescent nerve fibers) and the sympathetic cholinergic marker vasoactive intestinal peptide (VIP – the red immunofluorescent stain). The nerve fibers that are sympathetic cholinergic will be visualized by both their red VIP and their green PGP 9.5 immunostaining, thus will appear gold in this picture. The image is taken by confocal microscopy. The epidermal layer (blue unbroken arrow) can be visualized by the faint red tinge and has green intra-epidermal nerve fibers piercing through from the dermis (blue dashed arrows). The dermal layer (blue unbroken arrows) contains hair follicles (HF), sweat glands (SG), blood vessels (BV) and neural bundles that run below the epidermal layer called the subdermal plexus.

Arrector Pilorum Muscles

Blood Vessels

Arrector pili muscles anchor hair follicles by attaching the shaft of the hair follicle to the dermal tissue. Upon stimulation, the contracting muscles cause piloerection with the formation of cutis anseri, or goose bumps. Pili muscles contain a relatively simple pattern of innervation when compared to other dermal organelles. Nerve fibers travel in parallel with the pili muscle and are easily identified through either light or confocal microscopy using a pan-axonal marker, such as PGP 9.5 [5]. Selective immunohistochemical stains specific for sympathetic adrenergic innervation reveal that the majority of nerve fibers within pili muscles are sympathetic adrenergic (Fig. 85.3A–C) [5,6]. In the first report of pilomotor quantitation, the number of nerve fibers per milimeter running in parallel with the length of the muscle is counted in cross-section [5]. The density of nerve fibers is reduced in diabetic subjects compared to controls using the pan-axonal marker PGP 9.5, a sympathetic adrenergic marker, dopamine beta hydroxylase and the sympathetic cholinergic marker vasoactive intestinal peptide. The authors also report selective loss of cholinergic or adrenergic nerve fiber innervation in patients with diabetes. Further studies confirming neurophysiologic outcomes and clinical correlations in patients with diabetes and other diseases are warranted.

Blood vessels of differing size are present throughout the dermal layer but do not extend into the epidermal region. The blood vessels pass randomly through the dermal tissue but are present in dense vascular networks within and around other dermal organelles (hair follicles, sweat glands and arrector pili muscles). Due to the complexity of cutaneous vasomotor innervation, the specific subtypes of nerve fibers that surround blood vessels have not been fully identified. Evidence from pathologic and physiologic studies suggest there is sensory, sympathetic (cholinergic and adrenergic) and parasympathetic fiber expression within cutaneous blood vessels (Fig. 85.4A & B) [7,8].

SKIN BIOPSIES TO EVALUATE SPECIFIC AUTONOMIC DISORDERS While initial reports of skin biopsies over the last two decades have focused on pathologic changes to sensory nerve fibers, more recent reports have examined autonomic innervation. Most of the literature on autonomic innervation in disease comes from patients with diabetes. Sudomotor and pilomotor nerve fiber densities have been

VI. CARDIOVASCULAR DISORDERS

SkIn BIoPSIES To EvAluATE SPECIfIC AuTonomIC DISoRDERS

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FIGURE 85.2 Sweat glands. In (A), a sweat gland (green) with accompanying innervation is seen. The red/gold nerve fibers are stained by from PGP 9.5, a pan-axonal marker, showing the innervation around the sweat gland. In (B), the vascular system through a typical skin biopsy is shown. The blood vessels (shown in green) are stained by the endothelial marker CD31. The sweat gland tubules are shown as a faint green, and are much larger than the blood vessels. The nerve fibers, shown in red, are stained with the pan-axonal marker PGP 9.5. In (C), the sympathetic cholinergic innervation (green) stained with vasoactive intestinal peptide is seen. In (D), the same sweat gland as (C) is shown, but the fibers are stained with tyrosine hydroxylase, a sympathetic adrenergic marker. Note that very little of the total innervation of the sweat gland has adrenergic innervation. White scale bars at the bottom of each image indicate 100 μm.

reported in healthy subjects and in patients with diabetes [2,3,5]. The data show a length dependent pattern to autonomic nerve fiber damage in diabetes, with the distal sites exhibiting the earliest and most severely reduced nerve fiber densities. The decline in both sudomotor and pilomotor nerve fiber density parallels overall neuropathy progression and is associated with longer duration of diabetes and higher glycosylated hemoglobin levels. These data suggest that structural studies of autonomic nerves may provide a clinical biomarker to track disease progression, monitor response to treatment and can be repeated over time with reproducibility. Autonomic cutaneous innervation has also been assessed in other diseases. For example, immunofluorescent imaging of autonomic nerve fibers was able to differentiate subjects with Ross syndrome from subjects with Holmes–Adie syndrome [9]. The patients with Ross syndrome had a complete absence of sympathetic cholinergic innervation in anhidrotic skin (when stained with the cholinergic marker vasoactive intestinal peptide), reduced sensory innervation and an absence of sympathetic cholinergic innervation within blood vessels, hair follicles and arrector pilorum muscles in both anhidrotic and hyperhidrotic skin.

In contrast, the patients with Holmes–Adie syndrome had numerous morphologic abnormalities to sympathetic cholinergic nerves but had preserved sympathetic cholinergic nerve fiber density [9]. This study suggested that while Ross and Holmes–Adie syndromes have clinical similarities they may be distinguished by skin biopsy. Sudomotor nerve fiber density has been examined in patients with the neurodegenerative diseases multiple system atrophy and Parkinson’s [10–13]. These diseases may have significant clinical overlap, both in motor and autonomic symptoms and signs, but different clinical courses and outcomes. The autonomic dysfunction in subjects with multiple system atrophy is believed to be preganglionic dysfunction, while in Parkinson’s patients it is postganglionic [14]. Skin biopsies in these two diseases lend credence to this theory; patients with Parkinson’s disease appear to have greater autonomic nerve fiber density reduction than those with multiple system atrophy [10,13]. The rare disorder of cold induced sweating type 1 (CISS1) results in profuse sweating in cold environments. The mutation in CISS1 has been isolated to the cytokine receptor-like factor 1 [15]. A report of a patient with CISS1 documents diffuse anhidrosis by thermoregulatory sweat

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85. CuTAnEouS AuTonomIC InnERvATIon: ASSESSmEnT By SkIn BIoPSy

FIGURE 85.4 Blood vessels. In (A), the network of blood vessels within a skin biopsy is shown. The blood vessels, shown in green, are stained with the endothelial marker CD31. In (B), a blood vessel with accompanying innervation is seen. The blood vessel, shown in green, is stained with CD31, while the nerve fiber, shown in red, is stained with the sympathetic adrenergic marker tyrosine hydroxylase. White scale bars at the bottom of each image indicate 100 μm.

FIGURE 85.3 Arrector pili muscles. In (A–C), the same arrector pilorum muscle is seen. In (A), the red nerve fibers are stained with the sympathetic adrenergic marker tyrosine hydroxylase. In (B), the green nerve fibers are stained with vasoactive intestinal peptide, a sympathetic cholinergic marker. In (C), the adrenergic (red) and cholinergic (green) fibers are seen in overlay. The nerve fibers travel in parallel to the muscle and are predominantly sympathetic adrenergic. White scale bars at the bottom of each image indicate 100 μm.

testing and skin biopsy findings of absent sudomotor cholinergic innervation and increased sudomotor adrenergic innervation. Taken together, these data suggest an embryologic failure of transition of sudomotor fibers from adrenergic to cholinergic. This brief case report highlights the value of skin biopsy in defining the structural underpinnings, pathogenesis and embryologic development of this disease. Patients with familial dysautonomia have normal postganglionic sweat production [16]. However, skin biopsies in these individuals reveals an appropriate number of sweat glands, although many are atrophic in appearance [17]. The sweat gland nerve fiber density is reduced by more than 50% compared to control subjects, despite a normal sweat response to acetylcholine iontophoresis.

VI. CARDIOVASCULAR DISORDERS

SummARy

The reason for the discrepancy between measures of sudomotor structure and function in familial dysautonomia is not known.

SUMMARY The addition of skin biopsies to existing measures of autonomic function will provide a valuable addition to structural assessment of the autonomic nervous system. Autonomic nerve fibers can be sampled over different regions to monitor focal damage, they can be easily repeated over time to document disease progression and response to treatment and may act as novel biomarkers in the study of disease modifying agents. Skin biopsies are simple to obtain although they do require advanced laboratory processing for immunohistochemical staining. The number of sites performing skin biopsies is increasing with several centers now investigating autonomic innervation in addition to traditional intra-epidermal nerve fiber density. Despite the recent advances, there are still many diseases of the autonomic nervous system with minimal or no structural information from skin biopsies. In addition, the standardization of techniques across different laboratories is necessary for results to be comparable. There is also a need for normative control data to adequately describe the range of expression in healthy individuals. As this data becomes available, the utility of skin biopsy for evaluation of autonomic structure will lead to novel understanding of pathophysiology and the development of treatments for diseases of the autonomic nervous system.

References [1] Kennedy WR, Wendelschafer-Crabb G, Brelje TC. Innervation and vasculature of human sweat glands: an immunohistochemistrylaser scanning confocal fluorescence microscopy study. J Neurosci 1994;14:6825–33. [2] Gibbons CH, Illigens BM, Wang N, Freeman R. Quantification of sweat gland innervation: a clinical-pathologic correlation. Neurology 2009;72:1479–86.

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[3] Gibbons CH, Illigens BM, Wang N, Freeman R. Quantification of sudomotor innervation: a comparison of three methods. Muscle & Nerve 2010;42:112–9. [4] Dabby R, Vaknine H, Gilad R, Djaldetti R, Sadeh M. Evaluation of cutaneous autonomic innervation in idiopathic sensory small-fiber neuropathy. J Peripher Nerv Syst 2007;12:98–101. [5] Nolano M, Provitera V, Caporaso G, Stancanelli A, Vitale DF, Santoro L. Quantification of pilomotor nerves: a new tool to evaluate autonomic involvement in diabetes. Neurology 2010;75:1089–97. [6] Gibbons CH, Wang N, Freeman R. Capsaicin induces degeneration of cutaneous autonomic nerve fibers. Ann Neurol 2010;68:888–98. [7] Ruocco I, Cuello AC, Parent A, Ribeiro-Da-Silva A. Skin blood vessels are simultaneously innervated by sensory, sympathetic, and parasympathetic fibers. J Comp Neurol 2002;448:323–36. [8] Ramien M, Ruocco I, Cuello AC, St-Louis M, Ribeiro-Da-Silva A. Parasympathetic nerve fibers invade the upper dermis following sensory denervation of the rat lower lip skin. J Comp Neurol 2004;469:83–95. [9] Nolano M, Provitera V, Perretti A, et al. Ross syndrome: a rare or a misknown disorder of thermoregulation? A skin innervation study on 12 subjects. Brain 2006;129:2119–31. [10] Dabby R, Djaldetti R, Shahmurov M, et al. Skin biopsy for assessment of autonomic denervation in Parkinson's disease. J Neural Transm 2006;113:1169–76. [11] Donadio V, Nolano M, Provitera V, et al. Skin sympathetic adrenergic innervation: an immunofluorescence confocal study. Ann Neurol 2006;59:376–81. [12] Nolano M, Provitera V, Estraneo A, et al. Sensory deficit in Parkinson's disease: evidence of a cutaneous denervation. Brain 2008;131:1903–11. [13] Donadio V, Nolano M, Elam M, et al. Anhidrosis in multiple system atrophy: a preganglionic sudomotor dysfunction? Mov Disord 2008;23:885–8. [14] Goldstein DS. Functional neuroimaging of sympathetic innervation of the heart. Ann NY Acad Sci 2004;1018:231–43. [15] Di LR, Nolano M, Boman H, et al. Central and peripheral autonomic failure in cold-induced sweating syndrome type 1. Neurology 2010;75:1567–9. [16] Bickel A, Axelrod FB, Marthol H, Schmelz M, Hilz MJ. Sudomotor function in familial dysautonomia. J Neurol Neurosurg Psychiatry 2004;75:275–9. [17] Hilz MJ, Axelrod FB, Bickel A, et al. Assessing function and pathology in familial dysautonomia: assessment of temperature perception, sweating and cutaneous innervation. Brain 2004;127:2090–8.

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86 Pheochromocytoma Graeme Eisenhofer, Jacques W.M. Lenders, William M. Manger Pheochromocytomas are rare but treacherous catecholamine-producing neuroendocrine tumors that arise from chromaffin cells of the adrenal medulla in about 85% of patients, and from extra-adrenal chromaffin tissue in the remaining cases. The latter are referred to as extra-adrenal pheochromocytomas or paragangliomas; these mainly form in the abdomen, but also occur at numerous other locations such as in the urinary bladder, mediastinum and head and neck regions. Head and neck paragangliomas rarely produce catecholamines. Most catecholamine-producing pheochromocytomas and paragangliomas are sporadic, but at least 30% result from germ-line mutations of several tumor-susceptibility genes (Table 86.1). Mutations of the rearranged during transfection (RET) gene in multiple endocrine neoplasia type 2 (MEN 2), of the von Hippel–Lindau (VHL) gene in VHL syndrome, the neurofibromatosis type 1 (NF1) gene in von Recklinghausen disease and of genes encoding succinate dehydrogenase (SDH) subunits B (SDHB) and D (SDHD) are the most well known causes of hereditary tumors. Mutations of the gene for SDH subunit C (SDHC) are a less frequent cause of paragangliomas. Mutations of genes encoding the SDH complex assembly factor 2 (SDHAF2), transmembrane protein 127 (TMEM127) and SDH subunit A (SDHA) were more recently identified as other hereditary causes of adrenal and extra-adrenal pheochromocytomas. This brings together a total of nine tumor susceptibility genes now recognized to be responsible for hereditary chromaffin cell tumors.

Recognition of the significant hereditary contribution to pheochromocytoma is leading to new understanding about how the tumors develop and the basis for their highly variable phenotypic features and clinical manifestations (Table 86.1). Pheochromocytomas in MEN 2 and NF1 form almost exclusively within the adrenals, produce mixtures of epinephrine and norepinephrine and are rarely malignant. Those in VHL syndrome also form mainly in the adrenals, but can present at extra-adrenal locations and do not produce significant amounts of epinephrine. In contrast tumors due to mutations of SDH genes have mainly extra-adrenal locations and produce either exclusively norepinephrine, mixtures of dopamine and norepinephrine or in some cases exclusively dopamine. Those due to mutations of the SDHB gene are associated with a particularly high risk of malignancy. Both hereditary and sporadic epinephrine-producing pheochromocytomas generally present at later ages than tumors that do not produce epinephrine. The latter tumors, such as those due to VHL and SDH mutations, are characterized by immature (undifferentiated) catecholamine biosynthetic and secretory pathways and show extensive differences in gene expression profiles compared to epinephrine-producing tumors. These distinguishing features appear to reflect activation of distinct tumorigenic pathways and development from different chromaffin progenitors with variable susceptibility to disease-causing mutations. The introduction of screening programs in patients with identified mutations of susceptibility genes for

TABLE 86.1 Phenotypic Features of Hereditary Pheochromocytomas and Paragangliomas According to Affected Gene Gene

Chromosomal Location

Catecholamine Phenotype

Adrenal Tumors

Extra-adrenal Tumors

Predisposition to Malignancy (%)

VHL

3p25-26

NE





5%

RET

10q11.2

NE/EPI



–

3%

NF-1

17q11.2

NE/EPI



–

11%

SDHB

1p36.13

NE/DA





60–85%

SDHD

11q23

NE/DA





3–15%

SDHC

1q23.3

Unknown

–



Unknown

SDHA

5p15

Unknown

–



Unknown

SDHAF2

11q12.2

Unknown

–



Unknown

TMEM127

2q11.2

Unknown



Unknown

Unknown

Catecholamine phenotypes are shown according to whether the tumors produce combinations of norepinephrine and epinephrine (NE/EPI), predominantly norepinephrine (NE) or combinations of norepinephrine and dopamine (NE/DA). Relative frequencies of adrenal and extra-adrenal tumors: – rare,  occasional,  moderate, and  predominant presence. Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00086-X

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86. PHEOCHROMOCyTOMA

pheochromocytoma is now leading to diagnosis of many cases of the tumor before increases in blood pressure or other signs and symptoms of catecholamine excess become apparent. Similarly, an increasing proportion of pheochromocytomas are now being diagnosed as a consequence of screening for the tumors in patents with abdominal masses detected incidentally during imaging procedures carried out for unrelated conditions. These patients are also often asymptomatic and normotensive. Pre-test prevalences of pheochromocytomas in the above patients are much higher than in those in whom suspicion is based on clinical manifestations of catecholamine excess. Patients presenting with signs and symptoms of catecholamine excess are nevertheless still by far the group most often tested for pheochromocytoma and in whom the diagnosis remains the most problematic. Due to the high prevalence of hypertension, but the rarity of the tumor and the non-specific and highly variable nature of manifestations of catecholamine excess, pheochromocytoma is frequently searched for in this group, but only occasionally found. More often the tumors remain unsuspected. In such patients sudden excess catecholamine secretion can lead to a very sudden appearance of clinical manifestations and lethal complications. Adding to the difficulties in diagnosis, the clinical presentation of pheochromocytoma can be highly variable, with similar signs and symptoms produced by numerous other clinical conditions (Box 86.1). Occasionally some relatively common disorders, rather than having the more usual etiologies, result from a catecholamine-producing tumor. Furthermore, differential diagnosis of these clinical chameleons can be further complicated by increases in plasma catecholamines. The crucial initial step for diagnosis is to first consider the possibility of the tumor. Thereafter it is important to

select the most appropriate diagnostic test. Because missing a tumor can have deadly consequences, one of the most important considerations in choice of diagnostic test is a high level of reliability that the selected test will provide a positive result in that rare patient with the tumor. This conversely also provides confidence that a negative result reliably excludes the tumor, thereby avoiding the need for multiple or repeated biochemical tests and sometimes costly and unnecessary imaging studies to rule out the presence of a tumor. Therefore, the initial work-up of a patient with suspected pheochromocytoma should include a suitably sensitive biochemical test. Development of appropriately sensitive diagnostic tests has followed from advances in understanding catecholamine metabolism. These advances included observations that adrenal medullary cells and pheochromocytoma tumor cells contain catechol-O-methyltransferase, the enzyme that converts norepinephrine to normetanephrine, epinephrine to metanephrine, and dopamine to methoxytyramine (Fig. 86.1A). Thus, the adrenal glands, not the more commonly considered liver and kidneys, represent the single largest site of catecholamine O-methylation, accounting for over 90% of all circulating metanephrine and about 23% of normetanephrine. Normally the O-methylation pathway represents a minor route of catecholamine metabolism; deamination of norepinephrine within sympathetic nerves is the major pathway (Fig. 86.1A). Intraneuronal deamination is followed by O-methylation of the deaminated metabolite in extraneuronal tissues and finally oxidation in the liver to vanillylmandelic acid, the major urinary metabolite of norepinephrine and epinephrine. In patients with pheochromocytoma, intra-tumoral O-methylation becomes a dominant pathway of catecholamine metabolism. Consequently, the presence of the tumor leads to relatively

BOX 86.1

DIFFERENTIAL DIAGNOSIS OF PHEOCHROMOCYTOMA

Cardiovascular Ischemic heart disease Heart failure* Cardiac arrhythmias

Autonomic or neurologic Baroreflex failure** Stroke* Diencephalic (autonomic) epilepsy Migraine or cluster headaches

Endocrine Systemic mastocytosis Menopause Hyperthyroidism

Hypoglycemia (e.g., due to insulinoma)* Medullary thyroid carcinoma Carcinoid*

Miscellaneous Anxiety, panic attacks Factitiously-produced hypertension (e.g., self-injection of epinephrine)** Monoamine oxidase inhibitors Illicit drugs (e.g., amphetamines, cocaine)* Over-the-counter sympathomimetics (e.g., ephedrine)* Porphyria Clinical conditions associated with mild to moderate* or occasionally severe** increases in catecholamines.

VII. CATECHOLAMINE DISORDERS

PHEOCHROMOCyTOMA

large increases in production of the O-methylated metabolites, compared to minor increases in deaminated metabolites (Fig. 86.1B). Due to the continuous high rate of intra-tumoral catecholamine O-methylation, and because some tumors secrete catecholamines episodically or in low amounts, patients with pheochromocytoma usually have relatively larger and more consistent increases of plasma normetanephrine or metanephrine than of the parent catecholamines. The above understanding has led to a paradigm shift in laboratory testing for pheochromocytoma away from

423

measurements of catecholamines to a focus on their O-methylated metabolites. The higher diagnostic sensitivity for measurements of plasma metanephrines than urinary or plasma catecholamines has been confirmed by numerous independent studies. Measurements of urinary fractionated metanephrines also offer relatively high diagnostic sensitivity. Provided measurements are accurate and appropriate reference intervals are employed, findings of normal plasma concentrations or urinary outputs of metanephrines effectively rule out a catecholamine-producing tumor so that no further testing is required.

FIGURE 86.1 Pathways for metabolism of norepinephrine (NE) and epinephrine (EPI) derived from sympathetic nerves and the adrenal chromaffin or pheochromocytoma tumor cells are shown in panel A. Intraneuronal deamination by monoamine oxidase (MAO) of NE leaking from storage granules or recaptured after release by sympathetic nerves is the primary pathway of catecholamine metabolism leading to formation of dihydroxyphenylglycol (DHPG), then methoxyhydroxyphenylglycol (MHPG) in extraneuronal cells and finally vanillylymandelic acid (VMA) in the liver. Extraneuronal metabolism is a relatively minor pathway, but is important for processing metabolites produced in sympathetic nerves and adrenal chromaffin cells. Presence of catechol-O-methyltransferase (COMT) in adrenal chromaffin or pheochromocytoma tumor cells leads to formation of normetanephrine (NMN) and metanephrine (MN) from catecholamines leaking from storage granules into the cytoplasm. The MN and NMN produced in extraneuronal tissues or adrenal chromaffin cells are either further metabolized by deamination or sulfate conjugation, the latter catalyzed phenolsulfotransferase type 1A3 (SULT1A3). Corresponding diagnostic signal strengths for NE, EPI, NMN, MN, DHPG and VMA in plasma or urine of 365 patients with pheochromocytoma compared to a reference group of 846 patients are shown in panel B.

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86. PHEOCHROMOCyTOMA

For patients with elevations of plasma or urinary metanephrines there still remain problems in distinguishing false- from true-positive test results. Therefore, followup testing to confirm or exclude the tumor is required in most patients with a positive result for these and other measurements of catecholamine excess. Due to the rarity of the tumor among patients tested, false-positive results can be expected to far outnumber true-positive results. Considerations should therefore be given to the extent of increase, which correlates positively with increasing probability of a tumor, and sources of false-positive results. Sources of false-positive results include the clinical condition of the patient, inappropriate sampling conditions (e.g., seated rather than supine position), laboratory error and medications likely to interfere with analytical results or increase levels of metanephrines. Once sources of false-positive results are excluded, the clonidine-suppression test provides a useful method for distinguishing false- from true-positive elevations of plasma normetanephrine. The test was first developed to distinguish tumoral from sympathoneuronal sources of elevated plasma norepinephrine. Further refinements involving additional measurements of plasma normetanephrine showed that the test could also be used in patients with elevated plasma concentrations of normetanephrine but normal or only slightly increased plasma concentrations of norepinephrine. A fall in plasma normetanephrine of more than 40% from baseline or to below the upper limits of reference intervals indicates sympathetic activation and excludes pheochromocytoma, whereas lack of suppression of normetanephrine or norepinephrine provides strong evidence for the tumor. Because of the high diagnostic sensitivity of present day biochemical tests there is now general agreement that tumor localization should usually only be initiated after confirmation of disease by biochemical testing. Localization is best first carried out by computed tomography or magnetic resonance imaging. Functional imaging may also be carried out, usually by scintigraphy with 123I-meta-iododenzylguandidine. There are also now a variety of other functional ligands that may be used in conjunction with single photon emission tomography or positron emission tomography. Once a pheochromocytoma is conclusively diagnosed, appropriate medical treatment should be initiated to block the actions of catecholamines before and during surgery. The alpha-adrenoceptor blocker, phenoxybenzamine, represents the drug most commonly used to prepare patients with pheochromocytoma for surgery. Beta-adrenoceptor blockers should never be employed without first blocking alpha-adrenoceptor mediated vasoconstriction. The goal of preoperative management is relaxation of the constricted vasculature, expansion of the reduced plasma volume, and normalization of blood pressure. The laparoscopic approach for tumor removal is now recommended over laparotomy as the method of choice for surgical resection of most abdominal

pheochromocytomas. Advantages of the latter include less postoperative pain, a shortened hospital stay and convalescent period, and improved cosmetic result. Pheochromocytomas of the chest, neck, and urinary bladder require special surgical procedures; otherwise, management is similar to that of abdominal tumors. Completeness of tumor resection should be confirmed after recovery from surgery (4 to 6 weeks) by biochemical testing with a return of previously elevated test results to normal. About 17% of patients operated for the tumor develop recurrent disease, and about half of these have evidence of malignancy. Due to this relatively high risk, biochemical testing should continue at yearly intervals. There continues to be no reliable histopathological methods for distinguishing benign from malignant pheochromocytoma; only the presence of metastases at sites where no chromaffin tissue should be expected (i.e., bones, liver, lungs and lymph nodes) establishes a definitive diagnosis of malignant pheochromocytoma. Although there are no markers for reliably predicting development of malignant pheochromocytoma, there are several factors associated with increased risk of malignancy, including presence of an SDHB mutation, large tumor size and extraadrenal location. There also continues to be no effective cure for malignant pheochromocytoma. Once malignancy is confirmed, the 5-year survival rate is about 50%. Thus, most treatments for malignant pheochromocytoma are palliative, with therapy generally directed at maintaining normal blood pressure. Radiopharmaceutical therapy using high doses of 131I-MIBG, which is transported into pheochromocytoma tumor cells via the cell membrane norepinephrine transporter, provides the most commonly utilized therapy. Overall about 75% of patients treated with 131I-MIBG show improvement in manifestations, 50% have reductions in hormonal activity, and 22% show objective tumor responses. Complete remissions are rare and progressive disease following 131I-MIBG therapy is common.

Further Reading Amar L, Bertherat J, Baudin E, Ajzenberg C, Bressac-de Paillerets B, Chabre O, et al. Genetic testing in pheochromocytoma or functional paraganglioma. J Clin Oncol 2005;23:8812–8. Eisenhofer G, Keiser H, Friberg P, Mezey E, Huynh TT, Hiremagalur B, et al. Plasma metanephrines are markers of pheochromocytoma produced by catechol-O-methyltransferase within tumors. J Clin Endocrinol Metab 1998;83(6):2175–85. Eisenhofer G, Lenders JW, Linehan WM, Walther MM, Goldstein DS, Keiser HR. Plasma normetanephrine and metanephrine for detecting pheochromocytoma in von Hippel–Lindau disease and multiple endocrine neoplasia type 2. N Engl J Med 1999;340:1872–9. Eisenhofer G., Pacak K., Huynh T.T., Qin N., Bratslavsky G., Linehan W.M., et al.. 2011. Catecholamine metabolomic and secretory phenotypes in phaeochromocytoma. Endocr Relat Cancer 2011; 18:97–111. Hensen E.F., Bayley J.P. 2011 Recent advances in the genetics of SDHrelated paraganglioma and pheochromocytoma. Fam Cancer 2011; 10:355–63.

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PHEOCHROMOCyTOMA

Lenders JW, Eisenhofer G, Mannelli M, Pacak K. Phaeochromocytoma. Lancet 2005;366:665–75. Lenders JW, Pacak K, Walther MM, Linehan WM, Mannelli M, Friberg P, et al. Biochemical diagnosis of pheochromocytoma: which test is best? JAMA 2002;287:1427–34. Manger WM. An overview of pheochromocytoma: history, current concepts, vagaries, and diagnostic challenges. Ann NY Acad Sci 2006;1073:1–20.

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Pacak K, Eisenhofer G, Ahlman H, Bornstein SR, Gimenez-Roqueplo AP, Grossman AB, et al. Pheochromocytoma: recommendations for clinical practice from the First International Symposium. October 2005. Nat Clin Pract Endocrinol Metab 2007;3:92–102. Scholz T, Eisenhofer G, Pacak K, Dralle H, Lehnert H. Current treatment of malignant pheochromocytoma. J Clin Endocrinol Metab 2007;92:1217–25.

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C H A P T E R

87 Deficiencies of Tetrahydrobiopterin, Tyrosine Hydroxylase and Aromatic L-Amino Acid Decarboxylase Keith Hyland, Lauren A. Hyland BIOCHEMISTRY Tetrahydrobiopterin (BH4) is the cofactor for tyrosine hydroxylase (TH) and tryptophan hydroxylase (TRYPH), the rate limiting enzymes required for the synthesis of the catecholamines (dopamine, norepinephrine and epinephrine) and serotonin (5HT). BH4 is formed from GTP in a multistep pathway and defects in its biosynthesis have been described at the level of GTP cyclohydrolase, 6-pyruvoyltetrahydropterin synthase (6PTPS) and sepiapterin reductase (SR). In addition, deficiencies of dihydropteridine reductase (DHPR) and pterin-4α-carbinolamine dehydratase lead to the inability to regenerate BH4 following its oxidation in the hydroxylase reactions (Fig. 87.1). All the defects affecting BH4 metabolism are inherited in an autosomal recessive fashion, except GTP cyclohydrolase deficiency. In this disorder there is also an autosomal dominant form. The various defects of BH4 metabolism that occur within the CNS lead to a deficiency of 5HT and the catecholamines. There are, however, peripheral forms of 6PTPS deficiency in which central neurotransmitter metabolism is normal [1]. BH4 is also the cofactor for phenylalanine hydroxylase (PH); hence, defects in BH4 metabolism generally lead to hyperphenylalaninemia. Dominantly inherited GTP cyclohydrolase deficiency and SR deficiency, however, only cause a lack of BH4 within the CNS so phenylalanine metabolism is unaffected. Pterin-4α-carbinolamine dehydratase, is also involved in the hydroxylation of phenylalanine; deficiency again may lead to hyperphenylalaninemia in early life but effects on central 5HT and catecholamine metabolism are minimal, if present at all [1]. L-dopa and 5-hydroxytryptophan (5HTP), the products of the TH and TRYPH reactions, are decarboxylated by vitamin B6 requiring aromatic L-amino acid decarboxylase (AADC) to form the respective neurotransmitters (Fig. 87.1). Deficiency of TH leads to decreased concentrations of catecholamines [1] whereas a defect at the level of AADC leads to deficiencies of both 5HT and the catecholamines [2]. A defect of TRYPH has not been described.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00087-1

The main catabolites of 5HT and dopamine are 5-hydroxyindoleacetic acid (5HIAA) and homovanillic acid (HVA) respectively. The concentration of these metabolites in cerebrospinal fluid is used as an estimation of the turnover of the neurotransmitters and as a diagnostic tool.

PRESENTATION AND NEUROLOGICAL SYMPTOMS The neurological symptoms of the central BH4 defects (except dominantly inherited GTP cyclohydrolase deficiency) and AADC deficiency are very similar and reflect a combined lack of both 5HT and the catecholamine neurotransmitters. Patients generally present between 2 and 8 months of age with a fairly well characterized syndrome. Symptoms may include, hypersalivation, temperature instability, pinpoint pupils, ptosis of the eyelids, oculogyric crises, hypokinesis, distal chorea, truncal hypotonia, swallowing difficulties, drowsiness and irritability [3]. Autonomic features are variable with symptoms including ptosis, miosis, a reverse Argyll Robertson pupil, chronic or paroxysmal nasal congestion, paroxysmal sweating, temperature instability, gastrointestinal reflux, and constipation. In the first described cases of AADC deficiency, postural drop in blood pressure was not present at 9 months but was noted at 1 year of age [2]. Some patients with DHPR deficiency develop long tract signs associated with multifocal perivascular calcification located mainly in the basal ganglia and to a lesser degree in areas of white and gray matter [4]. These changes are thought to occur as a result of an insidious folate deficiency that has been postulated to arise as a result of inhibition of folate metabolism by the unusual forms of biopterin that accumulate in this disease. The neurological signs associated with peripheral forms of 6PTPS deficiency and pterin-4αcarbinolamine dehydratase deficiency are minimal and disappear following correction of the hyperphenylalaninemia. A peripheral form of 6PTPS deficiency progressing to give a central phenotype has been reported, thus all

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87. DEfICIEnCIEs of TETRAHyDRobIoPTERIn, TyRosInE HyDRoxyLAsE AnD ARomATIC L-AmIno ACID DECARboxyLAsE

3-OMD ↑ 11

L-dopa ↑

8

Tyrosine

Dopamine

11

HVA ↓

10

5HIAA ↓

9

Tryptophan

7

5HTP ↑

6

Tyrosine

Serotonin

**

Phenylalanine

5

7-Biopterins GTP

1

NH2 TP

2

6PTP

3

4

* B

N

B

N

qBH2

BH4

B

B

FIGURE 87.1 Biosynthesis of serotonin and dopamine, showing sites of metabolic block and abnormal metabolite profiles. GTP, guanosine triphosphate; NH2TP, dihydroneopterin triphosphate; 6PTP, 6-pyruvoyltetrahydropterin; BH4, tetrahydrobiopterin; qBH2, quinonoid dihydrobiopterin; 5HTP, 5-hydroxytryptophan; HVA, homovanillic acid; 5HIAA, 5-hydroxyindoleacetic acid. 3OMD, 3-O-methyldopa. 1, GTP cyclohydrolase, 2, 6-pyruvoyltetrahydropterin synthase, 3, sepiapterin reductase, 4, dihydropteridine reductase, 5, pterin-4α-carbinolamine dehydratase, 6, phenylalanine hydroxylase, 7, tryptophan hydroxylase, 8, tyrosine hydroxylase, 9, aromatic L-amino acid decarboxylase, 10, monoamine oxidase, 11, catechol-O-methyltransferase. B, biopterin, N, neopterin, ↑ & ↓ represent changes seen in aromatic L-amino acid decarboxylase deficiency. ↑ & ↓ represent changes seen in the central forms of tetrahydrobiopterin deficiency.  shows the position of a metabolic block. * Change only seen in the CNS. ** Excluding dominantly inherited GTP cyclohydrolase deficiency and sepiapterin reductase deficiency.

patients should be re-evaluated in terms of their central neurotransmitter status at a later age. Dominantly inherited GTP cyclohydrolase deficiency, otherwise known as Segawa’s disease or dopa responsive dystonia does not present as in the other defects of BH4 metabolism. Normal presentation is the appearance of a dystonic gait disorder at around 4–6 years of age, however, the age of presentation and the spectrum of clinical manifestations is broad. Occasionally onset has been with arm dystonia, retrocollis, torticollis, poor coordination or slowness in dressing before the development of leg signs. There may also be hyperreflexia and apparent extensor plantar responses, as well as other clinical features suggesting spasticity. The symptoms often, but not always, show a marked diurnal variation [5]. There is variable penetrance with females being more affected than males. Clinical presentation in TH deficiency is also varied. A recent review of 36 cases demonstrated that patients can be divided into two subgroups. In type A there is an infantile presentation, with a progressive, hypokinetic-rigid syndrome with dystonia. In Type B, onset is in the neonatal period with the presence of a complex encephalopathy [6]. Patients appear to have a paucity of autonomic features, suggesting a compensatory peripheral mechanism.

DIAGNOSIS Tetrahydrobiopterin Deficiencies Defective BH4 metabolism should be considered in all cases of hyperphenylalaninemia and in any child who

presents with the above neurological syndrome. Methods for diagnosis rely initially on the appearance of characteristic HPLC profiles of neopterins and biopterins in urine [1]. The changes expected in each condition are marked on Figure 87.1. It is likely within the near future that pterin analysis in dried blood spots will become the optimum initial screening method for diagnosing these conditions [7]. A BH4 loading test can also help distinguish between BH4 defects in which there is hyperphenylalaninemia and primary PH deficiency. Administering 2–20 mg/kg orally leads to a drop in plasma phenylalanine in deficiencies of GTP cyclohydrolase, pterin-4α-carbinolamine dehydratase and 6PTPS; however, some cases of DHPR deficiency fail to respond and many cases of BH4 responsive PH deficiency have been reported which may complicate the interpretation of test results [8]. Further tests are required in suspected cases of dominantly inherited GTP cyclohydrolase deficiency, 6PTPS, SR and DHPR deficiency. The urine biopterin, neopterin profiles are similar in DHPR deficiency and PH deficiency, therefore it is necessary to measure DHPR activity in blood spots or erythrocytes. Urine biopterin and neopterin analysis also cannot distinguish between the peripheral and central forms of 6PTPS deficiency; here it is necessary to measure CSF levels of 5HIAA and HVA. These are normal in the peripheral condition but reduced in the central forms of the disease [1]. Patients with dominantly inherited GTP cyclohydrolase deficiency and SR deficiency do not develop hyperphenylalaninemia, diagnosis in these cases is again dependent on CSF analyses. Neurotransmitter metabolite levels are low and there are also characteristic abnormal pterin patterns (Fig. 87.1) [1].

VII. CATECHOLAMINE DISORDERS

TREATmEnT

In particular there is an elevation of sepiapterin in CSF in patients with sepiapterin reductase deficiency. Molecular diagnostic tests are available for all the defects affecting BH4 metabolism and details of disease causing mutations can be found at the BIOMDB database (www.biopku.org).

Tyrosine Hydroxylase Deficiency The deficiencies of TH and AADC do not (with one exception – see below) lead to abnormal profiles using traditional screening methods (blood spot screening, organic acid or amino acid analyses etc.) and biopterin and neopterin levels are also normal. Recognition requires the clinician to consider an abnormality in biogenic amine metabolism in a child with clinical signs similar to those described above. Diagnosis of TH deficiency requires the analysis of catecholamine metabolites in CSF where concentrations of HVA and 3-methoxy-4-hydroxyphenylglycol are low [1]. Confirmation of the diagnosis relies on mutation analysis as there is no easily available peripheral tissue that can be used for enzyme analysis. Molecular diagnostic testing is available and details of disease causing mutations can be found in the BIOMDB database (www.biopku.org) and a recent review has been published [6].

Aromatic L-Amino Acid Decarboxylase Deficiency The pattern of biogenic amine metabolites in AADC deficiency is very characteristic. There is marked elevation of L-dopa, 5HTP and 3-O-methyldopa in CSF, plasma and urine. HVA and 5HIAA concentrations are greatly reduced in CSF, as are the levels of 5HT, catecholamines and their catabolites in blood. Positive diagnosis is accomplished by analysis of AADC activity in plasma [2]. Evidence for AADC deficiency can be found by analysis of urine organic acids. In this disorder there is an elevation of vanillactic acid, a metabolite of 3-O-methyldopa. Molecular diagnostic testing is available and details of disease causing mutations can be found in the BIOMDB database (www.biopku.org) and a recent review has been published [9].

TREATMENT

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effect. Adverse symptoms due to overtreatment are sometimes similar to the disease symptoms, therefore monitoring by measuring CSF levels of HVA and 5HIAA is crucial. Folinic acid (3 mg/d) should be administered in DHPR deficiency, with CSF 5-methyltetrahydrofolate levels measured at the same time as the neurotransmitter metabolites to ensure the adequacy of the dose.

TH Deficiency Treatment of TH deficiency aims to correct the abnormal catecholamine levels. Treatment with low-dose L-dopa/carbidopa (3 mg/kg and 0.75 mg/kg, respectively), three times daily has been extremely effective in some cases. In others, very slow institution of small doses of L-dopa/carbidopa 0.5–1 mg/kg), along with selegiline (MAO-B inhibitor) and an anticholinergic agent such as trihexyphenidyl has been much more beneficial than L-dopa/carbidopa alone. In type A patients response to L-dopa/carbidopa is in most cases excellent. In type B patients response may be minimal or it may take months for an effect to be observed. In addition, hypersensitivity to L-dopa in this group [9] may be a problem requiring initiation of therapy at an extremely low dose with levels being increased as tolerated [6].

AADC Deficiency Treatment in the index cases of the disease consisted of bromocriptine (dopamine agonist, 2.5 mg bid), tranylcypromine (non-selective monoamine oxidase inhibitor, 4 mg bid) and pyridoxine (100 mg bid) in combination. Therapy led to a marked clinical and biochemical improvement [2]. Pergolide and other dopamine agonists have since been shown to be effective alternates to bomocriptine in some cases. In a few cases there has been considerable response to L-dopa. Mutations affecting the L-dopa binding site provides the explanation for this effect. As AADC is a B6 requiring enzyme, high dose pyridoxine mono-therapy should be tried in all cases. Overall, the response to treatment has been disappointing. Of 78 patients reviewed, treatment in 60 cases was essentially without benefit, only 15 with a relatively mild presentation clearly improved with a mixture of vitamin B6, dopamine agonists and monoamine oxidase inhibitors [9].

References

BH4 Deficiencies Hyperphenylalaninemia may be corrected using a low phenylalanine diet or, in deficiencies of GTP cyclohydrolase, 6PTPS and pterin 4α-carbinolamine, by administration of oral BH4 (0.5–40 mg/d). BH4 is usually ineffective in DHPR deficiency. Central neurotransmitter deficiency is corrected by oral administration of the precursors, L-dopa (2–20 mg/kg/d) and 5HTP (0.8–12 mg/kg/d), in conjunction with carbidopa (0.3–4.0) mg/kg/d). Initial doses should be low, with clinical monitoring of the therapeutic

[1] Blau N, Thony B, Cotton RGH, Hyland K. Disorders of tetrahydrobiopterin and related biogenic amines. In: Scriver CR, Beaudet AL, Sly WS, Valle D, Childs B, Vogelstein D, editors. The Metabolic and Molecular Basis of Inherited Disease (8th edition). New York: McGraw-Hill; 2001. p. 1725–76. [2] Hyland K, Surtees RAH, Rodeck C, Clayton PT. Aromatic L-amino acid decarboxylase deficiency: Clinical features, diagnosis and treatment of a new inborn error of neurotransmitter amine synthesis. Neurology 1992;42:1980–8. [3] Hyland K. Abnormalities of biogenic amine metabolism. J. Inherit. Metab. Dis. 1993;16:676–90.

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[4] Smith I, Hyland K, Kendall B, Leeming R. Clinical role of pteridine therapy in tetrahydrobiopterin deficiency. J. Inherit. Metab. Dis. 1985;8(Suppl. 1):39–45. [5] Nygaard TG, Snow BJ, Fahn S, Calne DB. Dopa-responsive dystonia: Clinical characteristics and definitions. In: Segawa M, editor. Hereditary Progressive Dystonia with Marked Diurnal Fluctuation. Lancaster, UK: Parthenon; 1993. p. 3–13. [6] Willemsen MA, Verbeek MM, Kamsteeg E, de Rijk-van Andel JF, Aeby A, et al. Tyrosine hydroxylase deficiency: a treatable disorder of brain catecholamine biosynthesis. Brain 2010;133:1810–22.

[7] Nenad Blau, personal communication. [8] Blau N, Bélanger-Quintana A, Demirkol M, Feillet F, Giovannini M, et al. Optimizing the use of sapropterin (BH(4)) in the management of phenylketonuria. Mol. Genet. Metab 2009;96:158–63. [9] Brun L, Ngu LH, Keng WT, Ch’ng GS, Choy YS, et al. Clinical and Biochemical features of aromatic L-amino acid decarboxylase deficiency. Neurology 2010;75:15–17.

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88 Dopamine β-Hydroxylase Deficiency Emily M. Garland Dopamine beta-hydroxylase (DBH) deficiency, also known as norepinephrine deficiency, is a rare congenital disorder caused by the absence of the enzyme, DBH, which converts dopamine to norepinephrine (Fig. 88.1). It was simultaneously reported by investigators in Rotterdam and Nashville in the mid-1980s. Without DBH, patients are unable to produce norepinephrine or epinephrine, so that these catecholamines are undetectable in plasma, urine, and cerebrospinal fluid. As a result of enhanced tyrosine hydroxylase activity, dopamine levels are remarkably elevated. The disorder is characterized by a lack of sympathetic noradrenergic function, and affected individuals exhibit substantial deficits in autonomic regulation of cardiovascular function. The prevalence of DBH deficiency is unknown. Only 20 affected individuals, all of Western European descent, have been reported in the literature.

been systematically studied, although at least one female patient has delivered a healthy baby. Some biochemical abnormalities have also been reported in DBH deficiency. Since dopamine inhibits both the synthesis and secretion of prolactin, some degree of hypoprolactemia in these individuals is expected. Elevated blood urea nitrogen has been noted in five affected individuals in the United States. This may be evidence of a loss of renal function. Clinical features common in DBH deficiency are listed in Box 88.1. The full clinical spectrum of this disorder is not known because of the limited number of cases reported.

CLINICAL DESCRIPTION In patients with DBH deficiency, the perinatal period has been complicated by vomiting, dehydration, hypotension, hypothermia, and profound hypoglycemia requiring repeated hospitalization. Opening of the eyes has been delayed, and ptosis of the eyelids is seen in most affected infants. During childhood, exercise capacity is markedly reduced, perhaps because of hypotension provoked by physical exertion. Mental and physical development are normal. Orthostatic symptoms generally worsen in late adolescence and it is at this time that the diagnosis is commonly recognized. By early adulthood, affected individuals have profound orthostatic hypotension accompanied by presyncopal symptoms that include dizziness, blurred vision, dyspnea, nuchal discomfort, and occasionally chest pain. As a result, patients are often unable to stand for more than a few seconds. Exercise intolerance remains a problem and ptosis persists. Patients have a tendency for nasal stuffiness, particularly when lying down. Atrial fibrillation developed in one individual. Another patient had a T wave with reduced amplitude, possibly reflecting an electrolyte abnormality. In males, ejaculation is retrograde or unachievable, although sexual function in females appears normal. The reproductive behavior of DBH-deficient patients has not

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00088-3

FIGURE 88.1 Catechol synthesis pathway. Patients with dopamine β-hydroxylase (DBH) deficiency lack DBH as a result of mutations in the DBH gene and are unable to produce norepinephrine or epinephrine. The prodrug, droxidopa, is converted to norepinephrine by AADC, bypassing DBH, and is effective at relieving orthostatic hypotension and symptoms as it restores norepinephrine toward normal levels.

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BOX 88.1

PHYSICAL FINDINGS IN DBH DEFICIENCY

Feature

Severe orthostatic hypotension with an inadequate compensatory rise in heart rate Ptosis of eyelids Somewhat small pupils that respond to light and parasympatholytics but not to hydroxyamphetamine Hyperflexible or hypermobile joints Sluggish deep-tendon reflexes Intact sweating Nasal stuffiness Increased plasma creatinine and blood urea nitrogen High palate Anemia Impaired ejaculation

Despite the lack of norepinephrine, persons with DBH deficiency apparently have relatively normal mental status. Results from five patients revealed no substantial deficits in performance on a battery of cognitive tasks that have been proposed to depend on normal noradrenergic function. The investigators did not rule out that subtle cognitive deficits may exist. These findings suggest that other neuromodulators, including dopamine, have taken over the function of norepinephrine in the brains of these patients. Magnetic resonance imaging scans of the brain revealed that the patients had a smaller total brain volume, but the relative proportions of grey matter, white matter and cerebrospinal fluid, and the distribution of grey matter volume across the brain did not deviate from that of the normal population. Four patients with DBH deficiency have died. Three died from natural causes at ages 28, 57, and 63 years. One died at age 20 years, possibly by suicide. From the autopsy of the 28-year-old, the absence of DBH immunoreactivity in neurons of the ventrolateral medulla indicate that impaired central catecholaminergic neurotransmission likely contributed to the clinical picture in this patient. Additional findings included scattered pyknotic cerebral neurons, isolated microfoci of cortical gliosis, cardiac arteriolar smooth muscle hypertrophy, scattered fibrosis in the cardiac conduction system, and sclerotic renal glomeruli.

DIAGNOSIS Patients with DBH deficiency are unique in that they have virtually no norepinephrine or epinephrine or their metabolites. Without norepinephrine-mediated negative feedback on tyrosine hydroxylase, cytosolic dopamine production is amplified. Dopamine and its metabolites are therefore greatly elevated above levels found in healthy control individuals. The most helpful diagnostic test for DBH deficiency is measurement of the

plasma concentrations of norepinephrine and dopamine and their metabolites, dihydroxyphenylglycol and dihydroxyphenylacetic acid. This analysis is commonly performed with a high performance liquid chromatography (HPLC) procedure and electrochemical detection. This should be the first test undertaken in individuals who describe life-long orthostatic hypotension and profound symptoms that inhibit the ability to stand. Plasma norepinephrine concentration should be below the limits of detection (25 pg/mL or 0.15 nmol/L). Plasma dopamine concentration is frequently 100 pg/mL (0.65 nmol/L). Perhaps a more discriminating diagnostic measurement is the ratio of dihydroxyphenylacetic acid to dihydroxyphenylglycol. In individuals with DBH deficiency, this ratio is at least 100 and may exceed 1000, in contrast to 5 in healthy controls. Although catecholamines must be assayed to verify the diagnosis of DBH deficiency, a number of physiological and pharmacological tests can be used to assess autonomic function in these patients. Results of autonomic function testing in DBH deficiency are consistent with sympathetic noradrenergic denervation and adrenomedullary failure but intact vagal and sympathetic cholinergic function. For example, the vasopressor response is blunted in the isometric handgrip and cold pressor rests, and in phase II of the Valsalva maneuver, the drop in blood pressure is exaggerated but the heart rate increases normally. The plasma dopamine concentration in DBH deficiency responds to various physiological and pharmacological stimuli similarly to norepinephrine in normal individuals. For example, a change from supine to upright posture normally doubles plasma norepinephrine levels, but in patients with DBH deficiency, plasma norepinephrine remains undetectable while the plasma dopamine concentration increases 2–3 fold. These patients have no response even to high doses of tyramine, which normally increases blood pressure by releasing neuronal norepinephrine. Norepinephrine remains undetectable after tyramine

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MOuSE MODEL OF DBH DEFICIENCY

administration, while dopamine increases. These results are consistent with the hypothesis that dopamine, instead of norepinephrine, is being stored in and released by noradrenergic neurons in these patients. There is a severalfold hypersensitivity to α1-adrenoceptor agonists and β-adrenoceptor agonists in DBH deficiency. Consistent with absent noradrenergic function, propranolol, a β-adrenergic antagonist, does not lower heart rate, whereas intravenous atropine raises heart rate by 40–60 beats per minute, since parasympathetic innervation is intact. A DBH enzymatic assay is not an appropriate diagnostic test for DBH deficiency. DBH enzyme activity varies over a wide range in healthy individuals, and most individuals with low plasma DBH enzyme activity do not have DBH deficiency. Not only is DBH enzyme activity undetectable in the blood of individuals with DBH deficiency, but immunoassay and immunocytochemistry also indicate an absence of DBH protein.

DIFFERENTIAL DIAGNOSIS The striking catecholamine abnormalities and severe orthostatic hypotension distinguish DBH deficiency from other congenital disorders. Familial dysautonomia (FD) is another congenital disorder in which patients may have debilitating orthostatic hypotension. However, this disease affects sensory and parasympathetic function in addition to sympathetic neurons, and the clinical picture includes gastrointestinal dysfunction, vomiting crises, recurrent pneumonia, and altered sensitivity to pain and temperature besides blood pressure instability. Menkes disease and occipital horn syndrome are disorders of copper transport caused by mutations in the copper-transporting ATPase gene. Since DBH is a copper-dependent enzyme, DBH activity is depressed in affected individuals, leading to high plasma and cerebrospinal fluid concentrations of DOPA, dopamine, and dihydroxyphenylacetic acid but measurable dihydroxyphenylglycol and norepinephrine. Although severe orthostatic hypotension has been reported, individuals with Menkes disease and occipital horn syndrome can be differentiated from those with DBH deficiency by clinical findings. In another catecholamine disorder, aromatic amino acid decarboxylase deficiency, patients have developmental and motor problems as well as reduced dopamine levels.

GENETICS DBH deficiency is inherited in an autosomal recessive manner. The DBH gene, which maps to chromosome 9q34, is the only gene known to be associated with DBH deficiency. A number of variants in both the coding and non-coding regions of the DBH gene have been identified in patients and in unaffected family members but not in unrelated control individuals or in patients with

other autonomic disorders. Five putative disease mutations for DBH deficiency have been identified in patients in the USA, a splice donor site mutation in intron 1 and four missense mutations. The splicing mutation and three additional missense mutations were identified in patients in the Netherlands. The splice donor site mutation interferes with normal mRNA splicing, leading to undetectable levels of DBH protein. Based on studies with three of the missense mutations, it has been proposed that DBH deficiency is caused by the combination of abnormal mRNA processing and defective trafficking and trapping of DBH protein in the endoplasmic reticulum.

MANAGEMENT For the most part, treatment for DBH deficiency is supportive and directed at relieving orthostatic symptoms. Once the specific enzymatic defect for DBH deficiency had been elucidated, L-threo-3,4-dihydroxyphenylserine (DOPS or droxidopa) emerged as the treatment of choice. This agent is a prodrug acted upon by endogenous dopa decarboxylase in sympathetic nerves and in non-neuronal cells to yield norepinephrine, bypassing DBH. The administration of 100 to 500 mg droxidopa orally twice or three times daily to these patients results in dramatic increases in blood pressure and in the restoration of plasma and urinary levels of norepinephrine toward normal. Plasma epinephrine concentration remains below a detectable level. Droxidopa administration restores DOPA to within the normal range and reduces dopamine, but plasma concentrations of dopamine and its metabolites remain somewhat elevated. Individuals with DBH deficiency respond somewhat to standard therapeutic approaches for autonomic failure but not nearly as well as they respond to droxidopa.

MOUSE MODEL OF DBH DEFICIENCY DβH-knockout mice that lack DBH due to a targeted disruption of the DβH gene resemble humans with DBH deficiency in their lack of norepinephrine and epinephrine and elevated dopamine. Blood pressure is low in these mice. Since DBH deficiency is so rare, these mice are especially valuable in helping us to better understand this disorder, as well as the roles of norepinephrine and dopamine in physiology and in the actions of drugs.

Further Reading Biaggioni I, Robertson D. Endogenous restoration of noradrenaline by precursor therapy in dopamine-beta-hydroxylase deficiency. Lancet 1987;2:1170–2. Cheshire Jr. WP, Dickson DW, Nahm KF, Kaufmann HC, Benarroch EE. Dopamine beta-hydroxylase deficiency involves the central autonomic network. Acta Neuropathol 2006;112:227–9.

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Deinum J, Steenbergen-Spanjers GC, Jansen M, Boomsma F, Lenders JW, van Ittersum FJ, et al. DBH gene variants that cause low plasma dopamine beta hydroxylase with or without a severe orthostatic syndrome. J. Med. Genet. 2004;41:e38. Goldstein DS. L-Dihydroxyphenylserine (L-DOPS): a norepinephrine prodrug. Cardiovasc Drug Rev. 2006;24:189–203. Jepma M, Deinum J, Asplund CL, Rombouts SARB, Tamsma JT, Tjeerdema N, et al. Neurocognitive function in dopamine-βhydroxylase deficiency. Neuropsychopharmacology 2011;36:1608–19. Kim CH, Leung A, Huh YH, Yang E, Kim DJ, Leblanc P, et al. Norepinephrine deficiency is caused by combined abnormal mRNA processing and defective protein trafficking of dopamine {beta}hydroxylase. J Biol. Chem. 2011;286:9196–204. Kim CH, Zabetian CP, Cubells JF, Cho S, Biaggioni I, Cohen BM, et al. Mutations in the dopamine beta-hydroxylase gene are associated with human norepinephrine deficiency. Am J Med Genet 2002;108:140–7. Man In’t Veld AJ, Boomsma F, Moleman P, Schalekamp MA. Congenital dopamine-beta-hydroxylase deficiency. A novel orthostatic syndrome. Lancet 1987;1:183–8. Man In’T Veld AJ, Boomsma F, Van Den Meiracker AH, Schalekamp MA. Effect of unnatural noradrenaline precursor on sympathetic control

and orthostatic hypotension in dopamine beta-hydroxylase deficiency. Lancet 1987;2:1172–5. Mathias CJ, Bannister RB, Cortelli P, Heslop K, Polak JM, Raimbach S, et al. Clinical, autonomic and therapeutic observations in two siblings with postural hypotension and sympathetic failure due to an inability to synthesize noradrenaline from dopamine because of a deficiency of dopamine beta hydroxylase. Q J Med 1990;75:617–33. Robertson D, Garland EM. Dopamine beta-hydroxylase deficiency. 2010 GeneReviews. www.ncbi.nlm.nih.gov/books/NBK1474/. Robertson D, Goldberg MR, Onrot J, Hollister AS, Wiley R, Thompson Jr. JG, et al. Isolated failure of autonomic noradrenergic neurotransmission evidence for impaired beta hydroxylation of dopamine. New Engl J Med 1986;314:1495–7. Swoap SJ, Weinshenker D, Palmiter RD, Garber G. Dbh(–/–) mice are hypotensive, have altered circadian rhythms, and have abnormal responses to dieting and stress. Am J Physiol Regul Integr Comp Physiol 2004;286:R108–13. Thompson JM, O’Callaghan CJ, Kingwell BA, Lambert GW, Jennings GL, Esler MD. Total norepinephrine spillover, muscle sympathetic nerve activity and heart-rate spectral analysis in a patient with dopamine beta-hydroxylase deficiency. J Auton Nerv Syst 1995;55:198–206.

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89 Menkes Disease and Other ATP7A-Related Phenotypes Stephen G. Kaler Menkes disease (MD) is an inborn disorder of copper metabolism with multisystem ramifications [1]. It is caused by defects in an X-chromosomal gene that encodes an intracellular copper-transporting ATPase, ATP7A [2]. This gene product normally mediates incorporation of copper into secreted copper enzymes, including dopamine-β-hydroxylase (DBH), which are processed in the trans-Golgi network. Clinical autonomic abnormalities in MD and one of its allelic variants, occipital horn syndrome (OHS) [3], reflect partial deficiency of this enzyme. Levels of plasma and cerebrospinal fluid catechols influenced by DBH activity are distinctively abnormal and provide a highly sensitive and specific diagnostic marker for these disorders [4–6] (Fig. 89.1). Plasma catechol analysis is arguably the best current diagnostic test for at-risk newborns during the first month of life since other biochemical tests are unreliable then, and molecular analysis is less rapid. Early identification of affected infants remains a fundamental requirement for successful medical intervention, underscoring this assay’s importance [5]. Interestingly, a recently discovered third allelic variant, ATP7A-related distal motor neuropathy (DMN) resembling Charcot–Marie–Tooth disease type 2, does not involve any clinical or biochemical manifestations of DBH deficiency [7].

conjunction with neurological findings, often suggest the diagnosis [1]. Occipital horn syndrome (OHS) shares the hair and connective tissue abnormalities observed in classic MD [2,3]. However, since the neurological phenotype in OHS is mild (dysautonomia including syncope, orthostatic hypotension, and chronic diarrhea), affected individuals often escape detection until mid-childhood or later. The natural history of patients with OHS not well understood, owing to the scarcity of patients for whom longterm follow-up has been reported. The newly described ATP7A-related DMN phenotype features progressive distal motor neuropathy with mild sensory loss and no central nervous system effects [7]. Symptoms begin with distal muscle weakness and atrophy of the lower extremities, followed by upper limb involvement, reduction in tactile and vibratory sensation, and loss of deep tendon reflexes. Foot and hand deformities such as pes cavus, hammer toes and curled fingers are typical. The age of onset varies from the first to sixth decade of life, with the majority of cases presenting between 10–35 years of age. Nerve conduction studies are consistent with an axonopathy rather than a demyelinating process. The delayed-onset (often in adulthood) character of ATP7Arelated distal motor neuropathy implies that the mutations associated with this disease have subtle effects that require years to provoke pathological consequences [7].

EPIDEMIOLOGY

BIOCHEMICAL PHENOTYPES

The incidence of MD is estimated to be 1 per 50,000 to 100,000 live births. Approximately one third of these are predicted to involve de novo mutations. The clinical and biochemical features important for the diagnosis of MD are summarized in Table 89.1. Incidence data for occipital horn syndrome and ATP7A-related DMN are not available.

CLINICAL PHENOTYPE As an X-linked recessive condition, MD occurs in males who present at 2–3 months with loss of early neurodevelopmental skills, hypotonia, failure to thrive, and seizures. Characteristic physical changes of the hair and facies, in

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00089-5

The biochemical phenotype in MD involves low levels of copper in plasma, liver, and brain due to impaired intestinal absorption, reduced activities of numerous copper-dependent enzymes, and paradoxical accumulation of copper in certain tissues (i.e., duodenum, kidney, spleen, pancreas, skeletal muscle, placenta) [1]. The copper-retention phenotype is also evident in cultured fibroblasts in which reduced egress of radiolabeled copper is demonstrable in pulse-chase experiments [3]. OHS patients have low to normal levels of serum copper and ceruloplasmin, and abnormal plasma and CSF catecholamine levels, which reflect deficiency in DBH activity, as in MD [3].

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FIGURE 89.1 Diagnostic value of plasma neurochemicals for neonatal detection of MD. (A) is a scatter plot of the ratio of plasma dopamine to norepinephrine versus the ratio of plasma dihydroxyphenylacetic acid to dihydroxyphenylglycol in 36 newborns at risk. Partial deficiency of dopamineβ-hydroxylase in MD predicts a buildup of proximal metabolites in the normal catecholamine biosynthetic pathway and decreased levels of the distal metabolites. The ratios of dopamine to norepinephrine and of dihydroxyphenylacetic acid to dihydroxyphenylglycol reflect these alterations and distinguish the 14 affected infants from the 22 unaffected infants. The normal pathway of catecholamine biosynthesis is shown below the graph. (B) shows a receiver-operating-characteristic (ROC) curve for plasma dihydroxyphenylacetic acid in 36 newborns at risk for MD. ROC curves show the relationship between true positive and false positive rates for a test across various threshold values used to diagnose a condition. The upper curve, plotted by the locally weighted scatter-plot smoothing technique, represents the sensitivity and specificity for the diagnosis of MD when different cutoff values for dihydroxyphenylacetic acid are applied. The area under the curve (C statistic) for the ROC shown is 0.96. The diagonal line indicates where the curve would rest if a test were completely unreliable (area under the curve, 0.5). The C statistic for plasma dopamine levels in this sample was 1.0 (data not shown). Reprinted with permission obtained from the Massachusetts Medical Society © Kaler, S.G. et al. N. Engl. J. Med. 358:605-614, 2008.

TABLE 89.1 Diagnosis of Classical Menkes Disease CLINICAL CHARACTERISTICS Age/sex: History:

Birth to 6 months/male Neonatal hypothermia, hypoglycemia Loss of early developmental milestones Poor weight gain Seizures

Physical exam:

Profound hypotonia Abnormal hair Loose skin Pectus excavatum

AUTONOMIC MANIFESTATIONS Clinical Signs of Dysautonomia in MD and OHS

LAB FINDINGS Local hospital or clinic:

Serum copper 70 μg/L* Serum ceruloplasmin 200 mg/L* Pili torti on microscopic examination of hair

Specialized testing:

Copper egress in cultures fibroblasts Plasma catecholamine analysis** Placental copper level** Mutation analysis**

*Unreliable in newborns during first 6 to 8 weeks of life. **Rapid diagnostic test in newborns.

However, individuals with ATP7A-related distal motor neuropathy do not have biochemical findings similar to those in Menkes or OHS [7]. Specifically, no patients with ATP7A-related distal motor neuropathy have shown low serum copper, or neurochemical evidence of DBH deficiency.

Clinical features of MD conceivably attributable to DBH deficiency include temperature instability, hypoglycemia and eyelid ptosis, autonomic abnormalities that may result from selective loss of sympathetic adrenergic function. Similar clinical problems have been described in patients with congenital absence of DBH [7]. In patients with MD who have survived due to early diagnosis and pre-symptomatic initiation of copper treatment and in patients with OHS, orthostatic hypotension and chronic diarrhea are not uncommon [2].

VII. CATECHOLAMINE DISORDERS

TREATMEnT

Neurochemical Abnormalities Partial deficiency of DBH is responsible for a distinctively abnormal plasma and CSF neurochemical pattern in Menkes patients [4–6]. In our experience, the ratio of a proximal compound in the catecholamine biosynthetic pathway, dihydroxylphenylalanine (DOPAC), to a distal metabolite dihydroxyphenylglycol (DHPG), provides a better index of DBH deficiency in Menkes patients than norepinephrine (NE) levels alone. Plasma and especially CSF levels of NE, the direct product of DBH, are relatively well maintained in some Menkes patients, presumably due to compensatory mechanisms [4].

MOLECULAR DIAGNOSIS Rapid and robust molecular diagnosis of MD and OHS is available [8] although this approach typically is still not as fast as neurochemical analysis.

TREATMENT Early diagnosis and institution of subcutaneous copper injections has been successful in about 20% of MD infants treated within the first several weeks after birth [5]. The type and severity of the underlying ATP7A mutation appear to be important factors in the degree of responsitivity to early treatment. The autonomic symptoms that inconvenience many successfully treated MD patients (and which also occur in patients with OHS) should be

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amenable to treatment with l-threo-3,4-dihydroxyphenylserine (L-DOPS, Droxidopa), a compound that is converted to norepinephrine via decarboxylation by 1-aromatic-amino-acid decarboxylase in a copper-independent manner. A pilot clinical trial to confirm this hypothesis is scheduled to open shortly.

References [1] Kaler SG. Menkes disease. Barnes LA, editor. Advances in Pediatrics, Volume 41: CV Mosby; 1994. p. 263–304. [2] Kaler SG. ATP7A-related copper transport diseases - emerging concepts and future trends. Nat Rev Neu 2011;7:15–29. [3] Kaler SG, Gallo LK, Proud VK, Percy AK, Mark Y, Segal NA, et al. Occipital horn syndrome and a mild Menkes phenotype associated with splice site mutations at the MNK locus. Nat Genet 1994;8:195–202. [4] Kaler SG, Goldstein DS, Holmes C, et al. Plasma and cerebrospinal fluid neurochemical pattern in Menkes disease. Ann Neurol 1993;33:171–5. [5] Kaler SG, Holmes CS, Goldstein DS, Tang JR, Godwin SC, Donsante A, et al. Neonatal diagnosis and treatment of Menkes disease. N Engl J Med 2008;358:605–14. [6] Goldstein DS, Holmes CS, Kaler SG. Relative efficiencies of plasma catechol levels and ratios for neonatal diagnosis of Menkes disease. Neurochem Res 2009;34:1464–8. [7] Kennerson ML, Nicholson GA, Kaler SG, et al. Missense mutations in the copper transporter gene ATP7A cause X-linked distal hereditary motor neuropathy. Am J Hum Genet 2010;86:343–52. [8] Liu PC, McAndrew PE, Kaler SG. Rapid and robust screening of the MD/occipital horn syndrome gene. Genet Test 2002;6:255–60. [9] Robertson D, Goldberg MR, Ornot J, et al. Isolated failure of autonomic noradrenergic neurotransmission. N Eng J Med 1986;314:1494–7. [10] Biaggioni I, Goldstein DS, Atkinson T, et al. Dopamine-betahydroxylase deficiency in humans. Neurol 1990;40:370–3.

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90 Norepinephrine Transporter Deficiency Maureen K. Hahn Norepinephrine (NE) is the major neurotransmitter in postganglionic sympathetic synapses and is also released at synaptic terminals in the brain, including neurons that control cardiovascular function and mediate cognition, learning, memory, emotions and stress responses. The presynaptically localized NE transporter (NET) retrieves released NE via active transport into terminals, to limit the spread and duration of synaptic excitability and allow repackaging of NE into synaptic vesicles [1]. NET recaptures as much as 90% of released NE in the heart, making it a critical mediator of NE inactivation and presynaptic catecholamine homeostasis. NET is also a target for tricyclic antidepressants, NET-selective reuptake inhibitors (NSRIs), and psychostimulants, including cocaine, methylphenidate and amphetamine. NET is a member of the SLC6 family of Na/Cl-dependent transporters, with a predicted protein topology of 12 transmembrane domains with intracellularly localized amino- and carboxy-termini [2]. NET complexes with proteins of the synaptic terminal that regulate its activity and trafficking in response to signal transduction events. NET, like other SLC6 family members, can form homomultimers, which influence intracellular trafficking and functional properties of these transporters [3]. The importance of NET in the regulation of NE signaling mechanisms suggests that genetic variability that directs NET expression and activity contributes to individual differences in vulnerability to disease.

NET DEFICIENCY IN CARDIOVASCULAR DISEASE A loss of NET at postganglionic sympathetic nerve terminals, especially in the heart, occurs in a number of diseases of the autonomic system. Diminished NE uptake sites and activity have been observed in hypertension, diabetes, cardiomyopathy and heart failure. Ischemiainduced efflux of nonvesicular, cytoplasmic NE via NET may also contribute to fatal arrhythmias. Salient features of heart failure include marked increases in sympathetic nervous system activity and NE levels in the heart. Heart failure patients and animal models of heart failure demonstrate loss of NET [4]. Decreased NET expression may contribute to both the elevated NE levels and the eventual

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decrease in NE stores observed in heart failure. Indeed, NET knockout mice demonstrate enhanced stimulusevoked cardiovascular response and indices of reduced NE stores [5,6]. It remains to be determined to what extent NET is a mediator or sequela of heart failure.

HUMAN NET GENE POLYMORPHISMS NET (SLC6A2) is a single-copy gene located on chromosome 16; thus, opportunity for compensation by other gene products in response to NET genetic insult is limited (Fig. 90.1). This is supported by studies of NET knockout mice in which phenotypes of cardiovascular physiology and NE metabolism emerge [5,6]. Alternative splicing of exon 16 in the human NET gene yields two protein variants that confer differing levels of transport activity in heterologous cells. Although evidence of in vivo expression of variants has yet to be demonstrated, polymorphisms that direct a favored use of one splice pattern over others pose a potential influence on NET function. Single nucleotide polymorphisms (SNPs) are the most common type of genetic variability, and SNPs that generate changes to protein sequence and structure would be predicted to induce significant functional consequences in NET. Approximately 25 SNPs that produce amino acid substitutions that have been reported in NET, many identified in psychiatric and cardiovascular phenotypes (Fig. 90.2). Promoter region polymorphisms are another common source of genetic variation in NET with potential to influence expression and activity via control of NET transcription.

THE NET A457P VARIANT AND ORTHOSTATIC INTOLERANCE A form of orthostatic intolerance (OI) is characterized by a postural tachycardia that is not accompanied by hypotension. In some patients elevated plasma NE is a feature of the disorder that may be caused by increased sympathetic outflow or NE release. Dysfunction of NET could achieve the same endpoint, by preventing uptake of

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FIGURE 90.1 Exon/intron structure of the NET gene depicting the corresponding regions of the protein encoded by each exon. Untranslated regions of exons are shown in grey and protein transmembrane domains (TMD) are shown in black. The protein variants generated by alternative splicing of the NET gene are depicted.

FIGURE 90.2 NET amino acid variants generated by nonsynonymous SNPs from published reports or deposited at www.ncbi.nlm.nih.gov/SNP. NET is depicted as a 12 transmembrane domain spanning protein with intracellular amino- and carboxy-termini. The approximate positions of the variant residues are shown. The number refers to the amino acid position in the protein and is preceded by the single-letter code for the amino acid commonly at that position followed by the single letter code for the less common variant. Synonymous SNPs and SNPs in introns and flanking regions are also deposited at NCBI.

NE and prolonging sympathetic nervous system response. A portion of OI patients also exhibit an insensitivity to a tyramine-induced increase in plasma NE, an effect requiring uptake of tyramine through NET into terminals to displace NE from vesicular stores. Treatment of normal individuals with reboxetine, a selective NET blocker, recapitulates many of the salient characteristics of OI, including increased upright heart rate and plasma NE levels [7]. Taken together, these findings suggest NET deficiency

contributes to OI symptoms in a segment of this patient population. A proband with OI was identified who demonstrated standing-induced increased NE spillover and decreased NE clearance, decreased intraneuronal metabolism of NE, as measured by decreased dihydroxyphenylglycol (DHPG) to NE ratios, and decreased sensitivity to tyramine [8]. The proband and other family members harbor a heterozygous, nonsynonymous mutation of the NET

VII. CATECHOLAMINE DISORDERS

oTHER NET GENE PolymoRPHIsms AND CARDIovAsCulAR DIsEAsE

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OTHER NET GENE POLYMORPHISMS AND CARDIOVASCULAR DISEASE

FIGURE 90.3 Schematic representation of A457P hypothesized function at a noradrenergic synapse. Normally, NET takes up released NE that once in the cytoplasm can be repackaged into vesicles for exocytosis or metabolized by monoamine oxidase (MAO) to dihydroxyphenylglycol (DHPG). Some NE escapes the reuptake process and appears in plasma. DHPG can diffuse out of the neuron and also be measured in plasma. The failure of A457P to transport NE generates greater than normal spillover and diminished clearance of NE from the plasma. The lack of uptake diminishes the flux of NE through the MAO degradation pathway and, following release of NE, plasma DHPG levels do not increase as compared to normal. Tyramine is a substrate for NET and once inside the cell displaces NE from vesicles thereby increasing cytoplasmic NE levels and resulting in the exit of NE from the neuron via a reverse transport mechanism. The effect of tyramine to induce release of NE is blunted in the presence of A457P.

gene that produces the amino acid substitution A457P in transmembrane domain 9 (Fig. 90.2) [8]. A457P associates with elevations in standing-induced heart rate and plasma NE, and decreased plasma DHPG to NE ratio (Fig. 90.3) [8]. Transient transfection studies reveal A457P to be devoid of NE transport activity and greatly diminished in the mature, fully glycosylated form of the transporter and its surface expression [3,8]. A457P interacts in a complex with NET wild-type protein and exerts a dominant negative effect on NET wild-type plasma membrane expression and uptake [3]. The loss of wild-type function through A457P suggests that individuals heterozygous for A457P or other dominant negative transporter polymorphisms suffer from greater disruption of NET activity and subsequent symptomatology than would be predicted from the loss of one allele. Although A457P is a rare variant that has not been found in other kindreds, the study of its consequences will have implications for understanding the contribution of NET deficiency, derived through other means, to OI and other autonomic disorders. Indeed, a group of OI patients was recently demonstrated to express severely reduced NET protein levels in forearm venous tissue [9]. Our laboratory has recently generated a knockin mouse model of A457P to determine the extent to which genetically-driven NET deficiency drives altered NE homeostasis and heart rate regulation observed in human A457P carriers.

A promoter polymorphism, 3081 A/T, was identified in NET, in which the presence of the minor T allele creates a transcriptional repressor cis-element that results in decreased NET transcription in in vitro systems, although the extent to which this occurs in vivo remains to be studied [10]. 3081 A/T and 182 T/C, a promoter SNP in proximity to and demonstrating high linkage disequilibrium with 3081 A/T, are associated with exerciseinduced increases in blood pressure [11]. Interestingly, there is also a seemingly paradoxical association with decreased resting levels of plasma NE. However, although increased plasma NE has been observed in cases of extreme NET deficiency, such as the A457P phenotype and the response to pharmacological NET blockade, it is not clear what more subtle changes in NET might produce. Modest decreases in NET expression might reset the synaptic content of NE to a lower level, thus lowering basal plasma levels, whereas, following activation of the sympathetic nervous system and increased NE release, NET deficiency would contribute to an enhanced cardiovascular response. Small decreases in NET might also influence central NE neurons that regulate sympathetic outflow that, without dramatic changes in the periphery, might shift the balance to one of lower sympathetic tone. Overall, the consequences of NET SNPs associated with diminished NET activity are to alter NE homeostasis and cardiovascular responses, perhaps dependent upon the level of engagement of sympathetic nervous system activity. Several nonsynonymous SNPs that alter NET amino acid structure, and were identified in studies of blood pressure variance and long QT syndrome, were examined for their effects on NET expression and transport (Fig. 90.2) [12]. Strikingly, the majority of the SNPs have functional effects, including changes in expression and surface trafficking, substrate selectivity, regulation by protein kinase C activation and antidepressant potency. For example, the R121Q variant demonstrates a decrease in NET cell surface expression, a decrease in the VMAX for substrate transport with a greater deficit in NE versus DA transport. NET F528C demonstrates increased surface protein expression and NE transport and is, to our knowledge, the first example of a naturally occurring gain of function NET variant. The changes in F528C and R121Q trafficking and substrate-specific transport properties suggested that there might be compromised regulation by second messengers signaling. Indeed, F528C is resistant, and R121Q more sensitive to, the effects of protein kinase C activation by the phorbol ester, β-PMA to down-regulate NET activity. These data demonstrate multiple and complex influences of NET genetic variation on transporter function. Such findings underscore the potential of NET variants to influence norepinephrine homeostasis in people that carry these polymorphisms, and the critical need to explore such effects under dynamic physiological states in these individuals.

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NET AND COMORBIDITY OF CARDIOVASCULAR AND NEUROBEHAVIORAL DISORDERS NE signaling plays a role in cognitive and affective psychiatric disorders, including attention deficit hyperactivity disorder (ADHD), depression, and anxiety. Mood disorders are accompanied by altered indices of noradrenergic function, including NET levels. NE also plays an important role in attention, vigilance, learning, and memory and is hypothesized to contribute to ADHD. Therapeutics with NET-blocking activity are routinely used to treat these disorders. An association of the 3081 A/T promoter polymorphism with ADHD was found, and a novel NET protein variant, T283M, with diminished transport activity was also identified in this group of patients [10,13]. 3081 A/T and 182 T/C are also associated with phenotypes in depression [13]. Comorbidity of cognitive and affective disorders with cardiovascular disease is a serious and prevalent problem, but not well understood. Disproportionate levels of cognitive dysfunction are present in heart failure patients. Patients with anxiety complain of heart palpitations and patients with tachycardia disorders, including OI, have a higher incidence of anxiety. Activation of the sympathetic nervous system has been associated with depression and patients with major depression are at an increased risk for heart disease. Furthermore, a cardiac NE uptake deficiency has been observed in a subset of depressed patients, indicated by increased NE spillover and decreased extraction of radiolabeled NE across the heart, and decreased DHPG and DHPG to NE ratios in overflow from heart [14]. Thus, comorbidity of cardiovascular and psychiatric disease may involve common underlying genetic determinants, and NET is a strong candidate for such a mechanism.

References [1] Iversen LL. Role of transmitter uptake mechanisms in synaptic neurotransmission. Br J Pharmacol 1971;41(4):571–91. [2] Pacholczyk T, Blakely RD, Amara SG. Expression cloning of a cocaine- and antidepressant-sensitive human noradrenaline transporter. Nature 1991;350(6316):350–4.

[3] Hahn MK, Robertson D, Blakely RD. A mutation in the human norepinephrine transporter gene (SLC6A2) associated with orthostatic intolerance disrupts surface expression of mutant and wild-type transporters. J Neurosci 2003;23(11):4470–8. [4] Haider N, Baliga RR, Chandrashekhar Y, Narula J. Adrenergic excess, hNET1 down-regulation, and compromised mIBG uptake in heart failure poverty in the presence of plenty. JACC Cardiovasc Imaging 2010;3(1):71–5. [5] Keller NR, Diedrich A, Appalsamy M, Miller LC, Caron MG, McDonald MP, et al. Norepinephrine transporter-deficient mice respond to anxiety producing and fearful environments with bradycardia and hypotension. Neuroscience 2006;139(3):931–46. [6] Keller NR, Diedrich A, Appalsamy M, Tuntrakool S, Lonce S, Finney C, et al. Norepinephrine transporter-deficient mice exhibit excessive tachycardia and elevated blood pressure with wakefulness and activity. Circulation 2004;110(10):1191–6. [7] Schroeder C, Tank J, Boschmann M, Diedrich A, Sharma AM, Biaggioni I, et al. Selective norepinephrine reuptake inhibition as a human model of orthostatic intolerance. Circulation 2002;105(3):347–53. [8] Shannon JR, Flattem NL, Jordan J, Jacob G, Black BK, Biaggioni I, et al. Clues to the origin of orthostatic intolerance: a genetic defect in the cocaine- and antidepressant sensitive norepinephrine transporter. New Eng J Med 2000;342(8):541–9. [9] Lambert E, Eikelis N, Esler M, Dawood T, Schlaich M, Bayles R, et al. Altered sympathetic nervous reactivity and norepinephrine transporter expression in patients with postural tachycardia syndrome. Circ Arrhythm Electrophysiol 2008;1(2):103–9. [10] Kim CH, Hahn MK, Joung Y, Anderson SL, Steele AH, MazeiRobinson MS, et al. A polymorphism in the norepinephrine transporter gene alters promoter activity and is associated with attention-deficit hyperactivity disorder. Proc Natl Acad Sci USA 2006;50:19164–19169. [11] Kohli, U., Hahn, M.K., English, B.A., Sofowora, G.G., Muszkat, M., Li, C., et al. Genetic variation in the presynaptic norepinephrine transporter is associated with blood pressure responses to exercise in healthy humans. Pharmacogenetics and Genomics 2005;21(4): 171–8. [12] Hahn MK, Mazei-Robison MS, Blakely RD. Single nucleotide polymorphisms in the human norepinephrine transporter gene affect expression, trafficking, antidepressant interaction, and protein kinase C regulation. Mol Pharmacol 2005;68(2):457–66. [13] Hahn MK, Blackford JU, Haman K, Mazei-Robison M, English BA, Prasad HC, et al. Multivariate permutation analysis associates multiple polymorphisms with subphenotypes of major depression. Genes Brain Behav 2008;7(4):487–95. [14] Barton DA, Dawood T, Lambert EA, Esler MD, Haikerwal D, Brenchley C, et al. Sympathetic activity in major depressive disorder: identifying those at increased cardiac risk? J Hypertens 2007;25(10):2117–24.

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91 Monoamine Oxidase Deficiency Jacques W.M. Lenders, Graeme Eisenhofer Monoamine oxidase A (MAO-A) and B (MAO-B) catalyze the oxidative deamination of biogenic monoamines. Both mitochondrial isoenzymes are widely expressed among tissues, but some cell types selectively express MAO-B (platelets) and others MAO-A (skin fibroblasts, catecholaminergic neurons). The two isoenzymes share 70% homology in amino acid sequence and are encoded by two distinct genes with adjacent locations on the human X-chromosome (Xp11.23). The genes encoding MAO-A and MAO-B are located close to the gene for Norrie disease (ND). Exclusive deletion of the ND gene results in X-linked congenital blindness, and in 30–50% patients, to progressive hearing loss and mild mental retardation. Several patients have been described with a contiguous chromosomal deletion of MAO-A, MAO-B and ND genes resulting in a clinical syndrome characterized by severe mental retardation, seizures, hypotonic crises, impaired somatic growth and altered peripheral autonomic function. Loss of MAO-A and MAO-B activity was reflected neurochemically by severely reduced plasma and urinary concentrations of catecholamine deaminated metabolites, including dihydroxyphenylglycol (DHPG), the deaminated metabolite of norepinephrine and epinephrine. In contrast, concentrations of the O-methylated metabolites, normetanephrine and metanephrine, were increased. Thus, ratios of plasma normetanephrine to DHPG, which are increased by over 1000fold, provide a useful marker for the deficiency (Fig. 91.1). Also increased are platelet contents of 5-hydroxytryptamine and urinary concentrations of phenylethylamine (PE), the respective substrates of MAO-A and MAO-B. In 1993, a family with X-linked selective and complete MAO-A deficiency was described. The clinical phenotype involved borderline mental retardation and impaired impulse control, including stress-induced aggressive behavior. Carrier female subjects in this syndrome appeared phenotypically normal. The responsible point mutation consisted of a single base pair substitution in the MAO-A gene, which introduced a stop codon leading to complete loss of MAO-A activity. The neurochemical phenotype closely resembled that in patients with deletion of both MAO-A and MAO-B genes. In particular, ratios of plasma normetanephrine to DHPG were substantially increased, although less so than in patients with

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00091-3

the combined MAO-AB deficiency (Fig. 91.1). The distinguishing feature in patients with selective MAO-A deficiency was normal urinary PE excretion. The frequency of this mutation must be extremely rare since no other patients with selective MAO-A deficiency have so far been described. Two patients have been identified with deletions of MAO-B and ND genes, but with the MAO-A gene intact. These subjects showed the usual clinical features of ND and were of normal intelligence and without behavioral abnormalities. The neurochemical phenotype was similarly mild with normal levels of 5-HT, catecholamines and catecholamine metabolites, but elevated urinary levels of PE. The above clinical observations of MAO-A and MAO-B deficiency states closely agree with observations in MAO-A and MAO-B knock-out mice. MAO-A knock-out mice demonstrated aggressive behavior, probably related to impaired metabolic degradation of 5-HT. These mice showed significantly increased 5-HT levels in several brain areas and architectural alterations in the somatosensory cortex. Both phenomena were reversed by postnatal administration of a 5-HT synthesis inhibitor. Recently it was also shown in MAO-A knock-out mice that regulation of peripheral 5-HT by MAO-A plays a role in ventricular remodeling through activation of 5-HT (2A) receptors. As expected from the human findings, MAO-B deficient mice showed only mild phenotypic changes, including behavioral disinhibition and decreased anxiety-like responses, presumably due in part to regional increases of PE levels, the main neurochemical abnormality. From the above data, it is clear that MAO-A and MAO-B do not share equal or complementary capacities for the deamination of biogenic monoamines. In particular, catecholamines, including dopamine, are metabolized in vivo mainly if not exclusively by MAO-A. Apart from different affinities for monoamine substrate, the importance of MAO-A for catecholamine metabolism likely reflects selective expression of MAO-A in catecholaminergic neurons, the cellular compartment where most catecholamine deamination takes place. Several polymorphisms of MAO-A or MAO-B genes have been described. Some studies have reported positive relations between MAO-A polymorphisms and psychiatric or neurological conditions, including alcoholism, affective

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Further Reading

FIGURE 91.1 Plasma normetanephrine to DHPG ratios in control subjects, in two patients with MAO-A deficiency (A), in two patients with MAO-B deficiency (B), and in five patients with MAO-AB deficiency (AB). The dashed lines represent the 2.5 and 97.5 percentile values of the control subjects. Plasma normetanephrine was measured as the sulphate-conjugated metabolite.

disorders, panic disorder, stress-induced aggression antisocial behavior and Parkinson’s disease. There are no studies suggesting any link of MAO-B polymorphisms to a specific behavioral phenotype. It remains unclear whether any of the reported polymorphisms are of functional significance to monoamine metabolism, as reflected by altered plasma or urinary levels of monoamine metabolites, in particular ratios of plasma normetanephrine to DHPG.

Berry MD, Juorio AV, Paterson IA. The functional role of monoamine oxidases A and B in the mammalian central nervous system. Progr Neurobiol 1994;42:375–91. Bortolato M, Godar SC, Diavarian S, Chen K, Shih JC. Behavioral disinhibition and reduced anxiety-like behaviors in monoamineoxidase B-deficient mice. Neuropsychopharmacology 2009;34:2746–57. Brunner HG, Nelen M, Breakefield XO, Ropers HH, van Oost BA. Abnormal behavior associated with a point mutation in the structural gene for monoamine oxidase A. Science 1993;262:578–80. Brunner HG, Nelen MR, van Zandvoort P, Abeling NGGM, van Gennip AH, Wolters EC, et al. X-linked borderline mental retardation with prominent behavioral disturbance: phenotype, genetic localization, and evidence for disturbed monoamine metabolism. Am J Hum Genet 1993;52:1032–9. Cases O, Seif I, Grimsby J, Gaspar P, Chen K, Pournin S, et al. Aggressive behavior and altered amounts of brain 5-HT and norepinephrine in mice lacking MAO-A. Science 1995;268:1763–6. Kochersperger LM, Parker EL, Siciliano M, Darlington GJ, Denney RM. Assignment of genes for human monoamine oxidases A and B to the X-chromosome. J Neurosci 1986;16:601–16. Lairez O, Calise D, Bianchi P, Ordener C, Spreux-Varoquaux O, GuilbeauFrugier C, et al. Genetic deletion of MAO-A promotes serotonindependent ventricular hypertrophy by pressure overload. J Mol Cell Cardiol 2009;46:587–95. Lenders JWM, Eisenhofer G, Abeling NGGM, Berger W, Murphy DL, Konings CH, et al. Specific genetic deficiencies of the A and B isoenzymes of monoamine oxidase are characterized by distinct neurochemical and clinical phenotypes. J Clin Invest 1996;97:1–10. Murphy DL, Sims KB, Karoum F, de la Chapelle A, Norio R, Sankila E-M, et al. Marked amine and amine metabolite changes in Norrie disease patients with an X-chromosomal deletion affecting monoamine oxidase. J Neurochem 1990;54:242–7. Shih JC, Ridd MJ. Monoamine oxidase: from genes to behavior. Ann Rev Neurosci 1999;22:197–217. Weyler W, Hsu Y-PP, Breakefield XO. Biochemistry and genetics of monoamine-oxidase. Pharmacol Ther 1990;47:391–417.

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92 Congenital Central Hypoventilation Syndrome (CCHS) and PHOX2B Mutations Debra E. Weese-Mayer, Pallavi P. Patwari, Casey M. Rand, André Diedrich, Nancy L. Kuntz, Elizabeth M. Berry-Kravis Congenital central hypoventilation syndrome (CCHS) is characterized by disordered respiratory control (alveolar hypoventilation) and autonomic nervous system (ANS) dysregulation. Diagnosis is made in the absence of primary lung, cardiac, or neuromuscular disease or an identifiable brainstem lesion that might account for the entire phenotype inclusive of ANS dysregulation (ANSD) [1]. Alveolar hypoventilation, as demonstrated by diminutive tidal volumes and monotonous respiratory rates, results in hypoxemia and hypercarbia. Disordered respiratory control, as demonstrated by absent/severely attenuated ventilatory, behavioral, and arousal responses to endogenous/exogenous hypoxemia/hypercarbia occurring at rest or in activities of daily living results in severe physiologic compromise [1]. Mutations in the paired-like homeobox 2B (PHOX2B) gene confirm the clinical phenotype of CCHS. Because of the role of PHOX2B in early embryologic development of the ANS, it is not unexpected to identify multiple, characteristic features of physiologic ANSD and pathologic abnormalities in CCHS (Fig. 92.1 and table within).

PAIRED-LIKE HOMEOBOX 2B (PHOX2B) GENE The inheritance pattern and genetic basis of CCHS became more focused after segregation analysis of ANSD symptoms in a case-control family study [2] and demonstration of parent-to-child transmission [1]. Early genetic studies focused on Hirschsprung disease (HSCR)-related genes, but with limited results. In 2003 and 2004, several groups undertaking ANS-focused, candidate gene analysis reported variations in the PHOX2B gene [3–5], ultimately confirming PHOX2B as the disease-defining gene for CCHS [1]. To clarify, PHOX2B encodes a highly conserved homeodomain transcription factor which plays a key role in early embryologic development of ANS reflex circuits in mice [6,7] and has expression in both central autonomic neuron circuits and peripheral neural crest derivatives in the human embryo [3] and in the rodent [6–9].

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00092-5

The PHOX2B gene is located at chromosome 4p12 from base pair 41,746,098 to base pair 41,750,986. In the third of three exons, the larger of two polyalanine repeat regions normally has 20 alanines on both chromosomes; thus the PHOX2B genotype in a normal subject would be 20/20. Approximately 90% of individuals with CCHS will be heterozygous for a polyalanine repeat expansion mutation (PARM) (Fig. 92.2), with expansions to 24-33 alanine repeats on the affected allele [1]; genotypes of 20/24 to 20/33. The remaining individuals, typically with the most severe CCHS phenotypes, will be heterozygous for a nonPARM (mutation that is not a polyalanine repeat expansion, NPARM) in PHOX2B. The 76 reported NPARMs include 78% frameshift, 4% nonsense (resulting in stop codon), 16% missense mutations, and 3% missense with stop codon alteration (Fig. 92.2). Recently, deletions of/or in PHOX2B have been identified as CCHS-causing in a small subset (1%) of patients [10]. Among PARMS, the 20/25, 20/26, and 20/27 genotypes, and among NPARMs, a 38 bp deletion at the site of the polyalanine repeat, remain the most frequently identified. CCHS-related PHOX2B mutations have not been found in control populations. While de novo germline mutations cause the majority of CCHS cases, somatic mosaicism has been identified in a subset (5–10%) of parents of CCHS probands [5,11]. An autosomal dominant inheritance from these mosaic parents [5], as well as from probands [5,11], has been established, with stability of the specific PHOX2B mutation. This knowledge has led to improved educational efforts and genetic counseling in CCHS families regarding reproductive risks (Fig. 92.3).

PHOX2B GENOTYPE AND CCHS PHENOTYPE ASSOCIATION In recent years, growing evidence has demonstrated that the type of PHOX2B mutation is associated with severity of respiratory control deficit and with type and severity of ANSD features in the CCHS individual. This

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FIGURE 92.1 Signs and symptoms of CCHS-related ANS dysregulation.

allows for anticipatory guidance and an opportunity to provide improved clinical care [1,4,5,12].

Continuous Ventilatory Dependence and Other Studies Pertinent to Respiratory Control A relationship between the PHOX2B genotype and need for continuous ventilatory dependence has been reported [1,4,5,12]. Individuals with the 20/25 genotype have the mildest hypoventilation, typically requiring ventilatory support during sleep only [1]. A subset of patients with the 20/25 genotype and all known patients with the 20/24 genotype present outside of the newborn period, often after exposure to sedation, respiratory depressants or severe pneumonia. Individuals with the 20/26 genotype have variable awake needs depending upon activity level. Individuals with genotypes 20/27–20/33 typically require continuous ventilatory support, though the paucity of cases in the 20/28–20/33 genotype group prevents rigorous analysis. All patients with CCHS will hypoventilate during sleep, necessitating artificial ventilation.

Hirschsprung Disease (HSCR) HSCR is more prevalent among individuals with NPARMs than PARMs. HSCR is reported in 87–100% of NPARMs in contrast to 13–20% of PARMs [1,11,12]. Notably, HSCR has not been reported in individuals with the 20/25 genotype and only rarely with the 20/26

genotype. A high occurrence of HSCR in individuals with the 20/27 genotype has been found and anecdotally in ~30% of individuals with the 20/27–20/33 genotype.

Tumors of Neural Crest Origin Extracranial solid tumors of neural crest origin have been reported in CCHS, including neuroblastomas, ganglioneuromas, and ganglioneuroblastomas. These are found in locations with sympathetic nervous tissue such as the chest and abdomen in paraspinal ganglia or the adrenal glands. In individuals with NPARMs, neuroblastoma is the predominant tumor type (~50% are affected). In individuals with PARMs, ganglioneuromas and ganglioneuroblastomas have been reported in children with the 20/29–20/33 genotypes, though the risk for neuroblastoma in PARMs remains unexplored [1].

Cardiac Asystoles A correlation exists between the most common PARMs and length of longest R-R intervals on Holter monitoring: 0%, 19%, and 83% of individuals with the 20/25, 20/26, and 20/27 genotypes, respectively, had 3-second or longer pauses [1]. Individuals with the 20/25 genotype may be unaffected during childhood, but demonstrate prolonged asystoles in adulthood [1]. Risk of cardiac asystoles to individuals with NPARMs remains unascertained.

VII. CATECHOLAMINE DISORDERS

PHOX2B GENOTYPE AND CCHS PHENOTYPE ASSOCIATION

447

FIGURE 92.2 Schematic of the PHOX2B gene with location of all CCHS-associated mutations described to date. All polyalanine repeat expansion mutations (PARMs) are located within the second polyalanine expansion region of exon 3 (shown in red). Nearly all NPARMs identified thus far have been found at the extreme 3 end of exon 2 or in exon 3. (Reproduced with permission from reference [14]).

Facial Dysmorphology Characteristic features are described for children and young adults with CCHS, primarily those with PHOX2B PARMs [1]. The face is not dysmorphic, but is generally shorter and flatter with resulting effect of a boxy-shape. The “lip trait” includes an inferior inflection of the lateral {1/3} of the upper vermilion border. Using five variables to characterize facies (upper lip height, biocular width, upper facial height, nasal tip protrusion and the lip trait), 86% of the CCHS cases and 82% of the controls were predicted.

Phenotype Specific to NPARMs While there is overlap in all areas of the phenotypic profile of PARMs and NPARMs, disease tends to be more severe in cases with NPARMs. However, severity of the NPARM phenotype will be mutation-specific and vary greatly by the effect of frameshift, missense, or nonsense mutations on DNA transcription and, ultimately, on protein formation and function. NPARMs generally occur de novo producing severe respiratory and autonomic

dysfunction with HSCR and/or extensive intestinal dysmotility, need for continuous ventilatory support, and an increased tumor risk [1,4,5,11,12]. Some NPARMs are associated with a very high incidence of HSCR but a milder physiologic CCHS phenotype, and incomplete penetrance. Most NPARMS causing frameshift in the PHOX2B protein, especially recurrent 38- and 35-base pair deletions, produce very severe disease. However, a few frameshift mutations located early in exon 3 of PHOX2B have been inherited and are variably penetrant, suggesting that frameshifts in this area may produce a milder functional deficit than other frameshift mutations. Missense mutations in exon 2 have been found in several unrelated cases of CCHS and are the only missense mutations yet identified as causative in CCHS. Some of these, located at the 3’ end of the exon, may exert their effects by altering splicing.

Later-onset CCHS (LO-CCHS) Individuals presenting later than 1 month of age whose symptoms are otherwise compatible with CCHS and who have a PHOX2B mutation, are termed later-onset CCHS

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FIGURE 92.3 (A) Algorithm to determine when and what type of PHOX2B genetic testing should be performed in various clinical scenarios in which CCHS and LO-CCHS are suspected or confirmed. (B) Algorithm to determine when and what type of PHOX2B genetic testing should be performed in parents of CCHS proband. *The PHOX2B Sequencing Test will not identify low level mosaicism [15].

(LO-CCHS) [1]. LO-CCHS appears to reflect the variable penetrance of the PHOX2B genotypes 20/24 and 20/25 or rarely an NPARM; a subset of these mutations may require environmental cofactors to elicit the hypoventilation. LO-CCHS should be considered in the event of centrally mediated alveolar hypoventilation, cyanosis, or seizures following: (i) anesthetics or CNS depressants; (ii) severe pulmonic infection; or (iii) obstructive sleep apnea intervention. The suggested evaluation should commence with PHOX2B testing from a peripheral blood sample (Fig. 92.3). Autonomic function tests are useful to detect early autonomic dysfunction in adult-onset CCHS. Reduced heart rate variability, cardiac baroreflex sensitivity, blunted sympathetic responses to Valsalva maneuver, to hypoxemia, to isometric exercise, and to cold pressor have been reported in adult-onset PHOX2B mutation-confirmed

CCHS. Abnormal maturation of carotid body and visceral sensory ganglia are hypothesized to be the cause of the observed autonomic dysfunction [13].

COMPREHENSIVE CLINICAL EVALUATION Annual in-hospital comprehensive physiologic assessment for children three years and older, and bi-annually in the first three years of life, includes assessment of ventilatory needs during varying levels of activity and concentration while awake, ventilatory needs during all stages of sleep, exogenous ventilatory response to hyperoxia, hypoxia, hypercarbia, and combinations of these, along with continuous recording of multiple respiratory and cardiovascular variables [1]. Further, these studies are

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COMPREHENSIVE CLINICAL EVALUATION

accompanied by audiovisual surveillance with continuous recording (at a minimum) of the following variables: respiratory inductance plethysmography (chest, abdomen, sum), ECG, hemoglobin saturation with pulse waveform, end tidal carbon dioxide with waveform, sleep state staging, blood pressure, and temperature. CCHS is a life-long disease for which early initiation of chronic home ventilator support is essential. For infants and young children, optimal ventilation is provided via tracheostomy with portable home mechanical ventilation. In the older child, non-invasive positive or negative pressure ventilation may be appropriate. Non-invasive positive pressure ventilation is difficult to implement long-term and not optimal in young infants with their pliable and incompletely developed facial structures. Diaphragmatic pacing is useful in patients who are mobile and require daytime ventilatory support in addition to sleep support. All ventilatory support should be accompanied by home pulse oximetry and end tidal carbon dioxide monitoring to allow for precise control of gas exchange, and continuous attendance by a highly trained registered nurse. In CCHS, peripheral and central chemoreception is affected resulting in insufficient modulation of ventilatory response to derangements in O2 and CO2. Therefore, individuals with CCHS have monotonous respiratory rates with an attenuated or absent increase in tidal volume and respiratory rate, absence of perception of asphyxia, and potentially devastating consequences in at-risk situations (sedation, swimming, exertion, etc.). Care for individuals with CCHS is ideally provided through centers with extensive expertise in CCHS, working in close partnership with parents and regional pediatric pulmonologists and pediatricians, to provide consistent, state-of-the-art management guidance and to provide thorough, up-to-date education. The concept of centers is based on an understanding that management of children with CCHS is more time-intensive and complex than the care of other ventilator-dependent children. With modern technology for home ventilation, most children with CCHS can have prolonged survival with a good quality of life. At present, the oldest neonatally-identified patients with CCHS are graduating from college, marrying, and maintaining employment. It behooves the family and medical personnel to provide optimal ventilation and oxygenation to assure maximization of neurocognitive potential. Aggressive educational intervention coupled with careful ventilatory and cardiovascular management is essential to promote favorable neurocognitive and quality of life outcomes [1].

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References [1] Weese-Mayer DE, Berry-Kravis EM, Ceccherini I, Keens TG, Loghmanee DA, Trang H. An official ATS clinical policy statement: Congenital central hypoventilation syndrome: genetic basis, diagnosis, and management. Am J Respir Crit Care Med 2010;181(6):626–44. [2] Weese-Mayer DE, Silvestri JM, Huffman AD, Smok-Pearsall SM, Kowal MH, Maher BS, et al. Case/control family study of autonomic nervous system dysfunction in idiopathic congenital central hypoventilation syndrome. Am J Med Genet 2001;100(3):237–45. [3] Amiel J, Laudier B, Attie-Bitach T, Trang H, de Pontual L, Gener B, et al. Polyalanine expansion and frameshift mutations of the pairedlike homeobox gene PHOX2B in congenital central hypoventilation syndrome. Nat Genet 2003;33(4):459–61. [4] Matera I, Bachetti T, Puppo F, Di Duca M, Morandi F, Casiraghi GM, et al. PHOX2B mutations and polyalanine expansions correlate with the severity of the respiratory phenotype and associated symptoms in both congenital and late onset Central Hypoventilation syndrome. J Med Genet 2004;41(5):373–80. [5] Weese-Mayer DE, Berry-Kravis EM, Zhou L, Maher BS, Silvestri JM, Curran ME, et al. Idiopathic congenital central hypoventilation syndrome: analysis of genes pertinent to early autonomic nervous system embryologic development and identification of mutations in PHOX2b. Am J Med Genet A 2003;123A(3):267–78. [6] Pattyn A, Morin X, Cremer H, Goridis C, Brunet JF. The homeobox gene Phox2b is essential for the development of autonomic neural crest derivatives. Nature 1999;399(6734):366–70. [7] Pattyn A, Morin X, Cremer H, Goridis C, Brunet JF. Expression and interactions of the two closely related homeobox genes Phox2a and Phox2b during neurogenesis. Development 1997;124(20):4065–75. [8] Dubreuil V, Hirsch MR, Pattyn A, Brunet JF, Goridis C. The Phox2b transcription factor coordinately regulates neuronal cell cycle exit and identity. Development 2000;127(23):5191–201. [9] Stornetta RL, Moreira TS, Takakura AC, Kang BJ, Chang DA, West GH, et al. Expression of Phox2b by brainstem neurons involved in chemosensory integration in the adult rat. J Neurosci 2006;26(40):10305–10314. [10] Jennings LJ, Yu M, Rand CM, Kravis NS, Weese-Mayer DE. Analysis of PHOX2B gene for whole exon and gene deletion/duplication in 181 cases and controls. Am J Respir Crit Care Med 2010;181(A2282) [11] Trochet D, O’Brien LM, Gozal D, Trang H, Nordenskjold A, Laudier B, et al. PHOX2B genotype allows for prediction of tumor risk in congenital central hypoventilation syndrome. Am J Hum Genet 2005;76(3):421–6. [12] Berry-Kravis EM, Zhou L, Rand CM, Weese-Mayer DE. Congenital central hypoventilation syndrome: PHOX2B mutations and phenotype. Am J Respir Crit Care Med 2006;174(10):1139–44. [13] Diedrich A, Malow BA, Antic NA, Sato K, McEvoy RD, Mathias CJ, et al. Vagal and sympathetic heart rate and blood pressure control in adult onset PHOX2B mutation-confirmed congenital central hypoventilation syndrome. Clin Auton Res 2007;17(3):177–85. [14] Weese-Mayer DE, Rand CM, Berry-Kravis EM, Jennings LJ, Loghmanee DA, Patwari PP, et al. Congenital central hypoventilation syndrome from past to future: model for translational and transitional autonomic medicine. Pediatr Pulmonol 2009;44(6):521–35. [15] Jennings LJ, Yu M, Zhou L, Rand CM, Berry-Kravis EM, WeeseMayer DE. Comparison of PHOX2B testing methods in the diagnosis of congenital central hypoventilation syndrome and mosaic carriers. Diagn Mol Pathol 2010;19(4):224–31.

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93 Multiple System Atrophy David Robertson, Sid Gilman Multiple system atrophy (MSA) is an adult onset, steadily progressive neurodegenerative disease that presents with autonomic failure in combination with parkinsonism or cerebellar ataxia. Patients are designated as having MSA-P when parkinsonian features predominate and as MSA-C if cerebellar ataxia predominates. The neuropathological features consist of neuronal loss in the basal ganglia, cerebellum, pons, inferior olivary nuclei, and the intermediolateral column and Onuf’s nucleus of the spinal cord, typically accompanied by gliosis. The neuropathological hallmark of MSA is the glial cytoplasmic inclusion (GCI), particularly prominent in the oligodendroglia, but seen also in neurons. These structures contain misfolded, hyperphosphorylated, fibrillar α-synuclein as their main component. Thus the GCIs of MSA, like the Lewy bodies of Parkinson’s disease (PD) and dementia with Lewy bodies (DLB), contain deposits of α-synuclein. Unlike PD and DLB, however, MSA is a primary oligodendroglial disorder with an α-synucleinopathy that leads to neurodegeneration.

BACKGROUND The concept of MSA as a disease emerged slowly, and terminology has evolved. Cases of MSA were reported initially as olivopontocerebellar atrophy (OPCA), then as Shy–Drager syndrome, later as striatonigral degeneration, and finally as MSA, a term that emphasizes the combination of autonomic failure with parkinsonism, cerebellar ataxia, or all three features together. The term pure autonomic failure (PAF) emerged as the major severe idiopathic dysautonomia in which extrapyramidal and cerebellar symptoms are absent. The neuropathological changes underlying PAF, however, consist of Lewy bodies within the CNS, sympathetic and parasympathetic ganglia, and the pre- and postganglionic autonomic neurons. The hereditary forms of OPCA are now termed the spinocerebellar ataxias and are designated by number, which currently includes 30 genetically distinct forms. Among the non-hereditary (sporadic) forms of OPCA, only about 30% of cases evolve into MSA.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00093-7

EPIDEMIOLOGY The mean age of onset of MSA is about 53 years, and no proven case has had onset earlier than age 30 years. Onset after age 70 years is uncommon. Both sexes are affected equally. The rate of progression varies widely; in large series, survival rates at five years are about 84% and at ten years around 40%. The median survival time from symptom onset varies in different studies, ranging from 5 to 9 years. The annual incidence rate for all subjects is 0.6 cases per 100,000, but for subjects above age 50 years, the incidence rate is 3 cases per 100,000. The population prevalence of MSA has been estimated at 4 to 8 per 100,000, although with wide confidence intervals. In case series from Europe and North America, MSA-P cases predominate in a ratio of about 4:1; however, in Japan, MSA-C cases predominate. MSA is relentlessly progressive, and significantly shortens life expectancy. Although MSA is a sporadic disease, multiplex families with MSA have been described, principally in reports from Japan, but also from Europe. An association between single nucleotide polymorphisms within the SNCA gene and risk of MSA has been found in a large number of individual cases of MSA, which is not surprising, as the SNCA gene influences alpha-synuclein levels in both blood and brain.

PATHOPHYSIOLOGY The pathophysiology of MSA results from neuropathological changes in oligodendroglial cells consisting of GCIs composed principally of α-synuclein. The phosphoprotein-25-alpha (p25α; also known as tubulin polymerization promoting protein) accumulates early in oligodendrocytes. This protein cannot be found in normal oligodendroglia, but it can be found in myelin basic protein (MBP)-immunopositive sheaths and has been linked to myelination. p25α stimulates the aggregation of α-synuclein, indicating that some process interferes with the normal cellular function of p25α. The interference may come from the translocation of p25α within oligodendroglial cell bodies, which favors deposition and fibrillation

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of α-synuclein. SNCA gene expression signals cannot be found in oligodendroglia of either healthy adults or MSA patients, suggesting that α-synuclein derived from sources outside the oligodendroglia may represent the primary target in the disease process. There is also an association between α-synuclein and MBP. Currently it appears that in the MSA brain, the metabolism of p25α in myelin is altered, leading to partial sequestration within α-synuclein-positive GCIs. Although not emphasized in neuropathological studies, myelin degeneration is an important component of the pathophysiology in MSA. In transgenic mouse models of human SNCA, accumulation of α-synuclein causes degeneration of neurons as well as glial cells, perhaps because of mitochondrial impairment and oxidative stress. There is growing evidence that α-synuclein in diseased tissue can spread to nearby healthy tissue. For example, α-synuclein pathology in a rodent model of MSA injures new striatal transplants. Also, oligomeric α-synuclein can induce conformational changes in nearby cells. The basis of α-synuclein cell-to-cell transfer has not been elucidated, but is a key aspect of the disease. These findings indicate that MSA is a primary oligodendrogliopathy in which early myelin dysfunction is followed by α-synuclein deposition and axonal damage, and later secondary neurodegeneration occurs. It appears that GCIs induce glial cell degeneration and secondarily promote neuronal cell death. Misprocessing of p25α and MBP currently represent the earliest detectable change in MSA, indicating the key importance of the oligodendroglia-myelin-axon interface.

CLINICAL FEATURES Current diagnostic criteria derived from the Second Consensus Conference require neuropathological examination of brain tissue for a definite diagnosis of MSA.This conference established two additional levels of diagnostic certainty as well, probable MSA and possible MSA (Boxes 93.1–93.3, Table 93.1). Major clinical features supporting a

diagnosis of probable MSA include parkinsonism (usually poorly levodopa-responsive), cerebellar features, pyramidal signs, and urogenital (male erectile dysfunction, incontinence, incomplete bladder emptying and retention) and cardiovascular autonomic (particularly orthostatic hypotension) dysfunction. The parkinsonism is often symmetric, and although accompanied by a tremor in two thirds of cases (usually irregular and postural/action), fewer than 10% of cases display a pill-rolling tremor of the hands. Other clinical features (“red flags”) commonly occur, and may help to point towards a clinical diagnosis of MSA. These include the frequent, and early, occurrence of REM sleep behavior disorder (RBD); the presence of sleep apnea, increased snoring, nocturnal or daytime stridor; inspiratory sighs; contractures; disproportionate antecollis or truncal deviation (Pisa syndrome); myoclonic jerks of the fingers; cold, violaceous extremities; sweating disturbances; and volatile emotionality, commonly brief crying, or more rarely inappropriate laughter.

DIFFERENTIAL DIAGNOSIS The commonest cause for misdiagnosis of MSA in life is PD, as mild to moderate autonomic insufficiency can occur. The diagnostic criteria for probable MSA, principally the level of orthostatic hypotension required and the presence of urinary incontinence, are highly likely to exclude most cases of PD. Cases of progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), and cerebrovascular disease, particularly when it co-exists with PD, can be difficult to differentiate from possible MSA. When the presentation is mainly cerebellar, other conditions entering into the differential diagnosis are the spinocerebellar ataxias, especially SCA 1, 2-3, and 6 with an apparently negative family history, late-onset Friedreich’s ataxia, and demyelinating disease, especially primary progressive multiple sclerosis.

BOX 93.1

CRITERIA FOR THE DIAGNOSIS OF PROBABLE MSA

A Sporadic, Progressive, Adult (30 years) Onset Disease Characterized By: l

Autonomic failure involving urinary incontinence (inability to control the release of urine from the bladder, with erectile dysfunction in males) or an orthostatic decrease of blood pressure within 3 min of standing by at least 30 mmHg systolic or 15 mmHg diastolic and

l

l

Poorly levodopa-responsive parkinsonism (bradykinesia with rigidity, tremor, or postural instability) or A cerebellar syndrome (gait ataxia with cerebellar dysarthria, limb ataxia, or cerebellar oculomotor dysfunction).

MSA, multiple system atrophy.

VIII. AUTONOMIC SYNUCLEINOPATHIES

DIffERENTIAL DIAgNOSIS

455

BOX 93.2

CRITERIA FOR POSSIBLE MSA

A Sporadic, Progressive, Adult (30 years) Onset Disease Characterized By: l

l

l

Parkinsonism (bradykinesia with rigidity, tremor, or postural instability) or A cerebellar syndrome (gait ataxia with cerebellar dysarthria, limb ataxia, or cerebellar oculomotor dysfunction) and

l

At least one feature suggesting autonomic dysfunction (otherwise unexplained urinary urgency, frequency or incomplete bladder emptying, erectile dysfunction in males, or significant orthostatic blood pressure decline that does not meet the level required in probable MSA) and At least one of the additional features shown in Box 93.3.

MSA, multiple system atrophy.

BOX 93.3

A D D I T I O N A L F E AT U R E S O F P O S S I B L E M S A

Possible MSA-P or MSA-C l l

l

Babinski sign with hyperreflexia Stridor.

Possible MSA-C

Possible MSA-P

l l

l l l l

l l

Hypometabolism on FDG-PET in putamen, brainstem, or cerebellum.

Rapidly progressive parkinsonism Poor response to levodopa Postural instability within 3 years of motor onset Gait ataxia, cerebellar dyarthria, limb ataxia, or cerebellar oculomotor dysfunction Dysphagia within 5 years of motor onset Atrophy on MRI of putamen, middle cerebellar peduncle, pons, or cerebellum

l l

Parkinsonism (bradykinesia and rigidity) Atrophy on MRI of putamen, middle cerebellar peduncle, or pons Hypometabolism on FDG-PET in putamen Presynaptic nigrostriatal dopaminergic denervation on SPECT or PET.

MSA, multiple system atrophy; MSA-P, MSA with predominant parkinsonism; MSA-C, MSA with predominant cerebellar ataxia; FDG, [18F] fluorodeoxyglucose.

TABLE 93.1 features Supporting (Red flags) and not Supporting a Diagnosis of MSA Supporting Features

Nonsupporting Features

Orofacial dystonia

Classic pill-rolling rest tremor

Disproportionate antecollis

Clinically significant neuropathy

Camptocormia (severe anterior flexion of the spine) and/or Pisa syndrome (severe lateral flexion of the spine)

Hallucinations not induced by drugs

Contractures of hands or feet Inspiratory sighs Severe dysphonia

Onset after age 75 years Family history of ataxia or parkinsonism Dementia (on DSM-IV) White matter lesions suggesting multiple sclerosis

Severe dysarthria New or increased snoring Cold hands and feet Pathologic laughter or crying Jerky, myoclonic postural/action tremor MSA, multiple system atrophy; DSM-IV, Diagnostic and Statistical Manual of Mental Disorders, 4th Edition.

VIII. AUTONOMIC SYNUCLEINOPATHIES

456

93. MULTIPLE SYSTEM ATROPHY

SPECIAL TESTS MRI often reveals supratentorial putaminal atrophy, posterior putaminal hypointensity or a hyperintense rim at the lateral putaminal border, and infratentorially cerebellar and pontine atrophy, a hyperintense “hot-cross bun” appearance in the pons, and hyperintensity of the middle cerebellar peduncles. Nevertheless, a normal scan, especially early in the disease course, does not rule out the diagnosis, and a hot-cross bun sign may also be seen in SCAs 1 and 3, and pontine atrophy in SCA2. MR spectroscopy tends to show reduced N-acetyl aspartate (NAA) signal in the lentiform nucleus, but this is not sufficiently specific to assist diagnosis within individual patients. 18F-fluorodopa PET, or dopamine transporter SPECT, scans cannot reliably distinguish between PD, MSA and PSP. Reduced D2 receptor binding in striatum on 11 C-raclopride PET or IBZM-SPECT may suggest degeneration of D2 receptors, indicating MSA or PSP, but only in L-dopa naïve patients, and is much less helpful in treated cases due to down-regulation of D2 receptors by levodopa treatment. Clear cut striatal hypometabolism on 18F-FDG PET scanning is unlikely to be found in PD, and the finding of striatal with cerebellar hypometabolism is much more likely to result from MSA than from PD. According to classical concepts, the cardiac sympathetic defect in the Lewy body diseases PD and PAF, is postganglionic, whereas that in MSA is preganglionic. Many clinicians believed that, if cardiac scintigraphy with 123-I-metaiodobenzylguanidine (MIBG), a precursor of norepinephrine, reveals a clear deficit, Lewy body pathology (PD or PAF) is much more likely than MSA. Recent studies with 11C-hydroxyephedrine (HED)-PET, however, revealed partial or complete cardiac post-ganglionic sympathetic innervation in 4 of 10 patients with MSA, one of whom was verified by neuropathological examination. Hence the classical differentiation of PD and PAF as postganglionic diseases and MSA as preganglionic has not held up. Conventional cardiovascular autonomic function tests (AFTs) can demonstrate autonomic failure, but may not differentiate definitively between PD and MSA. Supine plasma norepinephrine levels under 100 pg/ml are uncommon in MSA, but common in PAF. Supine plasma norepinephrine levels that are in the normal range (150–300 pg/ml) but that change little (25%) with upright posture are found commonly in MSA. Olfactory function is characteristically lost in PD, DLB, and in PAF, but not in MSA. Therefore preservation of olfactory function increases the likelihood of MSA. Recordings of urethral and anal sphincter EMG can be helpful in differentiating MSA from PD. Thus, loss of specialized anterior horn cell neurons in MSA (and also PSP) leads to denervation and re-innervation of the striatal external sphincter muscles, manifesting as increased amplitude, polyphasia, and duration of sphincter muscle potentials. This can be diagnostically helpful provided the investigator avoids a number of potential pitfalls. The

growth hormone response to a brief intravenous clonidine infusion is impaired in MSA, but also in some cases with PD. Currently it appears that a normal response argues against MSA, but a defective response cannot distinguish between MSA and PD.

MANAGEMENT It is important to make a definitive diagnosis. Unfortunately, especially early in the disease course, this may not yet be possible, and the diagnosis may need to be reviewed at intervals. If parkinsonism is prominent, a trial of a levodopa preparation is required. If this causes unacceptable side effects, for example, dystonias/dyskinesias or worsening of postural hypotension, a trial of a dopamine agonist may be warranted. About 20% of patients may benefit from amantadine. There is no effective medical treatment for the cerebellar symptoms. Spasticity and myoclonus rarely need treatment with baclofen, or clonazepam or valproate, respectively. REM sleep behavior disorder (RBD) may be ameliorated by clonazepam. Breathing disorders should be evaluated by polysomnograms in a sleep laboratory and may require CPAP, vocal cord lateralization, or tracheostomy. Emotional incontinence can be helped by SSRIs, SNRIs, or the newly FDA-approved combination of dextromethorphan and quinidine, marketed as Nuedexta. Male erectile dysfunction is rarely responsive to sildenafil and this agent can aggravate postural hypotension. Detrusor hyperexcitability can be eased by a peripherally acting anticholinergic such as oxybutynin, but if the residual volume is above 100 ml, intermittent self-catheterization may be needed as well. For many patients, practical intervention may give the most benefit – physical therapy, occupational therapy (including a home visit to ensure safety), speech therapy (including attention to swallowing difficulty), social work, and expert assessment of the patient’s capacity to utilize a walker safely and if not, then assessment for a wheelchair, with determination regarding the need for a motorized wheelchair. Patient organizations such as the Shy–Drager Syndrome/Multiple System Atrophy Support Group (www.shy-drager.com) and the Sarah Matheson Trust for MSA (www.msaweb.co.uk) can provide information and support. In the later stages of the disease, outreach and inpatient care in hospice/palliative care facilities can provide great relief to overburdened patients and families.

NEUROPROTECTIVE THERAPY Currently two clinical trials are underway to seek a neuroprotective therapy for MSA. The first will examine Rasagiline, which is a selective irreversible inhibitor of type B monoamine oxidase that has been approved for the treatment of PD. Clinical trials of Rasagiline in PD patients

VIII. AUTONOMIC SYNUCLEINOPATHIES

NEUROPROTECTIvE THERAPY

utilizing the delayed start methodology provided evidence suggesting that Rasagiline may delay disease progression. Moreover, Rasagiline has neuroprotective effects in a transgenic mouse model of MSA. A multicenter double blind, placebo-controlled phase II trial is ongoing in patients with MSA to determine whether Rasagiline possesses diseasemodifying properties (NCT00977665, www.clinicaltrials. gov). The trial will also evaluate safety and tolerability of the medication and includes only patients with possible or probable MSA-P who are less than 3 years from the time of MSA diagnosis. Treatment is with Rasagiline 1 mg per day or placebo. The primary outcome measure is the change from baseline to Week 48/Termination visit in the total UMSARS score. The secondary outcome measures are: (a) change from baseline to week 24 in total UMSARS score; (b) ambulation endpoint: whether or not a subject achieves a score of 3 in UMSARS question 7; (c) the score of the COMPASS Select Change scale at week 48/Termination visit assessed with respect to baseline; and (d) change from baseline to week 48/Termination visit in the MSA-QoL scale. The estimated enrollment is 140 subjects; the study began in November 2009 and is expected to end in November 2011. The second medication is Rifampicin, an antibiotic used for leprosy and tuberculosis, which has been shown to remove aggregated α-synuclein and thus to modify both neurodegenerative changes and behavior in a transgenic mouse model of MSA. The purpose of this phase III study is to determine whether Rifampicin is effective in slowing or reversing the progression of the clinical symptoms and autonomic disorders of MSA (NCT01287221, www.clinicaltrials.gov). The study is being performed on participants with early MSA, preferably those with possible MSA of either the MSA-C or the MSA-P type. The study is a randomized, double-blind, placebo-controlled evaluation of the drug. The drug is administered in a dose of 300 mg twice daily for 12 months. Participants will undergo an evaluation of symptoms and function and will receive neurologic examination at the beginning of the study, at 6 months and at 12 months. They will also be contacted at 3 and 9 months by telephone. Studies will be performed at 10 participating sites. The primary outcome measures are the rate of change from baseline to 12 months in the total Unified Multiple System Atrophy Rating Scale (UMSARS_ part I score). The secondary outcome measures are (a) change from baseline to completion in total UMSARS score; (b) a slope analysis of the rate of progression in total UMSARS score from baseline to 12 months; (c) change from baseline to 12 months in UMSARS subscores; (d) whether or not a participant

457

achieves a score of 3 on each of the following UMSARS questions: #1 (Speech impairment), #2 (Swallowing impairment), and #8 (Falling); (e) change from baseline to 12 months in the COMPASS_Select scale; and (f) improvement in COMPASS_Select_change scale. The estimated enrollment is 100 subjects with MSA. The study began in March 2011 and should end in February 2014.

Further Reading Bower JH, Maraganore DM, McDonnell SK, et al. Incidence of progressive supranuclear palsy and multiple system atrophy in Olmsted County, Minnesota, 1976 to 1990. Neurology 1997;49:1284–8. Chrysostome V, Tison F, Yekhlef F, et al. Epidemiology of multiple system atrophy: a prevalence and pilot risk factor study in Aquitaine, France. Neuroepidemiology 2004;23:201–8. Danzer KM, Krebs SK, Wolff M, et al. Seeding induced by alpha-synuclein oligomers provides evidence for spreading of alpha-synuclein pathology. J Neurochem 2009;111:192–203. Fowler CJ, O’Malley KJ. Investigation and management of neurogenic bladder dysfunction. J Neurol Neurosurg Psychiatry 2003;74 (Suppl. 4):iv27–31. Freeman R. Current pharmacologic treatment for orthostatic hypotension. Clin Auton Res 2008;18(Suppl. 1):14–18. Gilman S, Little R, Johanns J, et al. Evolution of sporadic olivopontocerebellar atrophy into multiple system atrophy. Neurology 2000;55:527–32. Gilman S, Wenning GK, Low PA, et al. Second consensus statement on the diagnosis of multiple system atrophy. Neurology 2008;71:670–6. Hsu LJ, Sagara Y, Arroyo A, et al. alpha-synuclein promotes mitochondrial deficit and oxidative stress. Am J Pathol 2000;157:401–10. Kahle PJ, Neumann M, Ozmen L, et al. Hyperphosphorylation and insolubility of alpha-synuclein in transgenic mouse oligodendrocytes. EMBO Rep 2002;3:583–8. Lindersson E, Lundvig D, Petersen C, et al. p25alpha Stimulates alphasynuclein aggregation and is co-localized with aggregated alphasynuclein in alpha-synucleinopathies. J Biol Chem 2005;280:5703–15. Luk KC, Song C, O’Brien P, et al. Exogenous alpha-synuclein fibrils seed the formation of Lewy body-like intracellular inclusions in cultured cells. Proc Natl Acad Sci USA 2009;106:20051–6. Raffel DM, Koeppe RA, Little R, et al. PET measurement of cardiac and nigrostriatal denervation in Parkinsonian syndromes. J Nucl Med 2006;47:1769–77. Scholz SW, Houlden H, Schulte C, et al. SNCA variants are associated with increased risk for multiple system atrophy. Ann Neurol 2009;65:610–14. Stefanova N, Poewe W, Wenning GK. Rasagiline is neuroprotective in a transgenic model of multiple system atrophy. Exp Neurol 2008;210:421–7. Ubhi K, Rockenstein E, Mante M, et al. Rifampicin reduces alphasynuclein in a transgenic mouse model of multiple system atrophy. Neuroreport 2008;19:1271–6. Winner B, Jappelli R, Maji SK, et al. In vivo demonstration that α-synuclein oligomers are toxic. Proc Natl Acad Sci USA 2011;108:4194–99.

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C H A P T E R

94 Parkinson’s Disease John Y. Fang, Thomas L. Davis Parkinson’s disease (PD) is a neurodegenerative movement disorder defined by its motor features: asymmetric resting tremor, rigidity, bradykinesia, and postural instability. Typically, patients present in middle age, and respond well to dopamine replacement therapy (DRT). However, despite treatment, severity of the disease slowly increases. Frequently, motor fluctuations characterized by shorter duration of action of medication and choreatic dyskinesias or dystonia develop over several years. Pathologically, PD brains show eosinophilic cytoplasmic inclusions called Lewy bodies, concentrated within dopaminergic neurons of the substantia nigra. Although characteristic of PD, Lewy bodies also occur in other conditions, and small numbers may even be seen in persons who appear asymptomatic. PD also appears to be associated with several non-motor symptoms including anosmia, depression, cognitive dysfunction, sleep abnormalities and dysautonomia. Autonomic symptoms may be present to varying degrees in patients with PD, but very severe dysautonomia in patients with mild motor findings suggests a diagnosis of multiple system atrophy (MSA). In addition, because dysautonomia can be present in many elderly individuals without a movement disorder, it can be difficult to establish the precise relationship between dysautonomia and PD. Although DRT is very effective for the treatment of the motor disability from PD, autonomic symptoms can be exacerbated by standard treatments. Occasionally, some autonomic symptoms may respond favorably and other symptoms may be worsened by the same medication. For instance, anticholinergic agents often decrease drooling, but worsen constipation. There are several differing criteria in use to diagnose PD and parkinsonian syndromes [1]. Histopathology of post-mortem tissue is typically required for definitive diagnosis. Thus, the clinical diagnosis using history, physical examination and sometimes imaging tests is not always accurate. Commonly, separating PD from parkinsonian syndromes rests largely upon the response of the physical examination to DRT. However, since there are also varying methods for using dopaminergic medications in this context, there may be additional inaccuracy in the diagnosis. It is also possible that mortality could be increased with certain combinations of medications. Measuring cardiac postganglionic sympathetic

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00094-9

denervation using MIBG (123I-meta-iodobenzylguanidine) SPECT may be a useful diagnostic test for PD or MSA, but falsely positive and negative results are common [2]. Whether other imaging modalities, such as measuring uptake of levodopa analogs or dopamine transporter activity, can be effective in diagnosing PD also remains unclear [3]. Since assessing the clinical response to DRT is a key element in both the clinical diagnosis and treatment of PD, proficiency in using dopaminergic medications is an essential competency for clinicians. In patients without dysautonomia, titrating up from a low dose of carbidopa/ levodopa, pramipexole, or ropinirole as shown in Box 94.1 is generally limited only by transient nausea which can be controlled by any combination of additional carbidopa, domperidone, or 5-HT3 antagonists such as ondansetron. However, in patients with dysautonomia, severe orthostatic hypotension can occur. This effect has been largely attributed to decreased systemic vascular resistance and can be induced by either dopamine agonists or levodopa formulations. Non-pharmacologic interventions such as compression stockings may be helpful for many patients. In some cases, prophylactic treatments with oral water or midodrine, an alpha1-adrenergic receptor agonist, may also be necessary. Although mineralocorticoids and high salt intake can raise blood pressure, there is actually a decreased gastric pressor response compared to plain water [4]. A similar blunting of the blood pressure response to water has also been observed with glucose [5]. These measures to control blood pressure can be continued as long as there continues to be a beneficial clinical response. Interestingly, because many PD patients with dysautonomia also have hypertension in the supine position, the hypotensive effect of DRT can actually lower the requirement for antihypertensive therapy. However, once significant orthostatic hypotension occurs, antihypertensive treatment should be restricted to bedtime, when the patient is supine. Additional drug-induced autonomic effects can also negatively impact PD therapy. Type B monoamine oxidase (MAO-B) inhibitors, used to prolong the duration of action of dopamine, may cause severe hypertension. This risk is increased if taken with foods containing tyramines and certain medications such as some opioids and antidepressants. Since MAO-B inhibitors increase dopamine

459

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460

94. PARkInson’s DIsEAsE

BOX 94.1

I N I T I AT I N G D O PA M I N E R E P L A C E M E N T T H E R A P Y l

Carbidopa/levodopa 25/100 – Start with #{1/2} pill up to three times a day and titrate every 3–7 days until there is either a positive motor response, significant side effect or at least #10 pills per day. If there is no biological effect at this dose, advise patients to take medication at least two hours after food and to wait at least one hour until eating, as protein can interfere with absorption. Because of variable absorption, extended release forms of carbidopa/ levodopa are not recommended for initial therapy.

For all of these suggested dopamine agonist dosage algorithms, if the highest dosage is reached without either beneficial motor efficacy or significant toxicity, patients should be switched to carbidopa/levodopa. l

Pramipexole 0.125 mg or 0.25 mg – Start with 0.125 mg pill three times a day with food and titrate approximately every seven days until there is either a positive motor response, significant side effect or dose of 1.5 mg three times a day.

levels, these agents may also cause hypotension and many other adverse autonomic effects. Some studies suggest that there may also be beneficial effects of MAO-B inhibitors on the underlying disease processes in PD, but changes on autonomic function were not specifically analyzed [6]. Anticholinergic medications can be useful to reduce tremor in PD, but may also cause decreased salivation and constipation plus many other antimuscarinic side effects. Antidepressants, particularly non-specific MAO inhibitors and tricyclics, can also cause several anticholinergic effects including potentially dangerous interference with cardiac conduction. In some cases, the known “side effects” of anticholinergics can be used as a therapeutic effect as well, such as in drooling. Another aspect of dysautonomia in PD relates to effects on thermoregulation. In many patients, excessive sweating (hyperhidrosis) or decreased sweating (hypohidrosis) occurs in conjunction with motor fluctuations and responds to adjustments in DRT [7]. and perhaps deep brain stimulation [8]. Although very rare, neuroleptic malignant syndrome or parkinsonism hyperpyrexia syndrome can be a life-threatening event characterized by fever, muscle rigidity and mental status abnormalities. This condition can be precipitated by the use of dopamine receptor antagonists or the rapid withdrawal of dopaminergic therapy. Even in the absence of treatment-limiting iatrogenic effects on autonomic function, assessment of dysautonomia can be useful in counseling patients about prognosis. In order to quantify the severity of autonomic dysfunction, several test interventions can be helpful.

l

l

l

l

Pramipexole extended-release 0.375 mg – Start with #1 pill per day and titrate approximately every seven days by #1 pill per day until there is either a positive motor response, significant side effect or dose of 4.5 mg a day. Ropinirole 0.25 mg – Start with #1 or #2 pills three times a day with food and titrate approximately every seven days until there is either a positive motor response, significant side effect or dose of 8 mg three times a day. Ropinirole extended-release 2 mg – Start with #1 pill per day and titrate approximately every seven days by #1 pill per day until there is either a positive motor response, significant side effect or dose of 24 mg a day. Rotigotine patch 2 mg/day – Start with #1 patch to clean skin (upper body preferred) and titrate by 2 mg/day every week until there is either a positive motor response, significant side effect or dose of 6–8 mg/day. (Note that rotigotine is currently not available in the USA due to manufacturer recall.)

Although measurement of supine and standing blood pressure and heart rate remain common diagnostic tests for orthostatic hypotension, the actual protocol and positive and negative predictive values can be difficult to evaluate. Generally, a drop of 20 Torr in the systolic blood pressure and a drop of 10 Torr in the diastolic is considered diagnostic of orthostatic hypotension. Although tilt-table or head-tilt maneuvers may detect more cases of orthostatic hypotension, how this maneuver affects clinical management has not been carefully studied. The optimal amount of time after standing to measure blood pressure is also unclear [9], but three minutes has been commonplace [10]. Other commonly employed tests of autonomic function include sympathetic skin responses (SSRs) for sympathetic sudomotor function and heart rate variation with valsalva for vagal parasympathetic integrity. There is still some uncertainty as to whether dysfunction of these systems correlates with diagnosis, or with parkinsonian severity, a finding which would suggest more than a coincidental relationship between dysautonomia and PD. The PIGD (postural instability and gait difficulty) form of PD may be more associated with dysautonomia [11]. The evaluation and treatment of other symptoms of dysautonomia in PD also overlaps substantially with non-PD patients. Non-pharmacological treatments can be effective in many cases, and treatments without systemic toxicity are often preferred. When systemic medications are necessary, careful monitoring for unintended effects is of paramount importance. Constipation can often be managed with increased dietary intake of fluids and fiber plus increased physical

VIII. AUTONOMIC SYNUCLEINOPATHIES

ConClusIon

461

BOX 94.2

ANTICHOLINERGIC AGENTS USEFUL IN PD

For Drooling: l

l

Glycopyrrolate 1–2 mg up to three times a day.

For Tremor:

For Overactive Bladder: l

Tolterodine 1–2 mg twice a day or long-acting 2–4 mg once per day.

Oxybutynin 2.5–5 mg up to three times a day or longacting formulations 5–15 mg per day.

exercise. Stool softeners such as docusate 100–200 mg daily can often facilitate bowel movements, as can osmotic laxatives such as polyethylene glycol 3350 17–34 grams per day or lactulose 10–20 grams per day, mixed with water. For drooling which is mainly due to decreased swallowing of saliva as opposed to abnormal production, timed swallowing or chewing gum or candy (preferably sugarless to minimize dental effects) can be effective during waking hours. During sleep, an anticholinergic agent may be helpful (Box 94.2). Another option is to administer injections of botulinum toxins to the salivary glands. Doses of rimabotulinum toxin B from 2000 to 2500 units and onabotulinum toxin A up to 100 units have been shown to be effective. Overactive neurogenic bladder may also respond to some combination of behavioral therapy and medications. Timed voids and anticholinergic agents can be effective in many cases (Box 94.2). Botulinum toxins injected into the detrusor muscle and electrical stimulators have also been utilized in some patients. A small study of deep brain stimulation (targeting bilateral subthalamic nuclei) also showed some benefit on bladder abnormalities. Unfortunately, some types of neurogenic bladder may not respond adequately to any of these modalities. In severe cases of dyssynergia between the external sphincter and detrusor muscle, intermittent catheterization plus diapers may be the only alternative to an indwelling catheter. Sexual dysfunction in PD is a complicated problem due to multiple potentially conflicting underlying etiologies. Dysfunction of the peripheral autonomic nerves can lead to impotence, possibly responsive to sildenafil 50–100 mg in men [12]. Hypersexuality and other impulse control problems can be a side of effect of dopaminergic therapy [13]. Dementia can also be a factor. Changing treatments, particularly removing dopamine agonists, appears to be beneficial for hypersexuality. Treating dementia and hormonal therapy can also be effective.

CONCLUSION Differentiating PD from other parkinsonian syndromes can be very difficult and typically requires the use of

l l

Benztropine 1–4 mg up to three times a day. Trihexyphenidyl 1–4 mg up to three times a day.

dopaminergic therapy. Addressing complications of dopaminergic therapy is an important aspect of the management of both PD and parkinsonian syndromes, with or without dysautonomia. Although making a definitive diagnosis of PD and parkinsonian syndromes may require pathologic confirmation, premorbid clinical diagnosis can help with establishing a prognosis and in setting realistic therapeutic goals.

References [1] Suchowersky O, Gronseth G, et al. Practice parameter: neuroprotective strategies and alternative therapies for Parkinson disease (an evidence-based review): report of the Quality Standards Subcommittee of the American Academy of Neurology. Neurology 2006;66(7):976–82. [2] Nagayama H, Hamamoto M, et al. Reliability of MIBG myocardial scintigraphy in the diagnosis of Parkinson’s disease. J Neurol Neurosurg Psychiatry 2005;76(2):249–51. [3] Ravina B, Eidelberg D, et al. The role of radiotracer imaging in Parkinson disease. Neurology 2005;64(2):208–15. [4] Raj SR, Biaggioni I, et al. Sodium paradoxically reduces the gastropressor response in patients with orthostatic hypotension. Hypertension 2006;48(2):329–34. [5] Lu CC, Li MH, et al. Glucose reduces the effect of water to promote orthostatic tolerance. Am J Hypertens 2008;21(11):1177–82. [6] Olanow CW, Rascol O, et al. A double-blind, delayed-start trial of rasagiline in Parkinson’s disease. N Engl J Med 2009;361(13):1268–78. [7] Sage JI, Mark MH. Drenching sweats as an off phenomenon in Parkinson’s disease: treatment and relation to plasma levodopa profile. Ann Neurol 1995;37(1):120–2. [8] Trachani E, Constantoyannis C, et al. Effects of subthalamic nucleus deep brain stimulation on sweating function in Parkinson’s disease. Clin Neurol Neurosurg 2010;112(3):213–7. [9] Jamnadas-Khoda J, Koshy S, et al. Are current recommendations to diagnose orthostatic hypotension in Parkinson’s disease satisfactory? Mov Disord 2009;24(12):1747–51. [10] Lahrmann H, Cortelli P, et al. EFNS guidelines on the diagnosis and management of orthostatic hypotension. Eur J Neurol 2006;13(9):930–6. [11] Allcock LM, Kenny RA, et al. Clinical phenotype of subjects with Parkinson’s disease and orthostatic hypotension: autonomic symptom and demographic comparison. Mov Disord 2006;21(11):1851–5. [12] Zesiewicz TA, Helal M, et al. Sildenafil citrate (Viagra) for the treatment of erectile dysfunction in men with Parkinson’s disease. Mov Disord 2000;15(2):305–8. [13] Weintraub D. Dopamine and impulse control disorders in Parkinson’s disease. Ann Neurol 2008;64(Suppl. 2):S93–S100.

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95 Dementia with Lewy Bodies Sylvia Stemberger, Michaela Stampfer, Gregor K. Wenning CLINICAL ASPECTS AND DIFFERENTIAL DIAGNOSIS

PRACTICAL MANAGEMENT

Dementia with Lewy bodies (DLB) represents the second most common cause of neurodegenerative dementia in the elderly after Alzheimer’s disease (AD) [1]. DLB is still somewhat a controversial entity defined by coexistent parkinsonism and progressive cognitive decline accompanied by spontaneous recurrent visual hallucinations and conspicuous fluctuations in alertness and cognitive performance [1]. Affected patients at post-mortem examination show numerous Lewy bodies in brainstem, subcortical nuclei, limbic cortex and neocortex, with a more diffuse distribution compared to Parkinson’s disease (PD) [2]. Progressive cognitive decline with particular deficits of visuospatial ability as well as frontal executive function is accompanied by usually only mildly to moderately severe parkinsonism, which is often bilateral akinetic-rigid without the classical rest-tremor. Gait abnormalities and postural instability can be prominent. Moreover the effectiveness of levodopa (L-DOPA) on motor symptoms is less pronounced in DLB than in PD [3]. Recurrent visual hallucinations may occur without exposure to dopaminergic antiparkinsonian agents. Marked diurnal fluctuations in cognitive performance have been the most difficult to define but are often conspicuous to the environment. The McKeith criteria restrict the diagnosis of DLB to patients with parkinsonism who develop dementia within 12 months of the onset of motor symptoms. Parkinsonian patients who develop dementia after 12 months of motor onset should be labeled PD dementia (PDD). DLB and PDD appear to reflect similar LB pathologies according to recent clinicopathological studies. Compared to AD, DLB is characterized by early visuospatial and constructural dysfunction, together with neuropsychiatric features and intense fluctuations [4]. In order to improve the accuracy of premortem diagnosis the McKeith criteria discriminate two levels of clinical certainty: possible DLB is a useful category for screening purposes at the expense of accuracy, probable DLB instead shows limited sensitivity but higher specificity in the order of 85% (Table 95.1) [5].

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Management of patients with DLB has to be based on a multidimensional approach taking into account the cognitive decline and dementia that form the core clinical syndrome, the characteristic hallucinations and visual delusions present in a majority of cases as well as dementia associated behavioral symptoms and depression. Furthermore, parkinsonism is a therapeutic issue in these patients as are symptoms and signs of autonomic dysfunction and – not infrequently – sleep disorders like REM associated behavior disorder (RBD) (Table 95.2).

TABLE 95.1 Consensus Criteria for Dementia with Lewy Bodies [5] CENTRAL FEATURE Progressive cognitive decline CORE FEATURES Fluctuating cognition with pronounced variations in attention Recurrent visual hallucinations Parkinsonism SUGGESTIVE FEATURES REM sleep behavior disorder Severe neuroleptic sensitivity Low dopamine transporter activity in the basal ganglia by SPECT or PET SUPPORTIVE FEATURES Repeated falls and syncope Transient, unexplained loss of consciousness Severe autonomic dysfunction Hallucinations in other modalities Systemized delusions Depression Reduced occipital activity on SPECT/PET Low uptake MIGB myocardial scintigraphy REM, rapid eye movement; SPECT, single photon emission computed tomography; PET, positron emission tomography; MIGB, I-metaiodobenzylguanidine.

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95. DEmEnTIA wITH LEwy BoDIEs

TABLE 95.2 Practical management of Dementia with Lewy Bodies DEMENTIA Rivastigmine 3–12 mg/day (↑↑) Donepezil 5–10 mg/day (↔) Memantin 10–20 mg/day (↔) HALLUCINATIONS AND PSYCHOSIS Rivastigmine (↑↑) or Donezepil (↔) Clozapin, dose range starting with 6.25 mg/night, occasional increase up to 50 mg; during the first 18 weeks of treatment, the number of leucocytes has to be determined on a weekly basis, upon consecutive therapy on a monthly basis Quetiapin, alternative to Clozapin, no blood monitoring necessary, starting with 25 mg/day up to 50–150 mg/day (↔) Classical neuroleptics, as well as Risperidon or Olanzapin should not be considered PARKINSONISM L-dopa 300–500 mg/day, if tolerated (↔) In addition Entacapon 200 mg with each L-dopa dose, on occasion of wearing-off (↔) Dopamine agonists as well as anticholinergics should be avoided DYSAUTONOMIA Orthostatic hypotension Raising the head of a patient’s bed by 30 degrees Compression stockings Liberalizing the use of fluids and salts L-threo-DOPS 300 mg b.i.d. (↑↑) Midodrin 2.5 10 mg t.i.d. (↑↑) Fludrocortisone 0.1–0.3 mg/day Ephedrin 15–45 mg t.i.d. (↔) Postprandial hypotension Octreotid 25–30 mg s.c. 30 minutes before meal Nocturnal polyuria Desmopressin spray: 10–40 mcg/night or tablet form 100–400 mcg/night Bladder dysfunction Oxybutinin 2.5–5 mg b.i.d.–t.i.d. or Solifenacinsuccinat 5–10 mg/day Intermittent self-catheterization upon retention or rest volume 100 ml Erectile dysfunction Sildenafil 50–100 mg (↑↑) Obstipation Macrogel solution one sachet/day REM SLEEP BEHAVIOR DISORDERS Clonazepam 0.25–0.5 mg/night Melatonin 2 mg/day, 1–2 hours before going to sleep DEPRESSION SSRIs: Sertralin 50 mg/day (↔); Fluoxetine 20 mg/day (↔); Paroxetin 20 mg/day (↔); Mirtazapin 15–30 mg/day (↔) avoid Trizyklika OTHER THERAPEUTIC OPTIONS Physiotherapy Logopaedia Occupational therapy Percutaneous endoscopic gastrostomy (unusual, only at progressed disease stage) Wheel chair b.i.d., bis in die; t.i.d., ter in die; s.c., subcutaneous, REM, rapid eye movement; SSRIs, selective serotonin reuptake inhibitors.

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THERAPy

THERAPY Dementia Patients with DLB have a pronounced cholinergic deficit and the treatment of DLB generally parallels that for AD. Currently there are no neuroprotective agents for DLB. Acetylcholinesterase (AChE) inhibitors, originally developed to treat AD, improve cognitive, neuropsychiatric and functional symptoms in DLB and PDD [6]. However, there is only one double-blind placebocontrolled, multicenter trial that has evaluated the safety and efficacy of AChE inhibitor rivastigmine in the treatment of DLB [7], other studies have focused more on PDD [8]. At present, AChE inhibitors are the mainstay of DLB treatment. These medications, including donezipil, rivastigmine and galantamine, block the breakdown of acetylcholine within the synapse, thereby prolonging its effect on postsynaptic receptors. AChE inhibitors, are usually well tolerated at their standard dosings with slight side effects (e.g., nausea, vomiting, diarrhea, weight loss, leg gramps, urinary frequency) [9]. Doses for rivastigmine range between 3 and 12 mg/day (mg/d) with a usual mean target dose close to 10 mg. Donezipil, although not tested in a randomized controlled fashion, is also likely efficacious in a similar way as rivastigmine. Donezipil is started at a dose of 5 mg/d and can be increased to 10 mg/d. Tacrine has been very poorly studied in DLB and, due to its worse safety profile compared to rivastigmine and donezipril, is not generally recommended. Due to abnormal metabotropic glutamate receptor expression in patients with DLB, a prospective randomized placebo-controlled multicenter study for memantine, an uncompetitive glutamatergic antagonist with low to moderate affinity for the N-methyl D-aspartate receptor, was recently launched showing improvement of global clinical status and behavioral symptoms of patients with mild to moderate DLB [10].

Hallucinations and Psychosis Visual hallucinations, delusions, and psychotic behavior may improve when DLB patients are put on cholinesterase inhibitors like rivastigmine or donepezil. Still add-on treatment with antipsychotics is frequently needed. As in PD patients classic neuroleptics should be avoided due to their potential to significantly worsen motor symptoms. Unfortunately, this is also true for the atypical neuroleptics risperidone and olanzapine. Clozapine with starting bedtime doses of 6.25 mg/d and incomense in a range between 6.25 and 50 mg/d (rarely 75–150 mg/d) is probably the currently best option although it may be less well tolerated in DLB patients compared to psychotic PD patients. Weekly blood count monitoring is cumbersome but inevitable for the first 4 months (to be follow by monthly blood counts). Quetiapine may therefore prove to be an easier to use

465

option with a starting dose of 25 mg which generally has to be increased in a range between 50 and 150 mg/d. The clinical study data on the use of quetiapine in DLB, however, are very limited so far.

Parkinsonism L-dopa is the gold standard of symptomatic efficacy in PD and is also the drug of first choice to treat parkinsonism in DLB. Doses are usually in the low middle range between 300 and 500 mg of L-dopa plus decarboxylase inhibitor per day, but may be increased if clinically required. Visual hallucinations and psychotic behavior can be dose limiting. The best compromise between need for improvement of akinesia and rigidity and the risk of increasing psychotic behavior has to be sought. Dopamine agonists do not offer significant advantages over L-dopa in DLB but have a greater risk to induce psychotic side effects.

Dysautonomia Orthostatic hypotension may be a disabling feature of DLB that if present frequently exacerbates the disability arising from progressive motor disturbance. A number of simple non-pharmacological strategies such as a glass of water ingested while sitting or lying 5 to 10 minutes before rising, elastic support stockings or tights, a high-salt diet, frequent small meals, head-up tilt of the bed at night, and rising slowly from a sitting to a standing position, may all improve orthostatic symptoms and should be tried before resorting to drug therapy. If these measures fail, the mineralocorticoid fludrocortisone may be given at night (0.1–0.3 mg). If orthostatic blood pressure drop persists sympathomimetics such as ephedrine (15–45 mg t.i.d.) or midodrine (2.5–10 mg t.i.d.) should be added to fludrocortisone. Alternatively, droxidopa, the physiological pro-drug of noradrenaline, can be tried at a dose of up to 300 mg t.i.d. Frequency and urge incontinence are often helped by oxybutynin (2.5–5 mg b.i.d. to t.i.d.), but this peripherally acting anticholinergic may precipitate urinary retention. A substantial postmicturition residue of 100 ml is an indication for intermittent self-catheterization (or catheterization by the spouse). In the advanced stages of DLB, a urethral or suprapubic catheter may become necessary. Erectile failure can be improved by oral yohimbine (2.5–5 mg t.i.d.) or sildefanil (50–100 mg) or by intracavernosal injection of papaverine or a penis implant. REM-sleep associated behavior disorder (RBD) is a common cause for disturbed night sleep in patients with DLB. When reasonable suspicion for the presence of RBD emerges from interview with spouses or sleep laboratory studies a trial of clonazepam starting with 0.5 mg/d may be tried. As with other benzodiazepines patients should be closely monitored for possible paradoxical reactions and increased anxiety agitation or confusion.

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95. DEmEnTIA wITH LEwy BoDIEs

References [1] McKeith IG, Galasko D, Kosaka K, et al. Consensus guidelines for the clinical and pathologic diagnosis of dementia with Lewy bodies (DLB): report of the consortium on DLB international workshop. Neurology 1996;47:1113–24. [2] Walter BL. Cardiovascular autonomic dysfunction in patients with movement disorders. Cleve Clin J Med 2008;75(Suppl. 2):S54–8. [3] Molloy S, McKeith IG, O'Brien JT, Burn DJ. The role of levodopa in the management of dementia with Lewy bodies. J Neurol Neurosurg Psychiatry 2005;76:1200–3. [4] Geser F, Wenning GK, Poewe W, McKeith I. How to diagnose dementia with Lewy bodies: state of the art. Mov Disord 2005;20(Suppl. 12):S11–20. [5] McKeith IG, Dickson DW, Lowe J, Emre M, et al. Diagnosis and management of dementia with Lewy bodies: third report of the DLB Consortium. Neurology 2005;65:1863–72.

[6] Marsh L. Treatment of Lewy-body dementias and psychopathology. Lancet Neurol 2010;9:943–5. [7] McKeith IG, Grace JB, Walker Z, Byrne EJ, Wilkinson D, Stevens T, et al. Rivastigmine in the treatment of dementia with Lewy bodies: preliminary findings from an open trial. Int J Geriatr Psychiatry 2000;15:387–92. [8] Emre M, Aarsland D, Albanese A, Byrne EJ, Deuschl G, De Deyn PP, et al. Rivastigmine for dementia associated with Parkinson's disease. N Engl J Med 2004;351:2509–18. [9] Weisman D, McKeith I. Dementia with Lewy bodies. Semin Neurol 2007;27:42–7. [10] Emre M, Tsolaki M, Bonuccelli U, Destee A, Tolosa E, Kutzelnigg A, et al. Memantine for patients with Parkinson's disease dementia or dementia with Lewy bodies: a randomised, double-blind, placebocontrolled trial. Lancet Neurol 2010;9:969–77.

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C H A P T E R

96 Pure Autonomic Failure Horacio Kaufmann, Irwin J. Schatz Pure autonomic failure (PAF) was first described by Bradbury and Eggleston in 1925 [1]. The disorder has been most frequently referred to as idiopathic orthostatic hypotension. The name pure autonomic failure was introduced by Oppenheimer as one of the primary autonomic failure syndromes. PAF is a sporadic, adult-onset, slowly progressive neurodegenerative disorder characterized pathologically by the abnormal accumulation of the protein alpha synuclein in peripheral autonomic neurons and clinically by symptomatic orthostatic hypotension, variable bladder and sexual dysfunction and no somatic neurological deficits [2,3]. PAF affects men slightly more often than women. Its onset is slow and insidious and usually in middle age; the patient may recall that symptoms first came on several years prior to seeking help. Unsteadiness, lightheadeness, or faintness upon standing, that is worse in the morning, after meals, exercise or in hot weather usually causes the patient to seek medical advice. Questioning often reveals aching in the neck or occiput only when standing. Lying down relieves all symptoms. A decreased ability to sweat may be apparent. Men found to have PAF may have sought advice about urinary tract symptoms, including hesitancy, urgency, and dribbling, and occasional incontinence. Other signs of autonomic disturbance including erectile and ejaculatory dysfunction, an inability to appreciate orgasm and retrograde ejaculation may also occur. Women may experience urinary retention or incontinence as early symptoms. In contrast to the nausea and pallor that occur before losing consciousness in patients with vasovagal syncope, which are prominent signs of autonomic activation, in patients with PAF these signs are noticeably absent and consciousness is lost with little or no warning. Definitive diagnosis of neurogenic orthostatic hypotension as the cause of symptoms is made when symptoms are reproduced while documenting a decline in systolic blood pressure of at least 20 mmHg and diastolic blood pressure of at least 10 mmHg, within three minutes of standing. Despite the fall in blood pressure, heart rate increases little [3,4]. Patients with PAF also have decreased respiratory sinus arrhythmia and absent blood pressure overshoot during phase IV of Valsalva maneuver, indicating parasympathetic and sympathetic efferent dysfunction. PAF mainly affects efferent postganglionic neurons. Afferent pathways and somatic neurons are spared.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00096-2

DIFFERENTIAL DIAGNOSIS PAF should be distinguished from other forms of neurogenic orthostatic hypotension, such as peripheral somatic neuropathies with autonomic involvement (e.g. diabetes and amyloid), multiple system atrophy (MSA) and Parkinson’s disease (PD). Patients with PAF have no sensory, cerebellar, pyramidal or extrapyramidal dysfunction. In general, this allows clinical distinction with other forms of neurogenic orthostatic hypotension. Early in the course of the disease, the diagnosis of PAF is always tentative. After a few years, it is not uncommon that a patient presumed to have PAF develops extrapyramidal, cerebellar or cognitive deficits and turns out to have MSA or less frequently PD or dementia with Lewy bodies [2,3,5]. Therefore, a diagnosis of PAF may require a fiveyear history of isolated autonomic dysfunction because other neurological deficits may develop and thus cause reclassification of the patient’s disorder [2,3,5]. Physicians should make a meticulous search for CNS disorders before diagnosing PAF. In PD, clinical evidence of parkinsonism usually precedes the symptoms of autonomic failure by several years but this is not always the case [2,5]. Conversely, in patients with MSA, autonomic symptoms frequently precede parkinsonism or cerebellar ataxia. Hoarseness (due to dystonia of the vocal cord abductor) is highly suggestive of MSA. Sleep apnea and REM behavior disorder suggest MSA or PD. PAF is less progressive and less disabling than the other synucleinopathies. Patients with PAF most often will have a prolonged and sometimes stable course.

CATECHOLAMINE STUDIES PAF patients usually have very low plasma norepinephrine levels when recumbent, whereas plasma norepinephrine levels when recumbent are normal in MSA and variable in PD patients [6]. Upon standing, neither PAF, nor MSA nor some cases of PD with autonomic failure, have the expected increase in plasma norepinephrine levels indicating an inability to normally stimulate the release of catecholamines by baroreflex activation in all these disorders. When norepinephrine is infused into PAF patients, there is an exaggerated increase in blood

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96. PuRE AuTonomIC FAIluRE

TABLE 96.1 Plasma Concentrations of norepinephrine (nE) and Vasopressin and Response to Exogenous Catecholamines in PAF and mSA NE and Vasopressin

PAF

MSA

Plasma NE with patient recumbent

Very low

Normal

Plasma NE when patient stands

Minimal or no increase

Subnormal increase

NE infusion

Very marked increase Modest increase in in blood pressure blood pressure

Plasma vasopressin with patient recumbent

Normal

Normal

Plasma vasopressin during hypotension

Increases normally

No increase

generalized reduction in glucose utilization rate, indicating hypometabolism; most prominently in the cerebellum, brainstem, striatum, and frontal and motor cortices. None of these findings are present in patients with PAF. Sympathetic cardiac innervation, visualized with PET or SPECT after intravenous infusion of 6-[18F] fluorodopamine, or metaiodobenzylguanidine (MIBG), tracers taken up by sympathetic postganglionic neurons and handled similarly to norepinephrine, is selectively affected in patients with PD and in those with PAF but is intact in patients with MSA [2]. Thus, in a patient with presumed PAF finding normal sympathetic cardiac innervation indicates a likely development of MSA.

NEUROPATHOLOGY pressure, reflecting an excessive sensitivity of postsynaptic alpha-adrenergic receptors to exogenous catecholamines. MSA and PD patients have only a mild increase in blood pressure in response to infused norepinephrine (Table 96.1), without leftward shift in the dose-response curve. Similarly, beta-adrenergic receptor supersensitivity is present to a greater degree in PAF patients than in those with MSA [6].

NEUROENDOCRINE STUDIES PAF selectively involves efferent autonomic neurons with the postganglionic neurons mainly affected. Afferent pathways are spared. Baroreceptor-mediated vasopressin release – a measurement of afferent baroreceptor function, is normal in PAF, and presumably in PD, but is blunted in MSA [7]. Intravenous clonidine, a centrally active alpha 2-adrenoceptor agonist that stimulates growth hormone secretion, also tests the function of hypothalamic-pituitary pathways. Clonidine raised serum growth hormone in patients with PD and patients with PAF but did not in those with MSA [8]. In sum, neuroendocrine responses to hypotension or centrally acting adrenergic agonists are blunted in patients with MSA but preserved in patients with PD and PAF because brainstem-hypothalamic-pituitary pathways are only affected in MSA.

DIAGNOSTIC IMAGING TECHNIQUES Magnetic resonance imaging (MRI) of the brain and positron emission tomography (PET) of the brain and the heart may help distinguishing between PAF, MSA and PD patients. In patients with MSA, MRI of the brain frequently shows signal hypointensity in the putamen (relative to pallidum) on T2 weighted images and atrophy of the brainstem and cerebellum. Conversely, MRI of the brain in PAF is normal. PET in MSA patients shows a

In patients with PAF, intracytoplasmic eosinophilic inclusions with the histologic appearance of Lewy bodies, similar to those found in patients with PD, are identified mostly in neurons of autonomic ganglia and postganglionic nerves. Although patients with PAF have no movement disorder, a few Lewy bodies are frequently found in the CNS, including in the substantia nigra, locus coeruleus, thoracolumbar and sacral spinal cord [2,9,10]. There is, however, little neuronal loss in these structures, which explains the lack of CNS clinical deficits. As alpha-synuclein, a neuronal protein of unknown function, is a major component of the Lewy bodies and Lewy neurites, the term “Lewy body synucleinopathies” seems fitting. Lewy body synucleinopathies can be thought of as central and peripheral neurodegenerative disorders with distinct but overlapping motor, cognitive, and autonomic phenotypes. Clinical manifestations depend on the predominant central and peripheral sites of Lewy body formation and neuronal loss: nigrostriatal in PD, cortical in dementia with Lewy bodies, and peripheral sympathetic in PAF.

MANAGEMENT Patient education is an important aspect of treatment. Patients should be encouraged by the relatively benign nature of PAF. Treatment of orthostatic hypotension with volume expansion and alpha-adrenergic agonist agents allows for increased standing time and improved quality of life. Bladder management may require intermittent catheterization.

References [1] Bradbury S, Eggleston C. Postural hypotension: A report of three cases. Am Heart J 1925;I:75–86. [2] Kaufmann H, Goldstein DS. Pure autonomic failure: A restricted Lewy body synucleinopathy or early Parkinson disease? Neurology 2010;74(7):536–7.

VIII. AUTONOMIC SYNUCLEINOPATHIES

mAnAgEmEnT

[3] Schatz IJ, Bannister R, Freeman RL, Goetz CG, Jankovic J, Kaufmann HC, et al. Consensus statement on the definition of orthostatic hypotension, pure autonomic failure, and multiple system atrophy. Neurology 1996;46:1470. [4] Téllez MJ, Norcliffe-Kaufmann LJ, Lenina S, Voustianiouk A, Kaufmann H. Usefulness of tilt-induced heart rate changes in the differential diagnosis of vasovagal syncope and chronic autonomic failure. Clin Auton Res 2009;19(6):375–80. [5] Kaufmann H, Nahm K, Purohit D, Wolfe D. Autonomic failure as the initial presentation of Parkinson disease and dementia with Lewy bodies. Neurology 2004;63:1093–5. [6] Polinsky RJ, Kopin IJ, Ebert MH, Weise V. Pharmacologic distinction of different orthostatic hypotension syndromes. Neurology 1981;31:1–7.

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[7] Kaufmann H, Oribe E, Miller M, Knott P, Wiltshire-Clement M, Yahr MD. Hypotension-induced vasopressin release distinguishes between pure autonomic failure and multiple system atrophy with autonomic failure. Neurology 1992;42:590–3. [8] Kimber JR, Watson L, Mathias CJ. Distinction of idiopathic Parkinson's disease from multiple-system atrophy by stimulation of growth-hormone release with clonidine. Lancet 1997;349:1877–81. [9] Hague K, Lento S, Morgello S, Caro S, Kaufmann H. The distribution of Lewy bodies in pure autonomic failure: Autopsy findings and review of the literature. Acta Neuropathologica 1997;94:192–6. [10] Kaufmann H, Hague K, Perl D. Accumulation of alpha synuclein in autonomic nerves in pure autonomic failure. Neurology 2001;56:980–1.

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C H A P T E R

97 Diagnostic Workup of Peripheral Neuropathies with Dysautonomia Amanda C. Peltier Evaluation of the patient with peripheral neuropathy for associated dysautonomia may require a different focus than in patients presenting with purely autonomic complaints. Diagnostic evaluation of patients with peripheral neuropathy may be performed for three reasons: (i) as part of the diagnostic workup of the etiology of the peripheral neuropathy; (ii) for additional autonomic complaints cited by the patient, usually after etiology is established for the peripheral neuropathy; and (iii) for prognostic reasons (such as in diabetes mellitus or amyloidosis where autonomic neuropathy is known to confer increased mortality risk). One of the challenges of managing patients with peripheral neuropathy is that there are over 100 causes, and not every patient will have an identifiable etiology, even after extensive investigation. The history and physical exam and other electrodiagnostic, biopsychophysical tests such as nerve conduction studies and quantitative sensory tests may be helpful in determining whether autonomic testing is needed or indicated for a given patient. For the purpose of this chapter, diagnostic workup will be the primary focus, and will refer the reader to other chapters for evaluation and prognosis (clinical evaluation of autonomic disorders, diabetic autonomic neuropathy and amyloidosis for prognosis). In history taking with the patient presenting with peripheral neuropathy, it is helpful to determine whether there are autonomic symptoms present pertaining to the cardiovascular, gastrointestinal, sudomotor, genitourinary systems, and whether those symptoms may be indicative of a coexistent dysautonomia or whether medication treatment may be the cause. Tricyclic antidepressants such as amitriptyline, nortriptyline, or imipramine are typical therapeutic agents used for neuropathic pain which have multiple autonomic effects such as orthostatic hypotension, urinary retention, constipation, erectile dysfunction, anhidrosis, etc. Sudomotor impairments may be common with multiple types of polyneuropathy and are less specific than significant cardiovascular impairment such as orthostatic hypotension. Pain is also an important symptom to elucidate, as severe burning neuropathic pain is often indicative of C-fiber involvement and may be the only complaint of patients with small fiber neuropathy.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00097-4

Establishment of the time course of autonomic symptoms as compared to the development of sensory or motor symptoms is helpful determining etiology. Patients may have preexisting erectile dysfunction prior to the development of somatic neuropathies in the setting of diabetes. Patients with amyloidosis often present with sensory impairment and painful dysesthesias but occasionally may present with orthostatic hypotension and other autonomic system based complaints [1]. Patients with acute inflammatory demyelinating polyradiculoneuropathy (AIDP) often develop dysautonomia over the course of their disease but it is rarely the presenting symptom. Patients with painful idiopathic small-fiber predominant neuropathy may have coexistent autonomic symptoms but typically the autonomic involvement is subclinical or minimal at presentation. Patients with hereditary sensory and autonomic neuropathy, toxic neuropathies, or Fabry’s disease often manifest autonomic symptoms alongside symptoms of somatic C-fiber impairment [2]. Patients with paraneoplastic polyneuropathy may have coexistent paraneoplastic autonomic neuropathy due to the same autoantibody or a second autoantibody. Subclinical autonomic impairment is frequent in diabetes, impaired glucose tolerance, idiopathic small fiber polyneuropathy, and in the setting of connective-tissue diseases such as Sjögren’s syndrome, systemic lupus erythematosis, and mixed connective tissue disease [3–5]. Physical examination and neurophysiologic testing may be helpful in determining whether large fiber (Aα or Aβ) or small fiber (C) involvement is predominant. Decreased vibratory sensation at the toes using a 120 Hz tuning fork, Rydel–Seiffer fork or quantitative sensory testing, decreased deep tendon reflexes and weakness are indicative of large fiber involvement. Relative preservation of vibration and proprioception with significant loss of pin or temperature sensation is suggestive of predominant C fiber derangement. Normal nerve conduction studies also suggest that significant large fiber involvement is not present. Few large fiber predominant polyneuropathies, such as inherited demyelinating disorders, have significant autonomic involvement making formal autonomic testing less helpful diagnostically in those situations (see Table 97.1).

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97. DIAgNOSTIC WORkUP Of PERIPHERAL NEUROPATHIES WITH DySAUTONOMIA

TABLE 97.1 Peripheral Neuropathies with Significant Dysautonomia Large Fiber Peripheral Neuropathy

Mixed Fiber Peripheral Neuropathy

Small Fiber Peripheral Neuropathy

AIDP

Diabetes mellitus

Inherited amyloidosis

Cisplatinum induced neuropathy

Leprosy

Fabry’s disease

Anti-Hu related neuropathy

Toxic neuropathy (vincristine, paclitaxel, thalidomide, Hereditary sensory and autonomic neuropathy organic solvents, acrylamide, etc.) Plasma cell dyscrasia associated polyneuropathy Tangier disease Neuropathy in connective tissue disorders HIV neuropathy

BOX 97.1

R E C O M M E N D AT I O N S O N S P E C I F I C AU T O M AT I C T E S T I N G

Cardiovagal or Parasympathetic tests l l

Sinus arrhythmia† Heart rate response to Valsalva maneuver

Sudomotor Tests l l l

Adrenergic or Sympathetic Tests l l

l l l l

l

Orthostatic blood pressure measurements† Passive tilt study with beat to beat hemodynamic monitoring† Blood pressure response to Valsalva maneuver Sustained isometric exercise (hand grip) Cold Pressor Test Hyperventilation for 30 seconds

Formal autonomic testing may be helpful in establishing the diagnosis of peripheral neuropathy in small fiber predominant neuropathies where nerve conduction testing is normal. Quantitative sudomotor axon reflex testing (QSART) has become a recognized ancillary test in evaluation of small fiber predominant polyneuropathy [6]. Thermoregulatory sweat testing will also show a length dependent loss of sweat in small fiber polyneuropathy [7]. Because damage to sudomotor fibers often occurs in parallel with other C-fibers in diabetes, and small fiber polyneuropathy, tests of sudomotor function can provide up to 80% sensitivity in diagnosis of small fiber neuropathies. However, studies comparing autonomic tests to other tests of small fiber involvement such as skin biopsy with intraepidermal nerve fiber density (IENFD) measurement or quantitative sensory testing using cold thresholds typically do not show good correlation between measures[8–10]. Cardiac autonomic testing utilizing sinus arrhythmia or heart rate response during Valsalva maneuver can be performed easily in most laboratories and may provide up to 60% sensitivity in small fiber neuropathy diagnosis. Data on sensitivity and specificity of specific autonomic tests beyond QSART and heart rate variability is sparse, limiting recommendations on specific testing (see Box 97.1). In addition, many small fiber neuropathies are caused by relatively

Quantitative sudomotor axon reflex test† Thermoregulatory sweat test† Quantitative direct and indirect reflex test testing Quantitation of sudomotor innervation (skin biopsy)

†

Included in Practice parameter statement of American Academy of

Neurology [6].

rare genetic disorders such as Fabry disease or hereditary sensory and autonomic neuropathy, and comparison of patients may be difficult because of the small numbers. Advancements in treatment of disorders such as HIV infection related neuropathy has altered the prevalence of neuropathy caused by medications and previous studies of prevalence of autonomic function may be outdated [11]. Unlike evaluation of patients with primary autonomic complaints, additional studies such as brain imaging, cardiac nuclear imaging and microneurography are typically not warranted unless another primary autonomic disorder such as multiple system atrophy is suspected, or they are being used for prognostic testing such as in diabetic CAN. Some tests, such as laser Doppler probes, may be useful as endpoints in clinical research trials but there is limited data on their usefulness in peripheral neuropathy diagnosis. Pupillometry may be helpful in diabetes but is not typically used clinically for diagnosis in peripheral neuropathies. There is little literature on additional tests performed in some autonomic laboratories such as blood pressure response to isometric exercise, cold pressor test or hyperventilation. Further research is needed on the sensitivity and specificity of autonomic testing in peripheral neuropathy. In addition, the role of frequencydomain indexes has not been formerly evaluated for most peripheral neuropathies other than diabetes mellitus.

IX. PERIPHERAL AUTONOMIC DISORDERS

DIAgNOSTIC WORkUP Of PERIPHERAL NEUROPATHIES WITH DySAUTONOMIA

References [1] Matsuda M, Gono T, Morita H, Katoh N, Kodaira M, Ikeda S. Peripheral nerve involvement in primary systemic AL amyloidosis: A clinical and electrophysiological study. Eur J Neurol 2010;18(4):604–10. [2] Freeman R. Autonomic peripheral neuropathy. Lancet 2005;365(9466):1259–70. [3] McCombe PA, McLeod JG, Pollard JD, Guo YP, Ingall TJ. Peripheral sensorimotor and autonomic neuropathy associated with systemic lupus erythematosus. Clinical, pathological and immunological features. Brain 1987;110(Pt 2):533–49. [4] Mellgren SI, Goransson LG, Omdal R. Primary Sjogren's syndrome associated neuropathy. Can J Neurol Sci 2007;34(3):280–7. [5] Stacher G, Merio R, Budka C, Schneider C, Smolen J, Tappeiner G. Cardiovascular autonomic function, autoantibodies, and esophageal motor activity in patients with systemic sclerosis and mixed connective tissue disease. J Rheumatol 2000;27(3):692–7. [6] England JD, Gronseth GS, Franklin G, Carter GT, Kinsella LJ, Cohen JA, et al. Practice Parameter: Evaluation of distal symmetric polyneuropathy: Role of autonomic testing, nerve biopsy, and skin biopsy (an evidence-based review). Report of the American Academy of Neurology, American Association of Neuromuscular and Electrodiagnostic Medicine, and American Academy of Physical Medicine and Rehabilitation. Neurology 2009;72(2):177–84.

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[7] Low PA, Opfer-Gehrking TL. The autonomic laboratory. Am J Electroneurodiagnostic Technol 1999;39(2):65–76. [8] Novak V, Freimer ML, Kissel JT, Sahenk Z, Periquet IM, Nash SM, et al. Autonomic impairment in painful neuropathy. Neurology 2001;56(7):861–8. [9] Periquet MI, Novak V, Collins MP, Nagaraja HN, Erdem S, Nash SM, et al. Painful sensory neuropathy: Prospective evaluation using skin biopsy. Neurology 1999;53(8):1641–7. [10] Smith AG, Russell J, Feldman EL, Goldstein J, Peltier A, Smith S, et al. Lifestyle intervention for pre-diabetic neuropathy. Diabetes Care 2006;29(6):1294–9. [11] Evans SR, Ellis RJ, Chen H, Yeh TM, Lee AJ, Schifitto G, et al. Peripheral neuropathy in HIV: Prevalence and risk factors. AIDS 2011.

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C H A P T E R

98 Diabetic Autonomic Dysfunction Amanda C. Peltier, Stephen N. Davis All organ systems innervated by the autonomic nervous system may be impaired in patients with diabetes. The extent of impairment is highly variable from patient to patient, but it is generally related to duration of diabetes as with other complications of diabetes. Nerve impairment is further complicated by the normal decline in function with age, but also grossly correlates with degree of hyperglycemia in large epidemiological studies [1–3,3]. Cardiac autonomic neuropathy (CAN) in diabetes is particularly important to recognize because of the numerous studies confirming elevated mortality in diabetic patients with CAN. Autonomic testing in the diabetic population may be helpful for prognosis and may guide physicians to more aggressively control cardiovascular risk factors in this population. Currently, the only preventative treatment for diabetic autonomic dysfunction is maintenance of near-normal blood glucose. Each affected system is reviewed later in this chapter, but in general, manifestations of autonomic dysfunction are treated symptomatically when appropriate. Often, patient education can avoid potentially lifethreatening problems.

IRIS Sympathetic nerves, which dilate the iris, show earlier and more extensive impairment than the parasympathetic nerves that constrict the iris. Pupillometry, which is used to measure the ability of pupils to change size, suggests that pilocarpine hypersensitivity demonstrates early parasympathetic autonomic dysfunction in diabetes mellitus before evidence of sympathetic defects. The imbalance between parasympathetic and sympathetic nerves causes an inability to respond quickly to a dark stimulus. Patients will complain of inability to see in dark places such as the movie theater and will have difficulty driving at night. On clinical examination, small, poorly dilated pupils in a dark room combined with the above complaints are indicative. Impaired pupil dilation can be observed as early as childhood in type I diabetes and may improve with metabolic control [4]. Education about recognition of the problem, reassurance, and proper precautions when in darkness are adequate in most cases. Treatment with sympathetic stimulants or parasympathetic blockers generally is not necessary.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00098-6

ESOPHAGUS Dysphagia is the most common presenting symptom of diabetic autonomic dysfunction. This is often confused with cardiac pain, gastric atony, or both. Diagnosis of esophageal and pharyngeal motility dysfunction through barium swallow generally is adequate. Other sources of pain should be ruled out. Currently, an effective drug therapy does not exist.

STOMACH Signs and symptoms of gastric emptying abnormalities include early satiety, nausea, vomiting, “brittle” diabetes, large fluctuations in blood glucose, and weight loss. The prevalence of gastroparesis is 30–50% of patients with longstanding type I and type II diabetes [5]. Some patients may be asymptomatic. The current gold standard for evaluation of gastric emptying abnormality is scintigraphy with a low fat, egg white meal labeled with 99mTcsulfur colloid with measurement of gastric emptying for 4 hours [6]. Scintigraphy can also measure both liquid and solid phase emptying, unlike upper gastrointestinal X-ray series, which measure only liquid phase gastric emptying. This may provide added diagnostic value given that gastric emptying of solids and nutrient liquids is poorly correlated in diabetes. The etiology of diabetic gastroparesis is likely multi-factorial, with decreased numbers of interstitial cells of Cajal which generate electrical slow waves in the stomach, deficiency of inhibitory neurotransmission, reduced numbers of vagal postganglionic neurons, smooth muscle fibrosis and abnormalities in immune cells. In addition, a reduction of nitric oxide is observed, which may reflect loss of neuronal nitric oxide synthase (nNOS) expression within myenteric neurons, and inhibition of nNOS by advanced glycation end products [7]. Gastroparesis, unlike cardiac autonomic neuropathy, does not appear to portend any increased mortality risk nor does it appear to significantly worsen over time [8,9]. Successful treatment of gastric emptying abnormalities may be complex. Hyperglycemia, per se, may result in atonic stomach. Improvement in glucose control has the potential to correct the problem. However, it is difficult, if

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98. DIAbETIC AuTonomIC DysfunCTIon

not impossible, to improve glucose control because of the mismatch between caloric absorption and insulin action onset. Therefore, it may be necessary to initially improve gastric emptying with pharmacologic agents, and then attempt to improve glucose control. Once glucose control is within acceptable limits, it is reasonable to discontinue pharmacologic therapy and evaluate whether glucose control alone is adequate therapy. Effective pharmacologic therapies with prokinetic action include metoclopramide, bethanechol, cisapride, erythromycin, domperidone, octreotide, and famotidine. Metoclopramide inhibits the dopaminergic pathway, which inhibits gastric emptying. By inhibiting this pathway, endogenous peristalsis may occur uninhibited. Severe vagal impairment will obviate its efficacy. The usual dose is 10 mg, 30 minutes before meals. Side effects include extrapyramidal symptoms, including risk of neuroleptic malignant syndrome, irreversible tardive dyskinesia, drooling, and nystagmus. Domperidone is not available in the United States but may be compounded by pharmacies, and in doses of 10–30 mg four times daily 30 minutes before meals may be helpful. Domperidone is a dopamine-2 antagonist and may have side effects of increased prolactin, gynecomastia, and galactorrhea in less than 5% of patients. Bethanechol directly stimulates the stomach through the muscarinic receptors. Thus, it is generally effective in increasing gastric emptying even in cases in which metoclopramide is not effective. To overcome an atonic stomach, bethanechol is given subcutaneously for the first 2 weeks. Beginning dose is 2.5 mg subcutaneously, 30 minutes before each meal, for approximately 3 to 4 days. Subcutaneous bethanechol is increased for the remainder of the 2 weeks. At the end of this period, patients are switched to 50 mg oral bethanechol, 30 minutes before each meal and snack. Side effects include urinary urgency, sweating, and occasional nausea. If given after or too soon before the meal, it may cause regurgitation. Cisapride increases smooth muscle activity of the intestines and has some direct ganglionic activity. About 10 to 20 mg is given before meals. Cisapride is no longer available within the United States, because it may cause QT prolongation leading to lethal cardiac arrhythmias, except through an investigational limited access program with strict entry criteria. Erythromycin works by mimicking motilin. Typically, 250 mg is given every 6 hours. The drawback to this drug is that it often interferes with antibiotic coverage. Octreotide decreases gut hormone motility inhibitors such as gastric inhibitory polypeptide. As a promotility drug, it is not as good as cisapride or bethanechol, but it is not worse than metoclopramide. Typically, 100 µg is given subcutaneously every 6 hours. Famotidine is an unusual H2 blocker in that it has a neutral effect or some prokinetic effect unlike other H2 blockers. Doses of 20 to 40 mg are given every 12 to 24 hours. This drug must be adjusted for renal function.

It is recommended that gastric emptying studies be repeated after pharmacologic therapy has begun to verify that emptying has improved. If not, other pharmacologic therapies should be considered. An efficient method to measure efficacy of therapy is to give the drug the gastric emptying study and measure the response. If a patient is unresponsive to any of these therapies, frequent small meals, six times a day, with high-calorie liquid food may be necessary to maintain adequate nutrition.

GALLBLADDER Diarrhea and the development of gallstones are symptoms of gallbladder atony. Gallstones are much more likely to develop in patients with hypercholesterolemia, and often are seen in patients with diabetes. Gallbladder disease is evaluated by observing the response of the gallbladder to a fatty meal or cholecystokinin. Cholecystectomy may be indicated in some cases.

COLON The most common gastrointestinal symptom of autonomic neuropathy is constipation. It is evaluated by clinical history. Constipation is treated with a variety of medications including high-fiber products such as psyllium, which is bulk-forming fiber acting to encourage peristaltic activity. Mineral oil and bisacodyl may also be effective. Bisacodyl acts directly on the colonic mucosa to produce normal peristalsis throughout the large intestine. Diabetic diarrhea is a common result of autonomic dysfunction. However, other causes of diarrhea need to be ruled out before making a diagnosis. Diabetic diarrhea is characterized by frequent (8–20 bowel movements per day, 300 g of stool per day), watery, persistent bowel movements and is often nocturnal. Mild steatorrhea is common. Treatment is aimed at the cause. Initially a broad-spectrum antibiotic is used to eliminate bacterial overgrowth. Inappropriate spillage of bile salts into the intestines also is a common cause of diarrhea among patients with diabetes. If spillage of bile salts is suspected, bile salt binders should be tried. Pharmacologic therapies include methylcellulose, diphenoxylate hydrochloride with atropine sulphate (10 ml, 6–24 hours). Octreotide (100 µg subcutaneously every 4–6 hours) has been effective when other treatments have failed.

BLADDER Both afferent and efferent nerves to the bladder may be affected in patients with diabetes. Afferent neuropathy results in the inability to feel the need to void. Therefore there is a decrease in frequency of urination, which may be misinterpreted as an improvement in glucose control.

IX. PERIPHERAL AUTONOMIC DISORDERS

479

ADREnAl mEDullA

The decreased frequency causes bladder stasis and may lead to an increase in the occurrence of urinary tract infections (UTIs). Efferent neuropathy generally occurs later in the course of diabetes causing incomplete voiding, dribbling, and frequent UTIs due to atonic bladder [10]. Incontinence is a late finding and is rare. Evaluation for bladder dysfunction is considered in male individuals with more than two UTIs per year and in female individuals with more than three UTIs per year. Cystometrogram effectively evaluates both afferent and efferent neuropathy. Having the patient schedule urination every 4 hours generally can treat afferent neuropathy. Efferent neuropathy is treated with bethanechol given orally at a dosage of 30 to 50 mg four times a day. Incontinence may require placement of a suprapubic catheter, permanent catheterization, or both.

PENIS The incidence of erectile dysfunction (ED), or impotence, which may result from autonomic neuropathy, is nearly three times more common in male patients with diabetes than in age-matched male individuals without diabetes. Signs and symptoms include decreased tumescence and rigidity. Retrograde ejaculation occurs rarely and may be related to sympathetic dysfunction. The diagnosis of neuropathic ED is one of exclusion (Fig. 98.1). Diagnosis of ED may be important, as studies suggest ED is independently associated with elevated cardiovascular risk, most likely due to coexistent endothelial dysfunction [11]. Treatment is aimed at the cause. Alternative medications for antihypertensive and psychotropic therapy should always be initiated before pharmacologic or mechanical measures are used. Phosphodiesterase inhibitors have been found to be effective in alleviating ED in 50–60% of patients with types I or II diabetes mellitus [12,13]. If the cause is neuropathic, yohimbine occasionally Psychogenic

Normal

is helpful for patients with early loss of penile rigidity or with pelvic steal syndrome. Suction devices or injections with phentolamine, prostaglandins, or papaverine can be useful. Contraindications for use of phosphodiesterase inhibitors include prior heart attack, atherosclerosis, angina, arrhythmia, chronic low blood pressure problems, and use of organic nitrates. Complications with yohimbine include hypertension. Injections may cause bruising, pain, and priapism. Prostheses often fail to meet the patient’s expectations and may be painful.

VAGINA A dry, thin, atrophic vaginal wall and lack of lubrication characterize vaginal autonomic neuropathy resulting in painful intercourse. Many female diabetic patients cite lack of arousal or decreased orgasm, which may improve with phosphodiesterase inhibitors [14]. Over-the-counter lubricants help to decrease pain during intercourse. However, an estrogen cream not only adds moisture but helps thicken the vaginal wall, thus preventing tearing.

ADRENAL MEDULLA In patients with severe neuropathy, there is a loss of the adrenal output of epinephrine and probably of sympathetic tone to the liver resulting in a decreased counter-regulatory response to hypoglycemia. Without the adrenergic signs and symptom, hypoglycemia may become severe before it is treated. Diagnosis of hypoglycemic unawareness is based on the lack of adrenergic responses such as tachycardia and blood pressure when blood glucose is less than 40 mg/dl. Effective management of this condition requires that the patient, family members and co-workers are taught to recognize the subtle signs

Nocturnal penile tumescence

Abnormal

History

Organic

Physical examination and laboratory

All other Femoral Testos- Sonogram causes ruled pulses terone Illness: out Trauma: Surgery in Prolactin Penile Drug Use: genitourinary Chronic Penile blood AntihyperDebilitating organs Pelvic pressure tensives Vascular Spinal Antiinsult depressants cord Tranquilizers Neuropathic Arterial Hormonal Venous

FIGURE 98.1 The diagnosis of impotence secondary to diabetic autonomic neuropathy is one of exclusion. The above algorithm is suggested to evaluate the other possible etiologies of impotence.

IX. PERIPHERAL AUTONOMIC DISORDERS

480

98. DIAbETIC AuTonomIC DysfunCTIon

and symptoms of hypoglycemia (mood changes, confusion, slurred speech, fugue-like state, and memory lapses) and how to treat it with glucagon injections and glucose.

Evaluate heart rate Increase or no change

SUDOMOTOR

Decrease

Vaso-vagal reflex

No change or slight increase

Evaluate vestibular system

Evaluate diastolic blood pressure

Abnormal sweating patterns are commonly found in patients with diabetes. Sweat glands are innervated by sudomotor fibers which are postganglionic, unmyelinated cholinergic sympathetic C-fibers. Sudomotor dysfunction tends to occur in a proximal to distal fashion, but can also occur in bands around the trunk from thoracic radiculopathy. Over time, both upper and lower extremities fail to sweat, whereas the trunk overcompensates. Eventually, complete anhidrosis may occur. The patient generally complains of excessive sweating in the trunk area and face. Abnormal temperature regulation predisposes to heat stroke and heat exhaustion. Treatment is limited to education. Anhidrosis is also more likely in patients on anticholinergic medications. Patients should be warned that they are at increased risk for heat stroke and heat exhaustion, and that necessary precautions should be taken [15].

CARDIOVASCULAR Clinical manifestations of cardiovascular autonomic neuropathy (CAN) include exercise intolerance and painless myocardial ischemia. Patients with CAN also tend to have attenuation of nocturnal blood pressure fall, which may contribute to cardiovascular risk. Postural hypotension as a result of vascular autonomic dysfunction is characterized by postural dizziness, nausea, vertigo, weakness, presyncope, or syncope. These signs and symptoms can be misinterpreted as hypoglycemia. Measurement of change in heart rate and blood pressure when moving from sitting or lying to a standing position provide evidence of this condition. This is a diagnosis of exclusion. Orthostatic hypotension from CAN is typically a late occurring event, so early orthostatic hypotension within the first five years of diabetes diagnosis should prompt evaluation for other etiologies (Fig. 98.2). Treatment of postural hypotension resulting from autonomic neuropathy can be complex because supine hypertension and upright hypotension often occur concurrently. Both the supine hypertension and upright hypotension cannot be treated in the same patient because treatment of one may aggravate the other and vice versa. Keeping the head of the bed in an upright position during sleep often helps the patient adjust to change in position and lessons supine hypertension. Fludrocortisones (0.1–0.5 mg) increases plasma volume and catecholamine sensitivity and often is effective. Sympathetic stimulants (ephedrine and midodrine) also are useful. Midodrine, a selective alpha-1 agonist is often efficacious at doses of 5–10 mg up to three times a day. The simplest and most efficacious treatment is an atomized spray of 10% phenylephrine

Decrease

Evaluate plasma norepinephrine response Decrease or normal response

Supraphysiologic response

Decrease circulary volume

Impaired vascular sympathetic nervous system

FIGURE 98.2 Postural hypotension as a result of vascular autonomic dysfunction is characterized by symptoms that may be misinterpreted as hypoglycemia. The above algorithm may be used to evaluate the etiology of postural hypotension.

hydrochloride (neo-synephrine). The spray is used approximately every 2 to 4 hours, 3 to 4 sprays per nostril at each dosing. This nearly always results in an improvement in upright hypotension; rare complications include septum perforation and ulceration. Cardiac denervation may also cause exercise intolerance in patients with diabetes. Symptoms may be vague and the patient may present with only fatigue. The American Diabetes Association recommends that all patients with diabetes who are new to an exercise program and are older than 40 years with type II diabetes or have had type I diabetes for more than 15 years should have an exercise tolerance test before initiating an exercise program. There is no known treatment for exercise intolerance, although one study has shown that aldose reductase inhibitors may improve intolerance [16]. CAN may be evaluated with simple noninvasive tests such as heart rate variability (R-R variation), heart rate changes during deep breathing at 6 breaths per minute (15 beat variation is normal for patients younger than 60 years), heart rate ratio during and after a standardized Valsalva maneuver of 40 mm for 10 seconds (1.21) is normal for patients younger than 60 years. Cardiac parasympathetic dysfunction may be evident by decreased heart rate variation to these maneuvers. These heart rate variations decrease with level of dysfunction and age. Scintigraphy studies with [123I]-metaiodobenzylguanidine [MIBG] and single photon emission computed tomography, and quantitative assessment [11C]-hydroxyephedrine [HED] and positron emission tomography can be used to assess cardiac sympathetic integrity. Frequency domain indexes from spectral analysis of heart rate variability from short and long ECG recordings have also been used to measure parasympathetic and sympathetic modulation in research studies, although these methods require further investigation.

IX. PERIPHERAL AUTONOMIC DISORDERS

481

CARDIovAsCulAR

Type I patients whose duration is >15 years Type II patients whose age is >50 years

No history of cardiac disease No symptoms of cardiac disease

Two or more risk factors RR-Variation > 10 (denervated or or of CHD disease* heart)

Stage III neuropathy

FIGURE 98.3 BThe above criteria are used to identify the patients at risk for painless myocardial ischemia. *, series of risk factors.

Awareness of CAN may decrease long-term morbidity and mortality rates. Cardiac denervation syndrome, a total loss of innervation to the heart, is the most severe complication of CAN, and results in exercise intolerance, poor anesthesia outcome, pregnancy complications, sudden death and probably cardiomyopathy and painless myocardial ischemia [17]. Painless myocardial ischemia, associated with increased morbidity and mortality in patients with diabetes, is more common than in the nondiabetic population and is likely secondary to CAN. An algorithm to identify those at risk for painless ischemia is provided in Figure 98.3. A study in a small number of patients has shown that approximately 66% of patients screened via this algorithm had ischemia confirmed by stress thallium testing. Unfortunately, there is no evidence that the occurrence of sudden death and cardiomyopathy can be prevented. The ACCORD study (Action to Control Cardiovascular Risk in Diabetes) suggested that intensive glycemic control worsened cardiac outcomes [18]. This suggests that early focus on glycemic control to prevent the occurrence of CAN is likely more helpful than later attention to hyperglycemia once CAN is already present.

References [1] Dyck PJ, Davies JL, Clark VM, et al. Modeling chronic glycemic exposure variables as correlates and predictors of microvascular complications of diabetes. Diabetes Care Oct, 2006;29(10):2282–8. [2] Tesfaye S, Chaturvedi N, Eaton SE, et al. Vascular risk factors and diabetic neuropathy. N Engl J Med Jan, 2005;352(4):341–50. [3] The effect of intensive treatment of diabetes on the development and progression of long-term complications in insulin-dependent diabetes mellitus. The Diabetes Control and Complications Trial Research Group. N Engl J Med 1993;329(14):977–86. [4] Karavanaki K, Baum JD. Coexistence of impaired indices of autonomic neuropathy and diabetic nephropathy in a cohort of children with type 1 diabetes mellitus. J Pediatr Endocrinol Metab 2003;16(1):79–90. [5] Horowitz M, O'Donovan D, Jones KL, Feinle C, Rayner CK, Samsom M. Gastric emptying in diabetes: clinical significance and treatment. Diabet Med March, 2002;19(3):177–94. [6] Abell TL, Camilleri M, Donohoe K, et al. Consensus recommendations for gastric emptying scintigraphy: a joint report of the American Neurogastroenterology and Motility Society and the Society of Nuclear Medicine. J Nucl Med Technol March, 2008;36(1):44–54.

[7] Chang J, Rayner CK, Jones KL, Horowitz M. Diabetic gastroparesis-backwards and forwards. J Gastroenterol Hepatol Jan, 2011;26 (Suppl. 1):46–57. [8] Jones KL, Russo A, Berry MK, Stevens JE, Wishart JM, Horowitz M. A longitudinal study of gastric emptying and upper gastrointestinal symptoms in patients with diabetes mellitus. Am J Med Oct, 2002;113(6):449–55. [9] Kong MF, Horowitz M, Jones KL, Wishart JM, Harding PE. Natural history of diabetic gastroparesis. Diabetes Care Dec, 1999;22(3):503–7. [10] Daneshgari F, Liu G, Birder L, Hanna-Mitchell AT, Chacko S. Diabetic bladder dysfunction: current translational knowledge. J Urol Dec, 2009;182(6 Suppl):S18–26. [11] Lee JH, Ngengwe R, Jones P, Tang F, O'Keefe JH. Erectile dysfunction as a coronary artery disease risk equivalent. J Nucl Cardiol Nov, 2008;15(6):800–3. [12] Goldstein I, Young JM, Fischer J, Bangerter K, Segerson T, Taylor T. Vardenafil, a new phosphodiesterase type 5 inhibitor, in the treatment of erectile dysfunction in men with diabetes: a multicenter double-blind placebo-controlled fixed-dose study. Diabetes Care March, 2003;26(3):777–83. [13] Stuckey BG, Jadzinsky MN, Murphy LJ, et al. Sildenafil citrate for treatment of erectile dysfunction in men with type 1 diabetes: results of a randomized controlled trial. Diabetes Care Feb, 2003;26(2):279–84. [14] Schoen C, Bachmann G. Sildenafil citrate for female sexual arousal disorder: a future possibility? Nat Rev Urol Apr, 2009;6(4):216–22. [15] Low PA. Clinical Autonomic Disorders, Second ed. Philadelphia: Lippincott-Raven Publishers; 1997. [16] Johnson BF, Nesto RW, Pfeifer MA, et al. Cardiac abnormalities in diabetic patients with neuropathy: effects of aldose reductase inhibitor administration. Diabetes Care Feb, 2004;27(2):448–54. [17] Ewing DJ, Campbell IW, Clarke BF. The natural history of diabetic autonomic neuropathy. Q J Med 1980;49(193):95–108. [18] Pop-Busui R, Evans GW, Gerstein HC, et al. Effects of cardiac autonomic dysfunction on mortality risk in the Action to Control Cardiovascular Risk in Diabetes (ACCORD) trial. Diabetes Care 2010;33(7):1578–84.

Further Reading Kirby RS, Carson CC, Webster GD, editors. Impotence: diagnosis and management of male erectile dysfunction. Oxford: Butterworth Heinemann; 1991. Low PA, editor. Clinical Autonomic Disorders: evaluation and management. Boston: Little, Brown; 1991. Ward J, Goto Y, editors. Diabetic Neuropathy. New York: Wiley; 1990.

IX. PERIPHERAL AUTONOMIC DISORDERS

C H A P T E R

99 Amyloidotic Autonomic Failure Yadollah Harati, Cecile L. Phan The term amyloid was coined by Rudolph Virchow in 1854 to describe a macroscopic tissue deposit that displayed a positive reaction on staining with iodine. Since then, our knowledge about amyloid and amyloidosis has dramatically expanded. There are now more than 20 different protein molecules that are known to undergo conformational changes leading to the generation of amyloid deposits. These are homogeneous insoluble protein deposits, consisting of polypeptide fibrils, 7.5 to 10 nm in width, arranged in highly ordered aggregates of β-pleated sheets, conferring to them a high degree of stability. Amyloid deposits display a characteristic apple green birefringence when examined under polarized light after staining with Congo red (Fig. 99.1). Amyloidosis refers to a wide spectrum of disorders that result from abnormal extracellular deposits of amyloid. These deposits may be widespread (systemic amyloidosis) or restricted to certain organs (localized amyloidosis). Previous classification of amyloidosis was confusing and ever changing. Refined knowledge about the chemical composition of the amyloid deposits, as well as the recognition of hereditary amyloidosis, has resulted in a much more rational classification (Table 99.1). The current nomenclature system is based on the type of protein forming the amyloid deposits. The clinical presentation of amyloid deposits depends on the organs involved and the size of the amyloid fibrils. Involvement of the peripheral nervous system is an important feature of the systemic amyloidoses and typically presents with the classical clinical triad of small fiber neuropathy, autonomic neuropathy (AN), and carpal tunnel syndrome (CTS). Autonomic failure is an important feature of immunoglobulin amyloidosis and hereditary systemic amyloidosis and must be considered in all patients with familial or paraproteinemic neuropathies having autonomic dysfunction (especially when CTS coexists). A recent retrospective chart review of 65 patients with biopsy proven amyloidosis and autonomic function testing showed that the majority of patients (94%) have symptoms of generalized autonomic failure, and moderately severe failure on autonomic function testing was also found in patients who were asymptomatic. The mechanism of nerve injury is probably the same in all amyloidoses, and it involves physical pressure exerted by amyloid deposits on dorsal root ganglia or autonomic ganglia, or

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00099-8

directly on nerve fibers, leading to loss of normal tissue elements and disorientation of tissue architecture. Toxic, ischemic, and immunologic mechanisms have been suggested as alternatives. The following sections discuss amyloidoses that may demonstrate autonomic involvement.

IMMUNOGLOBULIN AMYLOIDOSIS The amyloid fibrils in immunoglobulin amyloidosis amyloidopathy are composed of immunoglobulin light chain proteins or their degradation products. AL is the most common form of systemic amyloidosis. The monoclonal immunoglobulin light chains are produced by monoclonal plasma cells, which may be derived from nonproliferative populations or from a malignant clone, such as in multiple myeloma (MM), Waldenstrom’s macroglobulinemia, non-Hodgkin’s lymphoma, and solid tumors like hypernephroma. In amyloidosis associated with nonmalignant AL, the peripheral nervous system is involved in more than 50% of patients and is the presenting symptoms in 40% of those cases. In amyloidosis associated with MM, clinical and electrophysiologic evidence of polyneuropathy develops in 13% and 40% of patients, respectively. The peripheral neuropathy in AL is axonal, distal, symmetric, and sensory with impaired pinprick and temperature more than vibratory and proprioceptive sensations, which is consistent with a small fiber neuropathy. As the disease progresses, large fiber involvement appears. Subtle autonomic abnormalities, as detected by autonomic function tests, seem to be prevalent even in asymptomatic patients. Both sympathetic and parasympathetic systems are affected to different extents. Signs of AN include orthostatic hypotension (OH) with inappropriate heart rate response, impotence, dry mouth, gastrointestinal (GI) autonomic disturbances (dysphagia, early satiety, diarrhea, and constipation), sluggish pupillary action, impairment of sweating, and bladder on. Autonomic dysfunction, rather than deposition of amyloid in the mucosa, seems to be a more frequent cause of GI symptoms in these patients. Target organs other than in the peripheral nervous system include the heart, skeletal muscles, liver, intestine, spleen, kidney, tongue, and skin.

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99. AmyloIdoTIC AuTonomIC FAIluRE

TABLE 99.1 Peripheral Autonomic Failure Classification of Amyloidosis

Systemic amyloidosis

Type

Previous Name

Subunit Protein

PNS Involvement

Ig derived (AL)

Primary/myeloma associated

Ig light chain

PN, AN, CTS

Reactive (AA)

Secondary (acquired)

A protein

PN, AN

Dialysis associated

B2-microglobulin

Hereditary

Heredofamilial

TTR

CTS

FAP I,II

Apolipoprotein I, II

PN, AN, CTS

FAP III

Gelsolin

PN

FAP IV

Fibrinogen

PN, CN

–

Lysosome

None

– Localized amyloidosis

None

Alzheimer’s disease: (H,A), HCHA(H), CAA(A), Genitourinary (A) Lichen (A) Cutaneous (H,A) IBM(H,A)

A, acquired; AN, autonomic neuropathy; CAA, cerebral amyloid angiopathy; CN, cranial neuropathy; CTS, carpal tunnel syndrome; FAP, familial amyloid neuropathy; H, hereditary; HCHA, hereditary cerebral hemorrhage with amyloidosis; IBM, inclusion body myopathy; Ig, immunoglobulin; PNS, peripheral nervous system; PN, peripheral neuropathy.

FIGURE 99.1 Amyloid deposits. (A) Hematoxylin and eosin staining. (B) Crystal violet. (C) Teased nerve preparation. (D) Congo red under fluorescence.

Pathogenesis

Diagnosis

Pathologic investigations have shown infiltration of dorsal root and sympathetic ganglia in patients with amyloidotic neuropathy early in the course of the disease. Intermediolateral cell column neurons at the T7 level of the spinal cord, which contains the cell bodies of preganglionic sympathetic neurons, may also be reduced.

Tests of autonomic function are often abnormal. Various methods are employed to identify and quantify the amount of free light chain in serum and urine; the most two common tests are protein electrophoresis and immunofixation with the latter one being more sensitive at detecting paraproteins at smaller quantities.

IX. PERIPHERAL AUTONOMIC DISORDERS

ImmunoglobulIn AmyloIdosIs

Nephelometric free light chain assay is a newer technique which is reported to be even more sensitive than immunofixation. Serum and urine protein electrophoresis detect a monoclonal gammopathy (M protein) in 80% of cases. Lambda light chains are three times as frequent as kappa. The presence of monoclonal protein helps to distinguish patients with AL amyloidosis from the hereditary type. Erythrocyte sedimentation rate is commonly increased. Anemia and proteinuria are common. To identify amyloid on biopsy specimens, Congo red is the most specific stain, whereas electron microscopic examination of the amyloidladen tissues is the most sensitive method of recognizing this disorder. Abdominal fat aspiration and rectal biopsy show amyloid in 80% of cases, and bone marrow stains positive for amyloid in 50% of cases. The bone marrow also provides an estimate of the number of plasma cells. In AL without MM, 3 to 5% of the bone marrow cells typically are plasma cells with no malignant features. This is increased in AL with MM to more than 50%, and many plasma cells display malignant features. Minor salivary gland biopsy also has a sensitivity of about 86%. Sural nerve biopsy provides another tissue source for diagnosis. In patients suspected of having amyloidosis, any tissue obtained during any surgery (e.g., flexor retinaculum during carpal tunnel surgery) may contain amyloid and should be specifically examined for amyloid deposit. Iodine-123 serum amyloid P component (123I-SAP) scintigraphy allows for safe and reliable diagnosis and monitoring of progression of disease and response to treatment. Although SAP-scintigraphy negates the need for invasive biopsy procedures, it might not be sufficient to replace pathologic evidence of amyloid on tissue samples. Additionally, SAP-scintigraphy does not detect cardiac amyloid. Once amyloid deposits are identified, further tissue typing to determine the specific amyloid forming protein can also be done with immunohistochemistry using specific anti-sera, although this method is not very reliable. Recently, a new technique using laser microdissection and tandem mass spectrometric-based proteomic analysis reportedly carries a 100% sensitivity and specificity for detecting specific types of amyloid in nerve biopsy specimens independent of clinical information.

Treatment Treatment approaches to AL amyloidosis are either specific, geared to suppress or reverse the disease process, or symptomatic. Specific treatments include high dose immunosuppressive or cytotoxic agents to suppress the production of light chains by the plasma cell clone. High dose intravenous melphalan with autologous blood stem cell transplantation results in hematologic remission, improved five-year survival, and reversal of amyloid-related disease in a significant number of patients when compared to no treatment. It should be cautioned, however, that patients with AL amyloidosis have multisystem disease especially cardiac involvement which puts

485

them at risk for serious toxicity and adverse events from these treatments. In patients unable to tolerate autologous stem cell transplantation, the combination of high dose dexamethasone and mephalan is an effective therapeutic option. Etanercept may also be used in patients refractory or ineligible to other modalities. In some select cases with specific organ involvement (heart or liver), organ transplantation may be successfully attempted, with recovery of organ function. Thalidomide, and its less toxic analog, lenalidomide, have been tried with some promise, but thalidomide is poorly tolerated in patients with polyneuropathy. Bortezomib, a proteosome inhibitor, together with dexamethasone were able to produce hematologic response a large number of patients. A murine derived antibody against human light chain was able to induce regression in amyloid formation in a mouse model. Symptomatic approaches to treatment target various complaints, mostly those associated with autonomic dysfunction. Metoclopramide has been used to treat early satiety, whereas octreotide, a somatostatin analog, has been successfully used to treat amyloidosis-associated diarrhea. Cisapride, because of its arrhythmias risk, is currently not widely available although it was reported to be useful in intestinal pseudo-obstruction in AL associated with MM. Its use in this particular population of patients with potential cardiac amyloidotic involvement should probably be best avoided altogether. Although fludrocortisone is the mainstay of treatment of OH, midodrine, pyridostigmine, erythropoietin, and l-threo-3,4-dihydroxyphenylserine (DOPS) are also reported to be helpful. Other therapies for OH include elastic support extending to the waist and instructions to the patients to rise slowly from the lying position and to sit on the edge of the bed for a few minutes before walking. In patients with syncope, investigation of cardiac arrhythmias or heart block can be life-saving. Treatment of the non-neurologic manifestations of this condition also is symptomatic (diuretics, antiarrhythmics, etc.).

Prognosis The prognosis of AL remains poor. The 5-year survival rate is only 20%. Patients with MM have a poor prognosis with a median survival of 24 to 36 months. Death is attributed to cardiac involvement from-congestive heart failure or arrhythmias in 50% of patients. The median survival of patients with AL who have congestive heart failure is only 4 months. The impact of more aggressive therapies, such as autologous stem cell transplantation or organ transplantation, on prognosis is still unclear. AL amyloidosis presenting solely as neuropathy has a better prognosis with a median survival of more than 5 years. The influence of the AN on the evolution of the disease has been evaluated. Patients with AL with AN was 7.3 months versus 14.8 months for those without AN. Patients with prolonged QT interval on electrocardiogram had even shorter survival periods. The prolonged QT is believed to result from autonomic dysfunction rather than a primary cardiac disorder.

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99. AmyloIdoTIC AuTonomIC FAIluRE

REACTIVE AMYLOIDOSIS Reactive amyloidosis (AA) is associated with several chronic inflammatory diseases like rheumatoid arthritis, systemic lupus erythematosus, chronic inflammatory bowel disease (especially Crohn’s disease), tuberculosis, leprosy, osteomyelitis, Castleman’s disease, and suppurative infections. The amyloid in these conditions contains a degradation product of an acute phase serum protein (A protein), which is a nonimmunoglobulin. Renal insufficiency and nephrotic syndrome is seen in 90% of patients. AN has been reported only rarely in association with this type of amyloidosis.

HEREDITARY AMYLOIDOSIS Hereditary amyloidoses have been linked to various proteins, including transthyretin (TFR), gelsolin, apolipoprotein AI and AII, lysozyme, and fibrinogen. The most common hereditary amyloidosis is familial amyloid polyneuropathy (FAP), associated with mutated TTR, apolipoprotein A, or gelsolin. The latter two subtypes are exceedingly rare, mostly restricted to two kindreds in the United States (in Iowa) and Italy (both apolipoprotein AI), and a few kindreds in Finland (gelsolin). Significant autonomic dysfunction has only been described in TTR amyloidosis. A few patients with apolipoprotein Al amyloidosis have mild gastroparesis as only sign of autonomic involvement. Transthyretin FAP, an autosomal dominant disorder, is the most common type of familial amyloidotic polyneuropathy. TTR is a normal plasma protein tetramer. Each TTR monomer is composed of 127 amino acids synthesized in the liver and choroid plexus. TTR is involved in the transport of vitamin A and thyroid hormones. It is coded by a single gene located on chromosome 18. Most patients with TTR amyloidosis are heterozygous for that gene. Currently, more than 100 TTR point mutations have been described, the most common being the one that substitutes methionine to valine at position 30 (Val30Met). Clusters of the Val30Met mutation are found in Portugal, Sweden, and Japan, although TTR FAP is found worldwide. The most common clinical manifestations of TTR-FAP are peripheral neuropathy and cardiomyopathy. However, phenotypic variations are known to be quite common, and can mimic common neuropathies including chronic inflammatory demyelinating polyneuropathy (CIDP). This is especially true for the sporadic cases of TTR-FAP without any documented family history which tend to have later onset and milder autonomic dysfunction. Delay in diagnosis also delays important treatment of potentially life-threatening autonomic dysfunction and more definitive treatment such as orthotopic liver transplant. Autonomic involvement in TTR amyloidosis is common and occurs early with both sympathetic and parasympathetic systems being affected. The severity of autonomic involvement generally correlates with the progression of the disease. Autonomic symptoms include alternating diarrhea

and constipation, palpitation caused by cardiac dysrhythmias, anorexia, nausea and vomiting, OH, impotence, urinary and fecal incontinence, and hypohidrosis. Delayed gastric emptying may cause organ distension and anorexia with subsequent cachexia, which may be an important factor in mortality. GI dysfunction is caused by autonomic involvement and direct deposition of amyloid in the intestinal wall. The scalloped pupil deformity has been described in Portuguese and Swedish kindreds and is probably caused by involvement of the ciliary nerves. AN may also cause severe urinary retention with secondary renal damage. Although diagnosis should not be difficult in typical cases, other hereditary neuropathies like hereditary motor and sensory neuropathies and hereditary sensory and autonomic neuropathies should be excluded. The late onset, predominant sensory symptoms, prominent early autonomic involvement, and frequent association with CTS strongly favor the diagnosis of FAP. Nevertheless, nerve biopsy and DNA analysis may be required for confirmation of diagnosis.

Pathogenesis Normal TTR is not amyloidogenic, although it is arranged in an extensive β-sheet conformation. Most amyloidogenic mutations increase the content of the β-pleated sheets and result in the production of an insoluble amyloidogenic monomer. Met30Val TTR, which is the most common mutation, demonstrated amyloidogenicity in transgenic mice. Amyloid deposits were eventually noted in intestine, kidney, and heart, but not in peripheral nerves (the most common site of involvement in humans). Demonstration of amyloid deposits in dorsal root ganglia and sympathetic chains in FAP is, perhaps, because of the absence of blood–nerve barrier in these structures, allowing easy access of amyloidogenic protein. The mechanism of OH in patients with FAP is not fully understood. Low levels of plasma norepinephrine in patients with OH without significant plasma norepinephrine response to postural changes and low serum dopamine-hydroxylase activity suggest depletion of peripheral norepinephrine secondary to adrenergic denervation.

Laboratory Data and Diagnosis Autonomic function tests are abnormal frequently and early in the course of the disease. Amyloid tissue diagnosis is not different from AL. DNA analysis allows the detection of specific mutations and provides valuable information for genetic counseling of family members at risk. This test can be applied to chorionic villi samples for prenatal diagnosis.

Treatment and Prognosis If left untreated, TTR amyloidosis is invariably fatal, with death occurring within 5 to 15 years. The only

IX. PERIPHERAL AUTONOMIC DISORDERS

HEREdITARy AmyloIdosIs

available therapeutic approach is orthotopic liver transplantation (OLT). Because the liver is the main source of TTR, OLT is supposed to eliminate the bulk of mutated TTR. More than 1500 patients worldwide had undergone OLT for TTR amyloidoisis with a 5-year survival rate of approximately 80% for the Val30Met mutations. A recent electrophysiologic study looking at the 10-year follow-up post OLT of 8 patients with Val30Met mutation showed that OLT can halt the progression of the neuropathy. Survival, however, is much lower for the other TTR mutations (50–60%). Many of these patients continue to have progression of systemic amyloid deposits after OLT, presumably due to ongoing deposition of wild-type TTR. Patients who have a combined liver and heart transplant do not have cardiac deposits at time of death but combined transplant does not prevent amyloid deposit in peripheral nerves. Due to obvious challenges of organ transplantations, other methods of treatment are being investigated. Recent studies have focused on using small molecules which bind to the transthyretin tetramer, stabilize it, and prevent it from dissociating into partially unfolded and toxic monomeric components. Diflunisal, an NSAID, is one such molecule and early studies are encouraging. Gene therapy with small interfering RNAs, antisense oligonucleotides, and single-stranded oligonucleotides in animal studies are underway.

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Benson M, Kincaid J. The molecular biology and clinical features of amyloid neuropathy. Muscle & Nerve 2007;36:411–23. Gertz M, Zeldenrust S. Treatment of immunoglobulin light chain amyloidosis. Curr Hem Ma Rep 2009;4:91–8. Hund E, Linke RP, Willig F, Grau A. Transthyretin associated neuropathic amyloidosis, pathogenesis and treatment. Neurology 2001;56:434–5. Klein C., Vrana, J., Theis, J., Dyck, P., Dyck, P.J.B., Spinner, R., et al., 2010. Mass spectrometric-based proteomic analysis of amyloid neuropathy type in nerve tissue. Arch Neurol. Epub ahead of print. Liepnieks J, Zhang L, Benson M. Progression of transthyrein amyloid neuropathy after liver transplant. Neurology 2010;75(4):324–7. Merlini G, Bellotti V. Molecular mechanisms of amyloidosis. N. Engl J. Med. 2003;349:583–96. Pepys MB. Pathogenesis, diagnosis and treatment of systemic amyloidosis. Phil. Trans. R. Soc. Lond 2001;356:203–11. Perfetto F, Moggi-Pignone A, Livi R, Tempestini A, Bergesio F, MatucciCerinic M. Systemic amyloidosis: a challenge for the rheumatologist. Nat Rev Rheum 2010;6:417–29. Plante-Bordeneuve V, Ferreira A, Lalu T, Zaros C, Lacroix C, Adams D, et al. Diagnostic pitfalls in sporadic transthyretin familial amyloid polyneuropathy (TTR-FAP). Neurology 2007;69(7):693–8. Planté-Bordeneuve V, Said G. Transthyretin related familial amyloid polyneuropathy. Curr Opin. Neurol. 2000;13:569–73. Shimojima Y, Morita H, Kobayashi S, Takei Y, Ikeda S. Ten year follow up of peripheral nerve function in patients with familial amyloid polyneuropathy after liver transplantation. J Neurol 2008;255:1220–5. Skinner M, Sanchorawala V, Seldin DC, Dember LM, Falk RH, Berk JL, et al. High dose melphalan and autologous stem-cell transplantation in patients with AL amyloidosis: An 8-year study. Ann. Intern. Med. 2004;140(2):85–93.

Further Reading Adams D, Samuel D, Goulon-Goeau C, Nakazato M, Costa PM, Feray C, et al. The course and prognostic factors of familial amyloid polyneuropathy after liver transplantation. Brain 2000;123:1495–504.

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100 Autoimmune Autonomic Ganglionopathy Steven Vernino, Phillip A. Low Acquired autonomic failure can result from dysfunction of peripheral autonomic nerves or ganglia. Chronic progressive autonomic neuropathy may occur in the context of a more diffuse peripheral neuropathy associated with diabetes, amyloidosis or other definable causes. A chronic idiopathic dysautonomia without sensorimotor neuropathy has customarily been called pure autonomic failure (PAF, first described by Bradbury and Eggleston), but this arbitrary classification probably encompasses disorders with several different pathophysiologies including chronic immunemediated autonomic failure. On the other hand, most cases of subacute autonomic failure (onset to peak deficit within 3 months) are attributable to neurological autoimmunity. This is also true for acute autonomic neuropathy when toxic and metabolic causes are excluded. Subacute autonomic neuropathies can be further divided into dysautonomia associated with sensory and motor neuropathy (acute inflammatory neuropathies such as Guillain–Barré syndrome), dysautonomia associated with malignancy (paraneoplastic autoimmune autonomic neuropathy) or idiopathic autoimmune autonomic ganglionopathy (AAG).

AUTOIMMUNE AUTONOMIC GANGLIONOPATHY Description The typical presentation of AAG (formerly known as acute pandysautonomia or idiopathic subacute autonomic neuropathy) is highly characteristic [1]. The typical syndrome occurs in a previously healthy individual in whom autonomic failure develops over the course of a few days or weeks. The onset of symptoms may follow a viral prodrome, minor surgical procedure, or routine immunization. There is a slight female predominance. The average age of onset in reported cases is around 55 years, but there is a wide age range. The most common syndrome is severe generalized sympathetic and parasympathetic autonomic failure. Sympathetic failure is manifested as orthostatic hypotension and anhidrosis, and parasympathetic failure as dry mouth, dry eyes, sexual dysfunction, constipation, impaired pupillary light response, and fixed heart rate. Gastrointestinal dysmotility is common (70% of patients) and presents with symptoms of anorexia, early satiety,

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00100-1

postprandial abdominal pain and vomiting, constipation or diarrhea [1]. In severe cases, objective autonomic abnormalities are obvious and include orthostatic hypotension, tonic pupils, and a fixed heart rate. The spectrum and severity of dysautonomia, however, varies from patient to patient. Less common presentations are selective cholinergic failure, selective adrenergic neuropathy, or isolated gastrointestinal dysmotility. About 25% of patients report neuropathic symptoms, such as tingling in the distal extremities, but sensory examination and nerve conduction studies are normal. Patients with objective evidence of sensory neuropathy may be better classified as subacute sensory and autonomic neuropathy.

Pathogenesis The most convincing evidence of an autoimmune pathogenesis for AAG is the demonstration of high titers of ganglionic nicotinic acetylcholine receptor (AChR) antibodies in the serum of about 50% of patients [2]. A number of studies have shown that the serum level of this antibody correlates with severity of symptoms [2,3]. For orthostatic hypotension, there is a sigmoidal relationship [4,5]. Orthostatic hypotension becomes prominent when antibody levels are greater than 1 nmol/L, worsens as antibody levels rise, and then reaches a maximal severity. Similar antibody thresholds likely exist for other autonomic symptoms in AAG, such as sicca symptoms and bladder dysfunction. In some series, only patients with antibody levels higher than 3 nmol/L had clinically evident impairment in pupillary light reflex [4]. As further proof that AAG is an antibody-mediated disorder, experimental AAG can be produced in animal models. Rabbits immunized against the ganglionic AChR or mice given ganglionic AChR antibody by passive transfer develop autonomic dysfunction similar to patients with AAG [6,7]. Antibodies from AAG patients specifically inhibit membrane currents through α3-type ganglionic AChR [8]. In addition, ganglionic AChR antibodies rapidly inhibit ganglionic synaptic transmission in isolated sympathetic ganglia [9]. After passive transfer of antibodies to mice, quantal size (the responsiveness of the postsynaptic neuron) is reduced for up to 2 weeks [9]. These antibody effects do not require complement and appear

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to result from antibody crosslinking and internalization of synaptic ganglionic AChR [10]. Patients with AAG and rabbits with EAAG have a unique laboratory profile consistent with impaired ganglionic synaptic transmission. Levels of plasma catecholamines are low while imaging of cardiac sympathetic innovation is normal [11,12]. This pattern of normal cardiac sympathetic innervation with reduced sympathetic activity is unique, and best explained by impairment of neurotransmission at the level of the autonomic ganglia.

Diagnosis The diagnosis of idiopathic AAG is suspected in cases of acquired autonomic failure without somatic neuropathy when toxic or paraneoplastic causes have been excluded. Patients with paraneoplastic autonomic neuropathy may be clinically indistinguishable from idiopathic AAG until a cancer, usually small-cell carcinoma of the lung, is detected. History of an antecedent event (viral syndrome or surgical procedure) or a personal or family history of other autoimmune disorders (such as autoimmune thyroiditis, pernicious anemia, type I diabetes, or myasthenia gravis) supports the diagnosis of AAG. A high serum level of ganglionic AChR antibody confirms the diagnosis of AAG [2,13]. Based on a review of autonomic findings in seropositive patients [3], a particular combination of “cholinergic” autonomic symptoms (neurogenic bladder, impaired pupillary function, gastroparesis and dry eyes and dry mouth) are most suggestive of AAG. Lower antibody values show more diverse clinical associations, including limited or chronic forms of autonomic failure (Table 100.1). Slowly progressive AAG may resemble pure autonomic failure (PAF), an insidious TABLE 100.1

presumed degenerative autonomic disorder with predominant sympathetic adrenergic and sudomotor failure. Gastrointestinal dysmotility and impaired pupillary responses common in AAG are not seen in PAF [14]. A comparison of the clinical features of AAG, PAF and multiple system atrophy is presented in Table 100.2. Although antibody testing is diagnostically important, many patients with the clinical features of AAG do not have ganglionic AChR antibodies, and the assessment of these seronegative cases remains difficult. Nevertheless, it is reasonable to consider an autoimmune etiology for cases of subacute autonomic failure. Several case reports describe patients with seronegative AAG that improved after treatment with immunotherapy [15]. Further investigations of seronegative AAG looking for novel diagnostic markers that can predict response to immunotherapy are still needed.

Clinical Course The original case of acute pandysautonomia [16] was remarkable for highly selective autonomic involvement and complete recovery. Subsequent case series have documented that most patients have a distinct clinical course with monophasic worsening followed by stabilization or remission without recurrences [1]. Patients can improve spontaneously, but recovery is typically incomplete. Only a third of patients experience major functional improvement of autonomic deficits. A subset of patients present with a more insidious and progressive form of AAG rather than the typical subacute monophasic presentation. Some of these patients are readily identified as AAG by their pattern of autonomic symptoms or by their serological profile (ganglionic AChR and

Association of Ganglionic AChR Antibody with Dysautonomia and other Disorders*

Diagnostic Group

% Seropositive

Antibody Levels

50%

0.5–41.0 nM/L

AUTONOMIC DISORDERS Subacute AAG [2,8]

**

Chronic AAG[3]

20–40%

0.2–5.0 nmol/L

Paraneoplastic AAG [2]

20%

0.2–20.0 nmol/L

Postural tachycardia syndrome (POTS) [23]

10%

0.25 nmol/L

Idiopathic gastrointestinal dysmotility [2]

5–10%

0.4 nmol/L

Diabetic autonomic neuropathy

10%

Multiple system atrophy [2]

0

OTHER DISORDERS Lambert–Eaton syndrome [2]

5–10%

0.06–0.4 nmol/L

Myasthenia gravis without thymoma[8]

3%

0.25 nmol/L

Paraneoplastic disorders with thymoma [24]

15–20%

0.06–2.0 nmol/L

Paraneoplastic disorders with SCLC [13]

5–10%

*Less than 0.5% of healthy controls are seropositive for ganglionic AChR antibodies [13]. **The exact frequency is not known since antibody status may affect case definition.

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PARAnEoPlAsTIC AuToImmunE AuTonomIC nEuRoPATHy

491

TABLE 100.2 Differentiation of Autoimmune Autonomic neuropathy (AAG), Pure Autonomic Failure (PAF) and multiple system Atrophy (msA) Parameter

AAG

PAF

MSA

Onset

Subacute or insidious

Insidious

Insidious

First symptom

Multiple

Orthostatism

Neurogenic bladder

Gastrointestinal Sx

Common

Absent

Uncommon

Pupillary involvement

Common

Absent

Uncommon

CNS involvement

Absent

Absent

Present

*

Somatic neuropathy

Mild/minimal

Absent

Present in 15–20%

Pain

Often present

Absent

Absent

Autonomic findings

Widespread

Limited

Relatively widespread

Progression

Often monophasic

Slow

Inexorably progressive

Prognosis

Relatively good

Relatively good

Poor

Lesion

Postganglionic

Postganglionic

Preganglionic; central

Supine plasma

Reduced

Markedly reduced

Normal

Nerve conduction studies

Usually Normal

Normal

Usually normal

Ganglionic AChR antibody

Positive (50%)

Negative†

Negative

norepinephrine

*More common in paraneoplastic cases. † Chronic AAG may be indistinguishable from PAF [7].

other organ-specific antibodies). Despite a long course of illness, chronic AAG patients may improve significantly when immunomodulatory treatment is initiated [17].

Treatment Treatment for AAG has largely been symptomatic. The convincing evidence that AAG is an antibody-mediated channelopathy supports the use of immunomodulatory therapy for AAG. A number of observational studies report benefit with IVIG, plasma exchange, mycophenolate, prednisone, azathioprine and rituximab alone or in combination [5,15,18–21]. Although patients with ganglionic AChR antibody positive AAG respond to either IVIG or plasma exchange, the benefits are often transient. Anecdotally, a combination of immunomodulatory treatment with chronic immunosuppression appears to be more effective.

PARANEOPLASTIC AUTOIMMUNE AUTONOMIC NEUROPATHY Subacute autoimmune autonomic neuropathy clinically indistinguishable from idiopathic AAG can occur as a remote effect of malignancy (most commonly small-cell carcinoma of the lung, less commonly thymoma or other neoplasm). Among paraneoplastic syndromes associated with small-cell lung carcinoma, autonomic neuropathy is far less common than sensorimotor neuropathy, polyradiculoneuropathy, or sensory neuronopathy [22]. Paraneoplastic autoimmune autonomic neuropathy may

manifest as pandysautonomia or as severe isolated gastrointestinal dysmotility. More commonly, dysautonomia in these patients is accompanied by symptoms of limbic encephalitis or other elements of a multifocal autoimmune neurological disorder. Several different autoantibody specificities may be encountered in patients with paraneoplastic autonomic neuropathy. As a group, cation channel autoantibodies are most common (ganglionic or muscle AChR antibodies, voltage-gated N-type or P/Q-type calcium channel antibodies, or neuronal potassium channel antibody). Next most common is anti-neuronal nuclear antibody type 1 (ANNA-1, also known as anti-Hu). A minority of patients will lack any currently recognized autoantibody marker. Small-cell lung carcinoma is found in at least 80% of patients who are seropositive for ANNA-1 [22].

References [1] Suarez GA, Fealey RD, Camilleri M, Low PA. Idiopathic autonomic neuropathy: Clinical, neurophysiologic, and follow-up studies on 27 patients. Neurology 1994;44:1675–82. [2] Vernino S, Low PA, Fealey RD, Stewart JD, Farrugia G, Lennon VA. Autoantibodies to ganglionic acetylcholine receptors in autoimmune autonomic neuropathies. N. Engl. J. Med 2000;343(12):847–55. [3] Klein CM, Vernino S, Lennon VA, Sandroni P, Fealey RD, BenrudLarson L, et al. The spectrum of autoimmune autonomic neuropathies. Ann. Neurol 2003;53(6):752–8. [4] Gibbons CH, Freeman R. Antibody titers predict clinical features of autoimmune autonomic ganglionopathy. Autonomic Neuroscience 2009;146:8–12. [5] Imrich R, Vernino S, Eldadah BA, Holmes C, Goldstein DS. Autoimmune autonomic ganglionopathy: treatment by plasma exchanges and rituximab. Clin. Auton. Res 2009;19(4):259–62.

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[6] Vernino S, Ermilov LG, Sha L, Szurszewski JH, Low PA, Lennon VA. Passive transfer of autoimmune autonomic neuropathy to mice. J. Neurosci 2004;24(32):7037–42. [7] Vernino S, Low PA, Lennon VA. Experimental autoimmune autonomic neuropathy. J. Neurophysiol. 2003;90(3):2053–9. [8] Vernino S, Lindstrom J, Hopkins S, Wang Z, Low PA. Characterization of ganglionic acetylcholine receptor autoantibodies. J. Neuroimmunol 2008;197:63–9. [9] Wang Z, Low PA, Vernino S. Antibody-mediated impairment and homeostatic plasticity of autonomic ganglionic synaptic transmission. Exp. Neurol 2010;222(1):114–9. [10] Wang Z, Low PA, Jordan J, Freeman R, Gibbons CH, Schroeder C, et al. Autoimmune autonomic ganglionopathy: IgG effects on ganglionic acetylcholine receptor current. Neurology 2007;68(22):1917–21. [11] Vernino S, Hopkins S, Wang Z. Autonomic ganglia, acetylcholine receptor antibodies, and autoimmune ganglionopathy. Auton Neurosci 2009;146(1–2):3–7. [12] Goldstein DS, Holmes C, Imrich R. Clinical laboratory evaluation of autoimmune autonomic ganglionopathy: Preliminary observations. Auton Neurosci 2009;146(1–2):18–21. [13] McKeon A, Lennon VA, Lachance DH, Fealey RD, Pittock SJ. Ganglionic acetylcholine receptor autoantibody: oncological, neurological, and serological accompaniments. Arch. Neurol. 2009;66(6):735–41. [14] Sandroni P, Low PA. Other autonomic neuropathies associated with ganglionic antibody. Auton Neurosci 2009;146(1–2):13–17. [15] Iodice V, Kimpinski K, Vernino S, Sandroni P, Fealey RD, Low PA. Efficacy of immunotherapy in seropositive and seronegative

[16] [17]

[18]

[19]

[20]

[21]

[22]

[23]

[24]

putative autoimmune autonomic ganglionopathy. Neurology 2009;72(23):2002–8. Young RR, Asbury AK, Corbett JL, Adams RD. Pure pan-dysautonomia with recovery. Brain 1975;98:613–36. Schroeder C, Vernino S, Birkenfeld AL, Tank J, Heusser K, Lipp A, et al. Plasma exchange for primary autoimmune autonomic failure. N. Engl. J. Med. 2005;353(15):1585–90. Gibbons CH, Vernino SA, Freeman R. Combined immunomodulatory therapy in autoimmune autonomic ganglionopathy. Arch. Neurol 2008;65(2):213–7. Kondo T, Inoue H, Usui T, Mimori T, Tomimoto H, Vernino S, et al. Autoimmune autonomic ganglionopathy with Sjögren’s syndrome: Significance of ganglionic acetylcholine receptor antibody and therapeutic approach. Auton Neurosci 2009;146(1–2):33–5. Peltier AC, Black BK, Raj SR, Donofrio P, Robertson D, Biaggioni I. Coexistent autoimmune autonomic ganglionopathy and myasthenia gravis associated with non-small-cell lung cancer. Muscle Nerve 2010;41(3):416–9. Modoni A, Mirabella M, Madia F, Sanna T, Lanza G, Tonali PA, et al. Chronic autoimmune autonomic neuropathy responsive to immunosuppressive therapy. Neurology 2007;68(2):161–2. Lucchinetti CF, Kimmel DW, Lennon VA. Paraneoplastic and oncologic profiles of patients seropositive for type 1 antineuronal nuclear autoantibodies. Neurology 1998;50(3):652–7. Thieben MJ, Sandroni P, Sletten DM, Benrud-Larson LM, Fealey RD, Vernino S, et al. Postural orthostatic tachycardia syndrome: The mayo clinic experience. Mayo Clin. Proc 2007;82(3):308–13. Vernino S, Lennon VA. Autoantibody profiles and neurological correlations of thymoma. Clin. Cancer Res 2004;10:7270–5.

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101 Guillain–Barré Syndrome Phillip A. Low, James G. McLeod CLINICAL FEATURES The core features of Guillain–Barré syndrome are the acute onset of an ascending, predominantly motor polyradiculoneuropathy associated with areflexia and elevated cerebrospinal fluid protein. The disease follows an antecedent bacterial or viral infection in about two-thirds of cases. Campylobacter jejuni infections are implicated in 17–39% of cases in North America and Europe and the frequency is higher in Asian countries [1]. Loss of tendon reflexes is usual. The ascending weakness progresses usually within 1 to 2 weeks followed by a plateau phase of about 3 weeks [2]. Severe weakness including respiratory paralysis is common. Although autonomic involvement is not a requirement for diagnosis, autonomic over or under activity is common and occurs in about two-thirds of patients [3–6]. During the acute phase, dysautonomia is dominated by sympathetic overactivity, sometimes manifested as episodes of hypertension, hyperhidrosis, and tachycardia (autonomic storms) [6], while parasympathetic failure is more evident during recovery [3–5]. Resting tachycardia is common, and orthostatic hypotension may alternate with hypertension. Bladder or bowel involvement tends to be mild compared to somatic involvement. Some patients with GBS have life-threatening dysautonomia, which presents most frequently in the acute evolving phase of the disease and tends to correlate with the severity of somatic involvement, being especially common in patients with respiratory failure. In addition to orthostatic hypotension, hypertensive episodes can occur and are usually paroxysmal but may occasionally be sustained [6]. Microneurographic evidence of sympathetic over-activity has been documented, which subsides with recovery from the neuropathy [7]. Sinus tachycardia is present in over 50% of patients with severe GBS. Less common autonomic symptoms include constipation, fecal incontinence, gastroparesis, ileus, erectile failure, and pupillary abnormalities. Gastrointestinal dysmotility is fairly common but rarely progresses to severe adynamic ileus. In a recent study, adynamic ileus occurred in 17 of 114 patients (15%) with severe GBS [8]. Cardiovascular dysautonomia coincided with ileus in only five patients. Ileus was thought to be related to mechanical

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00101-3

ventilation and immobilization and pre-existing conditions such as prior abdominal surgery or incremental doses of opioids. Related entities are acute motor axonal neuropathy (AMAN), acute motor and sensory axonal neuropathy (AMSAN), and the Miller–Fisher syndrome (characterized by the triad of ophthalmoplegia, ataxia, and areflexia). Another related entity is autoimmune autonomic ganglionopathy with sensory neuronopathy. These are best considered autoimmune neuropathies with different targets.

INVESTIGATIONS Tests of autonomic function are often abnormal. Orthostatic hypotension, impaired cardiovagal, sudomotor, and adrenergic vasomotor function are relatively common [6]. Abnormalities of cardiac rhythm include sinus tachycardia, bradyarrhythmias, heart block and asystole and may necessitate a cardiac pacemaker. Sinus arrest may occur following vigorous vagal stimulation, usually caused by tracheal suction.

ETIOLOGY OR MECHANISMS The disorder is likely to be immune-mediated although the antigen is unknown. The evidence is based on the sural nerve biopsy finding of perivascular round cell infiltration, the presence of relevant serum antibodies causing demyelination, frequent antecedent infections, the selectivity of involvement by fiber type, and response to immunotherapy and the overlap with cases of pandysautonomia. The autoimmune basis of GBS and its variants is slowly being unraveled. For instance, the Miller Fisher syndrome is known to have immunoglobulin G autoantibodies directed at the ganglioside GQ1b in more than 90% of patients [9]. Although GBS has been described after vaccination, vaccines developed subsequent to the swine influenza vaccine used in 1976–1977 have not been associated with an increased risk of GBS [10,11]. Indeed the risk of developing GBS following influenza significantly exceeds the risk of GBS following vaccination [10].

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COURSE AND PROGNOSIS Septic and autonomic complications are the major causes of death in GBS as respiratory failure is usually successfully managed with mechanical ventilation. Typically, autonomic neuropathy in GBS improves in concert with improvement in motor and sensory nerve function. Although functional recovery is typical, and autonomic recovery parallels somatic recovery, long-term autonomic sequelae are common; these include impairment of cardiovascular adrenergic function (including mild orthostatic hypotension, sudomotor, and cardiovagal impairment) [3,4,6,12]. Age at onset, need for mechanical ventilation, and duration of the plateau phase correlated with severity of neurological residua at follow-up [2].

MANAGEMENT Patients with autonomic instability require close monitoring of heart rate and BP. Paroxysmal hypertension may alternate with hypotension and patients may be supersensitive to pressor and depressor agents. For these patients, the use of hypotensive drugs is usually best avoided. When sustained hypertension develops, combined alpha and beta adrenergic blockade has been suggested and bradyarrythmias are probably best treated by the use of a demand pacemaker. The mainstay of treatment is supportive for the management of orthostatic hypotension and bowel and bladder symptoms. Since there is suggestive evidence of an immune-mediated process, treatment with intravenous immunoglobulin or plasma exchange should be undertaken in patients who have moderate to severe

weakness, respiratory compromise, or continue to worsen. Intravenous immunoglobulin is given as 0.4 g/kg (over 4 hours) daily for 5 days, but it has little if any effect if administered later than 14 days after onset of the illness [6].

References [1] Koga M, Ang CW, Yuki N, Jacobs BC, Herbrink P, van der Meche FG, et al. Comparative study of preceding Campylobacter jejuni infection in Guillain–Barre syndrome in Japan and The Netherlands. J. Neurol. Neurosurg. Psychiatry 2001;70:693–5. [2] Koeppen S, Kraywinkel K, Wessendorf TE, Ehrenfeld CE, Schürks M, Diener HC, et al. Long-term outcome of Guillain–Barré syndrome. Neurocrit Care 2006;5:235–42. [3] Flachenecker P, Hartung HP, Reiners K. Power spectrum analysis of heart rate variability in Guillain–Barré syndrome. A longitudinal study. Brain 1997;120:1885–94. [4] Flachenecker P, Wermuth P, Hartung HP, Reiners K. Quantitative assessment of cardiovascular autonomic function in Guillain–Barre syndrome. Ann. Neurol 1997;42:171–9. [5] Zochodne DW. Autonomic involvement in Guillain–Barre syndrome: a review. Muscle Nerve 1994;17:1145–55. [6] Low PA, Sandroni P. Autonomic neuropathies. In: Low PA, Benarroch EE, editors. Clinical Autonomic Disorders. Philadelphia: Lippincott Williams & Wilkins; 2008. p. 400–22. [7] Fagius J, Wallin BG. Sympathetic reflex latencies and conduction velocities in patients with polyneuropathy. J. Neurol. Sci 1980;47:449–61. [8] Burns TM, Lawn NC, Low PA, Camilleri M, Wijdicks EFM. Adynamic ileus in severe Guillain–Barre syndrome. Muscle Nerve 2001;24:963–5. [9] Yuki N, Sato S, Tsuji S, Ohsawa T, Miyatake T. Frequent presence of anti-GQ1b antibody in Fisher's syndrome. Neurology 1993;43:414–7. [10] Price LC. Should I have an H1N1 flu vaccination after Guillain– Barre syndrome? BMJ 2009;339:b3577. [11] Haber P, Sejvar J, Mikaeloff Y, DeStefano F. Vaccines and Guillain– Barré syndrome. Drug Saf 2009;32:309–23. [12] Lyu RK, Tang LM, Hsu WC, Chen ST, Chang HS, Wu YR. A longitudinal cardiovascular autonomic function study in mild Guillain– Barré syndrome. Eur. Neurol 2002;47:79–84.

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102 Hereditary Autonomic Neuropathies Yadollah Harati, Shahram Izadyar There are a few inherited peripheral neuropathies in which autonomic dysfunction, whether clinical or subclinical, is detected (Box 102.1). Autonomic abnormalities are frequent and prominent in familial dysautonomia and amyloidosis (see separate reviews in Chapters 103 and 99). This chapter discusses the remaining inherited neuropathies with autonomic involvement.

with the onset of disease manifestation in childhood or adolescence. Variants and milder forms of the disease in which residual enzyme activity may be detected have been reported. The gene for the enzyme is located on the long arm of chromosome X, in position Xq22. Currently more than 580 different mutations in this gene have been reported.

Clinical Manifestations of Fabry’s Disease FABRY’S DISEASE Fabry’s disease or Anderson–Fabry’s disease (angiokeratoma corporis diffusum) is an X-linked inherited slowly progressive metabolic disorder with protean and nonspecific clinical manifestations. Although the skin, kidney, heart, and peripheral and central nervous system are the most frequently involved organs, autonomic dysfunction also may be present. The clinical manifestations of the disease in the affected hemizygous male individuals result from progressive and widespread accumulation of neutral glycosphingolipids, particularly globotriaosylceramide (Gb3), caused by α-galactosidase A deficiency, in the lysosomes of vascular endothelial cells, smooth muscle, skin, cornea, neural cells, perineural cells of the autonomic nervous system, ganglia, and body fluids. Heterozygous female carriers are usually asymptomatic; 15%, however, have severe involvement of one or more organs, usually a decade later than males. The classically affected male individuals have no detectable α-galactosidase A activity

Unlike many other lysosomal storage diseases, most patients remain clinically silent during the first few years of life. The most prominent and frequent clinical presentation of Fabry’s disease includes bouts of severe painful burning sensation in the hands and feet; chronic pain characterized by burning and tingling paresthesia; a reddish purple maculopapular rash (angiokeratoma) of lower abdomen, pelvic, genital, upper thigh regions and, at times, in oral mucosa and conjunctive; hypohydrosis; heat intolerance; and lenticular and corneal opacities. Any boy or young man with severe painful sensory neuropathy should be suspected as having Fabry’s disease, and careful scrutiny for the skin lesions should be conducted because typical skin lesions are usually sparse and may be easily overlooked. Triggering factors for the episodic pain include fatigue, exercise, fever, emotional stress, and rapid changes in temperature and humidity. The pathophysiologic events leading to the incapacitating episodes of pain or acroparesthesias have not been clarified.

BOX 102.1

I N H E R I T E D P E R I P H E R A L N E U R O PAT H I E S I N W H I C H AU T O N O M I C DYSFUNCTION IS DETECTED 1. Hereditary sensory and autonomic neuropathies types I, II, III* (see Chapter 103), IV and V. 2. Hereditary motor and sensory neuropathy types I and II (Charcot–Marie–Tooth disease 1, 2). 3. Fabry’s disease.*

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4. Multiple endocrine neoplasia type 2B. 5. Amyloidosis* (see Chapter 99). 6. Porphyrias.* *Dysautonomia is prominent and clinically significant.

© 2012 Elsevier Inc. All rights reserved.

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Early albuminuria, uremia, renal failure, cardiomyopathy, cardiac hypertrophy, conduction abnormalities, aortic degeneration, hypertension, mitral valve thickening, atrioventricular block and supraventricular arrhythmias may occur. Short P-R interval, ST-T changes, and left ventricular hypertrophy are common electrocardiogram abnormalities of Fabry’s disease. Although cardiac involvement is a constant feature of Fabry’s disease, most patients do not experience cardiac symptoms until late in the disease course. Cerebrovascular disease secondary to multifocal abnormalities of large and small vessels, including transient ischemic attacks and stroke, may also occur. In young male patients with unexplained cardiovascular abnormalities and a history of acroparesthesia, angiokeratoma, and ophthalmologic findings, the diagnosis of Fabry’s disease should always be considered. Ocular manifestations involving the cornea, lens, conjunctiva, and retina are early and prominent, but require slit lamp microscopic evaluation. Involvement of many other tissues and organs results in a variety of symptoms and signs including gastrointestinal (episodic diarrhea, abdominal cramp, and achalasia), musculoskeletal (bony deformities), endocrine (hypothyroidism, osteopenia), hematopoietic (anemia, foamy macrophages and iron deficiency), pulmonary, vestibular and auditory symptoms and signs.

Autonomic Involvement Although there are many reports of structural abnormalities of the autonomic nervous system in Fabry’s disease, overt clinical autonomic dysfunction is not commonly observed. The symptoms that have been attributed to autonomic neuropathy in Fabry’s disease include anhidrosis or hypohydrosis, decreased tear and saliva formation, disturbances in cardiac rhythm, gastrointestinal dysmotility and abnormal cerebrovascular reactivity. Unlike other diseases with autonomic neuropathy, orthostatic intolerance and male sexual dysfunction are less frequent and less severe in patients with Fabry’s disease. Autonomic symptoms and possibly the episodic pain are probably caused by involvement of sympathetic ganglion cells and degeneration of unmyelinated nerve fibers. Both glycosphingolipid accumulation and vascular ischemia appear to play a role in the abnormalities of autonomic ganglia. There is a preferential loss of small myelinated and unmyelinated fibers in the sural nerve biopsies. Recently, it has been suggested that anhidrosis or hypohydrosis may be related to sweat gland dysfunction rather than autonomic neuropathy. Detailed clinical autonomic testing has shown involvement of both sympathetic and parasympathetic systems, the latter being more readily demonstrated. Sympathetic dysfunction is evident by the loss of the cutaneous flare response to scratch and histamine and the absence of thermal fingertip wrinkling. Parasympathetic dysfunction is evident by decreased tear

and saliva production, impaired pupillary response to pilocarpine and abnormal gastrointestinal function. Blood pressure, heart rate, and plasma norepinephrine responses to tilt are usually intact. Accumulation of Gb3 in the myenteric nerve plexuses throughout the gut results in disturbances in peristalsis and gastrocolic reflexes in more than 60% of hemizygous patients. Intestinal dysmotility may produce areas of high intraluminal pressure, allowing the formation of diverticula. Intestinal stasis and bacterial overgrowth will result in diarrhea. Diagnosis in male patients can be established by measuring α-galactosidase A activity in plasma or leukocytes. On the other hand, the enzyme activity level may fall within the normal range in affected females; therefore, their status should be determined by genotyping. Almost a decade ago, enzyme replacement therapy (ERT) using recombinant α-galactosidase A was introduced as the main disease-specific treatment for Fabry’s disease. Intravenous agalsidase beta (Fabrazyme) was approved by FDA in 2003 and should be initiated soon after diagnosis in males over 16 years of age. In younger males, it is recommended to initiate the therapy at the time of development of significant symptoms, or at age 7–10 years if asymptomatic. Females are recommended to start the treatment at the time of symptoms or evidence of progression of organ involvement. In clinical trials, ERT has been associated with clearance of microvascular endothelial deposits of glycosphingolipid from the kidneys, heart, and skin, reversing the pathogenesis of the chief clinical manifestations of this disease. There is evidence that sustained ERT is effective in improving peripheral nerve and sweat function, reducing pain and improving quality of life in patients with Fabry’s neuropathy. However, it is still unknown whether agalsidase beta therapy can reduce or prevent the cerebrovascular complications of this disease. It has been reported that carbamazepine used in the treatment of episodic pain may result in a dose-dependent aggravation of autonomic dysfunction including urinary retention, nausea, vomiting, and ileus. In patients that orthostatic hypotension and syncopal events are clinically significant, fludrocortisone acetate therapy can result in a reduction in the frequency of syncopal events.

PORPHYRIA Acute hepatic porphyrias (acute intermittent porphyria, variegate porphyria, and hereditary coproporphyria) are a group of autosomal dominant inherited metabolic disorders that manifest as acute or subacute, severe, lifethreatening motor neuropathy, abdominal pain, autonomic dysfunction, and neuropsychiatric manifestations. Its gene is thought to be present in 1/80,000 of people, although only one-third of affected persons ever manifest symptoms of the disease. The basic defect is a 50% reduction in hydroxymethylbilane synthase, also known as porphobilinogen deaminase activity (acute intermittent

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mulTIPlE ENdoCRINE NEoPlAsIA TyPE 2B

porphyria), protoporphyrinogen IX oxidase (variegate porphyria), and coproporphyrinogen oxidase (coproporphyria), resulting in abnormalities of heme synthesis. In the presence of sufficient endogenous or exogenous stimuli (e.g., drugs, hormones, menstruation, and starvation), this partial deficiency may lead to clinical manifestations.

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The neurologic manifestations of all forms of the acute porphyrias are identical. Symptoms of the acute attack include severe abdominal pain, nausea, vomiting, constipation, diarrhea, urinary frequency and hesitancy, urine discoloration, labile hypertension, tachycardia, excessive sweating, pain in limbs and back, and convulsions. Abdominal pain and ileus may occur several days before neurologic manifestations. In 20–30% of patients, signs of mental disturbance such as anxiety or confusion are present. Porphyria affects predominantly motor nerves, often leading to proximal, facial, and bulbar weakness. It usually develops within 2 to 3 days of the onset of abdominal pain and psychiatric symptoms and may begin in the upper limbs with wrist and finger extensor weakness or with cranial nerves dysfunction. The progression of weakness to trunk and respiratory muscles may resemble Guillain–Barré syndrome, but the ascending pattern of weakness is rare, cerebrospinal fluid is usually normal, and, in some patients, the reflexes remain intact. The nerve conduction velocities in porphyria are not slowed to the levels observed in demyelinating lesions. The mechanism of the neuropathic changes is poorly understood. Two hypotheses – the possible neurotoxicity of heme pathway intermediates such as delta-aminolevulinic and porphyrins, and heme deficiency in nervous tissue – have been suggested.

tests. Early parasympathetic dysfunction is detected during remission and in late asymptomatic patients. The immediate response of heart rate to standing (30:15 ratio) or to the Valsalva maneuver may be mildly abnormal in asymptomatic subjects with acute intermittent porphyria, suggesting the occurrence of a subclinical autonomic neuropathy in latent porphyrias. Gastrointestinal disturbances including abdominal pain, severe vomiting, obstinate constipation, intestinal dilation, and stasis may be explained by the impaired gut motor activity caused by autonomic or enteric nerved damage, or both. Studies on impaired proximal gastrointestinal tract motility and reduced circulating gut peptides in a few patients with acute intermittent porphyria tend to support this hypothesis. Other autonomic dysfunctions observed in acute intermittent porphyrias include sweating disturbances, pupillary dilation, and hesitancy in micturition and bladder distension. Limited pathologic studies of the autonomic nervous system in acute intermittent porphyrias have revealed lesions of vagus nerve including axonal degeneration and demyelinations, and chromatolysis of dorsal nuclei and sympathetic chain, as well as the splanchnic motor cells of the lateral horns and cells of the celiac ganglion. The sympathetic chain ganglia show a decrease of 50% in the density of ganglion cells and myelinated axons compared with control subject. The Watson–Schwartz test is used as a screening test for the detection of porphobilinogen (PBG) in urine during the attack. The laboratory diagnosis of porphyria depends on the measurement of porphyrin precursors in urine. To evaluate the porphyria type, measurement of porphyrins in both urine and feces is required. Enzyme measurements are used to identify asymptomatic family members whose quantitative excretions of porphyrins are normal.

Autonomic Involvement in Porphyria

Treatment of Porphyria

Autonomic disturbances in sympathetic and parasympathetic systems are prevalent immediately before and during the attacks of porphyrias, suggesting a greater and earlier susceptibility of the autonomic nerves. Persistent sinus tachycardia invariably precedes the development of peripheral neuropathy and respiratory paralysis and, together with labile hypertension, may be explained by damage to vagus or glossopharyngeal nerves, their nuclei, or central connections. Tachycardia and hypertension may be associated with increased catecholamine release and urinary excretion, suggesting increased peripheral sympathetic activity. Patients may have chronic hypertensions between the attacks leading to renal function impairment. Orthostatic hypotension may occur in acute attacks of intermittent or variegate porphyria. Use of a battery of baroreflex tests during the acute attack reveals mostly reversible parasympathetic or sympathetic dysfunction. The parasympathetic tests, however, become abnormal earlier and more frequently than sympathetic

The most effective treatment of porphyria attacks is the administration of hematin intravenously. With modern intensive care techniques and the advent of hematin therapy, the mortality rate for acute intermittent porphyrias is less than 10%. Porphyrinogenic drugs avoidance, high glucose intake, vitamin B6, β-blockers, analgesics, selective anticonvulsants, and hematin therapy (2–5 mg/kg per day intravenously for 3–14 days) are the mainstays of therapy. Phenlyephrine and phentolamine are reported to restore normal blood pressure. Recovery from psychiatric and autonomic dysfunction is usually rapid. All at-risk relatives should be screened for the latent disease.

Clinical Manifestations of Porphyria

MULTIPLE ENDOCRINE NEOPLASIA TYPE 2B Multiple endocrine neoplasia type 2 (MEN 2) syndromes are neural crest disorders. The MEN 2 syndromes

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comprise clinically related autosomal dominant cancer syndromes. MEN 2A (Sipple syndrome) is characterized by medullary thyroid carcinoma (MTC), pheochromocytoma in about 50% of cases, and parathyroid hyperplasia or adenoma in about 25% of cases. MEN 2B is similar to MEN 2A but is characterized by earlier age of tumor onset and the developmental abnormalities, which included intestinal ganglioneuromatosis, atypical facies with mucosal neuromas of distal tongue and subconjunctive, marfanoid body habitus, muscle underdevelopment, and bony deformities. Autonomic manifestations of MEN 2B are not prominent and are generally overshadowed by its other symptoms and signs. They include impaired lacrimation, orthostatic hypotension, impaired reflex vasodilation of skin and parasympathetic denervation supersensitivity of pupils, with intact sweating and salivary gland function. There are gross and microscopic abnormalities of the peripheral autonomic nervous system with both sympathetic and parasympathetic systems affected. There is disorganized hypertrophy and proliferation of autonomic nerves and ganglia (ganglioneuromatosis). Neuronal proliferation of the alimentary tract (Auerbach and Meissner’s plexus), upper respiratory tract, bladder, prostate, and skin may also be seen. Nerve biopsy shows degeneration and regeneration of unmyelinated fibers. Genetic linkage studies of MEN have mapped the gene responsible for this syndrome to the pericentromeric region of chromosome 10. This region also contains the RET (REarranged during Transfection) proto-oncogene, which codes for a receptor tyrosine kinase. Different missense mutations within RET proto-oncogene is thought to be responsible for MEN 2A and 2B. Molecular genetic testing of the RET gene is now available and confirms the diagnosis. Biochemical screening for neoplasias in patients with MEN 2A and 2B can be performed by measurement of serum calcitonin level at baseline or after pentagastrin stimulation test for MTC; 24-hour urinary excretion of catecholamines and metanephrines for pheochromocytoma; and serum calcium and parathyroid hormone levels for parathyroid hyperplasia. The prognosis in the MEN 2B syndrome is generally poor as a result of an aggressive medullary thyroid carcinoma that develops earlier in life for patients with MEN 2B than those with MEN 2A (5-year survival rate of 78% for MEN 2B and 86% for MEN 2A). This can be improved by regular screening of patients at risk; early diagnosis allows thyroidectomy, adrenalectomy, or both, which are likely to be curative.

HEREDITARY MOTOR AND SENSORY NEUROPATHIES TYPE I AND II (CHARCOT–MARIE–TOOTH 1 AND 2) Clinically significant autonomic dysfunction is not a common feature of Charcot–Marie–Tooth disease types 1 and 2 (CMT 1 and 2). When a battery of autonomic

function tests are systematically given to patients with well-established CMT abnormalities of sudomotor and local vasomotor responses, heart rate and blood pressure changes, pupillary abnormalities, impairment of sweating, and impaired tear production may be observed, suggesting involvement of postganglionic sympathetic and parasympathetic nerve fibers. Sural nerve biopsies may show abnormalities of unmyelinated nerve fibers, which explain the frequently observed abnormalities of sweat function tests. Pupillary abnormalities are secondary to a parasympathetic denervation of the iris sphincter and ciliary muscle, as shown by a positive methacholic test. Several patients with myotonic pupils received symptomatic relief from 0.025% pilocarpine. A particular point mutation in myelin protein zero (MPZ) in some families with CMT has been found to be associated with severe dysautonomia including bladder dysfunction and hypotension.

TYPE I, II, IV, AND V HEREDITARY SENSORY AND AUTONOMIC NEUROPATHY Type I hereditary sensory and autonomic neuropathy (HSAN) with an autosomal dominant gene on chromosome 9q22 and type II with a probable autosomal recessive inheritance pattern exhibit no significant autonomic dysfunction except for hypohydrosis. Types III and IV are autosomal recessive. Type III has preserved and, at times, excessive sweating. Type IV shares some of the features of types II and III, but the patients may have episodes of fever, low IQ, severe hypohydrosis with abnormal or absent sympathetic skin response test, and markedly reduced pain perception. There is a marked loss of small myelinated and unmyelinated nerve fibers. Type V presents with selective loss of extremity pain perception and thermal discrimination and impaired sudomotor function. Sural nerve biopsy reveals selective loss of small myelinated fibers with only a slightly decreased number of larger myelinated fibers.

Further Reading Albers JW, Fink JK. Porphyric neuropathy. Muscle Nerve 2004;30(4):410–22. Biegstraaten M, et al. Autonomic neuropathy in Fabry disease: a prospective study using the Autonomic Symptom Profile and cardiovascular autonomic function tests. BMC Neurol 2010;10:38. Eng CM, et al. Fabry disease: guidelines for the evaluation and management of multi-organ system involvement. Genet Med 2006;8(9):539–48. Germain DP. Fabry disease. Orphanet J Rare Dis 2010;5(1):30. Puy H, Gouya L, Deybach JC. Porphyrias. Lancet 2010;375(9718):924–37. Schiffmann R, et al. Enzyme replacement therapy improves peripheral nerve and sweat function in Fabry disease. Muscle Nerve 2003;28(6):703–10. Stojkovic T, et al. Autonomic and respiratory dysfunction in Charcot– Marie–Tooth disease due to Thr124Met mutation in the myelin protein zero gene. Clin Neurophysiol 2003;114(9):1609–14. Wohllk N, et al. Multiple endocrine neoplasia type 2. Best Pract Res Clin Endocrinol Metab 2010;24(3):371–87.

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103 Familial Dysautonomia (Riley–Day Syndrome) Horacio Kaufmann, Lucy Norcliffe-Kaufmann, Felicia B. Axelrod INTRODUCTION Familial dysautonomia (FD), also known as Riley–Day syndrome or hereditary sensory and autonomic neuropathy type III, is an autosomal recessive disease caused by mutations in the gene that encodes for I-κ-B kinase complex associated protein (IKAP) [1]. FD was first described by Riley and collaborators in 1949 [2]. Affected patients have a complex neurological phenotype. First, because of a congenital abnormality in the afferent baroreflex pathways, their blood pressure is extremely labile with severe episodic hypertension and orthostatic hypotension [3]. In addition to the autonomic cardiovascular abnormalities, patients with FD also have decreased pain and temperature perception, impaired sense of taste, abnormal swallowing, gait ataxia, decreased/absent myotatic reflexes and decreased ventilatory responses to hypoxia and hypercapnia [4]. There is no cure for FD. Life expectancy is reduced and treatments are supportive.

CLINICAL FEATURES Typically, affected patients present at birth with hypotonia, episodic skin blotching and irritability. Infants do not respond to pain or cold stimuli. Oral incoordination and abnormal swallowing reflexes lead to feeding difficulties and recurrent bouts of aspiration pneumonia. Temperature instability, particularly hypothermia and excessive perspiration, are distinctive features [5]. There is a characteristic muted response to intradermal histamine injection with an absent axon flare. Other early diagnostic clues include the lack of fungiform papillae and taste buds that gives the tongue a peculiar smooth appearance [6] and absent overflow tears [2]. Developmental milestones are usually delayed throughout childhood. Emotional lability with severe anxiety and behavioral problems including breath-holding spells are common. Learning difficulties are not infrequent [4]. Blood pressure instability occurs in all patients from birth. Cortical arousal results in hypertension and tachycardia. Conversely, when the patient is calm, hypotension

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and bradycardia occur with standing and physical exercise. A distinctive feature of FD is the severely disabling retching and vomiting attacks accompanied by marked hypertension, tachycardia, diaphoresis and blotching of the skin. These hyperadrenergic symptoms, frequently referred as dysautonomic crises, can occur in response to emotions, illness or on awakening [3]. Spinal deformities (scoliosis and kyphosis) are commonly present in adolescence and by adulthood stature is usually small. Frequently, spinal deformities and recurrent aspiration pneumonias result in severe chronic lung disease with hypoxia and hypercapnia. Charcot (neuropathic) joints can also develop as a consequence of pain insensitivity and impaired proprioception. Corneal analgesia can lead to abrasions, ulceration and scarring [4]. Gastrointestinal bleeds are a frequent cause for hospital admissions [7]. Many patients with FD have sleepdisordered breathing with obstructive and central sleep apneas. With advancing age, optic atrophy frequently leads to blindness and gait ataxia worsens significantly. The longterm consequences of labile hypertension include chronic kidney disease, end-stage renal disease and left ventricular hypertrophy. Respiratory and cardiovascular complications remain the leading cause of death.

GENETICS Over 99% of FD cases are homozygous for a single point mutation in the IKBKAP gene, located on the long arm of chromosome 9q. The gene encodes for the I-κ-B kinase complex associated protein (IKAP or Elp1) [1]. IKAP is a highly conserved protein found in all eukaryotic cells. It is believed to play a role in stem cell migration, neuronal development and myelination during embryogenesis, as well as being a regulator for gene transcription and elongation. Complete lack of the functional IKAP protein is fatal [8]. The most common FD mutation occurs at the beginning of intron 20. As a consequence, during splicing, a cellular process in which the non-encoding introns are removed and the encoding exons are joined together, the pre-mRNA sequence is often misread. The splicing mistake effectively

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BOX 103.1

N E U R O L O G I C A L A N D M U S C O L O S K E L E TA L F E AT U R E S I N F A M I L I A L D Y S AU T O N O M I A

Somatic Sensory and Motor Systems l l l l

Impaired pain and temperature perception Decreased/absent deep tendon reflexes Hypotonia Gait ataxia.

Cranial Nerves l l l l l

Autonomic System l

l l

l l

l

l

l

Afferent baroreflex failure causing extremely labile blood pressure. Orthostatic and exercise-induced hypotension Episodic hyperadrenergic crises with retching and vomiting, cutaneous blotching, hypertension and tachycardia, and irritability Episodic hyperhidrosis Oropharyngeal esophageal and gastrointestinal dysmotility Impaired hypoxic and hypercapnic ventilatory drive (hypoventilation) Sleep disordered breathing (central and obstructive apneas).

l

Psychiatric/Cognitive l l l l

CARDIOVASCULAR AUTONOMIC ABNORMALTIES Patients with FD have blunted baroreceptor afferents and, as a consequence, their blood pressure is extremely labile. Gravitational stimuli, normally sensed by baroreceptors in the vasculature, fail to elicit appropriate

Mild to severe cognitive impairment Anxiety/phobias/obsessive traits Emotional lability Breath-holding episodes.

Musculoskeletal l l l l

misses out exon 20 from the mRNA sequence, which is then translated into a short unstable protein that is quickly degraded. The protein deficiency, however, does not occur to the same degree in all tissues. As the mutation appears in a non-coding sequence, some cells are capable of producing relatively normal amounts of the mRNA message and the functional IKAP molecule. Other cells, like neurons in the CNS produce mostly mutant mRNA and little protein [1,8]. The FD mutation is largely confined to the Ashkenazi (European) Jews. The carrier rate varies from 1 in 17 to 1 in 32, with an incidence of around 1 in 3000 live births. However, with the introduction of widespread population screening, prenatal testing and selective abortion this incidence has since declined. Diagnosis is now based on molecular confirmation.

No overflow tears Depressed corneal reflexes Optic nerve atrophy Frequent exotropia Diminished or absent sense of taste (absent fungiform papillae) Dysarthric nasal speech Sialorrhea.

Frequent spinal curvature Increased risk of Charcot (neuropathic) joints Frequent bone fractures Short stature and decreased muscle mass.

adjustments of heart rate and vascular resistance resulting in supine hypertension and orthostatic hypotension. Vasopressin release during hypotension, a response solely dependent on the afferent baroreceptor pathways, is absent in patients with FD [3]. Although reduced in number, efferent sympathetic nerves are functional. Supine plasma norepinephrine levels are normal in FD. Stimuli that activate efferent sympathetic neurons, independently of baroreceptor afferent pathways, such as cognitive tasks and emotional arousal, dramatically increase blood pressure, heart rate and circulating norepinephrine levels (Fig. 103.1) [3]. The reduced number of sympathetic neurons likely results in vascular “denervation supersensitivity”, further increasing the hypertensive responses to sympathetic surges. Conversely, sedation and sleep markedly reduces blood pressure and heart rate. Blood pressure variability throughout the day is dramatically increased. Ambulatory blood pressure recordings reveal hypertensive peaks occurring on awakening, while eating and at times of emotional arousal (including anxiety, excitement and mental tasks requiring concentration) [3]. Significant hypotension can occur when the patient is calm, during sleep, or while exercising. Hypertension and excessive blood pressure variability

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PATHology

FD patient A

FD patient B 150 HR (bpm)

HR (bpm)

150 100

50

Phone call

300

300

250

250 BP (mmHg)

BP (mmHg)

50

100

200 150 100

Patient upset

200 150 100

50

50 0

1

2

3 4 Time (min)

5

6

0

7

1

FD patient C

4

5

40 75 50 200

Study nurse enters room to draw blood

150

100

Δ BP(mmHg) / Δ HR(bpm)

HR (bpm)

3 Time (min)

Response to cognitive stimuli

100

BP (mmHg)

2

30

BP

20

10

BP

HR BP

50

0

1

2

3

0 FD

PAF

HR HR Normals

FIGURE 103.1 Blood pressure (BP) and heart rate (HR) during emotional arousal and cognitive stimuli. (A) Beat-to-beat BP and HR in a patient with familial dysautonomia (FD) who received an unexpected phone call from a friend. Her BP and HR rose in parallel. She was then tilted to the upright position (min 1). BP initially fell, but quickly rose again. (B) BP and HR in a boy with FD during an argument with his mother. (C) BP and HR in a patient with FD when the study nurse enters the room to draw blood. (D) Changes in BP and HR in patients with FD, pure autonomic failure (PAF), and normal controls induced by a cognitive stimulus (mental arithmetic). Reproduced from ref [3].

frequently results in chronic kidney disease and left ventricular hypertrophy. Many patients with FD suffer periodic hyperadrenergic crises (also referred to as dysautonomic crises) characterized by nausea, retching and vomiting as well as skin blotching, hypertension and tachycardia. These crises are due to sudden surges in sympathetic activity unrestrained because of impaired baroreceptor feedback (Fig. 103.2) [3].

PATHOLOGY Neuropathology studies of the disease are limited. Tissue samples from FD patients showed reduced number of primary sensory neurons in dorsal root ganglia [9] and

fiber loss in spinothalamic and spinocerebellar tracts as well as in posterior columns of the spinal cord [10]. These findings provided an explanation for the impaired pain and temperature perception as well as the characteristic decreased/absent myotatic reflexes and gait ataxia. Sympathetic ganglia at the cervical and thoracic levels are reportedly reduced in size and contain fewer cells and there are also fewer preganglionic neurons in the intermediolateral column [9]. It is likely, therefore, that patients with FD also have vascular “denervation supersensitivity”, further increasing the hypertensive responses to sympathetic surges when the remaining neurons are activated. Only a few neuropathological studies have included the brain and have shown abnormalities in the brainstem, particularly medullary regions [10].

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FIGURE 103.2 Blood pressure (BP), heart rate, and plasma catecholamine levels during typical crisis triggered by emotionally charged situations. Left bar charts show systolic BP, middle bar chart shows heart rate, and right bar chart shows plasma norepinephrine concentration at baseline (open squares) and during a typical crisis provoked by an emotionally charged situation (black squares). Reproduced from ref [3].

TREATMENT As a first line, feeding difficulties in infants are managed with thickening feeds to prevent aspiration. Severe cases may require fundoplication to prevent vomiting and gastrostomy liquid feeding into the stomach. Because of the indifference to pain and temperature, precautions should be taken to avoid injuries, such as corneal abrasions or burns. Pharmacologic treatment of the labile blood pressure in patients with FD is complex and frequently unsuccessful, because drugs that increase standing blood pressure worsen hypertension and drugs that lower blood pressure may worsen orthostatic hypotension. Acutely, the hyperadrenergic crises with retching and vomiting are usually treated with a combination of benzodiazepines and central sympatholytic agents like the alpha-2-agonist clonidine. These drugs, however, are not always effective and may leave the patient extremely sedated and severely hypotensive. Patient education about physical counter maneuvers, volume expansion with salt, adequate hydration and head-up sleeping are the cornerstone of treatment for orthostatic hypotension. The alpha-1-agonist midodrine may be required to prolong standing time or maintain blood pressure at times of physical activity. Antihypertensive medications are required in many patients. Because of the lack of hypoxic ventilatory drive, extreme care should be taken when using sedative medications and with exposure to ambient hypoxia, for example at high altitudes or on airline flights. Positive pressure ventilation is recommended for the treatment of sleep apnea and hypoventilation.

Recently, the plant hormone kinetin has been shown to correct the protein deficiency in FD-derived cell lines. Whether this potential genetic therapy has an impact of the natural history of the disease remains to be seen.

References [1] Slaugenhaupt SA, Blumenfeld A, Gill SP, et al. Tissue-specific expression of a splicing mutation in the IKBKAP gene causes familial dysautonomia. Am J Hum Genet 2001;68(3):598–605. [2] Riley CM, Day RA, Greeley DM, Landford WS. Central autonomic dysfunction with defective lacrimation: I. Report of five cases. Pediatrics 1949;3(4):468–78. [3] Norcliffe-Kaufmann L, Axelrod F, Kaufmann H. Afferent baroreflex failure in familial dysautonomia. Neurology 2010;75(21):1904–11. [4] Riley CM, Freedman AM, Langford WS. Further observations on familial dysautonomia. Pediatrics 1954;14(5):475–80. [5] Geltzer AI, Gluck L, Talner NS, Polesky HF. Familial dysautonomia; studies in a newborn infant. N Engl J Med 1964;271:436–40. [6] Smith A, Farbman A, Dancis J. Absence of taste-bud papillae in familial dysautonomia. Science 1964;147:1040–1. [7] Wan DW, Levy J, Ginsburg HB, Kaufmann H, Axelrod FB. Complicated peptic ulcer disease in three patients with familial dysautonomia. J Clin Gastroenterol. 2010. [8] Mezey E, Parmalee A, Szalayova I, Gill SP, Cuajungco MP, Leyne M, et al. Of splice and men: what does the distribution of IKAP mRNA in the rat tell us about the pathogenesis of familial dysautonomia? Brain Res 2003;983(1-2):209–14. [9] Pearson J, Pytel BA, Grover-Johnson N, Axelrod F, Dancis J. Quantitative studies of dorsal root ganglia and neuropathologic observations on spinal cords in familial dysautonomia. J Neurol Sci 1978;35(1):77–92. [10] Brown WJ, Beauchemin JA, Linde LM. A neuropathological study of familial dysautonomia (Riley–Day syndrome) in siblings. J Neurol Neurosurg Psychiatry 1964;27:131–9.

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104 Autonomic Disturbances in Spinal Cord Injuries Christopher J. Mathias, David A. Low Normal functioning of the autonomic nervous system is critically dependent on integrity of the spinal cord, as the entire sympathetic outflow (T1-L2/3) and the sacral parasympathetic outflow travel and synapse within the spinal cord, before supplying various target organs (Fig. 104.1). In spinal cord injuries, therefore, autonomic impairment usually occurs, and this depends upon the site and the extent of the lesion. In cervical and high thoracic transection, the entire or a large part of the sympathetic outflow, together with the sacral parasympathetic outflow, is separated from cerebral control. Autonomic malfunction may affect the cardiovascular, thermoregulatory, sudomotor, gastrointestinal, urinary and reproductive systems [1]. The problems are usually worse in those with higher lesions. Soon after cord injury, there is a transient state of hypoexcitability, described as “spinal shock” [2]. There is flaccid paralysis of muscles, lack of tendon reflexes, and impairment of spinal autonomic function with atony of the urinary bladder and large bowel, dilatation of blood vessels and lack of spinal autonomic reflexes. This may last from a few days to a few weeks, following which activity in the isolated cord returns. In the chronic phase, with return of isolated spinal function, a different set of autonomic abnormalities occurs.

CARDIOVASCULAR SYSTEM In recent high injuries, basal blood pressure, especially diastolic, is usually lower than normal. Plasma norepinephrine and epinephrine levels are low, as in the chronic phase. Basal heart rate is usually below normal. In patients with high cervical lesions, who need artificial ventilation because of diaphragmatic paralysis, severe bradycardia and cardiac arrest may occur during tracheal stimulation (Fig. 104.2). This results from increased vagal activity, as efferent muscarinic blockade with atropine prevents bradycardia. Vagal activity is increased by hypoxia and by the absence of sympathetic reflexes; furthermore, there is an inability to reduce vagal activity through the pulmonary inflation reflex because of the inability to breathe. It is necessary to prevent such episodes by adequate oxygenation,

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treatment of respiratory infection and pulmonary emboli (which contribute to hypoxia), avoidance of cholinomimetic agents such as neostigmine and carbachol, and, if necessary, the use of parenteral atropine or a demand cardiac pacemaker. In the chronic stage, the levels of basal systolic and diastolic blood pressure are related closely to the level of the spinal lesion, being lower in the high lesions, and rising towards normal as the lesion descends. In tetraplegic subjects, plasma norepinephrine levels are about 25% of the levels observed in normal subjects, and they have reduced basal muscle sympathetic nerve activity, as measured by microneurography. Complicating factors, such as renal damage and failure, can elevate blood pressure, regardless of the lesion. In high lesions the blood pressure is sensitive to a number of physiological stimuli. Postural (orthostatic) hypotension is a particular problem in the early stages (Fig. 104.3). Plasma norepinephrine levels are low and do not rise with head-up postural change, unlike normal subjects. There is a marked rise in levels of plasma renin, aldosterone, and vasopressin, which may contribute to the recovery of blood pressure and account for other symptoms, such as reduced urine output. Improvement in symptoms and in postural blood pressure follows repeated head-up tilting, which presumably improves cerebral autoregulation and releases the various hormones which constrict blood vessels, raise intravascular volume and thus reduce the postural blood pressure fall. Drugs such as the sympathomimetics ephedrine or midodrine may need to be used. The reverse, severe hypertension, may occur during autonomic dysreflexia following stimulation below the level of the lesion, predominantly, but not always, through noxious stimuli [3]. This may occur through the skin (such as from complicating pressure sores), from abdominal and pelvic viscera (by contraction of the urinary bladder or irritation from a urethral catheter) (Figs 104.4 and 104.5) or via skeletal muscles (during muscle spasms). The paroxysmal rise in blood pressure is the result of increased spinal sympathetic neural activity causing constriction of both resistance and capacitance vessels. These changes occur below the level of the lesion, while above the lesion

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FIGURE 104.1 Schematic outline of the major autonomic pathways controlling the circulation. The major afferent input into the central nervous system is through the glossopharyngeal (CR,9) and vagus (CR,10) nerves by activation of baroreceptors in the carotid sinus and aortic arch. Chemoreceptors and low pressure receptors also influence the efferent outflow. The latter consists of the cranial parasympathetic (PS) outflow to the heart via the vagus nerves, and the sympathetic outflow from the thoracic and upper lumbar segments of the spinal cord. Activation of visceral, skin, and muscle receptors, in addition to cerebral stimulation influences the efferent outflow. In high spinal cord lesions, therefore, the input from chemoreceptors and baroreceptors is preserved along with the vagal efferent outflow, but there is no connection between the brain and the rest of the sympathetic outflow. The spinal sympathetic outflow may be activated through a range of afferents (visual, skin, muscle). This occurs through isolated spinal cord reflexes, not controlled by cerebral pathways, as seen normally.

0.6 mg IV

6 hr post atropine (0.6 mg IV)

FIGURE 104.3 (a) Blood pressure (BP) and heart rate (HR) in a tet20 min post atropine (0.6 mg IV)

FIGURE 104.2 (a) The effect of disconnecting the respirator (as required for aspirating the airways) on the blood pressure (BP) and heart rate (HR) of a recently injured tetraplegic patient (C4/5 lesion) in spinal shock, 6 hours after the last dose of intravenous atropine. Sinus bradycardia and cardiac arrest, also observed in the electrocardiograph, were reversed by reconnection, intravenous atropine and external cardiac massage (from Frankel et al. Lancet 1975;ii:1183–1185). (b) The effect of tracheal suction, 20 minutes after atropine in the same patient. Disconnection from the respirator and tracheal suction did not lower either heart rate or blood pressure. (from Mathias Eur J Int Care Med 1976;2:147–156).

raplegic patient before and after head-up tilt, in the early stages of rehabilitation, where there were few muscle spasms and minimal autonomic dysreflexia (from Mathias and Frankel Handbook of Clinical Neurology 1992;17:435–56). (b) Blood pressure (BP) and heart rate (HR) in a chronic tetraplegic patient before, during and after head-up tilt to 45°. Blood pressure promptly falls but with partial recovery, which in this case is linked to skeletal muscle spasms (S) inducing spinal sympathetic activity. Some of the later oscillations may be due to the rise in plasma renin, which was measured where there were interruptions in the intra-arterial record. In the later phases of head-up tilt, skeletal muscle spasm occur more frequently and further elevate the blood pressure. On return to the horizontal, blood pressure rises rapidly above the previous level and then returns slowly to horizontal levels Heart rate usually moves in the opposite direction, except during muscle spasms when there is an initial increase. (from Mathias and Frankel Ann Rev Physiol 1999;50:577–592).

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FIGURE 104.4 Blood pressure (BP), heart rate (HR  intravesical pressure (IVP), plasma norepinephrine (NE) (open histograms) and plasma epinephrine (E) (filled histograms) in a tetraplegic patient before during and after urinary bladder stimulation induced by suprapubic percussion of the anterior abdominal wall. The rise in BP is accompanied by a fall in heart rate as a result of increased vagal activity in response to the elevated blood pressure. Plasma NE but not E levels rise, suggesting an increase in sympathetic neural activity, independently of adrenomedullary activation . (from Mathias and Frankel J Autonom Nerv Syst Supple 1986; 457–64).

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normal subjects; this differs markedly from the levels seen in hypertensive crises due to a pheochromocytoma. During autonomic dysreflexia muscle sympathetic nerve activity measured by sympathetic microneurography shows only a modest increase [4], suggesting that the pressor response may be due to alpha-adrenergic receptor sensitivity. This also may explain the increased pressor response to intravenously infused norepinephrine. Other factors, such as impaired baroreflex activity, may be of importance as there is pressor hypersensitivity to a wide range of vasoactive agents of different chemical structures which act on a variety of receptors. Tetraplegic subjects also have an enhanced depressor response to vasodilator agents which, furthermore, favors baroreflex impairment. The contribution of sympathetic nonadrenergic transmission to increases in vascular resistance during autonomic dysreflexia has also recently been proposed [5]. Experimental studies indicate the importance of certain neuronal cells with activated nerve growth factors (NGF); furthermore neutralizing intraspinal NGF prevents the development of autonomic dysreflexia [6,7]. Autonomic dysreflexia is a serious problem. It may result in considerable morbidity, with severe sweating and a throbbing headache, and even mortality as a result of intracranial hemorrhage. The management consists of preventing the initiating factor caused increased sympathoneuronal activity. If necessary, a variety of drugs to reduce sympathetic efferent activity can be used [8].

CUTANEOUS CIRCULATION

FIGURE 104.5 Changes in mean blood pressure (MBP) and heart rate (HR) in patients with spinal cord lesions at different levels (cervical and thoracic) after urinary bladder stimulation induced by suprapubic percussion of the anterior abdominal wall. In the cervical and high thoracic lesions, there is a marked elevation in blood pressure and a fall in heart rate. In patients with lesions below T5 there are minimal cardiovascular changes . (from Mathias and Frankel J Autonom Nerv Syst Supple 1986; 457–64).

there may be sweating and dilatation of cutaneous vessels over the face and neck. Autonomic dysreflexia is accompanied by increased levels of plasma norepinephrine. Plasma norepinephrine levels, even at the height of hypertension, however, are increased only 2- or 3-fold above the low basal levels, and are still within the range of basal levels in

In higher lesions, the skin below the lesion is usually warmer and veins appear dilated. There may be extravasation of fluid into subcutaneous tissue which could contribute to skin breakdown and pressure sores. Vasodilation often occurs in the nose (Guttmann’s sign); similar changes occur after alpha antagonist, reserpine and guanethidine in hypertensive patients. In spinal shock the cutaneous responses to the triple or Lewis response are exaggerated, hence, the term dermatographia rubra. With the return of spinal cord reflex activity, in the chronic phase, there is sympathetic vasoconstriction and skin pallor, hence the term dermatographia alba.

THERMOREGULATION AND SUDOMOTOR FUNCTION Hypothermia may readily occur in high lesions as shivering is diminished and they may be unable to vasoconstrict the cutaneous circulation. The reverse may occur causing hyperthermia because of the inability to sweat and to reflexly vasodilate in the periphery, so as to lose heat [9].

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Maintenance of environmental temperature therefore is of critical importance. With hyperthermia tepid sponging, increasing air flow with a fan to accelerate heat loss and in severe cases ice cooled saline by intravenous infusion or urinary bladder irrigation may be needed. The sympathetic skin response (SSR), a technique that records neurogenic activation of sweat glands, is abnormal in spinal injuries, depending on the level of lesion [10]. Activation of supraspinal centers and descending sudomotor neural pathways in the spinal cord are necessary for the SSR, which is absent in the plantar region in low injuries, and absent in the palmar region in high spinal injuries. Importantly, the presence or absence of the SSR can be a useful marker of spinal cord autonomic involvement, in addition to motor and sensory evaluation, and may improve classification of the extent of spinal functional deficits [11]. The studies also exclude spinal cord sudomotor centers, isolated from the brainstem, that are capable of generating an SSR [10].

GASTROINTESTINAL SYSTEM In the early stages of spinal cord injury, there is vagal hyperactivity which may contribute to acid hypersecretion, with gastric ulceration and hemorrhage. H2 receptor antagonists, or allied agents, need to be used prophylactically. In high lesions paralytic ileus may occur; the mechanisms are unclear and often follow ingestion of solid food, which should be avoided. Large bowel dysfunction is common and adequate training, together with the use of an appropriate diet, mild laxatives and stool softeners may be needed [12].

URINARY SYSTEM In the early stages bladder atony occurs, with urinary retention, bladder distention and urinary overflow. With recovery of isolated cord function, the bladder can be trained to be an automated reflex or neurogenic bladder. Catheters should ideally be used intermittently in the early stages. Urinary infection in skin, bone and other tissues may cause secondary amyloidosis, with renal infiltration and serious sequelae.

REPRODUCTIVE SYSTEM In the male, sexual function is affected, especially in the early stages, with both erectile and ejaculatory failure. In the chronic phase, priapism may occur during autonomic dysreflexia. Ejaculation, if it occurs, is often retrograde. Various approaches which include electrical stimulation and collection of seminal fluid have been used for artificial insemination [13]. The phosphodiesterase inhibitor sildenafil (Viagra) is an effectively used drug in spinal injuries; whether it also lowers blood pressure excessively, as it

does in other groups with autonomic failure, such as multiple system atrophy, is not known [14]. In women, menstrual cycle disruption often occurs in the early stages. There is usually recovery within a year, and successful pregnancy has occurred in both tetraplegics and paraplegics. In high lesions, severe autonomic dysreflexia may accompany uterine contractions. Such patients are particularly prone, with the elevation of blood pressure, to epileptic seizures and cerebral hemorrhage. It is essential to lower their blood pressure. A combination of anticonvulsant (such as phenytoin) and agents to reduce spinal cord activity (such as spinal anesthetics) may be needed along with other agents to control blood pressure. Recent developments in clinical practice and scientific research have led to interventions that may facilitate recovery from spinal cord injury [15]. Functional recovery from spinal cord injury could be achieved by interventions that re-innervate disconnected systems or promote the natural plasticity of the central nervous system to facilitate the actions of surviving neurons that have retained axonal connections with their targets [16]. Interventions to produce functional improvements in spinal cord injury include repetitive transcranial magnetic stimulation to the motor cortex [17], tele-rehabilitation using functional electrical stimulation together with exercise and weightassisted treadmill walking therapy [18,19], which result in improved functional outcome of upper limb function and changes to sensorimotor systems.

References [1] Alexander MS, et al. International standards to document remaining autonomic function after spinal cord injury. Spinal Cord 2009;47:36–43. [2] Ditunno JF, Little JW, Tessler A, Burns AS. Spinal shock revisited: a four-phase model. Spinal Cord 2004;42(7):383–95. [3] Burton AR, Brown R, Macefield VG. Selective activation of muscle and skin nociceptors does not trigger exaggerated sympathetic responses in spinal-injured subjects. Spinal Cord 2008;46(10):660–5. [4] Stjernberg L, Blumberg H, Wallin BG. Sympathetic activity in man after spinal cord injury. Outflow to muscle below the lesion. Brain 1986;109(4):695–715. [5] Groothuis JT, Rongen GA, Deinum J, Pickkers P, Danser AH, Geurts AC, et al. Sympathetic nonadrenergic transmission contributes to autonomic dysreflexia in spinal cord-injured individuals. Hypertension 2010;55(3):636–43. [6] Krenz NR, Meaking SO, Krassioukov AV, Weaver LC. Neutralizing intraspinal nerve growth factor blocks autonomic dysreflexia caused by spinal cord injury. J Neurosci 1999;19:7405–14. [7] Krenz NR, Weaver LC. Nerve growth fact in glia and inflammatory cells of the injured rat spinal cord. J Neurochem 2000;74:730–9. [8] Krassioukov A, Warburton DE, Teasell R, Eng JJ. A systematic review of the management of autonomic dysreflexia after spinal cord injury. Arch Phys Med Rehabil 2009;90(4):682–95. [9] Price MJ. Thermoregulation during exercise in individuals with spinal cord injuries. Sports Med 2006;36(10):863–79. [10] Cariga P, Catley M, Savic G, Frankel HL, Mathias CJ, Ellaway PH. Organisation of the sympathetic skin response in spinal cord injury. J Neurol Neurosurg Psychiatry 2002;72:356–60. [11] Nicotra A, Catley M, Ellaway PH, Mathias CJ. The ability of physiological stimuli to generate the sympathetic skin response

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[12]

[13]

[14]

[15]

[16]

in human chronic spinal cord injury. Restor Neurol Neurosci 2005;23(5-6):331–9. Chung EA, Emmanuel AV. Gastrointestinal symptoms related to autonomic dysfunction following spinal cord injury. Prog Brain Res 2006;152:317–33. Consortium for Spinal Cord Medicine Sexuality and reproductive health in adults with spinal cord injury: a clinical practice guideline for health-care professionals. J Spinal Cord Med 2010;33(3):281–336. Hussain IF, Brady C, Swinn MJ, Mathias CJ, Fowler C. Treatment of erectile dysfunction with sildenafil citrate (Viagra) in parkinsonism due to Parkinson’s disease or multiple system atrophy with observations on orthostatic hypotension. J Neurol Neurosurg Psychiatry 2001;71:371–4. Ellaway PH, Anand P, Bergstrom EM, Catley M, Davey NJ, Frankel HL, et al. Towards improved clinical and physiological assessments of recovery in spinal cord injury: a clinical initiative. Spinal Cord 2004;42(6):325–37. Ellaway PH, Kuppuswamy A, Balasubramaniam AV, et al. Development of quantitative and sensitive assessments of physiological and functional outcome during recovery from spinal cord injury: A Clinical Initiative. Brain Res Bull 2011;84(4–5):343–57.

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[17] Belci M, Catley M, Husain M, Frankel HL, Davey NJ. Magnetic brain stimulation can improve clinical outcome in incomplete spinal cord injured patients. Spinal Cord. 2004;4:417–9. [18] Popovic MR, Thrasher TA, Adams ME, Takes V, Zivanovic V, Tonack MI. Functional electrical therapy: retraining grasping in spinal cord injury. Spinal Cord. 2006;44:143–51. [19] Dobkin B, Barbeau H, Deforge D, Ditunno J, et al. The evolution of walking related outcomes over the first 12 weeks of rehabilitation for incomplete traumatic spinal cord injury: the multicenter randomized spinal cord injury trial. Neurorehabil. Neural Rep 2007;21:25–35.

Further Reading Mathias C, Frankel HL. Autonomic disturbances in spinal cord lesions. In: Bannister R, Mathias. CJ, editors. Autonomic Failure: A Textbook of Clinical Disorders of the Autonomic Nervous System (4th Edition). Oxford: Oxford University Press; 1999. p. 494–513.

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105 Drug-Induced Autonomic Dysfunction James M. Luther INTRODUCTION Adverse effects of prescription, over-the-counter, and herbal medications contribute to significant morbidity and mortality in the United States [1]. Medications frequently alter blood pressure regulation, which may interfere with daily activities or contribute to cardiovascular complications. Although hypotension is frequently accompanied by symptoms, hypertension may go unrecognized until serious complications develop, prompting medical attention. This chapter provides examples of some offending medications, with extra attention given to non-antihypertensive agents. Further examples may also be found in other Primer chapters and in the literature [2]. It should be obvious from these examples that a thorough medication history is needed, or an opportunity to prevent complications could be missed.

DRUG-INDUCED HYPERTENSION Although an identifiable secondary cause is present in ~1–5% in the hypertensive population, drug-induced hypertension is often unrecognized or unreported. Physicians are more likely to recognize medications which increase blood pressure due to on-target effects (e.g., phenylephrine) than off-target effects. Patients frequently omit over-the-counter and herbal medications from their medication lists (e.g., NSAIDs, decongestants, appetite suppressants, nutritional supplements), whether intentionally or not. Patients often overlook vasoactive drugs in cold-and-sinus preparations or other combination preparations. A history of illicit drug or alcohol use may be withheld due to embarrassment or for legal reasons. Therefore, physicians should inquire about these possibilities in the absence of family members, after a solid patient–physician relationship has been established. Patients rarely report recent cessation or intermittent use of a drug unless specifically asked, which becomes relevant due to withdrawal-related hypertension. Withdrawal from ethanol, opioids, benzodiazepines, or other drugs of abuse produces hyperadrenergic symptoms including tachycardia, sweating, and hypertension. Excessive alcohol intake is associated with hypertension in epidemiologic

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studies, although episodic alcohol-withdrawal syndromes in this population may contribute to this association. Although the diagnosis of alcohol withdrawal is usually evident from the clinical situation, a history of alcoholism is often withheld and a high index of suspicion must be maintained. The opioid antagonist naloxone, or the combined agonist-antagonist pentazocine can also induce hypertension by inducing opioid-withdrawal [3]. Beta-blocker and clonidine withdrawal cause a syndrome with markedly elevated catecholamines and prominent hyperadrenergic symptoms, typically lasting for a few days. Some patients may be particularly sensitive to clonidine and develop daily rebound when used at inappropriate dosing intervals, such as once daily or on “as-needed” basis. Beta-blockers may worsen vasoconstriction in hyperadrenergic states, such as pheochromocytoma or clonidine-rebound, by blocking β2 mediated vasodilation and allowing unopposed α1-mediated vasoconstriction. Clonidine rebound can be minimized by tapering gradually, avoidance of concurrent beta-blockers, or by the addition of an α-blocking agent. Tizanidine is a muscle relaxant with a mechanism-of-action similar to clonidine, which may cause rebound hypertension when administered at high dose, taken once daily, or stopped abruptly. Tizanidine is often used or abused in conjunction with sedative-hypnotics, and convincing patients to discontinue it can be difficult. Although α-methyldopa is also an α2-agonist, rebound is less common due to its longer half-life and active metabolites. Antihypertensive medications can also have paradoxical effects on blood pressure. When given parenterally or in overdose, clonidine initially produces hypertension due to peripheral α2-receptor activation, followed by the centrally-mediated hypotensive effects. Dexmedetomidine is an α2-agonist used intravenously for its sedativeanalgesic properties, which also causes initial hypertension and bradycardia when given as an intravenous bolus. Clonidine’s peripheral vasoconstrictor effects may also predominate in patients with autonomic dysfunction, and discontinuation may actually improve blood pressure control [4]. As mentioned above, beta-blockers can increase blood pressure in the setting of excess catecholamines due to unopposed alpha effects. Drugs which cause hypertension via an off-target or unknown mechanism may be recognized after a

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medication has been marketed for many years. For example, clonidine-withdrawal hypertension was discounted for many years before its existence was generally accepted [5]. Although hypertension clearly complicates angiogenesis inhibitor (VEGF antagonist and receptor tyrosine kinase inhibitor) administration, the clinical importance is becoming increasingly recognized as the targeted population broadens. Cylcooxygenase (COX) inhibitors have long been recognized to raise blood pressure, but the adverse cardiovascular effects of these drugs have only recently been established. Other agents such as glucocorticoids and androgens can clearly cause hypertension, although the precise mechanisms remain unclear. Pharmacologic weight-loss agents have repeatedly targeted adrenergic pathways to treat obesity, producing hypertension as a common side effect. The serotonin and norepinephrine reuptake inhibitor sibutramine may promote weight loss, but is associated with an increased cardiovascular event rate, which prompted its withdrawal from the market in 2010. Sibutramine increases peripheral norepinephrine turnover and increases peripheral vasoconstriction, but also acts centrally to inhibit sympathetic activation. Due to this complex interaction, sibutramine may have pro-hypertensive or antihypertensive effects depending on the clinical substrate [6]. Ephedra (or ma huang) is a sympathomimetic herbal extract used for asthma treatment, weight loss, and enhanced athletic performance, which is associated with severe hypertension, cardiovascular events, and even death in young, apparently healthy individuals. Caffeine co-administration likely exacerbates ephedra-related complications [1]. Ephedra-caffeine also causes hypokalemia, hyperglycemia, and QT prolongation, contributing to the adverse cardiovascular profile. The FDA banned ephedra in 2004 due to these concerns and a lack of clinical benefit, a remarkable action considering the “herbal” designation of this agent. However, the lack of nutritional supplement regulation and standardization is an ongoing concern for similar agents. Ephedra-like compounds may still be found in supplements under various names, but the risks of these compounds are unclear. Although sibutramine and ephedra have been withdrawn from the market, herbal supplements are occasionally “adulterated” with these compounds or similar substitutes, and these drugs could also be illegally obtained over the internet from international sources [7]. A high index of suspicion should be maintained for any weight-loss supplement due to the recurring link to hypertension and cardiovascular events. Performance athletes or enthusiastic weight lifters may also take sympathomimetic supplements, which comprise many of the medications banned by the World AntiDoping Agency [8]. Another herbal supplement, glycyrrhizin, can produce hypertension with hypokalemic metabolic alkalosis via inappropriate mineralocorticoid receptor activation. Herbal glycyrrhizin is marketed for liver detoxification, gastrointestinal health, anti-inflammatory effects, and

other purposes. It is also found in true “black” licorice, in parts of Europe, and is used as a tobacco sweetener (chewing tobacco, “snuff”, “dip”, etc.). “Snuff” is a form of tobacco inserted along the gum line or intranasally, and is usually omitted from the initial tobacco abuse questioning. Licorice in the United States typically contains an artificial sweetener rather than glycyrrhizin. This supplement inhibits the enzyme 11-beta-hydroxysteroid dehydrogenase type II (11βHSD-2), which is co-expressed in renal epithelial tissues with the mineralocorticoid receptor (MR). 11βHSD-2 inactivates cortisol and prevents inappropriate MR activation by glucocorticoids. Carbenoxolone also inhibits 11βHSD-2, and is marketed for its gastrointestinal effects. In addition to hypertension, severe hypokalemia may cause arrhythmia, paralysis, rhabdomyolysis, and respiratory collapse. Laboratory features include suppressed plasma aldosterone and renin activity, and an increased urinary cortisol/cortisone ratio. Ophthalmic drops, intra-articular injections, intranasal sprays, topical ointments, and rectal suppositories are systemically absorbed and may produce systemic effects. Blood pressure effects have not been extensively studied for most agents, and practitioners should not reflexively disregard these agents as a contributing factor in hypertension. Inhaled epinephrine is available over-the-counter in the United States, and inappropriate usage produces episodic hypertension. Intra-articular glucocorticoids enter the systemic circulation and suppress the hypothalamicpituitary-adrenal axis even after a single injection, and may affect blood pressure [9]. Topical NSAIDs are commonly used for pain relief (i.e., as creams or lotions), and this route can rarely produce serious toxicity or even fatal overdose. Other agents and drug combinations which may worsen hypertension are provided in Boxes 105.1 and 105.2.

DRUG-INDUCED HYPOTENSION Although many drugs affect a single determinant of the blood pressure (vascular tone, heart rate, volume status, etc.), a clinically evident change in blood pressure is normally blunted by compensatory responses. This is observed during treatment with vasodilator agents (e.g., minoxidil) in the treatment of hypertension, which cause activation of sympathetic nervous system and the reninangiotensin-aldosterone system. This leads to resistance to their anti-hypertensive effect, unless a beta-blocker and diuretic are added. Similarly, an offending drug is more likely to reduce blood pressure during blockade of compensatory pathways, use of diuretics, or in the setting of certain comorbidities. Clinically asymptomatic autonomic dysfunction is particularly prevalent in the elderly and patients with vascular disease or diabetes, increasing this risk. Other factors may also increase drug exposure via drug interactions or pharmacogenetic effects. Patients may take drugs with anti-hypertensive effects for another indication, unaware of the effect on blood

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BOX 105.1

D R U G S W H I C H P R O D U C E H Y P E RT E N S I O N

Sympathetic Activation a. b. c. d. e. f. g. h.

i. j. k.

Mineralocorticoid Receptor (MR) Mediated

Illicit drugs: amphetamine, MDMA (“Ecstasy”), cocaine Wakefulness agents: modafanil, adrafanil, armodafinil ADHD treatment: dextroamphetamine, methylphenidate Over the counter: pseudoephedrine, nasal decongestants, eye drops Herbal drugs: ephedra (herbal formulations: ma huang, Metabolife, RippedFuel, etc.)† Weight loss agents: phenylpropanolamine, phentermine, dexfenfluramine Central α2-antagonists: yohimbine Central α2-agonists: i. Clonidine, given parenterally, in overdose, or in autonomic failure ii. Dexmedetomidine intravenous bolus Ergot alkaloids: ergotamine, dihydroergotamine Caffeine Water ingestion.

Drug Withdrawal Syndromes a. Ethanol, opioid, or benzodiazepine withdrawal b. Opioid antagonist-induced: naloxone, pentazocine c. Central α2-agonist withdrawal: clonidine, tizanidine, α-methyldopa d. Beta-blocker withdrawal: e.g. atenolol, metoprolol.

a. Indirect (11βHSD-2 inhibition): glycyrhizzic acid (in licorice, tobacco), carbenoxolone b. Direct MR agonists: fludrocortisone c. Altered steroid degradation: ketoconazole.

Miscellaneous a. Steroids: glucocorticoids, oral contraceptives, androgens b. Non-steroidal anti-inflammatory drugs: ibuprofen, naproxen, etc. c. Angiogenesis inhibitors: i. VEGF antagonists: bevacizumab ii. Receptor tyrosine kinase inhibitors: sunitinib, sorafenib d. Ophthalmic agents: α-agonists e. Adulterated herbal supplements f. Dopaminergic antagonists: metoclopramide g. Cholesterol-ester transport inhibition with torcetrapib h. Calcineurin inhibitors: tacrolimus, cyclosporine i. Erythropoietin j. Hypoventilation. †

Ephedra contains multiple compounds including ephedrine, pseudoephedrine, and phenylpropanolamine.

Abbreviations: ADHD, attention deficit hyperactivity disorder; MDMA, 3,4-Methylenedioxymethamphetamine; TCA, tricyclic antidepressant; MAOI, monoamine oxidase inhibitor; MR, mineralocorticoid receptor; 11-βHSD-2, 11-beta-hydroxysteroid dehydrogenase type II; NSAID, non-steroidal anti-inflammatory drugs; VEGF, vascular endothelial growth factor.

Antidepressant Agents a. Norepinephrine reuptake inhibitors: atomoxetine, reboxetine b. Serotonin/norepinephrine reuptake inhibitors: venlafaxine, sibutramine, duloxetine c. Tricyclic antidepressants.

BOX 105.2

C O M P L E X D R U G I N T E R A C T I O N S W H I C H M AY W O R S E N H Y P E RT E N S I O N 1. Monoamine oxidase inhibitor plus tyramine containing foods 2. Norepinephrine reuptake inhibitor (e.g. atomoxetine, venlafaxine) plus yohimbine 3. Beta-blocker during clonidine-withdrawal, pheochromocytoma, or hyperadrenergic state

4. Selective serotonin reuptake inhibitor plus dextromethorphan 5. Adulterated herbal supplements (e.g., with sympathomimetic) 6. Sympathomimetics with caffeine.

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BOX 105.3

D R U G S W H I C H R E D U C E B L O O D P R E S S U R E O R I M PA I R AU T O R E G U L A T I O N 1. Antihypertensive agents 2. Anti-Parkinsonian drugs: a. Dopamine precursors: levodopa b. vvCOMT inhibitors: tolcapone, entacapone c. Dopamine agonists: bromocriptine, pramipexole, ropinrole, apomorphine, rotigotine 3. Atypical muscle relaxants: tizanidine 4. Prostatic hypertrophy/α1-antagonists: terazosin, doxazosin

pressure. For example, α1-antagonists such as terazosin are commonly prescribed to increase urinary flow, rather than as an anti-hypertensive agent. Anti-Parkinsonian drugs commonly lower blood pressure and cause orthostatic hypotension. Tizanidine is a short-acting, central α2-adrenergic agonist similar to clonidine, prescribed for the treatment of muscle spasticity, or off-label for migraine headache, fibromyalgia, insomnia, and other disorders. As mentioned above, rebound hypertension is possible during withdrawal from tizanidine. However, a more frequent, dose-dependent side effect is hypotension, and factors which increase tizanidine drug exposure increase this risk. CYP1A2 inhibitors (e.g., ciprofloxacin, fluvoxamine, and verapamil) drastically increase tizanidine drug concentration and risk of hypotension by altering drug metabolism. Ciprofloxacin causes a 10-fold, and fluvoxamine a 33-fold increase in tizanidine drug exposure [10]. Reports of illegally-adulterated herbal medications are now fairly widespread, and could contribute to the problem. For example, herbal supplements for erectile dysfunction may contain phosphodiesterase inhibitors, and lower blood pressure [7]. Additional drugs which impair the protective response against hypotension are detailed in other chapters, and in Box 105.3.

SUMMARY The frequent use of over-the-counter, off-label, and herbal medications may contribute to drug-induced dysregulation of blood pressure control. Physicians should

5. Phosphodiesterase type 5 inhibitors: sildenafil, tadalafil, vardenafil 6. Adulterated herbal supplements (with anti-hypertensive agents, PDE-5 inhibitors, etc.) 7. Narcotic agents (opioids, benzodiazepines) 8. Beta2-agonists in autonomic failure 9. Miscellaneous: a. Food ingestion b. Hyperventilation.

extend medication inquiries to detect these specific classes of medications, and be cautious of any new drugs. A wise approach would be to discontinue any unnecessary drugs, especially those not providing their intended benefit.

References [1] Haller CA, Benowitz NL. Adverse cardiovascular and central nervous system events associated with dietary supplements containing ephedra alkaloids. N Engl J Med 2000;343:1833–8. [2] Grossman E, Messerli FH. Secondary hypertension: interfering substances. J Clin Hypertens (Greenwich) 2008;10:556–66. [3] Challoner KR, McCarron MM, Newton EJ. Pentazocine (Talwin) intoxication: report of 57 cases. J Emerg Med 1990;8:67–74. [4] Robertson D, Goldberg MR, Hollister AS, Wade D, Robertson RM. Clonidine raises blood pressure in severe idiopathic orthostatic hypotension. Am J Med 1983;74:193–200. [5] Reid JL, Wing LM, Dargie HJ, Hamilton CA, Davies DS, Dollery CT. Clonidine withdrawal in hypertension. Changes in blood-pressure and plasma and urinary noradrenaline. Lancet 1977;1:1171–4. [6] Birkenfeld AL, Schroeder C, Boschmann M, Tank J, Franke G, Luft FC, et al. Paradoxical effect of sibutramine on autonomic cardiovascular regulation. Circulation 2002;106:2459–65. [7] Bogusz MJ, Hassan H, Al-Enazi E, Ibrahim Z, Al-Tufail M. Application of LC-ESI-MS-MS for detection of synthetic adulterants in herbal remedies. J Pharm Biomed Anal 2006;41:554–64. [8] Docherty JR. Pharmacology of stimulants prohibited by the World Anti-Doping Agency (WADA). Br J Pharmacol 2008;154:606–22. [9] Habib GS. Systemic effects of intra-articular corticosteroids. Clin Rheumatol 2009;28:749–56. [10] Granfors MT, Backman JT, Neuvonen M, Ahonen J, Neuvonen PJ. Fluvoxamine drastically increases concentrations and effects of tizanidine: a potentially hazardous interaction. Clin Pharmacol Ther 2004;75:331–41.

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C H A P T E R

106 Postural Tachycardia Syndrome (POTS) Phillip A. Low, Paola Sandroni INTRODUCTION

complaint in about half the patients. Episodic, nonorthostatic symptoms of autonomic surges are common.

Orthostatic hypotension is well recognized but is relatively uncommon. For every patient seen with orthostatic hypotension, there are approximately 5–10 patients with orthostatic intolerance, defined as the development upon standing of symptoms of cerebral hypoperfusion (e.g., lightheadedness, weakness, blurred vision) associated with those of sympathetic activation (such as tachycardia, nausea, tremulousness) and an excessive heart rate increment (30 bpm). Female:male ratio is about 4–5:1 and most cases occur between the ages of 15 and 50 years [1,2].

CLINICAL FEATURES AND PHENOTYPES POTS is best considered a condition rather than a disease so that the same patient may or may not fulfill criteria at different times. There is considerable heterogeneity. Some of the disparate reports likely relate to different phenotypes, with different underlying mechanisms. There are, however, sufficient features in common to describe typical clinical features. Two studies best characterize the Mayo experience. First is a prospective evaluation of a cohort of 108 patients with POTS using a structured and validated autonomic symptom profile, consisting of 167 questions encompassing 10 autonomic categories of symptoms [3]. The second is a review of 152 patients over 10 years seen by two investigators (Low and Sandroni) with uniform testing and evaluation [2] (Table 106.1). Fifty percent of patients have an antecedent viral illness. Symptoms of dysautonomia were frequent or persistent (64%) and at least moderately severe in the majority and were either unchanged or getting worse in almost all patients (93%) at presentation. Positive family history of similar complaints occurred in 25%. The following orthostatic symptoms occurred in 75% of subjects: lightheadedness/dizziness, lower extremity or diffuse weakness, disequilibrium, tachycardia, shakiness. Hence the symptoms were due to a combination of hypoperfusion and autonomic activation. These symptoms were most commonly aggravated by ambient heat, meals, and exertion. Other autonomic symptoms were dry eyes or mouth, gastrointestinal complaints of bloating, early satiety, nausea, pain, and alternating diarrhea and constipation. Fatigue is a significant

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00106-2

PHENOTYPES OF POTS Three common phenotypes are sufficiently distinct to warrant separate descriptions. Their main distinguishing features are summarized in Table 106.2 and are described in more detail below.

Neuropathic POTS Our first contribution on POTS focused on the neuropathic basis of a significant subset of this condition [4]. Onset of POTS was often post-viral and peripheral autonomic denervation was manifest as loss of sweating on QSART and thermoregulatory sweat test distally. Adrenergic denervation was evident from loss of vasoconstrictor reflexes during the Valsalva maneuver. Our subsequent experience [2] supports this notion. Ganglionic A3 acetylcholine receptor antibody [5]found in 14% of patients [2] adds further support for functional sympathetic denervation. A study by Jacob et al. (2000) additionally implicates peripheral adrenergic denervation in a group of patients with POTS [6]. Plasma norepinephrine increment to sympathetic activation was intact in blood drawn from arm veins but reduced from corresponding leg veins. Our finding [2] of an increase in CASS-adrenergic score (presumed to reflect impaired baroreflex-mediated vasoconstriction to the Valsalva maneuver or HUT) is also consistent with this report. Impaired limb arteriolar vasoconstriction and increased venous compliance have been reported [7]. Gastrointestinal complaints as bloating, nausea, abdominal pain and constipation are reported to be more frequent in POTS with denervation [8].

Hyperadrenergic POTS Three features characterize hyperadrenergic POTS. First, patients have an excessive increment in plasma norepinephrine (NE) with standing values 600 pg/ml [2,9–11]. Second, there is sustained or fluctuating pressor response to head-up tilt so that systolic BP increases during HUT. Third, these patients have symptoms of sympathetic overactivity, which can be episodic or sustained.

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106. POSTuRAL TACHyCARdIA SyNdROmE (POTS)

These are characterized by episodes of tachycardia, hypertension, and hyperhidrosis [12]. When these are prominent, they comprise autonomic storms. The episodes can be spontaneous or provoked. Stimuli include stress, activity, and orthostatic stress.

POTS Associated with Poor Conditioning It has been argued, with significant evidence, that deconditioning of some degree is present in virtually all patients with POTS. However, poor conditioning is especially evident in two groups of patients. The first comprise previously well patients who undergo a period of bedrest

TABLE 106.1 Orthostatic Symptoms as Frequency (%) in Patients with POTS ORTHOSTATIC SYMPTOMS Light-headed or dizziness Palpitations Presyncope Exacerbation by heat Exacerbation by exercise Sense of weakness Tremulousness Shortness of breath Chest pain Exacerbation by meals Exacerbation associated with menses Hyperhidrosis Loss of sweating NON-ORTHOSTATIC SYMPTOMS Nausea Bloating Diarrhea Constipation Abdominal pain Bladder symptoms Vomiting Pupillary symptoms (glare) DIFFUSE ASSOCIATED SYMPTOMS Fatigue Sleep disturbance Migraine headache Myofascial pain Neuropathic type pain

FREQUENCY 118 114 92 81 81 76 57 42 37 36 22 14 8

% 78 75 61 53 53 50 38 28 24 24 15 9 5

FREQUENCY 59 36 27 23 23 14 13 5

% 39 24 18 15 15 9 9 3

FREQUENCY 73 48 42 24 3

% 48 32 28 16 2

From Thieben MJ, Sandroni P, Sletten DM, Benrud-Larson LM, Fealey RD, Vernino S, Lennon VA, Shen WK, Low PA. Postural Orthostatic Tachycardia Syndrome: The Mayo Clinic Experience. Mayo Clinic Proceedings 82:308–313, 2007. Modified with permission.

associated with some illness. These patients develop the type of deconditioning akin to that in astronauts following extended space flight. Symptoms of orthostatic intolerance tend to be mild and the prognosis is excellent [13]. A second group of patients have life-long orthostatic intolerance, poor exercise tolerance, and significant fatigue.

FOLLOW-UP On follow-up, 80% of patients were improved and 90% were able to return to work, although only 60% were functionally normal [3]. Some symptoms tend to persist and exercise tolerance is impaired. Patients who had an antecedent event appeared to do better than those with spontaneous POTS. Salt supplementation and beta-blockers were the most efficacious therapies. In a prospective study, this trend is seen but the improvement is less evident [14].

MANAGEMENT A variety of approaches have been used to alleviate symptoms in POTS. All patients need volume expansion and a high salt/high fluid regimen [1,11]. Certain drugs appear to be beneficial, at least temporarily. The most widely used drugs are midodrine, propranolol, and fludrocortisone. Other measures used include body stockings and physical counter maneuvers. Acutely, significant improvement has been described following treatment with midodrine and saline and pyridostigmine [15]. Pyridostigmine, in short term studies, improves both orthostatic tachycardia and their associated symptoms [16,17]. There is significant uncertainty on the benefits of drugs in long-term therapy of POTS. The concept that at least some of the chronic symptoms in POTS are due in part to secondary deconditioning raises the possibility that a “lifehardening” exercise training program might have utility in this condition. This approach has been used in a number of studies of fibromyalgia patients and has been successful in reducing disability and clearly warrants serious consideration in POTS [18]. A recent well-done study in POTS supports this approach [19]. An exercise program, provided it is sustained and adequate for several months can be very helpful [19]. Based on the small study noted above, the physiological data, and anecdotal reports, larger clinical trial of exercise training in POTS may be warranted.

TABLE 106.2 Some distinguishing Features of Common Types of POTS POTS Category

HR on HUT

BP on HUT

Plasma NE

QSART/TST

Autonomic “storms”

Neuropathic Hyperadrenergic Deconditioned

↑↑ ↑↑↑ ↑

Mild ↓ BP↑ PP ↓

N 600 pg/ml N or ↑

Distal anhidrosis Normal Normal

Absent/infrequent Common Absent

HUT, head-up tilt; PP, pulse pressure; QSART (quantitative sudomotor axon-reflex test); TST (thermoregulatory sweat test).

XI. ORTHOSTATIC INTOLERANCE

SummARy

CONCLUSIONS POTS is common and is heterogeneous in presentation and pathophysiologic mechanisms. Common mechanisms are denervation (neuropathic POTS), hyperadrenergic state, and deconditioning. Management always involves expansion of plasma volume with high salt and high fluid intake. Additional steps include compression garments. Medications that are commonly used include betablockers, midodrine, and fludrocortisone. Exercise training and reconditioning is emerging as a very important strategy in the deconditioned subject.

SUMMARY Postural tachycardia syndrome (POTS) is a syndrome of orthostatic tachycardia associated with symptoms of cerebral hypoperfusion and/or autonomic activation. The patient is usually a female between the ages of 20 and 50 years. This chapter will focus on phenotypes of POTS and an introduction to management. Chapter 107 will focus specifically on the mechanisms of POTS. Some patients have a limited autonomic neuropathy. An antecedent event such as a viral infection occurs in approximately half the patients. Evidence of peripheral denervation of sudomotor fibers includes sweat loss of the legs on thermoregulatory sweat test or QSART. Peripheral adrenergic denervation can be present, resulting in impairment of reflex vasoconstriction with baroreflex unloading. In hyperadrenergic POTS, sympathetic tone is increased, manifest as orthostatic hyperadrenergic response and sometimes as spontaneous episode of excessive sympathetic activity. POTS is usually associated with deconditioning and in POTS with deconditioning, this can be the dominant feature. The linchpins of management of POTS comprise volume expansion, education, and conditioning exercise. Medications such as beta-adrenoreceptor blockers, midodrine, and pyridostigmine have a limited role in management.

References [1] Low PA, Opfer-Gehrking TL, Textor SC, Benarroch EE, Shen WK, Schondorf R, et al. Postural tachycardia syndrome (POTS). Neurology 1995;45:S19–25.

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[2] Thieben M, Sandroni P, Sletten D, Benrud-Larson L, Fealey R, Vernino S, et al. Postural orthostatic tachycardia syndrome – Mayo Clinic experience. Mayo Clin Proc 2007;82:308–13. [3] Sandroni P, Opfer-Gehrking TL, McPhee BR, Low PA. Postural tachycardia syndrome: Clinical features and follow-up study. Mayo Clin Proc 1999;74:1106–10. [4] Schondorf R, Low PA. Idiopathic postural orthostatic tachycardia syndrome: an attenuated form of acute pandysautonomia? Neurology 1993;43:132–7. [5] Vernino S, Low PA, Fealey RD, Stewart JD, Farrugia G, Lennon VA. Autoantibodies to ganglionic acetylcholine receptors in autoimmune autonomic neuropathies. N Engl J Med 2000;343:847–55. [6] Jacob G, Costa F, Shannon JR, Robertson RM, Wathen M, Stein M, et al. The neuropathic postural tachycardia syndrome. N Engl J Med 2000;343:1008–14. [7] Stewart JM. Pooling in chronic orthostatic intolerance: arterial vasoconstrictive but not venous compliance defects. Circulation 2002;105:2274–81. [8] Al-Shekhlee A, Lindenberg JR, Hachwi RN, Chelimsky TC. The value of autonomic testing in postural tachycardia syndrome. Clin Auton Res 2005;15:219–22. [9] Garland EM, Raj SR, Black BK, Harris PA, Robertson D. The hemodynamic and neurohumoral phenotype of postural tachycardia syndrome. Neurology 2007;69:790–8. [10] Jordan J, Shannon JR, Diedrich A, Black BK, Robertson D. Increased sympathetic activation in idiopathic orthostatic intolerance: role of systemic adrenoreceptor sensitivity. Hypertension 2002;39:173–8. [11] Low PA, Sandroni P, Joyner MJ, Shen WK. Postural tachycardia syndrome (POTS). J Cardiovasc Electrophysiol 2009;20:352–8. [12] Figueroa JJ, Sandroni P, Singer W, Basford JR, Sletten D, Gehrking TL, et al. Hyperadrenergic postural tachycardia syndrome with sympathetic storms. Clin Auton Res 2010;20:305. [13] Masuki S, Eisenach JH, Johnson CP, Dietz NM, Benrud-Larson LM, Schrage WG, et al. Excessive heart rate response to orthostatic stress in postural tachycardia syndrome is not caused by anxiety. J Appl Physiol 2007;102:896–903. [14] Kimpinski K, Iodice V, Sletten DM, Singer W, Sandroni P, Lipp A, et al. Prospective analysis of postural tachycardia syndrome. Clin Auton Res 2010;20:291. [15] Gordon VM, Opfer-Gehrking TL, Novak V, Low PA. Hemodynamic and symptomatic effects of acute interventions on tilt in patients with postural tachycardia syndrome. Clin Auton Res 2000;10:29–33. [16] Raj SR, Black BK, Biaggioni I, Harris PA, Robertson D. Acetylcholinesterase inhibition improves tachycardia in postural tachycardia syndrome. Circulation 2005;111:2734–40. [17] Singer W, Opfer-Gehrking TL, Nickander KK, Hines SM, Low PA. Acetylcholinesterase inhibition in patients with orthostatic intolerance. J Clin Neurophysiol 2006;23:476–81. [18] Gowans SE, Dehueck A, Voss S, Silaj A, Abbey SE. Six-month and one-year followup of 23 weeks of aerobic exercise for individuals with fibromyalgia. Arthritis Rheum 2004;51:890–8. [19] Winker R, Barth A, Bidmon D, Ponocny I, Weber M, Mayr O, et al. Endurance exercise training in orthostatic intolerance: a randomized, controlled trial. Hypertension 2005;45:391–8.

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C H A P T E R

107 Mechanisms of Postural Tachycardia Syndrome Satish R. Raj It is best not to think about postural tachycardia syndrome (POTS) as a single disease. Rather, it is best viewed as a “disorder” or a syndrome. Excessive orthostatic tachycardia can be a final common pathway of many underlying pathophysiological processes. Many disorders with orthostatic tachycardia, a hallmark of POTS, have been described. Over the last 15 years, many different pathophysiological features have been described in different subgroups of patients with POTS. These subtypes may be of value in trying to understand the pathophysiology of POTS, and may help to develop rational therapeutic approaches. However, it is currently quite difficult to characterize an individual patient as belonging to one particular subtype. The putative pathophysiological mechanisms mentioned below are not mutually exclusive, and can co-exist in a particular individual. Even with insights into the pathophysiology underlying POTS, the etiology of POTS is usually unclear in any individual. Many patients with POTS describe an acute onset shortly following a viral illness. This suggests a possible autoimmune etiology to POTS, although a specific antibody has not been identified. Another large cohort of patients describe their symptoms as developing around the time of puberty, perhaps suggesting an intrinsic problem brought out by physiological changes. A small number of patients have described their POTS symptoms as developing some time following physical trauma, although a convincing putative mechanism has not been proposed.

HYPERADRENERGIC POTS

Central Hyperadrenergic POTS There are some cases of POTS in which the primary underlying problem seems to be excessive sympathetic discharge (Fig. 107.1A,B). These patients often have very high levels of upright plasma norepinephrine. Whereas the upright norepinephrine level is 600 pg/ml for hyperadrenergic POTS, the primary hyperadrenergic subgroup often has upright norepinephrine level 1000 pg/ml or even 2000 pg/ml. These patients can also have large increases in blood pressure on standing. Central hyperadrenergic POTS is much less common than neuropathic POTS, comprising only ~5–10% of patients. Therapy in these cases usually targets a decrease in sympathetic tone both centrally and peripherally. Central sympatholytic medications such as clonidine or alpha-methyldopa or beta-adrenergic blockers [3] can be used.

Norepinephrine Transporter Deficiency

Why do patients with POTS have an excessive heart rate on standing? One possible explanation is that excessive sympathoneural tone is driving the heart rate, and that this is particularly excessive with upright posture. This pathophysiological concept is the basis for the POTS subtype known as “hyperadrenergic POTS”. Norepinephrine is the primary neurotransmitter among post-ganglionic neurons in the sympathetic nervous system, and plasma norepinephrine can provide a crude

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00107-4

biochemical estimate of sympathetic nervous system activity. One suggested definition of hyperadrenergic POTS is a standing plasma norepinephrine  600 pg/ml (3.54 nM). Using this definition, the prevalence of hyperadrenergic POTS at American POTS referral centers range from under 30% [1] to over 60% [2] of patients with POTS. Using this definition, hyperadrenergic POTS may be the primary underlying problem (excessive central sympathetic outflow; “central hyperadrenergic POTS”) or may be secondary to another underlying problem. Some of these problems that can secondarily result in increased sympathoneural tone are outlined below.

The norepinephrine transporter (NET) is a presynaptic transporter in sympathetic neurons that is important for the clearance of synaptic norepinephrine. A specific genetic abnormality has been identified in a kindred with hyperadrenergic POTS [4]. These individuals have a single point mutation that causes a loss of function in the norepinephrine transporter, and the resultant inability to adequately clear norepinephrine produces a state of excess sympathetic activation.

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107. MECHAnISMS of PoSTuRAl TACHyCARdIA SyndRoME

Although this functional NET mutation is infrequent, pharmacological NET inhibition is very common. Many psychotropic and fibromyalgia medications inhibit NET. This includes traditional drugs such as tricyclic antidepressants (e.g., desipramine), and newer medications which are pure NET inhibitors (e.g., atomoxetine or reboxetine). Pharmacological NET inhibition can recreate an orthostatic tachycardia phenotype in otherwise healthy volunteer subjects [5,6].

A-Normal

Mast Cell Activation Disorder Some patients with POTS have co-existent mast cell activation disorder (MCAD). These patients often present with episodic flushing. Around episodes, they will have abnormal increases in urinary methylhistamine (the primary urinary metabolite of histamine) [7]. This assessment should ideally be measured in a 4-hour aliquot at the time of a flushing episode and not just in a random 24-hour period (which has a very low yield). Other associated symptoms can include shortness of breath, headache, excessive diuresis, and gastrointestinal symptoms such as diarrhea, nausea, and vomiting. Flushing can be triggered by long-term standing, exercise (including sexual intercourse), premenstrual cycle, and meals. These patients often have a hyperadrenergic response to posture, with both orthostatic tachycardia and orthostatic hypertension. It is not clear whether the primary event in this disorder is the mast cell activation (release of vasoactive mediators), or sympathetic activation (release of norepinephrine, neuropeptide Y and ATP) [8]. In patients with MCAD, beta-adrenergic blockers can actually trigger a flushing episode and worsen symptoms. Centrally acting sympatholytic agents (e.g., alpha-methyldopa or clonidine) may prove effective. Alternatively, treatment could target mast cell mediators with a combination of antihistamines (H1- and H2antagonists) and with the cautious use of non-steroidal agents (high dose aspirin) in refractory cases.

B-Neuropathic POTS

C-Central Hyperadrenergic POTS

NEUROPATHIC POTS

FIGURE 107.1 Pathophysiological schema in POTS. There are multiple distinct pathophysiological subtypes within the postural tachycardia syndrome (POTS). (A) Shows a basal situation with a normal amount of sympathetic nervous system outflow from the brain that activates receptors in the blood vessels (vascular tone and venous return), heart (heart rate and contractility) and kidney (blood volume regulation through renin). (B) Shows a schematic of neuropathic POTS. There is patchy denervation of the sympathetic innervation of the blood vessels in the extremities (especially the legs) and the kidney with subsequent hypovolemia and increased orthostatic venous pooling. This feeds back to the brain to increase sympathetic nervous system outflow in a compensatory effort. This increased sympathoneural flow is sensed most in the heart where there is no denervation. (C) Shows a schematic of central hyperadrenergic POTS. In this case, the underlying problem is excessive sympathetic nervous outflow from the brain that affects the blood vessels, kidneys and the heart. In addition to tachycardia, this form of POTS is often associated with orthostatic hypertension. Figures reprinted with permission from Raj SR et al., Indian Pacing Electrophysiol. J. 2006;6:84–99 [22].

Some POTS patients have a form of dysautonomia, with preferential denervation of sympathetic nerves innvervating the lower limbs [9–11]. There have been several findings consistent with this hypothesis. The results of sudomotor axon reflex testing [9] and galvanic skin stimulation [10] support this as well as skin biopsy results [12]. Further, these patients have been found to be hypersensitive to infusions of norepinephrine and phenylephrine into veins of the foot [11], suggesting denervation hypersensitivity of the leg veins. Jacob et al. [13] used a segmental norepinephrine spillover approach to demonstrate that some patients with POTS had diminished norepinephrine release (and thus less sympathetic activation) in their lower body, but normal levels in their arms. Some patients

XI. ORTHOSTATIC INTOLERANCE

low STRokE VoluME

with neuropathic POTS can also have excess sympathetic tone and be hyperadrenergic (Fig. 107.1C).

HYPOVOLEMIA AND BLOOD VOLUME REGULATION Many patients, but not all, with POTS have low plasma volumes [14–17]. Using 131-I labeled human serum albumin and a dye dilution technique, we have consistently found that a majority of patients with POTS had a deficit of their plasma volume, red cell volume and total blood volume compared to the control group. The renin-angiotensin-aldosterone system plays a key role in the neurohormonal regulation of plasma volume in humans and may be a part of the problem in POTS. We have found that many patients with POTS who were also hypovolemic have low levels of standing plasma renin activity and aldosterone compared to normovolemic patients [18,19]. These data suggest that abnormalities in the renin-angiotensin-aldosterone axis might have a role in the pathophysiology of POTS by contributing to hypovolemia and impaired sodium retention. Such hypovolemia could be accounted for by a neuropathic process involving the kidney. A significant modulator of renin release is the sympathetic nervous system. Thus perturbations in the renin-aldosterone system might result from partial sympathetic denervation involving the kidney.

LOW STROKE VOLUME Patients with POTS have been found to have a low stroke volume and reduced left ventricular mass, especially in the upright posture [20,21]. In an effort to maintain cardiac output and blood pressure, one would expect a physiological increase in heart rate. These findings are suggestive of deconditioning, which can quickly develop in patients with POTS due to the fatigue and exercise intolerance regardless of the initial underlying pathophysiology. Fu et al. [21] have recently shown that a structured 3-month exercise intervention could reduce the orthostatic tachycardia and improve symptoms in patients with POTS by reversing some of this deconditioning.

References [1] Thieben MJ, Sandroni P, Sletten DM, Benrud-Larson LM, Fealey RD, Vernino S, et al. Postural orthostatic tachycardia syndrome: the Mayo clinic experience. Mayo Clin Proc 2007;82:308–13. [2] Garland EM, Raj SR, Black BK, Harris PA, Robertson D. The hemodynamic and neurohumoral phenotype of postural tachycardia syndrome. Neurology 2007;69:790–8.

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[3] Raj SR, Black BK, Biaggioni I, Paranjape SY, Ramirez M, Dupont WD, et al. Propranolol decreases tachycardia and improves symptoms in the postural tachycardia syndrome: less is more. Circulation 2009;120:725–34. [4] Shannon JR, Flattem NL, Jordan J, Jacob G, Black BK, Biaggioni I, et al. Orthostatic intolerance and tachycardia associated with norepinephrine-transporter deficiency. N Engl J Med 2000;342:541–9. [5] Vincent S, Bieck PR, Garland EM, Loghin C, Bymaster FP, Black BK, et al. Clinical assessment of norepinephrine transporter blockade through biochemical and pharmacological profiles. Circulation 2004;109:3202–7. [6] Schroeder C, Tank J, Boschmann M, Diedrich A, Sharma AM, Biaggioni I, et al. Selective norepinephrine reuptake inhibition as a human model of orthostatic intolerance. Circulation 2002;105:347–53. [7] Shibao C, Arzubiaga C, Roberts LJ, Raj S, Black B, Harris P, et al. Hyperadrenergic postural tachycardia syndrome in mast cell activation disorders. Hypertension 2005;45:385–90. [8] Arzubiaga C, Morrow J, Roberts LJ, Biaggioni I. Neuropeptide Y, a putative cotransmitter in noradrenergic neurons, induces mast cell degranulation but not prostaglandin D2 release. J Allergy Clin Immunol 1991;87:88–93. [9] Schondorf R, Low PA. Idiopathic postural orthostatic tachycardia syndrome: an attenuated form of acute pandysautonomia?. Neurology 1993;43:132–7. [10] Hoeldtke RD, Davis KM. The orthostatic tachycardia syndrome: evaluation of autonomic function and treatment with octreotide and ergot alkaloids. J Clin Endocrinol Metab 1991;73:132–9. [11] Streeten DH. Pathogenesis of hyperadrenergic orthostatic hypotension. Evidence of disordered venous innervation exclusively in the lower limbs. J Clin Invest 1990;86:1582–8. [12] Singer W, Spies JM, McArthur J, Low J, Griffin JW, Nickander KK, et al. Prospective evaluation of somatic and autonomic small fibers in selected autonomic neuropathies. Neurology 2004;62:612–8. [13] Jacob G, Costa F, Shannon JR, Robertson RM, Wathen M, Stein M, et al. The neuropathic postural tachycardia syndrome. N Engl J Med 2000;343:1008–14. [14] Jacob G, Robertson D, Mosqueda-Garcia R, Ertl AC, Robertson RM, Biaggioni I. Hypovolemia in syncope and orthostatic intolerance role of the renin-angiotensin system. Am J Med 1997;103:128–33. [15] Fouad FM, Tadena-Thome L, Bravo EL, Tarazi RC. Idiopathic hypovolemia. Ann Intern Med 1986;104:298–303. [16] Streeten DH, Thomas D, Bell DS. The roles of orthostatic hypotension, orthostatic tachycardia, and subnormal erythrocyte volume in the pathogenesis of the chronic fatigue syndrome. Am J Med Sci 2000;320:1–8. [17] Raj SR, Robertson D. Blood volume perturbations in the postural tachycardia syndrome. Am J Med Sci 2007;334:57–60. [18] Raj SR, Biaggioni I, Yamhure PC, Black BK, Paranjape SY, Byrne DW, et al. Renin-aldosterone paradox and perturbed blood volume regulation underlying postural tachycardia syndrome. Circulation 2005;111:1574–82. [19] Mustafa HI, Garland EM, Biaggioni I, Black BK, Dupont WD, Robertson D, et al. Abnormalities of angiotensin regulation in postural tachycardia syndrome. Heart Rhythm 2011 [20] Masuki S, Eisenach JH, Schrage WG, Johnson CP, Dietz NM, Wilkins BW, et al. Reduced stroke volume during exercise in postural tachycardia syndrome. J Appl Physiol 2007;103:1128–35. [21] Fu Q, Vangundy TB, Galbreath MM, Shibata S, Jain M, Hastings JL, et al. Cardiac origins of the postural orthostatic tachycardia syndrome. J Am Coll Cardiol 2010;55:2858–68. [22] Raj SR. The postural tachycardia syndrome (POTS): pathophysiology, diagnosis and management. Indian Pacing Electrophysiol J 2006;6:84–99.

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C H A P T E R

108 Symptoms and Signs of Postural Tachycardia Syndrome (POTS) Julian Stewart POTS is a form of orthostatic intolerance associated with excessive sinus tachycardia. Orthostatic intolerance is defined by symptoms and signs that make upright posture unsustainable over a period of time and are relieved by recumbence [1]. Orthostatic intolerance in POTS can be loosely partitioned among findings related to cerebral malperfusion and those related to sympathoexcitation and parasympathetic withdrawal; a partial list of symptoms is shown in Box 108.1. While a diagnosis of POTS specifically includes the signs of upright tachycardia and often circulatory insufficiency, it specifically excludes early orthostatic hypotension; however, progressive hypotension may appear late during an orthostatic challenge. In our experience hypotension severe enough to produce loss of consciousness does not generally occur except following extended, stressful laboratory conditions. Importantly, in POTS patients, symptoms of cerebral malperfusion may be associated with significant reductions in upright cerebral blood flow and impairment of cerebral autoregulation, compared to healthy subjects [2]. These cerebrovascular findings can relate to hyperpnea and hypocapnia caused by baroreflex unloading, but can also occur in the absence of significant respiratory findings. Our typical POTS patient does not faint in real life, perhaps due in part to positional and other aversive strategies adopted to maintain consciousness. Being defined by an array of symptoms and the sign of excessive upright tachycardia ensures that POTS remains a syndrome, and that its pathophysiology remains heterogeneous. Within certain limits discussed below any condition that excessively reduces central blood volume when upright can produce POTSlike findings.

HEMODYNAMICS – TACHYCARDIA A chapter concerning hemodynamics in POTS implicitly poses a hypothesis of circulatory abnormality in POTS. A first consideration in examining this hypothesis is to consider whether the defining tachycardia is physiological or pathophysiological. Tachycardia could be considered pathological if it directly causes disease or results from

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00108-6

disease affecting the sinoatrial node. Tachycardia could be considered physiological if it were a normal reflex response of the sinoatrial apparatus to changing cardiovascular loading conditions. Does sinus tachycardia present a direct risk to cardiovascular health? This is a difficult question since reports of illness caused by sinus tachycardia are sparse. However, it is true that abnormal atrial and ventricular tachycardia comprise symptomatic illness, and rapidly conducted atrial flutter with AV-nodal conduction can produce a form of “pacemaker-induced cardiomyopathy” in the young akin to experimentally produced pacemaker heart failure models. The arrhythmic origin of cardiomyopathy may not be readily apparent if the flutter rhythm is intermittent. Even so, very high and often unremitting heart rates are required to exert such deleterious effects. Such sustained tachycardia is not characteristic of POTS. Tachycardic cardiomyopathy has not been evident in any of our POTS patients. Does sinus tachycardia present an indirect risk to cardiovascular health? A categorization of the origins of tachycardia is shown in Box 108.2. Tachycardia may occur because of intrinsic structural or functional abnormalities of the sinoatrial (SA) node. The SA node is an autonomous oscillator capable of intrinsic rhythmicity preprogrammed by ionic channels and receptors and supine tachycardia could result from malfunction of this intrinsic apparatus. Indeed, the separate but similar problem of inappropriate sinus tachycardia (IST) may represent one such disorder of intrinsic SA node function. It is nearly impossible to separate a pure intrinsic defect from autonomic influence in man; the sinus node does not exist in isolation and even in a tonic resting state is subject to external modulation via paracrine, endocrine and perhaps autocrine neurotransmitters which can interact with intrinsic heart rate regulators. These transmitters are predominantly contained within and subserve sympathomimetic and parasympatholytic nerves. Thus, for example, the IST patient is responsive to norepinephrine from sympathetic activation and will demonstrate excessive postural as well as supine tachycardia. The in vivo human situation is made even more complex because of interactions between the

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BOX 108.1

S Y M P T O M S O F O RT H O S TAT I C I N T O L E R A N C E Lightheadedness/Dizziness Headache Fatigue Neurocognitive/Sleep Disorders Exercise Intolerance Shortness of Breath Chest pain (ST changes) Weakness Nausea/abodominal pain Sweating, Tremulousness Tachycardia Hypotension

}

}

cerebral malperfussion

Sympathoexcitatory Vagolytic

BOX 108.2

O R I G I N S O F S I N U S TA C H Y C A R D I A l l

l

Intrinsic sinoatrial node resting tachycardia Reflex excessive upright thoracic hypovolemia – distributive or absolute Autonomic – sinus node impact l Sympathomimetic: CNS or peripheral transduction (NE synthesis, NET, NPY, receptors, Ang, NO deficit)

sympathetic and parasympathetic nerves innervating the sinus node, because of feedback and nonlinearity within the autonomic nervous system, and because numerous neurotransmitters are involved within autonomic nerves. These include not only norepinephrine and acetylcholine but also neuropeptide Y, vasoactive intestinal polypeptide, angiotensin-II, and nitric oxide, to name a few.

REFLEX TACHYCARDIA IN POTS – A CLASSIFICATION BASED ON PERIPHERAL BLOOD FLOW While alterations in autonomic transmitters, their receptors and second messengers almost certainly have a distinct role in the tachycardia of POTS, hemodynamic disturbances which reflexively contribute to tachycardia also appear to exert an effect in most forms of POTS. Virtually all forms of POTS have a contribution arising from central hypovolemia which produces “physiological” tachycardia and has effects on neurovascular function, blood flow, blood volume, and baroreflex unloading. Our group has developed a scheme of classification of POTS based on measurements of supine peripheral blood

Parasympathetic withdrawal: tonic vs. orthostatic alterations in HR due to cholinergic and nitrergic (NO) mechanisms Altered ganglionic transmission Deconditioning. l

l l

flow which includes contributions from sympathoexcitation and parasympathetic withdrawal during orthostasis related to either an absolute or redistributive form of central thoracic hypovolemia. Thus, at least for the young patient, the pathophysiology of POTS includes abnormalities in blood volume, vascular regulation and sympathoexcitation resulting in reflex as well as directly mediated tachycardia. The flow regime classification which can encompass most forms of POTS evolved from measurements of leg venous pressure, Pv. Initially we grouped patients by Pv and examined their calf blood flow as shown in Figure 108.1. Three flow regimes emerged which have since formed the basis of our POTS classification [3]. These are: “low flow”, “high flow” and “normal flow” POTS. While these categories comprise fuzzy sets, this classification scheme has been helpful in shaping thoughts and formulating hypotheses.

Low Flow POTS Patients are characterized by supine vasoconstriction, increased peripheral venous pressure, reduced stroke volume and cardiac output, blunted orthostatic vasoconstrictive responses, often supine tachycardia and reduced

XI. ORTHOSTATIC INTOLERANCE

REflEx TACHyCARdIA In POTS – A ClASSIfICATIOn BASEd On PERIPHERAl BlOOd flOw

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norepinephrine in venous blood making this group similar to the “hyperadrenergic” POTS originally proposed by Streeten [5]. There are many candidate mechanisms for peripheral vasoconstriction; among these are abnormalities of norepinephrine reuptake (defects in the norepinephrine transporter or NET deficiency) which occur in some patients including those with heterozygous NET mutation [6] or with lesser partial deficiencies of norepinephrine reuptake [7]. Our group has also demonstrated a subset of low flow patients with increased plasma angiotensin-II (Ang-II) despite reductions in plasma renin and serum aldosterone due to a defect in angiotensin converting enzyme 2 (ACE2 deficiency) [8]. This is the primary catabolic pathway for Ang-II through an intermediate step involving the vasodilator angiotensin-(1–7). Ang-II has pleiotropic effects and causes vasoconstriction through potentiation of norepinephrine, overproduction of reactive oxygen species and decreasing the bioavailability of cutaneous nitric oxide (NO). Suppression of neuronal nitric oxide synthase (nNOS) is a fundamental mechanism that centrally enhances sympathoexcitation. Increased sympathetic activity has been demonstrated by peroneal nerve microneurographic assessment of muscle sympathetic nerve activity (MSNA) in some patients but NET deficiency, Ang-II excess, NO deficiency affect neurovascular transduction as well as neural-sinoatrial node signal transduction. In low flow POTS subjects, tonic increases in MSNA are sometimes found while supine, but mildly reduced MSNA also occurs which is consistent with the central alpha-2 adrenergic sympatholytic effects of norepinephrine characteristic of NET deficiency. Increased MSNA when upright appears to be more common and may be potentiated by reflex sympathoexcitation.

High Flow POTS

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FIGURE 108.1 The figure shows frequency histograms of leg blood flow. The top panel depicts control subjects. POTS patients are shown in the middle and lower panels segregated by venous pressure (Pv). Low flow associates with increased Pv while the normal Pv is bimodal representing the normal and high flow POTS subsets.

blood volume of uncertain origin [4]. The low flow POTS phenotype is distinguished by generalized pallor, cool skin, and other findings suggesting circulatory insufficiency. Cardiac size may be reduced by the trophic effects of hypovolemia and may contribute to its pathophysiology. There is marked female gender preference. There is also an inverse relationship with body mass index. Patients often have increased upright concentrations of

Patients are characterized by normovolemia and reduced total peripheral resistance while supine due to reduced peripheral vasoconstriction in the lower extremities. There is increased supine cardiac output compared to healthy volunteers. Relative lower extremity vasodilation persists during orthostatic stress causing venous pooling in the legs. This is not caused by abnormality of leg venous capacitance properties but rather by decreased release of norepinephrine from post ganglionic sympathetic nerves evidenced by a decrease in radioactive norepinephrine spillover [9]. Blood flow and blood volume are redistributed to the lower extremities and enhanced microvascular filtration as well as dependent venous pooling account for the central hypovolemia and postural tachycardia. A peripheral neuropathy is also evidenced by reduced distal sweating and some patients may have a variant of autonomic autoimmune neuropathy. Thus high flow POTS corresponds closely to neuropathic POTS and evidence suggests a mechanism involving partial sympathetic denervation.

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Normal Flow POTS Patients are characterized by normovolemia and normal hemodynamics while supine including normal heart rate, normal peripheral resistance, normal regional blood flows and normal cardiac output. Some of these patients have orthostatic vagal withdrawal with little evidence of a hyperadrenergic state or regional venous pooling. The postural tachycardia is modest in such patients and heart rates approximate heart rates achieved during chemical or cardiac transplant vagolysis. The most common finding in normal flow patients is abnormal vascular regulation when upright resulting in abdominal vascular pooling, intense peripheral vasoconstriction, and dependent acrocyanosis caused by venous stasis [10]. Findings are based on electrical impedance measurements which are not anatomically specific, but generally, excess blood volume shifts from the thoracic to the abdominal compartment while blood pooling within the legs remains the same as control; pelvic vessels could be involved as well. However, the splanchnic vasculature is the single largest venous reservoir in mammals with diverse innervation and established capability for direct autonomic venous regulation. Venous capacitance may dynamically increase in some patients as shown by “splanchnic filling” during a standard quantitative Valsalva maneuver. A relationship with hypermobility syndromes such as Ehlers–Danlos syndrome may exist. Echocardiographic evidence of faulty splanchnic vasoconstriction exists but data are sparse. Comparable findings of splanchnic pooling have been found in patients with simple faint. Recent evidence implicates an up-regulation of the NET transporter in certain forms of reflex fainting and one might speculate a similar effect in POTS with splanchnic pooling. However, cutaneous studies in normal flow POTS have shown increased receptor mediated NO release using intradermal acetylcholine. The rich nitrergic parasympathetic innervation of the splanchnic vasculature offers great potential for the participation of nitric oxide in mesenteric blood volume regulation. One further analogy might provoke thought: microgravity causes transient orthostatic intolerance. There are numerous reasons why this can happen, including reduced blood volume and postural muscle mass (skeletal muscle pump). Also, astronauts and bedrested

humans demonstrate defective vasoconstriction caused by defective neurovascular transduction. The hindlimb suspension rat models microgravity and has demonstrated specifically attenuated vasoconstrictor responsiveness of mesenteric resistance arteries in vivo and in vitro.

SUMMARY In summary, the hemodynamics of POTS comprises direct, modulated and reflex autonomic acceleration of the sinoatrial node with at least upright sympathoexcitation. Central hypovolemia is a feature common to most forms. While the mechanisms of POTS are general, the specifics of mechanism are likely to become more diversified.

References [1] Schondorf R, Low PA. Idiopathic postural orthostatic tachycardia syndrome: an attenuated form of acute pandysautonomia? Neurology 1993;43:132–7. [2] Ocon AJ, Medow MS, Taneja I, et al. Decreased upright cerebral blood flow and cerebral autoregulation in normocapnic postural tachycardia syndrome. Am J Physiol Heart Circ Physiol 2009;297:H664–73. [3] Stewart JM, Montgomery LD. Regional blood volume and peripheral blood flow in postural tachycardia syndrome. Am J Physiol Heart Circ Physiol 2004;287:H1319–H1327. [4] Raj SR, Biaggioni I, Yamhure PC, et al. Renin-aldosterone paradox and perturbed blood volume regulation underlying postural tachycardia syndrome. Circulation 2005;111:1574–82. [5] Streeten DH. Pathogenesis of hyperadrenergic orthostatic hypotension. Evidence of disordered venous innervation exclusively in the lower limbs. J Clin Invest 1990;86:1582–8. [6] Shannon JR, Flattem NL, Jordan J, et al. Orthostatic intolerance and tachycardia associated with norepinephrine-transporter deficiency. N Engl J Med 2000;342:541–9. [7] Lambert E, Eikelis N, Esler M, et al. Altered sympathetic nervous reactivity and norepinephrine transporter expression in patients with postural tachycardia syndrome. Circ Arrhythm Electrophysiol 2008;1:103–9. [8] Stewart JM, Ocon AJ, Clarke D, et al. Defects in cutaneous angiotensin-converting enzyme 2 and angiotensin-(1–7) production in postural tachycardia syndrome. Hypertension 2009;53:767–74. [9] Jacob G, Costa F, Shannon JR, et al. The neuropathic postural tachycardia syndrome. N Engl J Med 2000;343:1008–14. [10] Stewart JM, Medow MS, Glover JL, et al. Persistent splanchnic hyperemia during upright tilt in postural tachycardia syndrome. Am J Physiol Heart Circ Physiol 2006;290:H665–73.

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109 Delayed Orthostatic Hypotension Christopher H. Gibbons, Roy Freeman Orthostatic hypotension (OH), as defined by a multispecialty consensus statement, is a reduction in systolic blood pressure of 20 mmHg or diastolic blood pressure of 10 mmHg within 3 minutes of standing or upright tilt table testing to 60 degrees [1] However, not all falls in blood pressures of this magnitude occur within this time frame. A revision of this consensus statement included an orthostatic hypotension variant called delayed orthostatic hypotension in which a fall in systolic blood pressure of 20 mmHg or diastolic blood pressure of 10 mmHg occurs beyond 3 minutes of standing or upright tilt table testing [2]. Streeten first drew attention to delayed OH when he described seven patients with symptoms of orthostatic intolerance. He reported symptomatic falls in blood pressure after 13 to 30 minutes of standing [3]. The orthostasis in these patients was associated with normal or elevated catecholamine release and the fall in blood pressure was prevented through use of external compression. There are a number of physiologic challenges to the maintenance of an upright posture. Immediately upon standing there is a shift of up to 1 L of blood to the dependent vasculature of the legs and abdomen. The gravitational shifts in fluid result in decreased venous return, cardiac output and blood pressure [4]. Under normal conditions the decreased blood pressure is detected by the carotid baroreceptors which mediate an increase sympathetic outflow and vagal inhibition thus increasing heart rate, cardiac output, peripheral vascular resistance, that results in the maintenance of blood pressure and cerebral perfusion [4]. More prolonged periods of orthostatic stress result in increased pressure on the venous capacitance vessels and transudation of fluid into the dependent interstitial space. The blood volume in healthy individuals can be reduced by 1–1.5 L over a period of 10 minutes during upright standing [4,5]. Slow relaxation by dependent capacitance vessels can result in additional pooling over time. These gradual changes cause decreased venous return to the heart and reduced cardiac output. Mechanisms to maintain blood pressure with more prolonged standing include a progressive increase in muscle sympathetic nerve activity, activation of the renin-angiotensin-aldosterone system, release of vasopressin and inhibition of atrial natriuretic peptide [4,5]. Failure of these compensatory mechanisms may be implicated in delayed orthostatic hypotension.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00109-8

The patients with delayed orthostatic hypotension in the Streeten series had a variety of proposed pathogenetic mechanisms including extra-adrenal pheochromocytoma, hyperadrenergic orthostatic hypotension, hyperbradykinism, primary hyperepinephrinemia (a relative of this patient had multiple endocrine neoplasia type IIA), and hypoaldosteronism with baroreflex impairment. Streeten concluded, based on the favorable therapeutic response to external compression and supersensitivity of foot veins to norepinephrine in two patients, that excessive gravitational pooling in dependent veins was the most likely defect underlying delayed orthostatic hypotension. He hypothesized also that reduced red blood cell mass, which was present in two patients, might be an additional contributing factor. In a study of 108 patients with sustained falls in blood pressure (20 mmHg systolic or 10 mmHg diastolic), we also found significant clinical heterogeneity [6]. Less than 50% had orthostatic hypotension within the first 3 minutes of head up tilt while ~40% had orthostatic hypotension after 10 minutes of tilt-table testing. In some cases, criteria for orthostatic hypotension were met only after 20 minutes. Thus, it seems reasonable to extend the duration of evaluation to at least 20 minutes (a sensitivity of 80% in our population), to increase the diagnostic yield in patients suspected to have some form of orthostatic hypotension [6]. We found an association between severity of blood pressure fall, time to onset of blood pressure fall and other abnormalities of autonomic testing. There were greater abnormalities of both phase 2 and 4 of the Valsalva maneuver in patients with earlier and greater falls in blood pressure compared to those with later and milder drops in blood pressure. There were also more likely to be abnormalities detected in heart rate variability in patients with earlier and more severe falls in blood pressure [6]. In addition, the patients with earlier and greater drops in blood pressure tended to be older than those with later and milder drops in blood pressure. Taken together, these findings suggest that delayed orthostatic hypotension may be a mild or early manifestation of sympathetic adrenergic failure. Symptoms of orthostatic intolerance in patients with delayed orthostatic hypotension appear similar to those with orthostatic hypotension that occurs within the first 3 minutes of upright posture. Patients report postural

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lightheadedness, dizziness, weakness, neck pain, platypnea, syncope and visual blurring [3,7]. There is no relationship between the timing or severity of the drop in blood pressure and associated symptoms of orthostatic hypotension [6]. In addition, symptoms of orthostatic intolerance appear to have no relation to the magnitude of blood pressure fall and by extension the severity of the autonomic failure. A recent study highlighted two patterns of orthostatic hypotension within the first 5 minute period of head up tilt. The first pattern consisted of an immediate and sustained drop in blood pressure during the first minute of head up tilt (see Fig. 109.1A). The second pattern, which was associated with more severe autonomic deficits, consisted of a more gradual fall in blood pressure during the first two minutes that continued to progress during the 5-minute period (see Fig. 109.1B) [8]. By extending the duration of testing beyond five minutes we have highlighted an additional pattern, delayed orthostatic hypotension (see Fig. 109.1C) [6]. The frequency of delayed OH in the general population has not been identified, although it is likely to be underdiagnosed [6]. Patients with delayed OH are at risk for syncope and may be injured as a consequence of a fall, however, it does not identify which patients with late falls in blood pressure require treatment. Therapeutic interventions, when necessary, are similar to treatments as for orthostatic hypotension. The long-term consequences of delayed OH are not known.

References

120 100 80 60 40 20 0 0

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FIGURE 109.1 Variations in orthostatic hypotension. (A) There is an immediate drop in blood pressure that then stabilizes and is sustained for the duration of the head up tilt. There is no change in heart rate with the drop in blood pressure, and only minimal heart rate variability during tilt. (B) There is a more gradual but more severe and progressive decrease in blood pressure that continues over the first 5 minutes of testing. There is no change in heart rate and less heart rate variability than in (A). (C) Orthostatic hypotension is delayed beyond 3 minutes. There is a gradual fall in blood pressure after approximately 22 minutes that progresses to a maximal drop in blood pressure of 40 mmHg after 37 minutes. There is a gradual rise in heart rate during the duration of testing.

[1] Anon Position paper: Orthostatic hypotension, multiple system atrophy (the Shy Drager syndrome) and pure autonomic failure. J Auton Nerv Syst 1996;58:123–4. [2] Freeman R, Wieling W, Axelrod FB, et al. Consensus statement on the definition of orthostatic hypotension, neurally mediated syncope and the postural tachycardia syndrome. Auton Neurosci 2011. [3] Streeten DH, Anderson GHJ, 1992. Delayed orthostatic intolerance. Arch Intern Med 1992;152:1066–72. [4] Smit AA, Halliwill JR, Low PA, Wieling W. Pathophysiological basis of orthostatic hypotension in autonomic failure. J Physiol 1999;519 (Pt 1):1–10. [5] Joyner MJ, Shepherd JT, Seals DR. Sustained increases in sympathetic outflow during prolonged lower body negative pressure in humans. J Appl Physiol 1990;68:1004–9. [6] Gibbons CH, Freeman R. Delayed orthostatic hypotension: a frequent cause of orthostatic intolerance. Neurology 2006;67:28–32. [7] Streeten DH, Anderson Jr. GH. The role of delayed orthostatic hypotension in the pathogenesis of chronic fatigue. Clin Auton Res 1998;8:119–24. [8] Gehrking JA, Hines SM, Benrud-Larson LM, Opher-Gehrking TL, Low PA. What is the minimum duration of head-up tilt necessary to detect orthostatic hypotension?. Clin Auton Res 2005;15:71–5.

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110 Chronic Fatigue Syndrome and the Autonomic Nervous System Luis E. Okamoto, Satish R. Raj, Italo Biaggioni Chronic fatigue syndrome (CFS) is a complex disorder characterized by severe, disabling fatigue, associated with several nonspecific systemic symptoms. Despite 25 years of research, the cause of this syndrome remains unknown, and there are no validated diagnostic tests or biomarker available. Several case definitions of CFS have been developed as operational criteria for research. The most widely supported case definition is the 1994 definition from the US Centers for Disease Control and Prevention (Box 110.1) [1], which is now considered the standard, and it is used by many clinicians to diagnose CFS. The prevalence of CFS varies widely depending on the case definition used, the type of population and the study methods. Community and primary care based studies have reported a prevalence between 0.007% and 2.8% in the general adult population, and from 0.006% to 3.0% in the primary care or general practice. CFS affects persons of all races, ages and socioeconomic groups. Adults from 20 to 40 years are more frequently affected, and it is about twice as common in women. The clinical presentation of CFS is heterogeneous, but certain common clinical features are important to recognize. The onset of illness is often acute or subacute and, in many patients, is preceded by an infectious illness. The fatigue in CFS is characterized by persisting or relapsing chronic fatigue lasting at least 6 months in the absence of any definable medical diagnosis. It is not improved by bed rest. It may be worsened by physical or mental activity, and directly results in substantial reduction in previous degrees of activity. The fatigue is accompanied by characteristic physical, constitutional, and neuropsychological complaints (Box 110.1), but many patients may experience other symptoms that are also common in other conditions (Box 110.2). The clinical picture often fluctuates, following a pattern of relapse and remission. The diagnosis of CFS is one of exclusion. To be diagnosed with CFS, patients must undergo a thorough clinical evaluation to exclude other mental and physical causes of their symptoms, and to ascertain if their symptoms fit the case definition [1]. The etiology and pathophysiology of CFS remains unclear. Several hypotheses have been proposed: alterations in neuroendocrine and immune systems, infectious

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00110-4

illnesses, and autonomic dysfunction. None of the proposed mechanisms, however, have been sufficient to explain the symptoms of CFS in all patients. Moreover, many of these hypotheses have yielded contradictory results. It is possible; therefore, that CFS represents a common endpoint of disease resulting from multiple causes and complex interactions between these systems.

CHRONIC FATIGUE SYNDROME AND AUTONOMIC DYSFUNCTION In addition to the case-defining symptoms, many CFS patients experience a constellation of other debilitating symptoms and signs consistent with autonomic dysfunction, particularly those associated with orthostatic intolerance. These autonomic features include orthostatic symptoms and tachycardia, increased sweating, pallor, sluggish pupillary responses, gastrointestinal symptoms, and frequency micturition. The true prevalence of autonomic dysfunction in CFS remains unknown. Few studies have systematically explored widespread autonomic dysfunction in CFS, and the results have been contradictory. Newton et al. [2] observed that 75% of patients with CFS exhibited symptoms suggesting autonomic dysfunction as measured by the Composite Autonomic Symptoms Scale (COMPASS). They found that the association between autonomic dysfunction and CFS, although strong, did not cover the full range of autonomic symptoms. A particularly strong association was seen between symptoms of orthostatic intolerance and fatigue severity, suggesting a possible role of autonomic dysfunction in the pathophysiology of CFS.

CHRONIC FATIGUE SYNDROME AND ORTHOSTATIC INTOLERANCE Orthostatic intolerance (OI) is defined as the development of characteristic symptoms while standing, which are significantly improved by recumbency. It refers to a group of clinical conditions that includes neurally mediated hypotension (NMH) and postural tachycardia

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110. CHRoNIC FATIguE SyNdRomE ANd THE AuToNomIC NERvouS SySTEm

BOX 110.1

C D C C R I T E R I A F O R D I A G N O S I S O F C H R O N I C F AT I G U E S Y N D R O M E A case of chronic fatigue syndrome must satisfy two criteria: 1. Have clinically evaluated, unexplained, persistent or relapsing fatigue for at least 6 months, that is of new or definite onset; is not the result of ongoing exertion; is not alleviated by rest; and results in substantial reduction of previous levels of occupational, educational, social, or personal activities; and 2. Four or more of the following symptoms, concurrently present for  6 consecutive months of illness that not predate the fatigue: l

l l l l l l l

Sore throat Tender cervical or axillary lymph nodes Muscle pain Multi-join pain without swelling or redness Headaches of a new type, pattern, or severity Unrefreshing sleep Post-exertional malaise lasting  24 hours.

Adapted from: Fukuda K, et al. Ann Intern Med 1994;121:953.

Self-reported impairment in short-term memory or concentration

BOX 110.2

A D D I T I O N A L S Y M P T O M S A S S O C I AT E D W I T H C H R O N I C F AT I G U E S Y N D R O M E Gastrointestinal disturbances (constipation, diarrhea) Visual disturbances (blurring, light sensitivity, eye pain) Chills and night sweats Difficulty maintaining upright posture, dizziness, balance problems or fainting

syndrome (POTS). Orthostatic intolerance has been associated with CFS in both adults and children [3–8]. The connection between OI and CFS was first proposed in 1992 by Streeten and Anderson [8]; who suggested that fatigue and exhaustion might be caused, in some patients with CFS, by failure to maintain blood pressure in the erect posture. Rowe et al. [4] were first to study systematically the relationship between OI and CFS proposed by Streeten and Anderson. They suggested that neurally mediated hypotension might be an unrecognized cause of chronic fatigue in patients with CFS. Since then, several studies have attempted to document the presence of OI in patients with CFS. Many, but not all [9], have demonstrated a frequent occurrence (up to 97%) of OI in CFS patients [4,7,10–12]. The prevalence, however, greatly differed among studies. Possible explanations for these discrepancies include differences in tilt-test protocols, clinical heterogeneity of CFS patients, and lack of delineation of selection criteria for subjects and controls, including age and gender. To date, no epidemiological studies have defined the prevalence of OI in CFS.

Cognitive impairment or brain fog Allergies and sensitivities to foods, odors, chemicals, medications Psychological problems (irritability, mood swings, anxiety, panic attacks).

Many clinical features of OI are common in CFS. Both conditions often have an acute or subacute onset of symptoms, which may be preceded by a viral-like infection. The age of presentation is similar in OI and CFS, and women are disproportionately affected. Orthostatic symptoms experienced by individuals with OI, such as lightheadedness, dizziness, nausea, fatigue, tremors, palpitations, chest discomfort, abdominal pain, headache, visual disturbances, sweating, neurocognitive impairment, anxiety and syncope; are also observed in patients with CFS. In both conditions, symptomatology can be exacerbated by physiological stressors such as exercise, heat exposure or meals. Some patients may experience prolonged fatigue following an episode of lightheadedness or fainting, which can last from hours to days. Both conditions follow a cyclical course of symptoms. Moreover, some early studies in small populations of CFS patients have shown that effective treatment of OI resulted in substantial improvement of their chronic fatigue. These studies, however, were not placebo-controlled, blinded or randomized [4,8,10], and were not confirmed by others.

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CoNCluSIoNS

POSTURAL TACHYCARDIA SYNDROME (POTS) Postural tachycardia syndrome (POTS) is a relatively common and heterogeneous condition. It is defined as the development of orthostatic symptoms associated with an increase in heart rate from the supine to upright position of  30 bpm, or to  120 bpm on standing. Several studies have shown an overlap between CFS and POTS. The first observation of POTS in patients with CFS was reported by De Lorenzo et al. [13]. They found postural tachycardia and orthostatic symptoms in three of five CFS patients during head-up tilt. This association has also been confirmed by others [3,6,7,11]. In these studies, the prevalence of POTS in CFS patients has ranged from 19% [11] to 70% [6]. On the other hand, studies in cohorts of patients selected for POTS have also shown a high prevalence of chronic fatigue (up to 77%) [14]. The prevalence of subjects fulfilling the Fukuda criteria for CFS in these cohorts has ranged from 17 to 23% [14,15]. It is not known, however, whether the clinical and autonomic features of POTS patients with and without CFS differ. One study in adolescents found no differences in the clinical features of orthostatic tachycardia and autonomic function between POTS with and without CFS. Both conditions showed a similar degree of vagal withdrawal with intact vasoactive baroreflexes and sympathoexitation [7].

not completely understood. A detailed discussion of the current theories concerning their pathogenesis is reviewed elsewhere (see Chapters 70 and 107). Several studies have addressed the pathophysiology of OI in patients with CFS. Streeten and colleagues suggested that delayed orthostatic hypotension and/or tachycardia are caused by excessive gravitational venous pooling into the lower body, which could be corrected by external lower-body compression [12]. In a recent study, Hurwitz et al. [16] observed that CFS patients had reduced total blood volume, plasma volume and red cell mass volume when compared to healthy controls. Moreover, they suggested that this hypovolemic state was the cause of the diminished cardiac volume observed in severe CFS patients. Although the authors did not assess autonomic function and, therefore, it is not known how many of these subjects had OI; these findings overlap with those reported in POTS (see Chapter 107). In addition, minor abnormalities of cardiovascular autonomic function tests have been observed in some studies [3,7,9], but there is no evidence of widespread dysautonomia. Cardiovascular deconditioning is often present in these patients. However, it seems to be a contributor rather than the cause of OI in CFS. It is not known how these pathophysiologies contribute to the clinical features of CFS, or which of them is more relevant in CFS patients with OI.

CONCLUSIONS NEURALLY MEDIATED HYPOTENSION Neurally mediated hypotension and neurally mediated syncope have been associated with chronic fatigue and CFS by several investigators. Rowe and colleagues [4] were first to demonstrate this association when they reported a series of seven adolescents with chronic fatigue, each of whom had evidence of neurally mediated hypotension. They suggested that a predisposition to neurally mediated hypotension may underlie some of the symptoms of CFS, and found that treatment of NMH resulted in substantial improvement of chronic fatigue [4,10]. Since then, other investigators have confirmed that there is a substantial overlap in the clinical features of CFS and neurally mediated hypotension [3,6,11]. In these studies, the incidence of NMH in patients with CFS ranged from 8% [6] to 96% [10]. NMH and POTS are not mutually exclusive diagnoses, the two conditions often are found together.

PATHOPHYSIOLOGY The different patterns of orthostatic intolerance in CFS suggest that CFS is heterogeneous in presentation and pathophysiological mechanisms. It is likely that there is no unique abnormality underlying the orthostatic intolerance in CFS. The pathophysiology of NMH and POTS is

CFS is a fairly common, incompletely understood disorder, which overlaps clinically with autonomic dysfunction, frequently manifested as orthostatic intolerance. The basis of this relationship is still unknown. Nonetheless, it seems likely that autonomic dysfunction plays a role in its pathophysiology in some patients. It is more difficult to determine if the autonomic dysfunction is the primary event, or if it is an appropriate (or exaggerated) response to other pathyphysiological processes (e.g., a reduction in blood volume or deconditioning). Further research is required to elucidate the pathophysiology of autonomic dysfunction in CFS, and thereby contribute to improve our understanding of this disorder.

References [1] Fukuda K, Straus SE, Hickie I, Sharpe MC, Dobbins JG, Komaroff A. The chronic fatigue syndrome: a comprehensive approach to its definition and study. International Chronic Fatigue Syndrome Study Group. Ann Intern Med 1994;121:953–9. [2] Newton JL, Okonkwo O, Sutcliffe K, Seth A, Shin J, Jones DE. Symptoms of autonomic dysfunction in chronic fatigue syndrome. QJM 2007;100:519–26. [3] Freeman R, Komaroff AL. Does the chronic fatigue syndrome involve the autonomic nervous system? Am J Med 1997;102:357–64. [4] Rowe PC, Bou-Holaigah I, Kan JS, Calkins H. Is neurally mediated hypotension an unrecognised cause of chronic fatigue? Lancet 1995;345:623–4. [5] Schondorf R, Freeman R. The importance of orthostatic intolerance in the chronic fatigue syndrome. Am J Med Sci 1999;317:117–23.

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[6] Stewart JM, Gewitz MH, Weldon A, Munoz J. Patterns of orthostatic intolerance: the orthostatic tachycardia syndrome and adolescent chronic fatigue. J Pediatr 1999;135:218–25. [7] Stewart JM. Autonomic nervous system dysfunction in adolescents with postural orthostatic tachycardia syndrome and chronic fatigue syndrome is characterized by attenuated vagal baroreflex and potentiated sympathetic vasomotion. Pediatr Res 2000;48:218–26. [8] Streeten DH, Anderson GH. Delayed orthostatic intolerance. Arch Intern Med 1992;152:1066–72. [9] Soetekouw PM, Lenders JW, Bleijenberg G, Thien T, Van Der Meer JW. Autonomic function in patients with chronic fatigue syndrome. Clin Auton Res 1999;9:334–40. [10] Bou-Holaigah I, Rowe PC, Kan J, Calkins H. The relationship between neurally mediated hypotension and the chronic fatigue syndrome. JAMA 1995;274:961–7. [11] Schondorf R, Benoit J, Wein T, Phaneuf D. Orthostatic intolerance in the chronic fatigue syndrome. J Auton Nerv Syst 1999;75:192–201.

[12] Streeten DH, Thomas D, Bell DS. The roles of orthostatic hypotension, orthostatic tachycardia, and subnormal erythrocyte volume in the pathogenesis of the chronic fatigue syndrome. Am J Med Sci 2000;320:1–8. [13] De Lorenzo F, Hargreaves J, Kakkar VV. Possible relationship between chronic fatigue and postural tachycardia syndromes. Clin Auton Res 1996;6:263–4. [14] Jacob G, Shannon JR, Black B, Biaggioni I, Mosqueda-Garcia R, Robertson RM, et al. Effects of volume loading and pressor agents in idiopathic orthostatic tachycardia. Circulation 1997;96:575. [15] Jacob G, Costa F, Shannon JR, Robertson RM, Wathen M, Stein M, et al. The neuropathic postural tachycardia syndrome. N Engl J Med 2000;343:1008–14. [16] Hurwitz BE, Coryell VT, Parker M, Martin P, Laperriere A, Klimas NG, et al. Chronic fatigue syndrome: illness severity, sedentary lifestyle, blood volume and evidence of diminished cardiac function. Clin Sci 2010;118:125–35.

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111 Joint Hypermobility Syndrome and Dysautonomia Giris Jacob, Blair P. Grubb INTRODUCTION Joint hypermobility (JH), also known as benign joint hypermobility syndrome (BJHS), is a common heritable connective tissue disorder. Its main clinical feature is joint laxity, which causes articular dislocations, subluxations and arthralgia, in the absence of evidence for any rheumatologic disorder [1]. Extra-articular manifestations of JH involve mild skin hyperextensibility, pelvic floor insufficiency with uterine and rectal prolapse, ocular (lid laxity), defective proprioception, chronic pain and autonomic dysfunction [2].

EPIDEMIOLOGY AND PATHOPHYSIOLOGY Physicians may be unaware of the prevalence of the JH. Asymptomatic patients are rarely diagnosed and those with symptoms are commonly misdiagnosed. Another overlapping syndrome that has similar clinical features, hypermobile Ehlers–Danlos syndrome (formerly hypermobile EDS III), is frequently difficult to distinguish from JHS. The main clinical mark of EDS III, other than joint hypermobility, is skin extendibility and widened atrophic scarring, which may, to some extent, be present in patients with JHS. Therefore, in the absence of genetic typing, biomarkers and pathognomotic features, both syndromes could be considered interchangeable [3]. While there has been considerable progress in identifying the genetic basis of related collagen disorders (such as osteogenesis imperfecta, Marfan’s and Ehlers–Danlos syndrome), the genetic basis of JH remains poorly understood. Recently, it has been found that some patients with JHS have a mutation in the non-collagenous molecule tenascin-X, a large extracellular matrix glycoprotein (as do some patients with hypermobile EDS). However, this accounts for a small portion of JH patients, suggesting that other many genetic factors could affect the pathophysiology of the syndrome. For instance, other considerations, such as environmental influences, muscle composition and bone structure may predispose to the presence of JH. Moreover, some rheumatologists view JH as a variant of

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normality on the edge of a Gaussian distribution of the normal joint elasticity. Accordingly, the exact prevalence is not known. However, based on clinical diagnostic criteria, JH appears not uncommon in countries such Iraq, Nigeria and some Asian populations (with an estimated incidence of 10–20%). In the occidental countries it is estimated that ~5% of the population are affected, with evident ethnic influences and female to male ratio 3:1 [4].

CLINICAL MANIFESTATION AND DIAGNOSIS The diagnosis of JH is based on a combination of symptoms, historical features and physical findings. Joint related findings, such as dislocation, arthralgia, and premature osteoarthritis are common. These often occur in conjunction with extra-articular symptoms such as hyperextensibility of skin, atrophic dermal scarring, varicose veins, uterine and rectal prolapse, as well as signs and symptoms of autonomic dysfunction of the cardiovascular and gastrointestinal systems. Due to lack of a specific biomarker, the physical examination remains the cornerstone for JH diagnosis. The current diagnostic criteria are elaborated in the Brighton revised criteria [5]. These include the previous Beighton Diagnostic Score that employed a 9-point system (with 4/9 points required for a diagnosis JH) as shown in Boxes 111.1 and 111.2. While joint findings are the characteristic feature of JH, it is often non-articular symptoms that dominate the clinical picture. In addition to musculoskeletal and cutaneous findings in JH, several other physiologic abnormalities may occur. Problems such as chronic pain, fibromyalgia, headache, chronic fatigue and autonomic dysfunction are frequently reported.

AUTONOMIC DYSFUNCTION IN JHS A constellation of many symptoms, such as dizziness, palpitation and chest discomfort suggest the presence

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of disturbed autonomic function in patients with JH. Historically, these symptoms were thought to be related to the presence of mitral valve prolapse (MVP syndrome) in some JH patients [6]. Later, increasingly sophisticated echocardiography imaging along with more rigorous diagnostic criteria for MVP demonstrated that it was present less frequently in JH than previously thought. Subsequently a report by Row and colleagues found that dysautonomia was a prominent feature in children who suffered from JH [7]. Moreover, epidemiological data have demonstrated that some of the most common functional syndromes that have related to autonomic dysfunction, such as fibromyalgia and chronic fatigue syndromes, are frequently reported to occur in association with JH syndrome.

BOX 111.1

THE BEIGHTON SCORING SYSTEM FOR JOINT HYPERMOBILITY

Scoring One Point for Each Side l l

l l

Passive dorsiflexion of the fifth MCP to 90o Apposition of the thumb to the flexor aspect of the forearm Hyperextension of the elbow beyond 90o Hyperextension of the knee beyond 90o.

2 2 2 2

Scoring 1 point l

Forward trunk flexion placing hands flat on floor with knees extended.

1

Recently, a study was conducted in a large group of patients with JH, selected using stringent criteria (Beighton score, median 6.5) and compared to healthy controls (Beighton score 2, median 0), with the aim of estimating the frequency and pathophysiology of autonomic disorders in JH. Symptoms related to autonomic nervous system dysfunction were reported by all patients with JH. Cerebral hypoperfusion, cardiorespiratory and gastrointestinal related symptoms were definitively higher in patients with JH compared to healthy controls. Hypermobile patients reported a higher rate of presyncope (83% vs. 23%), syncope (56% vs. 10%), fatigue, impaired concentration, palpitations (90% vs. 27%), chest discomfort (65% vs. 3%), nausea, stomach discomfort, diarrhea, heat intolerance and subjective orthostatic intolerance (56% vs. 10%). Furthermore, the objective mean time standing was significantly lower in these patients (14 vs. 19 minutes). Patients with JH showed a mean drop in systolic BP of 9 mmHg without significant change in diastolic BP after 3 minutes of standing. Twenty percent of patients and none of the controls had mild orthostatic hypotension (a drop of SBP/DBP 20/10 mmHg). The mean increase in heart rate after standing for at least 3 minutes was higher in patients compared to controls (22 vs. 15 bpm). Fifteen percent of hypermobile patients and none of the controls fulfilled the stringent criteria of postural tachycardia syndrome (POTS  five characteristic orthostatic symptoms for at least 6 months, and upright increase in HR 30 bpm after 5 minutes of standing) [8]. The high frequency of syncope (vaso-vagal type), and POTS in patients with JHS was also reported by other authors [9].

BOX 111.2

THE BRIGHTON CRITERIA FOR DIAGNOSIS OF JOINT HYPERMOBILITY

Major Criteria l

l

l

Beighton score of 4/9 or greater (either currently or historically) [see Box 111.1] Arthralgia for longer than 3 months in 4 or more joints.

Minor Criteria l l

l

l

l

l

l

Beighton score of 1–3/9 (0–3 if aged 50) Arthralgia (3 mo) in 1–3 joints, or back pain (3 mo), spondylosis, spondylolysis/spondylolisthesis Dislocation/subluxation in more than 1 joint, or in 1 joint or more on more than 1 occasion Soft tissue rheumatism 3 lesions (e.g. epicondylitis, tensosynovitis, bursitis)

Marfanoid habitus (tall, slim, span : height ratio 1.03, upper segment : lower segment ratio 0.89, arachnodactily [Steinberg/wrist sign]) Abnormal skin: striae, hyperextensibility, thin skin or papyraceous scarring Eye signs: drooping eyelids or myopia or antimongoloid slant. Varicose veins or hernia or uterine/rectal prolapsed.

JH Diagnosis Requires l

l

Two major criteria or one major  two minor criteria or four minor criteria or two minor criteria and unequivocally affected first-degree relative JH is excluded by presence of Marfan or Ehler–Danlos syndromes (as defined by the Berlin nosology).

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mAnAgEmEnT

Autonomic function test in these patients reveals that vagal cardiovascular control is within the normal range including the vagal baroreflex arm. On the other hand, cardiovascular sympathetic control showed significant abnormalities. Hypermobile patients in comparison to controls show exaggerated decrease and augmented increase in systolic BP during hyperventilation and cold pressor test, respectively. Valsalva maneuver phase IIa demonstrated exaggerated decrease in systolic BP in patients with JH (–19 vs. –7 mmHg). Furthermore, hypermobile patients were more sensitive to the heart rate increase effect of isoproterenol (β-adrenoreceptor agonist) and similarly showed hyper-responsiveness to the phenylephrine (α1adrenorecptor agonist) ability to increase BP [8]. These finding are compatible with the data previously described by different authors in patients with idiopathic POTS [10]. We believe that the association between JH and autonomic dysfunction makes biological sense. First of all, the phenomena of orthostatic tachycardia and mild hypotension can be prevented by applying military anti-shock trousers. This is consistent with the hypothesis that excessive venous pooling is an important contributor to the pathophysiology of postural tachycardia and hypotension [11]. In the standing position, patients with excessively compliant connective tissue would be expected to have an increased distensibility of the dependent veins in response to an ordinary degree of hydrostatic pressure. This would predispose those with JH (and mainly with EDS) to excessive pooling of blood, and thus to orthostatic intolerance. Second, affected proprioception, abnormal electromyographic recordings (EMG) [12], chronic pain syndrome (e.g. fibromyalgia), and the above described cardiovascular sympathetic abnormalities indicate that the neurophysiologic abnormalities exist in patients with JH. Accordingly, secondary (neuropathic) POTS could be part of JH, but this needs further confirmation [13]. Finally, some patients with JHS are limited in their daily physical activity due to arthralgia, chronic pain syndrome, and chronic fatigue syndrome causing deconditioning. It may cause aberrant cardiovascular regulation, hypovolemia, baroreflex sensitivity changes and adrenoreceptor hyperresponsiviness, as reported above. According to some authors cardiovascular deconditioning could be behind the pathophysiology of POTS, at least in a subgroup of patients [14]. We summarize that patients with JH have a higher incidence of autonomic dysfunction related syndromes compared to subjects without JH. Syncope, POTS, and mild orthostatic hypotension are now being frequently considered among the frequent extra-articular manifestations of JH. These dysautonomias seem to have multiple pathophysiologies that ought to be tested in future studies as stated above. However, the frequency of JH in patients with “primary” orthostatic syndromes (POTS and other autonomic dysfunction) remains unknown. Therefore, we suggest that each patient with orthostatic intolerance be tested for JH by a simple physical examination for fulfilling the clinical Brighton criteria.

MANAGEMENT The hypermobile syndrome is widely neglected by rheumatologists and therefore any qualified physician may encounter it. Management should involve a qualified physiotherapist adapted for the treatment of joint laxity and fragile tissue. Specifically, peri-articular muscle reinforcement exercises are required in order to overcome kinesiphobia (fear of pain) and eventually improve the joint proprioception acuity. As we mentioned before, patients with JH present mild-to-moderate orthostatic intolerance syndromes that are easily managed with high salt and water intake together with the adherent physiotherapy program. In refractory cases low doses of midodrine and mineralcorticoids are sufficient to control orthostatic symptoms. Chronic pain syndrome or fibromyalgia syndrome are possibly manageable by centrally acting analgesics and eventually with anxiolytic medications.

References [1] Ross J, Grahame R. Joint hypermobility syndrome. BMJ 2011;342:c7167. [2] Mishra MB, Ryan P, Atkinson P, et al. Extra-articular features of benign joint hypermobility syndrome. Br J Rheumatol 1996;35:861–6. [3] Malfait F, Wenstrup RJ, De PA. Clinical and genetic aspects of Ehlers–Danlos syndrome, classic type. Genet Med 2010;12:597–605. [4] Malfait F, Hakim AJ, De PA, Grahame R. The genetic basis of the joint hypermobility syndromes. Rheumatology (Oxford) 2006;45:502–7. [5] Grahame R, Bird HA, Child A. The revised (Brighton 1998) criteria for the diagnosis of benign joint hypermobility syndrome (BJHS). J Rheumatol 2000;27:1777–9. [6] Grahame R, Child A. Mitral valve prolapse. Br Med J (Clin Res Ed) 1984;289:317. [7] Rowe PC, Barron DF, Calkins H, Maumenee IH, Tong PY, Geraghty MT. Orthostatic intolerance and chronic fatigue syndrome associated with Ehlers–Danlos syndrome. J Pediatr 1999;135:494–9. [8] Gazit Y, Nahir AM, Grahame R, Jacob G. Dysautonomia in the joint hypermobility syndrome. Am J Med 2003;115:33–40. [9] Kanjwal K, Saeed B, Karabin B, Kanjwal Y, Grubb BP. Comparative clinical profile of postural orthostatic tachycardia patients with and without joint hypermobility syndrome. Indian Pacing Electrophysiol J 2010;10:173–8. [10] Jacob G, Shannon JR, Costa F, et al. Abnormal norepinephrine clearance and adrenergic receptor sensitivity in idiopathic orthostatic intolerance. Circulation 1999;99:1706–12. [11] Streeten DH, Scullard TF. Excessive gravitational blood pooling caused by impaired venous tone is the predominant noncardiac mechanism of orthostatic intolerance. Clin Sci (Lond) 1996;90:277–85. [12] Ferrell WR, Tennant N, Baxendale RH, Kusel M, Sturrock RD. Musculoskeletal reflex function in the joint hypermobility syndrome. Arthritis Rheum 2007;57:1329–33. [13] Jacob G, Costa F, Shannon JR, et al. The neuropathic postural tachycardia syndrome. N Engl J Med 2000;343:1008–14. [14] Levine BD, Zuckerman JH, Pawelczyk JA. Cardiac atrophy after bed-rest deconditioning: a nonneural mechanism for orthostatic intolerance. Circulation 1997;96:517–25.

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112 Neuroleptic Malignant Syndrome Fenna T. Phibbs, P. David Charles INTRODUCTION Neuroleptic malignant syndrome (NMS) is a rare and potentially fatal syndrome of hyperthermia, rigidity, autonomic instability, and mental status derangement [1]. It is likely an idiosyncratic reaction to drugs that alter central nervous system dopaminergic pathways. Neuroleptics, including dopamine antagonist commonly prescribed for psychiatric disorders, are the most common inciting agents. Older estimates of the incidence of NMS in patients treated with neuroleptics reached 2.2% but more current estimates indicated a far lower incidence of 0.01 to 0.02%, with a mortality rate of 10% [2]. A similar syndrome may rarely occur in the setting of sudden withdrawal of dopamine agonists used in the treatment of Parkinson’s disease.

MEDICATIONS AND RISK FACTORS

CLINICAL FEATURES The clinical features of NMSD are distinctive but the cardinal findings need not occur in every patient. The considerable list of potential symptoms (Box 112.1) can be grouped into the four general areas of hyperthermia, muscular rigidity, mental status changes, and autonomic dysfunction. Hyperthermia is present in most all cases and can exceed 103.0°F. Muscular rigidity is severe and can be associated with tremor, dystonia and bradykinesia; all three features are caused by extrapyramidal dysfunction. Passive movement of the limbs in all directions (lead pipe rigidity) is resisted. Mental status changes include confusion, delirium, speech disorders, and altered states of consciousness. Autonomic dysfunction is usually characterized by rapid fluctuations in blood pressure and heart rate. Other autonomic features include sialorrhea, incontinence, and dysphagia. Tachypnea is probably caused by a combination of autonomic instability, muscular rigidity of the chest wall, and aspiration. The laboratory findings of NMS are useful in supporting the diagnosis. Creatine kinase (CK) may exceed 10,000 IU due to sustained muscular contraction causing muscle fiber necrosis. Other less specific features are a leukocytosis, myoglobinuria, and elevated serum concentrations of

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00112-8

transaminases, lactic acid dehydrogenase, aldolase, and peripheral catecholamines. The primary morbidity and mortality associated with NMS are irreversible brain injury from hyperthermia and renal failure from myoglobinuria secondary to rigidityinduced skeletal muscle necrosis. Exacerbating this are aspiration pneumonia, myocardial infarction, disseminated intravascular coagulation, and metabolic and electrolyte derangements. In 2000 the Diagnostic and Statistical Manual of Mental Disorders, IV, text revision (DSM-IV-TR) [3] recommended diagnostic criteria for NMS (Box 112.2) with the intention of creating more consistency in diagnosis and facilitating early recognition and intervention, thus reducing morbidity and mortality.

Neuroleptics are commonly prescribed drugs in the United States. Phenothiazines, thiothixine, and butyrophenones are the agents most commonly implicated in causing NMS, with haloperidol being the most commonly associated, most likely because it was one of the most commonly prescribed neuroleptics [4]. The atypical neuroleptics are felt to have a lower risk of extrapyramidal effects and NMS, however there are reports associated with these agents also [5]. Intramuscular injection of depot preparations of neuroleptics may increase the risk of NMS and definitely prolong recovery because prompt drug withdrawal is not possible. The expanding list of implicated drugs includes those with dopamine-blocking properties such as amoxapine, tetrabenazine, metoclopramide, monoamine oxidase inhibitors, and tricyclic antidepressants, and other medications with differing mechanisms of action, including lithium, clomipramine, and selective serotonin reuptake inhibitors (SSRIs). Abrupt withdrawal of levodopa, amantadine, and baclofen has also been reported to cause a similar syndrome. The proposed common mechanism is development of a relative hypodopaminergic state (Table 112.1) [6]. Individuals at increased risk for NMS cannot be identified prior to initiating therapy. The onset of symptoms is most often within the first 30 days of starting treatment,

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BOX 112.1

CLINICAL FINDINGS IN NEUROLEPTIC MALIGNANT SYNDROME l

l

l

l

Hyperthermia l Often 103°F, but can be as low as 99°F Rigidity l Lead pipe in nature Other extrapyramidal findings l Tremor l Bradykinesia l Dystonic posturing Mental status changes l Confusion l Obtundation l Mutism

l

l

Autonomic instability l Blood pressure lability l Tachycardia l Tachypnea l Sialorrhea l Incontinence l Diaphoresis Laboratory l CPK often 10,000 l Leukocytosis l Lactic acid dehydrogenase elevation l Aldolase elevation l Transaminase elevation.

BOX 112.2

RESEARCH CRITERIA FOR NEUROLEPTIC MALIGNANT SYNDROME A. The development of severe muscle rigidity and elevated temperature associated with the use of neuroleptic medication. B. Two (or more) of the following: 1. diaphoresis 2. dysphagia 3. tremor 4. incontinence 5. changes in level of consciousness ranging from confusion to coma 6. mutism 7. tachycardia 8. elevated or labile blood pressure

9. leukocytosis 10. laboratory evidence of muscle injury (e.g., elevated creatine phosphokinase [CPK]) C. The symptoms in Criteria A and B are not due to another substance (e.g., phencyclidine) or a neurologic or other general medical condition (e.g., viral encephalitis). D. The symptoms in Criteria A and B are not better accounted for by a mental disorder (e.g., mood disorder with catatonic features). From American Psychiatric Association. Diagnostic and Statistical Manual for Mental Disorders, 4th edn.–TR (2000). APA, Washington, DC, 798.

TABLE 112.1 Potential Precipitants: Neuroleptic Malignant Syndrome Precipitants

Examples

TYPICAL NEUROLEPTICS Phenothiazines Dibenzoxazepines Butyrophenones Dihydroindolones Thioxanthenes Dibenzoapines

Fluphenazine, chlorpromazine, thioridazine, promethazine, prochlorperazine, trifluoperazine Pimozide Haloperidol Molindone Thiothixene Loxapine

ATYPICAL NEUROLEPTICS Antiemetics Tricyclic antidepressants Benzodiazepines (in overdose and in pharmacy) Withdrawal of dopamine agonist Polypharmacy (in combination with neuroleptics)

Clozapine, risperidone, olanzapine, quetiapine, aripiprazole Metoclopramide Amitriptyline, imipramine, etc. Diazepam, lorazepam, etc. L-DOPA, pergolide, bromocriptine, amantadine, etc. Alcohol, lithium, cimetidine

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TREATMENT

with two-thirds presenting in the first week of treatment [3]. Rapid neuroleptic dose escalation, dehydration, psychomotor agitation, catatonia, and underlying organic brain disease probably increase the incidence of NMS in at-risk individuals. Human immunodeficiency virus (HIV) infection my increase risk, particularly if advanced to acquired immune deficiency syndrome (AIDS) with dementia [7]. There is evidence of a possible genetic predisposition to NMS in individuals with either the TaqI A or 141C Ins/Del polymorphism of the DRD2 gene [8,9]. Other gene polymorphisms have yet to be implicated in the pathogenesis of the syndrome. NMS recurs in onethird of patients when neuroleptics are reintroduced after recovery from the initial episode. In situations where the need for neuroleptic therapy outweighs the risk, the chance of a second NMS episode can be reduced by waiting at least 2 weeks after resolution of symptoms and then using the lowest possible dose of a low-potency agent [10].

DIFFERENTIAL DIAGNOSIS The differential diagnosis of NMS includes other syndromes with hyperthermia as a prominent feature, fever complicating Parkinson’s disease or other extrapyramidal syndromes, and lethal catatonia. Malignant hyperthermia and NMS have similar clinical features but are distinguishable because malignant hyperthermia is inherited as an autosomal dominant condition and is only triggered by the administration of anesthetic agents [6]. Heat stroke shares some features with NMS but lacks rigidity and sweating and is characterized by an abrupt onset following physical exercise or exposure to a high ambient temperature. Neuroleptics increase the risk of heat stroke; this may confuse diagnosis in individuals whose history is compatible with both disorders [10]. The clinical features of NMS can be mimicked in patients with Parkinson’s disease whose rigidity worsens at the time of concurrent fever or infection. The same is true for other neurodegenerative disorders of the extrapyramidal system. Common infectious and metabolic disorders must be ruled out prior to making the diagnosis of NMS. Lethal catatonia is a rare disorder of psychotic patients known years before the development of neuroleptics. Its clinical features are identical to those of NMS and the two conditions are only distinguished by the prior use of neuroleptics [6]

PATHOGENESIS The precise pathogenesis of NMS is not fully understood. The most commonly accepted theory is that central dopaminergic hypoactivity acts as a trigger for the condition. This supports a hypothesis that dopamine receptor blockade in the basal ganglia and hypothalamus results from treatment with neuroleptics [10]. Disruption of the dopaminergic pathways of the hypothalamus and basal

ganglia leads to hyperthermia and rigidity. The experimental infusion of a dopamine agonist on the thermoregulatory center of the hypothalamus causes a dose-dependent decrease in body temperature. Basal ganglia dysfunction commonly produces tremor and other extrapyramidal symptoms in addition to rigidity. The autonomic instability combined with sustained muscle contraction and impaired heat dissipation exacerbates hyperthermia. The cerebrospinal fluid of patients with NMS shows a reduction of homovanillic acid, the major metabolite of dopamine, supporting the dopamine-blocking hypothesis, but results are varied [7]. Sympathetic nervous system hyperactivity has been recorded in the NMS active phase implicating central noradrenergic activity dysregulation in the pathogenesis of NMS [11]. Other factors must be involved in the pathogenesis of NMS because its occurrence is rare, while the drugs known to induce the syndrome are prescribed to millions of people each year. Furthermore, the reintroduction of neuroleptic agents in patients with a history of NMS does not always cause a recurrence.

TREATMENT Patients with NMS should be treated in an intensive care unit. Treatment begins with the immediate withdrawal of the offending neuroleptic or the reintroduction of the antiparkinson agent that was discontinued. The average time for recovery from NMS is 10 days. Depot preparations may extend resolution by weeks depending upon the dose and timing of symptom onset [7]. Therapy with dopamine agonists, such as bromocriptine, reduces the recovery time of NMS by restoring the dopaminergic balance in the preoptic area of the hypothalamus and basal ganglia. [10]. Levodopa/carbidopa has also been used successfully to reverse the hypo-dopaminergic state and to reverse hyperthermia. Dantrolene, a skeletal muscle relaxant, reduces the muscle rigidity of malignant hyperthermia and is also useful in NMS [10]. Early therapy with bromocriptine and dantroline reduces the occurrence of aspiration pneumonia, one of the leading causes of death in NMS. Benzodiazepines are also useful to treat the agitation rigidity [7]. Early diagnoses and prompt supportive measures in combination with drug therapy significantly reduced mortality. Cooling blankets are effective in combating hyperthermia and vigorous fluid replacement is essential to prevent dehydration. Aspiration is a common complication of NMS because of decreased chest wall compliance and depressed level of consciousness. Therefore, mechanical ventilation when indicated combined with aggressive pulmonary toilet and appropriate antibiotic therapy in the event of pneumonia, is helpful. Renal failure, resulting from muscle breakdown, is the most common and severe complication of NMS. Fluid replacement and hemodialysis are the primary treatment modalities. The mortality from NMS increases to 50% if renal failure develops [4].

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Electroconvulsive therapy (ECT) is a last resort when lethal catatonia enters the differential diagnosis [7]. The two syndromes are clinically indistinguishable, even when treated with benzodiazepines, and lethal catatonia does not respond to dopaminergic treatment as does NMS.

References [1] Delay J, Deniker P. Drug-induced extrapyramidal syndromes, in Handbook of clinical neurology. In: Vinken EPJ, Bruyn GW, editors. Disease of the basal ganglia. Amsterdam: North-Holland; 1968. p. 248–66. [2] Stubner S, et al. Severe and uncommon involuntary movement disorders due to psychotropic drugs. Pharmacopsychiatry 2004;37(Suppl 1):S54–64. [3] American Psychiatric Association: Medication-induced movement disorders: neuroleptic malignant syndrome. In Diagnosis and Statistical Manual of Mental Disorders: DSM-IV-TR. 2000: Washington D.C. p. 795–98.

[4] Adnet P, Lestavel P, Krivosic-Horber R. Neuroleptic malignant syndrome. Br J Anaesth 2000;85(1):129–35. [5] Ananth J, et al. Neuroleptic malignant syndrome and atypical antipsychotic drugs. J Clin Psychiatry 2004;65(4):464–70. [6] Margetic B, Aukst-Margetic B. Neuroleptic malignant syndrome and its controversies. Pharmacoepidemiol Drug Saf 2010;19(5):429–35. [7] Factor SA. Neuroleptic malignant syndrome. In: Factor SA, Lang Anthony E, Weiner William J, editors. Drug Induced Movement Disorders. Malden, Massachusetts: Blackwell Publishing; 2005. p. 174–212. [8] Suzuki A, et al. Association of the TaqI A polymorphism of the dopamine D(2) receptor gene with predisposition to neuroleptic malignant syndrome. Am J Psychiatry 2001;158(10):1714–6. [9] Kishida I, et al. Association in Japanese patients between neuroleptic malignant syndrome and functional polymorphisms of the dopamine D(2) receptor gene. Mol Psychiatry 2004;9(3):293–8. [10] Strawn JR, Keck Jr. PE, Caroff SN. Neuroleptic malignant syndrome. Am J Psychiatry 2007;164(6):870–6. [11] Gurrera RJ. Sympathoadrenal hyperactivity and the etiology of neuroleptic malignant syndrome. Am J Psychiatry 1999;156(2):169–80.

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113 Migraine and the Autonomic Nervous System* Pietro Cortelli Migraine is a syndrome of recurrent headaches manifesting in attacks lasting 4 to 72 hours, with typical unilateral localization, pulsating quality, moderate to severe intensity of the pain, aggravation by or avoidance of routine physical activities, and association with nausea and/or vomiting and photo- and phono-phobia [1]. The migraine attack consists of several phases: prodromal symptoms, aura, headache phase with pain and nausea/vomiting, resolution, and recovery. Two main types of migraine are recognized, migraine with and without aura. Migraine is highly prevalent in the general, especially female population, and carries a substantial genetic predisposition. Migraine is no longer thought to be caused by a primary vascular event [2]. It involves integrated brain mechanisms among a number of central nervous system (CNS) structures (cortex, brainstem, trigeminal system, meninges) and has a complex pathophysiology. It is generally recognized that migraine arises from a primary brain dysfunction that leads to activation and sensitization of the trigeminal system.

FUNCTIONAL ANATOMY OF MIGRAINE The cerebral vessels and the meninges represent the main pain-sensitive structures of the head. Sensory fibers to cranial structures derive from the trigeminal nerve and ganglion. These trigeminal terminals contain the neuropeptides substance P (SP), calcitonin gene-related peptide (CGRP), and pituitary adenylate cyclase activating polypeptide (PACAP) and supply the meninges and their vessels, in particular the pial arterioles in the region of the pial/glial limitans. Activation of the trigeminovascular system results in the release of CGRP and PACAP in the blood of the jugular vein, and, since CGRP release in the external jugular vein is found during the attacks of migraine, this has been taken as evidence of the activation of the trigeminal system during the attack. Cerebral vessels, namely those of the circle of Willis and the major arteries, receive also sympathetic fibers from

the cervical ganglia and parasympathetic nerve fibers originating from the sphenopalatine and otic ganglia. Intraparenchymal brain vessels also receive fibers originating directly from the CNS, including cholinergic (basal forebrain and mesopontine tegmentum), noradrenergic (locus caeruleus), and serotonergic (raphe) regions, but they are unlikely to be involved in the phenomenology of migraine. The trigeminocaudal nucleus forms part of the so-called trigeminocervical complex, a structure equivalent to the dorsal spinal horn and extending down to the C1 and C2 spinal levels. Stimulations of the superior sagittal sinus, dura mater, and cerebral vessels all result in activation of the trigeminocervical complex, which also receives convergent afferents from face, teeth, oral mucosa, and even from the greater occipital nerve originating from the C2 root, possibly accounting for the pain referred from cervical structures. This nucleus thus probably represents for primates the nucleus that mediates the pain of migraine. Trigeminovascular mechanisms are closely integrated with autonomic control exerted at all levels of the central autonomic network (CAN). In particular, connections exist between the trigeminal nucleus caudalis and the superior salivatory nucleus, which supplies preganglionic parasympathetic fibers to the sphenopalatine ganglion, which in turn innervates the cerebral vessels and the lacrimal gland and mucosa.

AUTONOMIC SYMPTOMS IN THE COURSE OF THE MIGRAINE ATTACK Neurologic signs of enhanced parasympathetic outflow to the head are found during migraine attacks in 73% of subjects, often bilaterally [3]. Conjunctival injection, lacrimation, nasal congestion, periorbital swelling, rhinorrhea and salivation, and diarrhea and frequent urination all may accompany the attacks of migraine. The trigeminoparasympathetic reflex accounts for these autonomic symptoms and signs observed in migraine attacks and in the trigeminal autonomic cephalalgias

*In memory of Pasquale Montagna, Professor of Neurology at Alma Mater Studiorum – University of Bologna, who died prematurely on 9 December 2010.

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reflexes, and biochemical and pharmacologic responses. These studies aimed at establishing whether a pattern of autonomic dysfunction could be observed in migraine patients, consistent with the usual distinction between the sympathetic and the parasympathetic branch of the autonomic nervous system. The results of these studies are inconsistent and contradictory: sympathetic hypofunction or hyperfunction and normal findings have been reported for each of the techniques used. To date, no unambiguous autonomic deficit has emerged that can be considered intrinsic to migraine headaches in the interictal state. These studies however hardly take into consideration that migraine attacks may represent integrated behavioral responses in which pain perception and modulation is played as a behavioral motif while autonomic responses serve behavior.

THE PAIN OF MIGRAINE

FIGURE 113.1 Schematic figure of the human brain stem, depicting the trigeminovascular mechanisms and parasympathetic pathways thought to be involved in the origin of pain and autonomic symptoms during the migraine attack. IV, fourth ventricle; ACh, acetylcholine; CGRP, calcitonin gene-related peptide; LC, locus coeruleus; MRN, magnus raphe nucleus; NKA, neurokinin A; NO, nitric oxide; PAG, periaqueductal gray region; SP, substance P; SPG, superior sphenopalatine ganglion; SSN, superior salivatory nucleus; TG, trigeminal ganglion; TNC, trigeminal nucleus pars caudalis; VIP, vasoactive intestinal peptide. (Modified from Pietrobon D, Striessnig J. Neurobiology of migraine. Nat Rev Neurosci 2003;4:386–398.)

(Fig. 113.1). Involvement of the sphenopalatine ganglion in the production of these autonomic signs and also of migraine pain is implied by their abolition upon sphenopalatine ganglion blockade with intranasal lidocaine. This procedure, however, is ineffective in alleviating the cutaneous allodynia during the migraine attack, thus demonstrating that the parasympathetic outflow contributes to migraine pain but is not responsible for pain sensitization during the attack.

INTERICTAL AUTONOMIC DYSFUNCTION IN MIGRAINE Interictal autonomic dysfunction in migraine has been variously investigated by techniques measuring cerebrovascular reactivity, reactivity of the pupils, cardiovascular

The brain contains no pain fibers and the only way it may signal pain is through the trigeminovascular system. Migraine is conventionally portrayed as a pain disorder, even though pain represents just the tip of the migraine iceberg, may even be absent and it is preceded by prodromes separated from the aura [4]. Prodromes (somnolence, hyperphagia or food rejection, changes in fluid balance) implicate the concept of “complete migraine” that is better explained by “a diffuse cerebral sufferance spreading to the hypothalamus” [4]. The behavior of migraine patients during an attack is characteristic, with patients wanting to lie down and remain as immobile as possible, dozing if they are able, in order to avoid all kinds of physical and mental exercise; this may be defined as a sickness behavior. The resolutory phase is characterized by tiredness, fatigue, head pain, difficulties in concentrating, “hangover”, gastrointestinal symptoms, mood changes, yawning and somnolence, and may end with sleep. Since no structural lesion of the central or peripheral nervous system has been shown to underlie the migraine pain it is better conceptualized as visceral pain, thus a dysfunction of the interoceptive system that, associated with autonomic control, engenders distinct feelings in the human body [5]. A visceral system of feeling is essential for the representation of the physiological conditions of all tissues in the body including the brain and for the maintenance of homeostatic activity [6]. Indeed the so-called pain neuromatrix (i.e., the network of brain regions activated during pain) strikingly overlaps with that of visceral afferent information. Migraine as pain of the brain, may therefore be viewed as a visceral homeostatic emotion like hunger, thirst, itching, and temperature, and as such it may represents a monitor of the internal bodily world with a fundamental adaptive role, as well as a powerful motivator of behavior. The concept of migraine as a visceral pain serving a homeostatic function in the interoceptive brain by way of

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its association with specific behavioral responses, leads to the consideration that headache pain should be understood within the frame of its behavioral significance (i.e., its “subjective utility”).

MIGRAINE AS A BIO-BEHAVIORAL DISORDER Welch [7] proposed the so-called bio-behavioral model for migraine, positing that migraine represents a “biobehaviorally based dysautonomia involving principally the intrinsic noradrenergic system and its putative orbitofrontal connections”. This model unifies the multifaceted phenomena of the migraine attack and migraine natural history around a principal pathogenic role played by the CAN, responsible not only for the pain during the attack, but also for the prodromic and resolution symptoms so often overlooked in pathogenesis. Above all, the biobehavioral model may usefully incorporate the interictal traits characteristic of the migraine brain, be they the predisposing factors of hypoxia or deficient brain energy metabolism [8] or the deficient habituation in information processing demonstrated by the evoked potential studies [9]. Within this bio-behavioral frame, the migraine attack could be envisaged as a protective event, as befits the general function of pain as a sensory modality. That migraine pain serves a protective function seems suggested also by the persistence of migraine as a genetic trait frequent in the population of reproductive age, and by hints that migraineurs have an evolutionary advantage [10].

MIGRAINE AS A REFLECTION OF GENETICALLY DETERMINED ADAPTIVE DARWINIAN BEHAVIORAL RESPONSES Within the autonomic-behavioral responses, sickness behavior seems to be best suited to the attack of migraine. The sickness behavior represents an innate genetically determined response typical of all mammalian species, evolved for adaptive reasons in the animal confronted with stressors such as a predator or a bacterium/parasite that in different ways threatens the survival of the organism. These responses are evolutionarily conserved, and their adaptive role also applies to the other emotional and autonomic features that are aptly integrated into the defined behavioral program. Darwin was the first to apply an evolutionary perspective to emotions both in animals and humans, and to recognize patterns of “emotional” behaviors that could have an evolutionary meaning [11]. A speculative hypothesis views migraine attacks as the response to visceral afferent signals which signify inescapable visceral (brain) pain and which lead to a sickness response and antinociception that helps to terminate the attack [12]. This quiet coping strategy as

evolutionarily conserved adaptive Darwinian behavior, has an adaptive role and it helps the brain to recover to the homeostasis of the brain itself. These considerations are counterintuitive when we recall that migraine is classified as disease, but an evolutionarily conserved behavior may become inappropriate and maladaptive under determined circumstances, particularly in an allostatic perspective.

CONCLUSION In summary, the bio-behavioral theory expanded in a Darwinian perspective views a migraine attack as an adaptive behavioral response engendered out of a genetic (evolutionary conserved) repertoire by a network of pattern generators in the central autonomic motor system for the maintenance of brain homeostasis. Such a view leads to several considerations: l

l

l

l

It is futile to study autonomic functions during the headache attack in isolation and separated from the biobehavioral responses they contribute to. Pain in the different headaches may be variously perceived according to its behavioral significance. Migraine and more generally the behavioral responses associated with it may be actually adaptive and serve the purpose of recovering brain homeostasis. The challenge is to identify which primarily nervous deviance from brain homeostasis (abnormal reward processes? information overload? energy expenditure? cell hyper-excitability?) typifies the migraine brain.

References [1] Headache Classification Subcommittee of the International Headache Society The International Classification of Headache Disorders. Cephalalgia 2004;24:1–160. [2] Goadsby PJ, Lipton RB, Ferrari MD. Migraine – current understanding and treatment. N Engl J Med 2002;346:257–70. [3] Gupta R, Bhatia MS. A report of cranial autonomic symptoms in migraineurs. Cephalalgia 2007;27:22–8. [4] Blau JN. Migraine: theories of pathogenesis. Lancet 1992;339:1202–9. [5] Saper CB. Pain as a visceral sensationMayer E.A. Saper CB, editors. Progress in brain research, Vol 122. : Elsevier; 2000. p. 237–43. [6] Craig AD. A new view of pain as a homeostatic emotion. Trends Neurosci 2003;26:303–7. [7] Welch KM. Migraine: a bio-behavioural disorder. Cephalagia 1986;6:103–10. [8] Montagna P, Sacquegna T, Cortelli P, Lugaresi E. Migraine as a defect of brain oxidative metabolism: a hypothesis. J Neurol 1989;236:124–5. [9] Schoenen J. Pathogenesis of migraine: the biobehavioral and hypoxia theories reconciled. Acta Neurol Belg 1994;94:79–86. [10] Loder E. What is the evolutionary advantage of migraine? Cephalalgia 2002;22:624–32. [11] Darwin C. The expression of the emotions in man and animals. 1873. [12] Montagna P, Pierangeli G, Cortelli P. The primary headaches as a reflection of genetic Darwinian adaptive behavioral responses. Headache 2010;50:273–89.

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114 Epilepsy and Autonomic Regulation Matthias Dütsch, Max J. Hilz Hughlings Jackson – one of the pioneers of clinical neurology – describes numerous autonomic symptoms during epileptic seizures: “… as the seat of the discharging lesion varies symptoms of the paroxysm vary … increased flow of saliva; pallor of face; shivering, with sensation of cold; arrest of respiration with sensation of suffocation; colored vision; noises in the ears; nausea (and other less definite sensations referred to the epigastrium); movements of the eyes with vertigo; convulsions of the limbs etc” [1]. Based on his clinical observations of patients with seizures, brain tumors and other structural diseases of the brain, Jackson assumed a cortical representation of the sympathetic nervous system. Today, we know that autonomic symptoms occur depending on the brain region involved in the spread of epileptic discharges. Still, autonomic symptoms are not specific for a certain area within the central nervous system. A tightly interconnected central neuronal network causes a variety of clinical autonomic symptoms. Beyond their central role for autonomic control, specific parts of the central autonomic network (CAN), e.g., the insula, amygdala, cingulate gyrus, and prefrontal cortex, also represent the most common foci of partial epilepsy, a finding that might explain the frequent observation of autonomic changes in association with epileptic seizures.

SUPRATENTORIAL COMPONENTS OF THE CENTRAL AUTONOMIC NETWORK The insula, anterior cingulate gyrus, and ventromedial prefrontal cortex are key areas of central autonomic function [2,3]. The insula represents the primary viscerosensory cortex. The cingulate gyrus and prefrontal (frontopolar and orbitofrontal) cortices constitute the premotor autonomic region. Bilateral lesions in that area not only provoke intellectual and emotional disturbances but also compromise autonomic responses to emotional stimuli. Isolated anterior cingulate gyrus lesions may cause gastrointestinal and urogenitary dysfunction. Intraoperative cingulate gyrus stimulation resulted in changes of heart rate and blood pressure. The amygdala mediates autonomic responses to emotions. In animal experiments, electrical or chemical

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stimulation of the central nucleus of the amygdala induced changes of blood pressure, heart rate, respiration, gastrointestinal motility and secretion. Physiological stimulation of the amygdala may be involved in hypertension and fear-related cardiac arrhythmia. In addition to the functional specialization of specific CAN structures, there is hemispheric lateralization of cardiovascular autonomic control. In temporal lobe epilepsy patients, pre-surgical amobarbital induced, unilateral hemispheric inactivation shows sympathetic predominance of the right hemisphere and parasympathetic predominance with up-regulation of baroreflex sensitivity in the left hemisphere [4].

INFRATENTORIAL COMPONENTS OF THE CENTRAL AUTONOMIC NETWORK At the subcortical level, the hypothalamic region provides an interface with endocrine stimuli and triggers autonomic responses to maintain homeostasis [2,3]. The periaqueductal grey in the mesencephalon contributes significantly to autonomic and antinociceptive responses as well as behavioral responses to stressful stimuli (“fight or flight” reaction). The parabrachial region in the dorsolateral pontine region is an important relay involved in cardiorespiratory control. Stimulation of that area may yield blood pressure increase and baroreflex inhibition. The ventrolateral pontine region contains a group of norepinephrine synthesizing cells. Stimulation of this A5-region provokes complex cardiovascular responses, e.g., an initial drop with consecutive rise of blood pressure accompanied by tachycardia and increased sympathetic activity in splanchnic nerves and reduced sympathetic activity in lumbar nerves. The solitary tract in the dorsomedial medulla oblongata is involved in several medullary reflexes including the baroreflex. Lesions in this area provoke either excessive hypertension or instable blood pressure. The ventrolateral medulla oblongata controls vasomotor tone, cardiac and respiratory function. Bilateral lesions in this region cause extensive hypertension. Moreover, this group of neurons modulates rhythm and

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deviation of the eyes, do not relate to any specific region of EEG activation.

Urogenital Autonomic Dysfunction The most common urogenital symptom associated with tonic clonic seizures is urinary incontinence. Sexual thoughts or sensations and even sexual arousal can be part of the clinical presentation of simple partial seizures. These seizures often originate in temporal lobe areas although painful urogenital episodes and orgasms have also been observed in patients with symptomatic epilepsies due to parietal tumors. Complex partial seizures may provoke sexual automatisms such as rhythmic pelvic movements, masturbation or exhibitionistic behavior, especially in patients with frontal lobe epilepsies [3,6].

Ictal Cutaneous and Pupillary Changes FIGURE 114.1 Piloerection and goose bumps in a woman during a right temporal lobe seizure. (Courtesy of the Center for Epilepsy Erlangen, Department of Neurology, University of Erlangen-Nuremberg, Germany.)

depth of breathing. Superficial neurons of the ventrolateral medulla oblongata contain chemosensory neurons that – similar to the region of the area postrema near the 4th ventricle – are sensitive to changes in carbon dioxide or pH in the cerebrospinal fluid. These neurons influence vasomotor tone and breathing rhythm [2,3].

ICTAL AUTONOMIC DYSFUNCTION Gastrointestinal Autonomic Dysfunction Gastrointestinal symptoms such as nausea are frequent complaints during an aura (“epigastric aura”). Current findings favor an insular or mesial temporal lobe origin of the subjective discomfort attributed to the epigastrium [5]. “Abdominal epilepsy” is a condition that was described by Penfield and is mostly seen in children. Clinically, patients complain about (epi-)gastric pain, nausea, spasms, meteorism, diarrhea. The symptoms might be provoked by epileptic discharges in the insular region or medial frontal areas. A rare form of epilepsy is the “gastrointestinal reflex epilepsy” with intermittent crises of rectal pain after a phase of increased motility of the gut. Simultaneous video-EEG recordings showed that ictal vomiting was associated with an epileptic spread within the temporal lobe of the non-dominant hemisphere [3,6]. Panayiotopoulos syndrome is a common childhood susceptibility to autonomic seizures with vomiting, paling, sweating, drooling, mydriasis, and status epilepticus. Ictal vomiting and other autonomic manifestations, as well as

Facial flushing and pallor as well as piloerection, goose bumps, profuse sweating, and an increase of nasal or lacrimal secretion might occur during simple partial or complex partial seizures that often originate from temporal, but also frontal or parietal areas (Fig 114.1). Bilateral mydriasis with areactive pupils is often seen with tonic clonic seizures. Unilateral mydriasis is a rare symptom of simple or complex partial temporal lobe seizures [3,6].

Respiratory Autonomic Dysfunction Epilepsy associated respiratory autonomic dysfunction is less common than other autonomic symptoms. In newborns, however, apnea might be the sole epileptic symptom. Still, apnea, reduced respiratory frequency and volume or inspiratory stridor and coughing may be associated with seizures involving, e.g., (mesial-) temporal, amygdala or anterior cingulate regions [7]. Simple or complex partial seizures may be associated with emotional responses such as fear or panic reaction. These seizures often involve the amygdala and are associated with hyperventilation [3,8,6].

Cardiovascular Autonomic Dysfunction Cardiovascular ictal dysfunction is the most prominent autonomic feature of epileptic seizures. Simultaneous video/EEG recordings that include an ECG trace demonstrated seizure-associated cardiac arrhythmia. Sinus tachycardia with heart rates between 120 to 200 bpm is the most common ictal arrhythmia. Heart rate acceleration is not restricted to a certain seizure type but is often seen with generalized tonic clonic seizures. Apart from sinus tachycardia, increased sympathetic tone during seizure may manifest as (supra-)ventricular tachycardia and atrial or even ventricular fibrillation.

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Ictal bradyarrhythmia is a relatively rare symptom during seizures. However, different types of bradyarrhythmia, e.g., sinus bradycardia, atrioventricular blocks, sinus arrest and asystole, have been observed during seizures. The onset of cardiac arrhythmia might precede the clinical or electroencephalographic onset of seizures suggesting CAN involvement in epileptic discharges prior to spreading of epileptic activity to areas recorded with superficial electrodes. In some patients, ictal tachyarrhythmia is associated with symptoms such as headache, hypertension, sweating, and shivering, resembling pheochromocytoma, or with pain radiating towards the neck, jaw or left arm, resembling angina pectoris. During seizures, there may be – mostly benign – electrocardiographic (ECG) abnormalities; serious ECG changes such as ST segment depression or T-wave inversion have been reported in 6–13% of seizures [7]. Differential diagnosis of epilepsy induced cardiovascular abnormalities includes side-effects of anticonvulsant drugs, such as carbamazepine (bradycardia, atrioventricular block), or convulsive syncopes due to long QT-syndromes [3].

Sudden Unexpected Death in Epilepsy Patients (SUDEP) Mortality rate of epilepsy patients is two to five times higher than of the general population [9]. Sudden unexpected death in epilepsy (SUDEP) is considered the most common cause of death in epilepsy patients [9]. SUDEP is mostly defined as sudden, unexpected death of patients with epilepsy that is not due to trauma or drowning. It may occur with or without evidence of a seizure, excluding deaths attributed to status epilepticus [10]. In cases of definite SUDEP, postmortem examination does not reveal a specific structural or toxicologic etiology of death [3,10]. Data on SUDEP incidence vary and range from incidence rates of 0.1 to 2 per 1000 person-years but also seem to be higher in selected populations, with rates as high as 6 to 9 per 1000 person-years in epilepsy surgery patients [10]. The exact mechanisms leading to SUDEP are still unknown. It is hypothesized that autonomic cardiac dysregulation causes lethal cardiac arrhythmia. Autopsies of SUDEP patients showed subendocardial ischemia and significant myocardial fibrosis due to assumed recurrent ictal ischemia, possibly associated with ictal arrhythmia. Moreover, neurogenic pulmonary edema seems to be involved in the SUDEP pathomechanisms [3]. Neurogenic pulmonary edema might result from massive increase in sympathetic activity during seizures [3]. Animal experiments suggest a 1:1 synchronization between epileptic discharges and discharges in cardiac sympathetic (and sometimes parasympathetic) nerves. This synchronization might be the functional substrate of ictal life-threatening or even -terminating cardiac arrhythmia.

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Autonomic Dysfunction During Status Epilepticus Repeated or ongoing epileptic discharges provoke massive autonomic arousal. A massive increase of sympathetic activity in the early phase of status epilepticus increases insulin, glucagon and catecholamine levels. Clinically, the patients might have high blood pressure, tachycardia, hyperglycemia, increased temperature and even pulmonary edema. Simultaneous activation of central sympathetic and parasympathetic activity may cause cardiac ventricular arrhythmia. Prolonged status epilepticus with increasing lactate acidosis may induce arterial hypotonia. Cerebral autoregulation may be compromised during status epilepticus to the extent that cerebral perfusion is passively driven by systemic blood pressure [3].

INTERICTAL AUTONOMIC DYSFUNCTION Mostly, reports on interictal autonomic dysfunction imply cardiovascular abnormalities. Interictal findings include an increased heart rate and blood pressure variability as well as increased responses to sympathetic stimulation maneuvers such as orthostatic challenge and cold pressor testing. Mathematical methods derived from chaos theory showed significantly reduced heart rate modulation in patients with frequent interictal epileptic discharges [3]. Spectral analysis showed decreased total autonomic variability in medically refractory temporal lobe epilepsy patients with a relative increase of sympathetic tone. Baroreflex function was impaired in this patient group [11]. Reduced cardiac uptake of Iodine123 meta-iodobenzylguanidine (MIBG) has been demonstrated interictally in patients with chronic temporal lobe epilepsy. Prominent reduction in cardiac MIBG uptake of patients with ictal asystole indicates post-ganglionic cardiac catecholamine disturbance in this patient group. Impaired sympathetic cardiac innervation might compromise heart rate adjustment and modulation, and may thus increase the risk of asystole and ultimately sudden unexpected death in epilepsy [12,13]. After temporal lobe epilepsy surgery, there is a reduction of sympathetic cardiovascular modulation and baroreflex sensitivity that might result from decreased influences of interictal epileptogenic discharges on brain areas involved in cardiovascular autonomic control. Temporal lobe surgery might stabilize the cardiovascular control in epilepsy patients by reducing the risk of sympathetically mediated tachyarrhythmias with subsequent counter-regulation causing excessive bradycardia. In addition, these patients showed reduced sympathetic cerebrovascular modulation and improved cerebral autoregulation after surgery [14–16].

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Effects of Vagus Nerve Stimulation on Cardiovascular Function of Epilepsy Patients Vagus nerve stimulation (VNS) does not seem to compromise autonomic cardiovascular regulation in epilepsy patients. However, VNS has been reported to increase respiratory frequency and decrease respiratory amplitude in children during sleep resulting in variable effects on cardiac activity [17,18].

References [1] Jackson JH. Clinical and physiological researches on the nervous system.1. On the anatomical and physiological localization of movements in the brain. London: Low, Churchill; 1875. [2] Benarroch EE. Central autonomic network: functional organization and clinical correlations. Armonk, New York: Futura Publ. Comp. Inc.; 1997. 3–60. [3] Hilz MJ, Dütsch M, Kölsch C. Epilepsy and autonomic diseases. Fortschr Neurol Psychiatr 1999;67(2):49–59. [4] Hilz MJ, Dütsch M, Perrine K, Nelson PK, Rauhut U, Devinsky O. Hemispheric influence on autonomic modulation and baroreflex sensitivity. Ann Neurol 2001;49:575–84. [5] Santana MT, Jackowski AP, da Silva HH, Caboclo LO, Centeno RS, Bressan RA, et al. Auras and clinical features in temporal lobe epilepsy: a new approach on the basis of voxel-based morphometry. Epilepsy Res 2010;89(2–3):327–38. [6] Freeman R, Schachter SC. Autonomic epilepsy. Semin Neurol 1995;15(2):158–66. [7] Devinsky O. Effects of seizures on autonomic and cardiovascular Function. Epilepsy Curr 2004;4(2):43–6.

[8] Gerez M, Sada A, Tello A. Amygdalar hyperactivity, a fear-related link between panic disorder and mesiotemporal epilepsy. Clin EEG Neurosci 2011;42(1):29–39. [9] Tellez-Zentano JF, Ronquillo LH, Wiebe S. Sudden unexpected death in epilepsy: Evidence-based analysis of incidence and risk factors. Epilepsy Res 2005;65(1–2):101–15. [10] Surges R, Thijs RD, Tan HL, Sander JW. Sudden unexpected death in epilepsy: risk factors and potential pathomechanisms. Nat Rev Neurol 2009;5(9):492–504. [11] Dütsch M, Hilz MJ, Devinsky O. Impaired baroreflex function in temporal lobe epilepsy. J Neurol 2006;253:1300–8. [12] Druschky A, Hilz MJ, Hopp P, Platsch G, Radespiel-Tröger M, Druschky K, et al. Interictal cardiac autonomic dysfunction in temporal lobe epilepsy demonstrated by [(123)I]metaiodobenzylguanidine-SPECT. Brain 2001;124:2372–82. [13] Kerling F, Dütsch M, Linke R, Kuwert T, Stefan H, Hilz MJ. Relation between ictal asystole and cardiac sympathetic dysfunction shown by MIBG-SPECT. Acta Neurol Scand 2009;120:123–9. [14] Hilz MJ, Platsch G, Druschky K, Pauli E, Kuwert T, Stefan H, et al. Outcome of epilepsy surgery correlates with sympathetic modulation and neuroimaging of the heart. J Neurol Sci 2003;216:153–62. [15] Hilz MJ, Devinsky O, Doyle W, Mauerer A, Dütsch M. Decrease of sympathetic cardiovascular modulation after temporal lobe epilepsy surgery. Brain 2002;125:985–95. [16] Dütsch M, Devinsky O, Doyle W, Marthol H, Hilz MJ. Cerebral autoregulation improves in epilepsy patients after temporal lobe surgery. J Neurol 2004;251:1190–7. [17] Stemper B, Devinsky O, Haendl T, Welsch G, Hilz MJ. Effects of vagus nerve stimulation on cardiovascular regulation in patients with epilepsy. Acta Neurol Scand 2008;117:231–6. [18] Zaaimi B, Grebe R, Berquin P, Wallois F. Vagus nerve stimulation induces changes in respiratory sinus arrhythmia of epileptic children during sleep. Epilepsia 2009;50:2473–80.

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115 Disorders of Sweating Robert D. Fealey Sweating, an important thermoregulatory activity under autonomic control, can be disturbed by disorders resulting in excessive (hyperhidrosis) or deficient (hypohidrosis and anhidrosis) sweating. The distribution of abnormal sweating can be ascertained via established autonomic tests and inferences regarding the pathophysiology of the disorder made. Sweating excess of the hands (primary focal or essential hyperhidrosis) and global anhidrosis with heat intolerance and hyperthermia (chronic idiopathic anhidrosis) are examples of hyperhidrosis and anhidrosis respectively. Many of the disorders and tests of sweating are more thoroughly discussed in the key references [1–5]. This chapter comprises a detailed, organized overview making liberal use of tables to discuss sweating disorders. After several definitions the chapter discusses first hyperhidrotic and then hypohidrosis/anhidrotic conditions.

HYPERHIDROSIS Hyperhidrosis is defined as sweating that is excessive for a given thermoregulatory or emotional stimulus. Some of the physiologic factors that increase the sweat response normally are mentioned in Table 115.1. When sweating exceeds these physiologic considerations pathologic responses or conditions need to be considered. A simplified algorithm approach categorizing hyperhidrotic disorders is shown in Figure 115.1. Hyperhidrosis can be generalized or localized, presenting the physician a challenging differential diagnosis (see Tables 115.2 and 115.3). Generalized hyperhidrosis can be primary (as in episodic hypothermia with hyperhidrosis or Shapiro’s syndrome) or secondary and due to general medical disorders. The hyperhidrosis is usually episodic rather than continuous with most disorders. In pheochromocytoma paroxysmal sweating is associated with headache, hypertension and tachycardia and more often a norepinephrine secreting tumor. High circulating levels of catecholamines may stimulate normal thermoregulatory structures to produce cholinergic sudomotor activity. Hodgkin disease is characterized by the triad of fever, sweating, and weight loss; night sweats may be the only symptom and 31 of 100 patients present with “B”-cell

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00115-3

symptoms (fever, weight loss, sweating). The excessive production of IL-1 by activated macrophages is implicated as the cause of temperature instability. IL-1 is known to induce an abrupt increase in the synthesis of prostaglandin E2 in the preoptic anterior hypothalamic region, causing an elevation of the temperature “balance point”. Excessive production of IL-6 by Hodgkin lymphoma cells is also implicated as cause of fever and subsequent night sweating. Advanced solid tumors may also cause sweating via immunologic mechanisms relating to TNF-α and effect of ILs on central thermoregulation. Anticholinergics, H2 receptor antagonists, plasma exchange and prostaglandin inhibitor drug therapy can be effective in these disorders. In hyperthyroidism inappropriate heat production and increased autonomic nerve sensitivity to circulating epinephrine may cause elevated body temperature and sweating and B adrenergic blockade can be effective treatment. Localized hyperhidrosis of the palms, soles and axillae with normal sweating elsewhere is a common disorder known as primary focal or essential hyperhidrosis. Existing evidence suggests an abnormal, regional increased activity of both cholinergic and adrenergic component of sweat gland innervation, coupled with over activity of sympathetic fibers passing through the T2–4 ganglia, may be partially responsible. This is a disorder of adolescents and young adults. A familial tendency is present 25–50% of the time. Axillary hyperhidrosis may present without other areas involved and vice versa. Thermoregulatory sweating almost always normal over the remainder of the body and is abnormally augmented in the palms and soles. Very rarely these patients exhibit heavy acral (distal) sweating while other body parts exhibit little or no thermoregulatory sweating. These rare patients clinically present as classic Primary Focal Hyperhidrosis patients and do not have generalized autonomic failure. Primary focal hyperhidrosis is frequently socially and occupationally disabling and requires treatment. Commonly used therapeutic modalities with the details of treatment are given in Table 115.4. Another fairly common localized hyperhidrosis occurs in post-menopausal women and primarily affects the head and upper trunk. Multiple factors including age and hormonal-hypothalamic thermoregulatory balance-point alterations are likely involved. When given a heat stress the remainder of body surface sweats normally. A disorder

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TABLE 115.1 normal (Physiologic) factors Affecting Sweating Parameter

Comments

Age, race and sex effects

Threshold temperature lower and sweat gland output higher in men. Higher sweat output per gland in young individuals. No significant sweating differences based on race

Acclimatization

Increased sweat gland size and sweat output with endurance training, chronic heat or greater humidity exposure

Circadian rhythm

Threshold temperature to sweat varies frequently lowest from 12 midnight to 4 am

Posture

Lying on side produces a contralateral hyperhidrosis and ipsolateral hypohidrosis

Stress and eating

Stress can increase sweating in palms, axillae, feet and forehead; some healthy subjects have symmetric gustatory sweating of their face with spicy food

Skin temperature

When near threshold temperature generalized sweating can be augmented or inhibited by changes in local skin temperature without core temperature change

FIGURE 115.1 Algorithm categorizing hyperhidrotic disorders.

known as idiopathic paroxysmal localized hyperhidrosis, which occurs in men and women, may be related as both conditions are responsive to clonidine a centrally acting alpha-2 adrenergic receptor agonist. Localized hyperhidrosis can also occur in the rare genetic disorder cold-induced sweating syndrome types I and II. This disorder and the pediatric-neonatal Crisponi syndrome are associated with alterations of the composite cytokine CLCF1 gene. The cold-induced sweating involves the upper body segments, which paradoxically do not sweat in the heat. Otherwise, localized hyperhidrosis is usually the result of autonomic nervous system lesions which are associated with compensatory or perilesional hyperhidrosis; not uncommonly the patient’s attention is given to the excessive sweating area when the abnormality is the widespread anhidrosis elsewhere! Diabetic autonomic failure, pure autonomic failure (Bradbury–Eggleston syndrome), Ross syndrome, Harlequin syndrome, chronic idiopathic anhidrosis and autoimmune autonomic ganglionopathy are examples where the phenomenon may occur. Both gustatory sweating

and the auriculotemporal (Frey’s) syndrome represent examples of localized hyperhidrosis due to aberrant regeneration of autonomic nerves damaged either surgically or by neuropathy. Contralateral hyperhidrosis following cerebral infarction has been described as an uncommon complication possibly due to interruption of descending inhibitory pathways. Patients with cervical and upper thoracic complete spinal cord traumatic transections are frequently troubled with localized hyperhidrosis of the head and upper trunk when noxious stimuli below the level of their lesion cause autonomic hyperreflexia. When accompanied by paroxysmal hypertension this syndrome can be confused with pheochromocytoma. Often the causative stimulus is a distended bladder or rectum. Increased spinal alpha adrenoreceptors and peripheral micro vascular adrenoreceptors as well as accumulation of substance P and reduction of inhibitory transmitters below the cord lesion may be causative. Removing the stimulus is the most effective treatment, although antihypertensives (clonidine, α blockers, and calcium channel blockers) and anticholinergic agents (once any gastrointestinal or urinary obstruction is relieved) such as propantheline, or glycopyrrolate are sometimes employed to suppress sweating. In syringomyelia the excessive sweating is segmental and often appears in dermatomes where sensation is later disturbed. Hyperhidrosis with partial nerve trunk injury occurs as part of a complex regional pain syndrome and may be due to an obvious lesion or an occult problem such as paraspinal metastatic deposits or primary nerve sheath tumor affecting the sympathetic chain or white rami.

HYPOHIDROSIS AND ANHIDROSIS Hypohidrosis is reduction in sweating and anhidrosis is the absence of sweating in response to an appropriate thermoregulatory or pharmacological stimulus. Normally, small areas of anhidrosis can occur in skin over bony prominences, and physiologic hypohidrosis can occur in proximal extremities of the elderly and women and in abdominal skin of obese individuals. Dehydrated individuals may show delayed sweat onset and generalized hypohidrosis.

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TABLE 115.2 Some Causes of Pathological generalized Hyperhidrosis Condition

Pathophysiologic Mechanism

Pheochromocytoma

Physiologic response to inappropriate catecholamine-induced thermogenesis; inhibited by anticholinergics

Thyrotoxicosis

Physiologic response to inappropriate heat production-induced thermogenesis; inhibited by B-blockers

Acromegaly

Growth hormone induced increase in sweat gland secretion rate; suppressed by somatotatin analogs and dopamine agonists especially with prolactin co-secretion

Malignancy/chronic infection

Night sweats; related to altered hypothalamic balance point temperature and effects on prostaglandin E2 or other “thermogenic cytokines” like interleukin IL1B, tumor necrosis factor, IL6; Probable vagal afferent “pyrogenic” pathways activated by complement factors.

Other causes

Severe anxiety, hypoglycemia, hypotension, cholinergic agents

TABLE 115.3 Differential Diagnosis and Pathophysiology of Some Causes of Localized Hyperhidrosis Condition

Pathophysiologic Mechanism of Sweating

Primary focal (essential) hyperhidrosis

Excessive emotional and physiologic sweating affecting hands, feet and axillae; isolated craniofacial and axillary can occur; thermal sweating usually intact (Type 1); rarely acral sweating only (Type 2); abnormal regional increase of both cholinergic and adrenergic sweating postulated

Perilesional/compensatory hyperhidrosis

Central and/or peripheral denervation of large numbers of sweat glands produces increased sweat output in innervated glands, maximal in contiguous dermatomal regions; occurs in PAF, Ross syndrome, SCI and post surgical sympathectomy

Gustatory sweating

Resprouting of secremotor axons to supply denervated sweat glands; occurs in Frey syndrome and post-sympathectomy

Post cerebral infarct

Loss of contralateral inhibition with cortical and upper brainstem infarcts

Autonomic dysreflexia

Uninhibited, segmental somatosympathetic reflex; often accompanied by paroxysmal hypertension and triggered by distended colon or bladder or during bowel/bladder evacuation care; prophylaxis with anti-hypertensives usually effective for latter and relief of distended viscus for former

Paroxysmal localized hyperhidrosis

Idiopathic; may be segmental or unilateral; when segmental head/neck/upper trunk affected; seen in post-menopausal females; may reflect altered hypothalamic thermoregulation; clonidine and α2 centrally acting agonist is often effective Rx

Complex regional pain syndrome (type I/type II)

Localized sympathetic sudomotor hyperactivity; associated with edema and neuropathic pain; axon reflex, ephaptic spread, aberrant proliferation of peptide fibers may contribute; partial nerve root compression by nerve sheath or metastatic tumor may produce dermatomal hyperhidrosis

Other causes

Cold induced sweating syndrome; Chiari malformation; olfactory hyperhidrosis

TABLE 115.4 Some Treatment Measures for Primary Hyperhidrosis Treatment

Details of Treatment

Side Effects/Complications

Topical Rx

20% Aluminum chloride hexahydrate in anhydrous ethyl alcohol (Drysol); 12% aluminum chloride in sodium carbonate water. Apply to dry skin at bedtime; wrap in plastic (hands) washoff in am. Daily to weekly applications; follow Physicians’ Desk Reference directions carefully

Irritation of skin; less effect on palms and soles but good for axilla; can cause skin cracking, stinging

Tanning Rx.

Glutaraldehyde (2–10%) solution; apply 2-4 times weekly as needed

Stains skin brown; for soles of feet only

Iontophoresis

For palms/soles/axillae; 15-20 ma current patient controlled; 20-30 min sessions reverse polarity, use of dc current and adding glycopyrrolate to water (make 0.1% solution) may make more effective. Use 3-6 times/week for total of 10-15 times initially then 1-2 times/week maintainence

Shocks, tingling may occur Time consuming, benefit temporary Difficult to use in axillae

Anticholinergic

Glycopyrrolate (Robinul/Robinul Forte); 1-2 mg orally tid prn. For intermittent/adjuctive Rx

Dry mouth, blurred vision; contraindicated in glaucoma, GI or GU tract obstruction

Clonidine

Useful for paroxysmal, localized hyperhidrosis 0.1 to 0.3 mg orally thrice daily; or as a TTS patch 0.1mg/day up to 0.3mg/day

Somnolence, hypotension, constipation, nausea, rash, impotence, agitation, dry mouth

Botulinum A toxin

50 to 100 mouse units of Botulinum toxin A into each axilla or body area treated. High doses 200 mU give prolonged effect. Can be repeated

Injection discomfort, variable duration of effect 3-12 months; expensive; may cause weakness of intrinsic hand muscles; contraindicated in pregnancy, NMJ disease

Surgery

Endoscopic thoracic sympathectomy or sympathotomy (palmar hyperhidrosis) Axillary liposuction

Invasive procedures; local scaring, transient pain; post-op scar or infection; compensatory hyperhidrosis of trunk, pelvis, legs, feet; Horner’s syndrome, gustatory sweating; reserved for severe cases only

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TABLE 115.5 Some Tests of Sympathetic Sudomotor function Test

Method

Use

TST (Thermoregulatory Sweat Test)

Whole body heating; Alizarin Red indicator of sweating and non-sweating skin

Good screen for focal or generalized lesions; Determine pre vs. postganglionic when used with a postganglionic test; Body surface anhidrosis can be quantitated.

QSART/QSWEAT (Quantitative sudomotor axon reflex tests)

Iontophoresis of 1% acetylcholine; record indirect axon reflex response

Quantitative response from one or more sites; determine sweat volume and latency of response; test of postganglionic axons.

Pilocarpine Sweat Test

Iontophoresis of 1% pilocarpine solution

Quantitative response from one or more sites; use filter paper or capillary tube to collect and measure sweat volume.

Silastic Imprint Method

Iontophoresis of pilocarpine; count sweat droplet imprint from directly activated sweat glands

Quantitative response from one or more sites; determine sweat droplet volume distribution; test of postganglionic axons and sweat glands.

Q-TST (Quantitative Thermoregulatory Sweat Test)

Analyzes frequency of sweat expulsion and local sweat rate

Frequency of sweat expulsions with TRH and heating tests central control.

PASP (Peripheral Autonomic Skin Potential)

Measures change in sweating indirectly by change in skin resistance

Dynamic, semi-quantitative; adaptable to EMG equipment; complex multisynaptic somatosympathetic loop with CNS and PNS components.

QDIRT (Quantitative direct and indirect sweat test)

Ach iontophoresis; digital images of sweat spots; direct and indirect (axon reflex); timed sweat gland activity measured

Dynamic, quantitative response from one or more sites; may distinguish direct and indirect responses.

Ach, acetylcholine; TRH, thyrotropin releasing hormone; EMG, electromyography; CNS, central nervous system; PNS, peripheral nervous system.

Pathologic hypohidrosis and anhidrosis may produce symptoms of heat intolerance and dry skin; for example a patient may recognize that exercise in hot weather causes exhaustion, but not sweating or that their feet are dry and stockings no longer wet at day’s end. More often patients are unaware of specific symptoms and other signs and symptoms of autonomic neuropathy should be sought. Areas of compensatory, excessive sweating may be noted rather than the anhidrotic regions. The distribution, anatomical location and severity of pathologic sweat loss can be characterized by tests of sympathetic sudomotor function (see Table 115.5). Such may provide clues as to the underlying pathophysiology and clinical diagnosis. Sweat is readily visualized by a topical indicator such as iodinated starch or sodium alizarin sulfonate (alizarin Red S). These techniques are used to evaluate large body surfaces. Iodinated starch powder is prepared by adding 0.5 to 1.0 g of iodine crystals to 500 g of soluble starch in a tightly capped bottle. Sodium alizarin sulfate is mixed with an equal amount (by weight) of sodium carbonate anhydrous and twice the amount of corn starch. Both are applied to the skin and undergo a dramatic color change when moistened by the water (sweat) from activated sweat glands. Sweat gland activity can be studied quantitatively by a number of techniques including: filter paper collection, weighing and analyzing of sweat composition, quantitative sudomotor axon reflex tests (QSART/QSWEAT/ QDIRT), skin sympathetic potentials, silastic mold or iodine-impregnated paper imprint after pilocarpine stimulation, collection into a Wescor Macroduct coil, determining relative humidity changes using sensors within ventilated capsules and by determining total body surface anhidrosis to a maximal thermoregulatory stimulus during

a thermoregulatory sweat test (TST). Other, more sophisticated techniques use microdialysis membranes delivering minute quantities of transmitter substances to the dermis or use microcannulation of the sweat duct or coil or utilize confocal electron microscopy and immunohistochemical analysis of biopsied skin stained for peptides and proteins that comprise the structure and innervation of the sweat gland. Sweat gland nerve fiber density (SGNFD) has been developed into a commercially available test utilizing a 4 mm punch skin biopsy and pgp 9.5 axonal staining. It is desirable to combine several methods for determining the integrity of the eccrine sweat response. For example, a TST can be combined with tests of the sweat gland and/or its peripheral nerve innervation to localize a sweating disorder to the peripheral or central nervous system. Or, a volumetric technique can be combined with a sweat droplet distribution imprint to estimate the sweat volume per active gland. The composition of collected sweat can give critical information about eccrine function. For example, sweat chloride ion concentration can determine the integrity of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl channel and provide diagnostic information for cystic fibrosis. Common patterns of abnormal sweating are described next (Table 115.5). Illigens and Gibbons provide an excellent overview of current tests of sudomotor function [5].

Distal Anhidrosis This refers to sweat loss affecting the peripheral (acral) portions of the extremities, the lower anterior abdomen and the central forehead. The feet by far are the most commonly

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affected and the lesion is usually a postganglionic denervation as occurs in peripheral neuropathy.

A

B

Global Anhidrosis This pattern denotes near total body (80%) sweat loss. This can occur in central lesions (i.e., multiple system atrophy (MSA-Parkinsonism and MSA-cerebellar), hypothalamic tumor, cervical spinal cord transection); at times residual acral (distal) sweating will be present in MSA. Large areas of sweat loss that are shown to be preganglionic or central in origin characterize early MSA. Tests such as the TST show global anhidrosis, while postganglionic sweat tests such as QSART are often normal. Progressive sweat loss (both pre- and postganglionic) on consecutive testing is characteristic of MSA. Global anhidrosis combined with minute islands of preserved sweating and absent QSART is most often due to a widespread postganglionic lesion (i.e., panautonomic neuropathy). Most recently a partially reversible, widespread anhidrosis has been observed in some cases of autoimmune autonomic ganglionopathy associated with α3 AchR ganglionic antibodies. Immunotherapy of this disorder improves the sudomotor deficit.

C

D

F

E

G

Dermatomal, Focal, or Multifocal Anhidrosis This refers to sweat loss within the distribution of a peripheral nerve(s) or its branches or root(s) of origin (T1 to L2 or L3 ventral roots). Mononeuritis multiplex due to vasculitis and leprosy produces a multifocal pattern. Focal abnormalities can also occur with skin disorders that damage sweat glands or plug or destroy their ducts. Such can occur with the miliarias, atopic dermatitis, psoriasis, seborrhic dermatitis, lichen planus, ichthyosis, scleroderma, Fabry’s disease, congenital ectodermal dysplasia, Sjogren’s syndrome, radiation therapy skin injury, burns and pressure skin injury, scars and skin incisions.

Segmental Anhidrosis This pattern occurs when large, contiguous body areas of sweat loss with sharply demarcated borders conforming to sympathetic or somatic dermatomes are present. Sympathectomy, ganglionopathies and myelopathies produces such a pattern. When borders are not well defined and anhidrosis not contiguous, a regional pattern is said to exist. Both postganglionic and preganglionic lesions may produce these distributions.

Hemianhidrosis Sweat loss over one half of the body due to a lesion of the descending sympathetic efferents in the brainstem or upper cervical cord is called hemianhidrosis. Often the pattern is incomplete. Strokes, tumors, demyelinating lesions and trauma are frequently causative.

FIGURE 115.2 Thermoregulatory sweat test distributions. Examples from patients having diabetes mellitus (A, B, E, F); patient D had Pancoast’s syndrome (apical lung tumor) on the right. Shown are Distal (A), Segmental (B and D), Regional (C), Focal (multifocal, dermatomal) (E), Global (F), Normal (G) and Mixed (B, D, E) showing multifocal or segmental and distal patterns). Sweating in dark-shaded areas. Note how pattern of anhidrosis can suggest the pathophysiology: the glove and stocking pattern (A) in length-dependant neuropathy; the unilateral limb loss (B and D) with sympathetic chain lesions, the curvilinear dermatomal anhidrosis in diabetic truncal radiculopathy (E).

Mixed patterns of anhidrosis (i.e., distal with focal) often occur, e.g., in diabetic neuropathy. Examples of most of the distributions just described are shown in Figure 115.2. Many disorders cause disturbances in sweating and characteristic abnormalities of tests of sudomotor function. Primary autonomic disorders, central and peripheral nervous system lesions, iatrogenic causes and disorders of skin can be implicated. Table 115.6 summarizes some of these disorders. Hyperthermia, heat intolerance, heat prostration and heat stroke may occur with widespread failure of

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TABLE 115.6 Some Disorders of Sweating and Characteristic Abnormalities of Sudomotor Testing Clinical Disorder

Pattern/site of Lesion Based on TST/QSART Data

PRIMARY AUTONOMIC DISORDERS Pure autonomic failure (PAF); chronic idiopathic anhidrosis (CIA), Ross syndrome

Global (diffuse) or segmental anhidrosis with/without acral sparing. Segmental lesions often asymmetric and can alternate as to left or right side. Compensatory, segmentally preserved sweating often excessive and most noticeable to patient. Lesion usually ganglionic or postganglionic although some CIA cases can be preganglionic initially. Occasional peri-eccrine infiltrates in global CIA cases. Abnormal pupils/absent reflexes in Ross syndrome.

Multiple system atrophy (MSA-P, MSA-C and MSA-A)

Global, segmental or regional widespread anhidrosis with or without acral sparing. Early preganglionic involvement, both preganglionic and postganglionic later stages. Rapid progression of anhidrosis indicates poor prognosis.

Acute pandysautonomia (panautonomic neuropathy) Segmental or distal (or both) anhidrosis without acral sparing but with scattered Autoimmune autonomic ganglionopathy islands of preserved sweating. Global at times. Ganglionic, preganglionic (synaptic) Paraneoplastic and other autoimmune autonomic neuropathies and postganglionic axonal involvement occurs. Sometimes associated with alpha-3 ganglionic acetylcholine receptor antibodies. May show spontaneous improvement (over months) and improve after immunotherapy. Neurogenic chronic idiopathic intestinal pseudo-obstruction

Often normal studies; occasional distal or more widespread regional postganglionic involvement.

CENTRAL NERVOUS SYSTEM LESIONS Tumors: hypothalamic, parasellar, pineal, brainstem, spinal cord Global with hypothalamic involvement. Segmental and ipsolateral with cord/ brainstem lesions. Preganglionic involvement. Cerebral infarction

Contralateral hyperhidrosis occurs (most prominent acutely) with cortical lesions and ipsolateral anhidrosis with brainstem stroke; preganglionic involvement.

Spinal cord injury (SCI); syringomyelia; demyelinating myelopathy

Global anhidrosis with cervical cord level; segmental loss below level with thoracic lesion, little or no anhidrosis with lumbar complete cord lesions; preganglionic involvement.

Parkinson’s disease, progressive supranuclear palsy

Often sweating is normal; distal loss in feet and regional loss in lower extremities can occur and is usually a postganglionic lesion.

Dysautonomia of advanced age

Regional affecting proximal extremities, lower trunk; both pre and postganglionic and sweat gland involvement.

PERIPHERAL NERVOUS DISORDERS Diabetic neuropathy Primary systemic and familial amyloidosis Hereditary sensory and autonomic neuropathy

Distal (length dependant) postganglionic anhidrosis most common; focal and radicular (dermatomal) loss with truncal and radiculoplexus neuropathy. Segmental (head and neck) unilateral lower limb and global can occur; percent of anhidrosis correlates positively with degree of dysautonomia; usually a postganglionic lesion.

Guillain–Barre syndrome; Lambert–Eaton myasthenic syndrome (LEMS)

Can be global without acral sparing; regional, segmental and distal occur; postganglionic; partial recovery not unusual.

Vincristine, propafenone, heavy metal, uremic, nutritional

Distal (length dependant) anhidrosis; postganglionic.

idiopathic small fiber neuropathies Connective tissue diseases

Focal, multifocal, dermatomal anhidrosis (forearm common); postganglionic axonal and skin involvement.

Tangier and Fabry’s diseases

Distal, focal, multifocal; segmental affecting head and upper extremities; postganglionic except preganglionic in Tangier’s and sweat gland in Fabry’s.

Leprosy

Multifocal (affecting cooler areas of body); scattered islands of anhidrosis to widespread distal, postganglionic sweat loss depending on type.

IATROGENIC CAUSES Drug induced: (phenothiazines, butyrophenones, tricyclic antidepressants, anticholinergic-anti-parkinson drugs, nicotinic and muscarinic anticholinergics)

Block central and peripheral autonomic pathway receptors; antimuscarinics can reduce QSART responses for up to 48 hours, depending on half-life, dose TST shows mild, generalized, symmetrically reduced sweating.

Surgical sympathectomy/sympathotomy

Segmental loss – pre and postganglionic; compensatory hyperhidrosis elsewhere may occur.

Botulinum toxin injections

Rounded, sharply demarcated lesions, postganglionic-sweat gland involvement.

CUTANEOUS DISORDERS Cholinergic urticaria

Focal loss/postganglionic or sweat gland.

Psoriasis and miliaria rubra

Focal loss related to skin-sweat duct inflammation, blockage. (Continued)

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TABLE 115.6 (Continued) Clinical Disorder

Pattern/site of Lesion Based on TST/QSART Data

Hypohidrotic ectodermal dysplasia

Scattered areas to global anhidrosis; absence of sweat glands.

Radiation injury

Sharp bordered (often rectangular), areas of anhidrosis conforming to radiation ports; affects sweat glands.

MSA-P, Multiple system atrophy with Parkinsonian features; MSA-C, Multiple system atrophy with cerebellar dysfunction; AchR, acetylcholine receptor; QSART, Quantitative Sudomotor Axon Reflex Test.

TABLE 115.7 Heat Intolerance and Heat Stroke Management Management of Heat Intolerance

Heat Stroke Management

Avoid exercise in hot/humid conditions. Plan outdoor activities for early am or late pm.

Heat stroke is a medical emergency!

Wear head covering and loose, lightweight clothing outdoors.

Diagnosis: CBT  41 degree C. (105.8°F), central nervous system disturbances (incoordination, confusion, coma), ashen or pink skin.

Stay well hydrated (2–2.5 liters of fluid/day) water and electrolyte “sport-drinks”; an aspirin can lower core balance point temperature of sweat onset.

Immediately remove from hot environment, remove clothing.

Avoid alcohol, diuretics, caffeine, benzodiazepines, barbiturates, anticholinergic and neuroleptic drugs

Circulatory collapse, seizures, hypoxia are common; tracheal intubation, intravenous isotonic fluids via central line, IV anticonvulsants (diazepam 5–10 mg doses) may be necessary.

Gradually acclimate to a hot environment over weeks before attempting strenuous activity outdoors

Surface cooling using tepid water sponging and fanning is preferred method to cool core temperature to 39.0°C. Massaging skin with ice bags or ice-water immersion are less preferred methods.

Cease any activity and cool down if heat edema, heat syncope, heat cramps or heat exhaustion occur

Avoid shivering, use IV diazepam if shivering occurs. Give 5% dextrose in water in IV fluids as hypoglycemia is common; be prepared for hospital evacuation to treat disseminated intravascular coagulation, prolonged shock, renal failure, rhabdomyolysis. Phenothiazines and dantrolene have been advocated to reduce core temperature.

CBT, core body temperature.

thermoregulatory sweating, whereas local skin trophic changes occur with chronic postganglionic sudomotor neuropathy. Observing some preventative guidelines can lessen heat prostration and dangerous hyperthermia with heat stroke can be successfully treated. Table 115.7 provides the therapeutic measures for these clinical situations.

[3] Fealey RD. Thermoregulatory sweat test. In: Low B, Benarroch EE, editors. Clinical autonomic disorders (3rd ed.). Philadelphia, PA: Lippincott Williams & Wilkins; 2008. p. 244–63. [Chapter 18] [4] Fealey RD, Sato K. Disorders of the eccrine sweat glands and sweating. In: Wolff MLAGKlaus, Katz Stephen I, Gilchrest Barbara, Paller Amy S, Leffell DJ, editors. Fitzpatrick’s dermatology in general medicine. New York: McGraw-Hill; 2008. p. 1–27. [Section 14; Chapter 82] [5] Illigens BM, Gibbons CH. Sweat testing to evaluate autonomic function. Clin Auton Res 2009;19(2):79–87.

References [1] Fealey RD. Thermoregulatory failure. The autonomic nervous system II. O. Appenzeller, 75. Amsterdam: Elsevier; 2000. [Chapter 2: 53–84] [2] Cheshire WP, Freeman R. Disorders of sweating. Semin Neurol 2003;23(4):399–406.

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116 Male Erectile Dysfunction Douglas F. Milam Sexual dysfunction is a common, almost expected, consequence of autonomic dysfunction. Sexual dysfunction may present with decreased erectile rigidity in the man, decreased libido, absent or delayed orgasm in either sex, or rapid ejaculation. Here we focus on decreased erectile rigidity. Impotence is classically defined by inability to attain penile rigidity sufficient for vaginal penetration or inability to maintain rigidity until ejaculation. Individuals with autonomic dysfunction are markedly more prone to develop impotence than those without autonomic dysfunction. Of men in the population at large, at age 40, approximately 5% never have penile rigidity sufficient for vaginal penetration [1]. By age 70, at least 15% of men experience complete erectile dysfunction while approximately 50% have varying degrees of erectile dysfunction. Age and physical health are the most important predictors of the onset of erectile dysfunction. Smoking was the most important lifestyle variable and erectile dysfunction did not correlate with male hormone levels. These figures contrast sharply with the prevalence of impotence in the autonomic dysfunction population. Patients with Parkinson’s disease and MSA both have a high rate of impotence. Impotence is a common early finding in MSA, while Parkinson’s patients often develop impotence and other urologic problems such as bladder overactivity later in their disease [2,3]. Singer et al., in a population of older Parkinson’s patients demonstrated that 60% of men were affected compared to 37.5% of age matched controls [4]. More recently, Gao et al., using the 51,529 participant Health Professionals Follow-up Study, published that 14 years into the study, the prevalence of erectile dysfunction (ED) in men with Parkinson’s disease was 68.8% compared to 31.2% of men not having a diagnosis of Parkinson’s [5]. They also demonstrated that men with ED at the beginning of the study were 3.8 times more likely to develop Parkinson’s during the study period than men without ED, a finding that raises questions about whether more subtle ED presenting earlier could have been missed in smaller studies. Beck et al. evaluated 62 patients with MSA for impotence. Their data indicate that 96% of the men were impotent and that 37% appeared to have impotence as the initial symptom of autonomic dysfunction [6]. Other studies have demonstrated similar results [7,8].

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00116-5

MECHANISM OF ERECTION Sexual thoughts from the cerebral cortex, nocturnal cortical stimuli during REM sleep, and tactile sexual stimulation may trigger penile erection. Cortical areas involved in sexual function include the medial preoptic area, medial amygdala, paraventricular nucleus, periaqueductal gray, and the ventral tegmentum [9–11]. Dopaminergic and oxytocinergic neurons project into central and spinal nerve structures. Nerve signals are carried through the pelvic plexus, a portion of which condenses into the cavernous nerves of the penile corpora cavernosa. The pelvic plexus receives input from both the sympathetic and parasympathetic nervous system. Sympathetic fibers originate in the thoracolumbar (T12–L2) spinal cord; condense into the hypogastric plexus located immediately below the aortic bifurcation and course into the pelvic plexus. Parasympathetic fibers originate in sacral spinal cord segments 2–4 and join the pelvic plexus. Discrete nerves carrying both sympathetic and parasympathetic fibers innervate the organs of the pelvis. Nonadrenergic noncholinergic (NANC) nerves follow the pelvic plexus and secrete nitric oxide (NO) into the neuromuscular junction with cavernous artery smooth muscle [12,13]. In 1982 Walsh and Donker demonstrated that nerves coursing posterolateral to the seminal vesicles and prostate and immediately lateral to the membranous urethra continue on to innervate the corpora cavernosa [14]. It is now known that branches of those NANC nerves are the principal innervation of the neuromuscular junction where arterial smooth muscle controls penile blood flow. Sexual stimulation causes release into the cavernous neuromuscular junction of a number of neurotransmitters from cholinergic parasympathetic and NANC fibers. From the standpoint of erectile rigidity, nitric oxide NO from NANC nerves appears to be the principle trigger (Fig. 116.1). The erectile pathway begins with NO release from NANC nerves which increases corporal artery blood flow and begins erectile elongation. Adequate rigidity for vaginal penetration also requires substantial NO production from vascular endothelium lining the trabecula of the corpora cavernosa. This produces the feedback loop shown in Figure 116.1. NO activates guanylyl cyclase which converts guanosine-5-triphosphate (GTP) into cyclic Guanosine monophosphate (cGMP).

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Protein Kinase G

PDE-5

5-GMP

GTP

cGMP

Proteins

Guanylyl Cyclase

NO

Decreased Smooth Muscle Ca++

NO

nNOS

Protein-P

Cavernous Artery Smooth Muscle Relaxation eNOS

NANC Cavernous Nerve

Cavernous Vascular Endothelium

Increased Cavernous Blood Flow

Penile Erection

Sexual Stimulation FIGURE 116.1 Erectile pathway.

Protein kinase G (PKG) is activated by cGMP and in turn activates several proteins which decrease intracellular calcium (Ca2) concentration. Decreased smooth muscle Ca2 concentration causes muscular relaxation, cavernosal artery dilation, increased blood flow and subsequent penile erection. The control of blood flow on the venous outflow side is less well understood.

ETIOLOGY OF ERECTILE DYSFUNCTION The anatomical site felt to be the most common cause of erectile dysfunction in the general population is the neuromuscular junction where the NANC nerves meet the smooth muscle and vascular endothelium of the deep cavernous penile arteries. This is where nitric oxide and cGMP play a critical role in regulating penile blood flow [15]. The typical erectile dysfunction (ED) patient secretes less than the normal amount of NO into the neuromuscular junction. Neurologic disease can produce discrete lesions in central or peripheral nerves which cause erectile dysfunction by altering upstream nerve function, or can alter NO production at the neuromuscular junction and vascular endothelium. In particular, Parkinson’s disease, multiple system atrophy (MSA), multiple sclerosis, and processes affecting the spinal cord can produce decreased erectile rigidity, failure of emission, or retrograde ejaculation. Other causes of impotence include drug induced erectile dysfunction, endocrine disorders, vascular disease and venogenic erectile dysfunction.

NEUROGENIC ERECTILE DYSFUNCTION Pure neurogenic erectile dysfunction is a frequent cause of erectile failure. Interruption of either somatic

or autonomic nerves or their end units may cause erectile dysfunction. These nerves control the flow of blood into and likely out of the corpora cavernosa. Afferent somatic sensory signals are carried from the penis via the pudendal nerve to sacral segments 2–4. This information is routed both to the brain and to spinal cord autonomic centers. Parasympathetic autonomic nerves originate in the interomediolateral gray matter of sacral segments 2–4 [3]. These preganglionic fibers exit the anterior nerve roots to join with the sympathetic fibers of the hypogastric nerve to form the pelvic plexus and cavernosal nerves. The paired cavernosal nerves penetrate the corpora cavernosa and innervate the cavernous artery. Parasympathetic ganglia are located distally near the end organ. Sympathetic innervation also originates in the intermediolateral lateral gray matter but at thoracolumbar levels T10–L2. Sympathetic efferents course through the retroperitoneum and condense into the hypogastric plexus located anterior and slightly caudal to the aortic bifurcation. A concentration of post ganglionic sympathetic fibers forms the hypogastric nerve which is joined by parasympathetic efferents. Adrenergic innervation plays an important role in detumescence. High concentrations of norepinephrine have been demonstrated in the tissue of the corpora cavernosa and tributary arterioles. Additionally, the alpha-adrenergic antagonist phentolamine is routinely utilized for intracorporal injection therapy to produce erection. Afferent signals capable of initiating erection can either originate within the brain, as is the case with psychogenic erections, or result from tactile stimulation. Patients with spinal cord injury often respond to tactile sensation, but usually require medical therapy to maintain the erection through intercourse.

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MEDICAl AnD suRgICAl TREATMEnT

HYPOGONADISM Testosterone plays a permissive role in erectile function, but erectile response is not correlated with hormone level in patients within the wide range of normal. Androgen replacement with testosterone cipionate 200 mg q2–3 weeks, daily topical testosterone preparations, or long acting pellets are expected to induce return of erectile function in patients with very low or undetectable serum total testosterone concentrations due to hypogonadism. These patients are relatively uncommon, however. Most have pituitary or hypothalamic tumors. More commonly, the impotent patient will have normal or mildly decreased free or total testosterone levels. Testosterone replacement rarely restores erectile function in those patients and should not be routinely given for that indication. Testosterone supplementation is never indicated for patients with normal circulating androgen levels.

MEDICAL AND SURGICAL TREATMENT Objective assessment of erectile function is best done using validated patient questionnaires. The International Index of Erectile Function (IIEF-15) and a subset of the IIEF-15 questionnaire termed the Sexual Health Inventory for Men (SHIM) are particularly helpful [16,17]. We find the SHIM to be useful in routine practice as it has only five questions and has a convenient 1 to 25 scale. One important practical limitation of the SHIM is that patients need to be actively trying to have sex, otherwise the score will be disproportionately low. Most patients perceive a component of erectile dysfunction with SHIM scores below 20. Most sexually active patients desire treatment when the SHIM score is 16 or below, though many opt for treatment earlier. First line medical therapy for decreased erectile rigidity is based on inhibition of phosphodiesterase type-5 (PDE-5) [18]. Figure 116.1 illustrates that cGMP is broken down to inactive 5-GMP by PDE-5. Sildenafil, vardenafil, and tadalafil competitively inhibit PDE-5 breakdown of cGMP by binding to the catalytic domain of PDE-5. Use of a PDE-5 inhibitor results in improved erectile rigidity even in patients with decreased nitric oxide production or cGMP synthesis. Generally, 60% of patients overall, 80% with ED due to spinal cord injury respond to PDE-5 inhibition. Safarinejad et al. administered sildenafil 100 mg to 236 Parkinson’s disease patients in a randomized, doubleblind, placebo-controlled study [2]. A normal erectile function domain score at end point was reported in 56.9% of the sildenafil treated group and by 8.7% of patients on placebo. Hussain et al. examined the response to sildenafil in patients with Parkinson’s disease and MSA [19]. Parkinson’s disease patients experienced improved erectile rigidity similar to patients with idiopathic ED. None of the Parkinson's patients experienced significant orthostatic hypotension. One hour after medication, the standing

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mean blood pressure dropped 9 mmHg, compared to 6 mmHg in normal volunteers. Such was not the case for MSA patients, however. Six patients had been enrolled before the study was halted due to profound orthostatic hypotension. Three patients who had stable blood pressures at study entry experienced standing blood pressure drops of 128/85 to 65/55, 104/60 to 56/32, and 115/70 to 55/39 one hour after taking medication. PDE-5 inhibitors should be used very cautiously in patients with MSA. Profound hypotension with PDE-5 inhibition raises a number of questions. Gamboa et al. evaluated 20 patients with autonomic failure [20]. Somewhat to their surprise, the autonomic failure patients in their study, pure autonomic failure and MSA, had increased NO function. This may explain the marked hypotension with PDE-5 inhibition. Other treatment options include intracorporal injection (ICI) of prostaglandin E-1, papaverine, or phentolamine directly into the corpora cavernosa, intraurethral delivery of prostaglandin E-1, vacuum erection devices, and inflatable penile implants. ICI is an attractive second line therapy as smooth muscle dilators injected into the corpora directly bathe the corporal arteries causing increased blood flow and penile erection. Systemic side effects are rare. PGE-1 for instance, is metabolized first pass through the lungs and never reaches arterial systemic circulation. Inflatable penile implants have the second highest overall patient satisfaction rate, behind only PDE-5 inhibition in responsive patients. Individuals with ED and good overall performance status who desire sexual intercourse should be offered urologic referral if initial medical therapy fails to achieve adequate penile rigidity.

References [1] Feldman HA, Goldstein I, Hatzichristou DG, et al. Impotence and its medical and psychosocial correlates: results of the massachusetts male aging study. J Urol 1994;151:54–61. [2] Safarinejad MR, Taghva A, Shekarchi B, Safarinejad SH. Safety and Efficacy of sildenafil citrate in the treatment of Parkinson-emergent erectile dysfunction: a double-blind placebo-controlled, randomized study. Int J Impotence Res 2010;22:325–35. [3] Sakakibara R, Uchiyama T, Yamanishi T, Kishi M. Genitourinary dysfunction in Parkinson’s disease. Mov Disord 2010;25:2–12. [4] Singer C, Weiner WJ, Sanchez-Ramos JR, et al. Sexual dysfunction in men with Parkinson’s disease. J Neurol Rehab 1989;3:199–204. [5] Gao X, Chen H, Schwarzschild MA, Glasser DB, Logroscino G, Rimm EB, et al. Erectile function and risk of Parkinson’s disease. Am J Epidemiol 2007;166:1446–50. [6] Beck RO, Betts CD, Fowler CJ. Genitourinary dysfunction in multiple system atrophy: clinical features and treatment in 62 cases. J Urol 1994;151:1336–41. [7] Wenning G, Shlomo Y, Magalhaes M, et al. Clinical features and natural history of multiple system atrophy. Brain 1994;117:835–45. [8] Sakakibara R, Hattori T. Uchiyama, et al: Urinary dysfunction and orthostatic hypotension in multiple system atrophy: which is the more common and earlier manifestation? J Neurol Neurosurg Psychiatry 2000;68:65–9. [9] Chen KK, Chan SH, Chang LS, Chan JY. Participation of paraventricular nucleus of hypothalamus in central regulation of penile erection in the rat. J Urol 1997;158:238–44.

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116. MAlE ERECTIlE DysfunCTIon

[10] Giuliano F, Rampin O, Brown K, Courtois F, Benoit G, Jardin A. Stimulation of the medial preoptic area of the hypothalamus in the rat elicits increases in intracavernous pressure. Neurosci Lett 1996;209:1–4. [11] Heaton JP. Central neuropharmacological agents and mechanisms in erectile dysfunction. Neurosci Biobehav 2000;24:561–9. [12] Andersson KE, Wagner G. Physiology of erection. Physiol Rev 1995;75:191–236. [13] Gratzke C, Angulo J, Chitaley K, Dai Y, Kim NN, et al. Anatomy, physiology, and pathophysiology of erectile dysfunction. J Sex Med 2010;7:445–75. [14] Walsh PC, Donker PJ. Impotence following radical prostatectomy: insight into etiology and prevention. J Urol 1982;128:492. [15] Kim N, Azadzoi KM, Goldstein I, et al. A nitric oxide-like factor mediates nonadrenergic, noncholinergic neurogenic relaxation of penile corpus cavernosum smooth muscle. J Clin Inves 1991;88:112. [16] Rosen RC, Riley A, Wagner G, et al. The international index of erectile function (IIEF): a multidimensional scale for assessment of erectile dysfunction. Urology 1997;49:822–30.

[17] Rosen RC, Cappelleri JC, Smith MD, Lipsky J, Peña BM. Development and evaluation of an abridged, 5-item version of the International Index of Erectile Function (IIEF-5) as a diagnostic tool for erectile dysfunction. Int J Impotence Res 1999;11:319–26. [18] Turko IV, Ballard SA, Francis SH, Corbin JD. Inhibition of cyclic GMP-binding cyclic GMP-specific phosphodiesterase (Type 5) by sildenafil and related compounds. Mol Pharmacol 1999;56:124–30. [19] Hussain IF, Brady CM, Swinn MJ, Mathias CJ, Fowler CJ. Treatment of erectile dysfunction with sildenafil citrate (Viagra) in parkinsonism due to Parkinson’s disease or multiple system atrophy with observations on orthostatic hypotension. J Neurol Neurosurg Psych 2001;71:371–4. [20] Gamboa A, Shibao C, Diedrich A, Paranjape SY, Farley G, Cristman B, et al. Excessive nitric oxide function and blood pressure regulation in patients with autonomic failure. Hypertension 2008;51:1531–636.

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117 Sleep Apnea Tomas Konecny, Virend K. Somers AUTONOMIC FUNCTION DURING PHYSIOLOGIC SLEEP

OBSTRUCTIVE SLEEP APNEA

Normal human sleep can be divided into rapid eye movement sleep (REM) characterized by desynchronized EEG signals, muscle atony, and dreaming, and non-rapid eye movement (NREM) sleep characterized by synchronous EEG patterns [1]. Humans begin sleep with a transition from wakefulness into non-rapid eye movement (NREM) sleep, progress deeper through the NREM stages, and then reach the first episode of rapid eye movement (REM) sleep. Cycles of NREM and REM sleep which last approximately 90 minutes then continue during the remainder of the night. Physiologic NREM sleep is accompanied by a progressive increase in parasympathetic tone and decrease in sympathetic activity, together manifesting as a decrease in heart rate (HR), blood pressure (BP), stroke volume, cardiac output, peripheral vascular resistance, and minute ventilation (Fig. 117.1) [2]. Refractory period during NREM increases, and premature ventricular contractions (PVC) occur less frequently [3]. This decline in nighttime PVCs predicts the success of β-blockers in suppressing PVCs during the day, further supporting the notion that modulation of sympathetic nervous system activity is involved [4]. During physiologic REM sleep (which accounts for only 25% of total sleep time), breathing and HR become more irregular, and depend on the occurrences of phasic REM sleep events [5]. Mainly thanks to the NREM portion, normal sleep can be seen as a period of cardiovascular relaxation; however, sleep apnea with its autonomic consequences may change sleep into a time of increased strain [6]. (For additional discussion see Chapter 32: Circadian Rhythms and Autonomic Function.)

Obstructive sleep apnea (OSA) constitutes a highly prevalent sleep breathing disorder affecting an estimated 15 million adult Americans (at least mild OSA is found in approximately one in five adults) [7]. Its mechanism lies in the collapse of the pharyngeal airway from decreased muscle tone, which leads to partial or complete obstruction, and resultant decrease or cessation of airflow as seen in Figure 117.2 (complete cessation 10 seconds defines an apnea, while decreased ventilation with accompanying drop in peripheral oxygen saturation defines a hypopnea). The ineffective respiration during apneic and hypopneic episodes leads to compensatory large recovery breaths, referred to as hyperpnea. Severity of OSA can be expressed by the apnea hypopnea index (AHI) which describes an average number of apneic or hypopneic episodes per hour of sleep. An AHI of 5 is considered abnormal.

OSA AND THE AUTONOMIC NERVOUS SYSTEM Acute Changes During Apneic Episodes Cardiovascular responses to OSA can be explained in part by the diving reflex, a reflex response to prolonged breath hold, during which simultaneous increase in parasympathetic activity to the heart and sympathetic activity to the periphery lead to concomitant bradycardia as well as increased peripheral arterial resistance [8]. These together aim at decreasing myocardial oxygen consumption during times of low oxygen supply (most pronounced in sea mammals, and elite human divers). As seen in Figure 117.3, hopoxemia and apnea elicit both bradycardia

Abbreviations: BP Blood pressure; CSA Central sleep apnea; HR Heart rate; HRV Heart rate variability; NREM Non-rapid eye movement; OSA Obstructive sleep apnea; PVC Premature ventricular contraction; REM Rapid eye movement; SCD Sudden cardiac death; SNA, Sympathetic neural activity.

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117. SlEEP APnEA

Stage 4

Awake

SNA SNA

125

125

BP 0

0 Stage 2

REM

SNA SNA

125 BP 0

125 BP

Stage 3

0

SNA

10 sec

125 BP 0

FIGURE 117.1 Blood pressure (BP) and sympathetic nerve activity (SNA) in one subject during wakefulness, stages 2, 3, and 4 of non-rapid eye movement (NREM) sleep, and during rapid eye movement (REM) sleep. Reproduced with permission from Somers VK, Dyken ME, Mark AL, Abboud FM. Sympathetic-nerve activity during sleep in normal subjects. New England Journal of Medicine 1993;328(5):303–307) [2]

Air flow Tongue

Nasal Passage Soft palate Uvula

Hypopnea

Apnea

FIGURE 117.2 Schematic showing normal breathing during sleep in a supine position (above), reduced airflow through the partially collapsed airway during hypopnea (left below), and the full closure of airways leading to apnea (right below). Reprinted from Hahn PY, Somers VK. Sleep apnea and hypertension. In: Lip GYH, Hall JE, eds. Comprehensive Hypertension. St. Louis, MO: Mosby; 2007:201–207. Copyright Elsevier 2007. Used with permission.

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OSA And THE AuTOnOmIC nERvOuS SySTEm

FIGURE 117.3 (A) Recordings during sleep of patients with obstructive sleep apnea (OSA). Sympathetic neural activity (SNA) increases during the apneic episode and peaks at the time of the release. Blood pressure (BP) and heart rate (HR) decrease during the apnea but dramatically increase after the release of the obstruction. Adapted with permission from Somers VK, Dyken ME, Clary MP, Abboud FM. Sympathetic neural mechanisms in obstructive sleep apnea. Journal of Clinical Investigation. Oct 1995;96(4):1897–1904 [9]. (B) Recordings showing sinus arrest (absence of p waves on ECG) during OSA. Adapted with permission from Somers VK, Dyken ME, Mark AL, Abboud FM. Parasympathetic hyperresponsiveness and bradyarrhythmias during apnoea in hypertension. Clin Auton Res 1992;2: 171–176 [20]

and a gradual increase in peripheral sympathetic neural activity (SNA). SNA reaches maximum during the release of obstruction. Subsequently, both BP and HR dramatically increase, which coincides temporally with the hyperpneic compensatory breathing [9].

Chronic Changes with OSA Given the surges of SNA occurring with each apneic episode it is not surprising that patients with OSA manifest higher BP and HR during sleep compared to the nocturnal dip in HR and BP described in normal subjects. This autonomic derangement persists also during the daytime (Fig. 117.4), and may contribute to the development of hypertension and other cardiovascular morbidity in OSA patients [7]. Severity of OSA correlates with 24-hour BP recordings, and in a large prospective study from Wisconsin, OSA emerged as an independent predictor of essential hypertension [10]. Treatment of OSA by continuous positive airway pressure (CPAP) reduces SNA, seems to reduce overnight urinary catecholamine levels,

and may lower daytime BP in patients with resistant hypertension [6]. Whether CPAP treatment also improves morbidity and mortality is still yet to be established, but some patient groups with known high prevalence of OSA and already existing cardiovascular morbidity – such as patients after myocardial infarction – could benefit from more rigorous OSA screening [11].

Atrial Fibrillation and OSA The evidence linking atrial fibrillation to OSA is strong, and autonomic dysregulation is often postulated as a potential contributing mechanism. The incidence of atrial fibrillation has been independently correlated with the severity of nocturnal hypoxia in patients 65 years old [12]. Effort to decipher the exact network of interractions between atrial refractoriness, ganglionated plexi, pulmonary veins and the various parts of the intrinsic cardiac nervous system are still under way [13]. A recent study on dogs provides strong support to the notion that apnea related atrial fibrillation can be prevented by autonomic blockade via ablation

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568

117. SlEEP APnEA

(A) Control

AWAKE NORMAL

1 second

II

OSA

Hb GP neurons 200

0.5mV

BP 0 (B) 15 secs. Apnea II 10 sec Hb

FIGURE 117.4 Recordings of sympathetic nerve activity (SNA) during wakefulness in patients with obstructive sleep apnea matched controls showing high levels of SNA in patients apnea. Reproduced with permission from Somers VK, Dyken MP, Abboud FM. Sympathetic neural mechanisms in obstructive Journal of Clinical Investigation 1995;96(4):1897–1904 [9]

(OSA) and with sleep ME, Clary sleep apnea.

GP neurons 200

0.5mV

BP

of the right pulmonary artery ganglionated plexus [14]. Recordings from one of the experimental animals depict the onset of apnea-related atrial fibrillation (Fig. 117.5). The findings of this study confirm that the autonomic nervous system plays a key part in the adverse cardiovascular effects of OSA. In patients with hypertrophic cardiomyopathy, in whom atrial fibrillation is seen as one of the leading causes of morbidity and mortality, OSA and its severity positively correlate both with the presence of atrial fibrillation as well as with the left atrial size [15].

0 (C) 1min 36 secs. Apnea-Spontaneous AF

1 second

II

Hb Gb neurons

0.5mV

300

Sudden Cardiac Death and OSA

BP

OSA could theoretically act as a potentially lethal nocturnal stressor in susceptible patients. More than half of patients with a diagnosis of OSA suffer a fatal cardiac event between 10 pm and 6 am, which is in stark contrast to those without OSA who are more likely to die suddenly between 6 am and 11 am. Ventricular arrhythmias seem to be more common among OSA patients, and in most OSA patients these tend to occur most often during sleep [7]. As in the case of atrial fibrillation, the exact mechanism of this phenomenon is difficult to discern, and autonomic nervous system likely serves as one of the links.

Heart Rate Variability in OSA Studies focusing on heart rate variability (HRV) in OSA report reduction in high frequency power of HRV, while the low frequency power is increased. This suggests a decrease in parasympathetic and increase in sympathetic

0

FIGURE 117.5 Spontaneous onset of atrial fibrillation during induced apnea in dogs. Panel B shows decreased neural firing from the anterior right ganglionated plexus during induced apnea, compared to a control recording in panel A. Panel C then depicts the onset of apnea induced atrial fibrillation. Reproduced with permission from Ghias M, Scherlag BJ, Lu Z, Niu G, Moers A, Jackman WM, Lazzara R, Po SS. The role of ganglionated plexi in apnea-related atrial fibrillation. Journal of the American College of Cardiology 2009;54(22):2075–2083 [14]

modulation of HR in patients with OSA [6]. Recording of respiration is an important variable that should be added to studies of HRV because high frequency HRV is mostly a function of respiratory sinus arrhythmia, which could be affected by the breathing pattern in OSA. Treatment with CPAP restores HRV indices towards normal [16].

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CEnTRAl SlEEP APnEA

CENTRAL SLEEP APNEA The pathophysiology of central sleep apnea (CSA) lies in the decrease of ventilation during sleep as a result of decreased respiratory drive (unlike in OSA where respiratory drive is normal and the obstructed airway constitutes the problem). CSA can be defined as a pause in ventilation lasting 10 seconds with no associated respiratory effort [7]. More than five such episodes per hour are abnormal. Cheyne–Stokes respiration, which can be classified as a form of CSA, includes alternating apneic and hyperpneic phases during which the respiratory tidal volume has a crescendo–decrescendo type of pattern.

Heart Failure and CSA The association of heart failure (HF) with CSA has been clearly established [7]. Resting sympathetic activity – already elevated in patients with HF – is even higher in those with concomitant CSA and HF [17]. Nocturnal rostral fluid shift could contribute to the pathogenesis of CSA (and even OSA) in HF patients, and since the autonomic nervous system could be playing a role in this interaction, further studies focusing on this phenomenon are needed [18]. Studies of HRV in CSA patients show an overall lower HRV, especially in the high frequency band. Parasympathetic modulation is reduced in severe CSA patients both nocturnally, and during a 24-hour recording [19]. An increased attention to the abnormal respiratory patterns has to be paid when interpreting the HRV results as these patterns could potentially influence HRV independent of changes in autonomic regulation (similarly to that described in the OSA patients) [6].

Acknowledgments Supported by grants from Ministry of Health NoNS10098-4/2008 and by the European Regional Development Fund–Project FNUSA-ICRC (No. C.Z.1.05/1.1.00/02.0123).

References [1] Chatterjee K. Cardiology – an illustrated textbook. Section 15: Evolving concepts. 2011. [2] Somers VK, Dyken ME, Mark AL, Abboud FM. Sympatheticnerve activity during sleep in normal subjects. New Engl J Med 1993;328:303–7. [3] Kong Jr TQ, Goldberger JJ, Parker M, Wang T, Kadish AH. Circadian variation in human ventricular refractoriness. Circulation 1995;92:1507–16.

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[4] Pitzalis MV, Mastropasqua F, Massari F, Totaro P, Scrutinio D, Rizzon P. Sleep suppression of ventricular arrhythmias: A predictor of beta-blocker efficacy. Eur Heart J 1996;17:917–25. [5] Snyder F, Hobson JA, Morrison DF, Goldfrank F. Changes in respiration, heart rate, and systolic blood pressure in human sleep. J Appl Physiol 1964;19:417–22. [6] Leung RS. Sleep-disordered breathing: Autonomic mechanisms and arrhythmias. Prog Cardiovasc Dis 2009;51:324–38. [7] Somers VK, White DP, Amin R, Abraham WT, Costa F, Culebras A, et al. Sleep apnea and cardiovascular disease: An American Heart Association/American College Of Cardiology Foundation Scientific Statement from The American Heart Association Council for High Blood Pressure Research Professional Education Committee, Council on Clinical Cardiology, Stroke Council, and Council on Cardiovascular Nursing. J Am Coll Cardiol 2008;52:686–717. [8] Gooden BA. Mechanism of the human diving response. Integr Physiol Behav Sci 1994;29:6–16. [9] Somers VK, Dyken ME, Clary MP, Abboud FM. Sympathetic neural mechanisms in obstructive sleep apnea. J Clin Invest 1995;96:1897–904. [10] Peppard PE, Young T, Palta M, Skatrud J. Prospective study of the association between sleep-disordered breathing and hypertension. N Engl J Med 2000;342:1378–84. [11] Konecny T, Kuniyoshi FH, Orban M, Pressman GS, Kara T, Gami A, et al. Under-diagnosis of sleep apnea in patients after acute myocardial infarction. J Am Coll Cardiol. 56:742–743 [12] Gami AS, Hodge DO, Herges RM, Olson EJ, Nykodym J, Kara T, et al. Obstructive sleep apnea, obesity, and the risk of incident atrial fibrillation. J Am Coll Cardiol 2007;49:565–71. [13] Hou Y, Scherlag BJ, Lin J, Zhang Y, Lu Z, Truong K, et al. Ganglionated plexi modulate extrinsic cardiac autonomic nerve input: Effects on sinus rate, atrioventricular conduction, refractoriness, and inducibility of atrial fibrillation. J Am Coll Cardiol 2007;50:61–8. [14] Ghias M, Scherlag BJ, Lu Z, Niu G, Moers A, Jackman WM, et al. The role of ganglionated plexi in apnea-related atrial fibrillation. J Am Coll Cardiol 2009;54:2075–83. [15] Konecny T, Brady PA, Orban M, Lin G, Pressman GS, Lehar F, et al. Interactions between sleep disordered breathing and atrial fibrillation in patients with hypertrophic cardiomyopathy. Am J Cardiol 105:1597–602. [16] Roche F, Court-Fortune I, Pichot V, Duverney D, Costes F, Emonot A, et al. Reduced cardiac sympathetic autonomic tone after longterm nasal continuous positive airway pressure in obstructive sleep apnoea syndrome. Clin Physiol (Oxford, England) 1999;19:127–34. [17] van de Borne P, Oren R, Abouassaly C, Anderson E, Somers VK. Effect of cheyne-stokes respiration on muscle sympathetic nerve activity in severe congestive heart failure secondary to ischemic or idiopathic dilated cardiomyopathy. Am J Cardiol 1998;81:432–6. [18] Yumino D, Redolfi S, Ruttanaumpawan P, Su MC, Smith S, Newton GE, et al. Nocturnal rostral fluid shift: A unifying concept for the pathogenesis of obstructive and central sleep apnea in men with heart failure. Circulation 121:1598–605. [19] Lanfranchi PA, Braghiroli A, Bosimini E, Mazzuero G, Colombo R, Donner CF, et al. Prognostic value of nocturnal cheyne-stokes respiration in chronic heart failure. Circulation 1999;99:1435–440. [20] Somers VK, Dyken ME, Mark AL, Abboud FM. Parasympathetic hyperresponsiveness and bradyarrhythmias during apnoea in hypertension. Clin Auton Res 1992;2:171–6.

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118 Altered Adrenal Function and the Autonomic Nervous System Subbulaxmi Trikudanathan, Gordon H. Williams The adrenal gland consists of the outer cortex and the inner medulla. The adrenal cortex consists of the outer zona glomerulosa that secretes aldosterone which plays a major role in regulation of blood pressure, volume and potassium balance. The middle layer of the adrenal cortex, the zona fasiculata, is where cortisol is synthesized. Cortisol is responsible for mounting a stress response, modulates immune functions and intermediary metabolism. The inner cortical layer, the zona reticularis, produces adrenal androgens that primarily play a role in the development of secondary sexual characteristics in women. Hypoadrenocorticism can occur via one of two mechanisms: primary adrenal insufficiency that almost invariably leads to the loss of all three steroid types and secondary adrenal insufficiency that leads to the selective loss of one type. The most common secondary form is caused by the lack of pituitary adrenocorticotrophin hormone (ACTH) leading to cortisol deficiency. A second form is hyporeninemic hypoaldosteronism.

CROSSTALK BETWEEN ADRENAL CORTEX AND MEDULLA The adrenal cortex and medulla appear to be interlaced with multiple areas of contact that are not separated by connective tissue or interstitial membranes [1]. After cortisol is produced by the zona fasiculata it travels through the portal venous system to reach the medullary regions of the adrenal gland. The interstitial fluid around the medullary cells equilibrates with the venous blood creating high concentrations of cortisol within the adrenal medulla. This elevated cortisol concentration is required for the methylation of norepinephrine to epinephrine by the enzyme phenylethanolamine-N-methyltransferase (PNMT) [2–4]. Patients with adrenal insufficiency, either primary or secondary, causing cortisol deficiency, may have impaired catecholamine synthesis suggesting that there is crosstalk between the adrenal cortex and medulla [5]. The decreased epinephrine in patients with adrenal insufficiency makes them more susceptible to fasting or insulin induced hypoglycemia. Furthermore, cortisol and other

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00118-9

glucocorticoids have a permissive effect in potentiating the vasoconstrictive effects of catecholamines by acting on the endothelial and vascular smooth muscle cells [6].

PRIMARY ADRENAL INSUFFICIENCY Primary adrenal insufficiency results from destruction of the adrenal cortex. Autoimmune adrenalitis remains the predominant cause for primary adrenocortical dysfunction in the western world. Autoimmune destruction of the adrenal gland can be isolated or occurs as a part of autoimmune polyglandular syndrome (APSS) with multiorgan involvement. Infections particularly tuberculosis forms the leading cause in the developing world. The other uncommon etiologies of primary adrenocortical insufficiency are listed in Box 118.1. The clinical manifestations of primary adrenal insufficiency occur as a result of cortisol and aldosterone deficiencies and in women, androgenic steroids. The impaired cortisol production stimulates the pituitary gland to hypersecrete ACTH because of a negative feedback relationship. The clinical features of adrenal insufficiency vary according to the rate and extent of adrenocortical destruction. When the adrenal dysfunction occurs slowly, as in the case of autoimmune or infiltrative diseases, then the symptoms and signs are gradual and non specific. Basal cortisol secretion is initially normal but fails to respond to stressful situations. As the adrenal cortex progressively gets destroyed the baseline production of both cortisol and aldosterone decreases leading to the clinical manifestations of chronic adrenal insufficiency. The most common symptoms include weakness, malaise, fatigue, weight loss, anorexia, and nausea. The distinctive clinical features of chronic primary adrenal insufficiency include hyperpigmentation [from the elevated ACTH, pro-opiomelanocortin (POMC) and melanocortin stimulating hormone (MSH)] and salt craving (from the decreased aldosterone secretion). Hyperpigmentation is typically seen in oral mucosa, palmar creases, nails, flexural areas, elbows and knees. Increased tanning of scars, freckles of sun exposed areas; nipples, areolae, and genital skin

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118. AlTEREd AdRENAl FuNCTIoN ANd THE AuToNomIC NERvouS SySTEm

BOX 118.1

E T I O L O G Y O F P R I M A RY A D R E N O C O RT I C A L I N S U F F I C I E N C Y

Common l l

l l

Autoimmune (80%) Tuberculosis.

l l l

Uncommon l l

l

HIV, AIDS-related opportunistic infection Metastatic carcinoma and lymphoma Surgical adrenalectomy Medications like ketoconazole, metyrapone, mitotane Amyloidosis, sarcoidosis and hemachromatosis Congenital enzyme defects.

Adrenal hemorrhage and infarction Histoplasmosis and other granulomatous infections

have also been reported. Vitiligo can occur in autoimmune form of adrenal insufficiency but seldom from other causes. Hyperkalemia and hypovolemia occurs as a result of aldosterone deficiency leading to renal sodium loss, potassium retention and volume depletion. Hyponatremia is a reflection of lack of aldosterone and the inability to excrete water load from cortisol deficiency. Salt craving has been observed in 20% of patients with adrenal insufficiency. Postural hypotension especially dizziness is frequently noted as the result of aldosterone (volume depletion) and cortisol (impaired vasoconstriction) deficiencies. Abdominal pain, vomiting and behavioral disturbances frequently make it a challenging diagnosis. If an undiagnosed person with adrenal insufficiency is subjected to major stress, i.e., infection, major illness or surgery they develop adrenal crisis.

ADRENAL CRISIS Apart from the above mentioned situation, adrenal crisis may also be precipitated when a patient with known adrenal insufficiency does not increase cortisol replacement during a major illness. Acute adrenal insufficiency can be triggered from bilateral adrenal hemorrhage or infarction especially when the patient has a coagulation disorder like antiphospholipid syndrome, or with anticoagulant therapy, or as a complication of blunt trauma. The predominant clinical feature of acute adrenal insufficiency is hypotension or shock from aldosterone deficiency. The cortisol deficiency also reduces the vascular responsiveness to vasoconstrictors. Other features include abdominal tenderness, fever, nausea, vomiting, anorexia, malaise and lethargy [7].

MANAGEMENT In a non-acute situation morning plasma cortisol should be obtained. When the morning plasma cortisol is less than 3 µg/dL (83 nmmol/L) it is virtually diagnostic

of adrenal insufficiency and if more than 19 µg/dL (525 nmol/L) rules out the disorder. A short ACTH stimulation test should be performed for intermediate values of morning cortisol or when the clinical suspicion is high. 250 µg of cosyntropin (synthetic ACTH) is administered intramuscularly or intravenously. The cortisol response is measured at 0, 30, and 60 minutes. The normal response is a basal or peak cortisol response greater than 18 µg/dL (495 nmol/L). This test is useful in diagnosing primary adrenal insufficiency or longstanding secondary adrenal insufficiency when the adrenal glands are atrophied but will be normal in patients with mild or new onset secondary adrenal insufficiency. Plasma ACTH values will be elevated (usually above 100 pg/ml) in primary adrenal insufficiency and can be low or inappropriately normal in secondary adrenal insufficiency. In acute adrenal insufficiency prompt treatment with parenteral steroids and fluid resuscitation with normal saline is vital. As adrenal crisis is a life-threatening emergency, blood for serum cortisol, ACTH should be drawn but treatment with intravenous steroids should be initiated prior to the results. The resulting hypotension is usually unresponsive to intravenous infusions of norepinephrine unless corrected with intravenous steroids. In previously undiagnosed patients short ACTH stimulation test can be done soon after, if dexamethasone is used as it is not measured in the cortisol assay. In patients with known adrenal insufficiency intravenous hydrocortisone at 100 mg thrice daily should be used. Once these patients have stabilized long-term steroid replacement can be maintained with hydrocortisone, typically 15–20 mg in the morning and 5–10 mg in the evening to mimic cortisol’s diurnal rhythm. Patients with primary adrenal insufficiency will need mineralocorticoid replacement with fludrocortisone at doses of 0.05 to 0.1 mg per day.

SECONDARY ADRENAL INSUFFICIENCY Secondary adrenal insufficiency results from lack of ACTH secretion from the pituitary gland or corticotrophin

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releasing hormone (CRH) secretion from the hypothalamus. Isolated ACTH deficiency is rare. It usually occurs along with other pituitary hormone deficiencies. The other common cause of secondary adrenal insufficiency is longterm exogenous glucocorticoid therapy which leads to suppression of hypothalamic pituitary axis. Many of the clinical manifestations are similar to primary adrenal insufficiency with a few exceptions. Hyperpigmentation is absent as ACTH production is diminished. The mineralocorticoid axis is preserved as it is not regulated by ACTH. Hence hyperkalemia and volume depletion is not seen. Hyponatremia can occur from inappropriate vasopressin secretion due to the lack of cortisol. The diagnosis is established by low cortisol paired with low or normal ACTH. Deficiencies in other pituitary hormones may also be seen. The short ACTH stimulation test may be false-negative in new onset secondary adrenal insufficiency. In secondary adrenal insufficiency usually only glucocorticoid therapy is warranted.

HYPOALDOSTERONISM Hyporeninemic hypoaldosteronism is common among diabetic patients with mild renal impairment. There is diminished renin release and hence decreased aldosterone production. While it could be a part of primary adrenal insufficiency the other causes of hyporeninemic hypoaldosteronism include drugs like nonsteroidal antiinflammatory drugs, cyclosporine, heparin, trimethoprim and pentamidine. Hypoaldosteronism commonly presents with hyperkalemia and metabolic acidosis out of proportion to the renal impairment. Therapy includes stopping offending drugs and fludrocortisone therapy. In patients with concomitant hypertension and edema, low potassium diet with loop diuretic may be necessary to control the hyperkalemia.

INFLUENCE OF THE AUTONOMIC NERVOUS SYSTEM ON ADRENOCORTICAL FUNCTION Activation of α-1 adrenoreceptors in the hypothalamus promotes the secretion of ACTH through CRH and other hypothalamic peptides. Methoxamine (α-1 agonist) crosses the blood brain barrier and stimulates ACTH secretion in humans. However norepinephrine and epinephrine when infused in ranges seen in moderate stress do not cross the blood–brain barrier and hence do not stimulate ACTH production [8]. In humans the cortisol secretory patterns in the morning and the ACTH/cortisol response to food have been shown to be mediated by α-1 stimulatory adrenergic receptors. α-2 adrenergic receptors on the other hand have an inhibitory effect on ACTH secretion when studied in rats. In humans there

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has been no evidence for the role of β adrenergic receptors on ACTH secretion. In humans peripheral circulating catecholamines do not stimulate ACTH secretion thus showing that the ACTH response to stress is not a sympathoadrenal response. Patients with idiopathic orthostatic hypotension or multisystem atrophy have impaired adrenergic responses to insulin-induced hypoglycemia but normal increase in cortisol secretion [9]. This observation indicates that the adrenocortical/cortisol secretion in response to hypoglycemia is not affected by autonomic insufficiency. On the other hand aldosterone secretion is reduced in autonomic failure as there is inadequate renin release due to impaired beta adrenergic stimulation of the juxtaglomerular apparatus. Patients with multisystem atrophy or diabetic neuropathy may have hyporeninemic hypoaldosteronism leading to hyperkalemia and hypovolemia. Fludrocortisone which may be useful in such patients expands blood volume and enhances the vascular sensitivity to circulating catecholamines. Fludrocortisone also augments the release of norepinephrine from sympathetic neurons. The autonomic nervous system also may play a role in mediating the increased cardiovascular risk associated with aldosterone. Animal studies indicate that aldosterone can centrally increase the sympathetic activity in the brain. In dogs, aldosterone also inhibits carotid baroreceptor discharge thereby increasing sympathetic activity. Recent studies from patients with hyperaldosteronism show elevated sympathetic activity comparable to patients with essential hypertension that normalizes after unilateral adrenalectomy, thus supporting a sympathoexcitatory role for aldosterone [10]. In summary, adrenocortical deficiency has a profound effect on the efficacy of sympathetic nervous system in maintaining blood pressure especially during stress. Some of the clinical features of adrenal insufficiency like dizziness; hypotension can closely mimic autonomic dysfunction. Early recognition of adrenal insufficiency and prompt therapy with intravenous steroids and fluids can be lifesaving. Finally, an inappropriate increase in sympathetic activity may contribute to the accelerated cardiovascular complications among patients with relative or absolute aldosterone excess.

References [1] Schinner S, Bornstein SR. Cortical-chromaffin cell interactions in the adrenal gland. Endocr Pathol 2005;16(2):91–8. [2] Ehrhart-Bornstein M, Bornstein SR. Cross-talk between adrenal medulla and adrenal cortex in stress. Ann N Y Acad Sci 2008;1148(Dec):112–7. [3] Ehrhart-Bornstein M, Hinson JP, Bornstein SR, Scherbaum WA, Vinson GP. Intraadrenal interactions in the regulation of adrenocortical steroidogenesis. Endocr Rev 1998;19(2):101–43. [4] Wurtman RJ, Pohorecky LA, Baliga BS. Adrenocortical control of the biosynthesis of epinephrine and proteins in the adrenal medulla. Pharmacol Rev 1972;24(2):411–26.

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[5] Bornstein SR, Breidert M, Ehrhart-Bornstein M, Kloos B, Scherbaum WA. Plasma catecholamines in patients with Addison's disease. Clin Endocrinol (Oxf) 1995;42(2):215–8. [6] Yang S, Zhang L. Glucocorticoids and vascular reactivity 2004;2(1): 1–12. Curr Vasc Pharmacol 2004;2(1):1–12. [7] Williams GH, Dluhy RG. Chapter 336. Disorders of the adrenal cortex. In: Fauci AS, Braunwald E, Kasper DL, Hauser SL, Longo DL, Jameson JL, Loscalzo J, editors. Harrison’s Principles of Internal Medicine. 17th edition.

[8] al-Damluji S. Adrenergic control of the secretion of anterior pituitary hormones. Baillieres Clin Endocrinol Metab 1993;7(2):355–92. [9] Polinsky RJ, Kopin IJ, Ebert MH, Weise V, Recant L. Hormonal responses to hypoglycemia in orthostatic hypotension patients with adrenergic insufficiency. Life Sci 1981;29(4):417–25. [10] Kontak AC, Wang Z, Arbique D, Adams-Huet B, Auchus RJ, Nesbitt SD, et al. Reversible sympathetic overactivity in hypertensive patients with primary aldosteronism. J Clin Endocrinol Metab 2010;95(10):4756–61.

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119 Mastocytosis L. Jackson Roberts, II Although mastocytosis is not a disorder of the autonomic nervous system, some of the symptoms and signs of the disease may be interpreted as consistent with autonomic dysfunction. Thus, it is important for physicians involved in the evaluation of patients suspected of having disorders of the autonomic nervous system to recognize the hallmarks of the disease.

MASTOCYTOSIS AND ALLIED ACTIVATION DISORDERS OF THE MAST CELL Mastocytosis is a disease characterized by an abnormal proliferation of tissue mast cells. The cause of the overproliferation of mast cells remains unknown. Although unusual forms of the disease can occur primarily in children, e.g., a localized mastocytoma, the disease in adults exists primarily in two forms; one in which the abnormal proliferation of mast cells appears to be either limited to the skin (cutaneous mastocytosis) or involves multiple tissues throughout the body (systemic mastocytosis) [1,2]. In recent years, an allied activation idiopathic disorder(s) of mast cells has also been identified in which patients experience episodes of systemic mast cell activation in the absence of any evidence of abnormal mast cell proliferation [3]. An allergic basis for the activation of mast cells may be suspected in some patients whereas in others, the cause remains unclear. Whereas mastocytosis is an uncommon disease, idiopathic activation disorders of the mast cell are encountered more frequently.

SYMPTOMS AND SIGNS The symptoms of both mastocytosis and systemic activation disorders of the mast cell are attributed primarily to episodic release of mast cell mediators [3]. The episodes of mastocyte activation can be brief, lasting several minutes, or protracted, lasting a few hours. These episodes frequently occur without any identifiable inciting cause. However, exposure to heat, exertion, and emotional upset are commonly identified as precipitating factors by many patients. The major symptoms experienced by these

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00119-0

patients are listed in Box 119.1. Probably the most important clinical clue which should lead one to suspect a diagnosis of systemic mast cell disease is flushing. In some patients, cutaneous vasodilation is not appreciated but patients will usually note that they feel very warm. Unlike allergic anaphylaxis, bronchospasm, angioedema, and urticaria are uncommon manifestations. Characteristically, following an episode of mastocyte activation, patients experience extreme fatigue and lethargy, which can last for hours. Hemodynamic alterations frequently occur during episodes of systemic mast cell activation [3]. Characteristically, the blood pressure falls and the heart rate increases. At times the reduction in blood pressure can be profound, resulting in severe lightheadedness or frank syncope. The reduction in blood pressure in accentuated in the upright position and patients note that lightheadedness is improved upon assuming the supine position. However, in some patients the blood pressure increases, at times dramatically, during episodes of mast cell activation. The elevation in blood pressure is also accompanied by an increase in heart rate. The basis for the rise in blood pressure in some patients remains speculative.

MAST CELL MEDIATORS RESPONSIBLE FOR THE SYMPTOMS AND SIGNS The hemodynamic alterations and symptoms experienced by patients with mastocytosis had previously been attributed to the release of excessive quantities of histamine from mast cells. However, treatment with antagonists of histamine H1 and H2 receptors had not been found to prevent episodes of vasodilation in these patients. In 1980, the discovery of marked overproduction of prostaglandin D2, a potent vasodilator, in patients with mastocytosis was reported. Subsequently, it was found that treatment of patients with mastocytosis with inhibitors of prostaglandin biosynthesis in addition to antihistamines can be effective in ameliorating episodes of vasodilation in these patients [3]. However, a subset of patients with disorders of systemic mast cell activation are “aspirin hypersensitive” and administration of any prostaglandin inhibitor, even in small doses, can provoke

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119. MAsToCyTosIs

BOX 119.1

S Y M P T O M S O F S Y S T E M I C M A S T C E L L A C T I VA T I O N 1. 2. 3. 4. 5.

Flushing (and/or a feeling of warmth) Palpitations Dyspnea (usually without wheezing) Chest discomfort Headache

6. Lightheadedness and occasionally syncope 7. Gastrointestinal symptoms: (a) Nausea and occasionally vomiting (b) Abdominal cramps and occasionally diarrhea 8. Profound lethargy after the attack.

a severe episode of mastocyte activation. Thus, great caution must be exercised in treating patients with disorders of systemic mast cell activation with inhibitors of prostaglandin biosynthesis. The pathogenesis of “aspirin hypersensitivity” remains poorly understood.

DIAGNOSIS The diagnosis of systemic disorders of the mast cell is not always straightforward [4]. Patients with mastocytosis frequently have small pigmented cutaneous lesions, termed uriticaria pigmentosa. Urticaria pigmentosa lesions characteristically urticate when stroked (Darier’s sign). When visible cutaneous clues to the diagnosis are absent, the diagnosis relies on the recognition of a compatible clinical history. In patients with mastocytosis, a diagnosis may be made histologically by demonstrating abnormal mast cell proliferation in the skin or bone marrow. In patients with activation disorders of the mast cell in the absence of abnormal mast cell proliferation, the diagnosis relies entirely on demonstrating a release of increased quantities of histamine and prostaglandin D2 during episodes of suspected mastocyte activation. In patients with systemic mastocytosis, increased urinary excretion of metabolites of histamine and prostaglandin D2 can usually be demonstrated even at quiescent times. On the other hand, in patients with idiopathic activation disorders of the mast cell, increased excretion of metabolites of histamine and prostaglandin D2 can only be demonstrated in fractional urines collected during episodes of mastocyte activation.

SUMMARY Patients with disorders of systemic mast cell activation may be encountered more frequently than previously thought and some of the symptoms and signs manifested by these patients, e.g., orthostatic hypotension, during episodes of mastocyte activation, are not unlike some of those experienced by some patients with autonomic dysfunction. Specifically, a patient with “spells” characterized by flushing that may be precipitated by heat, exertion, or emotional upset accompanied by lightheadedness and either a reduction or increase in blood pressure and tachycardia should lead the astute clinician to consider the possibility of a disorder of systemic mast cell activation.

References [1] Arock M, Valent P. Pathogenesis, classification, and treatment of mastocytosis: state of the art in 2010 and future perspectives. Expert Rev Hematol 2010;3:497–516. [2] Soter NN. The skin in mastocytosis. J Invest Dermatol 1991;96:32S–9S. [3] Roberts II LJ, Oates JA. Disorders of vasodilator hormones: the carcinoid syndrome and mastocytosis. In: Wilson JD, Foster DW, editors. Williams textbook of endocrinology (8th ed.). Philadelphia: W.B. Saunders; 1992. p. 1619–33. [4] Roberts LJ, Oates JA. The biochemical diagnosis of systemic mast cell disorders. J Invest Dermatol 1991;96:19S–25S.

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120 Cocaine Overdose Andrew Kontak, Wanpen Vongpatanasin, Ronald G. Victor Cocaine addiction is increasing worldwide among all income strata [1]. Cocaine is the most common illicit drug causing life-threatening cardiovascular emergencies, including acute coronary syndrome, stroke, sudden cardiac death, hypertensive crisis, and aortic dissection. These cardiovascular complications of cocaine have been attributed primarily to inhibition of norepinephrine (NE) reuptake in peripheral sympathetic nerve terminals leading to increased [NE] in the synaptic cleft [2,3]. However, this hypothesis was based largely on ex vivo rodent experiments with little or no supporting evidence from studies in intact animals or humans. In contrast, a large series of clinical physiology studies by our group has demonstrated that cocaine stimulates the human cardiovascular system mainly by acting in the brain to increase central sympathetic nerve activity (SNA) [4–6]. Thus, SNA constitutes a putative new drug target for the acute management of cocaine-induced cardiovascular emergencies.

EFFECTS OF COCAINE ON THE PERIPHERAL CIRCULATION Inhibition of the NE transporter in the peripheral circulation cannot be the only mechanism by which cocaine acutely raises blood pressure. Tricyclic antidepressants, which are more potent NE transporter inhibitors than cocaine, do not raise blood pressure [7]. In healthy humans, intranasal cocaine raises blood pressure and activates baroreceptor reflexes thereby reflexively decreasing muscle SNA, the proximate neural stimulus for NE release [4]. If there is little stimulus for NE release into the synaptic cleft, there should be little NE available for reuptake by the transporter and thus little transporter activity to be blocked by cocaine. Indeed, small doses of intranasal cocaine (onehalf the local anesthetic dose used in rhinolaryngologic procedures) raised blood pressure but had no effect on plasma NE levels in cocaine-naïve healthy subjects; however, in the same subjects, venous forearm [NE] increased when cocaine was infused directly into the brachial artery, achieving the same venous concentration without systemic spillover [5]. Thus, with intranasal cocaine, plasma venous [NE] does not increase as baroreflex-mediated suppression of muscle SNA offsets the effect of NE transporter inhibition. When muscle SNA was clamped during systemic

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00120-7

cocaine administration by the concomitant infusion of intravenous nitroprusside to minimize the increase in BP and baroreflex activation the increase in venous [NE] was restored, matching the increase seen with intrabrachial cocaine (Fig. 120.1). Thus, our microneurographic studies in healthy cocaine-naïve subjects indicate that baroreceptor reflexes play a key role in buffering the sympathomimetic actions of cocaine on the human skeletal muscle circulation (Fig. 120.2). The degree of neurogenic vasoconstriction induced by a given dose of cocaine is critically dependent on the ambient level of central muscle sympathetic outflow, which is suppressed in individuals with intact baroreceptor function. In this regard, patients with impaired baroreceptor reflexes such as those with longstanding hypertension or heart failure would suffer augmented risk of hypertensive crisis and other catastrophic cardiovascular complications from cocaine because of unrestrained sympathetic outflow coupled with inhibition of peripheral NE reuptake.

AUTONOMIC EFFECTS OF COCAINE ON THE HEART Cocaine’s acute adverse effects on the heart – acute coronary syndrome due to increased myocardial oxygen demands from increased heart rate and blood pressure and decreased oxygen delivery due to coronary vasoconstriction – also have been attributed to inhibition of cardiac NE reuptake [1,2]. However, studies by Hillis, Lange and colleagues led to calling this hypothesis into question, as cocaine produces its cardiac effects only with systemic administration (allowing cocaine uptake into the brain) but not with intracoronary administration [8,9]. Intranasal cocaine increased heart rate and caused alpha-adrenergic coronary vasoconstriction in patients undergoing coronary angiography (for evaluation of chest pain), whereas infusion of cocaine directly into the coronary arteries, even at doses that produced a high cocaine concentration in the coronary sinus, did not produce these effects [9]. Similarly, intranasal cocaine increased left ventricular contractility, whereas intracoronary cocaine did not [8]. These data strongly suggest a central rather than peripheral site of action of cocaine – to increase sympathetic outflow to the heart.

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120. COCAInE OvERdOsE

MAP

SNA

% total activity 120

mmHg 100

80

80

80

FIGURE 120.1 The role of baroreflexes

[NE] pg/ml 120

40

40 40

60 0

Baseline Cocaine Cocaine +NTP

Baseline Cocaine Cocaine +NTP

0

Baseline Cocaine Cocaine +NTP

in buffering sympathomimetic effects of cocaine. Summary data of five healthy subjects showing mean arterial pressure (MAP), sympathetic nerve activity (SNA), and venous NE concentration (top panel) and original recordings of muscle SNA in one subject (bottom panel) at baseline, after intranasal cocaine alone, and after intranasal cocaine plus intravenous nitroprusside. With intranasal cocaine alone, MAP increased, muscle SNA decreased reflexively while forearm venous NE concentration was unchanged. When SNA was carefully returned to the baseline level by attenuating the cocaine-induced rise in MAP with nitroprusside, a significant increase in venous NE concentration was observed. *p  0.01 vs. baseline.

10 sec

COCAINE

ARTERIAL BARORECEPTORS

FIGURE 120.2 Diagram showing how cocaine

CNS

+

–

SKELETAL MUSCLE SNA

+

NERVE TERMINALS

raises BP in humans. Cocaine increases sympathetic nerve discharge to the heart causing tachycardia and elevated BP. The increase in BP activates arterial baroreceptors, which in turn trigger sympathetic withdrawal to the skeletal muscle vasculature. Although cocaine can inhibit the peripheral NE transporter, this mechanism is not effective in increasing NE levels at the synaptic cleft because the muscle SNA, the neural stimulus for NE release, is suppressed.

COCAINE

–

NE HR

BP

CARDIAC OUTPUT

Peripheral Resistance

Although microneurographic measurements of cardiac SNA are not possible to perform in human subjects, our microneurographic studies show that intranasal cocaine is a potent stimulus to skin SNA, a regional sympathetic outflow that is very sensitive to central neural stimuli but quite insensitive to baroreflex stimuli [6]. Low-dose intranasal cocaine causes a large and sustained increase in skin SNA lasting for 90 minutes after administration. The increase in SNA is accompanied by a parallel increase in heart rate that is abolished by β-adrenergic receptor blockade but

unaffected by muscarinic receptor blockade, indicating sympathetic rather than parasympathetic mediation. This sympathetically-mediated increase in the heart rate is a potent effect of cocaine because it can overcome the opposing influence of the arterial baroreflex to activate parasympathetic outflow to the sinus node. In humans, this central sympathoexcitatory effect of cocaine on heart rate (and thus cardiac output) is a major mechanism by which cocaine raises blood pressure, with total systemic vascular resistance and stroke volume remaining unchanged [8].

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EffECTs Of COCAInE On THERmOREgulATIOn

Cutaneous Vascular Conductance (% Maximum)

Sweat Rate (mg/cm /min) 2

1.2

80

60

Lidocaine

Lidocaine

0.8

40

0.4 20

Cocaine Cocaine

0

0.0 36.8

A

37.2

36.8

37.6

Esophageal Temperature (oC)

B

37.2

37.6

Esophageal Temperature (oC)

FIGURE 120.3 Effects of cocaine on autonomic adjustments to heat stress. Changes in cutaneous vascular conductance (A) and sweat rate (B) relative to esophageal temperature during both stresses (cocaine: closed circles; lidocaine or placebo: open squares). Data are mean SEM. Cocaine significantly increased the esophageal temperature threshold for the onset of cutaneous vasodilation (p  0.01) and sweating (p  0.001) without affecting the slope of the elevation in cutaneous vascular conductance or sweat rate relative to the elevation in esophageal temperature. At any given esophageal temperature, cutaneous vascular conductance and sweat rate were significantly reduced for the cocaine trial (*p  0.01 cocaine vs. lidocaine trials).

EFFECTS OF COCAINE ON THERMOREGULATION Hyperthermia is a specific complication of cocaine overdose, which further amplifies the drug’s cardiovascular toxicity. Fatal cocaine overdose typically is associated with high blood cocaine levels (3–6 mg/L) but cocainerelated deaths also can occur at 10–20 times lower blood levels when hyperthermia is present [10]. The hyperthermic properties of cocaine have been attributed largely to a hypermetabolic state (agitation with increased locomotor activity) that increases heat production. However, we found that impaired heat dissipation is another major mechanism by which cocaine elevates body temperature [10]. When healthy cocaine-naïve individuals were subjected to passive heating, pretreatment with even a small dose of intranasal cocaine impaired both sweating and cutaneous vasodilation, the major autonomic adjustments to thermal stress (Fig. 120.3) [10]. The precise mechanism by which cocaine impairs both cutaneous vasodilation and sweating is still unknown but likely has a central mechanism of action because heat perception was also impaired by cocaine. This was a dramatic effect because subjects experienced less thermal discomfort with cocaine even though core temperature was higher than with placebo (Fig. 120.4). In addition, heat perception is the key trigger for the behavioral adjustments such as seeking a cooler environment or adjusting the thermostat on the air conditioner which, in humans, constitute the most important thermoregulatory responses [10].

Rating of Perceived Heating Unbearably 8.0 Hot

Lidocaine

Very Hot 7.0

Hot 6.0

Cocaine Warm 5.0

Comfortable 4.0 36.8

37.2

37.6

38.0 o

Esophageal Temperature ( C)

FIGURE 120.4 Effects of cocaine on thermal perception. Esophogeal temperature at each rating of thermal sensation for both heat stresses. Values are displayed as mean SEM. At any given esophageal temperature above 37°C, the rating scores of perceived heating were significantly lower during cocaine trials (closed circles) than lidocaine or placebo trials (open squares) and these differences became even larger as esophageal temperature rose progressively, suggesting that cocaine impaired thermal perception (*p  0.05 vs. lidocaine).

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120. COCAInE OvERdOsE

Skin SNA

skin vascular resistance in response to cocaine and Dex intranasal cocaine increased skin sympathetic nerve activity (SNA), which was accompanied by a parallel increase in skin vascular resistance. Dexmedetomidine (Dex) reversed effects of cocaine on SNA and skin vascular resistance, beginning at the dose of 0.1 μg/kg/ min. Data are mean SE. *p  0.05 vs. baseline, †p  0.01 vs. baseline, ‡p  0.05 vs. saline, §p  0.01 vs. saline.

25

Resistance?units

Ln?%total nerve activity

6

5

FIGURE 120.5 Changes in skin SNA and

Skin vascular resistance

*

4

20

* 15 S S Dexmedetomidine (n=11) saline (n–8)

S S 3

Baseline Cocaine + Dex 0.1 + Dex 0.3 or saline or saline

10

Baseline Cocaine + Dex 0.1 + Dex 0.3 or saline or saline

TREATMENT OF COCAINE OVERDOSE Current treatment recommendations for cocaineinduced chest pain and acute hypertension are based on limited data [11]. Benzodiazepines (for agitation), nitroglycerin, and aspirin are recommended as first-line treatment of cocaine-induced chest pain/acute coronary syndrome, with primary angioplasty being indicated for the small fraction of patients who develop ST-elevation myocardial infarction. If blood pressure remains elevated, second-line treatment options are controversial and include calcium channel blockers, phentolamine, and labetalol (a combined alpha-/beta-blocker). Beta-blockers, which are anti-hypertensive and cardio-protective in most acute coronary syndromes, are thought to exacerbate cocaine-induced alpha-adrenergic coronary vasoconstriction and thus have been contraindicated; however, the guidelines are being re-evaluated in light of recent observational data showing improved clinical outcomes in patients who received beta-blockers in the setting of cocaine overdose [12]. Better treatment options are needed. If, as we postulate, cocaine acts in the brain to increase SNA to the human heart and peripheral circulation, then a central sympatholytic drug should eliminate the cardiovascular response to cocaine at its central origin. We showed that dexmedetomide, a central sympatholytic which is a much more potent alpha-2 adrenergic agonist than clonidine, effectively abolished the cocaine-induced increase in skin SNA and the corresponding increases in regional vascular resistance, blood pressure, and heart rate (Fig. 120.5) [13]. The alpha-2 agonist was effective in blocking these sympathomimetic actions of cocaine even in subjects who were homozygous for the Del322–325 polymorphism in the alpha-2C adrenoreceptor, a loss-of-function mutation that is highly enriched in black individuals.

Further research is warranted to move this work from the clinical physiology laboratory to the clinical setting – emergency management of cocaine overdose. The studies reviewed in this chapter involved cocaine-naïve subjects receiving a low-dose cocaine challenge, i.e., a dose that causes a measureable cardiovascular response without exposing cocaine-naïve subjects to cocaine intoxication. Thus, additional studies will need to determine if central sympatholytic agents such as dexmedetomide constitute an effective new treatment for acute coronary syndrome and acute hypertension in non-treatment seeking cocaineaddicted individuals.

Acknowledgements This work was supported by NIH RO1 DA010064 (RGV, WV) and by Ruth L. Kirschstein National Research Service Award F32DA027274 (AK).

References [1] Phillips K, Luk A, Soor GS, Abraham JR, Leong S, Butany J. Cocaine cardiotoxicity: a review of the pathophysiology, pathology, and treatment options. Am J Cardiovasc Drugs 2009;9(3):177–96. [2] Lange RA, Hillis LD. Cardiovascular complications of cocaine use. N Engl J Med 2001;345(5):351–8. [3] Maraj S, Figueredo VM, Lynn MD. Cocaine and the heart. Clin Cardiol 2010;33(5):264–9. [4] Jacobsen TN, Grayburn PA, Snyder RW, et al. Effects of intranasal cocaine on sympathetic nerve discharge in humans. J Clin Invest 1997;99(4):628–34. [5] Tuncel M, Wang Z, Arbique D, Fadel PJ, Victor RG, Vongpatanasin W. Mechanism of the blood pressure–raising effect of cocaine in humans. Circulation 2002;105(9):1054–9. [6] Vongpatanasin W, Mansour Y, Chavoshan B, Arbique D, Victor RG. Cocaine stimulates the human cardiovascular system via a central mechanism of action. Circulation 1999;100(5):497–502.

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REfEREnCEs

[7] Schroeder JS, Mullin AV, Elliott GR, et al. Cardiovascular effects of desipramine in children. J Am Acad Child Adolesc Psychiatry 1989;28(3):376–9. [8] Boehrer JD, Moliterno DJ, Willard JE, et al. Hemodynamic effects of intranasal cocaine in humans. J Am Coll Cardiol 1992;20(1):90–3. [9] Daniel WC, Lange RA, Landau C, Willard JE, Hillis LD. Effects of the intracoronary infusion of cocaine on coronary arterial dimensions and blood flow in humans. Am J Cardiol 1996;78(3):288–91. [10] Crandall CG, Vongpatanasin W, Victor RG. Mechanism of cocaine-induced hyperthermia in humans. Ann Intern Med 2002;136(11):785–91.

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[11] McCord J. Cocaine-associated chest pain and acute myocardial infarction. Rev Esp Cardiol 2010;63(9):1013–4. [12] Rangel C, Shu RG, Lazar LD, Vittinghoff E, Hsue PY, Marcus GM. Beta-blockers for chest pain associated with recent cocaine use. Arch Intern Med 2010;170(10):874–9. [13] Menon DV, Wang Z, Fadel PJ, et al. Central sympatholysis as a novel countermeasure for cocaine-induced sympathetic activation and vasoconstriction in humans. J Am Coll Cardiol 2007;50(7):626–33.

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121 Complex Regional Pain Syndrome Wilfrid Jänig

The term complex regional pain syndrome (CRPS) was introduced in 1995. In CRPS I (previously reflex sympathetic dystrophy) minor injuries at the limb or lesions in remote body areas precede the onset of symptoms. CRPS II (previously causalgia) develops after injury of a major peripheral nerve. The new terminology and diagnosis of CRPS are based on elements of history, symptoms and findings of clinical examination with no implied pathophysiological mechanism [9]. There are no gold standards to compare with and no absolute diagnostic tests that are specific for CRPS. It is difficult to distinguish CRPS from other extremity pain syndromes and to predict acutely after a trauma at an extremity who is going to develop CRPS. The clinical criteria for diagnosis of CRPS and the additional objective tests supporting the diagnosis have been described [1].

CRPS I IS A NEURONAL DISORDER INVOLVING THE CNS The sensory, sympathetic, somatomotor, and trophic changes (including swelling), observed in variable combinations in patients with CRPS are the results of changes and distorted processing of information in the CNS. Various levels of integration probably are involved such as spinal cord, brain stem, diencephalon (hypothalamus, thalamus), and telencephalon (cortex and limbic system). The upper part in Box 121.1 lists clinical and experimental observations on patients with CRPS that clearly support this contention. The lower part in Box 121.1 lists the peripheral changes in patients with CRPS that are related in some yet unknown way to the central changes. The scheme of Figure 121.1 outlines a general heuristic explanatory hypothesis [3,6]. This hypothesis postulates changes in the central representations of the somatosensory, autonomic and somatomotor systems to explain the clinical findings. The clinical symptoms mostly follow a trauma in the somatic domains at the extremities, but sometimes also trauma in the viscera or in the CNS. The changes developing after these triggering events usually outlast the trauma by orders of magnitude.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00121-9

SYMPATHETIC SYSTEMS SUPPLYING SKIN Cutaneous Vasoconstrictor Neurons and Blood Flow Through Skin Thermoregulatory reflexes to whole body heating and cooling are changed in the distal parts of the affected extremity of CRPS patients. Three reaction patterns of cutaneous vasoconstrictor neurons supplying the hand of the affected extremity during thermoregulatory load (whole body cooling and warming) occur: whole body cooling generates either a weaker than normal activation or no activation or a stronger than normal activation or an intermediate response pattern leading to the expected difference in temperature and blood flow through acral skin between the affected and the contralateral (control) side. These differences in blood flow and temperature are small at extreme thermoregulatory states (maximal cooling and maximal warming) and large at neutral thermoregulatory states [10]. In the early stages of CRPS, vasoconstriction and vasodilatation in fingers elicited by deep inspiration/ expiration may be attenuated. The respiration-induced vasoconstriction of the cutaneous vascular blood vessels of the hand is generated by activation of vasoconstrictor neurons related to the coupling between the neuronal networks in the lower brain stem regulating respiration and cutaneous blood flow. The changes occurring in cutaneous blood flow and temperature during these interventions can only be attributed to central changes, which are reflected in changes of activity in cutaneous vasoconstrictor neurons innervating the distal parts of the affected extremity [4]. The central changes reflected in the changed reflexes in cutaneous vasoconstrictor neurons may at least in part occur at the spinal cord level. Integration between supraspinally generated signals (e.g., in the hypothalamus and in the respiratory network) and signals in spinal circuits may have changed. This idea is consistent with the observation that the thermoregulatory changes in the CRPS patients is only present in the affected extremity but not in the contralateral one.

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BOX 121.1

ARGUMENTS FOR CENTRAL AND FOR PERIPHERAL CHANGES IN CRPS

Central changes

l

Changes of Regulation by Sympathetic Systems (1 in Fig. 121.1) l

l

l l

Thermoregulatory reflexes in cutaneous vasoconstrictor neurons reduced Respiration elicited reflexes (generated by deep in- and expiration) in cutaneous vasoconstrictor neurons reduced Changes of activity in sudomotor neurons (sweating) Swelling reduced by sympathetic blocks.

l

Peripheral changes Sympathetic-afferent Coupling (6 in Fig. 121.1) l

l

Sensory Changes (2 in Fig. 121.1) l l

l

Mechanical allodynia (quadrant, hemisensory) Hypoesthesias (mechanical, cold, warm; hemisensory, quadrant) Bilateral distribution of hypo- and hyperesthesias (mechanical, cold, warm, heat).

Somatomotor Changes (3 in Fig. 121.1) l l l

l l

Active motor force and active range of motion reduced Physiological tremor increased Poor motor control and coordination of movement; altered gait and posture Dystonia Sensory-motor body perception disturbance.

Initiating Events (4 in Fig. 121.1) l l

l

Pain Relief by Sympathetic Blocks with Local Anesthetics (5 in Fig. 121.1) l

l

l

Relief of pain outlasts conduction block by an order of magnitude, (i.e., a temporary block is followed by a long-lasting pain relief)

Sudomotor Neurons and Sweating Sweating is changed in the affected extremity (hypo- or hyperhidrosis) of CRPS patients. These changes can only be attributed to central changes reflected in changes of activity in sudomotor neurons. Reflex inhibition in cutaneous vasoconstrictor neurons induced by peripheral or central (warm) stimuli is always accompanied by reflex activation of sudomotor neurons implying a reciprocal reflex organization of both systems in the spinal cord, brainstem and hypothalamus [4].

After nerve lesion via noradrenaline and adrenoceptors (CRPS II) Indirectly via vascular bed and other mechanisms (CRPS I; deep somatic?) Indirectly via inflammatory mediators and neurotrophic factors Mediated by the adrenal medulla (adrenaline).

Inflammatory Changes and Edema (7 in Fig. 121.1) l

l

l

l

Out of proportion to pain disease (minor trauma) Events remote from affected extremity (e.g., in the visceral domain) Central (e.g., after stroke; related to endogenous control systems?).

A few temporary blocks are sometimes sufficient to generate permanent pain relief Sympathetic activity maintains a positive feedback circle (?)

Neurogenic inflammation (precapillary vasodilatation, venular plasma extravasation), involvement of peptidergic afferents (?) Sympathetic fibers mediating effects of inflammatory mediators (e.g., bradykinin) to venules leading to plasma extravasation (?) Involvement of inflammatory cells and immune system (?) Change of capillary filtration pressure (?)

Trophic Changes (8 in Fig. 121.1) l

l

l

Long-range consequences of inflammatory changes and edema (?) Direct (trophic?) effect of sympathetic and afferent fibers on tissue (?) Endothelial damage (?)

Modified from [6].

SYMPATHETIC NEURONS AND EDEMA, INFLAMMATION AND TROPHIC CHANGES Edema The edema in CRPS patients may be attenuated after sympathetic blocks showing its dependence on activity in sympathetic neurons. The decrease starts within one to two hours and the edema may disappear within days. The edema may also be generated by antidromic activity in peptidergic afferent neurons with unmyelinated (C) or

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585

SomATomoToR CHAngES

Chronic excitation of deep somatic or visceral afferents

Central events, central lesion 4

Trauma with/without peripheral nerve lesion

4

4 Abnormal state of afferent neurones

Pain, other sensory changes

2

CHANGED INFORMATION PROCESSING IN THE CNS

3

Movement disorders

Abnormal activity in motoneurones to skeletal muscle

Sympathetic block 5

6

7 Abnormal regulation of blood flow and sweating

1 Abnormal activity in sympathetic neurones (vaso-, sudomotor, other?)

7 8

7

8

Swelling inflammation Trophic changes

FIGURE 121.1 General explanatory hypothesis about the neural mechanisms of generation of CRPS following peripheral trauma with and without nerve injury, chronic stimulation of deep somatic afferents or rarely visceral afferents (e.g., during angina pectoris, myocardial infarction) or, rarely, central trauma. The clinical observations are put in bold lined boxes. An important component is the excitatory influence of postganglionic sympathetic axons on primary afferent neurons. The numbers indicate the changes occurring potentially in CRPS patients that have been quantitatively measured or postulated on the basis of clinical observations (see Box 121.1): 1, changes of activity in sympathetic neurons; 2, pain, other somatosensory changes; 3, changes of activity in somatomotoneurons; 4, initiating events; 5, consequences of sympathetic blocks or sympathectomy (dotted line); 6, sympathetic-afferent coupling (establishing the positive vicious feedback circle [in bold]); 7, “antidromically” conducted activity in peptidergic afferent C-fibers (dotted arrow) leading to increase of blood flow (arteriolar vasodilation) and venular plasma extravasation (in deep tissues), both hypothetically contributing to increase in blood flow, swelling/inflammation and trophic changes; 8, sympathetic postganglionic fibers hypothetically contributing to swelling/inflammation and trophic changes. Modified [5].

small-diameter myelinated (Aδ) fibers (see interrupted arrow in Fig. 121.1) producing arteriolar vasodilatation and venular plasma extravasation (in deep tissues).

Inflammation Already Sudeck believed that CRPS is an inflammatory bone atrophy (“entzündliche Knochenatrophie”). This is supported by animal studies showing that the sympathetic nervous system can influence the intensity of inflammatory processes and clinical studies showing that sympatholytic procedures can ameliorate inflammation and edema in humans. The underlying mechanisms are unclear [6].

Trophic Changes The underlying mechanisms of trophic changes are unclear. These changes may ameliorate after sympathetic blocks, indicating that they are related to the sympathetic innervation.

extremity showing increased thresholds to mechanical, cold, warm or heat stimuli compared with the contralateral healthy body side. Patients with these extended sensory deficits have longer illness duration, greater pain intensity, a higher frequency of mechanical allodynia, and a higher tendency to develop changes in the somatomotor system than patients with spatially restricted sensory deficits. The anatomical distributions of the changed painful and non-painful somatosensory perceptions are likely due to changes in the central representation of somatosensory sensations in the cortex as supported by magnetic encephalographic (MEG) and functional magnetic resonance imaging (MRI) studies. It is unclear to what degree these changes are specific for CRPS. CRPS I patients mostly locate their spontaneous pain into deep somatic structures of the affected extremity and have deep somatic mechanical hyperalgesia/allodynia [1,8].

SOMATOMOTOR CHANGES SENSORY SYSTEMS OF THE SKIN About 50% of patients with chronic CRPS I develop hypoesthesia and hypoalgesia on the whole half of the body or in the upper quadrant ipsilateral to the affected

About 50% of patients with CRPS I show a decreased active range of motion, increased amplitude of physiological tremor, and reduced active motor force in the affected extremity, the extremes being myoclonus and dystonia. These motor changes are generated by changes of activity

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121. ComPlEx REgIonAl PAIn SyndRomE

in the motoneurons, i.e., they have a central origin and are possibly related to plastic changes in the somatosensory, motor and premotor cortices. Functional MRI studies reveal pathological sensorimotor integration in the parietal cortex resulting in abnormal central programming and processing of motor tasks. A neglect-like syndrome may be involved in the disuse of the extremity and incongruence between central motor output and sensory input (body perception disturbance) is hypothesized as underlying mechanism in CRPS [3].

INITIATING EVENTS The clinical signs and symptoms in CRPS are disproportionate to the traumatic events initiating or triggering this syndrome. The local changes generated by the trauma often disappear, yet the syndrome persists. Furthermore, CRPS may be triggered by remote events (e.g., in the viscera) or by events in the CNS. These clinical observations argue that mechanisms operating in CRPS cannot be reduced to events in the periphery of the body related to the trauma.

maintains a positive feedback circle via the primary afferent neurons and a central state of hyperexcitability (e.g., of neurons in the spinal dorsal horn) (Fig. 121.1).

CONCLUSIONS Clinical observations, experimentation on humans and animals suggest that CRPS is a disorder of the CNS: 1. CRPS patients exhibit alterations of the somatosensory, sympathetic nervous and somatomotor systems indicating that the central representations of these systems are changed (see dots in Fig. 121.2 and bold-lined boxes in Fig. 121.1). Thus, CRPS is a disorder

Dynamic changes in central representations of Somato-sensory Somato-motor & Symp. systems

CRPS AND SYMPATHETICALLY MAINTAINED PAIN (SMP) Pain dependent on activity in the sympathetic neurons called sympathetically maintained pain (SMP) is present in about 60% of patients with acute CRPS but can persist for years. Sympathetic neurons, sympathetic-afferent coupling, afferent neurons and central neurons (spinal neural circuits and their supraspinal control) are postulated to form a positive feedback circle (Fig. 121.1). This concept is based on long standing clinical observations and experimental studies of CRPS patients [2,5]. 1. In CRPS patients with SMP it has been shown that spontaneous pain, mechanical allodynia and cold allodynia in the hand can be rekindled, under the condition of proximal sympathetic block, or enhanced by injection of norepinephrine into the skin area that is or was painful before sympathetic blockade. Furthermore, SMP is significantly reduced by the α-adrenoceptor blocker phentolamine infused intravenously [5]. 2. The intensity of spontaneous pain and the area of mechanical hyperalgesia/allodynia can be increased or decreased by increasing or decreasing the activity of the sympathetic outflow in CRPS patients with SMP, but not in CRPS patients without SMP [2]. 3. In CRPS patients with SMP blockade of sympathetic activity to the affected extremity by a local anesthetic generates pain relief in the affected extremity for significantly longer time periods than control saline injection, the duration of pain relief outlasting the duration of conduction block by orders of magnitude [7]. This suggests that activity in sympathetic neurons

SOMAT. SENS. Mech. Percept. Warm percept. Cold percept.

PAIN Spontaneous Hyperalgesia Allodynia

Forebrain

Brain stem

Initiation Maintenance

Dynamic changes of, spinal circuits

Spinal cord

Afferent

Sympathetic Somatomotor PERIPH. TISSUES Swelling & edema Inflammation Trophic changes

SYMPATHETIC NS Regulation of blood Flow & sweating Symp.-aff. Coupling

MOTOR SYSTEM Motor force Tremor Dystonia

FIGURE 121.2 CRPS is a disorder of the central nervous system (CNS): a hypothesis. Schematic diagram summarizing the sensory, autonomic and somatomotor changes in CRPS patients. The figure symbolizes the CNS (forebrain, brain stem and spinal cord). Changes occur in the central representations of the somatosensory, somatomotor and sympathetic nervous system (which include spinal circuits) and are reflected in the changes of perception of painful and non-painful stimuli, of cutaneous blood flow and sweating, and of motor performances. They are triggered and possibly maintained by nociceptive afferent inputs from the somatic and visceral body domains. It is unclear whether these central changes are reversible in chronic CRPS patients. The central changes affect the endogenous control system of nociceptive impulse transmission possibly too. Coupling between sympathetic neurons and afferent neurons in the periphery (see bold closed arrow) is one component of pain in CRPS patients with SMP. However, it seems to be unimportant in CRPS I patients without SMP. Modified from [6].

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ConCluSIonS

involving these neuronal systems and their central representations. 2. The peripheral changes (sympathetic-afferent coupling, vascular changes, inflammation, edema, trophic changes) cannot be seen independently of the central ones. It is postulated that a mismatch between the afferent and efferent signals occurring on different levels of integration in the afferent and efferent body maps in the CNS cause the changed autonomic, sensory and somatomotor reactions. 3. Using imaging techniques we will learn which cortical and subcortical changes are specific for CRPS and how these central changes are expressed in the efferent (somatomotor and autonomic) systems and the distorted sensory perceptions of the body.

Acknowledgement This work was supported by the Deutsche Forschungsgemeinschaft, the German Ministry of Research and Education within the German Research Network on Neuropathic Pain (BMBF, 01EM01/04).

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[2] Baron R, Schattschneider J, Binder A, Siebrecht D, Wasner G. Relation between sympathetic vasoconstrictor activity and pain and hyperalgesia in complex regional pain syndromes: a case-control study. Lancet 2002;359:1655–60. [3] Harden RN, Baron R, Jänig W, editors. Progress in pain research and management, vol. 22. Seattle: IASP Press; 2001. [4] Jänig W. The integrative action of the autonomic nervous system Neurobiology of homeostasis. Cambridge New York: Cambridge University Press; 2006. [5] Jänig W. Autonomic nervous system and pain. In: Basbaum AI, Bushnell MC, editors. In science of pain. San Diego: Academic Press; 2009. p. 193–225. [6] Jänig W, Baron R. Complex regional pain syndrome: a mystery explained? The Lancet Neurol 2003;2:687–97. [7] Price DD, Long S, Wilsey B, Rafii A. Analysis of peak magnitude and duration of analgesia produced by local anesthetics injected into sympathetic ganglia of complex regional pain syndrome patients. Clin J Pain 1998;14:216–26. [8] Rommel O, Malin J-P, Zenz M, Jänig W. Quantitative sensory testing, neurophysiological and psychological examination in patients with complex regional pain syndrome and hemisensory deficits. Pain 2001;93:279–93. [9] Stanton-Hicks M, Jänig W, Hassenbusch S, Haddox JD, Boas R, Wilson P. Reflex sympathetic dystrophy: changing concepts and taxonomy. Pain 1995;63:127–33. [10] Wasner G, Schattschneider J, Heckmann K, Maier C, Baron R. Vascular abnormalities in reflex sympathetic dystrophy (CRPS I): mechanisms and diagnostic value. Brain 2001;124:587–99.

References [1] Baron R. Complex regional pain syndromes. In: Basbaum AI, Bushnell MC, editors. In science of pain. San Diego: Academic Press; 2009. p. 909–18.

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122 Carcinoid Tumors Kenneth R. Hande Carcinoid tumors arise from neuroendocrine cells. They comprise part of a larger group of cancers referred to as neuroendocrine tumors, which include islet cell tumors, large cell neuroendocrine tumors, atypical carcinoids and small cell carcinomas [1]. The incidence of neuroendocrine tumors appears to be increasing with five cases/100,000 persons reported during 2004 [2]. Since the growth rate of carcinoid tumors is generally slow and patients may live with their disease for many years, the prevalence of carcinoid tumors in the population is higher (over 100,000 in the USA) than esophageal (28,000), gastric (65,000) or pancreatic cancer (65,000). Seventy percent of carcinoids originate within the lung and GI tract. Within the GI tract, 42% are in the small bowel, 27% in the rectum and 8% in the stomach [1,2]. Most small bowel carcinoids occur in the ileum. Carcinoid tumors can arise anywhere in the body. Typical carcinoids are composed of monotonously similar cells with round nuclei, pink granular cytoplasm and few mitosis. There is significant heterogeneity in carcinoid tumors with different pathological staining patterns, different rates of metastasis and differing ability for neuropeptide secretion depending on their anatomical location (Table 122.1). Pulmonary carcinoid tumors comprise 1–2% of lung tumors and 25% of all carcinoid tumors. The 5-year survival rate of resected typical pulmonary carcinoids is 90% which contrasts with a 50% cure rate for atypical carcinoids [3]. Pulmonary carcinoids can be incidentally found on routine radiographs or present with symptoms of bronchial obstruction (pneumonia, cough, hemoptysis). They can secrete ACTH, growth hormone or serotonin, but only rarely (5%) result in Cushing’s syndrome, acromegaly, or carcinoid syndrome. Gastric carcinoids are divided into three types: (i) those associated with chronic atrophic gastritis (CAG); (ii) those associated with MEN-1 or Zollinger–Ellison syndrome; and (iii) sporadic type [1,3,4]. CAG results in hypergastrinemia leading to hyperplasia of multiple benign carcinoids in the stomach. CAG associated carcinoids have an indolent course and rarely metastasize. Sporadic gastric carcinoids are more aggressive than CAG associated tumors and occasionally produce an atypical carcinoid syndrome mediated by histamine and 5-hydroxytryptophan. The flush consists of patchy, serpiginous areas of erythema with defined bonders rather than the common diffuse

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00122-0

rash of most carcinoids. Colon and rectal carcinoids do not secrete serotonin are not associated with the carcinoid syndrome. Rectal carcinoids are often small and have a low risk of metastasis while colon carcinoids have frequent metastasis at the time of diagnosis. Most physicians associate carcinoid tumors with welldifferentiated neuroendocrine tumors of the small bowel which can produce serotonin and other amines. Patients with small bowel carcinoids may present with episodic abdominal pain resulting from partial bowel obstruction caused by mesenteric fibrosis surrounding the carcinoid tumor. Patients may have abdominal symptoms for months to years (average 4–7 years) prior to diagnosis. Other patients may present with the carcinoid syndrome, usually associated with metastasis to the liver. The term “carcinoid syndrome” describes the humoral manifestations of carcinoid tumors (Table 122.2). The carcinoid syndrome results from release of various biologic amines into the systemic circulation. The association between hepatic metastases and carcinoid syndrome is due to efficient inactivation by the liver of amines released into the portal circulation by primary GI tumors. Electron microscopy of carcinoid tumors shows granules of various size and shape that contain many biologic amines including serotonin, tachykinins (such as substance P, neurokininns, and hemekinin), histamine, dopamine, prostaglandins, gastrin, bradykinin, kalikrein, somatostatin, corticotrophin, and neuron-specific enolase. When released into the systemic circulation, these neuropeptides may cause flushing, diarrhea, wheezing, heart disease, and rarely hypotension (the carcinoid syndrome). The released neuropeptides have partially overlapping biologic function. Serotonin, tachykinins and prostaglandins all activate receptors which result in smooth muscle contraction and vasodilatation. Secretion of these neuropeptides usually occurs in concert. For instance, the amount of tachykinin secretion in patients with carcinoid tumors correlates with the rate of serotonin secretion [5]. The biological processes mediating the specific symptoms of the carcinoid syndrome are likely multifactoral in the majority of cases. The typical paroxysmal flushing of the carcinoid syndrome is manifested by transient episodes of erythema usually limited to the face, neck and the upper trunk [6]. Patients may experience a sensation of warmth during

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122. CARCInoId TumoRs

TABLE 122.1 Characteristics of Carcinoid Tumors Foregut (respiratory tract, pancreas, stomach)

Midgut (jejunum, ileum, appendix, cecum)

Hindgut (distal colon, rectum)

Percent of carcinoids

25%

30%

20%

Argentaffin stain

Negative

Positive

/–

Urine excretion

5-HTP, histamine

5HIAA

Negative

Metastasis frequency

10–30%

10–35%

3–32%

Carcinoid syndrome

Atypical type

Typical type

Rare

5-HTP, 5-hydroxytryptophan; 5-HIAA, 5-hydroxyindolacetic acid.

TABLE122.2 Clinical Features of Carcinoid syndrome Feature

Frequency

Diarrhea

70–80%

Flushing

60–75%

Heart disease

20–40%

Telangiectasia

20–25%

Wheezing

10–20%

Pellagra

1–2%

flushing and sometimes palpitations. Although severe flushing and hypotension can occur, most episodes are brief (1–2 minutes) and do not cause dizziness or palpitations. Rare, severe attacks of flushing may be accompanied by shock and syncope. Flushing over a long period can cause a constant facial erythema and persistent cutaneous telangiectasia. Flushing usually occurs spontaneously in the absence of any evident precipitating cause. Serotonin is not the mediator of the flushing. Bradykinin is released in some patients during flushing, but the absence of detectable bradykinin release in other patients suggests that it is not a universal mediator of the flush. Production of the vasodilator prostaglandin E2 is not increased in patients with the carcinoid syndrome. With tumors of the midgut, tachykinins are believed to be mediators of the flushing. Tachykinins are a family of structurally related peptides that exert similar biologic effects, such as vasodilation and contraction of various types of smooth muscle. These peptides include substance P, substance K (neurokinin alpha), and neuropeptide K (an extended form of substance K). The diarrhea associated with the carcinoid syndrome varies from two to 30 stools a day. It is usually a discomfort and an annoyance but not disabling. Occasionally, voluminous diarrhea may cause malabsorption and fluid and electrolyte imbalance. Diarrhea is frequently accompanied by abdominal cramping. A variety of carcinoid

FIGURE 122.1 Metabolism and structure of serotonin and 5-hydroxy indolacetic acid.

tumor products (serotonin, substance P, histamine, etc.) stimulate peristalsis. Small and large bowel transit times are two and six times faster in patients with carcinoid syndrome than normals [7]. Bronchospasm may occur with episodes of flushing. Pellagra from excess tryptophan catabolism may occur but is uncommon (Fig. 122.1). Carcinoid heart disease is manifest pathologically by plaque-like thickening on the endocardium of heart leaflets. Tricuspid regulation occurs in 92% of patients with carcinoid heart disease, tricuspid stenosis in 27%. Leftsided lesions are rare. Carcinoid heart disease is associated with high plasma serotonin concentrations [8]. The most dramatic manifestation of carcinoid tumors is the carcinoid crisis. This is observed in patients who have the more intense syndromes associated with foregut carcinoids or patients who have greatly elevated 5HIAA levels (200 mg/24 hours). Carcinoid crisis is frequently precipitated by physically stressful situations, particularly by induction of anesthesia. Patients will develop an intense generalized flush that persists for hours or days. There may be severe diarrhea. CNS symptoms are common, ranging from mild light-headedness or vertigo through somnolence to deep coma. There are usually associated cardiovascular abnormalities including tachycardiac, rhythm irregularity, hypertension, or severe hypotension. Urinary excretion of 5HIAA, the end metabolite of serotonin metabolism (Fig. 122.1), is used to confirm the diagnosis of carcinoid syndrome. Normal excretion is less than 10 mg/24 hours. Values of more than 25 mg/24 hours are essentially diagnostic of carcinoid syndrome. A sensitivity of 73% and a specificity of 100% have been reported for elevated urinary 5-HIAA in metastatic carcinoid tumors. Serine chromagranin A is elevated in 60–80% of carcinoid tumors and appears to correlate with tumor mass. Somatostatin receptor scintography (111In-pentreotide) is useful in tumor localization with 65–80% sensitivity in detecting metastatic disease [9]. Treatment of localized carcinoid tumors is surgical resection [1,3,9,10]. Carcinoids 2 cm are usually cured. Therapy for patients with unresectable metastatic disease is usually palliative. However, it is important to remember that carcinoid tumors have an indolent

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REFEREnCEs

growth pattern and patients may live many years even with metastatic disease (median survival 5 years). Somatostatin analogues have a central role in the treatment of patients with carcinoid syndrome. Somatostatin is a 14-amino-acid peptide that inhibits the secretion of a broad range of hormones. Somatostatin binds to somatostatin receptors expressed on 80% of carcinoid tumors. Somatostatin analogs, such as octreotide, are effective in relieving the symptoms of carcinoid syndrome in 80% of patients and decrease urinary 5HIAA excretion in 70%. However, regression in tumor size is rarely (5%) seen. Long-acting preparations of somatostatin analogs (lanreotide, octreotide-LAR) can be administered once a month. Chemotherapy has shown only minimal benefit in treatment of carcinoid tumors. Hepatic-artery embolization can be used for hepatic metastases. Urinary 5HIAA excretion and symptoms improve in up to 80% of patients following tumor embolization. Survival benefit from embolization procedures has not been documented. Cytoreductive surgery for hepatic metastasis improves symptoms of carcinoid syndrome in 65–90% of patients [1,9].

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References [1] Pasieka JL. Carcinoid tumors. Surg Clin N Am 2009;89:1123–37. [2] Yao JC, Hassan M, Phan A, et al. One hundred years after “carcinoid”: Epidemiology of and prognostic factors for neuroendocrine tumors in 35,825 cases in the United States. J Clin Oncol 2008;26:3063–72. [3] Pinchot SN, Holen K, Sippel RS, et al. Carcinoid tumors. The Oncologist 2008;13:1255–69. [4] Kulke MH, Mayer RJ. Carcinoid tumors. N Engl J Med 1999;340:858–68. [5] Cunninham JL, Janson ET, Agarwal S, et al. Tachykinins in endocrine tumors and the carcinoid syndrome. Eur J Endocrinology 2008;159:275–82. [6] Moertel CG. An odyssey in the land of small tumors. J Clin Oncol 1987;10:503–1522. [7] Von Der Ohe MR, Camilleri M, Kvols LK, et al. Motor dysfunction of the small bowel and colon in patients with the carcinoid syndrome. N Eng J Med 1993;329:1077–8. [8] Waller JE, Connolly HM, Rubin J, et al. Factors associated with the progression of carcinoid heart disease. N Engl J Med 2003;348:1005–14. [9] Range JK, Davis AHG, Ardell J, et al. Guidelines for the management of gastroentero-pancreatic neuroendocrine (including carcinoid) tumors. Gut 2005;54S iv1-iv16 [10] Kvols LK, Revisiting CG. Moertel’s Land of Small Tumors. J Clin Oncol 2008;31:5005–7.

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123 Paraneoplastic Autonomic Dysfunction Ramesh K. Khurana Paraneoplastic autonomic dysfunction (PAD), as a result of the tumor’s remote effect upon the autonomic nervous system, is rare but extremely important to recognize because (1) it may be the initial manifestation of an underlying malignancy; (2) it may be more devastating than the tumor itself; (3) it may simulate metastatic disease or other conditions such as B12 deficiency and multiple system atrophy (MSA); (4) it may present with widespread or limited autonomic involvement; (5) it may be confirmed with serologic tests; (6) early treatment of the tumor may arrest the progression or improve the autonomic dysfunction; and (7) immunomodulatory therapy may be of benefit. PAD may be immune-mediated or non-immune mediated. The latter (inappropriate diuresis, Cushing’s syndrome, neoplastic fever, etc.), attributed to the ectopic production of hormones and cytokines, is not included in this review. Immune-mediated PAD (Box 123.1) is discussed below. For supplemental details, see Table 123.1. Dysautonomia may be the only manifestation of PAD, or it may occur in combination with recognizable syndromes such as subacute sensory neuropathy. Some of these syndromes are discussed.

presents with hallucinations, short-term memory loss, mood disturbances, and seizures. In women with NMDA receptor antibodies, limbic dysfunction may accompany movement disorder such as choreoathetosis, myoclonus, dyskinesias in face and arms, and opisthotonos-like posture. Autonomic instability may manifest as episodic mydriasis, tachycardia, tachypnea, diaphoresis, and hypertension. This combination of symptoms should arouse suspicion of ovarian teratoma. Excessive daytime sleepiness, narcolepsy-cataplexy, decreased CSF hypocretin, hyperthermia, hypothalamicpituitary dysfunction, gain in weight, and sexual dysfunction incriminate the diencephalic region. The disorder, associated with anti-Ma2 antibodies, targets the upper brainstem structures involving the supranuclear control of vertical gaze and oculomotor nuclei. Horizontal gaze, abducens nuclei, and dorsal nuclei of the medulla may be affected. Patients develop cranial neuropathy, nuclear or supranuclear ophthalmoparesis, dysarthria, dysphagia, and asymmetric mild ataxia. The constellation of upper brainstem and limbic structures is seen in patients with testicular germ-cell tumors.

ENCEPHALITIS

MORVAN’S SYNDROME AND NEUROMYOTONIA

Autoantibodies against multiple antigens including Hu, Ma2, CRMP-5, VGKC, and NMDA receptors may present with involvement of limbic system. Sometimes diencephalon or brainstem may be affected. Limbic encephalitis

Peripheral, central, and autonomic nervous systems may be affected in Morvan’s syndrome and neuromyotonia. Clinical features of peripheral involvement include muscle stiffness, cramps, myokymia, weakness, and

BOX 123.1

C L A S S I F I C AT I O N O F I M M U N E - M E D I AT E D PA R A N E O P L A S T I C AU T O N O M I C D Y S F U N C T I O N I. Involving the central nervous system Encephalitis: limbic, diencephalic, and brainstem II. Involving both central nervous system and peripheral nervous system A. Morvan’s syndrome B. Sensory neuronopathy C. Enteric neuronopathy

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00123-2

III. Involving the peripheral nervous system A. Autonomic neuropathy B. Enteric neuropathy IV. Involving the neuromuscular junction Lambert–Eaton myasthenic syndrome.

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TABLE 123.1 Paraneoplastic neurological syndromes: Clinical and Immunological features and Associated Tumors PNS

Distinguishing Clinical Feature

Antibodies

Common Tumor Type

Other Tumors

Encephalitis

Seizures, memory loss, psychiatric disturbance, dyskinesias, vertical gaze palsy

Anti-Hu, CRMP-5, anti-Ma2, anti-VGKC, anti-NMDAR

SCLC, testicular germ cell, ovarian teratoma

Thymoma, neuroblastoma, prostate, breast, Hodgkin’s lymphoma

Morvan’s syndrome

Neuromyotonia, myokymia, limbic encephalitis

Anti-VGKC

Thymoma

SCLC, Hodgkin’s disease

Sensory neuronopathy

Progressive and severe proprioceptive deficit

Anti-Hu/ANNA-1, anti-amphiphysin

SCLC

GI tract, breast, adrenal glands, uterus, prostate, lymphoma, neuroblastoma, testicular seminoma, embryonal carcinoma

Enteric neuronopathy

Intestinal pseudo-obstruction, gastroparesis

Anti-Hu/ANNA-1, enteric neuronal, ganglionic acetylcholine receptor, N-Type VGCC

SCLC

Pulmonary carcinoid, undifferentiated epithelioma, breast

Autonomic neuropathy

Widespread autonomic failure with little somatic involvement

Anti-Hu/ ANNA-1, CRMP-5

SCLC

Pancreas, bladder, rectum, prostate, thymus, Hodgkin’s disease

Lambert–Eaton myasthenic syndrome

Proximal weakness improves with exercise, decreased CMAP amplitude, abnormal repetitive nerve stimulation test

Anti-VGCC

SCLC

Breast, GI tract, GU tract, prostate, gall bladder, lymphosarcoma

ANNA-1, antineuronal nuclear antibody; CMAP, compound muscle action potential; CRMP 5, collapsin response-mediator protein; GI, gastrointestinal tract; GU, genitourinary; PNS, paraneoplastic neurological syndrome; SCLC, small cell lung cancer; VGCC, voltage-gated calcium channel; VGKC, voltage-gated potassium channel.

pseudomyotonia. Central involvement may manifest as hallucinations, fluctuating cognition, impairment of recent memory, complex nocturnal behavior, and insomnia. Autonomic involvement is characterized by hyperhidrosis, increased salivation and lacrimation, tachycardia, cardiac arrhythmias, severe constipation, and urinary incontinence. Paroxysms of sweating, piloerection, and salivation may occur. Electromyography shows spontaneous firing of motor units as doublet, triplet, or multiple discharges occurring at irregular intervals with a high intraburst frequency. The passive transfer of neuromyotonia IgG increases neuronal excitability in mice. Antibodies against voltage-gated potassium channels (VGKC) have been demonstrated in the serum and on neuronal dendrites in the hippocampus, thalamic neurons, and peripheral nerves. Patients show improvement following plasma exchange and immunosuppression with prednisone and azathioprine.

SUBACUTE SENSORY NEURONOPATHY First described by Denny Brown in 1948, it is characterized by subacute onset and asymmetric distribution of dysthesiae, lancinating pains, and numbness affecting the limbs, face, and tongue. Loss of proprioception, sensory ataxia, pseudo-athetosis, and areflexia are common, while motor strength is spared. This syndrome may mimic subacute combined degeneration or tabes dorsalis.

Electrophysiological studies show absent or markedly reduced sensory nerve action potentials. Autonomic dysfunction is present in over one third of cases. Tests show local or widespread sympathetic and parasympathetic insufficiency. Hu antigens are expressed throughout the central nervous system, sensory and sympathetic ganglia, and cancer cells. Anti Hu/ANNA-1 is the serologic marker.

ENTERIC NEURONOPATHY Pseudo-obstruction of bowels, a distinguishing feature of this illness, may precede or follow the diagnosis of tumor. Postural dizziness, syncope, and other symptoms may follow. Somatic neurologic findings are of variable severity and affect the peripheral or central nervous system. The syndrome may mimic MSA, but pseudo-obstruction of bowels in MSA is rare. Myenteric plexus neurons display lymphocytic infiltration and progressive loss. Serum antibodies from patients can induce cell death in cultured myenteric plexus neurons.

AUTONOMIC NEUROPATHY Autonomic dysfunction may occur with minimal or no somatic involvement. There is subacute onset of orthostatic, gastrointestinal, genitourinary, and pupillary

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DIAgnosIs

symptoms. Autonomic tests show widespread parasympathetic and sympathetic dysfunctions. The patients may be seropositive for ganglionic acetylcholine receptor (AchR) autoantibodies. Serum ganglionic AchR antibody levels correlate with severity of autonomic dysfunction. Administration of these antibodies to mice results in autonomic dysfunction. Experimental autoimmune autonomic ganglionopathy can be reproduced in rabbits by immunizing against AchR.

Lambert–Eaton Myasthenic Syndrome Lambert–Eaton myasthenic syndrome (LEMS) presents with symmetrical proximal weakness, reduced or absent reflexes, and ptosis. Autonomic symptoms (subclinical excepting dry mouth) occur in about 75% of patients. Autonomic tests show widespread cholinergic and adrenergic abnormalities. The electrophysiological hallmarks are low amplitude of the compound muscle action potentials, decremental responses on 2- to 5-Hz nerve stimulation, and augmentation 100% on 20- to 50-Hz stimulation or following 10 seconds of voluntary exercise. Almost all patients show autoantibody against PQ-type voltage-gated calcium channels (VGCC). Autoantibodies against N-type VGCC, important for transmitter release from autonomic nerve terminals are reported in about 30% of patients. Pathogenecity of the VGCC antibodies is proved by expression of VGCC antigen on cancer cells, passive transfer of the human disorder to mice with

the injection of LEMS IgG, antibody binding to the active zone particles (AZPs) of the presynaptic calcium channels, reduced number and disorganization of AZPs demonstrated by freeze-fracture studies of the patient’s neuromuscular junction, reduced presynaptic quantal release of acetylcholine, and relatively rapid improvement following removal of antibodies with plasma exchange. LEMS IgG, found to impair transmitter release from parasympathetic (bladder) and sympathetic (vas deferens) neurons, is probably responsible for autonomic dysfunction. However, contribution of other factors such as antibodies against neuronal ganglionic acetylcholine receptor cannot be excluded.

DIAGNOSIS Urgent evaluation is necessary to prevent progressive neuronal loss, especially in CNS syndromes. Paraneoplastic disorder should be suspected in patients presenting with acute or subacute onset of progressive autonomic symptoms such as orthostatic hypotension, dry mouth, urinary retention, and constipation (see algorithm in Box 123.2). Occurrence of recognizable syndromes such as LEMS, limbic encephalitis, myokymia, pseudoobstruction of bowels, or progressive proprioceptive deficit should strengthen this suspicion. It is important to be aware that the spectrum of neurological manifestations is broader and more multifocal than initially described

BOX 123.2

ALGORITHM Autonomic symptoms with or without somatic neurologic symptoms

↓ Neurological examination Brain MRI with contrast CSF examination, nerve conduction studies Autonomic evaluation

↓ Autonomic dysfunction with a distinguishing feature (e.g., intestinal pseudo-obstruction in patients with enteric neuropathy)

↓ Exclude other causes with similar presentation Use appropriate laboratory tests Obtain antibodies titers against the most frequent tumors

↓ If serology positive, work-up for commonly suspected tumor. If serology negative,

↓ Screen for other tumors

↓ If work-up negative, longitudinal follow-up and repeat studies every 6 months up to 4 years

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in “syndromic” patients due presumably to evolving immune responses to multiple onconeural antigens in a single tumor. The diagnosis is made by confirming severity and distribution of autonomic dysfunction, by exclusion of other causes of autonomic dysfunction, by electrophysiological studies in patients with neuromyotonia or suspected LEMS, and by serologic studies for onconeural antibodies. Since most syndromes and tumors are related to more than one antibody, screening for a panel of antibodies provides a better yield. These antibodies constitute a diagnostic tool and may suggest the likely type and location of the neoplasm, but their pathogenetic role (except in LEMS and Morvan’s syndrome) is still unclear. These antibodies can be absent in patients with paraneoplastic neurologic syndrome. Since tumors producing paraneoplastic manifestations are usually small, a diligent search with appropriate tests is mandatory. Close longitudinal follow-up every 3 to 6 months and repeat studies may be necessary for up to four years to diagnose the tumor.

TREATMENT A proper diagnosis can spare the patients unnecessary surgery in cases of pseudo-obstruction of the bowels. Treatment may be directed at the tumor, the antibodies, and the symptoms. An early surgical and cytotoxic reduction of the tumor may diminish autonomic dysfunction in some patients. Use of steroids, plasmapheresis, intravenous immunoglobulin, or immunosuppression with cyclophosphamide to reduce the antibody may provide benefit. Immune therapy has no or modest effect on CNS syndromes, whereas such therapy is beneficial for syndromes affecting the peripheral nervous system and neuromuscular junction. For example, plasma exchange results in clinical and electrophysical improvements in patients with LEMS or Morvan’s syndrome. The patients

with LEMS show symptomatic benefit from drugs such as 3,4-diaminopyridine, which enhances cholinergic function. Symptomatic improvement of orthostatic hypotension with fludrocortisone and pressor medications (Midodrine), dry mouth with artificial saliva, dry eyes with artificial tears, and neoplastic fever with naproxen can improve the quality of life. Acetylcholinesterase inhibitors such as pyridostigmine may improve orthostatic hypotension, salivation, lacrimation, and bowel hypomotility.

Further Reading Dalmau J, Graus F, Rosenblum MK, Posner JB. Anti-Hu associated paraneoplastic encephalomyelitis/sensory neuronopathy. A clinical study of 71 patients. Medicine 1992;71:59–72. Dalmau J, Rosenfeld MR. Paraneoplastic syndromes of the CNS. Lancet Neurol 2008;7:327–40. Dalmau J, Tüzün E, Wu H-Y, Masjuan J, Rossi JE, Voloschin A, et al. Paraneoplastic Anti-N-Methyl-D-aspartate receptor encephalitis associated with ovarian teratoma. Ann Neurol 2007;61:25–36. Dalmau J, Graus F, Villarejo A, Posner JB, Blumenthal D, Thiessen B, et al. Clinical analyses of anti-Ma2-associated encephalitis. Brain 2004;127:1831–44. Dhamija R, Tan KM, Pittock SJ, Foxx-Orenstein A, Benarroch E, Lennon VA. Serological profiles aiding the diagnosis of autoimmune gastrointestinal dysmotility. Clin Gastroenterol Hepatol 2008;6:988–92. Khurana RK. Neoplasia and the autonomic nervous system IN: Appenzeller, O, editor. Handbook of Clinical Neurology: The Autonomic Nervous System. Part II, vol. 75. Amsterdam: Elsevier Science BV; 2000. p. 527–49. Quartel A, Turbeville S, Lounsbury D. Current therapy for Lambert– Eaton myasthenic syndrome: development of 3,4-diaminopyridine phosphate salt as first-line symptomatic treatment. Current Medical Research and Opinion 2010;26:1363–75. Tan KM, Lennon VA, Klein CJ, Boeve BF, Pittock SJ. Clinical spectrum of voltage – gated potassium channel autoimmunity. Neurology 2008;70:1883–90. Titulaer MJ, Soffietti R, Dalmau J, Gilhus NE, Giometto B, Graus F, et al. (2010). Screening for tumors in paraneoplastic syndromes: report of an EFNS task force. European Journal of Neurology. doi:10.1111/j.1468-1331.2010.03220.x. Vernino S. Antibody testing as a diagnostic tool in autonomic disorders. Clin Auton Res 2009;19:13–19.

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124 Abdominal Pain and Cyclic Vomiting Gisela Chelimsky, Thomas Chelimsky Rising interest in the “brain–gut connection” originates from recent developments in functional gastrointestinal disorders (FGIDs). These highly prevalent disorders clearly involve both the autonomic and enteric nervous systems, yet the role of each in disease and symptom production remains unknown. FGIDs are evolving into a model for the study of the brain–gut connection, which is clearly bi-directional. The current bio-psychosocial conceptualization of FGIDs best describes the multiple factors involved in their pathogenesis. In a simplistic way, the brain–gut connection can be viewed as the central nervous system (brain) and the enteric nervous system (gut) being connected through both an afferent sensory system and an efferent system, the autonomic nervous system. Therefore, this model requires an understanding of factors pertaining to the brain, the gut, and this bidirectional connection. The model begins with predisposing contributions to the development of an FGID including both genes and early life experiences. Superimposed psychological factors (“the brain”) then modulate the individual’s coping strategies, to affect the gastrointestinal symptoms themselves, and their impact on the person’s experience. Importantly, this model proposes that psychological factors do not cause FGID, but only affect disease response. However, both the chronic illness itself and the medical system’s lack of preparation to handle this patient population, resulting in years of disregarded or disbelieved symptoms, inaccurate diagnoses, and ineffective management, may engender secondary psychological consequences on the individual’s response to the disease. This added burden may lead patients to “prove” to prospective health providers that their symptoms are “real”, altering the normal patient–physician dynamic at the outset. Subjects with FGID also harbor physiologic changes at the level of “the gut”. Hyperalgesia, allodynia, altered mucosa permeability [1], increased amount of bacteria in the intestine and small bowel bacterial overgrowth [2] sometimes with a component of altered motility [3] have been demonstrated in these subjects. For example, subjects with irritable bowel syndrome demonstrated lower visceral perceptual threshold to rectal balloon inflation than healthy controls. This hyperalgesia is not limited to the

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gastrointestinal tract as many of these subjects also have somatic hypersensitivity [4]. Finally, the link between the brain and the enteric nervous system may be affected. The enteric nervous system coordinates and affects the gastrointestinal motility, secretions and perfusion. Constant afferent information normally travels from the gut to the central nervous system. A healthy subject is completely unaware of the consequent homeostatic reflexes generated by the brain. However, in the disease state, subjects will perceive pain or discomfort even with physiological events [5]. This was observed often in females with irritable bowel syndrome, who will report pain when a balloon is inflated in the sigmoid, while the normal controls would not report the event as painful [6]. Evidence suggests both an alteration in the afferent signaling system itself and in the responsiveness of the CNS circuits [5]. Efferent gut control may also be abnormal, though it is not clear if this constitutes an intrinsic autonomic abnormality, or simply a consequence of CNS integrative dysfunction. Most patients with any type of an FGID will demonstrate a skew toward higher sympathetic tone, and will frequently have orthostatic intolerance or even a postural tachycardia syndrome (POTS) on formal autonomic testing [7]. Whether such a sympathetic skew mitigates or contributes to gut dysfunction is unknown. FGIDs are currently classified according the ROME III criteria (Box 124.1). In adults, there are six main groups of disorders (the subgroups have been omitted in groups not described in this chapter) [3]. This review addresses cyclic vomiting syndrome and chronic abdominal pain.

CYCLIC VOMITING SYNDROME (CVS) The current CVS definition requires two or more periods of unexplained intense nausea and unremitting vomiting or retching lasting hours to days with interictal return to usual state of health lasting weeks to months [3]. A typical episode is divided in four phases: (i) the interictal phase in which the subject is at baseline health;

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BOX 124.1 1. Functional esophageal disorders 2. Functional gastroduodenal disorders: (a) Functional dyspepsia (b) Belching disorders (c) Nausea and vomiting disorders: l Chronic idiopathic nausea l Functional vomiting l Cyclic vomiting syndrome (d) Rumination syndrome in adults 3. Functional bowel disorders

(a) Irritable bowel syndrome (b) Functional bloating (c) Functional constipation (d) Functional diarrhea (e) Unspecified functional bowel disorder 4. Functional abdominal pain syndrome (a) (No subgroups described) 5. Functional gallbladder and Sphincter of Oddi disorder 6. Functional ano-rectal disorder.

(ii) the prodrome phase, where the subject knows the episode is coming, may feel some nausea but is still able to keep food down; (iii) the emetic phase, in which the subject has intense nausea and vomiting; and (iv) recovery phase with return to baseline, with discontinuation of vomiting and development of hunger, which interestingly often happens quite suddenly “like a switch” [8]. The North American Society of Pediatric Gastroenterology, Hepatology and Nutrition (NASPGHAN) published a 2008 consensus statement on the diagnosis and treatment of CVS. In their criteria for CVS, NASPGHAN specifies that the episodes have to be stereotypical, at least 1 week apart and not produced by other causes [9]. Since no test can confirm the diagnosis of CVS, detailed history and physical examination are crucial. Many other CVS mimickers require exclusion, occurring in about 10% of children where the subject needs prompt evaluation and treatment for an underlying disorder [10]. The NASPGHAN consensus recommends performing upper gastrointestinal series at least to the angle of Treitz to rule out an anatomical problem like malrotation and checking electrolytes and glucose prior to starting intravenous fluid. Hyponatremia or hypoglycemia raises concern about Addison’s disease. Other diagnostic considerations include recurrent pancreatitis, uretero-pelvic junction obstruction, gallbladder disease, hepatitis and rarely porphyria [10]. Disorders of fatty acid oxidation, urea cycle defects and mitochondrial disorders can have also a similar pattern. Neurological disorders increasing intracranial pressure like posterior fossa or hypothalamic tumors, Chiari malformations, subdural hematomas and hydrocephalus are also in the list of differential diagnoses. The evaluation would depend on the associated symptoms, response to treatment and detailed general and neurological examination [9,10]. Idiopathic CVS is considered a migraine variant. In 82% of CVS cases, subjects either develop migraine headaches later in life or harbor a family history of migraines [11]. Further, Boles demonstrated a strong maternal inheritance in all subjects with CVS, reported a higher incidence of

migraine, irritable bowel syndrome, depression and hypothyroidism in the affected matrilineal relatives, and found that hypotonia and ADHD more commonly occurred in affected probands [12]. Twenty-five percent of his subjects with CVS suffered from co-existing neuromuscular problems such as cognitive disorders, skeletal myopathies, cranial nerve dysfunction, or seizure disorders (“CVS plus group”). This group also had earlier onset of symptoms than a CVS group without associated neuromuscular issues (“CVS minus”), more autonomic co-morbidities such as migraine headache, chronic fatigue and complex regional pain [13]. Both in subjects with migraine without aura and in some subjects with CVS 2 common mitochondrial DNA polymorphisms have been described which may play a pathogenetic role [14]. Cyclic vomiting syndrome has been associated with autonomic dysfunction. Several reports in adults and in children show a sympathetic nervous system impairment with postural tachycardia and/or sudomotor dysfunction [15–17].

CHRONIC ABDOMINAL PAIN Many FGIDs present with abdominal pain. All the diagnoses are defined based on ROME III criteria, which mandate 3 months of symptoms over the last 6 months prior to diagnosis. Functional dyspepsia requires one of the following criteria with a normal endoscopy: discomfort after eating, early satiety, epigastric pain or epigastric burning. Epigastric pain syndrome differs from functional dyspepsia in requiring no relationship of symptoms to eating, but this may be hard to sort out. Criteria include at least moderate intermittent epigastric pain or burning at least once per week, not relieved by defecation or passage of flatus and does not fulfill the criteria for gallbladder and sphincter of Oddi disorder. Irritable bowel syndrome can also present with abdominal pain with criteria including recurrent abdominal pain or discomfort at least 3 days per month in the last three months associated with two or more of

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the following: (a) improvement with defecation; (b) onset associated with a change in stool frequency; (c) onset associated with a change in form of stool. Continuous or nearly continuous pain is also a key component in functional abdominal pain syndrome, where the pain bears no or only occasional relationship with physiological events like eating, defecation or menses. In addition, the subject must experience some loss of daily function and obviously, the pain cannot be feigned. Symptoms cannot meet criteria for other FGID [3]. The role of the autonomic nervous system in the development of IBS is poorly understood, with several studies with inconsistent results. A general finding does support increased sympathetic nervous system activity on tilt table test. However, Aggarwal et al. found that subjects with IBS-constipation have vagal dysfunction and that the diarrhea predominant group had more sympathetic dysfunction [18,19]. In relation to functional dyspepsia, we have shown that children with POTS have worsening of the normal gastric electrical activity when in the upright position suggesting that orthostatic stress may produce either delayed gastric emptying or abnormal accommodation, which may explain the upper gastrointestinal symptoms [20]. Furthermore, both vomiting and upper abdominal discomfort are very common in children with POTS [21]. Treatment has not been standardized for any of these disorders. Frequent treatment regimens for functional dyspepsia include: (i) acid suppression; (ii) peppermint oil, a spasmolitic based on its calcium-antagonist properties; (iii) caraway oil hypothetically increases muscle tone; (iv) domperidone increases antroduodenal motility; (v) tegasarod, a 5-HT4 partial agonist and 5 HT2b antagonist that accelerates gastric emptying; it has been withdrawn from the market due to cardiac ischemia events; (vi) selective serotonin-reuptake inhibitors (SSRIs) relax the gastric fundus and modify gastric sensorimotor function; (vii) sumatriptan which is an 5HT 1B/D receptor antagonist may relax also the gastric fundus and affect the perception threshold; and (viii) antidepressants by affecting the central processing of pain, helping with psychiatric issues and improving sleep [22]. There is currently more literature addressing IBS treatment than functional dyspepsia or functional abdominal pain syndrome. The serotonin agents are also useful in IBS. Tegasarod was mainly used in IBS-constipation in women. Alosetron, a 5HT3 antagonist is used in women with severe IBS-diarrhea. Ischemic colitis and severe constipation have been reported with alosetron, therefore it is only used with great caution in specific circumstances. Antidepressants are also used here for the similar reasons to their use in functional dyspepsia, namely altering gastrointestinal sensation, while simultaneously addressing any psychiatric co-morbidities. In subjects with IBS-constipation, lubiproston may be useful. This medication works on the epithelial cell chloride channel increasing fluid and electrolyte secretion into the small intestine,

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resulting in an easier bowel movement. Dietary changes, increasing bulk and fiber in IBS-diarrhea predominant, avoiding fructose in the subgroup that is fructose intolerance may also help. Dietary changes should be first line, mainly when symptoms are not very severe. Laxatives can also be used in the constipation predominant group. The use of rifaximin, a non-absorbable antibiotic may also reduce symptoms in the IBS-diarrhea group by addressing a presumably dysfunctional balance of gut flora. This antibiotic is FDA approved for the treatment of traveler’s diarrhea [23]. In conclusion, FGIDs are highly complex disorders that are still not well understood, in part due to the number of gastrointestinal and non-gastrointestinal factors involved in the pathogenesis. The autonomic nervous system may be playing a role in the development of symptoms, but so far the data are limited and the clinical implications unclear.

References [1] Marshall JK, Thabane M, et al. Intestinal permeability in patients with irritable bowel syndrome after a waterborne outbreak of acute gastroenteritis in Walkerton, Ontario. Aliment Pharmacol Ther 2004;20(11–12):1317–22. [2] Pimentel M, Chow EJ, et al. Eradication of small intestinal bacterial overgrowth reduces symptoms of irritable bowel syndrome. Am J Gastroenterol 2000;95(12):3503–6. [3] Drossman D, Corazziari E, et al. ROME III: The Functional Gastrointestinal Disorders. McLean, Virginia: Degnon Associates, Inc; 2006. [4] Wilder-Smith CH, Robert-Yap J. Abnormal endogenous pain modulation and somatic and visceral hypersensitivity in female patients with irritable bowel syndrome. World J Gastroenterol 2007;13(27):3699–704. [5] Mayer EA, Tillish K. The brain–gut axis in abdominal pain syndromes. Annu Rev Med 2011(62):22.1–22.16. [6] Munakata J, Naliboff B, et al. Repetitive sigmoid stimulation induces rectal hyperalgesia in patients with irritable bowel syndrome. Gastroenterology 1997;112:55–63. [7] Chelimsky G, Boyle JT, et al. Autonomic abnormalities in children with functional abdominal pain: coincidence or etiology?. J Pediatr Gastroenterol Nutr 2001;33(1):47–53. [8] Fleisher DR, Gornowicz B, et al. Cyclic Vomiting Syndrome in 41 adults: the illness, the patients, and problems of management. BMC Med 2005;3:20. [9] Li BU, Lefevre F, et al. North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition consensus statement on the diagnosis and management of cyclic vomiting syndrome. J Pediatr Gastroenterol Nutr 2008;47(3):379–93. [10] Li BU, Murray RD, et al. Heterogeneity of diagnoses presenting as cyclic vomiting. Pediatrics 1998;102(3 Pt 1):583–7. [11] Li BU, Murray RD, et al. Is cyclic vomiting syndrome related to migraine?. J Pediatr 1999;134(5):567–72. [12] Boles RG, Adams K, et al. Maternal inheritance in cyclic vomiting syndrome. Am J Med Genet A 2005;133(1):71–7. [13] Boles RG, Powers AL, et al. Cyclic vomiting syndrome plus. J Child Neurol 2006;21(3):182–8. [14] Zaki EA, Freilinger T, et al. Two common mitochondrial DNA polymorphisms are highly associated with migraine headache and cyclic vomiting syndrome. Cephalalgia 2009;29(7):719–28. [15] Rashed H, Abell TL, et al. Autonomic function in cyclic vomiting syndrome and classic migraine. Dig Dis Sci 1999;44(Suppl. 8):74S–8S.

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[16] Chelimsky T, Chelimsky G. Autonomic Abnormalities in Cyclic Vomiting Syndrome. J Pediatr Gastroenterol Nutr 2007;44:326–30. [17] Venkatesan T, Prieto T, et al. Autonomic nerve function in adults with cyclic vomiting syndrome: a prospective study. Neurogastroenterol Motil 2010;22:1303–7 [18] Aggarwal A, Cutts T, et al. Predominant Symptoms in Irritable Bowel Syndrome Correlate With Specific Nervous System Abnormalities. Gastroenterology 1994;106:945–50. [19] Manabe N, Tanaka T, et al. Pathophysiology underlying irritable bowel syndrome – from the viewpoint of dysfunction of autonomic nervous system activity. J Smooth Muscle Res 2009;45(1):15–23.

[20] Safder S, Chelimsky TC, et al. Gastric Electrical Activity Becomes Abnormal in the Upright Position in Patients With Postural Tachycardia Syndrome. J Pediatr Gastroenterol Nutr 2010;51:314–8 [21] Ojha A, Chelimsky TC, et al. Comorbidities in pediatric patients with postural orthostatic tachycardia syndrome. J Pediatr 2011;158(1):119–22. [22] Brun R, Kuo B. Functional dyspepsia. Therap Adv Gastroenterol 3(3): 145–164. [23] Khan S, Chang L. Diagnosis and management of IBS. Nat Rev Gastroenterol Hepatol 7(10): 565–81.

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125 Fecal Incontinence James F.X. Jones PREVALENCE OF THE CONDITION Fecal incontinence has been reported to affect 11% of the adult general population in the UK [1]. But the prevalence of any condition depends on the definition of that condition. In clinical practice incontinence scores are required to grade severity of the disease and care is required to avoid conflating fecal and flatal incontinence. It can be quite difficult to ascertain the burden of disease because it is probably under reported due to patient embarrassment. Of particular note, the pudendal nerve is often injured after traumatic childbirth which may result in postpartum fecal incontinence [2].

THE PHYSIOLOGICAL MECHANISMS THAT SUSTAIN CONTINENCE OF FECES There are three barriers and one important reflex that provide continence to feces. The internal and external anal sphincters are arranged as concentric cylinders and the puborectalis forms a sling around the anal canal. During rectal distension an urge to defecate forms in the sensorium and a spinal reflex operates to increase tonus of the striated external anal sphincter (recto-anal excitatory reflex).

Internal Anal Sphincter (IAS) The IAS has inner circular and outer longitudinal coats and most attention has been paid to the inner circular layer because it generates the pressure in the anal canal. The major contribution of the IAS to resting tone (75%) was established by comparing conscious and curarized patients under general anesthesia. The origin of the IAS tone is both intrinsic (myogenic) and extrinsic (adrenergic). In the 19th century Langley demonstrated that lumbar sympathetic nerves constrict the IAS and cause pallor of anal mucosa. Many studies have demonstrated that isolated IAS increase tone in response to adrenoceptor agonists including human and the subtype is probably alpha1A/L [3]. Pelvic parasympathetic fibres can modulate the excitatory action of sympathetic hypogastric nerve stimulation by presynaptic cholinoceptor inhibition of noradrenaline release. Lestar et al. (1989) estimated the various contributions to resting anal pressure and found that 30% of total

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pressure was due to striated sphincter tonic activity, 45% of it via neurogenic IAS activity, 10% to purely myogenic IAS activity and 15% attributed to the expansion of the hemorrhoidal plexuses in the anal cushions [4].

External Anal Sphincter (EAS) The EAS is a striated muscle under voluntary control which is supplied by the inferior rectal nerve, a branch of the pudendal nerve. In the rat it differs considerably from its urinary counterpart, the striated external urethral sphincter, being much more fatigable and containing less oxidative enzymes [5]. In humans the EAS contains increasing proportions of slow twitch type I fibres with development but in the rat EAS, type 2A muscle fibers have the greatest areal density [5].

Puborectalis Although some radiologists and surgeons have proposed that the human puborectalis should be considered part of the external anal sphincter complex rather than the levator ani; the arguments against this ill-considered view are based on anatomy, embryology and innervation rather than proximity. The puborectalis has a vertical component near its insertion at the rectum which elevates the anal canal upon contraction. The length and vertical descent of the puborectalis during a Valsalva maneuver can provide useful information about pelvic floor laxity.

Recto-Anal Reflexes During defecation the recto-anal inhibitory reflex (RAIR) is triggered by rectal distension and evokes profound relaxation of the IAS. ATP and NO are responsible for the fast phasic and slow sustained relaxations, respectively, of strips of rat IAS. However, continence is sustained by a recto-anal excitatory reflex (RAER) which increases the tone of the EAS. During maximal voluntary activation of the EAS, the RAER can augment the canal pressures still further. Rectal distension can increase the maximal squeeze pressure of the anal canal by about onethird in both male and female human subjects [6]. This phenomenon may play an important role in fecal continence as it resembles the guarding reflex of the bladder;

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a sustained improvement of incontinence [8]. There was no effect on anal resting or squeeze pressures. A subsequent study by that group showed that there was no effect on the recto-anal angle either. The mechanism of action of sacral neuromodulation is probably sensory as posterior tibial nerve stimulation is also effective (a large randomized double blind trial was completed in 2010 by Peters et al. [9]).

ANIMAL MODELS FIGURE 125.1 The sphincters of continence. The puborectalis muscle forms a sling around the rectum to regulate the ano-rectal angle. The external anal sphincter (EAS) and internal anal sphincter (IAS) act in unison to create the resting anal pressure.

a phenomenon where the external urethral sphincter increases its activity on bladder distension.

RECTO-ANAL SENSATION AND THE URGE TO DEFECATE Small diameter mechanosensory fibres (Aδ and C) in the rectal mucosa signal the arrival of diverse agents of distension (solid, liquid or gas). According to the old rectoanal sampling hypothesis of Duthie and Bennett (1963) the RAIR that is evoked by the rectal afferents allows the contents to reach the anal canal for sampling by additional discriminative receptive fields. Mechanotransduction sites in the IAS are associated with fine varicose intramuscular arrays [7]. Gaseous and solid stimuli may be differentiated by this mechanism but little work has been carried out on the neural processing of such different stimuli.

FECAL INCONTINENCE If physiology is the basis of scientific medicine, pathology is just physiology with obstacles. Consideration of the physiological mechanisms of continence immediately suggests different mechanisms of disease. The possibilities include disorder of EAS, IAS, puborectalis or the RAER and there is evidence for such dysfunctions (e.g., the RAER can be abnormal in cases of idiopathic fecal incontinence). As soiling may occur without a concomitant urge to defecate, disordered ano-rectal sensation and/or perception may also be a major defect.

THE MECHANISM OF ACTION OF SACRAL NEUROMODULATION Chronic sacral nerve root stimulation is a minimally invasive, safe but expensive therapy for fecal incontinence. Uludag et al. (2010) followed a large cohort of patients treated with sacral neuromodulation for 7 years and found

The author has created an animal model of obstetric related injury to the pudendal nerve by the pelvic compression created by retro uterine balloon inflation. This model displays atrophy of EAS and IAS and signs of sphincter denervation. In addition small diameter sensory neurones are injured in the S1 dorsal root ganglia and there is diminution of somatosensory cortical potentials evoked by anal canal stimulation [10]. This last finding is of particular interest because recent experiments show that both sacral neuromodulation and posterior tibial nerve stimulation produce a long term potentiation of cortical evoked potentials. Work is ongoing to translate these findings back to the perineal clinic and human neurophysiological laboratory.

References [1] Buckley BS, Lapitan MC. Prevalence of urinary and fecal incontinence and nocturnal enuresis and attitudes to treatment and helpseeking amongst a community-based representative sample of adults in the United Kingdom. Int J Clin Pract 2009;63(4):568–73. [2] Snooks SJ, Setchell M, Swash M, Henry MM. Injury to innervation of pelvic floor sphincter musculature in childbirth. Lancet 1984;2(8402):546–50. [3] Mills K, Hausman N, Chess-Williams R. Characterization of the alpha1-adrenoceptor subtype mediating contractions of the pig internal anal sphincter. Br J Pharmacol 2008;155(1):110–7. [4] Lestar B, Penninckx F, Kerremans R. The composition of anal basal pressure. An in vivo and in vitro study in man. Int J Colorectal Dis 1989;4(2):118–22. [5] Buffini M, O'Halloran KD, O'Herlihy C, O'Connell R, Jones JF. Comparison of the contractile properties, oxidative capacities and fibre type profiles of the voluntary sphincters of continence in the rat. J Anat 2010;217(3):187–95. [6] Bajwa A, Thiruppathy K, Trivedi P, Boulos P, Emmanuel A. Effect of rectal distension on voluntary external anal sphincter function in healthy subjects. Colorectal Dis. 2010 Sep. 22. doi: 10.1111/j. 1463-1318.2010.02420.x. [Epub ahead of print]. [7] Duthie HL, Bennett RC. The relation of sensation in the anal canal to the functional anal sphincter: a possible factor in anal continence. Gut 1963;4(2):179–82. [8] Uludağ O, Melenhorst J, Koch SM, Van Gemert WG, Dejong CH, Baeten CG. Sacral neuromodulation: long term outcome and quality of life in patients with fecal incontinence. Colorectal Dis. 2010 Oct 19. doi: 10.1111/j.1463-1318.2010.02447.x. [Epub ahead of print]. [9] Peters KM, Carrico DJ, Perez-Marrero RA, Khan AU, Wooldridge LS, Davis GL, et al. Randomized trial of percutaneous tibial nerve stimulation versus Sham efficacy in the treatment of overactive bladder syndrome: results from the SUmiT trial. J Urol 2010;183(4):1438–43. [10] Peirce C, Healy CF, O'Herlihy C, O'Connell PR, Jones JF. Reduced somatosensory cortical activation in experimental models of neuropathic fecal incontinence. Dis Colon Rectum 2009;52(8):1417–22.

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126 Panic Disorder Murray Esler, Marlies Alvarenga, David Kaye, Gavin Lambert, Jane Thompson, Jacqui Hastings, Rosemary Schwarz, Margaret Morris, Jeff Richards*

Some people are subject to episodes of recurring, often inexplicable anxiety. These attacks typically are very unpleasant, and are accompanied by physical symptoms such as sweating, palpitations, tremor and a sensation of suffocation. There may be a precipitating cause, such as being present in a confining space (as in claustrophobia), or in public places (as in agoraphobia), but more commonly the occurrence of the panic attacks is unexpected. Recurring attacks over a period of months, or in many cases years, form the basis for the diagnosis of panic disorder [1]. This is a distressing and often very restricting condition, and is prone to lead to social avoidance behavior, in some instances so extreme that the sufferer never leaves the home. Until recently it has been felt that although panic disorder was distressing and disabling, it did not constitute a risk to life. Sufferers often fear that they have heart disease, because of the nature of their symptoms, but have been reassured that this is not the case. Epidemiological studies, however, now indicate that there is a 2–5 fold increase in the risk of myocardial infarction and sudden death in patients with panic disorder [2,3]. Although this is a significantly increased risk, it is of course still small in individual sufferers, most commonly younger women in whom rates of heart attack in general are very low. The cause is not known, but possibly involves activation of the sympathetic nerves of the heart, predisposing to disturbances of cardiac rhythm and possibly coronary artery spasm. Accordingly, studying the mediating autonomic mechanisms of cardiac risk in panic disorder patients is pertinent, both in terms of devising strategies for primary heart attack prevention [4,5], and in a broader context, for exploring the larger issue of the ways by which mental stress might contribute to cardiac risk.

* Deceased

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00126-8

RESTING SYMPATHETIC NERVOUS SYSTEM FUNCTION IN PANIC DISORDER Sympathetic Nervous Activity and Epinephrine Secretion Rates Multiunit sympathetic nerve firing rates measured directly by microneurography, in the sympathetic outflow to the skeletal muscle vasculature, and rates of norepinephrine spillover from the sympathetic nerves of the whole body are normal in untreated, resting patients with panic disorder, as is the spillover of norepinephrine measured selectively for the sympathetic nerves of the heart [4]. Similarly, adrenal medullary secretion of epinephrine, measured by isotope dilution, is typically normal [4]. Single fiber sympathetic nerve recording does, however, detect striking abnormalities in panic disorder, in the absence of an attack [6]. Single fiber firing rates are elevated, and multiple firings within a cardiac cycle (firing “salvos”) are commonly present. Whether the presence of these nerve firing salvos, which are known to cause high rates of transmitter release, are linked to heart attack risk is unknown. Single fiber sympathetic nerve firing salvos have come to be thought of as providing a “signature” of mental stress exposure [7].

Brain Serotonin Release What might be the CNS origins of this abnormality in central sympathetic outflow? In 34 untreated patients with panic disorder and 24 healthy volunteers a novel method utilizing internal jugular venous sampling, with thermodilution measurement of jugular blood flow, was used to directly quantify brain serotonin turnover, by measuring the overflow of serotonin metabolites from the brain [8]. CNS serotonergic neurons are known to influence sympathetic outflow. Brain serotonin turnover, estimated from jugular venous overflow

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Panic Disorder: Sympathetic Nerve Epinephrine Cotransmission

Faulty Neuronal Reuptake of Noradrenaline in Panic Disorder? Sympathetic Nerve Varicosity

Induction of PNMT in Heart, leading to local synthesis of epinephrine?

Sympathetic Nerves

3H NA

Cardiac release of epinephrine at rest (av. 2.2 ng/min)

MAO

3H DHPG

Neuronal Uptake

Uptake of epinephrine by symp. nerves of heart?

Tritiated DHPG Spillover (P<0.02) PD 1048 (SD 482) dpm/min NS 4231 (SD 2885) dpm/min

Adrenal medullary epinephrine secretion increases 2-6 fold during panic attacks

3H DHPG

3H NA

3H NA Extraction

Tritiated NE Extraction (P<0.01) PD 59 (SD 19) % NS 82 (SD 5) %

FIGURE 126.1 Adrenal medullary secretion of epinephrine increased two- to six-fold during a panic attack. Uptake of epinephrine from plasma into the sympathetic nerves of the heart during epinephrine surges increases the neuronal epinephrine stores, and is perhaps the basis of continuous release of epinephrine, as a sympathetic cotransmitter. An alternative possibility is that epinephrine is synthesised in situ in cardiac sympathetic nerves, by phenylethanolamine methyltransferase (PNMT).

of the metabolite, 5-hydroxyindole acetic acid, was increased approximately 4-fold in panic disorder. The marked increase in serotonin turnover, in the absence of a panic attack, possibly represents the underlying neurotransmitter substrate for the disorder. Support for this interpretation comes from the direct relationship which exists between serotonin turnover and panic disorder illness severity [8].

Epinephrine Cotransmission in Sympathetic Nerves

FIGURE 126.2 Representation of transcardiac processing of tritiated norepinephrine (3H NE) infused intravenously. The majority of tritiated norepinephrine is removed from plasma via a clearance mechanism involving neuronal uptake by sympathetic nerves. Within sympathetic nerves, 3H NE is metabolized to tritiated 3,4-dihydroxyphenylglycol (3H DHPG) by monoamine oxidase (MAO), with some subsequent release into the venous circulation. Tritiated norepinephrine uptake by the heart was reduced in 10 panic disorder patients (PD), 59% (SD 19%) compared with 82% (SD 5%) in healthy subjects (NS) (P  0.01). In parallel, release of tritiated DHPG into the coronary sinus venous drainage of the heart was lower, 1048 dpm/min (SD 482 dpm/min), than in healthy subjects, 4231 dpm/min (SD 2885 dpm/min) (P  0.02). This provides strong phenotypic evidence of impaired neuronal norepinephrine reuptake in panic disorder.

norepinephrine via the norepinephrine transporter [9]. The processes of neuronal reuptake of norepinephrine can be quantified in humans during the course of an infusion of tritiated norepinephrine by analysis of the disposition and intraneuronal processing of the tracer [9] (Fig. 126.2). In untreated patients with panic disorder neuronal reuptake of norepinephrine is impaired (Fig. 126.2). Such an abnormality would be expected to magnify sympathetically mediated responses, particularly in the heart where norepinephrine inactivation is so dependent on neuronal reuptake, causing sensitization to symptom development and predisposing to the development of panic disorder [9].

Release of epinephrine from the sympathetic nerves of the heart, as an accessory neurotransmitter, has been demonstrated in patients with panic disorder (Fig. 126.1). Sympathetic nerves have the capacity to extract circulating epinephrine from plasma, such that during the surges of epinephrine secretion accompanying panic attacks, this process might load sympathetic neuronal vesicles with epinephrine, to be continuously co-released with norepinephrine in the interim periods between attacks (Fig. 126.1) [9]. An alternative explanation is that adrenaline is synthesized within sympathetic nerves in patients with panic disorder [7]. In experimental animals exposed to mental stress, the epinephrine-synthesizing enzyme, phenylethanolamine methyltransferase (PNMT), is induced in sympathetic nerves. Sympathetic nerves of panic disorder patients, obtained from a subcutaneous vein biopsy, do contain PNMT [7], in contrast to healthy people.

Heart rate and blood pressure increase during a panic attack, primarily due to sympathetic nervous system activation and adrenal medullary secretion of epinephrine [4] (Figs. 126.1, 126.3).

Reduction in Neuronal Norepinephrine Reuptake by Sympathetic Nerves

Sympathetic Nerve Firing and Secretion of Epinephrine

Each pulse of the sympathetic neural signal is terminated primarily by reuptake of the released

When recorded directly by microneurography, the size of sympathetic bursts increases remarkably during a panic

AUTONOMIC NERVOUS CHANGES DURING A PANIC ATTACK

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AUTONOMIC NERvOUS CHANGES DURING A PANIC ATTACk

FIGURE 126.3 Top panel: Multifiber sym-

PANIC ATTACKS A. Sympathetic Nervous Activation

At Rest

Panic Attack

B. Cardiac Release of NPY

attack, without any increase in firing rate [4] (Fig. 126.3), presumably by recruitment of additional firing fibers. This response is qualitatively different from that seen during laboratory mental stress with stimuli such as difficult mental arithmetic, where muscle sympathetic nerve activity increases little if at all. During a panic attack, the secretion of epinephrine increases two- to six-fold [4].

Post-Panic

pathetic nerve firing in the sympathetic outflow to the skeletal muscle vasculature in a patient with panic disorder, measured by clinical microneurography. During a panic attack, there was a large increase in the amplitude of sympathetic “bursts”, without any increase in burst frequency. Bottom panel: The arterial and coronary sinus concentration of the sympathetic cotransmitter, neuropeptide Y (NPY), is shown in panic disorder patients at rest, and during a panic attack. There was no net release of NPY from the sympathetic nerves of the heart into the coronary sinus at baseline, but a panic attack evoked measurable release of the sympathetic cotransmitter. *P  0.05.

Coronary spasm during panic attack

Release of Neuropeptide Y With the pronounced activation of the cardiac sympathetic outflow occurring during a panic attack, neuropeptide Y (NPY) is co-released from the cardiac sympathetic nerves and appears in measurable quantities in coronary sinus venous blood (Fig. 126.3).

FIGURE 126.4 Coronary angiogram in a patient with panic disorder,

Mediating Autonomic Mechanisms of Cardiac Risk During a Panic Attack

performed because of recurrent angina. During a panic attack occurring during angiography, spasm occurred in the left anterior descending coronary artery (LAD). The arterial spasm was reversed by administration of glyceryl trinitrate (GTN).

Our own extensive clinical experience with the cardiological management of panic disorder sufferers has provided case material encompassing the range of cardiac complications which occur. Those patients with typical, severe anginal chest pain during panic attacks, who are in the minority, appear to be at cardiac risk. During panic attacks in such patients we have documented, variously, triggered cardiac arrhythmias, recurrent emergency room attendances with angina and ECG changes of ischemia,

coronary artery spasm during panic attacks occurring at the time of coronary angiography (Fig. 126.4) and myocardial infarction associated with coronary spasm and thrombosis. Our research findings suggest that release of epinephrine as a cotransmitter from cardiac sympathetic nerves and activation of the sympathetic nervous system during panic attacks may be mediating mechanisms.

Spasm in left anterior descending coronary artery

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Significant resolution following glyceryl trinitrate

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In this context, release of neuropeptide Y from the sympathetic nerves of the heart into the coronary sinus during the sympathetic activation accompanying panic attacks is an intriguing finding, given the capacity of NPY to cause coronary artery spasm [10]. A better understanding of the mechanism of coronary artery spasm in panic disorder would facilitate therapeutic intervention. At present we treat patients with panic disorder and clinical evidence of coronary spasm with drugs and other measures aimed at preventing or minimizing their panic attacks, a dihydropyridine calcium-channel blocker as a non-specific anti-spasm measure, and low-dose aspirin as prophylaxis against coronary thrombosis during spasm [5]. Neuropeptide Y antagonists are not yet available for clinical use.

References [1] American Psychiatric Association Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition. Washington, DC: American Psychiatric Association; 1994. [2] Kawachi I, Sparrow D, Vokanas PS, Weiss ST. Symptoms of anxiety and coronary heart disease: The normative aging study. Circulation 1994;90:2225–9. [3] Kawachi I, Colditz GA, Ascherio A, Rimm EB, Giovannucci E, Stampfer MJ, et al. Prospective study of phobic anxiety and risk of coronary heart disease in men. Circulation 1994;89:1992–7.

[4] Wilkinson DJC, Thompson JM, Lambert GW, Jennings GL, Schwarz RG, Jefferys D, et al. Sympathetic activity in patients with panic disorder at rest, under laboratory mental stress and during panic attacks. Arch Gen Psychiatry 1998;55:511–20. [5] Mansour VM, Wilkinson DJC, Jennings GL, Schwarz RG, Thompson JM, Esler MD. Panic disorder: Coronary spasm as a basis for coronary risk?. Med J Aust 1998;168:390–2. [6] Lambert E, Hotchkin E, Alvarenga M, Pier C, Richards J, Barton D, et al. Single-unit analysis of sympathetic nervous discharges in patients with panic disorder. J Physiol (Lond) 2006;570:637–43. [7] Esler M, Eikelis N, Schlaich M, Lambert G, Alvarenga M, Dawood T, et al. Chronic mental stress is a cause of essential hypertension: presence of biological markers of stress. Clin Exp Pharm Physiol 2008;35:498–502. [8] Esler M, Lambert E, Alvarenga M, Socratous F, Richards J, Barton D, et al. Increased brain serotonin turnover in panic disorder patients in the absence of a panic attack: Reduction by a selective serotonin reuptake inhibitor. Stress 2007;10:295–304. [9] Esler M, Eikelis N, Schlaich M, Lambert G, Alvarenga M, Kaye D, et al. Human sympathetic nerve biology: parallel influences of stress and epigenetics in essential hypertension and panic disorder. Ann N Y Acad Sci 2008;1148:338–48. [10] Hass M, Neuropeptide Y. A cardiac sympathetic cotransmitter? In: Goldstein DS, McCarthy R, editors. Catecholamines – Bridging Basic Science With Clinical Medicine. Academic Press; 1998. p. 129–32.

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127 Physical Measures Wouter Wieling, Roland D. Thijs PHYSICAL COUNTERMANEUVERS Specific treatment of the underlying disease in patients with autonomic failure and reflex syncope is usually not possible and consequently the goal in management is to obtain symptomatic improvement by other means. Physical maneuvers that are both easy to apply and effective in combatting orthostatic lightheadedness and eventually syncope in daily life are, therefore, of obvious importance. Patients with autonomic failure have discovered several such maneuvers themselves. Subsequent physiological investigations documented the beneficial of effects of leg-crossing, squatting, abdominal compression, bending forward, and placing one foot on a chair (Fig. 127.1). A great advantage of these maneuvers is that they can be applied immediately at the start of hypotensive symptoms. Physical countermaneuvers need to be related specifically to the individual patient. An interactive session assessing the efficacy and the practicality of the various counterpressure maneuvers is extremely valuable and will allow tailored therapy. Physical countermaneuvers may be difficult to perform in patients with multiple system atrophy, who may have motor disabilities and compromised balance. Leg-crossing is the simplest maneuver to increase the standing time in a patient with autonomic failure. It has the advantage that it can be performed without much effort and without bringing much attention to the patient's problem. The maneuver is performed by crossing one leg in direct contact with the other while actively standing on both legs (Fig. 127.1). The increase in mean arterial pressure and pulse pressure induced by leg-crossing can be attributed to compression of the muscles in the upper legs and abdomen with mechanical squeezing of venous vessels resulting in an increase in central blood volume and thereby in cardiac filling pressures and cardiac output. Tensing of leg and abdominal muscles can increase this effect considerably by a further increase in venous return. Leg-crossing can also be used for the prevention of orthostatic lightheadedness in the sitting position (Fig. 127.1). Although the increase in upright blood pressure induced by leg-crossing alone is relatively small with an average increase in mean arterial pressure of 10–15 mmHg

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00127-X

(Fig. 127.1), one should realize that medical treatment with fludrocortisone, erythropoietin, and midodrine results in similarly small blood pressure increases. Despite these small increases the standing time improves markedly by all four methods, because they shift mean arterial pressure from just below to just above the critical level of perfusion of the brain. A perfusion pressure of about 40 mmHg is needed to preserve consciousness in young adult subjects in supine posture. A mean arterial pressure of about 60 mmHg measured at heart level is needed in order to compensate for the effects of gravity on the cerebral circulation. Crossing one’s legs is often applied unintentionally also by healthy humans when standing for prolonged periods (cocktail party posture). Recent studies show that instruction to apply physical countermaneuvers is very helpful to otherwise healthy subjects with functional orthostatic disorders like the postural tachycardia syndrome. The combination of leg-crossing and tensing of leg and abdominal muscles can abort an impending vasovagal reaction and is now widely applied in patients with a tendency for vasovagal fainting (Fig. 127.2). Leg-crossing with muscle tension reduces the risk for further recurrence of vasovagal syncope by 39%. Lower body muscle tensing has also been documented to be very effective to counteract initial orthostatic hypotension.

Squatting Squatting increases arterial mean pressure and pulse pressure (Fig. 127.1) by two mechanisms. First, blood is squeezed from the veins of the legs and the splanchnic vascular bed, which increases cardiac filling pressures and cardiac output. Second, mechanical impediment of the circulation to the legs is thought to increase systemic vascular resistance. Squatting is an effective emergency mechanism to prevent loss of consciousness when presyncopal symptoms develop rapidly both in patients with autonomic failure and in patients with vasovagal episodes. Bending over as if to tie one’s shoes has similar effects and is simpler to perform by elderly patients. The beneficial effects of sitting in knee-chest position or placing one foot on a chair while standing (Fig. 127.1) are

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FIGURE 127.1 Physical countermaneuvers using isometric contractions of the lower limbs and abdominal compression. The effects of leg-crossing in standing and sitting position, placing a foot on a chair and squatting on finger arterial blood pressure (FINAP) in a 54-year-old male patient with pure autonomic failure and disabling orthostatic hypotension. The patient was standing or sitting quietly prior to the maneuvers. Bars indicate the duration of the maneuvers. Note the increase in blood pressure and pulse pressure during the maneuvers. (From Harms and Wieling, unpublished, with permission of the patient.)

comparable to squatting. Especially, when advising squatting, patients should be aware that the beneficial effects on blood pressure are only temporary: when arising again from the squatted position symptoms may recur and immediate lower body muscle tensing is advised to prevent hypotension.

EXTERNAL SUPPORT Applying external pressure to the lower half of the body substantially reduces venous pooling when upright, and consequently arterial pressure and cerebral perfusion are better maintained. External support can be applied by bandages firmly wrapped around the legs, or a snugly fitted abdominal binder, but is best accomplished by a custom-fitted counterpressure support garment, made of elastic mesh, which forms a single unit extending from the metatarsals to the costal margin. External support

garments are helpful in the treatment of a patient with incapacitating orthostatic hypotension, but have the disadvantage that the motivation of the patient must be strong, since they are uncomfortable to wear. In addition, counterpressure support garments prevent the formation of peripheral edema in the legs, which is considered to be an essential factor for effective therapy of orthostatic hypotension by acting as a perivascular water jacket that limits the vascular volume available for orthostatic pooling. We, therefore, only use an abdominal binder as a temporary external support expedient to achieve mobility in our most severely affected patients. A small lightweight portable fishing chair or a derby chair, which is a cane when folded but a seat when unfolded, are useful mechanical aids for severely affected patients. They enable the patients to sit for brief periods when presyncopal symptoms develop during standing. The lower the chair the more pronounced is the effect on blood pressure.

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611

ConClusIon

1.0 5 ∆ MAP (mmHg)

5 –5 –10 –15 0.5

∆ PetCO2(%) 0

–0.5

FIGURE 127.2 Aborting a vasovagal faint by the combination of legcrossing and muscle tensing. Typical vasovagal syncope in a 24-year-old male subject with recurrent syncope during orthostatic stress testing on a tilt-table. Note progressive fall in finger arterial pressure and heart rate. After crossing of the legs and tensing of leg and abdominal muscles, blood pressure and heart rate recover quickly. The delay in the increase in blood pressure is explained by transit time from the right to the left ventricle (about five beats). Bar indicates onset of leg-crossing and muscle tensing. (From Krediet and Wieling, unpublished.)

RESPIRATORY MANEUVERS The “respiratory pump” affects blood pressure in several ways, e.g., may augment venous return and thus blood pressure, if the intrathoracic pressure becomes more negative during inspiration. Building on this concept, selective increase of inspiratory impedance by means of an impedance threshold device (ITD) has been found to increase both supine and standing blood pressure in healthy controls. This hand-held device increases the inspiratory negative pressure gradient by 7 cmH2O. In patients with autonomic failure such a device augments standing blood pressure by ~8 mmHg. The thought that this additional circulatory pump is under volitional control prompted the study of other deviceless maneuvers including inspiratory sniffing and inspiration through pursed lips. With these interventions similar increases of blood pressure were recorded. However, in contrast to the impedance threshold device these maneuvers could also aggravate the low blood pressure in some patients (Fig. 127.3). This negative response was explained by concomitant hypocapnia due to hyperventilation. Therefore, the use of a device increasing inspiratory impedance is the most reliable respiratory maneuver. Inspiratory sniffing and inspiration through pursed lips can also reduce orthostatic hypotension with the important caveat that hyperventilation must be avoided.

–1

PLB

IS

MT

IO

FIGURE 127.3 Change of mean arterial pressure (ΔMAP) and endtidal CO2 tension (ΔCO2) observed with four maneuvers: breathing through pursed lips during inspiration (PLB); inspiratory sniffing (IS); muscle tensing of the legs (MT); and inspiratory obstruction through narrowing of an inspiratory valve (IO) in 10 patients with autonomic failure. Data are shown as differences from normal standing (mean SE). The high variability in the blood pressure response to each maneuver (as indicated by the bars) is best explained by a variable tendency to hyperventilate. Maneuvers tending to cause a decrease of end-tidal CO2 tension (IS or PLB) were likely to result in a lower MAP, whereas maneuvers with little change of end-tidal CO2 tension (MT or IO) where likely to increase MAP. (Thijs et al., Neurology 2007.)

CONCLUSION Mechanical maneuvers such as leg-crossing and squatting are simple to perform, and can increase the standing time of patients with orthostatic hypotension decisively. Their beneficial effect is an increase in mean arterial pressure, small in magnitude but sufficient to guarantee adequate cerebral blood flow. The underlying mechanism is an augmentation of thoracic blood volume. Instruction in these maneuvers should be part of a treatment program for patients with orthostatic hypotension due to autonomic failure and otherwise healthy subjects with a tendency for orthostatic reflex syncope. It is our experience that after proper instruction and training patients automatically apply leg-crossing in daily life.

Further Reading Krediet CTP, Van Dijk N, Linzer M, Van Lieshout JJ, Wieling W. Management of vasovagal syncope: controlling or aborting faints by the combination of leg-crossing and muscle tensing. Circulation 2002;106:1684–1689.

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Krediet CT, Go-Schön IK, Kim YS, Linzer M, Van Lieshout JJ, Wieling W. Management of initial orthostatic hypotension: lower body muscle tensing attenuates the transient arterial blood pressure decrease upon standing from squatting. Clin Sci (Lond) 2007;113(10):401–7. Krediet CT, de Bruin IG, Ganzeboom KS, Linzer M, van Lieshout JJ, Wieling W. Leg-crossing, muscle tensing, squatting, and the crash position are effective against vasovagal reactions solely through increases in cardiac output. J Appl Physiol 2005;99(5):1697–703. Melby DP, Lu F, Sakaguchi S, et al. Increased impedance to inspiration ameliorates hemodynamic changes associated with movement to upright posture in orthostatic hypotension: a randomized blinded pilot study. Heart Rhythm 2007;4:128–35. Smit AAJ, Halliwill JR, Low PA, Wieling W. Topical Review. Pathophysiological basis of orthostatic hypotension in autonomic failure. J Physiol 1999;519:1–10. Smit AA, Wieling W, Fujimura J, Denq JC, Opfer-Gehrking TL, Akarriou M, et al. Use of lower abdominal compression to combat orthostatic

hypotension in patients with autonomic dysfunction. Clin Auton Res 2004;14:167–75. Ten HarkeI ADJ, Van Lieshout JJ, Wieling W. Effects of leg muscle pumping and tensing on orthostatic arterial pressure; a study in normal subjects and in patients with autonomic failure. Clin Sci 1994;87:533–58. Thijs RD, Wieling W, van den Aardweg JG, van Dijk JG. Respiratory countermaneuvers in autonomic failure. Neurology 2007;69(6):582–5. van Dijk N, Quartieri F, Blanc JJ, Garcia-Civera R, Brignole M, Moya A, et al. PC-Trial Investigators. Effectiveness of physical counterpressure maneuvers in preventing vasovagal syncope: the Physical Counterpressure Manoeuvres Trial (PC-Trial). J Am Coll Cardiol. 2006;48(8):1652–7. Epub 2006 Sep 26 van Dijk N, de Bruin IG, Gisolf J, de Bruin-Bon HA, Linzer M, van Lieshout JJ, et al. Hemodynamic effects of leg-crossing and skeletal muscle tensing during free standing in patients with vasovagal syncope. J Appl Physiol 2005;98(2):584–90.

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128 Water and the Osmopressor Response Jens Jordan

In mammals the relationship between free water and solutes in extra- and intracellular compartments, which determines osmolality, is tightly regulated through adjustment in water ingestion and excretion. Even mild abnormalities in systemic osmoregulation can have grave consequences. Reflex mechanisms governed by osmoreceptive neurons located in brainstem nuclei with no blood–brain barrier maintain extracellular fluid osmolality within a tight range. In addition, there is evidence for hitherto poorly characterized peripheral osmosensitive neurons. Recent findings suggest that local changes in osmolality through water drinking may elicit a powerful pressor response and changes in energy metabolism through sympathetic nervous system activation. This osmopressor response may involve activation of transient receptor potential vanniloid 4 (TRPV4) receptors.

THE WATER-INDUCED PRESSOR RESPONSE The fact that water drinking may elicit acute cardiovascular responses was recognized in patients with severe orthostatic hypotension due to autonomic failure. Some autonomic failure patients reported rapid symptomatic improvement with water drinking. Subsequently, drinking 480 ml tap water was shown to elicit a profound pressor response in these patients. In patients with central autonomic failure due to multiple system atrophy, seated systolic blood pressure increased 33 mmHg. In patients with peripheral autonomic degeneration due to pure autonomic failure, seated systolic blood pressure increased 37 mmHg. The pressor response had an onset within 5 minutes after water drinking, reached a maximum in 30–40 minutes, and was maintained for more than one hour (Fig. 128.1). In elderly healthy subjects drinking 480 ml tap water, systolic blood pressure increased up to 11 mmHg. Water drinking in healthy young subjects did not elicit a pressor response. Increased systemic vascular resistance rather than cardiac output mediated the pressor response in autonomic failure patients. Similarly, healthy young subjects showed an increase in calf vascular

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00128-1

resistance while systemic vascular resistance and blood pressure remained unchanged. Together, these observations suggested that water drinking elicits changes in vascular tone that are unmasked in individuals with aging-associated changes in baroreflex regulation and more so in patients with profound baroreflex abnormalities due to neurodegenerative diseases.

EVIDENCE FOR WATER-INDUCED SYMPATHETIC ACTIVATION Even in severely affected autonomic failure patients, loss of efferent sympathetic function is rarely complete. Autonomic failure patients with a complete loss of sympathetic efferent function suggested by absence of a pressor response to yohimbine showed no changes in blood pressure after water drinking. Patients with residual sympathetic function indicated by a large response to yohimbine also showed a large water-induced pressor response. Furthermore ganglionic blockade with trimethaphan abolished the water-induced pressor response. Moreover, water drinking increased muscle sympathetic activity in healthy subjects in the absence of a pressor response. Finally, venous plasma norepinephrine concentrations

50

∆ mmHg or bpm

INTRODUCTION

40

n=19

30

SBP

20

DBP

10 0

HR

–10 –10

0

10

20

30

40

50

60

Time (min)

FIGURE 128.1 Changes in systolic blood pressure (SBP), diastolic blood pressure (DBP) and heart rate (HR) in patients with pure autonomic failure after ingestion of 480 ml tap water. Patients started drinking at 0 minutes. The blood pressure increase was evident within 5 minutes of drinking water, reached a maximum after approximately 20–30 minutes, and was sustained for more than 60 minutes. (From Jordan et al.)

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increased with water drinking in younger and in older healthy subjects as well as in autonomic failure patients. Because sympathetic nervous system activity has a central role in the regulation of energy expenditure, water drinking increases resting metabolic rate approximately 30%. With water drinking, a similar increase in endogenous norepinephrine produces a much greater blood pressure increase in autonomic failure patients compared with healthy subjects. Autonomic failure patients are also extremely hypersensitive to exogenous alpha adrenergic agonists. Loss of baroreflex blood pressure buffering and increased vascular sensitivity may contribute to the pressor hypersensitivity.

A SPINAL SYMPATHETIC REFLEX? The underlying pathology in water responsive patients helped localize the neural substrate for water-induced sympathetic activation. Both in multiple system atrophy and in high spinal cord injured patients, water drinking raises blood pressure. In multiple system atrophy patients, the lesion to the efferent part of the autonomic nervous system may lie in the brainstem. More distal efferent sympathetic structures are at least partially intact. In high spinal cord injured patients, spinal sympathetic neurons are intact but disconnected from brainstem input. Postganglionic sympathetic neurons can be activated by spinal reflexes but not by reflexes traveling through the brainstem including baroreflexes. Thus, water drinking engages a spinal reflex-like mechanism activating sympathetic efferent nerves. Indeed, bilateral subdiaphragmatic vagotomy did not abolish the pressor response to water drinking in sinoaortic denervated mice. Intragastric and intraduodenal water infusion induced an identical pressor response in sinoaortic denervated mice suggesting that the afferent structure responding to water is likely distally located from the stomach.

EVIDENCE FOR AN OSMOSENSITIVE MECHANISM INVOLVING Trpv4 Recent studies uncovered potential stimuli activating the spinal sympathetic response to water drinking. In autonomic failure patients, the magnitude of the

water-induced pressor response was not related to water temperature. Only one third of the increase in resting energy expenditure could be explained by warming the water from 22°C to 37°C. Even water drinking of 37°C increased metabolic rate. Gastric distention increases sympathetic activity in humans. Yet, the maximal response to water drinking was observed after approximately 40 minutes. At this time, only 25% of the ingested water remains in the stomach. Temperature or gastric distention is not sufficient to set off the water response, leaving local/ regional hyposmolarity as a likely trigger. Water elicited a much greater pressor response compared with isotonic saline in sinoaortic denervated mice. Moreover, in patients with autonomic failure due to multiple system atrophy, water given through a nasogastric tube increased blood pressure more than the same volume of normal saline. Similarly, addition of sodium chloride to drinking water attenuated the pressor response in autonomic failure patients. All these findings support the idea that hyposmolarity is the stimulus for water-induced sympathetic activation. Osmosensitive afferent neurons located in the portal tract might be involved. Indeed, in mice, osmolality changes in the portal vein coincide with changes in blood pressure. The molecular transduction mechanisms involved in sensing hypoosmotic signals in peripheral tissues are poorly understood. The transient receptor potential (Trp) channel family including the vanilloid subfamily (Trpv) is involved in recognition of noxious environmental stimuli including osmolality, temperature, and pain. Trpv4 is a prime suspect given its sensitivity to osmotic changes. Indeed, although portal osmolality decreases after water application in wild-type and Trpv4–/– mice, when all had undergone sinoaortic denervation, only wild-type animals showed a pressor response (Fig. 128.2). Thus, the presence of Trpv4 is required to express the osmopressor response. Trpv4 channels on hepatic and/or portal spinal afferent neurons could be involved.

THERAPEUTIC UTILITY OF WATER DRINKING Water drinking improves standing blood pressure and orthostatic tolerance in a large subgroup of patients with

FIGURE 128.2 Blood pressure changes after duodenal infusion of water (25 µl/g body weight) in anesthetized sinoaortic denervated Trpv4 knockout and wild-type mice. Absence of a pressor response in Trpv4 knockout animals suggests that the receptor is an essential mediator in the osmopressor response. (From McHugh et al. 2010)

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THERAPEuTIC uTIlITy Of WATER dRInkIng

autonomic failure. The maximal pressor effect to water is reached at a time when other pressor agents just begin to act. Furthermore, water drinking attenuates postprandial hypotension. Water should be ingested before meals and when orthostatic symptoms are worst. Water ingestion is particularly useful in the morning before arising with or without addition of pressor drugs. Patients with supine hypertension should avoid water drinking within one hour before bedtime. The effects of pressor agents, such as pseudoephedrine and phenylpropanolamine, are potentiated by water drinking. This “drug interaction” between water and pressor agents can be exploited in the treatment of orthostatic hypotension. However, the interaction can also lead to potentially dangerous blood pressure surges. Excessive water ingestion should be avoided, particularly in patients with multiple system atrophy because potentially life threatening hyponatremia could ensue. Water drinking could have a therapeutic benefit in patients with postural tachycardia syndrome (POTS, idiopathic orthostatic intolerance). A syndrome that is more common than autonomic failure. Drinking 480 ml water lowered upright heart rate 15 and 10 beats per minute after 3 and 5 minutes standing, respectively. Influences of water drinking on orthostatic tolerance in healthy subjects and in patients with neurally mediated (vasovagal) syncope have been studied using head-up tilt testing combined with lower body negative pressure or with regular head-up tilt testing. In young healthy subjects water drinking can delay or even prevent syncope during head-up tilt testing with or without lower body negative pressure. Patients with neurally mediated syncope showed similar improvements in orthostatic tolerance with ingestion of 500 ml water. Water drinking also decreases the risk for blood donation related vasovagal reactions and may be beneficial in individuals with post exercise syncope.

Further Reading Ando S, Kawamura N, Matsumoto M, Dan E, Takeshita A, Murakami K, et al. Simple standing test predicts and water ingestion prevents vasovagal reaction in the high-risk blood donors. Transfusion 2009;49(8):1630–6. Boschmann M, Steiniger J, Hille U, Tank J, Adams F, Sharma AM, et al. Water-induced thermogenesis. J. Clin. Endocrinol. Metab. 2003;88(12):6015–9. Cariga P, Mathias CJ. Haemodynamics of the pressor effect of oral water in human sympathetic denervation due to autonomic failure. Clin. Sci. 2001;101(3):313–9.

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Claydon VE, Schroeder C, Norcliffe LJ, Jordan J, Hainsworth R. Water drinking improves orthostatic tolerance in patients with posturally related syncope. Clin. Sci. (Lond) 2006;110(3):343–52. France CR, Ditto B, Wissel ME, France JL, Dickert T, Rader A, et al. Predonation hydration and applied muscle tension combine to reduce presyncopal reactions to blood donation. Transfusion 2010;50(6):1257–64. Jordan J, Shannon JR, Black BK, Ali Y, Farley M, Costa F, et al. The pressor response to water drinking in humans: a sympathetic reflex? Circulation 2000;101(5):504–9. Jordan J, Shannon JR, Diedrich A, Black B, Robertson D, Biaggioni I. Water potentiates the pressor effect of ephedra alkaloids. Circulation 2004;109(15):1823–5. Jordan J, Shannon JR, Grogan E, Biaggioni I, Robertson D. A potent pressor response elicited by drinking water. Lancet 1999;353(9154):723. Lipp A, Tank J, Franke G, Arnold G, Luft FC, Jordan J. Osmosensitive mechanisms contribute to the water drinking-induced pressor response in humans. Neurology 2005;65(6):905–7. Lu CC, Diedrich A, Tung CS, Paranjape SY, Harris PA, Byrne DW, et al. Water ingestion as prophylaxis against syncope. Circulation 2003;108(21):2660–5. McHugh J, Keller NR, Appalsamy M, Thomas SA, Raj SR, Diedrich A, et al. Portal osmopressor mechanism linked to transient receptor potential vanilloid 4 and blood pressure control. Hypertension 2010;55(6):1438–43. Newman B, Tommolino E, Andreozzi C, Joychan S, Pocedic J, Heringhausen J. The effect of a 473-mL (16-oz) water drink on vasovagal donor reaction rates in high-school students. Transfusion 2007;47(8):1524–33. Raj SR, Biaggioni I, Black BK, Rali A, Jordan J, Taneja I, et al. Sodium paradoxically reduces the gastropressor response in patients with orthostatic hypotension. Hypertension 2006;48(2):329–34. Routledge HC, Chowdhary S, Coote JH, Townend JN. Cardiac vagal response to water ingestion in normal human subjects. Clin. Sci. (Lond) 2002;103(2):157–62. Schroeder C, Bush VE, Norcliffe LJ, Luft FC, Tank J, Jordan J, et al. Water drinking acutely improves orthostatic tolerance in healthy subjects. Circulation 2002;106(22):2806–11. Scott EM, Greenwood JP, Gilbey SG, Stoker JB, Mary DASG. Water ingestion increases sympathetic vasoconstrictor discharge in normal human subjects. Clin. Sci. 2001;100(3):335–42. Shannon JR, Diedrich A, Biaggioni I, Tank J, Robertson RM, Robertson D, et al. Water drinking as a treatment for orthostatic syndromes. Am. J. Med. 2002;112(5):355–60. Stookey JD, Constant F, Popkin BM, Gardner CD. Drinking water is associated with weight loss in overweight dieting women independent of diet and activity. Obesity 2008;16(11):2481–8. Tank J, Schroeder C, Stoffels M, Diedrich A, Sharma AM, Luft FC, et al. Pressor effect of water drinking in tetraplegic patients may be a spinal reflex. Hypertension 2003;41(6):1234–9. Thijs RD, Reijntjes RHAM, Van Dijk JG. Water drinking as a potential treatment for idiopathic exercise-related syncope: A case report. Clin. Auton. Res. 2003;13(2):103–5.

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129 Droxidopa (L-DOPS) Horacio Kaufmann INTRODUCTION L-threo-dihydroxyphenylserine (L-DOPS) is a synthetic amino acid that, after oral administration, is converted to norepinephrine, via a single decarboxilation step catalyzed by the enzyme L aromatic aminoacid decarboxylase (LAAADC) (Fig. 129.1). As a precursor of norepinephrine, L-DOPS (droxidopa) has been approved in Japan since 1989 for the treatment of orthostatic hypotension in patients with chronic autonomic disorders. Large, multinational, phase III clinical trials are currently underway to obtain approval for the same indication in the US and Europe. L-DOPS crosses the blood–brain barrier and may have effects on mood, behavior and motor activity, but its ability to increase blood pressure in patients with chronic autonomic disorders is due to its conversion to norepinephrine outside the brain, both in neuronal and non neuronal tissues. Complete inhibition of LAAAD outside the CNS by carbidopa prevents the peripheral conversion of L-DOPS to norepinephrine and blunts its pressor effect. Incomplete inhibition of LAAAD, as in patients with PD receiving standard dosages of levodopa/carbidopa or benzerazide combinations, does not prevent the pressor effect of L-DOPS.

HISTORY DOPS is an artificial amino acid first synthesized in 1919. In the 1950s, it was shown that DOPS was converted to norepinephrine when incubated in extracts of guinea pig kidney and liver containing LAAAD [1]. Convincing evidence that this decarboxylation also occurred in vivo came from the finding that the urine from rabbits, which have been given DOPS, contained norepinephrine. In 1980, Araki et al. reported that given orally to rats L-DOPS had a slow-onset and long-acting pressor effect, suggesting a potential use as an oral pressor agent for the treatment of autonomic disorders with orthostatic hypotension [2]. The increase in blood pressure was markedly reduced by inhibition of peripheral decarboxylase, which also blunted the elevation of plasma norepinephrine, and by blockade of alpha-adrenoceptors. Its pressor effect was enhanced in rats made hypotensive by chemical

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00129-3

sympathectomy with 6-hydroxydopamine. L-DOPS produced the same increase in plasma NA concentrations in sympathectomized rats as in the controls.

PHARMACOLOGY Isomeric Structure 3,4-threo-dihydroxyphenylserine exists as four stereoisomers. Due to the stereospecificity of LAAAD, only L-threo-DOPS and L-erythro-DOPS are decarboxylated to form norepinephrine and only L-threo-DOPS is converted to biologically active L-norepinephrine. Of note, the D-stereoisomer of DOPS competitively inhibits the decarboxylation of the L-stereoisomer and thereby the formation of biologically active L-norepinephrine. Thus, the pure L isoform is the preferred formulation to use rather than the racemic mixture containing both the D- and L-isoforms.

Pharmacokinetics Because L-DOPS is structurally identical to L-DOPA with an additional beta hydroxyl group on the side chain (Fig. 129.1), levels in plasma are similar in magnitude and timing to those reported after administration of L-DOPA. L-DOPS, a neutral aminoacid, is well absorbed after oral administration. Peak plasma levels occur after around 3 hours and thereafter, the plasma concentration of L-DOPS decline monoexponentially, with a half time of 2 to 3 hours. Plasma levels of norepinephrine also peak at approximately 3 hours; almost concurrent with the L-DOPS peak, but at much lower concentration. Rather than low efficiency of conversion of L-DOPS to norepinephrine, the difference in plasma concentration is readily explained by the shorter half time of norepinephrine in plasma, which is 1.5 minutes, compared to the half time of L-DOPS in plasma, which is about 140 minutes. After they peak, norepinephrine levels decline multiexponentially, with an initial half time of around 9 hours [3]. All cells that express the neutral aminoacid transporter, including sympathetic postganglionic nerves and parenchymal cells in the liver and kidney, take up L-DOPS. These cells also express LAAAD in their cytoplasm so

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H H C C

HO

COOH

Tyrosine

H NH2 Tyrosine hydroxylase

H H C C COOH

HO HO

Levodopa

H NH2

Aromatic-L-amino acid decarboxylase H H C C

HO HO

H

Dopamine

H NH2 Dopamine-beta-hydroxylase

OH H C C

HO HO

OH H COOH

HO

H NH2

C C HO

H

Norepinephrine

H NH2 PhenylethanolaminoN-methyltransferase OH H C C

HO HO

H

Epinephrine

H NHCH2

FIGURE 129.1 Catecholamine pathway. Norepinephrine is synthesized from levodopa (L-DOPA) in two steps. The first step is the decarboxylation of levodopa to dopamine by L-aromatic amino acid decarboxylase. The second step is the hydroxylation of dopamine to norepinephrine by the ratelimiting enzyme dopamine beta hydroxylase. Droxidopa bypasses this rate-limiting enzyme and is converted to norepinephrine in a single step by L-aromatic amino acid decarboxylase.

that L-DOPS can be converted to norepinephrine, build up in the cytoplasm and be released via reverse transport through the Uptake 2 carrier or, in sympathetic nerves, through exocycitosis during nerve activation. The slow decline in plasma norepinephrine from the peak level probably reflects ongoing production of norepinephrine from L-DOPS within a cellular storage site and ongoing entry of the produced norepinephrine into the bloodstream. Analyzing the different pharmacokinetics of L-DOPS in patients with PAF and MSA is instructive. Patients with PAF have a profound loss of sympathetic nerves, whereas patients with MSA have intact sympathetic nerve

terminals. Norepinephrine in plasma remained higher than baseline for a longer time in MSA suggesting ongoing conversion to norepinephrine in patients with MSA. However, the magnitude of the increase in plasma norepinephrine and the slope of the relationship between L-DOPS and norepinephrine concentrations were similar in the PAF and MSA groups [3]. These similarities in the norepinephrine responses in PAF and MSA despite the differences in their sympathetic nerves, suggest that the production of norepinephrine from L-DOPS occurs mainly in non-neuronal cells. Persistent elevation of plasma NE in MSA suggests residual release of NE from sympathetic nerves in those with MSA.

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MECHANISM OF ACTION Pressor Effect The threshold norepinephrine venous plasma level required to increase blood pressure in patients with neurogenic orthostatic hypotension was 700 pg/ml [4] (Fig. 129.2). This is similar to the plasma concentration necessary to increase blood pressure when norepinephrine is infused. Carbidopa, a competitive inhibitor of LAAAD that does not cross the blood–brain barrier, was used as a pharmacological probe to determine whether the pressor action of L-DOPS was due to its conversion to norepinephrine inside or outside the central nervous system. The administration of 200 mg of carbidopa with L-DOPS blocked the increase norepinephrine in plasma and the pressor effect, indicating that the hypertensive effect of L-DOPS occurs after its conversion to norepinephrine in the periphery rather than in the brain [4]. Lower dosages of LAAAD inhibitors, such as those commonly used to treat Parkinson’s disease, do not abolish the pressor effect of L-DOPS [5]. Newly synthesized norepinephrine could act in two possible ways: 1. As a peripheral sympathetic neurotransmitter. L-DOPS is taken up by postganglionic sympathetic neurons, converted to norepinephrine in the cytoplasm, stored in vesicles, and released via exocytosis when sympathetic neurons are activated. This mechanism was shown in patients with dopamine beta hydroxylase (DBH) deficiency [6]. Therefore, the DOPS derived norepinephrine may act as a physiologic neurotransmitter in peripheral sympathetic neurons. 2. As a circulating hormone. As the neutral aminoacid transporter and the enzyme LAAAD are widely expressed in non neuronal cells throughout the body

50

Change in SBP (mmHg)

40 30 20 10 0 –10

200

400

600

800

1000

1200

1400

Norepinephrine (pg/mI)

FIGURE 129.2 The relationship between blood pressure and norepinephrine levels after droxidopa administration. Changes in systolic blood pressure (SBP) versus plasma concentration of norepinephrine after droxidopa administration. Solid curve represents the line of best fit. All data are mean  SEM (n  8).

(including the stomach, kidney and liver), L-DOPS could be taken up and converted to norepinephrine in different cell types. Newly formed norepinephrine would then be released into the blood stream to exert a pressor effect as a circulating hormone [7].

Other Actions L-DOPS crosses the blood–brain barrier and is converted to norepinephrine in the CNS. The possibility of increasing norepinephrine levels centrally has therapeutic potential for patients with a variety of disorders, including Parkinson’s disease in which there is degeneration of the norepinephrine producing neurons in the locus ceruleus.

CLINICAL STUDIES DBH Deficiency L-DOPS was first used successfully to treat patients with autonomic failure as result of a congenital deficiency of the enzyme DBH [6]. Because of the inability to convert dopamine to NE, these patients have severe orthostatic hypotension with high serum dopamine levels and undetectable levels of norepinephrine in plasma. Oral administration of L-DOPS had a dramatic effect completely abolishing the orthostatic fall in blood pressure [6]. Because L-DOPS is directly decarboxylated to NE it bypassed the missing enzyme (Fig. 129.1). After treatment with DOPS, norepinephrine increased upon standing, and infusion of tyramine produced release of NE, whereas before treatment, tyramine had induced release of dopamine [6].

Other Types of Autonomic Failure L-DOPS has now been used successfully to treat orthostatic hypotension in several autonomic disorders, including familial amyloid polyneuropathy, autoimmune autonomic ganglionopathy, PAF, Parkinson disease and multiple system atrophy [4,8,9]. Because patients with autonomic failure have different degrees of adrenergic denervation supersensitivity, individualized dosing for each patient is useful. The optimal dose of L-DOPS varied 10-fold, between 200 mg and 2000 mg [4]. Administration of L-DOPS significantly increased both supine and standing blood pressure in patients with chronic autonomic failure [4]. The pressor effect began 1 h after L-DOPS administration and lasted for 6 h while standing and for 8 h while supine [3]. The highest standing blood pressure occurred 3.5 h after L-DOPS administration [3]. Patients were less lightheaded, reported feeling better and were able to stand for longer. There were no significant changes in heart rate following administration of L-DOPS. Like all pressor drugs, supine hypertension is a side effect of treatment with L-DOPS, with the frequency of supine hypertension being similar in patients with MSA and PAF [4].

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Hyponatremia and transitory chest pain with electrocardiographic ST depression, without elevation of cardiac enzymes, were also reported. Large, multinational phase III clinical trials of L-DOPS in patients with autonomic failure are currently underway. Initial results were positive.

References [1] Blaschko H, Burn JH, et al. The formation of noradrenaline from dihydroxyphenylserine. Br J Pharmacol 1950;5(3):431–7. [2] Araki H, Tanaka C, et al. Pressor effect of L-threo-3,4-dihydroxyphenylserine in rats. J Pharm Pharmacol 1981;33(12):772–7. [3] Goldstein DS, Holmes C, et al. Clinical pharmacokinetics of the norepinephrine precursor L-threo-DOPS in primary chronic autonomic failure. Clin Auton Res 2004;14(6):363–8.

[4] Kaufmann H, Saadia D, et al. Norepinephrine precursor therapy in neurogenic orthostatic hypotension. Circulation 2003;108(6):724–8. [5] Mathias CJ. L-dihydroxyphenylserine (Droxidopa) in the treatment of orthostatic hypotension: the European experience. Clin Auton Res 2008;18(Suppl. 1):25–9. [6] Biaggioni I, Robertson D. Endogenous restoration of noradrenaline by precursor therapy in dopamine-beta-hydroxylase deficiency. Lancet 1987;2(8569):1170–2. [7] Kaufmann H. Could treatment with DOPS do for autonomic failure what DOPA did for Parkinson's disease?. Neurology 1996;47(6):1370–1. [8] Freeman R, Landsberg L, et al. The treatment of neurogenic orthostatic hypotension with 3,4-DL-threo-dihydroxyphenylserine: a randomized, placebo-controlled, crossover trial. Neurology 1999;53(9):2151–7. [9] Kaufmann H. L-dihydroxyphenylserine (Droxidopa): a new therapy for neurogenic orthostatic hypotension: the US experience. Clin Auton Res 2008;18(Suppl. 1):19–24.

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130 Midodrine, Adrenergic Agonists and Antagonists Janice L. Gilden The sympathomimetics used for treatment of neurogenic orthostatic hypotension (NOH) have either direct alpha-1 (e.g., midodrine) or mixed direct and indirect actions (e.g., ephedrine). Midodrine, a direct alpha-1agonist, produces vasoconstriction of the arteriolar and venous vasculature and decreases venous pooling. Since it does not cross the blood–brain barrier, there are no central nervous side effects. Midodrine does not cause cardiac stimulation. Midodrine, is a prodrug which is hydrolyzed in the liver to its active form, desglymidodrine. The peak activity is 1 hour after administration and its duration of action is 4–6 hours. Studies have demonstrated efficacy for improving both standing blood pressure and symptoms. Side effects are generally mild, dose-related, and include piloerection, urinary retention, and supine hypertension. Other sympathomimetics, such as ephedrine, pseudoephedrine, phenylpropanolamine, and methylphenidate have both direct and indirect actions and cross the blood– brain barrier. These pharmacologic agents have a short duration of action and are associated with marked central nervous system and cardiac toxic effects. Comparative studies with these mixed sympathomimetics show less efficacy in terms of blood pressure and symptomatic improvement. Adrenergic antagonists, such as clonidine and yohimbine may be used for treatment of orthostatic hypotension in selective cases.

Pharmacology Midodrine is well-absorbed after oral administration. In the liver, midodrine hydrochloride is converted via hydrolytic cleavage to its active metabolite, desglymidodrine, which is 93% bioavailable. Desglymidodrine is excreted primarily in the urine, whereas fecal elimination of the prodrug and its metabolite is not significant. The half life of desglymidodrine in normal subjects is 2–3 hours and of midodrine is 0.49 hours. The duration of action is longer than the other sympathomimetic agonists. The dose response curve for effect on the standing systolic blood pressure shows a log linear relationship [2]. In normal individuals without autonomic dysfunction, the maximal pharmacologic effect is found to be 1 hour after administration and duration of action is generally 4–6 hours. In patients with neurogenic orthostatic hypotension (NOH), these effects may be somewhat variable. This may reflect the differences in receptor and postreceptor impairments of the autonomic nervous system.

Efficacy

MIDODRINE Mechanism of Action Midodrine [(1-2,5-dimethoxyphenyl 1)-2 glycinamido-ethanol (1)-hydrochloride] (trade names: Amatine, Proamatine, Gutron) is a selective alpha-1 agonist and results in alpha adrenergic receptor activation of the arteriolar and venous vasculature. These actions produce vasoconstriction of the blood vessels and a decrease in venous pooling, thereby increasing blood pressure in the upright position. In addition, midodrine does not stimulate B-adrenergic receptors. This agent lacks central nervous system side effects, since it does not cross the blood–brain barrier. In contrast to the nonselective sympathomimetic

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00130-X

agents, midodrine also does not cause cardiac stimulation [1]. This drug has no known effects on the pulmonary, renal, or blood coagulation function, or changes in blood glucose and lipids [1].

Midodrine is effective for treating orthostatic hypotension in patients with either preganglionic (multiple system atrophy and Parkinson’s disease) or postganglionic (pure autonomic failure and diabetes mellitus) lesions. Earlier studies by Kaufmann et al. [3] evaluated seven patients after treatment with midodrine and found an increase in mean arterial pressure (MAP) of 15 mmHg compared to baseline levels in the three responders (2 MSA and 1 PAF) with decreased orthostatic symptoms. Other studies demonstrated maintenance of blood pressure improvements over 15 months [1]. In a later study of 97 patients with varying etiologies of NOH, the 10 mg three times daily dose significantly increased standing systolic blood pressure by 22 mmHg without affecting

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heart rate. There was significant improvement of symptoms (dizziness, lightheadedness, syncope, and depressed feelings), even at doses lower than that required to show significant improvement in systolic blood pressure (SBP). Global evaluation scores assessed by patients and investigators, also improved [4,5]. A second study by Gilden et al. [6] evaluated 53 patients with severe NOH treated in a double-blind fashion with three daily doses of 10 mg midodrine vs. placebo. There was not only an increase in standing SBP compared to placebo, but also improvement of symptoms, and a 50% increase in motionless standing time. In a 6-week double-blind study of 162 patients with severe (NOH) (10 mg three times daily), SBP significantly improved by 22 mmHg in all 4 weeks of treatment with symptomatic improvement, rated by patients, as well as investigators [7]. Another multicenter study of midodrine hydrochloride, using open titration and a placebo phase, has also demonstrated significant improvements in symptoms, quality of life, as measured by standard scales developed for orthostatic hypotension symptoms, and global improvement scores, rated independently by both subjects and investigators [8]. Other studies have demonstrated effectiveness for treating patients with familial dysautonomia [9], neurocardiogenic syncope, intradialytic orthostatic hypotension [10] and post-flight autonomic dysfunction with orthostatic hypotension (due to its ability to cause arterial and venous constriction leading to a decrease in heart rate by baroreceptor reflexes) [11]. There have been few well-controlled studies with midodrine treatment for orthostatic hypotension in spinal cord injury patients (SCI). Small studies have suggested improvements of blood pressure, as well as increases in exercise capacity and sexual function. In 2010, Wecht et al. reported that a 10 mg dose of midodrine resulted in significant increases in MAP and attenuation of decreases in middle cerebral artery blood flow during head up-tilt in two patients with minimal change in the remaining eight [12]. Their study concluded a beneficial effect for select SCI patients. However, further larger scale studies are needed. Other uses of midodrine therapy have included: chronic fatigue syndrome; amelioration of the hypotensive effects of various psychotropic drugs and infections (pneumonia, enteritis, and meningitis). There are other reports for benefits in female stress incontinence, and hepatorenal syndrome (due to its vasoconstrictor properties with reduction in refractory ascites from the reduction in plasma renin and aldosterone, and naturesis). In comparative studies with etilefrine, dimetofrine, and ephedrine, midodrine treatment resulted in a greater improvement in standing blood pressure. When compared to norefenefrine, midodrine therapy resulted in a greater improvement in symptoms [1]. Fouad-Tarazi et al. [13] compared midodrine to ephedrine and confirmed greater improvement in upright blood pressure and symptoms with midodrine. When midodrine was combined with the

B-adrenoreceptor agonist, denodopamine, postprandial hypotension was prevented by increasing cardiac output and peripheral vascular resistance [1]. In a prospective study of 30 patients with neurocardiogenic syncope, addition of midodrine to metoprolol therapy, resulted in 77% improvement in symptomatology and tilt-table testing, when compared to beta-blocker alone [14]. A subanalysis of the study by Low et al. [7] showed that the effect of midodrine was not altered by fludrocortisone or support garment use.

Other Beneficial Effects When midodrine hydrochlochoride was administered to anesthetized dogs with experimentally-induced postural hypotension, there were significant attenuations of decreases in blood pressure, as well as increases in cerebral tissue blood flow, vertebral arterial blood flow, cardiac output, and femoral arterial blood flow [15]. In 15 healthy male volunteers, a single dose of midodrine, resulted in decreases of norepinephrine levels and heart rate, with increases in atrial naturetic peptide, independent of blood pressures [16]. In a study of patients with NOH, midodrine treatment resulted in improvements of venous capacity during orthostasis, increased SBP, broadened amplitude, and decreased pulse rate upright and under ergometric stress, as well as improvements in well-being [17]. Other studies have confirmed significant hemodynamic improvement and less syncope during tilt table testing, more symptom free days, subjective therapeutic responses in all domains of quality of life, physical functioning, energy, and vitality, improvement in overall health status, and a lower thoracic fluid index (indicating increased venous return when supine and during head up tilt) vs. placebo [18]. In diabetic patients, improvements in sympathetic withdrawal have been reported after midodrine treatment [19]. Various studies by Gilden et al. have confirmed that midodrine therapy improves cerebral blood flow, cognitive function, as well as depression in patients with various etiologies of NOH [20,21].

Adverse Effects and Disadvantages The most common side effects are related to piloerection (paraesthesia, pruritis of the scalp, goosepumps, and chills), head pressure, flushing, nervousness, anxiety, and urologic (urgency, hesitancy, frequency, and retention). Therefore, this drug needs to be used with caution in patients with urinary retention, as well as in diabetics also taking fludrocortisone, since the latter is known to increase intraocular pressure and glaucoma. There is also the risk of supine hypertension, especially in higher doses [2,4,7]. Percentages for supine hypertension have generally ranged from 4–7% [4,7]. However, one study observed an 11% rate for blood pressures greater than 200 mmHg in response to a single dose of 10 mg [2]. It should also be noted that

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supine hypertension is a common feature of more severe autonomic failure, even prior to treatment, and hypertensive cerebrovascular accidents are not increased in autonomic neuropathy. Cerebral hypoperfusion with ischemic strokes due to the falls in blood pressure during the daytime may occur. Nevertheless, it is still recommended to administer the final dose 4 hours before bedtime. Side effects of midodrine are generally dose-related and can be reversed by standing or alpha-receptor antagonist phentolamine. Midodrine is contraindicated in patients with severe organic heart disease, pheochromocytoma, or thyrotoxicosis. Patients with renal failure may require smaller doses, since the active metabolite is renally excreted. Since this drug is metabolized by the liver, in liver disease or with concomitant drugs which have liver clearance, the effects may be increased due to this slower clearance.

ADVERSE EFFECTS AND PRECAUTIONS Patients treated with midodrine should be cautioned about not taking other vasoactive drugs, such as epinephrine, for colds, coughs. Furthermore, concurrent administration of vasoactive agents with anesthetic blocks, such as those used for dental procedures, can result in severe hypertensive episodes.

upon the clinical response. In addition, those patients with postprandial exacerbation may be given a dose prior to meals, so that the peak effect of the medication coincides with the maximal postprandial decrease in upright blood pressure. This medication can also be withheld during recumbency in patients with supine hypertension. Studies are currently in progress to determine the most effective, and optimal dose with the least side effects for specific etiologies of NOH.

Other Special Populations Although not officially approved by the United States Food and Drug Administration for patients under the age of 18, varying doses of midodrine have also been used to treat NOH, POTS, and familial dysautonomia in the pediatric and adolescent age group. There are no significant differences in dosing required for the older patient. It is also important to note that this drug is considered category C in patients during pregnancy (since animal studies have shown an adverse effect and there are no adequate studies in pregnant women) and there are no adequate studies in women during breast feeding.

EPHEDRINE/OTHER ALPHA AGONISTS

Dosing

Mechanism of Action

The dose of midodrine is quite variable and should be individualized. Since there is generally a marked variation in symptoms and decreases in blood pressures (generally more profound in the early morning hours), this agent may be administered orally from 2.5 to 10 mg every 3–4 hours up to a maximum of 50–60 mg per day, depending

The other sympathomimetics such as ephedrine and pseudoephedrine are non-selective alpha agonists, having both direct and indirect effects. A comparison of the various sympathomimetics is listed in Table 130.1. These agents stimulate both alpha and beta receptors. The effectiveness of these agents depends upon the increase in

TABLE 130.1 sympathomimetic Agents for Treatment of neurogenic orthostatic Hypotension Pharmacologic Agent

Mechanism of Action

Dose

Side Effects

Alpha-1-adrenergic agonist with activation of arteriolar and venous vasculature, and decreases venous pooling.

2.5–10 mg every 3–4 h, to maximal dose of 50–60 mg.

Piloerection, urinary retention, supine hypertension, anxiety.

Stimulation alpha/beta receptors Action depends upon receptors and baroreceptor defects. Action depends upon norepinephrine release from postganglionic neurons.

12.5–25 mg T.I.D., 30–60 mg T.I.D.

Nervousness, tremors, anxiety, insomnia, agitation, arrhythmias, supine hypertension. Nervousness, tremors, anxiety, insomnia. Agitation, arrhythmias, supine hypertension.

Clonidine

Antagonizes α-2 adreno-receptors.

Yohimbine

Antagonizes α-2 adreno-receptors.

0.1–0.8 mg in divided doses. 5.4 mg doses.

A. DIRECT ACTIONS Midodrine HCl

B. MIXED DIRECT AND INDIRECT ACTIONS Ephedrine Pseudoephedrine

Phenylpropanolamine Methylphenidate

12–25 mg T.I.D. 5–10 mg T.I.D. dose before 6 p.m.

C. ANTAGONISTS

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Dry mouth, tiredness, sedation, altered mental status, hypertension. Unpredictable responses, hypertension, anxiety, mood stimulation.

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130. MIdodRInE, AdREnERgIC AgonIsTs And AnTAgonIsTs

receptor number and affinity and the reduction in baroreflex modulation that accompanies autonomic failure. Ephedrine produces smooth muscle relaxation and cardiac stimulation. This results in blood pressure increases with increased cardiac output and to a lesser effect, peripheral vasoconstriction [22,23].

Adverse Events and Disadvantages Since ephedrine crosses the blood–brain barrier, central nervous system stimulation is similar to that produced by amphetamines. Frequent side effects, such as nervousness, anxiety, headaches, weakness, dizziness, tremulousness, and supine hypertension result from this class of pharmacologic agents. Furthermore, angina and potentially fatal arrhythmias may be provoked in patients with ischemic heart disease. Ephedrine may also constrict renal blood vessels, and decreased urine production. Other disadvantages include tachyphylaxis (tolerance) which often develops within weeks [22,23].

Dosing The doses required for effective treatment of NOH with these alpha sympathomimetic agents are often higher than those contained in commonly used decongestants. Ephedrine doses of 12.5 to 25 mg three times daily are the most commonly used. Other over-the-counter sympathomimetics, such as pseudoephedrine (30 to 60 mg three times daily) and phenylpropanolamine (12.5 to 25 mg three times daily) are also included in this class. Phenylpropanolamine has been found to be associated with a small but significant risk of cerebrovascular accident and has been removed from the market by the U.S. Food and Drug Administration. Methylphenidate and dextromethorphane sulfate also have indirect and amphetamine-like actions [22,23]. The vasoconstrictor effect results from norepinephrine release from postganglionic neurons, and may be more effective in patients with partial or incomplete lesions. Doses required for methylphenidate are 5–10 mg three times daily with meals, with the last dose being given before 6 pm. Common side effects include agitation, tremor, insomnia, supine hypertension. This pharmacologic agent also has central nervous system stimulation, which limits its use and it is also a controlled substance.

ANTAGONISTS Mechanism of Action Clonidine has a central action on alpha 2-adrenoreceptors, resulting in inhibition of sympathetic cardio-accelerator and vasoconstrictive centers. Clonidine increases baroreceptor activity and acts on peripheral postsynaptic α-2 adrenoreceptors, thus stimulating peripheral α1-adrenergic receptor, which causes vasoconstriction,

increases in venous return and blood pressure, and decreases sympathetic outflow from the central nervous system, peripheral resistance, and renal vascular resistance. The predominance of the pressor effect may be due to decreases in sympathetic outflow and low circulating catecholamine levels. The pressor response depends upon the degree of postjunctional hypersensitivity and autonomic insufficiency. Studies have shown that the change in blood pressure is inversely proportional to the supine level of norepinephrine [24]. This drug is useful for selected patients with NOH and supine hypertension (common in the elderly) or hyperadrenergic orthostatic hypotension. Yohimbine, blocks presynaptic α-2 adrenoreceptors and can increase sympathetic ouflow, potentiates release of norepinephrine from nerve endings, thus activating α-1 and β 1 receptors in the heart and peripheral vasculature, with a rise in blood pressure [25]. Yohimbine increases neurotransmitter release by blocking presynaptic α-2 receptors. Therefore, the efficacy depends upon the capacity of the sympathetic nervous system to be activated and release norepinephrine.

Adverse Events and Disadvantages Both drugs require careful monitoring, as responses depend upon the integrity of the autonomic nervous system. Side effects of clonidine include: dry mouth, sedation, altered mental status, and hypertension. Adverse effects of yohimbine include mood stimulation and anxiety.

Dosing Varying doses of clonidine may be required, from 0.1 to 0.8 mg per day. Yohimbine may be used in 5.4 mg doses.

References [1] McTavish D, Goa KL. Midodrine. A review of its pharmacological principles and therapeutic use in orthostatic hypotension and secondary hypotensive disorders. Drugs 1989;38:757–77. [2] Wright RA, Kaufmann HC, Perera R, Opfer-Gehrking TL, McElligott MA, Sheng KN, et al. A double-blind, dose response study of midodrine in neurogenic orthostatic hypotension. Neurology 1998;51:120–4. [3] Kaufmann H, Brannan T, Krakoff L, Yahr MD, Mandeli J. Treatment of orthostatic hypotension due to autonomic failure with a peripheral alpha-adrenergic agonist (midodrine). Neurology 1988;38:951–6. [4] Jankovic J, Gilden JL, Hiner BC, Kaufmann H, Brown DC, Coghlan CH, et al. Neurogenic orthostatic hypotension: a double-blind placebo controlled study with midodrine. Am J Med 1998;95:38–48. [5] Gilden JL. Midodrine in neurogenic orthostatic hypotension – a new treatment. Int Angiol 1993;12:125–31. [6] Gilden JL, Kaufmann H. Midodrine therapy for neurogenic orthostatic hypotension. A double-blind placebo controlled study [abstract]. Clin Auton Res 1994;4:203. [7] Low PA, Gilden JL, Freeman R, Sheng K, McElligott MA. Efficacy of midodrine vs. placebo in neurogenic orthostatic hypotension: a randomized, double-blind multicenter study for the Midodrine Study Group. J Am Med Assn 1997;277:1046–51.

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AnTAgonIsTs

[8] Malamut R, Freeman R, Gilden J, Tulloch SJ, Kaufmann H. A multicenter, double-blind, randomized, placebo controlled, crossover study to assess the clinical benefit of midodrine in patients with neurogenic orthostatic hypotension [abstract]. Clin Auton Res 2005;15(5):327. [9] Axelrod FB, Goldberg JD, Rolnitsky L, Mull J, Mann SP, Gold von Simson G, et al. Fludrocortison in patients with familial dysautonomia – assessing effect on clinical parameters and gene expression. Clin Auton Res 2005;15(4):249–50. [10] Prakash S, Garg AX, Heidenheim AP, House AA. Midodrine appears to be safe and effective for dialysis-induced hypotension: a systematic review. Nephrol Dial Transplant 2004;19(10):2253–8. [11] Piwinski SE, Jankovic J, McElligott MA. A comparison of postspaceflight orthostatic intolerance to vasovagal syncope and autonomic failure and the potential use of the alpha agonist midodrine for these conditions. J Clin Pharmacol 1994;34(5):466–71. [12] Wecht JM, Rosado-Rivera D, Handrakis JP, Radulovic M, Bauman WA. Effects of midodrine hydrochloride on blood pressure and cerebral blood flow during orthostasis in persons with chronic tetraplegia. Arch Phys Med Rehabil 2010;91(9):1429–35. [13] Fouad-Tarazi F, Okabe M, Goren H. Alpha sympathomimetic treatment of autonomic insufficiency with orthostatic hypotension. Am J Med 1995;99:604–10. [14] Klingenheben T, Credner S, Hohnloser SH. Prospective evaluation of a two-step therapeutic strategy in neurocardiogenic syncope: midodrine as second line treatment in patients’ refractory to betablockers. Pacing Clin Electrophysiol 1999;22(2):276–81. [15] Tsuchida K, Yamazaki R, Kaneko K, Alhara H. Effects of midodrine on experimentally-induced postural hypotension in dogs. Arzneimittelforschung 1986;36(12):1748–51. [16] Lamarre-Cliché M, Souich P, Champlain J, Larochelle P. Pharmacokinetic and pharmacodynamic effects of midodrine

[17]

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on blood pressure, the autonomic nervous system, and plasma naturetic peptides: a prospective, randomized, single-blind, two period, crossover, placebo-controlled study. Clin Ther 2008;30(9):1629–38. Scholing WE. Studies on the effect of the alpha-receptor stimulant, gutron, in the orthostatic syndrome. Wien Klin Wochenschr 1981;93(13):429–34. Ward CR, Gray JC, Gilroy JJ, Kenny RA. Midodrine: a role in the management of neurocardiogenic syncope. Heart 1998;79(1):45–9. Arora RR, Bulgarelli RJ, Ghosh-Dastidar S, Columbo J. Autonomic Mechanisms and therapeutic implications of postural diabetic cardiovascular abnormalities. J Diabetes Sci Technol 2008;2(4):645–57. Gilden JL, Hassan T, Yu A, Ho H, Singh SP. The effects of midodrine therapy on cerebral blood flow in diabetic neurogenic orthostatic hypotension [abstract]. Clin Auton Res 1997;7:247. Gilden JL, Hassan T, Rodriguez L, Singh SP. Depression and cognitive function in patients with orthostatic hypotension [abstract]. Clin Auton Res 1996;6:284–5. Grubb BP, Karas B. Clinical disorders of the autonomic nervous system associated with orthostatic intolerance: an overview of classification, clinical evaluation, and management. PACE 1999;22:798–810. Robertson D, Davis TL. Recent advances in the treatment of orthostatic hypotension. Neurology 1995;45(suppl 5):S26–32. Robertson D, Golderg MR, Tung CS, Hollister AS, Robertson RM. Use of alpha 2 adrenoreceptor agonists and antagonists in the functional assessment of the sympathetic nervous system. J Clin Invest 1986;78(2):576–80. Shibao S, Okamoto LE, Gamboa A, Yu C, Diedrich A, Raj SR, et al. Comparative efficacy of yohimbine against pyridostigmine for the treatment of orthostatic hypotension in autonomic failure. HTN 2010;56(5):847–51.

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131 Agents Potentiating Sympathetic Tone Cyndya Shibao, Luis Okamoto

The autonomic nervous system plays a crucial role in the regulation of acute blood pressure changes in response to upright posture. When this system fails as in patients with autonomic failure, the clinical presentation is often dominated by disabling orthostatic hypotension. The mechanism underlying orthostatic hypotension in autonomic failure is an impairment of autonomic reflexes that affects the ability to release norepinephrine from the postganglionic sympathetic neurons. The degree by which tonic norepinephrine release is impaired varies significantly among patients and depends in part on the severity of the autonomic damage. This residual sympathetic activity can be harnessed with pharmacological probes acting centrally or in peripheral autonomic nerves producing an increase in blood pressure which can be exploited for the treatment of orthostatic hypotension. Even small increases in plasma norepinephrine induced by these agents may lead to exaggerated rises in blood pressure because of autonomic denervation-induced upregulation of adrenoreceptors, and inability to buffer hemodynamic changes due to lack of baroreflex capacity. Among the different strategies to engage residual sympathetic activity, one can stimulate what is left of the patient’s own sympathetic nervous system, preventing the uptake of norepinephrine, hence increasing its availability in the synapse or enhancing sympathetic transmission through the autonomic ganglia. Although all these interventions have shown in clinical studies to increase orthostatic tolerance in autonomic failure, selected patients may have better response which depends mostly on the underlying pathophysiology of their autonomic impairment. There are at least two distinct forms of primary autonomic failure depending on the level of autonomic damage. In patients with central autonomic failure (multiple system atrophy, MSA) the damage resides within the central nervous system and involves the neural connections responsible for baroreflex modulation of sympathetic tone. The neurons that tonically discharge sympathetic activity and distal pathways appear to be intact, whereas in patients with peripheral autonomic failure (pure autonomic failure, PAF and Parkinson’s disease, PD) the neural damage involves more distal structures compared with MSA. In these patients there is a loss of postganglionic

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00131-1

noradrenergic fibers with preservation of the central autonomic pathways [1]. An example of how these different pathophysiologies can impact the response to targeted pharmacological probes is presented in Figure 131.1. Centrally, yohimbine antagonizes α2-adrenergic receptors in the brainstem that stimulate the rostroventrolateral medulla (RVLM) and thus increases central sympathetic outflow. An opposite effect of yohimbine can be found with the norepinephrine reuptake inhibitor atomoxetine that decreases sympathetic activation by enhancing endogenous norepinephrine release in the brainstem and therefore inhibiting the RVLM. In the periphery, these drugs have opposite actions; yohimbine inhibits the presynaptic α2-adrenergic receptors that normally inhibit norepinephrine release, and atomoxetine potentiates the effect of synaptic norepinephrine by inhibiting its reuptake, both resulting in potentiation of sympathetic activation. A delicate balance exists between these opposing central and peripheral actions that in normal individuals result in no changes in blood pressure. However, in autonomic failure patients depending on the level of the lesion, whether is central or peripheral, these drugs could have a totally different effect, a topic that we will discuss in the next part of this chapter.

YOHIMBINE Yohimbine is found in a variety of botanical sources and is the principal alkaloid extracted from the bark of the Pausinystalia yohimbe tree. It is rapidly absorbed after oral administration with a peak plasma concentration occurring at 30 minutes and a half-life of 5 hours [2]. This medication increases norepinephrine release from sympathetic nerves by augmenting central sympathetic outflow and by interfering with inhibitory modulation of presynaptic α2-adrenoreceptors. As expected from its pharmacological actions, which facilitate sympathetic activation, yohimbine increases plasma norepinephrine in a dose-dependent manner and this translates into an increase in blood pressure and heart

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131. AgEnTS PoTEnTIATIng SymPATHETIC TonE

FIGURE

131.1 Schematic representation of the site of action of different agents that potentiate sympathetic activity (atomoxetine, pyridostigmine, yohimbine). Centrally, yohimbine stimulates () central sympathetic tone by antagonizing α2-adrenergic receptors in the brain stem that stimulates the rostroventrolateral medulla (RVLM). The norepinephrine reuptake inhibitor, atomoxetine, inhibits () sympathetic activation by enhancing endogenous norepinephrine (NE) release in the brain stem and therefore inhibiting the RVLM. In the periphery, these drugs have opposite actions. Pyridostigmine stimulates () sympathetic tone by increasing endogenous acetylcholine (Ach), and enhancing sympathetic transmission through the autonomic ganglia.

rate. In normal volunteers this effect occurred at doses of 45 mg or higher, between 60 to 90 minutes after oral administration, lasting on average 4 to 5 hours. Similar effects can be achieved in autonomic failure patients at a very low dose (5.4 mg) given their denervation hypesensitivity [3]. The effect of yohimbine on orthostatic tolerance was first studied by Robertson and collaborators in 12 patients with autonomic failure. Intravenous administration of yohimbine (4–64 μg/kg/iv bolus) increased norepinephrine spillover and blood pressure and reduced the hypotensive response to head-up tilt in these patients [4]. In open label studies, oral administration of 5.4 mg of yohimbine increased blood pressure by 50 mmHg with a peak effect at 75 minutes [5]. This pressor effect was comparable to that of midodrine (a direct α1-adrenergic agonist) and phenylpropanolamine, another sympathomimetic. More recently, in a single-blind, placebo-controlled, crossover study, the same dose of yohimbine increased standing diastolic blood pressure by 11 mmHg and reduced presyncopal symptoms, particularly lightheadedness in 31 patients with autonomic failure (Fig. 131.2 A and B) [6]. The action of yohimbine on blood pressure is exclusively mediated by its interaction with the sympathetic nervous system. Yohimbine has no effect in patients with dopamine β-hydroxylase deficiency, presumably because the lack of norepinephrine in their neurons. These observations imply that even patients with severe forms of autonomic failure have some degree of residual sympathetic tone that can be potentiated by this drug. The pressor response to yohimbine differs depending on the underlying pathophysiology of their autonomic failure. Patients with MSA who had intact efferent sympathetic nerves had a greater pressor response than patients with PAF, who are affected by loss of efferent noradrenergic fibers. This difference is likely due to residual

sympathetic tone in MSA because the pressor response to yohimbine was strongly correlated with the decrease in blood pressure produced by trimethaphan. Also, the increase in blood pressure induced by yohimbine was also greater in MSA patients with supine hypertension, known to be mediated by residual sympathetic tone, than in those without supine hypertension. Yohimbine is well-tolerated in autonomic failure patients. There was no report of adverse events associated with oral administration of yohimbine in clinical studies. However, these trials were done in the acute setting. Caution should be used when given to autonomic failure patients with concomitant atrial fibrillation because there is a concern that it may decompensate this condition inducing rapid ventricular response.

ATOMOXETINE Atomoxetine is a selective norepinephrine transporter (NET) blocker commonly used for the treatment of attention deficit hyperactivity disorder in children and adults. After oral administration, the time to achieve peak plasma concentration is 1 to 2 hours and its elimination half-life is 5 hours [7]. These parameters change if the individual is a poor metabolizer for cytochrome P450 2D6 (CYP2D6) which happens in around 7% of Caucasians. Atomoxetine blocks the norepinephrine reuptake receptor and increases neurotransmitter concentrations in the neuroeffector junction, an effect that would potentially lead to a pressor response. However, this mechanism seems to be counteracted by a central sympatholytic action through activation of central α2-adrenergic receptors. The balance between central sympatholytic and peripheral sympathomimetic effects produces minimal, if any, increase in blood pressure in normal subjects.

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PyRIdoSTIgmInE

effect could be useful as a diagnostic tool to determine the level of the lesion within the autonomic nervous system in patients presenting with orthostatic hypotension and therefore enable identification of early forms of MSA prior to the development of motor deficits. Furthermore, the profound pressor response achieved with 18 mg of atomoxetine can be used in the treatment of refractory orthostatic hypotension in MSA patients. In this regard, 18 mg of atomoxetine can be given 1 hour prior to any activities, the effect lasting between 4–5 hours. Patients should be instructed to avoid an evening dose because of the risk of supine hypertension.

PYRIDOSTIGMINE

FIGURE 131.2 Panel A represents the changes in standing diastolic blood pressure 60 minutes after oral administration of placebo and 5.4 mg of yohimbine, in 31 patients with autonomic failure. Panel B represents changes in pre-syncopal symptoms with each intervention (placebo, yohimbine). The asterisk (*) indicates P   0.05 was considered statistical significant.

The effect of NET blockade can be dissected in humans by determining its pressor response in patients with distinct forms of autonomic impairment based on the level of the lesion. In theory NET blockade could induce a pressor effect in patients with intact peripheral sympathetic fibers (MSA, with central autonomic impairment), but not in patients with PAF and peripheral autonomic denervation. Indeed, pediatric dose of atomoxetine (18 mg) acutely increased seated and standing systolic blood pressure in patients with MSA by about 50 mmHg compared with placebo (Fig. 131.3). In contrast, in patients with peripheral autonomic failure, atomoxetine did not elicit any pressor response [8]. This diverge

Pyridostigmine inhibits the enzyme acetylcholinesterase and increases the transmission of impulses from cholinergic neurons across the synaptic cleft. After oral administration, pyridostigmine achieved peak plasma concentration around 1.7 hours and has a short half-life of 1–2 hours. Because ganglionic neurotransmission is cholinergic, it facilitates both sympathetic and parasympathetic function. The appeal of this pharmacological approach is that it will be silent under conditions of low sympathetic tone such as in supine posture, but will potentiate the sympathetic activation that occurs on standing. This drug has the potential, therefore, to improve orthostatic blood pressure without worsening supine hypertension. Two previous studies have reported the beneficial effect of pyridostigmine as a treatment for orthostatic hypotension in patients with autonomic failure. The effect of 60 mg of pyridostigmine on standing diastolic blood pressure was first tested in an open label study in 15 patients with autonomic failure. Pyridostigmine significantly increased orthostatic blood pressure and reduced the fall in blood pressure associated with head-up tilt [9]. In a subsequent doubleblind, randomized 4-way crossover study, pyridostigmine increased upright systolic blood pressure by only 4 mmHg compared with placebo; the combination with 5 mg of midodrine was slightly more effective [10]. The improvement in orthostatic blood pressure in both studies was associated with a significant improvement in orthostatic symptom without causing supine hypertension. A recent study, however, did not yield the same results; pyridostigmine did not improve orthostatic tolerance or symptoms, because autonomic failure patients were more severely affected with orthostatic hypotension than in previous studies [6]. This supports the notion that the response to pyridostigmine is proportional to the degree of residual sympathetic tone. Pyridostigmine should be considered part of the available therapy for the treatment of autonomic failure patients. As with other agents, therapy should be individualized because some patients may have only a modest response. It is probably more effective in patients with some degree of residual sympathetic tone.

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FIGURE 131.3 Panel A represents systolic blood pressure measurements with 18 mg of atomoxetine and placebo in patients with central autonomic failure (MSA). Systolic blood pressure increased significantly with atomoxetine 18 mg (P   0.001); the asterisk (*) indicates P   0.001. Panel B. Systolic blood pressure measurements with atomoxetine and placebo in patients with peripheral autonomic failure (PAF, PD). No significant changes in SBP were observed (P  0.08). (Reprinted with permission from Wolters Kluwer Health/Lippincott, Williams & Wilkins.)

References [1] Shibao C, Gamboa A, Diedrich A., Biaggioni I. Management of hypertension in the setting of autonomic failure. A pathophysiological approach. Hypertension. 2005;45:469–75. [2] Tam SW, Worcel M, Wyllie M. Yohimbine: a clinical review. Pharmacol Ther 2001;91:215–43. [3] Onrot J, Goldberg MR, Biaggioni I, Wiley R, Hollister AS, Robertson D. Oral yohimbine in human autonomic failure. Neurol 1987;37:215–20. [4] Robertson D, Goldberg MR, Tung CS, Hollister AS, Robertson RM. Use of α2 adrenoreceptor agonists and antagonists in the functional assessment of the sympathetic nervous system. J Clin Invest 1986;78:576–81. [5] Jordan J, Shannon JR, Biaggioni I, Norman R, Black BK, Robertson D. Contrasting actions of pressor agents in severe autonomic failure. Am J Med 1998;105:116–24.

[6] Shibao C, Okamoto LE, Gamboa A, Yu C, Diedrich A, Raj SR, et al. Comparative efficacy of yohimbine against pyridostigmine for the treatment of orthostatic hypotension in autonomic failure. Hypertension 2010;56:847–51. [7] Simpson D, Plosker GL. Atomoxetine: a review of its use in adults with attention deficit hyperactivity disorder. Drugs 2004;64:205–22. [8] Shibao C, Raj SR, Gamboa A, Diedrich A, Choi L, Black BK, et al. Norepinephrine transporter blockade with atomoxetine induces hypertension in patients with impaired autonomic function. Hypertension 2007;50:47–53. [9] Singer W, Opfer-Gehrking TL, McPhee BR, Hilz MJ, Bharucha AE, Low PA. Acetylcholinesterase inhibition: a novel approach in the treatment of neurogenic orthostatic hypotension. J Neurol Neurosurg Psychiatry 2003;74:1294–8. [10] Singer W, Sandroni P, Opfer-Gehrking TL, Suarez GA, Klein CM, Hines S, et al. Pyridostigmine treatment trial in neurogenic orthostatic hypotension. Arch Neurol 2006;63:513–8.

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132 Acetylcholinesterase and its Inhibitors Brett A. English, Andrew A. Webster

CHOLINESTERASES AND ACETYLCHOLINE METABOLISM

MOLECULAR PHARMACOLOGY OF ACETYLCHOLINESTERASE INHIBITORS

Cholinergic neurotransmission is mediated by the neurotransmitter acetylcholine (ACh). Upon release, acetylcholine (ACh) is rapidly hydrolyzed into choline and acetic acid (Chapter 14) by a family of enzymes called cholinesterase (ChE). There are two types of ChE enzymes in mammals, acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) each sharing ~65% amino acid sequence homology. In mammals, the AChE gene is encoded by a single gene, however, differences in mRNA splicing and post-translational modifications within their carboxy-termini result in three different isoforms. AChE is localized to the cytoplasm and outer cell membrane of blood and neural synapses allowing both intracellular and extracellular ACh metabolism. Within the CNS, AChE exists as a homomeric oligomer (catalytic subunits) and a heteromeric oligomer (catalytic and structural domains). Each active site of AChE contains a negatively charged anionic site and an esteratic site containing crucial residues for enzyme activity (Fig. 132.1) with six active sites per AChE molecule. Upon binding of ACh, the esteratic site binds with the acyl moiety of ACh, while the anionic site associates with the positively charged quaternary nitrogen, resulting in rapid hydrolysis of ACh achieving metabolic rates of 0.1 millisecond. Butyrylcholinesterase is found in plasma, liver and the CNS (neurons and glia) and differs in its substrate specificity for ACh. BuChE exhibits less specificity for the ACh substrate, with AChE hydrolyzing ACh at low substrate concentrations. Recently, BuChE has received increased interest in Alzheimer’s disease (AD) patients, as recent data from cortex of these patients has shown that as the disease progresses, concentrations of AChE decline dramatically, while levels of BuChE in certain brain remain relatively unchanged increasing their importance in modifying the disease.

Inhibition of AChE is accomplished by either reversible (competitive) or irreversible mechanisms involving binding to one of three domains within the AChE active site. Irreversible inhibitors (see below) form covalent bonds within the serine active center of AChE resulting in a stable complex, inactivating the enzyme. Reversible inhibitors such as donepezil and edrophonium bind to the active center and choline subsite of the active center respectively. Carbamate inhibitors bind to the serine active center producing a carbamoylated enzyme. After administration of an AChE inhibitor, hydrolytic activity of AChE is abolished, leading to increased levels of ACh at the synapse and potentiating postsynaptic cholinergic activity within the CNS, autonomic ganglia and neuromuscular junctions. The extent and duration of the postsynaptic potentiation depends on the AChE inhibitor used and dose administered. The specific actions and indications for each of the different classes of AChE inhibitors are described in further detail below. The most prominent effects of AChE inhibition outside the CNS are on organ systems that include cardiovascular, gastrointestinal, ocular and skeletal muscle systems and are summarized in Table 132.1.

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00132-3

Quaternary Amines Compounds within this chemical class consist of quaternary alcohols and include agents such as edrophonium and ambenonium, which form electrostatic and hydrogen bonds within the AChE active site resulting in a reversible inhibition of the enzyme with a relatively short duration of action (Table 132.1). Both agents are highly polar, exhibiting poor CNS penetration. Edrophonium is given intravenously for the differential diagnosis of myasthenia gravis and as a reversing agent for anticholinergic toxicity

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due to its short duration of action. Ambenonium is similar to neostigmine in that it is used to treat myasthenia gravis, however due to its short biologic half-life, it must be given several times a day.

Carbamates This group of compounds consists of carbamate esters that undergo two-step hydrolysis forming a covalently linked carbamoylated AChE–drug complex that is considerably more resistant to hydration, thus resulting in a reversible inhibition of AChE for up to 4 hours or more. Classical carbamate AChE inhibitors include physostigmine, neostigmine and pyridostigmine. A newer agent, rivastigmine, was developed in 1997 and indicated for cognitive dysfunction in 2006. Physostigmine is an alkaloid extract from the Calabar bean and has been primarily used to treat glaucoma. However, unlike physostigmine, the other classical inhibitors pyridostigmine and neostigmine lack CNS penetration due to the presence of a quaternary ammonium charge preventing blood–brain barrier

Gl

Hi

s

44

0

u

-C=O

32

7S

er

20

O

0

O

O-

N+

FIGURE 132.1 Molecular pharmacology of acetylcholinesterase inactivation. Active binding sites of AChE and hydrolysis of ACh. The positively charged quaternary nitrogen of ACh binds electrostatically to an anionic site of AChE. The acyl group of ACh undergoes nucleophilic attack forming a covalent bond resulting in metabolism of ACh at the esteratic site.

crossing. These agents have been primarily used in the treatment of myasthenia gravis. Rivastigmine (Exelon®) is considered a “pseudoirreversible” inhibitor, forming a carbamoylated complex resulting in equipotent inhibition of both AChE and BuChE. It is indicated for the treatment of mild-tomoderate AD and cognitive dysfunction associated with Parkinson’s disease (PD). Unlike other AChE inhibitors used in AD, rivastigmine does not undergo hepatic cytochrome p450 enzymes (CYP 450).

Phenanthrene and Piperidines The two representative agents in these chemical classes are galantamine (Reminyl®) and donepezil (Aricept®) respectively. Galantamine a selective, reversible inhibitor of AChE, also exerts allosteric, modulatory activity at nicotinic acetylcholine receptors (nAChRs) resulting in enhanced cholinergic functioning. Donepezil is a selective, reversible inhibitor of AChE, with little affinity for BuChE. Both drugs undergo hepatic metabolism via cytochrome P450 isoenzymes CYP2D6 and 3A4. Each agent is primarily indicated for the symptomatic treatment of mild-to-moderate AD, with clinical data for both agents demonstrating clinical benefits that were maintained for 36 months. Additionally, galantamine has been used to treat cognitive dysfunction associated with Parkinson’s disease and Lewy body dementia, while donepezil has been used to treat attention deficit-hyperactivity disorder (ADHD). While each of these compounds exert their precognitive effects in AD by the reversible inhibition of central AChE, their stabilization of cognitive deterioration associated with this disorder points to addition disease modifying mechanisms by non-cholinergic mechanisms. Suggested mechanisms include amyloid precursor protein (APP) processing through a non-amyloidogenic pathway and A-beta protein expression that is independent of AChE inhibition.

TABLE 132.1 Chemical Classes and Pharmacology of select Acetylcholinesterase Inhibitors Chemical class

Inhibitor

Site of action

Pharmacologic effect/indication

Duration of action

Mono-quaternary amine

Edrophonium

Neuromuscular junction

Myasthenia gravis

10 min

Bis-quaternary amine

Ambenonium

Neuromuscular junction

Myasthenia gravis

Carbamates

Physostigmine Pyridostigmine Neostigmine Rivastigmine

CNS, parasympathetic nervous system Neuromuscular junction Neuromuscular junction CNS

Glaucoma Myasthenia gravis Myasthenia gravis Alzheimer’s disease

1–5 h 2–4 h 1–2 h 10–12 h

Phenanthrene

Galantamine

CNS

Alzheimer’s disease

7–10 h

Piperidine

Donepezil

CNS

Alzheimer’s disease

Organophosphate

Parathion Malathion Sarin (GB) Soman (GD) VX

CNS, peripheral CNS, peripheral CNS, peripheral CNS, peripheral CNS, peripheral

Pesticide Pesticide Chemical warfare Chemical warfare Chemical warfare

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 100 h  100 h  100 h  100 h  100 h

633

THERAPEuTIC APPlICATIons

Organophosphates Organophosphates (OPs) are a group of phosphoric acid ester compounds that upon binding to AChE are hydrolyzed, producing phosphorylation of the AChE active site resulting in irreversible inactivation of AChE. This covalently bound enzyme–phosphorylation complex is highly stable, hydrolyzing only after several hundred hours. Additionally, some OP compounds undergo a process called aging. This process involves breaking one of the oxygen-phosphorous bonds, which results in further strengthening of the covalent bond between the OP and AChE. For example, the nerve agent soman “ages” within 10 minutes, while VX requires  48 hours. This group of irreversible AChE inhibitors has been used as insecticides and as highly toxic “nerve gases” in both warfare and terrorism. OPs that have been used as commercial insecticides include parathion, paraoxon (active metabolite of parathion), diazinon and malathion. Parathion is inactive at inhibiting AChE and must be metabolized into its active components of paraoxon and malaoxon. Commercial use of parathion has decreased dramatically due to its toxicity and risk accidental overdose in humans. Malathion has become increasing used in commercial spraying due to its increased safety in that it is rapidly detoxified by plasma carboxylesterases in mammals. A number of OP compounds have been used as chemical weapons and include classical agents such as tabun (GA), sarin (GB), soman (GD), cyclosarin (GF) and VX. All of these agents exist as liquids at standard temperature and pressure, and are highly volatile, evaporating at room temperature. Their volatility and persistence (T1/2 in ground or material) is why these agents have gained importance in both warfare and terrorism, as this property contributes to the differential clinical toxidrome with either liquid or vapor exposure. For instance, G-agents have the density and evaporation point similar to water, therefore constituting a greater vapor hazard compared to more persistent agents such as VX, which exhibits an oily consistency. As a result, the cholinergic crisis due to increased ACh levels occurs much more quickly with vapor exposure than dermal contact with liquid exposure.

THERAPEUTIC APPLICATIONS Inhibitors of AChE have been found useful in a number of clinical indications, however since these medications indirectly increase levels of ACh facilitating non-selective postsynaptic cholinergic transmission, they are associated with a number of off-target adverse reactions (Table 132.2). The carbamate compounds have found clinical utility in the treatment of acute-angle glaucoma and

TABLE 132.2 organ system Effects of Acetylcholinesterase Inhibitors Organ system

Effect

Cardiovascular

↑ Autonomic activity w/parasympathetic predominance ↓ Inotropic, chronotropic and dromotropic effects with associated ↓ cardiac output ↑ Systemic vascular resistance ↑ Contraction, secretions and motility Constriction of pupillae sphincter muscle (miosis) Prolonged strength of contraction ↑ Secretions from brochial, lacrimal, sweat and salivary glands ↑ Urination ↑ Cognition and attention Overdose results in convulsions and respiratory arrest

Gastrointestinal Ocular Neuromuscular junction Secretory glands Urinary Central nervous system

accommodative esotropia. Additionally these agents have been used successfully to treat postoperative ileus, urinary retention and dry mouth associated with Sjogren’s syndrome. The quaternary compounds have been used primarily in the diagnosis and treatment of myasthenia gravis, an autoimmune disorder affecting skeletal muscle neuromuscular junction. The carbamates and quaternary amines have been used to treat supraventricular tachycardias and antimuscarinic toxicity associated with atropine-like drugs. Lastly, the novel AChE inhibitors, rivastigmine, galantamine and donepezil have been used to primarily treat AD, however several have been used to successfully treat cognitive impairments associated with PD, ADHD, schizophrenia and delirium.

Further Reading Caldwell JE. Clinical limitations of acetylcholinesterase antagonists. J Crit Care 2009;24:21–8. Giacobini E. Cholinesterases: new roles in brain function and in Alzheimer’s disease. Neurochem Res 2003;28(3/4):515–22. Giacobini E. Do cholinesterase inhibitors have disease-modifying effects in Alzheimer’s disease?. CNS Drugs 2001;15(2):85–91. Jann MW, Shirley KL, Small GW. Clinical pharmacokinetics and pharmacodynamics of cholinesterase inhibitors. Clin Pharmacokinet 2002;41(10):719–39. Karczmar AG. Cholinesterases (ChEs) and the cholinergic system in ontogenesis and phylogenesis, and non-classical roles of cholinesterases – a review. Chemico-Biologic Interac 2010;187(1–3):34–43. Newmark J. Nerve agents. The Neurologist 2007;13(1):20–32. Pepeu G, Giovannini MG. Cholinesterase inhibitors and memory. Chemico-Biolog Interact 2010;187:403–8. Racchi M, Mazzucchelli M, Porrello E, Lanni C, Govoni S. Acetylcholinesterase inhibitors: novel activities of old molecules. Pharm Res 2004;50:441–51. Serotonin, acetylcholine and histamine (2001). In: Nestler EJ, Hyman SE and Malenka RC, editors. Molecular neuropharmacology: a foundation for clinical neuroscience. New York: McGraw-Hill; pp. 201–208.

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133 Fludrocortisone David Robertson, Rose Marie Robertson INTRODUCTION

CLINICAL PHARMACOLOGY

Fludrocortisone (9-alpha-fluorohydrocortisone) has been the most widely used drug in treating orthostatic hypotension in the United States since it was introduced for this purpose 50 years ago. It is less widely used in Europe. Studies by Frick and by Liddle in the 1960s demonstrated that the addition of a fluorine atom to the cortisol molecule fundamentally altered the pharmacodynamics of the parent compound, producing a drug with potent mineralocorticoid, but minimal glucocorticoid effect. The overall effect of this agent in patients with autonomic failure is a rise in blood pressure, both in the supine and the upright postures, and over time fluid retention based on sodium conservation at the level of the kidney. It has also been used in the management of those patients with vasovagal syncope in whom maintaining an adequate fluid balance is otherwise difficult. A controlled trial of fludrocortisone in neurally mediated syncope should be completed as this is going into press. It has been tested and found unhelpful in patients with the chronic fatigue syndrome without autonomic deficits.

Fludrocortisone is rapidly absorbed following oral administration and declines with a half-life of about two to three hours. With doses of 0.2 mg, fludrocortisone levels peaked at about 2.4 ng/mL after 1–2 hours and were undetectable after 12 hours. With 2 mg, the levels peaked at 17.6–24.5 ng/mL at 1 hour and were undetectable by 24 hours. With the 2 mg dose, plasma cortisol fell at 10 hours to 71–87% (14–23 ng/mL) of baseline. The estimated half-life for fludrocortisone is 1.6 hours for the 2 mg dose and 2.4 hours for the 0.3 mg dose. While this relatively short half-life was unexpected, because of the action of the drug in the nucleus to alter transcription, doses as low as 0.05 mg once daily may sometimes be efficacious. Some physicians give fludrocortisone in a twice daily regimen. There are several issues to consider in using fludrocortisone effectively. First, long-term studies of outcomes after fludrocortisone have not been undertaken, so there could be chronic untoward effects that have heretofore not been recognized. Second, since the full pressor action of fludrocortisone is not seen for 1–2 weeks, doses should not be altered more frequently than at weekly or biweekly intervals. The initial dose should usually be 0.05 to 0.1 mg po daily, with weekly or biweekly titration by 0.1 mg increments, aiming for improvement in symptoms of orthostatic hypotension without eliciting unacceptable degrees of supine hypertension. In severe orthostatic hypotension many patients have supine hypertension even before fludrocortisone is introduced. To obtain improved symptoms, we generally accept a weight gain of 3–5 pounds and mild ankle swelling, but in exceptional cases, greater weight gains may be justified. The patient should be educated about the expected time course of the effect. It will be rare to find additional benefit beyond 0.2 mg po qd, but doses as high as 2.0 mg/day have been reported; such doses when sustained commonly lead to severe potassium and magnesium depletion, with muscle damage and raised CPK. There is little if any glucocorticoid effect at doses in the range of 0.1 to 0.2 mg daily, but reduced cortisol level due to ACTH suppression has been seen following a single dose of 2.0 mg. Weight is a good guide to the

MECHANISM OF EFFECT The cardiovascular response to fludrocortisone is a gradual rise in blood pressure in patients with orthostatic hypotension. This is initially related to sodium retention with an increase in plasma volume. This effect, dependent on mineralocorticoid receptors within the nucleus and altered gene transcription, takes several days to weeks to reach its peak. Although the increased plasma volume may return to baseline subsequently, in many patients a residual beneficial pressor effect continues. At least in some patients, this has been demonstrated to be due to increased peripheral resistance, and the enhanced response to infused norepinephrine seen in several studies suggests a mechanism via cell surface receptors acting through second messengers. Such mechanisms would be expected to be activated more rapidly and to follow a different time course than those requiring alteration of cellular transcription.

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133. FludRoCoRTIsonE

required dose, and the weight gain due to fluid retention should be limited to 5 lbs. Because much of the blood pressure effect is related to this fluid retention, the addition of a waist-high compression garment helps to keep this fluid distributed in the most beneficial manner, if patients are willing to tolerate the discomfort and inconvenience of these garments. These garments are less tolerated in hot climates.

SIDE EFFECTS Fludrocortisone should not be used in patients who cannot tolerate increased fluid retention, for example those with heart failure, but this is rarely an issue, since patients with pre-existing congestive heart failure rarely have autonomic failure. In unusual cases patients with pure autonomic failure and on no drug therapy have experienced improvement in orthostatic hypotension when they develop congestive heart failure. If symptoms of pulmonary congestion or even of pulmonary edema do develop in an autonomic failure patient after a fludrocortisone-induced increase in plasma volume, these symptoms will respond very rapidly with assumption of the seated or upright posture. There are potential side effects and complications that can develop with the use of fludrocortisone. Nearly 50% of patients will develop hypokalemia, and this can appear within the first week of treatment. It will respond to oral supplementation with potassium, which usually must then be given chronically. A smaller group, perhaps 5%, will develop concomitant hypomagnesemia, and while correction of the hypokalemia will often lead to secondary correction of the hypomagnesemia, if this is not complete, small doses of magnesium sulfate can be added. Fludrocortisone therapy commonly produces the side effect of headache, especially initially and particularly in younger, healthier patients. For example, while 5 days of fludrocortisone therapy would seem to be an ideal approach for astronauts, who commonly experience orthostatic intolerance when they return from microgravity associated with space travel, the headache in this healthy and rather young population was a limitation, and fludrocortisone use in the final 5 days before return to earth was ultimately abandoned. Likewise, young patients using fludrocortisone to prevent vasovagal syncope may find headache a limiting factor. However, patients with severe autonomic failure, in whom it is most helpful, do not usually complain of headache with fludrocortisone. An additional important issue is the development of an excessive rise in blood pressure, especially in the supine posture. While supine hypertension can be lowered acutely by raising the head of the bed, by having the patient sit or stand, and/or by giving a carbohydrate snack, if it persists a reduction in dosage or discontinuation may be necessary. As with all drugs, interactions with other medications must be considered. While it is rare for

O HO

HO

OH

H F

H

O

FIGURE 133.1 Fludrocortisone structure.

patients with the potential for falling caused by orthostatic hypotension to be treated with warfarin, occasional patients will require the latter for valvular heart disease or another indication. In some of these patients, fludrocortisone will lead to an increased warfarin requirement to achieve the same INR. The institution of rifampicin can also lower fludrocortisone levels.

LONG-TERM EFFECTS OF FLUDROCORTISONE A theoretical concern with the long-term use of fludrocortisone has arisen in recent years with the description of novel effects of aldosterone. Classically, aldosterone has been known to act at the epithelial cell level to induce sodium reabsorption and potassium excretion, leading to intravascular volume expansion, and the antagonism of aldosterone by spironolactone has been used to good effect in the treatment of heart failure. However, in the Randomized Aldactone Evaluation Study (RALES), the addition of spironolactone to existing therapy in patients with heart failure led to a significant reduction in morbidity and mortality as well. Aldosterone has additional, non-epithelial effects in the kidney and in vascular smooth muscle, involving activation of the sodium/hydrogen antiporter, and it is possible that this can produce cardiovascular damage independent of the level of blood pressure. Since this cardiovascular damage in experimental settings can be prevented by administering a selective mineralocorticoid receptor antagonist, one must have some concern about the chronic use of a mineralocorticoid receptor agonist. However, fludrocortisone has been used chronically in patients with Addison’s disease with good results, and in patients with severe orthostatic hypotension, the benefit would seem to outweigh this theoretical risk. It is prudent in all patients to be certain that all appropriate measures of cardiovascular protection and prevention are being employed.

Further Reading Blockmans D, Persoons P, Van Houdenhove B, Lejeune M, Bobbaers H. Combination therapy with hydrocortisone and fludrocortisone does not improve symptoms in chronic fatigue syndrome: a randomized, placebo-controlled, double-blind, crossover study. Am J Med 2003;114:736–41.

XIII. MANAGEMENT OF AUTONOMIC DISORDERS

long-TERm EFFECTs oF FludRoCoRTIsonE

Chobanian AV, Volicer L, Tifft CP, Gavras H, Lian CS, Faxon D. Mineralocorticoid-induced hypertension in patients with orthostatic hypotension. N Engl J Med 1979;301:68–73. Frick MH. 9-alpha-fluorohydrocortisone in the treatment of postural hypotension. Acta Med Scan 1966;179:293–9. Frishman WH, Azer V, Sica D. Drug treatment of orthostatic hypotension and vasovagal syncope. Heart Dis 2003;5(1):49–64. Goldstein DS, Robertson D, Esler M, Straus SE, Eisenhofer G. Dysautonomias: clinical disorders of the autonomic nervous system. Ann Int Med 2002;137:753–63.

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Laviolle B, LeMaguel P, Verdier MC, Massart C, Donal E. Biological and hemodynamic effects of low doses of fludrocortisone in healthy volunteers with hypoaldosteronism. Clin Pharmacol Ther 2010;88:183–90. Mitsky VP, Workman RJ, Nicholson E, Vernikos J, Robertson RM, Robertson D. A sensitive radioimmunoassay for fludrocortisone in human plasma. Steroids 1994;59:555–8. Rocha R, Williams GH. Rationale for the use of aldosterone antagonists in congestive heart failure. Drugs 2002;62:723–31.

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134 Acarbose Cyndya Shibao Patients with severe autonomic failure have a high incidence of postprandial hypotension (PPH). This condition is defined as a fall in systolic blood pressure of more than 20 mmHg occurring within 2 hours after a meal [1]. Blood pressure measurements should be obtained while sitting or supine because the hypotensive effect after a meal might be influenced by the amount of blood pooling to the lower body due to gravitational forces. Postprandial hypotension may present with a variety of symptoms that range from feelings of lightheadedness, nausea, blurred vision and weakness to somnolence or syncope. Isolated cases of PPH precipitating transient ischemic attack, and/or angina pectoris have been reported. This is particularly worrisome in patients with existing carotid stenosis or coronary artery disease, in whom large blood pressure reductions after a meal could decrease arterial blood flow to target organs. In autonomic failure patients, systolic blood pressure falls on average 50 mmHg after a meal. The drop in blood pressure starts within the first 10 to 15 minutes after the meal is finished with a nadir between 30–60 minutes [1]. This hypotensive effect can last up to 2 hours and in severe cases, pre-syncopal symptoms can be so debilitating that patients prefer to fast to avoid them, inducing substantial weight loss in this already frail population. Furthermore, orthostatic and postprandial hypotension can co-exist and have additive effects in autonomic failure patients, potentially increasing their risk for syncope.

PATHOPHYSIOLOGY OF POSTPRANDIAL HYPOTENSION IN AUTONOMIC FAILURE The etiology of PPH is likely multifactorial. In normal subjects, food ingestion promotes biochemical and hormonal changes that result in blood pooling within the splanchnic circulation. Doppler ultrasonography demonstrates increase in the superior mesenteric artery blood flow [2]. In order to maintain blood pressure within a normal range, a variety of hemodynamic changes are necessary, including an increase in heart rate, stroke volume and cardiac output. These changes are orchestrated by activation of the sympathetic nervous system, characterized by increases in plasma norepinephrine and muscle sympathetic nerve activity [1]. Failure of these

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00134-7

compensatory mechanisms seems pivotal in the development of PPH, hence the greater prevalence of this condition in subjects with autonomic impairment. Gastrointestinal and pancreatic hormones appear to play a key role in the pathogenesis of PPH. These peptides are secreted in response to food ingestion and are known to have vasodilating properties particularly on the splanchnic circulation. Several attempts have been made to identify the specific hormones involved but results so far are inconclusive. For instance, both autonomic failure patients with PPH and normal controls have similar levels of plasma insulin, gastrin, vasoactive intestinal polypeptide, somatostatin, and cholecystokinin-8 before and after a standard meal [1]. This finding, however, does not exclude a role for these peptides, since patients with autonomic failure are known to be more sensitive and have enhanced hypotensive responses to vasoactive agents. The most important evidence of the role of gastrointestinal peptides in the pathogenesis of PPH derives from medication trials. Notably, drugs that blunt the release of these hormones, e.g., octreotide, or that antagonize their action, e.g., caffeine, attenuate PPH and are integral components of the treatment strategy for this condition [3–5]. Food composition is an important factor in the severity of postprandial hypotension. Postprandial reductions in blood pressure are more marked after carbohydrate rich meals than after fat and protein. The rate of nutrient delivery to the small intestine seems to be also an important determinant of blood pressure changes. The magnitude of the fall in systolic blood pressure is substantially greater when glucose is infused intraduodenally at a rate of ~3 Kcal/min compared with infusion at a rate of 1 Kcal/ min [6]. Of note, the hypotensive response to oral glucose is attenuated by the viscous polysaccharide guar, which slows gastric emptying [5].

ACARBOSE IN THE TREATMENT OF POSTPRANDIAL HYPOTENSION Treatment of PPH remains a challenge in patients with autonomic failure. Non-pharmacologic interventions have been proposed to prevent this condition. In mild cases a reduction in meal size from three large to six smaller meals, and/or a decrease in its carbohydrate content,

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134. ACARbosE

FIGURE 134.1 Change in systolic blood pressure (circles) and diastolic blood pressure (squares) measurements during placebo (open) and acarbose (filled) at baseline and for 90 minutes post-meal challenge. Postprandial hypotension was significantly attenuated with acarbose. The asterisk (*) indicates P  0.01. (Reprinted with permission from Wolters Kluwer Health/ Lippincott, Williams& Wilkins.)

may be useful. This latter approach, however, is difficult to adhere to, because carbohydrates represent 45 to 65% of the normal Western diet. Alternatively, water drinking (16 ounces of plain water) at least 20 minutes prior to meal ingestion can also attenuate the postprandial fall in blood pressure. Severe PPH cases are challenging to treat; these patients often do not respond to non-pharmacological agents. Postprandial pre-syncopal symptoms occur even while supine, because the fall in blood pressure decreases cerebral perfusion below a critical threshold. Therefore, a pharmacological intervention is often necessary. Caffeine (250 mg) or octreotide (25 µg subcutaneously) has been shown to prevent PPH [5]. The latter should be use only in refractory cases because of its route of administration and potent pressor effect in autonomic failure patients. Recent studies showed that acarbose, a medication used to control postprandial hyperglycemia in type 2 diabetes mellitus, could have an important therapeutic role in PPH. Acarbose is a pseudotetrasaccharide, a natural microbial product derived from cultures of Actinoplanes strain SE 50. This medication inhibits alpha-glucosidases in the brush border of the small intestine, delaying glucose absorption by decreasing the breakdown of complex carbohydrates. These actions also decrease the release of gastrointestinal hormones and slow gastric emptying [7]. Case reports have shown improvement in PPH with acarbose in patients with diabetes mellitus; in an open label study, voglibose, another alpha-glucosidase inhibitor, improved the blood pressure fall after an oral glucose load in patients with multiple system atrophy, Parkinson’s disease and diabetes mellitus [8]. Furthermore, the hemodynamic effect of acute administration of acarbose has been studied in patients with severe autonomic failure and healthy elderly in randomized, placebo-control clinical trials with favorable results [7,9]. Acute administration of 100 mg of acarbose, 20 minutes before a standardized meal reduced the postprandial fall

in systolic blood pressure by 17 mmHg compared with placebo in autonomic failure patients (Fig. 134.1). This effect was associated with an improvement in total peripheral resistance but no significant changes in cardiac output [7]. Moreover, Gentilcore and colleagues also observed an attenuation of the postprandial fall in blood pressure of about ~6 mmHg in healthy elderly individuals [9]. This improvement was associated with delayed gastric emptying. Even though these studies showed attenuation or even prevention of PPH and provide a proof-of-concept for the use of acarbose in this condition, further studies designed to assess the effect of chronic administration of acarbose for the treatment of PPH are needed. No serious side effects have been reported in these studies, but again, this medication was only used acutely. Chronic treatment with acarbose can be associated with an increased rate of flatulence and loose stools as a result of specific drug effects on carbohydrate digestion. To minimize gastrointestinal adverse effects, treatment should be initiated with a low dose and gradually increased until adequate effects are achieved. In patients with autonomic failure suffering from PPH, we start treatment with 50 mg of acarbose once a day, 20 minutes before eating their largest meal. The dose can be titrated up to 100 mg three times a day. That said, however, it is important to emphasize that this medication should be used as needed, preferably when patients eat large carbohydrate diets or when they need to be upright after a meal. Acarbose is contraindicated in patients with diabetic ketoacidosis, cirrhosis, inflammatory bowel disease, ulcerative colitis, intestinal obstruction or any chronic intestinal disease that could disrupt digestion or absorption. It is important to monitor serum transaminases within the first 3 months of treatment because acarbose has been associated with liver impairment [10]. In conclusion, postprandial hypotension is an important clinical condition in patients with autonomic failure

XIII. MANAGEMENT OF AUTONOMIC DISORDERS

ACARbosE In THE TREATmEnT of PosTPRAndIAl HyPoTEnsIon

and can cause severe disability. Acute administration of acarbose effectively attenuates the postprandial fall in blood pressure in these patients. Controlled clinical trials assessing the chronic use of acarbose in the treatment of PPH are needed. Nonetheless in a number of clinical observations acarbose can be helpful in patients with autonomic failure and postprandial hypotension.

[4]

[5] [6]

[7]

References [1] Mathias CJ. Postprandial hypotension. Pathophysiological mechanisms and clinical implications in different disorders. Hypertension 1991;18:694–704. [2] Gatt M, MacFie J, Anderson AD, Howell G, Reddy BS, Suppiah A, et al. Changes in superior mesenteric artery blood flow after oral, enteral, and parenteral feeding in humans. Crit Care Med 2009;37:171–6. [3] Onrot J, Goldberg MR, Biaggioni I, Hollister AS, Kingaid D, Robertson D. Hemodynamic and humoral effects of caffeine in

[8]

[9]

[10]

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autonomic failure. Therapeutic implications for postprandial hypotension. N Engl J Med 1985;313:549–54. Hoeldtke RD, Horvath GG, Bryner KD, Hobbs GR. Treatment of orthostatic hypotension with midodrine and octreotide. J Clin Endocrinol Metabol 1998;83:339–43. Luciano GL, Brennan MJ, Rothberg MB. Postprandial hypotension. Am J Med 2010;123:281–6. O'Donovan D, Feinle C, Tonkin A, Horowitz M, Jones KL. Postprandial hypotension in response to duodenal glucose delivery in healthy older subjects. J Physiol-London 2002;540:673–9. Shibao C, Gamboa A, Diedrich A, Dossett C, Choi L, Farley G, et al. Acarbose, an alpha-glucosidase inhibitor, attenuates postprandial hypotension in autonomic failure. Hypertension 2007;50:54–61. Maruta T, Komai K, Takamori M, Yamada M. Voglibose inhibits postprandial hypotension in neurologic disorders and elderly people. Neurol 2006;66:1432–4. Gentilcore D, Bryant B, Wishart JM, Morris HA, Horowitz M, Jones KL. Acarbose attenuates the hypotensive response to sucrose and slows gastric emptying in the elderly. Am J Med 2005;118:1289. Coniff R, Krol A. Acarbose: a review of US clinical experience. Clin Ther 1997;19:16–26.

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135 Erythropoietin in Autonomic Failure Italo Biaggioni Patients who suffer from severe autonomic failure have a high incidence of anemia, which may contribute to their symptoms. Sympathetic failure can contribute to this anemia by impairing erythropoiesis. Recombinant erythropoietin reverses the anemia of autonomic failure, improves upright blood pressure, and may ameliorate symptoms of orthostatic hypotension. There are, however, concerns about the long-term safety of this treatment, and its use should be limited to patients who fail other therapies.

MODULATION OF ERYTHROPOIETIN PRODUCTION BY THE AUTONOMIC NERVOUS SYSTEM Animal studies suggest that the sympathetic nervous system modulates erythropoiesis. The reticulocyte response to acute bloodletting was greatly diminished in rats with denervated kidneys. Intravenous administration of the β-adrenergic agonist salbutamol increased plasma concentrations of erythropoietin-like factor in rabbits and, conversely, β-blockers blunted the erythropoietin response to hypoxia.

THE ANEMIA OF AUTONOMIC FAILURE Patients with severe autonomic failure have an unusually high incidence of anemia; if World Heath Organization criteria are followed (hemoglobin 120 g/L for women and 130 g/L for men), up to 38% of patients are anemic [1]. The anemia of autonomic failure is mild to moderate and is not accompanied by a compensatory reticulocyte response, suggesting an inadequate erythropoiesis. Furthermore, an inappropriately low serum erythropoietin is evident in the patients with lower hemoglobin levels. Lack of sympathetic stimulation may thus result in decreased erythropoietin production and the development of anemia in patients with autonomic failure. In support of this hypothesis, the severity of the anemia correlates with the magnitude of sympathetic impairment. However, it is not certain if the anemia of autonomic failure is solely due to the lack of sympathetic input to erythropoietin-producing cells in the kidney, because acute renal denervation does not affect erythropoietin mRNA upregulation in animals.

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The fact that anemia can be associated with low erythropoietin levels in autonomic failure led to its treatment with recombinant erythropoietin (erythropoiesis-stimulating agents, ESA). This therapy has successfully corrected anemia [1–3], even when used at modest doses (epoetin alpha 25–50 units/kg body weight, subcutaneously, three times a week). ESA also increases supine and upright blood pressure in autonomic failure patients, and may be effective in ameliorating orthostatic hypotension. This pressor response is expected, since it is a well-documented side effect of ESA treatment in chronic renal failure.

RECOMBINANT HUMAN ERYTHROPOIETIN IN THE TREATMENT OF ORTHOSTATIC HYPOTENSION Treatment of orthostatic hypotension remains a challenge in autonomic failure patients and erythropoietin may provide a therapeutic alternative. These patients are very sensitive to volume changes, which explains why fludrocortisone is often the first step in their treatment. Fludrocortisone increases plasma volume only transiently and at the expense of expanding interstitial space; its effectiveness may be explained by potentiation of pressor hormones rather than by volume changes. Treatment with ESA has the theoretic advantage of selectively increasing intravascular volume, and through this mechanism improving venous return and blood pressure. However, ESA increases blood pressure in animals even if correction of anemia is prevented. It is likely, therefore, that other mechanisms contribute to the increase in blood pressure with ESA, including increased sensitivity to the pressor effects of norepinephrine and angiotensin II, increased plasma endothelin levels, enhanced renal tubular sodium reabsorption, impaired nitric oxide production, and increased cytosolic free calcium in vascular smooth muscle [4]. The relevance of these findings to its pressor effect in autonomic failure remains speculative. ESA has consistently improved orthostatic hypotension in the handful of reports that have been published, but long-term, placebocontrolled trials are lacking. In our anecdotal experience, treatment with ESA can be extremely successful, but in many patients there is only modest symptomatic improvement that does not justify the cost and inconvenience of

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the treatment. Furthermore, there are valid safety concerns about long-term treatment with ESA. Studies in renal failure patients have shown increased incidence of stroke in those receiving darbepoetin alpha compared to placebo [5]. Other studies have reported a higher risk of death and cardiovascular complications in patients randomized to have their anemia corrected to a higher hemoglobin target. It is not clear if this is related to hemoconcentration or the fact that higher doses of ESA were needed to reach that target. Indeed, patients resistant to treatment, who required higher doses of ESA to correct anemia, are at greater risk of adverse cardiovascular events [6]. It is not clear if the increased risk is attributable to a dose effect of the drug, but it is reassuring that autonomic failure patients generally require low doses to correct anemia. Given these safety concerns, the use of ESA should be limited to autonomic failure patients who fail other forms of treatment.

References [1] Biaggioni I, Robertson D, Krantz S, Jones M, Haile V. The anemia of autonomic failure: Evidence for sympathetic modulation of erythropoiesis in humans and reversal with recombinant erythropoietin. Ann Int Med 1994;121:181–6. [2] Hoeldtke RD, Streeten DHP. Treatment of orthostatic hypotension with erythropoietin. N Engl J Med 1993;329:611–5. [3] Perera R, Isola L, Kaufmann H. Effect of recombinant erythropoietin on anemia and orthostatic hypotension in primary autonomic failure. Clin Auton Res 1995;5:211–3. [4] Krapf R, Hulter HN. Arterial hypertension induced by erythropoietin and erythropoiesis-stimulating agents (ESA). Clin J Am Soc Nephrol 2009;4:470–80. [5] Pfeffer MA, et al. A trial of darbepoetin alfa in type 2 diabetes and chronic kidney disease. N Engl J Med 2009;361:2019–32. [6] Solomon SD, et al. Erythropoietic response and outcomes in kidney disease and type 2 diabetes. N Engl J Med 2010;363:1146–55.

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136 Somatostatin Agonists Robert Hoeldtke

INTRODUCTION Somatostatin was originally discovered when extracts of rat hypothalamic tissue were shown to inhibit growth hormone secretion by pituitary cells in vitro. Subsequent studies revealed the presence of five distinct somatostatin receptors, each of which is helical, transverses the plasma membrane seven times and is coupled to G protein receptors. The pituitary gland expresses receptors one, two, three and five whereas the first generation somatostatin agonists, octreotide and lantreotide, activate receptors two, three and five. Receptor two is probably the most important in the pituitary, and its activation by somatostatin agonists has revolutionized the treatment of growth hormone excess in patients with acromegaly. Somatostatin agonists have also been useful in the treatment of gastrointestinal neuroendocrine illnesses such as the carcinoid syndrome, vasoactive intestinal peptide secreting tumors (VIPomas), and glucagonomas.

SOMATOSTATIN AND AUTONOMIC NEUROPATHY Our interest in somatostatin and the autonomic nervous system was stimulated by a patient we encountered 25 years ago at Temple University. This 70-year-old white male with a prior history of alcoholism developed recurrent orthostatic syncope which persisted even after he stopped drinking. He complained of impotence and had fecal incontinence. Physical examination did not, however, reveal the usual stigmata of alcohol abuse and there was no evidence of Parkinson’s disease. He had decreased sensation in the feet, and nerve conduction velocity studies confirmed the presence of peripheral neuropathy. Profound orthostatic hypotension (an 85 mmHg drop in systolic blood pressure) occurred almost immediately after he assumed the upright posture, and there was little or no increment in his heart rate when he tried to stand. It was obvious he had autonomic neuropathy, which we attributed to neurotoxic effects of alcohol. We were surprised to discover, however, that he continued to have recurrent syncope even when he was completely supine. Continuous

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00136-0

monitoring of his blood pressure revealed that food ingestion (particularly carbohydrates) was the trigger responsible for the recurrent hypotension and syncope. Even a few slices of bread would trigger a hypotensive response. Robertson et al. at Vanderbilt University had previously reported that many patients with severe autonomic failure become hypotensive after meals, especially breakfast, and postulated that adenosine was the neurohumoral mediator of this response [1]. Blocking this response with caffeine was therapeutic. However, neither caffeine, nor fludrocortisone, nor indomethacin prevented postprandial syncope in our patient and he became increasingly disabled. We were forced to feed him parenterally in order to avoid malnutrition. We discovered that intravenous glucose and other nutrients decreased his blood pressure slightly, but did not cause the severe hypotension which occurred when he ate normally. This led us to postulate that the ingestion of food triggered the release of a vasodilating hormone, most likely a gut peptide that was the mediator of the hypotensive response. We then decided to treat the patient with somatostatin, since it is known to suppress the secretion of multiple gastrointestinal and pancreatic peptides. We discovered that the intravenous infusion of somatostatin temporarily prevented the hypotension associated with glucose ingestion. Although somatostatin also suppressed serum insulin, a known vasodilator, this did not explain the beneficial hemodynamic effects of somatostatin. Fortunately octreotide, which can be given subcutaneously, became available at the very time we discovered the somatostatin effect. Octreotide completely eliminated postprandial hypotension and recurrent syncope in our index case and several other patients, including those with pure autonomic failure and multiple system atrophy (Fig. 136.1). The mechanism of this effect of octreotide is poorly understood. Dudl et al. reported that octreotide stimulated norepinephrine secretion [2]. We found, to the contrary, that it suppressed plasma norepinephrine in patients with pure autonomic failure. We have found it difficult to elucidate a hormonal mediator for octreotide’s effect. Octreotide may be merely acting on the vasculature as a splanchnic vasoconstrictor, from which we infer that the hypothetical vasodilating gut peptide, which prompted us to use somatostatin, may not exist.

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FIGURE 136.1 Effect of octreotide on the blood pressure and plasma norepinephrine response to glucose ingestion in autonomic failure [8]. Patients with pure autonomic failure (n  5) and multiple system atrophy (n  6) were given either octreotide subcutaneously (0.8 microgram/kg) (closed circles) or a placebo (open circles) immediately before ingesting glucose (see hatched area) Drug treatment was different from placebo: *P  0.05; **P  0.01.

Although octreotide prevents postprandrial hypotension and may even increase the blood pressure after breakfast, it does not correct the powerful hypotensive effect of standing suffered by some patients with autonomic neuropathy. We found, however, that pretreatment of patients with midodrine, an alpha adrenergic agonist, potentiated the pressor effect of octreotide. We have described several patients, for example, who had a poor response to midodrine and a poor response to octreotide but, nevertheless, responded nicely to combination therapy. The major shortcoming of octreotide therapy for patients with autonomic neuropathy is that the therapeutic effect only lasts three or four hours, so multiple subcutaneous injections may be required daily. Some patients, however, do well with a single subcutaneous injection with breakfast, which prevents the typical midmorning nadir in blood pressure In order to achieve a more long-lasting pressor effect, we have treated four patients with autonomic neuropathy with octreotide LAR (long-acting release). This agent needs to be given intramuscularly once a month. Two of the patients had Parkinson’s disease and two had pure autonomic failure. One of the patients with Parkinson’s disease had debilitating postprandial and orthostatic hypotension, even more severe than our index case, and typically

had about 100 syncopal episodes a month. Subcutaneous octreotide helped slightly, but octreotide LAR was remarkably effective, and was free of side effects. The therapeutic effect was maintained for the last five years of his life. Two of the three other patients in this series also responded to the octreotide LAR but supine hypertension developed and the risk of this therapy outweighed its benefit.

POSTURAL TACHYCARDIA SYNDROME AND ORTHOSTATIC INTOLERANCE Octreotide has also been used to treat the postural tachycardia syndrome (POTS) and orthostatic syncope. Patients with these disorders are typically tall, slender young women who complain of chronic fatigue and experience dizziness, sweating and palpitations after shifting from the supine to the upright posture. The diagnosis is made if the heart rate increases by more than 30 beats/ minute during the first ten minutes of standing or headup tilt. Streeten observed that subjecting these patients to orthostatic stress led to excessive pooling of blood in the lower extremities and demonstrated the presence of denervation hypersensitivity to norepinephrine in the

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veins of the feet [3]. He interpreted this to signify the presence of occult sympathetic neuropathy specifically in the lower extremities. Jacob et al. accordingly demonstrated that patients with POTS had decreased norepinephrine secretion in the legs [4]. POTS, however, is a very heterogeneous disorder. Some patients have decreased blood flow in the legs (low flow POTS) whereas others are just the opposite (high flow POTS). Moreover, an association between POTS and hyperflexibility (Ehlers–Danlos and Marfan syndromes) has been described. These patients are not known have sympathetic dysfunction yet their hemodynamic response to standing is indistinguishable from that of POTS patients with normal flexibility. Finally, some patients with POTS provide a history of an acute viral illness which triggers the initiation of symptoms. This form of POTS may resolve spontaneously within a few weeks or months. This subset of patients may have an attenuated form of post-viral pandyautonomia. Our interest in orthostatic intolerance was rekindled by recent studies by Tani et al. [5] who have documented splanchnic hypervolemia in patients with POTS and Stewart et al. [6] who have documented that subjecting patients with POTS and orthostatic syncope to head up tilt causes pooling of blood in the splanchnic veins. As discussed above, octreotide is a splanchnic vasoconstrictor which made it a theoretically attractive treatment option. We compared octreotide (0.9 micrograms/kg) with midodrine (10 mg), and found that the two drugs suppressed postural tachycardia and improved orthostatic tolerance equally well (Fig. 136.2). Combination therapy was slightly better than monotherapy in a few patients but we did not see the striking synergism between the two drugs which we had documented in patients with autonomic neuropathy. The major shortcoming of octreotide therapy was its relatively short duration of action, as discussed above. We therefore explored the use of octreotide

LAR. We have given octreotide LAR to seven patients with orthostatic intolerance. Four had POTS and three had orthostatic syncope. Both groups responded to gradually increasing doses of octreotide LAR (Fig. 136.3). Kanjwal et al. have studied octreotide therapy in patients with orthostatic intolerance who have failed multiple other therapies [7]. They confirmed that octreotide therapy suppressed tachycardia and increased the blood pressure in this patient population. Syncope and palpitations were suppressed in about half of the patients.

POSTMENOPAUSAL HOT FLASHES Patients with postmenopausal hot flashes typically experience episodic sensations of warmth in the face, neck, and chest several times a day. The feeling of warmth is often associated with tachycardia, palpitations and sweating. This symptom complex shares similarities with the complaints associated with the postural tachycardia syndrome. This reasoning prompted us to study subcutaneous octreotide in patients with hot flashes. We observed that octreotide therapy suppressed the incidence, duration and intensity of postmenopausal hot flashes in short-term placebo controlled clinical trials. This approach was not practical, however, since patients needed to take four or five injections a day. Octreotide LAR needs to be studied in this population.

ADVERSE EFFECTS OF SOMATOSTATIN ANALOGS Octreotide causes abdominal cramps, nausea and diarrhea in about 50% of patients, and is rarely tolerated by patients with diabetic autonomic neuropathy or

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FIGURE 136.2 Effect of octreotide (0.9 microgram/kg) and midodrine (10 mg) alone or in combination on the heart rates of patients with POTS [9]. The patients stood up at time 0. *No treatment was different from each of the treatments at each minute (P  0.01). **Combination therapy was different from monotherapy for the first 10 minutes (P  0.01).

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hypotension, orthostatic hypotension, postural tachycardia, and orthostatic syncope. Chronic octreotide therapy is expensive, however, and may have unwanted consequences, including cholecystitis, so it should be reserved for patients with severe symptoms that are refractory to more conservative therapies.

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FIGURE 136.3 The effect of chronic octreotide LAR on the heart rate at the end of the therapeutic trial two weeks and four weeks after the maximal dose (30 mg) [10]. The patients stood up at time 0. The closed boxes represent the pretreatment heart rates. The closed circles represent the post-treatment heart rates at the 11th week (2 weeks after they had received 30 mg of octreotide LAR). The open circles represent the post-treatment heart rates at the 13th week (4 weeks after they had received the 30 mg dose). The slope of the heart rate change with time was decreased at both the 11th week (P  0.001) and the 13th week (P  0.001).

underlying gastrointestinal disease. Octreotide suppresses gallbladder emptying, and many patients develop sludge or gallstones during chronic therapy. These are generally asymptomatic but cholecystitis can occur. Postprandial hyperglycemia develops in about 40% of patients taking octreotide chronically. We recommend administering the drug 15 minutes after the beginning of breakfast. Sufficient insulin will be secreted under this circumstances to minimize postprandial hyperglycemia. We have never seen chronic octreotide therapy lead to an elevation in glycosylated hemoglobin concentrations or a diabetes related complication. Octreotide should not be given to patients who cannot tolerate it. Many patients with POTS, for example, have the irritable bowel syndrome, and octreotide worsens that. Similarly, administration of octreotide to patients with inflammatory bowel disease invariably exacerbates their symptoms. Inflammatory bowel disease is an absolute contraindication to octreotide therapy.

[1] Robertson D, Wade D, Robertson RM. Postprandial alterations in cardiovascular hemodynamics in autonomic dysfunctional states. Am J Cardiol 1981;48:1048–52. [2] Dudl RJ, Anderson DS, Forsythe AB, Ziegler MG, O’Dorisio TM. Treatment of diabetic diarrhea and orthostatic hypotension with somatostain analogue SMS-201-995. Am J Med 1987;83:584–8. [3] Streeten DH, Pathogenesis of hyperadrenergic orthostatic hypotension: evidence of disordered venous innervation exclusively in the lower limbs. J Clin Invest 1990;86:1582–8. [4] Jacob G, Costa F, Shannon JR, Robertson RM, Wathen M, Stein M, et al. The neuropathic postural tachycardia syndrome. New Engl J Med 2000;343:1008–14. [5] Tani H, Singer W, McPhee BR, Opfer-Gehrking TL, Haruma K, Kajiyama G, et al. Splanchnic-mesenteric capacitance bed in postural tachycardia syndrome (POTS). Auton Neurosci 2000;86:107–13. [6] Stewart JM, McLeod KJ, Snayal S, Herzberg G, Montgomery LD. Relation of postural vasovagal syncope to splanchnic hypervolemia in adolescents. Circulation 2004;100:2675–81. [7] Kanjwal K, Saeed B, Karabin B, Kanjwal Y, Grubb BR. Use of octreotide in the treatment of refractory orthostatic intolerance. Am J of Ther 2011 2010 Dec. [EPub ahead of Print] [8] Hoeldtke RD, Dworkin GE, Gaspar SE, Israel BC, Boden G. Effect of the somatostatin analogue SMS 210-995 on the adrenergic response to glucose ingestion in patients with postprandial hypotension. Am J Med 1989;86:673–7. [9] Hoeldtke RD, Bryner KD, Hoeldtke M, Hobbs G. Treatment of the postural tachycardia syndrome: a comparison of octreotide and midodrine. Clin Auton Res 2006;16:390–5. [10] Hoeldtke RD, Bryner KD, Hoeldtke M, Hobbs G. Treatment of autonomic neuropathy, postural tachycardia and orthostatic syncope with octreotide LAR. Clin Auton Res 2007;17:334–40.

Further Reading Hoeldtke RD, Horvath GG, Bryner KD, Hobbs G. Treatment of orthostatic hypotension with midodrine and octreotide. J Clin Endo Metab 1998;83:339–43. Hoeldtke RD, Bryner KD, Palmer HC, Stark L, Eddy L, Hobbs G. Effect of Octreotide on postmenopausal hot flushes. Clin Auton Res 2009;19:69–72.

SUMMARY The extraordinary diversity of somatostatin physiology has led to the discovery of new applications for somatostatin analogs. Octreotide can be used to treat several disorders of autonomic function, including postprandial

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137 Harnessing the Autonomic Nervous System for Therapeutic Intervention Murray Esler An important influence of the autonomic nervous system in homeostatic cardiovascular, metabolic and body temperature control is well-established, so it is not surprising that this system has been a target for therapeutic intervention, in many illnesses.

PHARMACOLOGICAL AND SURGICAL TARGETING OF THE AUTONOMIC NERVOUS SYSTEM Autonomic pathophysiology, principally that of the sympathetic nervous system, has been targeted in many illnesses, this system being antagonized or augmented according to the clinical need. Examples of therapeutic sympathetic inhibition are the prescribing of beta-adrenergic blocking drugs in patients with heart failure, in whom ongoing activation of the cardiac sympathetic outflow is harmful, and the use of drugs inhibiting the sympathetic nervous system, adrenergic blocking drugs and centrally acting sympathetic inhibitors, in patients with essential hypertension, in whom activation of the sympathetic outflows to the kidneys, heart and skeletal muscle blood vessels is common. Conversely, augmentation of sympathetic nervous actions is helpful in patients with circulatory control disorders leading to postural hypotension and syncope, with the use of indirect sympathetic agonists, such as ephedrine, and alpha-adrenergic agonists such as midodrine and dihydroergotamine. In other clinical circumstances the use of drugs acting on the autonomic nervous system is not to reverse existing pathophysiology, but to exploit helpful pharmacological drug actions on the autonomic nervous system and its receptors. Examples are the prescribing of beta-adrenergic stimulant inhalants for bronchial asthma, adrenergic agonists for circulatory collapse and shock, and alphaadrenergic blockers for chronic prostatic obstruction. Surgery targeting of the autonomic nervous system also has clinical applications. In the early decades of the 20th century, faced with the high mortality of severe hypertension and the absence of effective pharmacological

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00137-2

therapy, a number of non-selective operations on the sympathetic nervous system were devised in an attempt to lower blood pressure. Notable among these was radical lumbodorsal splanchnicectomy, developed by Smithwick [1], which lowered blood pressure and reduced mortality, but at the cost of often incapacitating side effects. By the 1950s ganglionic blocking drugs had been introduced, this drug class having a mode of action akin to surgical sympathectomy, although reversible. One contemporary application of surgical sympathectomy is laparoscopic thoracic sympathectomy for hyperhidrosis of the palms and face.

“HARNESSING” THE AUTONOMIC NERVOUS SYSTEM A recent initiative, differing from the above, has been the development of devices for the management of drugresistant hypertension [2,3]. These modify autonomic blood pressure control systems reversibly with one device [2], and permanently with the other [3]. But first we must consider the neural pathophysiology of essential hypertension, which underpinned the development of these two antiadrenergic antihypertensive devices.

Activation of the Sympathetic Nervous System in Essential Hypertension Application of sympathetic nerve recording and norepinephrine spillover methodology has demonstrated activation of sympathetic nervous outflows to the kidneys, heart and skeletal muscle vasculature, typically a doubling or trebling overall, in patients with essential hypertension [4,5]. The syndrome of neurogenic essential hypertension appears to account for no less than 50% of all cases of high blood pressure. This estimate is based on both the proportion of patients with essential hypertension who have demonstrable sympathetic excitation, and the number in whom substantial blood pressure lowering is achieved with antiadrenergic drugs or devices, described below.

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Does this sympathetic activation cause the blood pressure elevation? Once it was thought that the sympathetic nervous system exerts minute by minute circulatory control only, and was not of importance in the pathogenesis of hypertension. The regulatory effects of the renal sympathetic nerves on renin release, glomerular filtration rate and renal tubular reabsorption of sodium are, however, now seen to provide a range of potential hypertension-producing mechanisms [6]. Given that antihypertensive drugs antagonizing the renin-angiotensin system are the dominant therapy, how can a case for pre-eminence of sympathetic neural origins of human hypertension be sustained? Drug prescribing practices in hypertension, however, manifestly do not prove pathogenesis. In any era, the drug class used most widely for an illness commonly dictates which research stream is followed for the illness (especially if the drug patents have not expired!), determining the prevailing notions of pathophysiology. Despite the current dominance in therapy of drugs antagonizing the renin -angiotensin system, plasma renin levels in essential hypertension are often low, when plasma renin activity is high this typically has a neural mechanism, high sympathetic outflow to the kidneys stimulating renin release, and the clinical case made for specific therapeutic benefit with renin-angiotensin inhibitors has been claimed to have, in fact, been overstated [4].

Drug-Resistant Hypertension Successful treatment of hypertension has proven to be difficult, perhaps surprising given the availability of multiple antihypertensive drug classes, and the blood pressure lowering value of strategies to eliminate adverse lifestyle influences. In approximately 50% of hypertensive patients treatment goals are not met, despite apparently the best efforts of doctors and their patients. Drug-resistant hypertension is so common as to suggest that there may be multiple system failures in therapy: intrinsic limitations to existing antihypertensive pharmaceuticals, a sometimes high level of physician inertia, and reluctance of patients to adhere life-long to drug treatment. Perhaps the underlying pathophysiology is refractory to the most commonly used antihypertensive drugs, renin-angiotensin system antagonists, dihydropyridine calcium channel blockers and diuretics. The national and international society guidelines for treating hypertension do typically relegate antiadrenergic drugs to the third or fourth tier.

Anti-Adrenergic Devices for Treating Hypertension The sympathetic nervous system is the “forgotten pathway” in hypertension treatment. This may soon change, with the recent testing of two device-based

antiadrenergic therapies, the surgically implantable arterial barostimulator [2], and the radiofrequency renal nerve ablation catheter [3,7]. The implantable barostimulator In the setting of sustained elevation of blood pressure, the arterial baroreflex quickly resets. The set point for the reflex becomes the higher, prevailing blood pressure, so that there is no baroreflex drive operating to restore the normal pressure. In experimental models of hypertension, however, activation of central baroreflex pathways by electrical stimulation of carotid sinus nerves reduces sympathetic outflow from the central nervous system and lowers blood pressure, this effect persisting for weeks without adaptation [8]. Baroreflex stimulation devices have been developed for the treatment of patients with hypertension, and are currently undergoing clinical testing. The Rheos implantable carotid sinus stimulator (CVRx, Minneapolis, MN, USA) has been studied in patients with severe hypertension refractory to drug therapy [2,9]. In the implantation procedure both carotid sinuses are surgically exposed and electrodes placed bilaterally around the carotid adventitial surfaces. The leads are run subcutaneously to an implantable stimulation device placed subcutaneously on the anterior chest wall. Electrical baroreflex activation is then initiated on both carotid sinuses simultaneously, with incremental stimulation voltage increases until a tolerable chronic level of stimulation level is achieved. Short-term sympathetic inhibition and lowering of blood pressure is clearly evident [2]. Studies are currently being conducted to evaluate the efficacy and safety of the procedure, and to identify the hypertensive population who might receive greatest benefit. To this point the device has been used for the treatment of refractory hypertension only. A large-scale randomized study is in progress, with all patients having the device implanted, but in 50% without baroreflex stimulation. A recent preliminary report [9] has indicated that the key efficacy endpoint had not been met; the trial has been extended, with longer follow-up. The future of this procedure is therefore uncertain. Further evaluation will establish whether blood pressure lowering benefit is sufficient to outweigh the cost and invasive nature of the procedure.

Endovascular Renal Sympathetic Nerve Ablation The other revolutionary treatment principle involves ablation of the renal sympathetic nerves with a radiofrequency emitting catheter inserted percutaneously into the femoral artery in the groin, and advanced to lie, in turn, in the lumen of both renal arteries [3,7]. Sympathetic nerves enter the human kidneys in the walls of the renal arteries, within reach of ablative energy delivery (Fig. 137.1). In earlier times, prior to the availability of antihypertensive drugs, extensive surgical sympathectomy was used as a treatment of severe hypertension [1]; survival benefit was

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FIGURE 137.1 As the renal sympa-

Anatomical Location of Renal Sympathetic Nerves

thetic nerves pass from the sympathetic chain to the kidneys they lie in the wall of the renal arteries, where they are within reach of radiofrequency energy delivery from a radiofrequency ablation catheter in the artery lumen.

Arise from T10-L1 Follow the renal artery to the kidney Primarily lie within the adventitia Vessel Lumen

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demonstrated but complication rates were high, as was morbidity from the extensive denervation, which did not specifically target the kidneys. In many experimental models of hypertension, the sympathetic outflow to the kidneys is activated, and renal sympathectomy typically prevents the development of the hypertension [6]. Initiation of the new treatment strategy for hypertension was based on these experimental observations, and the demonstration that the renal sympathetic outflow is activated in essential hypertension [4]. For entry into the recently completed randomized trial [7], patients had to meet international criteria of uncontrolled essential hypertension (clinic blood pressure in excess of 160/90 mmHg on three or more drugs). The catheter is inserted percutaneously via the femoral artery to lie in the renal artery lumen, and radiofrequency energy is delivered in 90° quadrants in stepwise fashion to the full circumference of both renal arteries. Both efferent and afferent renal nerves traverse the renal arteries, lying primarily in the adventitia (Fig. 137.1). Preliminary experimental studies in pigs demonstrated that radiofrequency energy emission in the renal artery lumen, at sufficient and optimized level, could selectively ablate the renal nerves, which are more thermally sensitive than the remainder of the artery wall. The aims in the randomized trial [7] and a pilot study [3] were to establish that the procedure does produce renal sympathectomy in humans, that it is safe, and that blood pressure is lowered. All have been confirmed. To establish whether the catheter ablates renal sympathetic nerves, measurements of renal norepinephrine spillover were made at baseline and at follow-up. These measurements indicate that sympathetic denervation does, in fact, occur [3]. The mean level of blood pressure reduction achieved

–30 –40 –50 –60 –70

FIGURE 137.2 Reductions in systolic and diastolic blood pressure in relation to time elapsed since the renal denervation procedure. The patients shown are those participating in the initial denervation pilot study [3], with additional patients recruited subsequently. The blood pressure lowering effect of renal denervation is fully maintained through 18 and 24 months of follow-up. The numbers of patients evaluated at each time point is indicated.

in the randomized trial was 32/12 mmHg at 6 months (P  0.001). Longer follow-up, up to two years, has been possible in patients participating in the pilot study. The blood pressure reduction is durable, not diminishing over time (Fig. 137.2), suggesting that renal sympathetic reinnervation, if it has occurred, is insufficient to cancel out the blood pressure lowering benefit.

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Uncertain at present is the importance of destruction of renal afferent nerves in the antihypertensive effect achieved by the RF ablation procedure. An unexpected observation with the procedure is that sympathetic outflow from the CNS is reduced, evident in reduced wholebody norepinephrine spillover measured by isotope dilution and lowering of sympathetic nerve traffic to the skeletal muscle vasculature measured by microneurography [10]. There is conclusive evidence that renal afferent nerves exist, and that their projections to the hypothalamus can stimulate sympathetic outflow [11]. This CNS input from renal afferent nerves is critical in producing both the sympathetic activation and hypertension found in patients with end-stage renal disease. It is probable that RF deafferentiation of the kidney in the patients with previously resistant hypertension studied in the reported trial contributed to the blood pressure lowering observed. A cure for essential hypertension? It has been suggested that renal artery catheter-based renal denervation might, perhaps, provide a cure for essential hypertension in selected patients, those with milder hypertension than treated in the recent study. This speculation remains untested. For the procedure to be applied in milder forms of essential hypertension a very high level of safety would be mandatory, which does seem to apply. Some efferent sympathetic nerve re-growth is probable, although the degree to which this would fully restore sympatheticallymediated function in the kidneys, and perhaps cancel out the observed antihypertensive effect, is problematic. It is now certain that the radiofrequency procedure ablates renal afferent nerves [10], and that this, by inhibiting systemic sympathetic outflow, contributes to the blood pressure lowering observed. Regeneration of renal afferent

nerves is does not occur. Any blood pressure reduction attributable to renal differentiation is likely to be permanent.

References [1] Smithwick RH, Bush RD, Kinsey D, Whitelaw GP. Hypertension and associated cardiovascular disease; comparison of male and female mortality rates and their influence on selection of therapy. J Am Med Assoc 1956;160:1023–6. [2] Mohaupt MG, Schmidli J, Luft FC. Management of uncontrollable hypertension with a carotic sinus stimulation device. Hypertension 2007;50:825–8. [3] Krum H, Schlaich MP, Whitbourn R, Sobotka P, Sadowski J, Bartus K, et al. Catheter-based renal sympathetic denervation for resistant hypertension: a multicentre safety and proof-of-principle cohort study. The Lancet 2009;373:1275–81. [4] Esler M, Lambert E, Schlaich M. Point: Counterpoint. Chronic activation of the sympathetic nervous system is the dominant contributor to systemic hypertension. J Appl Physiol 2010;109:1996–8. [5] Grassi G, Colombo M, Seravalle G, Spaziani D, Mancia G. Dissociation between muscle and skin sympathetic nerve activity in essential hypertension, obesity, and congestive heart failure. Hypertension 1998;31:64–7. [6] DiBona GF, Kopp UC. Neural control of renal function. Physiol Rev 1997;77:75–197. [7] Symplicity HTN-2 Investigators Renal sympathetic denervation in patients with treatment-resistant hypertension (The Symplicity HTN-2 Trial): a randomized controlled trial. Lancet 2010;376:1903–9. [8] Navaneethan SD, Lohmeier TE, Bisignano JD. Baroreflex stimulation: A novel treatment for resistant hypertension. J Am Soc Hypertens 2009;3:69–74. [9] Nainggolan L. Future of Rheos system uncertain: company in discussion with FDA, June 21, 2010. Available at: http://www.theheart .org/article/1089587.co/. [accessed 18.08.10]. [10] Schlaich MP, Sobotka PA, Krum H, Lambert E, Esler MD. Renal sympathetic nerve ablation for treatment of uncontrolled hypertension. N Engl J Med 2009;361:932–4. [11] Campese VM, Kogosov E. Renal afferent denervation prevents hypertension in rats with chronic renal failure. Hypertension 1995;25:878–82.

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138 Acupuncture Regulation of Cardiovascular Function John C. Longhurst INTRODUCTION Acupuncture, practiced for over 3000 years in China [1], is an important component of Traditional Chinese Medicine (TCM) and frequently includes herbal therapy. Traditional physicians use acupuncture in a variety of disease conditions, which they describe as either having a deficiency or excess of Qi (pronounced “Chi”), their term for energy that flows through meridians and associated Chinese organ systems. Chinese organs named similarly but not anatomically identical to the Western equivalent include the heart, pericardium, lung, spleen, kidney, liver, gallbladder, stomach, small and large intestine, bladder and triple burner or tri heater (no Western correlate). The use of acupuncture spread to other countries in Asia, including Korea and Japan, and more recently to the United States and Europe. In practice, acupuncture is applied at one or more acupuncture points (also called acupoints) located along meridian channels using thin (~32 Ga) needles that are inserted, left in place without adjustment, manipulated manually, or stimulated electrically during electroacupuncture (EA). There is no documented anatomical substrate for meridians. In fact, methods used to detect meridians, and the acupoints, for example using skin resistance, have not been reliably demonstrated [2]. Thus, the meridian system is best considered as a roadmap directing practitioners where best to stimulate somatic regions of the body to achieve optimal clinical responses. The actual practice of acupuncture varies from country to country and from physician to physician, but generally one acupuncture point, or more commonly a combination of acupoints, is stimulated to restore circulation of Qi through the meridians and their associated organ systems. As alternatives to needling, acupressure or heat (called moxibustion) is applied to acupoints. In addition to the classical twelve meridians, ear or auricular acupuncture, developed in China and refined in France, is frequently used in treatment.

WESTERN UNDERSTANDING OF ACUPUNCTURE Many, if not all, meridians and associated acupoints are located over major peripheral somatic mixed nerve

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00138-4

pathways containing both afferent and efferent fibers that are stimulated by acupuncture. Stimulated action potentials are transmitted by afferent nerve fibers [3,4] to the central nervous system (CNS) where the sensory information is processed in a number of regions concerned with processing pain and cardiovascular input. Studies demonstrate that acupuncture lowers elevated blood pressure, for example during exercise and in patients with mild to moderate hypertension [5,6,7]. Acupuncture applied to normotensive human subjects or experimental preparations does not alter blood pressure [5,8]. Conversely, in several models of experimental hypotension, for example with vasodepressor reflex stimulation or following hemorrhage, acupuncture appears to be able to elevate blood pressure when it is low [9]. Thus, consistent with the Oriental philosophy of homeostasis, acupuncture’s main role is to normalize blood pressure, suggesting a potential role for this therapy in clinical conditions associated with abnormalities of blood pressure, including both high and low blood pressure. Low frequency electroacupuncture (2–6 Hz) stimulates the release of both excitatory and inhibitory neurotransmitters specific to each brain nucleus (Fig. 138.1) [10]. For example, in the rostral ventrolateral medulla (rVLM), which regulates activity of sympathetic premotor neurons, opioids are functionally important since naloxone reduces the acupuncturerelated reduction in demand-induced myocardial ischemia (Fig. 138.2). High frequency EA (100 Hz) also leads to the release of opioids, particularly dynorphin [11]. Low frequency EA is useful in treatment of pain and cardiovascular disease, although there also may be role for high frequency EA in pain.

NEUROLOGICAL SUBSTRATE Although acupuncture is not a painful stimulus, approximately {2/3} of fibers activated during stimulation are finely myelinated (Group III) and {1/3} are unmyelinated (Group IV); both groups appear to be essential to the EA-cardiovascular response [3,4]. Most patients report a sensation of heaviness, tingling, numbness, fullness, slight burning or mild pain that is called Deqi by

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138. ACuPunCTuRE REgulATIon oF CARdIovAsCulAR FunCTIon

ARC Hypothalamus Glu Β-End ACh

vlPAG

ARC

vlPAG

(+)

Endocannabinoids

Midbrain

(–)

vlPAG P5-P6

GABA

(+)

S36-S37

Glu

Groups III+IV

Medulla NR rVLM (–)

rVLM

(+)

vlPAG Enk B-End

IML

(–)

5HT

Glu GABA Nociceptin

N Raphe Glu

rVLM

FIGURE 138.1 Diagram of neural pathways and regions in central nervous system through which electroacupuncture (EA) modifies sympathetic outflow and cardiovascular function. EA at pericardial (P5 and 6) and stomach or S36 and 37, located over the median and deep peroneal somatic nerves, respectively, attenuates reflex-induced elevations in blood pressure. Somatic afferents stimulated by EA enter the spinal cord through the dorsal horn and ascend through polysynaptic connections to the hypothalamus, midbrain and medulla. Processing occurs in the arcuate nucleus (ARC) in the hypothalamus, midbrain ventral lateral periaqueductal gray (vlPAG) and rostral ventral lateral and raphé regions in the medulla (rVLM and NR). A number of neurotransmitter systems, specific to each nucleus (see text) contribute to acupuncture’s ability to modulate sympathetic outflow to the heart and vascular system from the intermediolateral columns in the thoracic spinal cord. Other abbreviations: Glu, glutamate; β-End, β-endorphin; ACh, acetylcholine; GABA, gamma amino butyric acid; Enk, enkephalin, 5HT, 5-hydroxytryptamine or serotonin; , excitation; , inhibition. (Modified with permission from Li, P. and Longhurst, J.C. Neural mechanism of electroacupuncture’s hypotensive effects. Auton Neurosci. 2010;157:24–30.)

TCM physicians. Sensations evoked during low frequency, low intensity EA represent a sensory paresthesia, signify the potential for a beneficial therapeutic response and provide strong evidence that a sensory neural component is essential for acupuncture’s clinical action [2]. Manual acupuncture involving needle manipulation without electrical stimulation and low frequency EA similarly stimulate somatic Group III and IV afferent nerve fibers and similarly lower elevated blood pressure [12].

Anatomical information from immunohistochemical studies employing light and florescent as well as confocal microscopy during acupuncture-induced somatic sensory nerve stimulation demonstrate a potential role for a number of regions in the central nervous system, including the arcuate nucleus in the ventral hypothalamus, the ventrolateral periaqueductal gray (vlPAG) in the midbrain and the nucleus raphé pallidus and rostral ventrolateral regions in the medulla [10]. Neurons in these nuclei

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nEuRologICAl subsTRATE

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FIGURE 138.2 Reflex increases in arterial blood pressure and regional myocardial function following visceral reflex stimulation (arrows). Regional function was measured with a single crystal sonomicrometer system that assessed wall thickening (WTh), measured online and expressed as percent WTh. Cardiovascular function was assessed during normal coronary artery flow conditions (a), following occlusion of a diagonal branch of the left anterior descending coronary artery to create ischemia (b), 30 minutes of bilateral electroacupuncture (EA) during ischemia at the P5 and P6 acupoints, situated over the median nerve (c) and following intravenous administration of naloxone (d). Naloxone was administered 5–10 minutes after termination of EA, at a time when EA significantly inhibits ischemic responses. Since naloxone reversed the effect of EA, an opioid mechanism underlies EA’s action to reduce the demand-induced myocardial ischemia. (Reproduced with permission from Chao, D.M., Shen, L.L., Tjen-A-Looi, S.C., Pitsillides, K.F., Li, P. and Longhurst, J.C. Naloxone reverses inhibitory effect of electroacupuncture on sympathetic cardiovascular reflex responses. Am J Physiol. 1999;276:H2127–134.)

process somatic input through the action of a number of excitatory and inhibitory neurotransmitters. Thus, these regions and associated neurotransmitter systems underlie acupuncture’s action on sympathetic outflow and hence cardiovascular function. Electrophysiological recordings of neural activity in each of these regions combined with pharmacological manipulation, microdialysis and high performance liquid chromatography to assess extracellular concentrations have confirmed the importance of several neurotransmitters in the long-loop pathway extending from the hypothalamus through the midbrain to medullary regions to ultimately regulate activity of sympathetic premotor fibers in the rVLM. In addition to opioids, inhibitory neurotransmitters that may play a role in the action of low frequency EA in the rVLM include gamma amino butyric acid (GABA), nociceptin and serotonin [10,13,14], while the excitatory neurotransmitter glutamate underlies hypotensive responses to acupuncture in the hypothalamic arcuate nucleus, vlPAG in the midbrain

and the nucleus raphé pallidus in the medulla [10,15]. Furthermore, acetylcholine in the arcuate nucleus serves as an excitatory neurotransmitter and endocannabinoids in the vlPAG presynaptically reduce GABA release to disinhibit activity during EA-induced sympathoinhibition [10,16,17]. Lastly, the dorsal horn and intermediolateral columns in the thoracic spinal cord, where somatic afferent information feeds into the CNS and where sympathetic motor nerves exit, also appear to be sites of cardiovascular regulation during EA [18]. Thus, acupuncture’s action to lower blood pressure is complex and involves a number of cardiovascular centers and neurotransmitter systems in the brainstem and spinal cord that process somatic evoked input and sympathetic outflow. While acupuncture lowers elevated blood pressure by inhibiting sympathoexcitation, its ability to elevate low blood pressure may involve actions on both sympathetic and parasympathetic nuclei in the brainstem. These studies are ongoing and require further validation.

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CLINICAL ROLE OF ACUPUNCTURE Acupuncture has a number of important attributes. The first is its relative point specificity (Fig. 138.3). Some acupoints, like P5 and P6 (referring to the pericardial meridian) located over the median nerve or S36 and S37 (referring to the stomach meridian) located over the deep peroneal nerve exert strong sympathoinhibitory responses, whereas others like LI6 and L7 (referring to large intestine and lung meridians), located over the superficial radial nerve have little or no cardiovascular influence [19]. The difference between these acupoints appears to be due to hard wiring of the system. In this regard, P5, P6, S36 and S37 provide strong input to cardiovascular centers like the rVLM in the brainstem, whereas L6 and L11 evoke little response in this region, and hence have little influence on cardiovascular function. A second unique aspect of acupuncture applied for 15–45 min, and one that differentiates it from brief somatosensory stimulation, is its slow onset and, more importantly its prolonged action. Experimental studies show

that 30 min of low intensity, low frequency EA reduces elevated blood pressure and myocardial ischemia, beginning after 10–15 min and lasting for up to 90 min [3]. Acupuncture’s cardiovascular actions can last for 10–12 hours in unanesthetized experimental studies and when applied repetitively on a weekly basis to patients with mild to moderate hypertension, hypotensive responses can be maintained for up to four weeks after an eight week course of treatment [6,10]. This prolonged action of acupuncture on blood pressure is due to several factors. First, involvement of the long-loop hypothalamic-midbrain-medullary pathway is critical. Reinforcing pathways between the vlPAG in the midbrain and the arcuate nucleus in the hypothalamus help to prolong its action for minutes or hours [10]. Additionally, repetitive acupuncture appears to “turn on” the genome in regions like the rVLM leading to enhanced and prolonged production of modulatory neurotransmitters precursors like preproenkephalin [20]. Thus, the clinical benefit of acupuncture is prolonged by appropriate (low frequency application for a number of minutes) and repeated application. FIGURE 138.3 Somatic regions identified as acupoints, designated according to the meridian (m), that have strong or weak influence on cardiovascular function, including increased and decreased blood pressure and myocardial ischemia. Acupoints exhibiting strong cardiovascular effects are designated active, while those demonstrating weak cardiovascular influence can be used as control acupoints. See text for discussion of meridians and point specific actions of acupuncture. Abbreviations: L, P, G, S, LI and H refer to lung, pericardium, gallbladder, stomach, large intestine and heart meridians. (Reproduced with permission from Li, P. and Longhurst, J.C. Neural mechanism of electroacupuncture’s hypotensive effects. Auton Neurosci. 2010;157:24–30.)

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ouTsTAndIng IssuEs In ACuPunCTuRE REsEARCH

OUTSTANDING ISSUES IN ACUPUNCTURE RESEARCH Studies of acupuncture present a number of challenges. Although we understand a great deal about this treatment modality, much remains to be learned about mechanisms by which acupuncture influences autonomic outflow and hence cardiovascular function. The system is complex, involving a number of neurotransmitters/neuromodulators and regions of the brain and spinal cord, with many interactions likely and with different regions playing a role depending upon the underlying condition as well as the specific acupoint and stimulus modality input. Most acupuncturists perform individualized therapy involving combinations of acupoints that vary between practitioners [1,11]. Point specificity, or the ability of specific acupoints or a combination of acupoints to cause selected responses needs further study. Another area of concern by many scientists and practitioners is that the acupuncture response may simply represent a placebo effect that is not much greater than the response to sham intervention [21]. In fact, placebo responses can occur in 30–40% of treated subjects and acupuncture leads to beneficial response in only 70–80% of cases, suggesting a narrow difference in response rate between the two treatments. Furthermore, placebo responses, like acupuncture, involves endogenous opiates [22]. This common underlying mechanism shared between acupuncture and placebo may explain why so many studies show that verum acupuncture is little better than a sham control [23]. However, an important issue to consider in any trial is the adequacy of the sham group. Control interventions employing stimulation outside the meridian system or a non-penetrating needle surrogate may not be as advantageous as stimulating inactive acupoints or placing a needle in the active acupoint that is not stimulated [12]. A final issue to address with acupuncture is the difficulty of blinding the subject and the acupuncturist [11]. Experimental studies are not complicated by this latter problem since anesthesia is employed. However, future clinical trials will need to consider each of these issues. Supported by NIH grants HL063313 and HL072125.

References [1] Longhurst J. Acupuncture’s beneficial effects on the cardiovascular system. Prev Cardiol 1998;1:21–33. [2] Longhurst JC. Defining meridians: A modern basis of understanding. J Acupunct Meridian Stud 2010;3:67–74. [3] Li P, Pitsillides K, Rendig S, Pan HL, Longhurst J. Reversal of reflexinduced myocardial ischemia by median nerve stimulation: A feline model of electroacupuncture. Circulation 1998;97:1186–94.

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[4] Tjen-A-Looi S, Fu LW, Zhou W, Longhurst JC. Role of unmyelinated fibers in electroacupuncture cardiovascular responses. Auton Neurosci 2005;118:43–50. [5] Li P, Ayannusi O, Reed C, Longhurst J. Inhibitory effect of electroacupuncture (EA) on the pressor response induced by exercise stress. Clin Auton Res 2004;14:182–8. [6] Longhurst JC. Acupuncture in cardiovascular medicine. In: O’Hara T, editor. Integrative cardiology. : Oxford University Press; 2010. pp. 100–116. [7] Flachskampf FA, Gallasch J, Gefeller O, Gan J, Mao J, Pfahlberg AB, et al. Randomized trial of acupuncture to lower blood pressure. Circulation 2007;115:3121–9. [8] Li P, Rowshan K, Crisostomo M, Tjen-A-Looi S, Longhurst J. Effect of electroacupuncture on pressor reflex during gastric distention. Am J Physiol 2002;283:R1335–R1345. [9] Syuu Y, Matsubara H, Hosogi S, Suga H. Pressor effect of electroacupuncture on hemorrhagic hypotension. Am J Physiol 2003;285:R1446–R1452. [10] Li P, Longhurst JC. Neural mechanism of electroacupuncture’s hypotensive effects. Auton Neurosci-Basic 2010;157:24–30. [11] Ernst E, White A. Acupuncture: A scientific appraisal, 2nd ed. Oxford: Butterworth Heinemann; 1999. [12] Zhou W, Fu LW, Tjen-A-Looi SC, Li P, Longhurst JC. Afferent mechanisms underlying stimulation modality-related modulation of acupuncture-related cardiovascular responses. J Appl Physiol 2005;98:872–80. [13] Tjen-A-Looi SC, Li P, Longhurst JC. Role of medullary GABA, opioids, and nociceptin in prolonged inhibition of cardiovascular sympathoexcitatory reflexes during electroacupuncture in cats. Am J Physiol 2007;293:H3627–H3635. [14] Moazzami A, Tjen-A-Looi SC, Longhurst JC. Serotonergic projection from nucleus raphe pallidus to rostral ventrolateral medulla modulates cardiovascular reflex responses during acupuncture. J Appl Physiol 2010;108:1336–46. [15] Li P, Tjen-A-Looi SC, Longhurst JC. Nucleus raphé pallidus participates in midbrain-medullary cardiovascular sympathoinhibition during electroacupuncture. Am J Physiol 2010;299:R1369–R1376. [16] Tjen-A-Looi S, Li P, Longhurst JC. Processing cardiovascular information in the vlPAG during electroacupuncture in rats: Roles of endocannabinoids and GABA. J Appl Physiol 2009:1793–9. [17] Fu LW, Longhurst JC. Electroacupuncture modulates vlPAG release of GABA through presynaptic cannabinoid CB1 receptor. J Appl Physiol 2009;106:1800–9. [18] Zhou W, Mahajan A, Longhurst JC. Spinal nociceptin mediates electroacupuncture-related modulation of visceral sympathoexcitatory reflex responses in rats. Am J Physiol 2009;297:H859–65. [19] Tjen-A-Looi SC, Li P, Longhurst JC. Medullary substrate and differential cardiovascular response during stimulation of specific acupoints. Am J Physiol 2004;287:R852–62. [20] Li M, Tjen-A-Looi SC, Longhurst JC. Electroacupuncture enhances preproenkephalin mRNA expression in rostral ventrolateral medulla of rats. Neurosci Lett 2010;477:61–5. [21] Mayer DJ. Acupuncture: An evidence-based review of the clinical literature. Ann Rev Med 2000;51:63. [22] Riet G, Craen A, Boer A, Kessels A. Is placebo analgesia mediated by endogenous opioids? A systemic review. Pain 1998;76:273–5. [23] Langevin H, Wayne PM, MacPherson H. Paradoxes in acupuncture research: Strategies for moving forward. Evid Based Complement Alternat Med 2011  http://www.ncbi.nlm.nih.gov/pmc/articles/ PMC2957136/pdf/ECAM2011-180805.pdf/ 

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139 Bionic Baroreflex Takayuki Sato, André Diedrich, Kenji Sunagawa A novel therapeutic strategy against central baroreflex failure is proposed based on bionic technology. The bionic baroreflex system is a recent innovation designed to revitalize baroreflex function. In the bionic baroreflex system, arterial pressure is sensed via a micromanometer placed in the aortic arch. Its output is fed into a computer that functions as an artificial vasomotor center. Based upon measured changes in arterial pressure, the artificial vasomotor center generates command signals to an electrical stimulator to provide an appropriate frequency of stimulation to vasomotor sympathetic nerves. Although the bionic baroreflex system is not currently available for clinical practice, its clinical use is expected after further development.

INTRODUCTION The arterial baroreflex is the most important negative feedback system to suppress rapid daily disturbances in arterial pressure [1]. Therefore, in patients with autonomic failure with dysfunctional baroreflex control of arterial pressure, the simple act of standing would cause a fall in arterial pressure, reducing perfusion of the brain, and resulting potentially in loss of consciousness. The functional restoration of the arterial baroreflex is essential in numerous patients groups (e.g., those with autonomic failure) for maintaining consciousness and a level of life quality. In patients with central baroreflex failure such as baroreceptor deafferentation, Shy–Drager syndrome, and spinal cord injuries, peripheral sympathetic nerves remain functional but are not controlled by the brain. A novel therapeutic strategy has been proposed to use a bionic baroreflex system (BBS) with a neural interface to control arterial pressure [2,3]. A bionic system is an artificial device for the functional replacement of a failed physiological system. It is should be able to mimic its static and dynamic characteristics. In the proposed BBS (Fig. 139.1), arterial pressure is sensed via a micromanometer placed in the aortic arch, and fed into a computer that functions as an artificial vasomotor center. Based upon measured changes in arterial pressure, the artificial vasomotor center

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00139-6

generates commands signals that trigger an electrical stimulator to provide a stimulus of the appropriate frequency to vasomotor sympathetic nerves. The BBS has been able to revitalize baroreflex function in an animal model of central baroreflex failure.

BIONIC BAROREFLEX SYSTEM Theoretical Background It is of critical importance to identify the algorithm of the artificial vasomotor center, i.e., how to determine the stimulation frequency (STM) of the vasomotor sympathetic nerves in response to a change in arterial pressure (AP). Based on expertise of bionics and systems physiology, the algorithm has been determined as a transfer function by using a white-noise identification method [2]. First, the functional characteristics have to be mimicked by the BBS, i.e., the open-loop transfer function of the native baroreflex

FIGURE 139.1 Central baroreflex failure and its functional replacement by a bionic baroreflex system.

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139. BIonIC BARoREflEx

(A) Native Baroreflex

HNative

To operate in real time as the artificial vasomotor center, the computer was programmed to automatically calculate instantaneous STM in response to instantaneous AP changes according to a convolution algorithm [2,3]:

AP

Pd

(B) Bionic Baroreflex HAP→STM

Implementation of Algorithm of Artificial Vasomotor Center in BBS

Pd

HSTM→AP

AP

FIGURE 139.2 Block diagrams of native and bionic baroreflex systems. HNative denotes the open-loop transfer function of the native baroreflex system. HAP→STM and HSTM→AP are the open-loop transfer functions from arterial pressure (AP) to the frequency of electrical stimulation (STM) and from STM to AP, respectively. Pd is an external disturbance in pressure. (Modified from Sato et al. (2002) with permission of American Heart Association.)

STM(t )

∞

∫0

h( ) ⋅ AP(t

)d

where h(t) is an impulse response function computed by an inverse Fourier transform of HAP→STM.

Efficacy of BBS In a prototype of the BBS for rats with central baroreflex failure, the celiac ganglion was selected as the sympathetic vasomotor interface [3]. The efficacy of the BBS against orthostatic hypotension during head-up tilting (HUT) is shown in Figure 139.3. Without the activation of the BBS, HUT produced a rapid progressive fall in AP by 40 mmHg in 2 seconds. In contrast, while the BBS was

FIGURE 139.3 Real-time operation of the bionic baroreflex system during head-up tilting (HUT) in the rat. In a model of central baroreflex failure (broken line), when the bionic baroreflex system was inactive, arterial pressure (AP) fell rapidly and severely immediately after HUT. On the other hand, while the bionic baroreflex system was activated (thick line), such an AP fall was buffered, which was comparable to the native baroreflex (thin line). While sensing changes in AP, the bionic baroreflex system automatically computes the frequency of electrical stimulation (STM) of the sympathetic nerves and drives a stimulator. (Modified from Sato et al. (2002) with permission of American Heart Association.)

(Hnative) is identified in normal subjects (Fig. 139.2). Second, the open-loop transfer function of the AP response to STM (HSTM→AP) is determined in patients with central baroreflex failure. Finally, a simple estimation process, HNative/ HSTM→AP, is used to yield the transfer function required for the artificial vasomotor center of the BBS, i.e., HAP→STM.

FIGURE 139.4 The response of arterial pressure (AP) to spinal cord stimulation at Th9-11 (A) and the step response estimated from the transfer function analysis (B).

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EPIduRAl CATHETER APPRoACH foR HumAn BBS

activated, it automatically computed STM and appropriately stimulated the sympathetic nerves to quickly and effectively attenuate the AP drop. Such an AP response to HUT during the real-time execution of the BBS was indistinguishable from that observed in a control rat with an intact baroreflex system. Therefore, the BBS was considered to revitalize the native baroreflex function.

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EPIDURAL CATHETER APPROACH FOR HUMAN BBS To apply BIONIC technology to patients, we need a neural interface with quick and effective controllability of AP in humans. Here, we proposed an epidural catheter approach for the human BBS [4]. We percutaneously placed an epidural catheter with a pair of electrodes at the

FIGURE 139.5 Clinical application of the bionic baroreflex system to a 55-year-old man with high cervical cord injury during head-up tilting (HUT). (A) Control; (B) Real-time operation of the bionic baroreflex system. STM, stimulation frequency; AP, arterial pressure.

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level of Th9-11, and then we randomly altered the stimulation frequency between 0 and 20 Hz (Fig. 139.4A). The step response computed by the transfer function analysis showed that AP quickly responded to the electrical stimulation and reached 90% of the steady-state response at 21 ± 5 sec (Fig. 139.4B). The gain was 1.0 ± 0.3 mmHg/ Hz. Therefore, the epidural approach would be a potential interface for the human BBS.

FEASIBLITY STUDY OF BBS IN PATIENTS WITH HIGH CERVICAL SPINAL CORD INJURY A prototype of the clinical BBS with the epidural catheter electrodes [4] was developed and tested in some patients with high cervical spinal injury. A representative example is shown in Figure 139.5. HUT induced severe hypotension and syncope after transient AP elevation due to short myoclonus; however, the BBS prevented hypotension and enabled the subject to maintain a sitting position. The efficacy and safety of long-term use of the BBS should be investigated in the future.

IMPLANTABLE BBS For practical use of the BBS for patients with central baroreflex failure, clinically applicable materials and devices should be developed, i.e., a pressure sensor, an implantable stimulator, and stimulating electrodes. Fortunately, certain difficulties posed by these challenges

have already been addressed in other areas of clinical practice, and may be readily adaptable for use with the BBS. For example, a tonometer has been developed as a noninvasive continuous monitor of AP [5]. Implantable pulse generators such as cardiac pacemakers can serve as an electrical stimulator. Also, implantable wire leads for nerve stimulation and epidural catheters for spinal stimulation have been approved for the long-term treatment of some neurological disorders [6]. A future advance in development of the implantable BBS for assisting patients with difficulties in maintaining blood pressure during orthostatic maneuvers is expected in the future.

References [1] Sato T, Kawada T, Inagaki M, Shishido T, Takaki H, Sugimachi M, et al. New analytic framework for understanding sympathetic baroreflex control of arterial pressure. Am J Physiol Heart Circ Physiol 1999;276:H2251–H2261. [2] Sato T, Kawada T, Shishido T, Sugimachi M, Alexander Jr. J, Sunagawa K. Novel therapeutic strategy against central baroreflex failure: A bionic baroreflex system. Circulation 1999;100: 299–304. [3] Sato T, Kawada T, Sugimachi M, Sunagawa K. Bionic technology revitalizes native baroreflex function in rats with baroreflex failure. Circulation 2002;106:730–4. [4] Yamasaki F, Ushida T, Yokoyama T, Ando M, Yamashita K, Sato T. Artificial baroreflex: Clinical application of a bionic baroreflex system. Circulation 2006;113:634–9. [5] Sato T, Nishinaga M, Kawamoto A, Ozawa T, Takatsuji H. Accuracy of a continuous blood pressure monitor based on arterial tonometry. Hypertension 1993;21:866–74. [6] Shimoji K, Kitamura H, Ikezono E, Shimizu H, Okamoto K, Iwakura Y. Spinal hypalgesia and analgesia by low-frequency electrical stimulation in the epidural space. Anesthesiology 1974;41:91–4.

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140 Anesthetic Management in Autonomic Disorders Chih Cherng Lu, Shung Tai Ho, Che Se Tung INTRODUCTION A major goal of anesthesia is to maintain optimum autonomic cardiovascular homeostasis in the surgical patient. Intelligent anesthesia care requires knowledge of autonomic nervous and cardiovascular pharmacology to achieve desirable interactions of anesthetics with the autonomic nervous system. Nowadays, the anesthesiologist is confronted with more surgical patients with autonomic disorders. Some of the most severely affected individuals with pure autonomic failure or multiple system atrophy often have both extremely low blood pressures upright and extremely high blood pressures supine [1]. They are perhaps the most vulnerable of all such patients to the complex and interacting responses of drugs and perturbations which occur during anesthetic care. The major risk factors for autonomic disorder undergoing surgery include vasovagal reflex activation, unexplained intraoperative hypotension, and unpredictable cardiovascular collapse [2]. The loss of cardiovascular reflexes in autonomic disorders can complicate anesthesia and predispose patients to life threatening changes in blood pressure [3]. This review will discuss the pathophysiology that underlies autonomic disorders with particular emphasize on those aspects most relevant to the pre-anesthetic evaluation and anesthetic care in the perioperative and intraoperative settings.

CLINICAL PRESENTATION AND ASSESSMENT Orthostatic hypotension (OH) is the core feature of severe autonomic failure and defined as a fall in blood pressure of 20/10 mmHg on standing [4]. Other common clinical manifestations of autonomic failure include postprandial hypotension, urinary bladder dysfunction with urinary retention, decreased GI motility (sometimes with severe constipation), and erectile dysfunction. Special physiological phenomena were first revealed in autonomic failure patients, e.g., food ingestion induces a profound

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00140-2

decrease in blood pressure in these patients, perhaps due to splanchnic blood pooling. Conversely, commonly used medications such as phenylpropanolamine, and evenly seemingly innocuous interventions such as water drinking, can produce dramatic increases in blood pressure in these patients. Autonomic function testing can help to determine the severity and extent of the autonomic impairment, and hence aid in the anticipation of potential complications during anesthesia and surgery. A useful panel of five simple tests of autonomic cardiovascular function includes heart rate responses to the Valsalva maneuver, standing up, and deep breathing, and blood pressure responses to standing up and sustained handgrip. The tests involving changes in heart rate permit assessment for impairment in the parasympathetic system, which precedes changes in the measures of blood pressure that reflect sympathetic injury (Table 140.1).

PATHOPHYSIOLOGY Proper functioning of the autonomic nervous system (ANS) requires that both afferent and efferent limbs are intact. Dysfunction of the afferent limb is typically associated with labile hypertension, as seen in baroreflex failure. Abnormalities of central autonomic pathways (e.g., multiple system atrophy), of efferent effector systems (e.g., deficiency of dopamine beta hydroxylase), or any combination thereof (e.g., diabetic autonomic neuropathy) can all lead to clinical autonomic failure and disabling orthostatic hypotension. Autonomic disorders (“dysautonomia”) refer to a condition in which altered autonomic function adversely affects health (Fig. 140.1). Orthostatic hypotension is ascribed to a defective increase in arterial resistance and an excessive venous pooling upon standing. Effective pharmacological treatment is now available, but may aggravate supine hypertension and have other undesirable effects. Non-pharmacological measures are regarded as a cornerstone in the treatment of orthostatic hypotension.

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PREOPERATIVE MANAGEMENT When performing a preoperative evaluation of a patient with an autonomic disorder, one should pay attention to those who are particularly sensitive to change in vascular tone or volume status. This information may help ease some concerns or allow us to better prepare for the upcoming anesthetic care. Basic evaluation of these patients includes measurement of blood pressure and heart rate while supine, and again after one and three minutes of standing. Ideally prepared and optimally managed before anesthesia, even patients with known autonomic disorders may, with optimal care, have comparable morbidity and mortality to those without autonomic disease. Through the measurement of specific autonomic nervous system parameters one can stratify the risk of cardiovascular and cerebrovascular events before surgery [5].

TABLE 140.1 Noninvasive Tests for Assessing the Autonomic Nervous System Clinical examination

Normal value Ratio of >1.21

HR response to standing

HR is measured as the subject moves Ratio of >1.04 from a resting supine position to standing. A normal tachycardic response is maximal around the 15th beat after rising. A relative bradycardia follows, that is most marked around the 30th beat after standing. The response to standing is expressed as a 30:15 ratio and is the ratio of the longest R-R interval around the 30th beat to the shortest R-R interval around the 15th beat

HR response to deep breathing

The subject takes six deep breaths in 1 minute. The maximum and minimum heart rates during each cycle are measured, and the mean of the differences (maximum HR – minimum HR) during three successive breathing cycles is taken as the maximum-minimum HR

Sympathetic BP response to standing

The subject moves from resting Difference supine to standing, and standing SBP <10 mmHg is subtracted from supine SBP

BP response to sustained handgrip

The subject maintains a handgrip of 30% of the maximum handgrip squeeze for up to 5 minutes. BP is measured every minute, and the initial DBP is subtracted from the DBP just before release

ANESTHETIC MANAGEMENT Currently, there are no systematic studies that have addressed the preference of general versus regional anesthesia in autonomic failure patients. The anesthesia technique should be selected on an individual basis because there is no single “recipe” that can guarantee hemodynamic stability during anesthetic induction. The anesthetic may interfere with cardiovascular performance either by direct effects on the heart and vasculature, or indirectly by modifying the neurohumoral control mechanisms of the circulation. More frequent and severe hypotension on induction of anesthesia must be anticipated in the surgical patient with an autonomic disorder because of the impaired autonomic cardiovascular compensatory mechanisms. Unexpected hypotension in autonomic disorders may be precipitated by a relatively hypovolemic state and vasodilatation secondary to a reduction in sympathetic tone induced by anesthetics. Volatile agents are often chosen as the primary maintenance anesthetic. The predominant effect of isoflurane, desflurane, and sevoflurane is dose-dependent vasodilatation with resultant decreases in systemic vascular resistance and blood pressure. Commonly, “transient sympathectomy” below the mid-thoracic level is achieved through spinal or epidural anesthesia, resulting in vasodilatation and decreases in blood pressure. Impairment of compensatory baroreflex control of upper extremity and cardiac function is complicated by regional and general anesthesia in autonomic disorders; which is usually preventable by adequate perioperative fluid replacement and postural manipulation [6]. Transition from spontaneous to controlled ventilation will acutely reduce venous return and, subsequently, cardiac filling in patients with autonomic disorders relies more heavily on adequate preload than normal subjects. Whenever practical, maintenance of spontaneous respiration is preferred to controlled ventilation. The combination of generous use of induction agents and aggressive

Technique

Parasympathetic HR response The seated subject blows into a to a Valsalva mouthpiece (while maintaining a pressure of 40 mmHg) for 15 maneuver seconds. The Valsalva ratio is the ratio of the longest R-R interval (which comes shortly after release) to the shortest R-R interval (which occurs during the maneuver)

Mean difference >15 beats/min

Difference >16 mmHg

BP, blood pressure; DBP, diastolic blood pressure; HR, heart rate; SBP, systolic blood pressure. (Reprint from David B. Glick, 2009 Miller Anesthesiology, 7th edn. Churchill Livingstone.)

ventilation may result in disastrous hypotension in patients and should be avoided [7].

SPECIAL CONSIDERATION FOR ANESTHESIA Airway Management Several features related to the pathophysiology of the autonomic disorder may complicate airway management in the anesthetic care setting. The combination of impaired gastrointestinal motility and laryngeal dysfunction make

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SPECIAL CONSIDERATION fOR ANESTHESIA

Noradrenergic inhibition Common

Rare

Prescribed drugs Neurocardiogenic syncope Drabetic autonomic neuropathy Alcohol Parkinson disease Hyperthyroidism Multiple system atrophy Multiple myeloma Quadriplegia Amyloidosis Pure autonomic failure Chagas disease Familial dysautonomia Dopamine β-hydrpxylase deficiency

Noradrenergic activation Common

Essential hypertension Congestive heart failure Myocardial infarction Postural tachycardia syndrome Panic disorder Carotid endarterectomy Intracranial bleeding Hyperdynamic circulation syndrome Renovascular hypertension Guillain-Barré syndrome Baroreflex failure “Autonomic epilepsy” Norepinephrine transporter deficiency

Rare

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pressure ventilation on venous return and cardiac output. In patients with autonomic failure, hyperventilation causes a rapid fall in blood pressure. Conversely, hypercapnea (such as hypoventilation or increased dead space ventilation) can rapidly raise blood pressure.

Hepatic Blood Flow and Lidocaine The liver blood flow can decrease as much as 30% with upright posture in patients with autonomic failure. In consequently, a hepatically cleared drug like lidocaine may display plasma levels that are dramatically posturedependent owing to their blood pressure dependence. When patients with orthostatic hypotension are receiving intravenous lidocaine, the plasma drug levels are almost twice as high when they are seated vs. supine, and this change in posture can result in a toxic seizure despite an unchanged infusion rate during anesthetic management.

Temperature Dysregulation Impaired perspiration can lead to an inability to dissipate heat adequately. This could also contribute to a persistent high fever postoperatively. With aggressive warming measures, there is sometimes also a thermally induced hypotension. Conversely, the inability of autonomic disorder to vasoconstrict in response to a cool environment may result in excessive heat loss and hypothermia and should be watched cautiously in anesthetic care.

Response to Infection

FIGURE 140.1 Dysautonomias featuring altered sympathetic noradrenergic function. (Reprint from David, S., Goldstein et al. Ann Intern Med. 2002;137:753–763.)

aspiration a particular concern in these patients. Rapid sequence induction should be considered. Mask induction with volatile anesthetics technique should be encouraged for those patients with autonomic disorders complicated by compromised cardiopulmonary reserve. Apnea in autonomic failure patients, either due to upper airway obstruction (such as that seen in obstructive sleep apnea) or due to impaired central regulation of respiration, may be aggravated postoperatively. In such instances it is likely due to disordered central control of respiration during anesthesia rather than extreme respiratory center sensitivity to the anesthetic agent [8].

Ventilatory Management The inability of patients with autonomic disorders to increase cardiac output through sympathetic activation makes them exquisitely sensitive to the effects of positive

Patients with autonomic disorder often have impaired capacity to manifest a fever in response to infection. Instead an infection may present with minimal fever or sometimes none at all. If a patient with autonomic failure experiences greater than usual hypotension or blood pressure lability, new-onset recurrent syncope, or an acute decrease in performance status, a search should be undertaken for occult infection, with emphasis on evaluation for urinary tract infection or aspiration pneumonia.

Diabetic Neuropathy Diabetes mellitus is the most commonly recognized cause of autonomic dysfunction. Unexplained tachycardia has been attributed to damage of parasympathetic fibers and it is thought to represent early cardiac involvement by diabetic peripheral neuropathy. When in the late stages, sympathetic impairment of cardiac innervation also occurs. Unexpected intraoperative bradycardia in patients with diabetic autonomic neuropathy may not respond to atropine (since vagal tone is already diminished in these patients), and alternatives such as isoproterenol or a temporary pacemaker can be life-saving.

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Uremia Uremic patients are sometimes encountered with afferent baroreflex impairment, and the patient’s blood pressure and heart rate are no longer buffered. In this circumstance, large excursions of blood pressure and heart rate may occur in response to stress (very high blood pressures) and sedation (low blood pressures).

SUMMARY Patients with autonomic disorders usually have significantly perturbed homeostatic responses to normal physiological and environmental stresses. Severe autonomic disorders increase in prevalence with age; aging is associated with reduced baroreflex responsiveness, decreased cardiac compliance, and attenuation of the vestibulosympathetic reflex. Most anesthetics remove an important compensatory mechanism to maintain blood pressure and exaggerate the response to vasoactive and cardiac depressant agents in human subjects [9]. Preoperatively, cardiovascular autonomic function should be carefully evaluated and should facilitate meticulous control of blood volume in patients with autonomic disorders. The pharmacologic complexities of anesthetic management in patients with

autonomic disorders suggest that all anesthetics should be administered judiciously. Postoperatively, orthostatic hypotension control necessitates the use of volume expansion, postural training and occasionally vasoconstrictors.

References [1] Goldstein DS, Sharabi Y. Neurogenic orthostatic hypotension: A pathophysiological approach. Circulation 2009;119:139–46. [2] Grenier Y, Drolet P. Asystolic cardiac arrest: An unusual reaction following iv metoclopramide. Can J Anaesth 2003;50:333–5. [3] Malan MD, Crago RR. Anaesthetic considerations in idiopathic orthostatic hypotension and the Shy-Drager syndrome. Can Anaesth Soc J 1979;26:322–7. [4] Freeman R. Clinical practice. Neurogenic orthostatic hypotension. N Engl J Med 2008;358:615–24. [5] Gunther A, Witte OW, Hoyer D. Autonomic dysfunction and risk stratification assessed from heart rate pattern. Open Neurol J 2010;4:39–49. [6] Tokuda K, Motoyama Y, Kai Y, Sakaguchi Y, Hoka S. Anesthetic management for a patient with significant orthostatic hypotension probably due to pure autonomic failure. Masui 2009;58:1010–3. [7] Shimazu T, Tamura N, Shimazu K. Aging of the autonomic nervous system. Nippon Rinsho 2005;63:973–7. [8] Drury PM, Williams EG. Vocal cord paralysis in the Shy-Drager syndrome. A cause of postoperative respiratory obstruction. Anaesthesia 1991;46:466–8. [9] Klein CM. Evaluation and management of autonomic nervous system disorders. Semin Neurol 2008;28:195–204.

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141 Evolution of the Cardiovascular Autonomic Nervous System in Vertebrates Tobias Wang The need for coordinated control of visceral functions to maintain homeostasis has undoubtedly been of paramount importance since the early evolution of animals. Thus, a functional analogy of the autonomic nervous system (ANS) is likely to have an ancient evolutionary history, and must have been essential as soon as multicellular animals divided specific physiological function to particular organs. Because of this evolutionary history, studies on different animal groups can provide fundamental insight to the foundations of both anatomy and function of the ANS in mammals, including humans. Many of the basic functions of the ANS were originally discovered in ectothermic vertebrates. As prominent examples, Gaskell’s demonstrations of the vagal inhibitory role on the heart were performed on turtles and frogs and Loewi’s demonstration of acethylcholine being the postganglionic neurotransmitter within the parasympathetic nervous system (“vagus stoff”) was based on studies in amphibians. These animals were chosen because of their enormous tolerance to hypoxia and low temperatures, rendering these animals resilient to experimentation and hence suitable for physiological studies before methods for anesthesia and mechanical ventilation of mammals. More recently, studies on ANS functions in ectothermic vertebrates were crucial for the discovery of non-adrenergic-non-cholinergic neurotransmitters within the sympathetic and parasympathetic nervous systems.

THE AUTONOMIC NERVOUS SYSTEM IN VERTEBRATES A phylogeny depicting extant (living) groups of chordates (animals that have a notochord during some stage within their life cycle) is shown in Figure 141.1, where the different vertebrate groups, i.e., animals with skeletal elements surrounding the spinal cord and notochord, are highlighted within the grey box [1]. While tunicates and amphioxus are endowed with a nervous system that may resemble the enteric nervous system, the equivalent of an ANS is not present [2,3]. Within hagfishes and lampreys the ANS is considered rudimentary, because either some organs are devoid

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of innervation or most organs lack dual innervation. In both groups, the left and right vagi unite to form a ramus intestinalis impar that innervates the intestine and the gallbladder. In hagfishes, the ramus intestinalis impar does not innervate the heart, whereas the heart of lampreys does receive vagal innervation. In both hagfishes and lampreys, spinal “sympathetic” nerves leave the dorsal as well as well as ventral spinal nerves, but there are no sympathetic chains or segmental ganglia. The sympathetic nerves innervate several visceral organs, but not the heart. It must be emphasized that hagfishes and lampreys have evolved over the past several hundred millions years and are unlikely to represent exact copies of their ancestors. The eyes of both groups, for example, are degenerated, which may relate to the poor innervation by the ocular nerve. Nevertheless, it seems very reasonable to conclude that the ANS was poorly developed in early vertebrates [2–4]. Sharks and rays (cartilaginous fishes) have segmentally arranged paravertebral ganglia that are linked by a loose plexus of nerve fibers [2–4]. The vast majority of the fibers arise from the ventral roots of the spinal nerves. These sympathetic nerves innervate most visceral organs with the notable exception of the heart, and do not appear to enter the head. Cranial autonomic fibers, i.e. the parasympathetic nervous system, occur within the oculomotor (III), facial (VII), glossopharyngeal (IX) and vagus (X) nerves. The vagal fibers reach the stomach and the anterior parts of the intestine. The vagus also innervates the heart and exerts both negative chronotropic and inotropic effects [2,3]. In bony fishes (teleosts), the sympathetic chain ganglia are well developed and form distinct sympathetic chains [2,3]. Some of these sympathetic fibers join the vagus, creating a vagosympathetic trunk. This is also the case in amphibians and reptiles. The overall pattern of innervation and the actions of the autonomic nervous system in bony fishes resemble that found amphibians, reptiles, birds and mammals (i.e., the tetrapods) is rather similar. Therefore, while the ANS of early vertebrates can be considered simple in comparison to subsequent groups of vertebrates, the basic foundations of the ANS were established by the time of fishes and evolved prior to the invasion of terrestrial habitats. The swimbladder of bony

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141. EVoluTIoN of THE CARdIoVASCulAR AuToNomIC NERVouS SySTEm IN VERTEbRATES

FIGURE 141.1 A phylogeny of the major

Cardiac innervation Vagal

groups of extant (i.e., living) craniates and vertebrates. Note that only birds and mammals are endothermic and that this trait appears to have evolved independently from two different groups of reptilian and ectothermic ancestors (modified from Liem et al., 2001 [1]). The figure includes a synopsis of the parasympathetic and sympathetic innervations of the heart and vasculature.

Sympathetic

Tunicates Amphioxus Hagfishes Excitatory

Lampreys

Coelacanth Lungfishes Amphibians Birds Crocodilians Lizards & snakes Turtles

A-and β adrenrgic innervation

Bony fishes

Excitatory sympathetic innervation of the heart

Inhibitory parasympathetic innervation of the heart

Sharks & rays

Mammals

fishes is used to control buoyancy, but probably evolved originally as a gas exchange organ. Therefore, the swimbladder can be considered homologous to the lungs of lungfish and tetrapods. Both organs form embryologically as an outpocketing of the gut; the swimbladder as a ventral extension, while the lungs derive from a dorsal extension. Both organs receive a dual innervation form the ANS in most species.

COMPARATIVE ASPECTS OF THE AUTONOMIC REGULATION OF THE CARDIOVASCULAR SYSTEM Anatomy of the Cardiovascular System in Vertebrates The morphology and the function of the heart and cardiovascular systems of vertebrates have undergone large evolutionary changes associated with the transition between water and air-breathing and during the evolution of endothermy within mammals and birds [5,6]. A schematic representation of these cardiovascular designs is shown in Figure 141.2, where all the different groups of

fish (hagfishes, lampreys, cartilaginous fishes, teleosts and the coelacanth) are presented as having a similar “piscine” circulatory design. Thus, while the cardiac morphology, the number of gills arches and other characters differ considerable between these groups, they all rely on gills for gas exchange [7]. With some variations, the hearts of all groups consist of a sinus venosus, a single atrium and a single ventricle as well as an outflow tract that may be both muscular and contractile as the conus arteriosus in sharks. This heart ejects the oxygen poor-blood that has returned from the body towards the gills, where the blood is oxygenated before it is delivered to the body. The branchial circulation and the perfusion of the systemic vascular beds of fish, therefore, occur in series and the emergence of parallel circulations arose with evolution of the lungs. In lungfishes (Dipnoi), the first group of vertebrates with real lungs, the common pulmonary vein draining the lungs empties into the sinus venosus in close proximity to the atrium. The single trabeculated atrium is partially divided by a fold that prevents admixture of the oxygenrich blood from the lungs and oxygen-poor blood from the systemic circulation. The oxygen-poor blood is preferentially directed through the posterior gill arches and can either enter the pulmonary artery or the dorsal aorta after having perfused the gills. The oxygen-rich blood primarily

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671

Body Body

Lungfish

Body

Amphibians

Lungs

Body

Birds

Body

Crocodiles

Body

Turtles, lizards and snakes

Body

Lungs Lungs Lungs

Skin

Lungs

Heart

Gills

Gills

“Piscine”

Lungs

Autonomic Regulation of the Heart

Mammals

FIGURE 141.2 The evolution of the cardiovascular system amongst the major groups of extant vertebrates. In this representation, the cardiovascular design of hagfish, lampreys as well as the cartilaginous and bony fishes are presented as a general “piscine design”. Note that only mammals and crocodiles have a complete division of the ventricle and that lungfishes, amphibians and reptiles have the capacity to mix oxygen-rich and oxygen-poor blood within the ventricle (modified from Jensen et al., 2010 [6]).

enters the anterior and degenerated gill arches to enter the dorsal aorta directly [7]. In amphibians, snakes, lizards and turtles there are two separate atria receiving the venous blood from the systemic and pulmonary circulations. While the ventricular anatomy differs enormously amongst these groups, they are characterized by having a single ventricle, but distinct outflows to either the systemic or pulmonary circulations [6]. In crocodiles, which in some ways are more closely related to birds than the other reptiles, a complete septum divides the ventricle, but because the left aorta arises from the right ventricle, blood can bypass the pulmonary circulation. Birds have a cardiac anatomy that closely resembles mammals, but an independent evolution of the divided ventricle is indicated by birds retaining the right aortic arch during ontogenetic development, whereas mammals retain the left aortic arch [1].

The aneural myogenic hagfish heart is devoid of any innervation and is remarkably insensitive to acetylcholine or cholinergic antagonists [4,7]. The hagfish heart, nevertheless, is under a tonic paracrine β-adrenergic tone that seems to derive from catecholamine stores within the heart [4,7]. It remains uncertain whether the cardiac release of catecholamines and hence the adrenergic tone is regulated. Heart rate increases markedly upon increased filling, a response that is independent of adrenergic stimulation and this may be important to increase cardiac output when venous return is elevated. The mechanism for this response still remains to be determined. Hagfish inhabit hypoxic sediments and exhibit a marked tolerance to oxygen deprivation, associated with a pronounced reduction in heart rate that may be direct effects of oxygen lack on the pacemaker cells. Consistent with the lack of innervation, hagfish exhibit only small changes in heart rate during exercise [7]. Lampreys represent the first group of chordates with a vagal innervation of the heart, where the vagus travels along the jugular vein to the sinus venosus where the primary pacemaker region is located. This innervation, however, differs fundamentally from the rest of the vertebrates by being excitatory and that cardiac acceleration can be blocked by nicotinic cholinoceptor antagonists. Although there is no sympathetic innervation, the lamprey heart, nevertheless, exhibit positive inotropic and chronotropic responses to β-adrenergic stimulation and heart rate. As in hagfish, catecholamines are released in a paracrine fashion from cardiac stores in lampreys and increased filling of the heart also causes tachycardia [4,7]. The typical inhibitory action of the vagus nerve did not appear until the evolution of cartilaginous fishes, while the opposing role of an excitatory sympathetic innervation evolved later with the emergence of bony fishes. This cardiac innervation has remained the same within all subsequent groups of vertebrates, although the lungfish heart lacks sympathetic innervation although it does respond with increased rate and contractility to β-adrenergic stimulation originating from catecholamines stored within the heart [7].

Innervation of the Systemic Vasculature In tunicates and Amphioxus, the endothelium within the blood vessel is either very poorly developed or absent, and there is no endothelium in innervation amongst protostomes. Also there is no evidence for adrenergic receptors affecting vascular tone. Sympathetic nerves appear to be present in both hagfish and lampreys and the systemic vasculature responds to adrenergic agonists as well as acetylcholine. In all other groups of vertebrates, there are both α- and β-adrenergic receptors causing constriction and dilatation, respectively. This innervation is present on both the arterial and venous side of the circulatory system, but

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141. EVoluTIoN of THE CARdIoVASCulAR AuToNomIC NERVouS SySTEm IN VERTEbRATES

FIGURE 141.3 The cardiovascular changes in a fresh-

ventilation

120 100 80 60 40 20 0 100 80 60 40 20 0 50 40 30 20 10 0

water turtle (Trachemys scripta) during the transition from apnea to ventilation of the lungs. Heart rate and pulmonary blood flow are low during breath-holding (apnea), but increase markedly during ventilation associated with a change in the cardiac shunt pattern (modified from Wang and Hicks, 1996 [9]).

pulmonary blood flow (ml min–1)

systemic blood flow (ml min–1)

hart rate (min–1) 2 min

very little is known about putative difference between the different vascular beds to specific organs. The sympathetic innervation therefore is involved in both blood pressure regulation and regulation of venous tone and cardiac filling. The endothelium of ectothermic vertebrates releases nitric oxide as in mammals, but endothelial nitric oxide synthase (eNOS) only seems to have evolved in tetrapods, so neural NOS is responsible for NO production in the endothelium of fish.

Cardiovascular Responses to Altered Pressure, Exercise and Hypoxia A barostatic regulation of heart rate mediated through the ANS is present in fishes. Being aquatic, the circulatory system of fishes is not influenced by gravity and blood pressure control therefore seems to have evolved prior to any orthostatic pressure changes. The ancestral evolutionary benefit of an inhibitory vagal effect on heart rate and an opposing positive chronotropic and inotropic responses from sympathetic stimulation (or release of catecholamines) is likely to have been the ability to accommodate changes in metabolism with appropriate changes in heart rate and stroke volume, while maintaining a stable perfusion pressure. In all of the ectothermic vertebrates, metabolism changes in accordance with altered body temperatures; typically metabolism increases two-tothreefold when temperature increases 10°C. In these cases, heart rate increases proportionally, primarily because of a direct effect of temperature on the cardiac pacemaker and the scope for autonomic regulation normally remains intact. Therefore, the animals are able to increase heart

rate during exercise while the capacity to increase aerobic metabolism remains intact over a broad range of temperatures. Similarly the ectothermic animals retain the capacity for heart rate to respond to altered blood pressure over a broad temperature range. Hypoxia is common in aquatic habitats. Given that the foundations of autonomic regulation evolved in vertebrates relying on aquatic respiration over the gills, it is interesting that cartilaginous and bony fishes typically respond to hypoxia with a reduction in heart rate. Cardiac output typically remains unchanged, because the bradycardia is accompanied by increased stroke volume. The hypoxic bradycardia does not appear to enhance gas exchange efficiency across the gills, and may have evolved to protect the heart from oxygen lack. Regardless of the functional benefit of the hypoxic bradycardia, it is interesting that mammals also exhibit bradycardia during hypoxic stimulation of peripheral chemoreceptors but only when afferent feedback from pulmonary stretch receptors is ablated. Hence, this somewhat “counter-intuitive” cardiovascular response to hypoxia is likely to reflect a piscine condition that evolved prior to pulmonary gas exchange.

Autonomic Regulation of the Pulmonary Circulation Lungs and the pulmonary circulation evolved prior to the separation of the ventricle by a complete ventricular septum that is a characteristic of mammals and birds (Fig. 141.2). The ventricle in lungfish, amphibians, snakes, lizards and turtles, accordingly, is not fully divided and systolic blood pressures in the systemic and

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ComPARATIVE ASPECTS of THE AuToNomIC REgulATIoN of THE CARdIoVASCulAR SySTEm

pulmonary circulations are identical [6]. The lack of division also implies that oxygen-rich blood from the lungs will, at least to some extent, be mixed with the oxygenpoor blood returning from the systemic circulation. As in the example presented in Figure 141.3, where blood flows were recorded in a freshwater turtle, the degree of mixing of oxygen-rich and oxygen-poor blood within the ventricle as well as the distribution of blood flows between the systemic and pulmonary circulations changes consistently with pulmonary ventilation [9]. Thus, during apnea, which in turtles and other diving species can last for more than one hour, heart rate and pulmonary blood flow are low and much of the oxygen poor blood returns directly into the systemic circulation (i.e., a large right-to-left cardiac shunt). As soon as pulmonary ventilation resumes pulmonary blood flow and heart rate then increase abruptly, while systemic blood flow is less affected. In several species, pulmonary blood flow exceeds systemic blood flow during ventilation, implying that a significant fraction of the blood returning from the lungs is returned directly to the pulmonary circulation (i.e., a left-to-right cardiac shunt).While the changes in cardiac shunt pattern act to provide a temporal matching of pulmonary ventilation and perfusion, the functional significance of these cardiovascular changes remain to be understood. The degree of oxygenated and deoxygenated blood mixing and the distribution of blood flows between the systemic and pulmonary circulations are dictated by the vascular resistance in the lungs and body [8]. Thus, when pulmonary vascular resistance is low compared to the systemic circulation, blood will primarily be distributed towards the lungs and vice versa [6,8]. The resistance of

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the pulmonary circulation is regulated by the ANS, where vagal innervation of smooth muscle in the pulmonary artery can cause constriction and reduce pulmonary blood flow. In mammals and birds, this vagal regulation of pulmonary blood vascular resistance is not present.

References [1] Liem KF, Bemis WE, Walker WF, Grande L. Functional anatomy of the vertebrates An evolutionary perspective. Philadelphia: Harcourt; 2001. [2] Burnstock G. Evolution of the autonomic innervation of visceral and cardiovascular systems in vertebrates. Pharmacol Rev 1969;21:247–324. [3] Gibbins I. Comparative anatomy and evolution of the autonomic nervous system. In: Nilsson S, .Holmgren S, editors. Comparative physiology and evolution of the autonomic nervous system.: Harwood Academic Publishers; 1994. p. 1–67. [4] Nilsson S, Holmgren S. The autonomic nervous system and chromaffin tissue in hagfishes. In: Jørgensen JM, Lomholt JP, Weber RE, Malte H, editors. Biology of Hagfishes. London: Chapman & Hall; 1998. p. 480–95. [5] Morris JL, Nilsson N. The circulatory system. In: Nilsson S, Holmgren S, editors. Comparative physiology and evolution of the autonomic nervous system. : Harwood Academic Publishers; 1994. p. 193–246. [6] Jensen B, Nielsen JM, Axelsson M, Pedersen M, Löfman C, Wang T. How the python heart separates pulmonary and systemic blood pressures and blood flows. J Exp Biol 2010;213:1611–7. [7] Farrell AP. Cardiovascular systems in primitive fishes. In: McKenzie DJ, Farrell AP, Brauner C J, editors. Fish physiology: Primitive fishes.: Elsevier; 2007. p. 53–120. [8] Taylor EW, Andrade D, Abe AS, Leite Cleo AC, Wang T. The unequal influences of the left and right vagi on the control of the heart and pulmonary artery in the rattlesnake, Crotalus durissus. J Exp Biol 2009;212:145–51. [9] Wang T, Hicks JW. Cardiorespiratory synchrony in turtles. J Exp Biol 1996;199:1791–800.

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142 Human Physiome Peter Hunter INTRODUCTION Computational biology and computational physiology are playing an increasingly important role in helping scientists understand and quantify the complex multiscale structure-function relations of biological systems. Interpreting the wealth of quantitative data now available on tissue and subcellular processes requires a new level of international collaboration between biological, physical (including engineering), mathematical, computer and computational scientists. Furthermore, biological processes operate primarily at the molecular scale (ligand/ protein/DNA/RNA interactions) but are influenced by, or in turn create an influence on, the physiological systems of cells, tissues, organs and whole body organ systems. This influence therefore encompasses a 109 range of spatial scales (from nm at the molecular scale to m at the human organ system level) and a 1015 range of temporal scales (from μs for molecular interactions to the 109 s of a human lifespan). The VPH/Physiome project is facilitating the multidisciplinary contributions and addressing the challenges of modeling the enormous range of spatial and temporal scales involved in physiological processes. In this brief overview of the human physiome we discuss the development of modeling standards, databases and computational tools by the VPH/Physiome project and illustrate these with reference to the heart. We then suggest how the framework could be applied to the autonomic nervous system and describe some initial steps that have been taken.

PHYSIOME STANDARDS As the computational models inevitably become more complex, it is increasingly difficult for anyone other than the author(s) of the publication describing the model to decipher, code and run the model in order to reproduce the results claimed in a publication. It can also be very

difficult to then use this model as one component of a more complex model. To address these challenges several groups have developed standards for encoding models over the past 10 years. These modeling standards typically use the eXtensible Markup Language (XML) developed by the w3c1 as well as a variety of other standards based on XML, such as MathML for encoding mathematics, as well as various metadata standards. Two XML-based model encoding standards are currently being developed under the IUPS Physiome Project [1,2] and the European Virtual Physiological Human (VPH) project (www.vph-noe.eu). CellML (www.cellml. org) is designed to encode lumped parameter biophysically based systems of ordinary differential equations (ODEs) and nonlinear algebraic equations (together called differential algebraic or DAE systems). FieldML (www. fieldml.org) is designed to encode spatially and temporally varying field information such as anatomical structure, the spatial distribution of protein density or computed fields such as the electrical potential or oxygen concentration throughout a tissue. A third markup language called the “systems biology markup language” or SBML (www.sbml. org) has been developed by the systems biology community. This is similar to CellML but targeted more specifically to representing models of biochemical reactions. CellML maintains a clean separation between the syntax of a model (e.g., the mathematical equations encoded in MathML) and the semantics (the biological and biophysical meaning of the model components and parameters) defined in the model metadata by reference to suitable ontologies. This facilitates building complex models by importing modular components defined in libraries. SBML is more closely tied to the concept of a biochemical “reaction”, and does not maintain a separation between the model syntax and the biological semantics. FieldML deals with the encoding of anatomy at multiple spatial scales by allowing hierarchies of material coordinate systems that preserve anatomical relationships (e.g., coronary arteries embedded in a deforming myocardial tissue that is itself part of a heart contained within a torso).

1

“w3c” refers to the worldwide web consortium that is developing and maintaining standards for information exchange via the internet – see www.w3c.org.

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142. HumAn PHysIomE

FIGURE 142.1 The multi-scale cardiac physiome modeling hierarchy from genes to the whole organism. Parameters used in a model at one scale can often be derived from a more detailed model at a lower spatial scale.

These three standards are supported by the US National Institutes of Health (NIH) and the European Commission’s funding agency (currently operating under Framework 7). Model repositories are available for all three markup languages – PMR2 (Physiome Model Repository 2, see models.cellml.org) for CellML and FieldML models and Biomodels (www.biomodels.net) for SBML models. Various minimum information standards are also available including MIRIAM (www.ebi.ac.uk/miriam) and MIASE (www .ebi.ac.uk/compneur-srv/miase). An illustration of the use of these standards, together with the computational software and model repositories associated with the CellML and FieldML standards, is given in Fig. 142.1. The heart physiome project is the most developed example of multi-scale modeling as it has been able to draw on decades of measurements and modeling of cardiac ion channels [3], myofilament mechanics [4] and myocardial structure [5]. The models link the biomechanics of the myocardial tissue and blood flow (ventricles and coronaries) to cellular processes that include the ion channel based electrophysiology, intracellular calcium transport, myofilament mechanics, control of pH and bicarbonate, metabolic pathways and signal transduction pathways. A key feature of physiome models is the incorporation of anatomical structure into the models in order to

accurately represent structure–function relations at the various spatial scales. For example, the 3D structure of tissue that gives rise to continuum properties (conductivity, elasticity, etc) is shown as part of the multi-scale framework in Figure 142.2. Note that there are characteristic material directions in the tissue that define structural features, such as the fiber direction or sheet orientation, and that the continuum properties are different in each direction (the tissue is said to be “anisotropic”). Note also that these characteristic material directions and the values of the continuum properties vary throughout the tissue (the tissue properties are said to be “inhomogeneous”). Thus the electrical and mechanical properties along the fiber direction in cardiac muscle are different from those properties measured transversely to the fibers and these properties are different in different material locations. Anisotropy and inhomogeneity are characteristics of all biological tissues. The mechanics equations are usually solved using finite element techniques and the electrical activation equations are solved using finite volume or similar techniques based on grid points defined as material points of the deforming mechanics grid. Other organ systems that have been modeled with anatomically and biophysically based multi-scale approaches have been the lungs [6], the digestive system [7], the neuro-musculo-skeletal system [8]. In fact there are now

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Organ model

Myocardial activation Ventricular mechanics Ventricular blood flow & heart valve mechanics

Continuum tissue model

Calcium transport models Myofilament mechanics

Composite lumped parameter

Discrete tissue structure model

Ion channel model of the Hodgkin-Huxley type

Signal pathway models

3D protein model

Metabolic pathway models Gene regulation models

Markov ion channel model

Coarse grained MD model 3D cell model Molecular dynamics model Quantum mechanics model

FIGURE 142.2 Types of model used in the multiscale modeling hierarchy. Models based on systems of ODEs and algebraic equations are shown in blue (so called ‘lumped parameter’ models) and these are encoded in CellML. Models that require the solution of partial differential equations are shown in pink and are encoded in FieldML. The FieldML models link to CellML models at material points in the tissue. The arrows above are shown as unidirectional but, in fact, information flows both ways. The models shown in gray will be linked into the cardiac modeling hierarchy in the future.

nascent physiome modeling efforts on some aspects of all of the body’s 12 organ systems.

MODELING THE AUTONOMIC NERVOUS SYSTEM There is of course a large neuroscience community involved in modeling many aspects of the brain but there has been surprisingly little engagement between neuroscientists and the physiome community over the challenges of connecting organ system models to their neural control or sensory systems. The autonomic nervous system would be a good place to start. Two steps that could be taken are as follows: 1. Develop a FieldML-encoded finite element model of the spinal chord and associated neural pathways throughout the body (not the brain) – see Figure 142.3. These would need to be defined anatomically with respect to the musculo-skeletal system so that they would be, for example, carried with the tissue during movement. 2. Encode, curate and annotate all published models of neural processes in the CellML model repository. These can then be used to create more integrated models that can be linked to organ system physiology. This process has been started and the models that are currently in the CellML model repository at

models.cellml.org/neurobiology are listed below. Literature references are also given for all of these models: Bertram, Rhoads, Cimbora, 2008 [9]. A phantom bursting mechanism for episodic bursting: (a) Original model. (b) Modified to include channel noise in the leak current. Butera, Rinzel, Smith, 1999 [10,11]. Models of respiratory rhythm generation in the pre-Botzinger complex: (a) Bursting Pacemaker Neurons: model 1 (does not include a slow potassium current). (b) Bursting Pacemaker Neurons: model 2 (includes a slow potassium current). (c) Populations of Coupled Pacemaker Neurons: 5 Cell Model. (d) Populations of Coupled Pacemaker Neurons: 10 Cell Model. (e) Populations of Coupled Pacemaker Neurons: Single Cell Model. (f) Populations of Coupled Pacemaker Neurons: Single Cell Model. Cloutier, Bolger, Lowry, Wellstead, 2009 [12]. An integrative dynamic model of brain energy metabolism using in vivo neurochemical measurements. Friel, 1995 [13]. [Ca2]i oscillations in sympathetic neurons: an experimental test of a theoretical model.

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FIGURE 142.3 (a) Neural pathways, and (b) a schematic mapping to organ systems. Figure 3a is from Rita Carter, ‘The Human Brain Book’, by permission Dorling Kindersley Ltd.

Li, Bertram, Rinzel, 1996 [14]. Modeling N-methyl-Daspartate-induced bursting in dopamine neurons: (a) Simple model based on the equations in the published paper. (b) Complex model based on the equations in the published paper. (c) Simple model based on the equations in the original code. Ostby, Omholt, Oyehaug, Einevoll, Nagelhus, Plahte, Zeuthen, Voipio, Lloyd, Ottersen, 2008 [15]. Astrocytic processes explaining neural-activity-induced shrinkage of extraneuronal space. Phillips, Robinson, 2007 [16]. A quantitative model of sleep-wake dynamics based on the physiology of the brainstem ascending arousal system.

Phillips, Robinson, 2008 [17]. Sleep deprivation in a quantitative physiologically based model of the ascending arousal system: (a) Baseline model. (b) Sleep deprivation model. Plant, 1981 [18]. Bifurcation and resonance in a model for bursting nerve cells. Purvis, Butera, 2005 [19]. Ionic current model of a hypoglossal motoneuron. Purvis, Smith, Koizumi, Butera, 2007 [20]. Intrinsic bursters increase the robustness of rhythm generation in an excitatory network: (a) Single pacemaker cell. (b) Single non-pacemaker cell.

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FIGURE 142.4 Schematic of the computational model of the brain stem respiratory network. The model includes interacting neuronal populations within the major brain stem respiratory compartments (Pons, BotC, pre-BotC, and rVRG). Spheres represent neuronal populations (excitatory, red; inhibitory, blue; motoneuronal, brown); green triangles represent sources of tonic excitatory drives (in pons, RTN/BotC, and pre-BotC compartments) to different neural populations. Excitatory and inhibitory synaptic connections are indicated by arrows and small circles, respectively. Simulated 'transections' (dashed lines) mimic those performed in situ.

Rubin, Shevtsova, Ermentrout, Smith, Rybak, 2009 [21]. Multiple rhythmic states in a model of the respiratory central pattern generator. Smith, Abdala, Koizumi, Rybak, Paton, 2007 [22]: (a) (b) (c) (d) (e)

Synaptic coupling. Preinspiratory neuron. Early inspiratory neuron. Augmenting expiratory neuron. Postinspiratory neuron.

Tabak, Mascagni, Bertram, 2010 [23]. Mechanism for the universal pattern of activity in developing neuronal networks. Vasalou, Henson, 2010 [24]. A multiscale model to investigate circadian rhythmicity of pacemaker neurons in the suprachiasmatic nucleus. The model by Smith et al. (2007) [21] provides an example of the use of the CellML framework. A schematic of the model is given in Figure 142.4 which shows interacting neuronal populations within the major brain stem respiratory compartments. The model deals with the brain stem respiratory central pattern generator (CPG) that produces rhythmic movements and shows how the normal

three-phase respiratory rhythm transforms to a two-phase and then to a one-phase rhythm as the network is reduced. Expression of the three-phase rhythm required the presence of the pons, generation of the two-phase rhythm depended on the integrity of Botzinger and pre-Botzinger complexes and interactions between them, and the onephase rhythm was generated within the pre-Botzinger complex. The model shows that the respiratory network has rhythmogenic capabilities at multiple levels of network organization, allowing expression of motor patterns specific for various physiological and pathophysiological respiratory behaviors.

CONCLUSIONS The VPH/Physiome project is providing a framework for multi-scale modeling of physiological processes, typically involving anatomically and biophysically based models that capture structure/function relations at multiple spatial (and sometimes temporal) scales. The framework has processes for ensuring model reproducibility, including markup languages for encoding models, data standards

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and model repositories, as well as open source software for authoring, visualizing and solving models. In this brief overview, we have described the markup languages CellML, SBML and FieldML and have illustrated the use of the physiome framework for modeling the heart. We have also proposed two steps that could now be taken to extend the physiome approach to neural processes, and in particular the autonomic nervous system.

References [1] Hunter PJ, Borg TK. Integration from proteins to organs: The physiome project. Nat Rev Mol Cell Biol 2003;4:237–43. [2] Hunter PJ, Nielsen PMF. A strategy for integrative computational physiology. Physiology 2005;20:316–25. [3] Noble D, Rudy Y. Models of cardiac ventricular action potentials: Iterative interaction between experiment and simulation. Phil Trans R Soc Lond A 2001;359(1783):1127–42. [4] Nash MP, Hunter PJ. Computational mechanics of the heart. J Elasticity 2001;61(1–3):113–41. [5] LeGrice IJ, Hunter PJ, Smaill BH. Laminar structure of the heart: a mathematical model. Am J Physiol 1997;272:H2466–H2476. [6] Tawhai M, Hoffman EA, Lin CL. The lung physiome: Merging imaging-based measures with predictive computational models of structure and function. Wiley Interdiscip Rev Syst Biol 2009;1(1): p. 61–72. [7] Cheng LK, O'Grady GB, Du P, Egbuji JU, Windsor JA, Pullan AJ. Gastrointestinal system. Wiley Interdiscip Rev Syst Biol Med 2010;2(1):p65–79. [8] Fernandez JW, Mithraratne P, Thrupp SF, Tawhai MH, Hunter PJ. Anatomically based geometric modeling of the musculoskeletal system and other organs. Biomech Model Mechanobiol 2004;2(3):139–55. [9] Bertram R, Rhoads J, Cimbora WP. A phantom bursting mechanism for episodic bursting. Bull Math Biol 2008;70:1979–93. [10] Butera RJ, Rinzel J, Smith JC. Models of respiratory rhythm generation in the pre-Botzinger complex. I. Bursting pacemaker neurons. J Neurophysiol 1999;81:382–97. [11] Butera RJ, Rinzel J, Smith JC. Models of respiratory rhythm generation in the Pre-Botzinger complex. II. Populations of coupled pacemaker neurons. J Neurophysiol 1999;82:398–415.

[12] Cloutier M, Bolger FB, Lowry JP, Wellstead P. An integrative dynamic model of brain energy metabolism using in vivo neurochemical measurements. J Comput Neurosci 2009;27:391–414. [13] Friel DD. [Ca2]i oscillations in sympathetic neurons: an experimental test of a theoretical model. Biophys J 1995;68:1752–66. [14] Li YX, Bertram R, Rinzel J. Modeling N-methyl-D-aspartateinduced bursting in dopamine neurons. Neuroscience 1996;71:397–410. [15] Ostby I, Oyehaug L, Einevoll GT, Nagelhus EA, Plahte E, Zeuthen T, Lloyd CM, Ottersen OP, Omholt SW. Astrocytic mechanisms explaining neural-activity-induced shrinkage of extraneuronal space. PLoS Comput Biol 2009;5:1. [16] Phillips AJK, Robinson PA. A quantitative model of sleep-wake dynamics based on the physiology of the brainstem ascending arousal system. J Biol Rhythms 2007;22:167–79. [17] Phillips AJK, Robinson PA. Sleep deprivation in a quantitative physiologically based model of the ascending arousal system. J Theor Biol 2008;255:413–23. [18] Plant RE. Bifurcation and resonance in a model for bursting nerve cells. J Math Biol 1981;11:15–32. [19] Purvis LK, Butera RJ. Ionic current model of a hypoglossal motoneuron. J Neurophysiol 2005;93:723–33. [20] Purvis LK, Smith JC, Koizumi H, Butera RJ. Intrinsic bursters increase the robustness of rhythm generation in an excitatory network. J Neurophysiol 2007;97:1515–26. [21] Rubin JE, Shevtsova NA, Ermentrout GB, Smith JC, Rybak IA. Multiple rhythmic states in a model of the respiratory central pattern generator. J Neurophysiol 2009;101:2146–65. [22] Smith JC, Abdala AP, Koizumi H, Rybak IA, Paton JF. Spatial and functional architecture of the mammalian brain stem respiratory network: A hierarchy of three oscillatory mechanisms. J Neurophysiol 2007;98:3370–87. [23] Tabak J, Mascagni M, Bertram R. Mechanism for the universal pattern of activity in developing neuronal networks. J Neurophysiol 2010;vol. 103:2208–21. [24] Vasalou C, Henson MA. A multiscale model to investigate circadian rhythmicity of pacemaker neurons in the suprachiasmatic nucleus. PLoS Comput Biol 2010;6

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143 Modeling the Autonomic Nervous System Ilya A. Rybak, Yaroslav I. Molkov, Julian F.R. Paton, Ana P.L. Abdala, Daniel B. Zoccal Sympathetic nerve activity normally exhibits respiratory modulation that suggests the existence of central interactions between the respiratory and sympathetic networks within the brainstem. A large-scale computational model of interacting respiratory and sympathetic circuits has been developed and used to investigate the possible mechanisms of sympatho-respiratory interactions and their role in the baroreceptor reflex control of sympathetic activity and in the elevated sympathetic activity following chronic intermittent hypoxia. Several model predictions have been formulated and tested experimentally. The model provides important insights into the role of sympatho-respiratory interactions in the control of sympathetic outflow and arterial blood pressure under different physiological and patho-physiological conditions.

LARGE-SCALE COMPUTATIONAL MODEL OF THE BRAINSTEM SYMPATHO-RESPIRATORY NETWORK A large-scale computational model of the brainstem sympatho-respiratory network has been developed to simulate the respiratory and sympathetic neural circuits interacting within the brainstem (Fig. 143.1A). The major circuits critically involved in generation of the respiratory rhythm and pattern are located in the ventral respiratory column (VRC) and include (rostral-to-caudal) the Bötzinger (BötC) and pre-Bötzinger (pre-BötC) complexes and the rostral (rVRG) and caudal (cVRG) ventral respiratory groups [1,2]. The core of the respiratory central pattern generator was proposed to include (i) an excitatory pre-inspiratory/inspiratory (pre-I/I) population of neurons with intrinsic bursting properties located in the pre-BötC and (ii) a ring of three mutually inhibiting neural populations: the post-inspiratory (post-I) and augmenting-expiratory (aug-E) populations of BötC, and the early-inspiratory (early-I(1)) population of pre-BötC (Fig. 143.1A, see [2,3]). The respiratory circuitry in the model also incorporates two neural populations within the rVRG, the bulbospinal ramp-inspiratory (ramp-I) neurons, projecting to phrenic motoneurons in the spinal cord that send their axons to the

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00143-8

phrenic nerve (PN), and the inhibitory early-I(2) neurons shaping the firing pattern of ramp-I neurons, and a population of bulbospinal premotor expiratory neurons of cVRG (bs-E) projecting to the abdominal motoneurons that define activity of the abdominal nerve (AbN) (Fig. 143.1A). The sympathetic circuits in the model include neurons located in the rostral (RVLM) and caudal (CVLM) ventrolateral medulla (VLM). Specifically, the RVLM neurons define the activity in the thoracic sympathetic nerve (tSN). In addition, the following populations were included in the model: two populations of 2nd order baroreceptor neurons in the nucleus tractus solitarii (NTS) receiving baroreceptor afferents and a population of phase-spanning inspiratory-expiratory neurons (IE) in the ventrolateral pons. (Fig. 143.1A, see also [4]). The model also incorporates a compartment known as the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG), containing neurons performing central chemoreceptor function whose activity is sensitive to CO2. This compartment includes a population of neurons with intrinsic bursting properties termed the late-expiratory (late-E) population (Fig. 143.1A). The inclusion of this population in the RTN/pFRG is based on the multiple experimental data that the late-E activity emerging in AbN during hypercapnia originates in this region [3,5]. In addition to multiple mutual interactions all respiratory neural populations in the model receive excitatory drives from the pons, RTN/pFRG and raphé [2,3].

MODELING THE EFFECTS OF BARORECEPTOR ACTIVATION ON THE RESPIRATORY PATTERN: INSIGHTS INTO RESPIRATORY–SYMPATHETIC INTERACTIONS The baroreceptor reflex is an important negative feedback mechanism controlling sympathetic outflow. The classical baroreflex controls tSN via 2nd order barosensitive neurons in the NTS that receive the direct excitatory inputs from baroreceptor afferents. It is suggested that excitatory NTS neurons project to CVLM neurons, which

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FIGURE 143.1 The computational model of sympatho-respiratory brainstem network. (A) Schematic of the model showing interactions between different populations of respiratory neurons within major brainstem compartments involved in the control of breathing and sympathetic activity. Abbreviations: brainstem compartments: BötC – Bötzinger complex; CVLM – caudal ventrolateral medulla; cVRG – caudal ventral respiratory group; NTS – nucleus tractus solitarii; pre-BötC – pre-Bötzinger complex; RTN/pFRG – retrotrapezoid nucleus/parafacial respiratory group; RVLM – rostral ventrolateral medulla; rVRG – rostral ventral respiratory group; VLM – ventrolateral medulla; VRC – ventral respiratory column; neural populations: aug-E – augmenting expiratory; early-I – early-inspiratory; IE – phase-spanning inspiratory-expiratory; late-E – late-expiratory; post-I – postinspiratory; post-I(e) – post-inspiratory (excitatory); pre-I/I – pre-inspiratory/inspiratory; ramp-I – ramp-inspiratory; motor outputs: PN – phrenic nerve; AbN – abdominal nerve; tSN – thoracic sympathetic nerve. Keys are shown in the right-bottom corner. Each population (large sphere) consists of 20–50 neurons modeled in the Hodgkin–Huxley style. All tonic drive sources (gray triangles) provide constant drive, except for RTN/pFRG which is CO2-dependent (see panel C). Not all connections from pontine and RTN/pFRG drive sources are shown. For details see [2,3]. (B) Conceptual model of interaction between VRC, PONS, NTS, and RVLM/CVLM. The sympathetic baroreceptor reflex operates via two pathways: one direct pathway (black solid arrows) includes baroreceptors, NTS (2nd order barosensitive cells) and CVLM, which inhibits RVLM and tSN; the other pathway goes from baroreceptors through NTS and VRC (black dashed arrows), whose post-inspiratory neurons inhibit RVLM and tSN. Gray dashed arrows show interactions between PONS and medullary compartments VRC and RVLM/CVLM. (C) RTN/pFRG tonic drive as function of CO2 for the control (solid curve) and CIH-conditioning (dashed curve) cases.

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inhibit RVLM neurons hence lowering the tSN activity (Fig. 143.1B). On the other hand, tSN has an obvious respiratory modulation that persists after vagotomy and decerebration, supporting the idea of central coupling between respiratory and sympathetic networks. In turn, the respiratory activity is known to be modulated by baroreceptor afferents through the same 2nd order barosensitive neurons of NTS. This suggests that central sympathorespiratory interactions may contribute to the dynamic control of sympathetic activity. Specifically, it has been hypothesized [4] that the sympathetic baroreceptor reflex has two major pathways (Fig. 143.1B): one direct path mentioned above, that is independent of the respiratorysympathetic interactions, and the other, operating via the baroreceptor modulation of the respiratory activity and respiratory-sympathetic interactions and hence dependent on the respiratory modulation of tSN activity. At rest, the tSN activity typically exhibits a well expressed positive modulation during inspiration and a negative modulation during post-inspiration (see Fig. 143.2A1–A3 before applied stimulations). The tSN respiratory modulation was significantly suppressed or eliminated after removal of the pons, when phrenic nerve (PN) activity transformed to an apneustic pattern with prolonged inspiratory bursts and shortened expiration durations (see Fig. 143.2A4 before applied stimulation), underlying a critical role of the pons in the tSN respiratory modulation [6]. In Figure 143.2A1–A5, the transient increases in the arterial pressure were induced in the arterially perfused in situ rat preparation [6]. Stimuli were delivered during inspiration, post-inspiration or late expiration and produced phase-dependent effects on both the phrenic nerve (PN) activity and the respiratory modulation of tSN (Fig. 143.2A1–A3). With pons intact, the applied barostimulation had almost no effect on the amplitude and duration of the phrenic bursts even when stimuli were delivered during inspiration (Fig. 143.2A1). At the same time, these stimuli suppressed or abolished inspiratory modulation of tSN activity. In contrast, the same stimuli delivered during post-inspiration (Fig. 143.2A2) or late expiration (Fig. 143.2A3) produced an increase in the expiration period and decreased the tSN activity. The barostimulation-evoked prolongation of expiration was greater if stimulation was applied later during the expiratory phase (compare Figs 143.2A2 and A3). Importantly, after pontine transection the respiratory modulation of tSN activity was greatly reduced [6]. In all cases, however, the sympathetic baroreflexinduced lowering of tSN persisted and the barostimulation shortened the apneustic inspiratory burst (see Fig. 143.2A4). Figure 143.2B1–B4 shows the results of our simulation of the effects of transient barostimulation during different phases of the respiratory cycle using the computational model described above (Fig. 143.1A) before (Fig. 143.2B1– B3) and after (Fig. 143.2B4) removal of the pontine compartment. The model generates a normal three-phase respiratory pattern with augmenting PN bursts (Fig. 143.2A1–A3). The sympathetic output in the model (tSN) exhibits a positive

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inspiratory modulation provided by the pontine IE population to RVLM and a negative post-inspiratory modulation resulting from the inhibitory inputs from the post-I population of BötC to the RVLM. Transient barostimulation applied to the barosensitive 2nd order NTS population produces a temporal reduction of tSN output via direct activation of the CVLM population that inhibits the activity of RVLM population. Stimulus application during inspiration (Fig. 143.2B1) does not affect respiratory (PN) activity. In contrast, stimuli applied during post-inspiration (Fig. 143.2B2) and late expiration (Fig. 143.2B3) prolong expiration via activation of post-I neurons of BötC that inhibit both the aug-E population and the RVLM. These interactions represent a second component of the sympathetic baroreflex involving interactions between the respiratory and sympathetic circuits. Similar to our experimental data (Fig. 143.2A2,A3), stimulation-evoked prolongation of expiration is greater if stimulation is applied later during the expiratory phase (Fig. 143.2B2,B3). Removing the pontine compartment in the model converts the normal eupnea-like respiratory pattern to the apneustic pattern characterized by prolonged PN busts (see Fig. 143.2B4). As demonstrated previously [2], this pattern is characterized by a lack of post-I activity that is strongly dependent on pontine drive. Therefore, with the pons removed, the respiratory modulation of tSN (formed by inputs from pontine IE and BötC’s post-I populations to RVLM) is abolished. Simultaneously, the central suppression of the baroreflex gain by the rVRG early-I(2) population, whose activity in the model is also dependent on the pontine drive, is eliminated with pontine removal. Therefore the applied barostimulation can activate post-I population during inspiration and produce an advanced termination of the apneustic inspiratory bursts hence shortening inspiration (compare Fig. 143.2B4 with Fig. 143.2A4). Figure 143.2B5 illustrates the neural mechanism by which the transient barostimulation applied during expiration prolongs this expiration in the intact model (see Fig. 143.2B3 for comparison). The post-I neurons of BötC when activated inhibit all inspiratory (and aug-E) neurons and initiate the post-inspiratory phase of expiration. During expiration, the activities of these neurons decrement hence releasing aug-E neurons form inhibition and allowing for their gradual activation (see unperturbed breathing cycles in Fig. 143.2B5). When a barostimulus occurs during expiration, the post-I population is activated and inhibits the aug-E population, hence producing a “resetting of expiration”. This resetting of expiration by the transient barostimulation provides a mechanistic explanation for expiratory period prolongation. To test this model prediction, extracellular recordings of post-I and aug-E neurons were made within the BötC of in situ rat preparations [4] (see example in Fig. 143.2A5). With the transient increase in perfusion pressure during expiration, the activity of post-I neurons increased and the activity of aug-E neurons decreased in full accordance with the model prediction (compare Fig. 143.2A5 with Fig. 143.2B5). In most cases,

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FIGURE 143.2 Effects of transient, respiratory phase-dependent baroreceptor stimulation on phrenic (PN) and sympathetic (tSN) nerve activities in situ and in the model. (A1–A5) Experimental results from the arterially perfused in situ preparation A1-A3: Stimulation was applied to the intact preparation during inspiration (A1), post-inspiration (A2) and late expiration (A3). After pontine transection (A4), the applied stimulus shortened the apneustic inspiratory (PN) burst. Traces from top to bottom: integrated sympathetic (tSN) activity, integrated phrenic (PN) activity; perfusion pressure (PP). In A5, top two traces show an extracellular recording from a post-I neuron in BötC and the histogram of its activity; the next pair of traces show a simultaneous extracellular recording from an aug-E neuron of BötC and the corresponding histogram; the remaining traces show the integrated activities of PN and tSN) and perfusion pressure (PP). (B1–B5) Corresponding simulation results. In simulations shown, the stimulus was applied during inspiration (B1), post-inspiration (B2) and late expiration (B3), and also after removal of the pontine compartment in the model (B4). In B5, the top pair of traces show membrane potential of a randomly chosen neuron from the post-I population of BötC and integrated spike histogram of the entire post-I population. Second pair of traces: membrane potential of a randomly chosen neuron from the aug-E population of BötC and spike histogram of the entire aug-E population.

the barostimulation applied during expiration resulted in prolongation of expiration, and this prolongation was greater when stimulation was applied later in expiration. These studies have clarified the role of baroreceptor input in activating post-I neurons and inhibiting aug-E neurons and demonstrated that even weak excitatory input from baroreceptors to the post-I neurons can account for the prolongation of expiration and the corresponding effect on tSN activity. In general, this demonstrates an important contribution of central sympatho-respiratory interactions to the baroreceptor control of arterial pressure.

SYMPATHETIC NERVE ACTIVITY FOLLOWING CHRONIC INTERMITTENT HYPOXIA-INDUCED SENSITIZATION OF CENTRAL CHEMORECEPTORS Recurrent episodes of hypoxia, such as observed in obstructive sleep apnea lead to the development of hypertension. It was shown that rats exposed to chronic intermittent hypoxia (CIH) exhibited higher levels of arterial pressure associated with an elevated sympathetic vasomotor tone [7,8] and an enhanced respiratory modulation

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143.3 Sympathetic and respiratory responses to hypercapnia before and after CIH-conditioning: experimental records and simulations. (A1– A5) Activity of phrenic (PN), abdominal (AbN), and thoracic sympathetic (tSN) nerves in the representative control preparation at 5% CO2 (base level, A3), during hypercapnia (7% CO2 in A4 and 10% CO2 in A5) and during hypocapnia (3% CO2 in A2 and 1% CO2 in A1). Note the skipping of some late-E bursts in both AbN and tSN at 7% CO2 (in A4). (B1–B5) PN, AbN, and tSN activities in the representative CIH-conditioned preparation at 5% CO2 (base level, B3), during hypercapnia (7% CO2 in B2 and 10% CO2 in B3) and hypocapnia (3% CO2 in B2 and 1% CO2 in B1). Note the presence of late-E bursts in both AbN and tSN at 5% CO2 (in B1). The activity of each nerve is represented by raw recording (bottom trace) and integrated activity (upper trace). (C–E) Model performance in simulated control (C) and CIH (E) cases. In both cases, integrated activity for phrenic (PN), abdominal (AbN) and thoracic sympathetic (tSN) outputs are shown. The CO2 level was changed in a step-wise manner from 1% (hypocapnia) to 10% (hypercapnia) which is shown in panel D. The dashed vertical arrows indicate CO2 levels for emerging late-E activity and for hypocapnic apnea, respectively.

of sympathetic activity [7]. This suggests that central coupling between respiratory and sympathetic circuits may contribute to hypertension in CIH-conditioned animals [8]. Typical patterns of respiratory (PN and AbN) and sympathetic (tSN) activities in the naïve rat (arterially perfused in situ preparation) are shown in Fig. 143.3A1–A5.

Under normal conditions (5% CO2), the integrated PN burst has an augmenting profile, AbN shows low-amplitude activity, and the tSN expresses an augmenting inspiratory modulation (Fig. 143.3A3). Hypercapnia (increase in CO2 level from 5%) evokes high-amplitude lateexpiratory (late-E) AbN discharges, which are phase-locked

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143. ModElINg THE AuToNoMIC NERvouS SySTEM

to PN bursts (Fig. 143.3A4,A5; see also [3,5]). With progressive development of hypercapnia, the ratio of AbN late-E burst frequency to PN burst frequency quantally (step-wise) increases from about 1:4/1:3 to 1:2 (at 7% CO2), when approximately each second respiratory cycle was skipping in AbN late-E activity (Fig. 143.3A4), and finally, to 1:1 as the CO2 level increases to 10% (Fig. 143.3A5), when the AbN late-E discharges preceding PN bursts are observed in every respiratory cycle [3]. This CO2-induced AbN late-E activity appears to originate in the RTN/ pFRG region [3,5,9] that is also known to be a major site for central chemoreception [10]. The CO2-censitive RTN/ pFRG neurons project to both VRC and RVLM. The latter is important for the sympatho-excitation and the corresponding increase of arterial pressure observed during hypercapnia. The hypercapnia-evoked late-E activity, quantally accelerated with an increase in CO2, has been observed in tSN (Fig. 143.3A4,A5). Moreover, the tSN lateE activity coincides with the AbN late-E bursts, suggesting a common source of late-E activities in AbN and tSN located the RTN/pFRG. The interesting effect of CIH conditioning is that it alters the AbN and tSN activities in both normocapnia and hypercapnia [7,8]. Figure 143.3B3 shows that the juvenile rats submitted to CIH for 10 days exhibits an expressed late-E activity in both AbN and tSN (with a frequency ratio to PN of about 1:3/1:2) in the baseline conditions (5% CO2), and at 7% CO2 this ratio already reaches 1:1, i.e. full synchronization (Fig. 143.3B4). We suggest that CIH conditioning augment the CO2 sensitivity of RTN/pFRG neurons, and hence reduces CO2 threshold for the emergence of late-E oscillations seen in both AbN and tSN. The computational model shown in Figure 143.1A was used for simulating the effect of CIH-induced sensitization of RTN/pFRG neurons on the respiratory and sympathetic activities and the sympatho-respiratory response to hyperand hypocapnic conditions. The model includes excitatory tonic drives from several sources including RTN/ pFRG that is considered to be a major central chemoreceptor site sensitive to CO2 [10]. In the model, we consider RTN/pFRG tonic drive to be not constant but dependent on the CO2 level as shown in Figure 143.1C (solid curve). As hypothesized above, the CO2 sensitivity of RTN/pFRG increases as a result of CIH exposure. This is simulated by the horizontal shift of the CO2-dependent RTN drive by 2% CO2 to the direction of lower CO2 values (to the left, see dashed curve in the Fig. 143.1C). Figure 143.3C shows the results of our simulations with CO2 step-wise increasing from 1% CO2 (hypocapnia) through 5% CO2 (normocapnia) to 10 % CO2 (hypercapnia) as illustrated in Figure 143.3D. In our simulations, progressive hypercapnia (Fig. 143.3C,D, right part of the graph) leaded to the emergence and quantal acceleration of late-E bursts in both AbN and tSN, which was consistent with experimental records (see panels A3– A5). Specifically, the late-E discharges in AbN and tSN

appeared at 7% CO2 and reached 1:1 ratio to the PN bursts at 9% CO2. Note also (see Fig. 143.3C,D, left part) that a reduction of CO2 below 3% caused “hypocapnic apnea” (a lack of PN activity). To simulate CIH conditions, the curve reflecting the CO2 dependence of RTN/pFRG drive was shifted to the left (Fig. 143.1C). As a result of this shifting, the late-E bursts in AbN and tSN emerged at 4% CO2, and in the normocapnic state (5% CO2) they showed a stable 1:2 ratio to the PN bursts (Fig. 143.3E); at 7% CO2 this ratio reached 1:1 (Fig. 143.3B4), which was consistent with our experimental observations (see Fig. 143.3B). The second observation from the above simulation is that “CIH conditioning” reduced the apneic threshold for hypocapnia by at least 2% CO2, since the PN bursts were still generated even at 1% CO2 (see Fig. 143.3E, left). To check this modeling prediction, the control (naïve) and CIH-conditioned rat preparations were exposed to progressive hypocapnia (from normal 5% CO2 to 3% and then to 1%). The naïve rat preparations exhibited a reduction in the integrated PN burst amplitude at 3% CO2 and a hypocapnic apnea at 1% CO2 (Fig. 143.3A2,A1). Importantly, these preparations never expressed late-E activity in AbN or tSN in either normocapnia or hypocapnia, and the respiratory modulation of tSN was reduced at 3% CO2 and absent at 1% during hypocapnic apnea (Fig. 143.3B2,B1). In CIH rat preparations, the expressed late-E activity in both AbN and tSN was already present during normocapnia (at 5% CO2) and disappeared from both nerves at 3% CO2 (Fig. 143.3B2). At the same time, PN activity with a reduced amplitude (and respiratory modulation of tSN) was still present even at 1% CO2 (Fig. 143.3B1), hence confirming modeling prediction on a reduction of apneic threshold for hypocapnia in CIH-conditioned rats. Our multidisciplinary investigation suggests that the arterial blood pressure elevation associated with CIH may result from an increased CO2 sensitivity of central chemoreceptors and early emergence of late-E oscillations in the RTN/pFRG.

References [1] Alheid GF, McCrimmon DR. The chemical neuroanatomy of breathing. Respir Physiol Neurobiol 2008;164:3–11. [2] Smith JC, Abdala APL, Koizumi H, Rybak IA, Paton JFR. Spatial and functional architecture of the mammalian brain stem respiratory network: a hierarchy of three oscillatory mechanisms. J Neurophysiol 2007;98:3370–87. [3] Molkov YI, Abdala APL, Bacak BJ, Smith JC, Paton JFR, Rybak IA. Late-expiratory activity: Emergence and interactions with the respiratory CPG. J Neurophysiol 2010;104:2713–29. [4] Baekey DM, Molkov YI, Paton JFR, Rybak IA, Dick TE. Effect of baroreceptor stimulation on the respiratory pattern: Insights into respiratory-sympathetic interactions. Respir Physiol Neurobiol 2010;174:135–45. [5] Abdala APL, Rybak IA, Smith JC, Paton JFR. Abdominal expiratory activity in the rat brainstem-spinal cord in situ: Patterns, origins and implications for respiratory rhythm generation. J Physiol 2009;587:3539–59.

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SyMPATHETIC NERvE ACTIvITy FollowINg CHRoNIC INTERMITTENT HyPoxIA-INduCEd SENSITIzATIoN oF CENTRAl CHEMoRECEPToRS

[6] Baekey DM, Dick TE, Paton JFR. Pontomedullary transection attenuates central respiratory modulation of sympathetic discharge, heart rate and the baroreceptor reflex in the in situ rat preparation. Exp Physiol 2008;93:803–16. [7] Zoccal DB, Simms AE, Bonagamba LG, Braga VA, Pickering AE, Paton JFR, Machado BH. Increased sympathetic outflow in juvenile rats submitted to chronic intermittent hypoxia correlates with enhanced expiratory activity. J Physiol 2008;586:3253–65. [8] Zoccal DB, Paton JFR, Machado BH. Do changes in the coupling between respiratory and sympathetic activities contribute

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to neurogenic hypertension?. Clin Exp Pharmacol Physiol 2009;36:1188–96. [9] Janczewski WA, Onimaru H, Homma I, Feldman JL. Opioidresistant respiratory pathway from the preinspiratory neurones to abdominal muscles: In vivo and in vitro study in the newborn rat. J Physiol 2002;545:1017–26. [10] Guyenet PG, Stornetta RL, Bayliss DA. Retrotrapezoid nucleus and central chemoreception. J Physiol 2008;586:2043–8.

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144 Optogenetics Sergey Kasparov Optogenetic Reporters for Ca2

INTRODUCTION Central regulation of autonomic outflows is a highly complex process coordinated by a network of neurons distributed throughout the whole extent of the brainstem. The spectrum of neuronal phenotypes involved in various autonomic responses is remarkably diverse and includes glutamatergic, GABA-ergic, acetylcholinergic, peptidergic, catecholaminergic, serotonergic and others. In many cases it is essential to be able to selectively study and manipulate these diverse groups of cells in order to reveal their respective contributions to the physiological process in question. Optogenetics provides a unique opportunity for this type of enquiries. The term “optogenetics” means experimentation using a combination of genetic manipulation and optics. Genetic engineering enables targeted expression of reporters and effectors (or actuators) in mammalian cells. Optogenetics can be used for studies of the autonomic areas of the brain and provides tools for specific control of phenotypically identified groups of neurons and more recently, astrocytes.

OPTOGENETIC REPORTERS Optical monitoring of various processes in cells of interest is advantageous compared to electrical recordings because it is non-invasive, allows much better spatial resolution, not limited to electrogenic processes and typically provides information from more than one cell, thus greatly increasing the throughput. Optogenetic reporters can be easily used in vitro in brain slices. Their in vivo application for studies of the autonomic centers of the brain has been hampered by the poor optical access to these areas which are all located in deep brain structures. Nevertheless, in vivo application of this approach is possible [1]. Multiphoton excitation is of little help here because its depth of penetration (typically 100–150 μm) is only sufficient for the upper layers of cortex. In the near future fluorescent imaging of the autonomic areas will be facilitated by the introduction of new fiber-based microscopes which use needle-like “objectives” and reveal fluorescence in any brain structure irrespective of its depth. Optogenetic reporters can also be effectively used in slices and slice cultures which contain autonomic nuclei [2,3].

Primer on the Autonomic Nervous System. DOI: 10.1016/B978-0-12-386525-0.00144-X

Genetically encoded calcium indicators (GECI) consist of one or two fluorescent protein(s) (FP) and a Ca2sensitive domain. GECI respond to Ca2 by altering their fluorescence intensity or by a wavelength shift. The most common single GFP-based biosensor platform relies on circularly permuted GFP. The N and C-termini of these GECI contain “sticky ends” – the Ca2 binding motif from calmodulin and its target binding protein, M13 (derived from myosin light chain kinase) which are attracted in the presence of Ca2. The most recently published GECI of that family is GCaMP3, which has an improved dynamic range, good maturation in mammalian neurons and is suitable for in vivo Ca2 imaging [4]. Another cpGFP indicator – Case 12 [5] has an exceptionally high contrast ratio (12 times) and proven very effective for imaging of [Ca2]i in astrocytes in vitro [3] and in vivo [1]. Förster resonance energy transfer (FRET) is a different principle applied to design of GECI. In this design two fluorescent proteins are used, typically cyan and yellow and Ca2 binding results in a spectral shift [6,7]. The recent constructs from this subfamily were reported to detect Ca2 transients triggered by single action potentials [8]. In combination with appropriate genetic targeting these and other newer probes will allow enquiries into the Ca2 signaling in identified populations of cells located in the autonomic centers of the brain. Optical monitoring using genetic probes is by no means limited to studies of [Ca2]i concentration. Optogenetic sensors for cAMP, Cl and other important indicators of cellular activity have been developed. Probes for membrane potential are of particular interest although their dynamic range is still limited [9].

Optogenetic Effector Proteins Controlling activity of neurons and astrocytes using optogenetic effectors is a new and very exciting technology which circumvents the limitations of all the previous methods used for the same purpose. In contrast to electrical current, light will only affect cells which specifically express optogenetic effectors and this will have no effect on other cells even if they are located nearby or on the fibers en-passant. Many optogenetic effectors operate with millisecond precision and therefore permit a high degree

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of temporal control over the activity of the neurons under study. Light-Sensitive Cation Channel Channelrhodopsin-2 and its Derivatives Channelrhodopsins (ChRs) are the only currently known family of ion channels that are directly gated by light. These proteins have seven transmembrane spanning domains and have been cloned from algae which use them for detection of light. ChRs are non-selective cation channels permeable to Na, K and Ca2 and when opened upon illumination depolarize the membrane [10]. ChRs have been extensively used in neuroscience as tools to depolarize neuronal membranes but they can be used to control other excitable cells such as cardiac myocytes or skeletal muscles. Presently used variants of ChR2 differ in their kinetics, dynamics of inactivation and recovery, channel conductance, spectral characteristics and membrane trafficking [11]. In almost all studies they are used as fusions with a fluorescent protein, such as yellow or red fluorescent protein to assist visualization [12]. Fusions with red fluorescent proteins are advantageous because they are visualized using green or yellow light to which ChRs are only minimally sensitive thus avoiding an unsolicited activation of ChRs. Because of the relatively modest conductance of ChRs, their successful application requires high level of expression in target cells which needs to be taken into account and controlled for its non-specific effects. Nevertheless, it proves a means to optogenetically control the excitability of various populations of neurons in vitro and in vivo [13,14]. Optogenetic excitation of the retrotrapezoid nucleus has recently been used to address its role in respiratory control [15]. Expression of ChR2 in astrocytes allows one to induce increases in [Ca2]i in these cells, the effect which has recently been used to demonstrate their involvement in central chemosensitivity [1]. All currently used variants of ChR are most sensitive to blue light (450–480 nm) which can be delivered from pulsing sources of light to deep structures via “optrodes” (Fig. 144.1). High levels of light (more than a few mW) and especially, continuous (not pulsing) light can evoke artifacts in vitro and in vivo and it is therefore essential to carefully control the power [14]. Optogenetic “Silencers” Two families of genes are currently being used for “silencing” of neurons with light on demand. The first one is the microbial halorhodopsin NpHR, a fast lightactivated electrogenic Cl pump and its mutants and derivatives [16]. The second is the light-activated proton pump “Arch” from Halorubrum sodomense [17]. Both genes are able to significantly hyperpolarize neurons and within milliseconds curtail their action potential activity. Interestingly, both NpHR and Arch are sensitive to yellow/orange light (550–590 nm) which makes it possible to combine them with ChR in the same experiment.

FIGURE 144.1 Optogenetic stimulation of a brainstem autonomic center combined with chronic direct measurement of arterial blood pressure in a rat. Viral vectors were used to introduce ChR into one of the autonomic nuclei of the brainstem. The animal was then implanted with a light guide (optrode) which is mounted into the skull and connected to the black light guide. Light guide is connected to a solid state pulsing laser controlled by a computer (not shown). Additionally, the rat was implanted with an arterial catheter (exteriorized on its back) to monitor blood pressure directly. The animal is kept in a small box to limit its activity and the light guide and catheter are kept out of its sight to prevent damage. This approach can be used to study the roles of diverse neuronal populations in acute and chronic control of blood pressure. Photo supplied by Dr. A.P.L. Abdala.

Light-Sensitive G-protein Coupled Receptors (GPCRs) Many essential processes in the autonomic centers of the brain are mediated via receptors coupled via G-proteins. Moreover, all known receptors for noradrenaline and the “M” type receptors for acetylcholine are GPCRs. These receptors belong to rhodopsin superfamily and have the same 7-transmembrane topology. This closeness has allowed generation of chimeras where the extracellular and membrane-spanning part of the receptor is taken from rhodopsin and the intracellular part from one of the “classic” G-protein coupled receptors, such as α1 or β2 adrenergic receptors [18]. These interesting optogenetic tools enable control of intracellular signaling by mimicking G-protein-mediated events. Using “optoGPCR” it is now possible to “switch on” with light all three main pathways employed by these ubiquitous regulators of cellular metabolism, including the signaling cascades affected by GαS, GαQ and GαI subunits [19]. It is difficult to overestimate the potential use of this technology for studies into the roles of specified populations of neurons and astrocytes in the autonomic nuclei of the brain.

MEANS OF GENE DELIVERY Successful gene delivery is the key to application of optogenetics. There are two basic technologies which can

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COnClusIOn

be used for that purpose: viral gene delivery and germline transgenesis (almost exclusively mice at present). Both approaches can also be combined.

Cell-Specific Viral Targeting Viral vectors are extremely powerful and versatile scientific tools. Their advantages are speed and flexibility, fairly low cost, applicability to any strain of rodents, ease of sharing between investigators. Targeting specified cell types with viral vectors always relies on isolation of a relatively short (typically 1–3 kilobases) DNA sequence which may be used as a cell-specific promoter. In some cases this approach has been highly successful, while for some cell types no such short promoters are known. The most commonly used types of vectors at present are lentiviral vectors (usually derived from HIV), adeno-associated virus vectors and adenoviral vectors [2].

Germline Transgenics Review of transgenic technology is outside of the scope of this chapter. Briefly, transgenic animals may also have cassettes with short promoters driving optogenetic transgenes incorporated in their genomes. However, for the most specific targeting bacterial artificial chromosomes (BAC) are used, because this allows incorporation of the whole transcriptional unit with all the elements potentially important for the correct expression of the transgene. BAC can either directly control expression of any of the optogenetic tools mentioned above or express Cre recombinase to allow activation of the target genes only in specified cells. This last approach is favored at present because it is highly flexible and usually gives higher level of gene expression as well as site-specificity.

CONCLUSION Almost all areas of the brain implicated in autonomic control contain heterogeneous populations of neurons with diverse and in some cases opposing functions, connectivity and activity patterns. In addition, astroglia have recently been recognized as another potentially important component of central information processing machinery. Optogenetics offers new opportunities for selective studies of genetically targetable autonomic cells in vitro and in vivo. The range of available optogenetic tools is rapidly expanding.

Acknowledgements

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References [1] Gourine AV, Kasymov V, Marina N, Tang F, Figueiredo MF, Lane S, et al. Astrocytes control breathing through pH-dependent release of ATP. Science 2010;329:571–5. [2] Teschemacher AG, Paton JFR, Kasparov S. Imaging living central neurones using viral gene transfer. Adv Drug Deliv Rev 2005;57:79–93. [3] Guo F, Liu B, Tang F, Lane S, Souslova EA, Chudakov DM, et al. Astroglia are a possible cellular substrate of angiotensin(1-7) effects in the rostral ventrolateral medulla. Cardiovasc Res 2010;87:578–84. [4] Tian L, Hires SA, Mao T, Huber D, Chiappe ME, Chalasani SH, et al. Imaging neural activity in worms, flies and mice with improved GCaMP calcium indicators. Nat Methods 2009;6:875–81. [5] Souslova EA, Belousov VV, Lock JG, Strömblad S, Kasparov S, Bolshakov AP, et al. Single fluorescent protein-based Ca2  sensors with increased dynamic range. BMC Biotechnol 2007;7:37. [6] Lutcke H, Murayama M, Hahn T, Margolis DJ, Astori S, Zum Alten Borgloh SM, et al. Optical recording of neuronal activity with a genetically-encoded calcium indicator in anesthetized and freely moving mice. Front Neural Circuits 2010;4:9. [7] Mank M, Santos AF, Direnberger S, Mrsic-Flogel TD, Hofer SB, Stein V, et al. A genetically encoded calcium indicator for chronic in vivo two-photon imaging. Nat Methods 2008;5:805–11. [8] Horikawa K, Yamada Y, Matsuda T, Kobayashi K, Hashimoto M, Matsu-Ura T, et al. Spontaneous network activity visualized by ultrasensitive Ca(2) indicators, yellow Cameleon-Nano. Nat Methods 2010;7:729–32. [9] Mutoh H, Perron A, Akemann W, Iwamoto Y, Knopfel T. Optogenetic monitoring of membrane potentials. Exp Physiol 2010;96:13–8. [10] Nagel G, Szellas T, Huhn W, Kateriya S, Adeishvili N, Berthold P, et al. Channelrhodopsin-2, a directly light-gated cation-selective membrane channel. Proc Natl Acad Sci U S A 2003;100:13940–13945. [11] Lin JY. A user’s guide to channelrhodopsin variants: features, limitations and future developments. Exp Physiol 2010;96:19–25. [12] Figueiredo M, Lane S, Tang F, Liu B, Hewinson J, Marina NG, et al. Optogenetic experimentation on astrocytes. Exp Physiol 2010;96:40–50. [13] Gradinaru V, Zhang F, Ramakrishnan C, Mattis J, Prakash R, Diester I, et al. Molecular and cellular approaches for diversifying and extending optogenetics. Cell 2010;141:154–65. [14] Zhang F, Gradinaru V, Adamantidis AR, Durand R, Airan RD, de LL, et al. Optogenetic interrogation of neural circuits: Technology for probing mammalian brain structures. Nat Protoc 2010;5:439–56. [15] Abbott SB, Stornetta RL, Fortuna MG, Depuy SD, West GH, Harris TE, et al. Photostimulation of retrotrapezoid nucleus phox2bexpressing neurons in vivo produces long-lasting activation of breathing in rats. J Neurosci 2009;29:5806–19. [16] Gradinaru V, Thompson KR, Deisseroth K. eNpHR: a Natronomonas halorhodopsin enhanced for optogenetic applications. Brain Cell Biol 2008;36:129–39. [17] Chow BY, Han X, Dobry AS, Qian X, Chuong AS, Li M, et al. Highperformance genetically targetable optical neural silencing by lightdriven proton pumps. Nature 2010;463:98–102. [18] Airan RD, Thompson KR, Fenno LE, Bernstein H, Deisseroth K. Temporally precise in vivo control of intracellular signalling. Nature 2009;458:1025–9. [19] Masseck OA, Rubelowski JM, Spoida K, Herlitze S. Light- and drug-activated G-protein coupled receptors to control intracellular signaling. Exp Physiol 2010;96:51–96.

Financial support from the British Heart Foundation and Royal Society is gratefully acknowledged.

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Index

A AA, see Arachidonic acid AAADC, see Aromatic amino acid decarboxylase AAG, see Autoimmune autonomic gangliopathy Abdominal pain, chronic, 598–599 Acarbose, postprandial hypotension management, 639–641 Acetylcholine (ACh) airway control, 202 autonomic neuron development role, 7 axon reflex, 409 cotransmission, 27, 29–32 metabolism, 71, 75, 631 synthesis, 71 transporters, 71 Acetylcholine receptor (AChR) autoimmunity, see Autoimmune autonomic gangliopathy; Paraneoplastic autoimmune dysfunction muscarinic receptors agonists, 76–77 antagonists, 77 overview, 71–72, 76 nicotinic receptors electrophysiology of activation, 80–81 overview, 72 structure, 79–82 subtypes, 79–80 optogenetics, 688 pharmacology, 72–73 tissue distribution, 75–76, 81 Acetylcholinesterase (AChE) functional overview, 631 inhibitors, see also Pyridostigmine carbamates, 632 organophosphates, 633 phenanthrene, 632 piperidines, 632 quaternary amines, 631–632 therapeutic applications, 633 ACh, see Acetylcholine AChE, see Acetylcholinesterase AChR, see Acetylcholine receptor ACTH, see Adrenocorticotrophic hormone Acupuncture clinical role, 656 neurological substrate, 653–655 principles, 653 prospects for study, 657 Western understanding, 653 AD, see Alzheimer's disease Adaptive immunity autonomic nervous system effects on cells, 326–328 overview, 325–326

Adenosine cardiovascular effects in nucleus of the solitary tract, 142 cardiovascular autonomic regulation, 96–97 central autonomic regulation, 96 formation, 95 metabolism, 95 neuroexcitatory actions, 96 postsynaptic anti-adrenergic effects, 95 presynaptic effects, 95–96 Adenosine receptor central autonomic regulation, 96 pharmacology, 91 ADHD, see Attention deficit hyperactivity disorder Adrenal gland, see also Pheochromocytoma autonomic control of adrenocortical function, 573 crosstalk between cortex and medulla, 571 diabetic autonomic dysfunction effects, 479 insufficiency management, 572 primary, 571–572 secondary, 572–573 sympathoadrenal axis, 20–21 Adrenaline, see Epinephrine α1-Adrenergic receptor functions, 52–54 ligand binding and activation, 51–52 regulation, 54 signaling, 54 structure, 51, 53 subtypes, 51–52 α2-Adrenergic receptor gene polymorphisms, 56–57 ligands, 55 protein–protein interactions, 56–57 signaling, 55–56 subtypes, 55–56 trafficking, 55–56 β-Adrenergic receptor aging and cardiac receptor response, 272–273 desensitization, 60 gene polymorphisms, 61 metabolism regulation, 253–254 signaling, 59–60 structure, 60 subtypes, 59 therapeutic targeting beta-agonists, 60–61 beta-blockers, 60 tissue distribution, 59 Adrenocorticotrophic hormone (ACTH), adrenal insufficiency, 571–573 Adrenomedullin (AM) apoptosis inhibition, 129

693

inflammation attenuation, 128–129 oxidative stress attenuation, 128 receptor and signaling, 128 structure, 127 synthesis, 127 tissue distribution, 127 Aging cardiac β-adrenergic receptor response, 272–273 cardiac baroreflex function effects, 271–272 cerebral autoregulation effects, 273 integration of autonomic control networks, 272 neurotransmitter changes, 272 parasympathetic activity response, 272 sympathetic activity response, 272 Airway innervation bronchodilator nerves inhibitory nerves, 203 sympathetic nerves, 202 comparative biology of pulmonary circulation autonomic regulation, 670–671 nerve types afferent nerves, 201 cholinergic efferents, 202 cholinergic nerves, 202 overview, 201 neuropeptides, 203 neurotransmitters, 201 pathology, 203 Aldosterone hypertension and excess, 117–118 hypoaldosteronism, 573 kidney effects, 117 receptor, see Mineralocorticoid receptor salt-sensitivity of blood pressure role, 314, 316 tissue injury mediation, 119 Altitude illness acute hypoxia effects, 281–282 chronic hypoxia effects, 282 high altitude cerebral and pulmonary edema, 281–282 Alzheimer's disease (AD), α-synuclein in pathology, 303 AM, see Adrenomedullin Ambenonium, 631–632 γ-Aminobutyric acid (GABA) receptors GABAA signaling, 109–110 subtypes, 110 therapeutic targeting, 110–111 GABAB, 109 synthesis, 109 transporters, 109

694 α-Amino-3-hydroxyl-5-methyl-4-isoxazolepropionate (AMPA) receptor, functional overview, 104 AMPA receptor, see -Amino-3-hydroxyl5-methyl-4-isoxazole-propionate receptor Amygdala, autonomic function, 9 Amyloidosis hereditary amyloidosis autonomic function testing, 486 diagnosis, 486 gene mutations, 486 pathogenesis, 486 prognosis, 486–487 treatment, 486–487 immunoglobulin amyloidosis diagnosis, 484–485 pathogenesis, 483 peripheral neuropathy, 483 prognosis, 485 treatment, 485 overview, 483 reactive amyloidosis, 486 Anemia, autonomic failure association, 643 Anesthesia, autonomic dysfunction of patients airway management, 664–665 assessment, 663–664 diabetic neuropathy, 665 infection response, 665 lidocaine and hepatic blood flow, 665 pathophysiology, 663 preoperative management, 664 temperature dysregulation, 665 uremia, 666 ventilatory management, 665 Angiotensin, see Renin-angiotensin system Anhidrosis distal anhidrosis, 556 focal anhidrosis, 557 global anhidrosis, 556–557 hemianhidrosis, 557 overview, 554–556 segmental anhidrosis, 557 treatment of heat intolerance and heat stroke, 559 Anterior cingulate cortex, autonomic function, 9 Anxiety disorder, autonomic function, 292 AP2, autonomic nervous system development and specification role, 5–6 Apraclonidine, pupil response, 240 Aquaporin, vasopressin regulation, 122 Arachidonic acid (AA), metabolites in saltsensitivity of blood pressure, 317 Arecoline, pupil response, 240 Arginine vasopressin (AVP) aquaporin regulation, 122 disorders of body water homeostasis, 122–126 orthostatic stress response, 195 receptor types, 121 secretion regulation, 121 Aromatic amino acid decarboxylase (LAAAD) deficiency clinical presentation, 427 diagnosis, 429 treatment, 429 functional overview, 38, 427

Index

functions, 427 gene therapy, 46–47 metabolomics, 42 Arrector pilorum muscle, innervation, 414 β-Arrestin, dopamine receptor signaling, 69 Asthma, neural control of airways, 203–204 Atomoxetine efficacy, 629–630 mechanism of action, 628–629 ATP cotransmission, 27–32 metabolism, 88 overview of neurotransmission, 87–89 receptors, see Purinergic receptors ATP7A, Menkes disease mutations, 435 Atrial fibrillation, obstructive sleep apnea association, 567–568 Atrioventricular (AV) node, 178–179, 181 Atropine, pupil response, 239 Attention deficit hyperactivity disorder (ADHD), norepinephrine transporter polymorphisms, 442 Autoimmune autonomic gangliopathy (AAG) clinical features, 489 course, 490–491 diagnosis, 490 pathogenesis, 489–490 Autonomic neuron, see Neuron, autonomic AV node, see Atrioventricular node AVP, see Arginine vasopressin Axon reflex, testing, 409–411

B Baroreceptor reflex, see also Bionic baroreflex system aging and cardiac baroreflex function, 271–272 arterial baroreflex, 161–162 baroreceptor activity determinants large artery compliance, 163 neuronal mechanisms mediating sensory transduction, 163 rate sensitivity, 161, 163 baroreflex sensitivity blood pressure control, 164 cardiovascular risk with decrease, 164 genetic determinants, 164, 167–169 heart rate control, 163–164 therapeutic targeting, 164–165 cardiopulmonary baroreflex, 161–162 failure autonomic failure comparison, 349–350 causes, 349 clinical presentation, 349–351 diagnosis, 351–352 treatment, 352 hypertension and baroreflex adaptation/ resetting, 163 modeling of baroreceptor activation effects on respiratory pattern, 679, 681–682 pregnancy and impairment, 265–267 stimulator therapy for hypertension, 650 BAT, see Brown adipose tissue BBS, see Bionic baroreflex system Benign joint hypermobility syndrome, see Joint hypermobility Beta-blockers, see β-Adrenergic receptor BH4, see Tetrahydrobiopterin

Bile, autonomic control of secretion, 205 Bionic baroreflex system (BBS) artificial vasomotor center algorithm, 660 efficacy, 660–661 epidural catheter approach, 661–662 implantable device, 662 rationale, 659 spinal cord injury application, 662 theory, 659–660 Biopsy, see Skin biopsy BK potassium channel, polymorphisms and baroreflex function, 168 Bladder, see Lower urinary tract Bladder pain syndrome (BPS), stress effects, 25 Blood pressure (BP), see also Baroreceptor reflex; Hypertension; Salt-sensitivity of blood pressure aging effects, 272–273 autonomic dysfunction evaluation, 377, 380 emotion studies, 297–298 head-up tilt table testing monitoring, 383 high frequency oscillation, 151–154 power spectrum analysis, 405–407 variability patterns baroreflex disorders, 357 body temperature, 356 diurnal rhythms, 355–356 emotional states, 356 exercise, 356–357 food intake and postprandial hypotension, 356 intraoperative variation, 357 medication effects, 357 postural adaptation, 356 respiratory variation, 355 salt sensitivity, 357 smoking effects, 357 white coat hypertension, 356 sympathetic nervous system outflow control, 355 Blood vessel comparative biology of innervation, 669–670 innervation, 414 BMPs, see Bone morphogenetic proteins BOLD signal, see Functional magnetic resonance imaging Bone anatomy, 257 muscle sympathetic nerve activity findings in loss, 396–397 ontogeny, 257 pathology, 258–269 remodeling effects, 257–258 sympathetic innervation Bone morphogenetic proteins (BMPs), autonomic neuron development role, 4 Bosentan, hypertension trials, 137 Botulinum toxin overactive bladder syndrome management, 228 pupil response, 239 BP, see Blood pressure BPS, see Bladder pain syndrome BQ-788, blood pressure response, 137 Bradykinin receptor, polymorphisms and baroreflex function, 168

Index

Brain autonomic output parasympathetic outputs, 11–12 sympathetic preganglionic units, 11 brainstem components in autonomic control caudal raphe nuclei, 11 caudal ventrolateral medulla, 11 functional magnetic resonance imaging, 13–16 nucleus of the solitary tract, 11 parabrachial complex, 11 periaqueductal gray, 10–11 rostral ventrolateral medulla, 11 circulation, see Cerebral blood flow computational modeling of brainstem sympatho-respiratory network, 679–680 forebrain components in autonomic control amygdala, 9 anterior cingulate cortex, 9 hypothalamus, 9–10 insular cortex, 9 leptin receptors, 132–133 Bromocriptine, neuroleptic malignant syndrome management, 543 Brown adipose tissue (BAT), thermoregulation, 243 BuChE, see Butylcholinesterase Butylcholinesterase (BuChE), functional overview, 631

C Calcitonin gene-related peptide (CGRP) airway control, 203 apoptosis inhibition, 129 cotransmission, 29 inflammation attenuation, 128–129 migraine role, 545 oxidative stress attenuation, 128 receptor and signaling, 128 release, 127–128 structure, 127 synthesis, 127 tissue distribution, 127 vasomotor control, 187, 190 Calcium, fluorescent genetic reporters, 687 CAN, see Cardiac autonomic neuropathy; Central autonomic nucleus Capacitance vessels, orthostatic reflex adjustment, 194–195 Carbachol, pupil response, 240 Carbon monoxide (CO), carotid body hypoxic sensing, 332 Carcinoid tumor carcinoid syndrome features, 589–590 epidemiology, 589 serotonin metabolism, 590 sites, 589 treatment, 590–591 Cardiac autonomic neuropathy (CAN), diabetic autonomic dysfunction, 476, 479–480 Cardiac norepinephrine spillover (CNES), heart failure studies, 367–369 Cardiac vagal ganglia abundance, 182 atrial ganglionated plexuses, 182 autonomic nervous system integration, 183–185

ventral ganglionated plexuses, 182–183 Cardiovascular-respiratory coupling, see Respiratory-cardiovascular coupling Carotid body gas messengers in hypoxic sensing, 332 hypoxic stimulus transduction, 331–332 molecular determinants of oxygen sensing, 332–333 morphology, 331 pathology, 333 unique aspects of oxygen sensing, 331 Carotid sinus hypersensitivity, 357 Carpal tunnel syndrome (CTS), amyloidosis, 483 Catechol-O-methyltransferase (COMT), functional overview, 41–42 Caudal pressor area (CPA), functional magnetic resonance imaging, 16 Caudal raphe nuclei, autonomic function, 11 Caudal ventrolateral medulla (CVLM) autonomic function, 11, 13 functional magnetic resonance imaging, 15–16 CBF, see Cerebral blood flow CCHS, see Congenital central hypoventilation syndrome CCK, see Cholecystokinin Central autonomic nucleus (CAN), sympathetic nervous system, 17 Central sleep apnea, see Sleep apnea Cerebral blood flow (CBF) aging effects, 273 autonomic innervation, 199 autoregulation, 197–199 imaging, 197 neurovascular coupling, 197 CFS, see Chronic fatigue syndrome CGRP, see Calcitonin gene-related peptide Channelrhodopsins, optogenetics, 688 Charcot–Marie–Tooth disease (CMT), autonomic dysfunction, 498 ChAT, see Choline acetyltransferase Cholecystokinin (CCK) bile secretion regulation, 205 pancreatic secretion regulation, 205 Choline acetyltransferase (ChAT), functional overview, 71 Chronic fatigue syndrome (CFS) autonomic dysfunction overview, 531 pathophysiology, 533 clinical presentation, 531 definition, 531 diagnosis, 531 epidemiology, 531 neurally mediated hypotension, 533 orthostatic intolerance, 531–532 postural tachycardia syndrome, 533 Chronic inflammatory demyelinating polyradiculopathy (CIDP), sympathetic microneurography, 393 Chronic kidney disease, muscle sympathetic nerve activity findings, 395–396 Chronic obstructive pulmonary disease (COPD), neural control of airways, 204 CIDP, see Chronic inflammatory demyelinating polyradiculopathy

695 Circadian rhythm cardiac events, 159 control of autonomic nervous system, 158–159 suprachiasmatic nuclei output and autonomic control, 157–158 CISS1, see Cold-induced sweating type 1 Clonidine adverse effects, 624 baroreflex failure management, 352 dosing, 624 mechanism of action, 624 CMT, see Charcot–Marie–Tooth disease CNES, see Cardiac norepinephrine spillover CO, see Carbon monoxide Cocaine autonomic effects on heart, 577–578 overdose management, 580 peripheral circulation effects, 577 pupil response, 240 thermoregulation effects, 579 Cold-induced sweating type 1 (CISS1), skin biopsy, 415 Colon, see Intestine Comparative biology, autonomic nervous system in vertebrates cardiovascular response to pressure alterations, exercise, and hypoxia, 670 cardiovascular system anatomy, 668–669 heart autonomic regulation, 669 overview, 667–668 pulmonary circulation autonomic regulation, 670–671 vasculature innervation, 669–670 Complex regional pain syndrome (CRPS) diagnosis, 381 edema, 584–585 inflammation, 585 initiating events, 586 skin sympathetic system cutaneous vasoconstrictor neurons and blood flow through skin, 583 sudomotor neurons and sweating, 584 sympathetically maintained pain, 586 trophic changes, 585 type I central changes, 584 peripheral changes, 584–585 somatomotor changes, 585–586 type II, 583 COMT, see Catechol-O-methyltransferase Congenital central hypoventilation syndrome (CCHS) gene mutation, see PHOX2B genotype–phenotype correlations cardiac asystole, 446 continuous ventilatory dependence, 446 facial dysmorphology, 447 frameshift mutations, 447 Hirschsprung disease, 446 later-onset disease, 447–448 tumors of neural crest origin, 446 management, 448–449 COPD, see Chronic obstructive pulmonary disease Coronary artery spasm, stress cardiomyopathy, 371–373

696

Index

Cotransmission cardiac neurons, 30 central nervous system, 27 enteric nervous system, 29–30 history of study, 27 parasympathetic nerves, 28–29 peripheral nervous system, 27 physiological significance different firing patterns, 30 excitation and inhibition, 32 false cotransmitters, 32 negative cross-talk, 32 neuromodulation, 30–31 neuropeptides, 32 postjunctional cell specificity, 30 synergism, 31 trophic factors, 32 plasticity, 32 sensory motor nerves, 29 sympathetic nerves, 27–28 CPA, see Caudal pressor area CRPS, see Complex regional pain syndrome CTS, see Carpal tunnel syndrome Cutaneous vasoconstriction (CVC), thermoregulation, 243–246 CVC, see Cutaneous vasoconstriction CVLM, see Caudal ventrolateral medulla CVS, see Cyclic vomiting syndrome Cyclic vomiting syndrome (CVS), 597–598 Cyclopentolate, pupil response, 239

D DA, see Dopamine Dantrolene, neuroleptic malignant syndrome management, 543 Darusentan, hypertension trials, 137 DBH, see Dopamine β-hydroxylase DCN, see Dorsal commissural nucleus Defecation, motor control, 209 Dementia with Lewy bodies (DLB) clinical features, 463 differential diagnosis, 463 management dementia, 465 dysautonomia, 465 hallucinations and psychosis, 465 overview, 463–464 Parkinsonism, 465 Detrusor overactivity (DO), incontinence, 231–232 DHPG, see Dihydroxyphenylglycol DI, see Diabetes insipidus Diabetes insipidus (DI), types and pathogenesis, 124 Diabetic autonomic dysfunction adrenal function, 479 bladder dysfunction, 478–479 cardiac autonomic neuropathy, 476, 479–480 colon dysfunction, 478 diabetic neuropathy and anesthesia precautions, 665 erectile dysfunction, 479 gallstones, 478 gastric emptying, 476–477 pupil findings, 476 sudomotor function, 479 vaginal function, 479

Dihydroxyphenylglycol (DHPG), biomarker of sympathetic innervation and function, 42 DLB, see Dementia with Lewy bodies DMV, see Dorsal motor nucleus of the vagus DO, see Detrusor overactivity Donepezil, 632 DOPA decarboxylase, see Aromatic amino acid decarboxylase Dopamine (DA) neuron classification and function, 63–65 synthesis, 38, 63 Dopamine β-hydroxylase (DBH) deficiency clinical features, 431–432 diagnosis, 432–433 differential dialysis, 433 genetics, 433 knockout mouse model, 433 management, 433, 619 Menkes disease, 435–437 expression regulation, 6 functional overview, 38, 431 metabolomics, 42 orthostatic hypotension mutations, 6–7 Dopamine receptor autonomic nervous system, 67 central nervous system, 67 classification, 67 functional selectivity, 69–70 gene structure, 67–68 ligand specificity, 69 oligomerization, 69 pharmacology, 69 signaling, 67–70 structure, 68–69 Dorsal commissural nucleus (DCN), sympathetic nervous system, 17 Dorsal motor nucleus of the vagus (DMV), central output, 12 Droxidopa (L-DOPS) autonomic failure indications, 619 dopamine β-hydroxylase deficiency management, 433 dopamine β-hydroxylase deficiency treatment, 619 history of study, 617 mechanism of action, 619 Menkes disease management, 437 norepinephrine synthesis, 617–618 pharmacokinetics, 617–618 structure, 617 Drug-induced autonomic dysfunction hypertension, 511–513 hypotension, 512, 514

E EAS, see External anal sphincter ECT, see Electroconvulsive therapy ED, see Erectile dysfunction Edrophonium, 631–632 EEG, see Electrencephalogram EGG, see Electrogastrogram Electrencephalogram (EEG), head-up tilt table testing monitoring, 384 Electroconvulsive therapy (ECT), neuroleptic malignant syndrome management, 544

Electrogastrogram (EGG), emotion studies, 296–297 Emotion, see Mind–body interactions Encephalitis, paraneoplastic autonomic dysfunction, 593 Endothelial function assessment nitric oxide activity, 320 pharmacologic testing, 320 physiologic testing, 320–321 autonomic interactions in cardiovascular pathophysiology, 322–323 biomarkers activation and dysfunction, 321 repair and regeneration, 321 consequences of dysfunction, 322–323 endothelial cell dysfunction, 319–320 normal function, 319 improvement strategies, 322 Endothelin (ET) aging effects, 273 animal models of pathophysiology, 136 endothelin-1 function cardiac effects, 136 essential hypertension role and therapeutic targeting, 136–137 renal effects, 136 expression regulation, 135 gene polymorphisms, 137 isoforms, 135 knockout mouse phenotypes, 135 nervous system distribution and pathology, 137–138 processing, 136 receptor polymorphisms and baroreflex function, 168 signaling, 135 salt-sensitivity of blood pressure role, 316 ENS, see Enteric nervous system Enteric nervous system (ENS), cotransmission, 29–30 Ephedrine adverse effects, 624 dosing, 624 mechanism of action, 623–624 Epilepsy central autonomic network infratentorial components, 549–550 supratentorial components, 549 ictal autonomic dysfunction cardiovascular, 550–551 cutaneous, 550 gastrointestinal, 550 pupil, 550 status epilepticus, 551 urogenital, 550 interictal autonomic dysfunction, 551–552 sudden unexpected death in epilepsy patients, 551 vagal nerve stimulation effects on cardiovascular function, 552 Epinephrine activation in stress, 42–43 panic disorder cotransmission in sympathetic nerves, 606 secretion, 605–607

697

Index

pupil response, 240 Erectile dysfunction (ED) associated conditions, 561 diabetic autonomic dysfunction, 479 epidemiology, 561 erection mechanism, 561–562 etiology, 562 hypogonadism, 562 neurogenic erectile dysfunction, 562 spinal cord injury, 508 treatment, 563 EROS-Clitoral Therapy Device, 238 Erythropoietin anemia in autonomic failure, 643 orthostatic hypotension management, 643–644 production modulation by autonomic nervous system, 643 ET, see Endothelin Evolution, see Comparative biology, autonomic nervous system in vertebrates Exercise benefits, 275, 277–278 blood pressure effects, 356–357 cardiovascular response acute exercise, 275–276 exercise training, 276–277 External anal sphincter (EAS), fecal continence, 601 External pressure, autonomic dysfunction management, 610

F

innervation, 236–237 sexual arousal afferent and sexual pathways, 235–236 neurotransmitters, 236 spinal cord injury, 508 FFT, see Fast Fourier transform FGIDs, see Functional gastrointestinal disorders Flow-mediated dilatation (FMD), endothelial function assessment, 320–321 Fludrocortisone history of study, 635 long-term effects, 636 mechanism of action, 635 neurally mediated syncope management, 342 pharmacology, 635–636 side effects, 636 structure, 636 Fluorescent probes, see Optogenetics FMD, see Flow-mediated dilatation fMRI, see Functional magnetic resonance imaging Functional gastrointestinal disorders (FGIDs) chronic abdominal pain, 598–599 classification, 597–598 cyclic vomiting syndrome, 597–598 overview, 597 treatment, 599 Functional magnetic resonance imaging (fMRI) brainstem sites in cardiovascular control, 13–16 neurovascular coupling, 197

Fabry's disease autonomic dysfunction, 496 clinical manifestations, 495–496 enzyme replacement therapy, 496 gene mutations, 495 Familial dysautonomia (FD) cardiovascular autonomic abnormalities, 500–501 clinical features, 499–500 genetics, 499–500 history of study, 499 pathology, 501 skin biopsy, 416–417 treatment, 502 Fast Fourier transform (FFT), power spectrum analysis, 405–407 FD, see Familial dysautonomia Fecal incontinence animal models, 602 physiology external anal sphincter, 601 internal anal sphincter, 601 puborectalis, 601 recto-anal reflexes, 601–602 prevalence, 601 sacral neuromodulation, 602 Female sexual function diabetic autonomic dysfunction and vaginal function, 479 dysfunction assessment, 237–238 classification, 235 etiologies, 237 management, 238 hormonal influences, 237

GABA, see γ-Aminobutyric acid Galantamine, 632 Gallstones, diabetic autonomic dysfunction, 478 Gastric emptying, see Stomach Gastric secretion, see Stomach GATA-3, autonomic nervous system development and specification role, 5 GBS, see Guillain–Barré syndrome GCHI, see GTP cyclohydrolase I GDNF, see Glial-derived neurotrophic factor Glial-derived neurotrophic factor (GDNF), autonomic neuron development role, 4 Glutamate autonomic function, 105–106 clearance, 105 excitotoxicity, 103 functional overview, 103 metabolism, 105 receptors, see specific receptors synthesis, 103 transporters, 103–104 GTP cyclohydrolase I (GCHI), gene therapy, 46–47 Guillain–Barré syndrome (GBS) clinical features, 493 course, 494 etiology, 493 investigations, 493 management, 494 prognosis, 494 sympathetic microneurography, 393

G

H Hair follicle, innervation, 413 Head-up tilt table testing (HUT) clinical applications, 384 indications, 384 monitoring, 383–385 neurally mediated syncope, 342, 384 orthostatic hypotension, 386–387 overview, 383 physiological basis, 383–384 postural tachycardia syndrome, 384, 386 types, 383 Heart, see also Cardiac vagal ganglia anatomy, 177–178 autonomic nervous control alterations, 179–180 overview, 178–179 beat control, 179 cocaine effects, 577–578 comparative biology of autonomic regulation, 669 conduction system, 178–179, 181 sympathetic imaging in Lewy body disease, 402 Heart failure central sleep apnea association, 569 individual variability, 367 muscle sympathetic nerve activity findings, 395 sympathetic activity, 367–370 Heart rate (HR) aging effects, 272 autonomic nervous system nitric oxide control, 101 baroreflex sensitivity control, 163–164 emotion studies, 297–298 power spectrum analysis, 405–407 variability in sleep apnea, 568–569 Heat stroke, management, 559 Hereditary sensory and autonomic neuropathy (HSAN) type III, see Familial dysautonomia types, 498 HfO, see High frequency oscillation HIF, see Hypoxia-inducible factor High altitude, see Altitude illness High frequency oscillation (HFO), blood pressure, 151–154 Hirschsprung disease, genotype–phenotype correlation, 446 Histamine, see Mastocytosis History taking autonomic disorder evaluation, 377 peripheral neuropathy with dysautonomia, 473 Hot flash, somatostatin analogs in management, 647 HR, see Heart rate HSAN, see Hereditary sensory and autonomic neuropathy HUT, see Head-up tilt table testing Hydrogen sulfide, carotid body hypoxic sensing, 332 Hydroxyamphetamine, pupil response, 240 11-β-Hydroxysteroid dehydrogenase type II, deficiency, 117 Hyperhidrosis causes, 553, 555

698

Index

Hyperhidrosis (continued) classification, 553–554 differential diagnosis, 555 treatment, 553–555, 649 Hypertension, see also Orthostatic hypertension; Preeclampsia aldosterone excess, 117–118 autonomic dysfunction complication by other disease, 346–347 evidence, 345 organ damage role, 345–346 therapeutic intervention, 347 baroreflex adaptation/resetting, 163 baroreflex stimulator therapy, 650 drug-induced, 511–513 drug-resistant hypertension, 650 endothelin-1 in essential hypertension and therapeutic targeting, 136–137 low-renin hypertension, 118–119 muscle sympathetic nerve activity, 395 obesity-associated hypertension and sympathetic activation, 360–361 renal dopamine role, 223 renal sympathetic nerve ablation for management, 650–652 sex differences, 263 spinal cord injury, 505 splanchic circulation, 212 sympathetic nervous system activation, 649–650 white coat hypertension, 356 Hyperthermia, thermoregulation, 288–289 Hypervolemia clinical manifestations, 123–124 etiology, 123 Hypoaldosteronism, 573 Hypohidrosis overview, 554–556 treatment of heat intolerance and heat stroke, 559 Hypotension, see also Neurally mediated hypotension; Orthostatic hypotension drug-induced, 512, 514 muscle sympathetic nerve activity testing, 393 postprandial hypotension, 356 Hypothalamus autonomic function, 9–10 thermoregulation, 245–245, 287 Hypothermia, thermoregulation, 288 Hypovolemia etiology, 123 postural orthostatic tachycardia syndrome, 523 Hypoxia, see also Altitude illness autonomic response acute hypoxia, 281–282 chronic hypoxia, 282 carotid body sensing, 331–332 comparative biology of response, 670 modeling sympathetic nerve activity following chronic intermittent hypoxia, 682–684 Hypoxia-inducible factor (HIF), carotid body hypoxic sensing, 332–333

I IAS, see Internal anal sphincter IBS, see Irritable bowel syndrome

IC, see Interstitial cystitis, Nucleus intercalatus spinalis ILF, see Intermediolateralis pars funicularis ILP, see Intermediolateralis pars principalis IML, see Intermediolateral cell column Immunoglobulin amyloidosis, see Amyloidosis Impedance threshold device (ITD), autonomic dysfunction management, 610 Incontinence, see Fecal incontinence; Lower urinary tract Inferior cervical ganglia, sympathetic nervous system, 17 Inflammation adrenomedullin attenuation, 128–129 autonomic nervous system effects on immune cells, 326–328 calcitonin gene-related peptide attenuation, 128–129 complex regional pain syndrome, 585 Innate immunity autonomic nervous system effects on cells, 326–328 overview, 325 Insular cortex, autonomic function, 9 Insulin, cardiovascular effects in nucleus of the solitary tract, 142 Insulin resistance (IR) complications, 307 sympathetic activity activation and resistance induction, 308–309 insulin resistance induction of activation, 307–308 overview, 307–308 therapeutic targeting, 309 Intermediolateral cell column (IML), sympathetic nervous system, 17 Intermediolateralis pars funicularis (ILF), sympathetic nervous system, 17 Intermediolateralis pars principalis (ILP), sympathetic nervous system, 17, 19, 21 Internal anal sphincter (IAS), fecal continence, 601 Interstitial cystitis (IC), stress effects, 25 Intestine autonomic control of secretion and absorption, 206 circulation, see Splanchic circulation diabetic autonomic dysfunction and colon function, 478 gut motility control, 206, 208 incontinence, see Fecal incontinence normal motor function, 208–209 spinal cord injury effects, 608 IR, see Insulin resistance Iris, see Pupil Irritable bowel syndrome (IBS), 599 ITD, see Impedance threshold device

J JH, see Joint hypermobility Joint hypermobility (JH) autonomic dysfunction, 535–537 clinical manifestations, 535 diagnosis, 535–536 epidemiology, 535

management, 537 pathophysiology, 535

K Kidney autonomic receptors, 215–217 blood volume and reflex regulation, 217–218 dopamine excretion, 221 hypertension role, 223 receptors expression, 221 sodium excretion regulation, 222 renin-angiotensin system interactions, 222–223–224 synthesis, 221 innervation, 215 pathophysiological states and autonomic control, 219–220 renal sympathetic nerve ablation for hypertension management, 650–652 renorenal reflex, 218–219

L LAAAD, see Aromatic amino acid decarboxylase Lambert–Eaton myasthenic syndrome (LEMS), 595 L-DOPS, see Droxidopa Leg crossing, autonomic dysfunction management, 609 LEMS, see Lambert–Eaton myasthenic syndrome Leptin functional overview, 131–132 interactions in hypothalamus melanocortin system, 133 neuropeptide Y, 133–134 receptor brain distribution, 132–133 isoforms, 131 signaling, 131–132 Lidocaine, see Anesthesia Liver circulation, see Splanchic circulation Lower urinary tract (LUT) central neural control, 227 clinical evaluation, 230 diabetic autonomic dysfunction and bladder dysfunction, 478–479 dysfunction bladder outlet obstruction, 232 incontinence, 230–232 neurology, 232 painful bladder syndrome, 232 treatment, 227–228 neuropathology, 227 neurotransmitter receptors, 227 normal sensory and motor properties, 229–230 parasympathetic pathways, 225–226 spinal cord injury effects, 508 structures, 226, 229 sympathetic pathways, 226–227 urethral sphincter somatic motor pathways, 227 Lumbodorsal splanchidectomy, 659 Lung, see Airway innervation LUT, see Lower urinary tract

Index

M Magnetic resonance imaging (MRI), see also Functional magnetic resonance imaging multiple system atrophy findings, 456 pure autonomic failure findings, 468 MAO, see Monoamine oxidase Mash1, autonomic nervous system development and specification role, 4–5 Mast cell activation disorder (MCAD), postural orthostatic tachycardia syndrome association, 522 Mastocytosis clinical features, 575–576 diagnosis, 576 mediators responsible for signs and symptoms, 575–576 MCAD, see Mast cell activation disorder MD, see Menkes disease Median preoptic nucleus (MnPO), thermoregulation, 243 Melanocortin receptor, types, 133 Menkes disease (MD) biochemical phenotype, 435–436 clinical features, 435 dysautonomia signs, 436–437 epidemiology, 435 molecular diagnosis, 437 neurochemical abnormalities, 437 Metabolic syndrome, muscle sympathetic nerve activity findings, 396 Metabolism regulation carbohydrate metabolism, 253 lipid metabolism, 253–254 protein metabolism, 254–255 resting metabolic rate and sympathetic stimulation contributions, 253 sypathoadrenal system dysregulation, 255 energy expenditure contribution, 253 Metabotropic glutamate receptor (mGluR), functional overview, 105 Methacholine, pupil response, 239–240 N-Methyl-D-aspartate (NMDA) receptor, functional overview, 104–105 mGluR, see Metabotropic glutamate receptor Microneurography, see Sympathetic microneurography Middle cervical ganglia, sympathetic nervous system, 17 Midrodine adverse effects, 622–623 dosing, 623 indications and efficacy, 621–622 mechanism of action, 621 pharmacology, 621 Migraine autonomic symptoms, 45–547 bio-behavioral model, 547 functional anatomy, 545 interictal autonomic dysfunction, 546 pain representation, 546–547 Mind–body interactions cardiovascular arousal studies of emotion, 297–298 electrogastrogram studies of emotion, 296–297 overview, 295

skin conductance response decision-making studies, 295 somatic marker hypothesis, 295 Mineralocorticoid receptor (MR), ligands, 117 MnPO, see Median preoptic nucleus Modeling, autonomic nervous system baroreceptor activation effects on respiratory pattern, 679, 681–682 brainstem sympatho-respiratory network, 679–680 sympathetic nerve activity following chronic intermittent hypoxia, 682–684 Virtual Physiological Human project/ Physiome project, 675–677 Monoamine oxidase (MAO) deficiency gene polymorphisms, 443–444 Norrie disease association, 443 functional overview, 41, 83 genes, 443 inhibitors and isoform specificity, 41 isoform function, 443 metabolomics, 42 Morvan's syndrome, 593–594 MR, see Mineralocorticoid receptor MRI, see Magnetic resonance imaging MSA, see Multiple system atrophy MSNA, see Muscle sympathetic nerve activity Multiple endocrine neoplasia type 2B, autonomic dysfunction, 497–498 Multiple myeloma, see Amyloidosis Multiple system atrophy (MSA) clinical features, 454 diagnosis, 454–456 differential diagnosis, 454–456 epidemiology, 453 history of study, 453 management, 456 muscle sympathetic nerve activity, 393 neuroprotective therapy, 456–457 pathophysiology, 303, 453–454 skin biopsy, 415 Muscarinic acetylcholine receptor, see Acetylcholine receptor Muscle sympathetic nerve activity (MSNA), see also Sympathetic microneurography applications bone loss, 396–397 chronic kidney disease, 395–396 heart failure, 367–369, 395 hypertension, 395 hypotensive attacks, 393 metabolic syndrome, 396 multiple system atrophy, 393 obstructive sleep apnea, 394–395 radiculoneuropathies, 393 insulin resistance studies, 307–310 power spectrum analysis, 406 sex differences, 261, 263

N NADPH oxidases, reactive oxygen species production, 336–337 ND, see Norrie disease NE, see Norepinephrine Nerve growth factor (NGF), autonomic neuron development role, 4

699 NET, see Norepinephrine transporter Neurally mediated hypotension, chronic fatigue syndrome, 533 Neurally mediated syncope (NMS) chronic fatigue syndrome, 533 diagnosis, 341–342 natural history, 342 pathophysiology, 341 syncope overview, 341 tilt table testing, 342, 384 treatment, 342–342 Neuroleptic malignant syndrome (NMS) clinical features, 541–542 differential diagnosis, 543 pathogenesis, 543 precipitants, 541–542 risk factors, 541, 543 treatment, 543–544 Neuromyotonia, paraneoplastic autonomic dysfunction, 593–594 Neuron, autonomic neural crest cell precursors, 3 neurotransmitters development acetylcholine, 7 noradrenaline, 6–7 phenotypes, 22–23 signaling in development, 3–4 transcription factors in autonomic nervous system development and specification AP2, 5–6 GATA-3, 5 Mash1, 4–5 Phox2 genes, 5 Neuropeptide Y (NPY) bone function, 258 cotransmission, 27–30, 32 leptin interactions in hypothalamus, 133–134 panic disorder and release, 607 vasomotor control, 187–190 Neurotrophin-3 (NT3), autonomic neuron development role, 4 NGF, see Nerve growth factor Nicotinic acetylcholine receptor, see Acetylcholine receptor Nitric oxide (NO) airway control, 203 autonomic nervous system nitric oxide heart rate control, 101 peripheral interactions, 100–101 cardiovascular effects in nucleus of the solitary tract, 141–142 central–autonomic nervous system interactions, 99–100 cotransmission, 27–31 endothelial function, 320–322 peroxynitrite formation, 335 salt-sensitivity of blood pressure role, 316–317 sympathetic function, 25 synthases, 99, 168 uncoupled nitric oxide synthase and reactive oxygen species production, 336 vasomotor control, 187, 190 NMDA receptor., see N-Methyl-D-aspartate receptor NMS, see Neurally mediated syncope; Neuroleptic malignant syndrome

700

Index

NO, see Nitric oxide Noradrenaline, see Norepinephrine Norepinephrine (NE) activation in stress, 42–43 autonomic function overview, 6 biomarkers of sympathetic innervation and function, 42 cardiovascular system neurons, 37–38 kidney function, 215–217 metabolism, 41–42 metabolomics, 42 panic disorder and reuptake, 606 release, 39 removal, 39–40 storage, 38–39 synthesis, 38, 617–618 transporter, see Norepinephrine transporter vasomotor control, 187–188 Norepinephrine transporter (NET) deficiency cardiovascular disease, 439 comorbidity of cardiovascular and neurobehavioral disorders, 442 gene polymorphisms, 439–442 orthostatic intolerance, 439–441 postural orthostatic tachycardia syndrome, 521–522 inhibition, see Atomoxetine regulation, 49–50 reuptake into nerve terminals, 39–40, 439 structure, 49 Norrie disease (ND), monoamine oxidase deficiency association, 443 NPY, see Neuropeptide Y NT3, see Neurotrophin-3 NTS, see Nucleus of the solitary tract Nucleus intercalatus spinalis (IC), sympathetic nervous system, 17 Nucleus of the solitary tract (NTS) anatomy, 141 autonomic function, 11, 13 cardiovascular effects adenosine, 142 angiotensin II, 142–143 insulin, 142 nitric oxide, 141–142 functional magnetic resonance imaging, 15–16

O Obesity, see also Hypertension; Insulin resistance epidemiology, 359 hypertension and sympathetic activation, 360–361 minority populations and sympathetic activation, 360 sympathetic activation, 359 Obstructive sleep apnea, see Sleep apnea Octreotide postprandial hypotension management, 640, 645–646 postural tachycardia syndrome management, 646–647 OI, see Orthostatic intolerance Optogenetics definition, 687 gene delivery, 688–689

reporters calcium, 687 effector proteins, 687–688 Organum vasculosum of the lateral terminalis (OVLT), osmoregulation, 121 Orthostatic hypertension associated conditions, 363–364 baroreflex failure, 350 overview, 363–364 Orthostatic hypotension aging, 271 definition, 529 delayed orthostatic hypotension, 529–530 dementia with Lewy bodies, 465 dopamine β-hydroxylase mutations, 6–7 erythropoietin management, 643–644 Guillain–Barré syndrome, 494 head-up tilt table testing, 386–387 midrodine efficacy, 621–622 Parkinson's disease, 460 pyridostigmine efficacy, 629 sex differences, 262–263 Orthostatic intolerance (OI), see also specific disorders chronic fatigue syndrome, 531–532 norepinephrine transporter defects, 439–441 space flight physiology, 284–285 Orthostatic stress arterial baroreceptor response, 193–194 capacitance vessels in orthostatic reflex adjustment, 194–195 humoral mechanisms, 195 local vasoconstrictor mechanisms, 194 skeletal muscle pump, 195 Orthostatic training, neurally mediated syncope management, 342–343 Osmoreceptors dysfunction, 124–125 Trpv4 role, 614 water drinking therapeutic utility, 614–615 water-induced pressor response evidence for sympathetic activation, 613–614 overview, 613 spinal sympathetic reflex, 614 Overactive bladder syndrome management, 227–228 overview, 231–232 OVLT, see Organum vasculosum of the lateral terminalis Oxidative stress adrenomedullin attenuation, 128 calcitonin gene-related peptide attenuation, 128 reactive oxygen species antioxidant defenses, 337 autonomic outflow, 337–338 biology, 335–336 sources mitochondrial respiration, 336 NADPH oxidases, 336–337 uncoupled nitric oxide synthase, 336 xanthine oxidase, 336 salt-sensitivity of blood pressure role, 316–317 Oxygen sensing carotid body gas messengers in hypoxic sensing, 332

hypoxic stimulus transduction, 331–332 molecular determinants of oxygen sensing, 332–333 morphology, 331 pathology, 333 unique aspects of oxygen sensing, 331 measures, 331 overview, 331

P PACAP, see Pituitary adenylate cyclaseactivating polypeptide PAD, see Paraneoplastic autonomic dysfunction PAF, see Pure autonomic failure PAG, see Periaqueductal gray Painful bladder syndrome, 232 Pancreas autonomic control of secretion, 205 circulation, see Splanchic circulation Panic disorder autonomic function, 292 cardiac risk, 607–608 neuropeptide Y release, 607 sympathetic nervous system function epinephrine cotransmission in sympathetic nerves, 606 secretion, 605–607 norepinephrine reuptake, 606 serotonin release, 605–606 Parabrachial complex (PBN) autonomic function, 11 thermoregulation, 243 Paraneoplastic autonomic dysfunction (PAD) classification, 593 clinical features, 593–594 diagnosis, 595–596 encephalitis, 593 enteric neuronopathy, 594 Lambert–Eaton myasthenic syndrome, 595 Morvan's syndrome, 593–594 neuromyotonia, 593–594 neuropathy, 491 treatment, 596 Parasympathetic nervous system (PNS) cotransmission, 28–29 functions, 24–25 neurotransmitter phenotypes, 22–23 overview, 18, 21–22 Parkinson's disease (PD) animal studies of autonomic function, 25 anticholinergic agent therapy, 461 denervation imaging, 402 diagnosis, 459 dopamine replacement therapy, 459–460 gene therapy targets, 46–47 L-DOPA therapy, 46 orthostatic hypotension, 460 skin biopsy, 415 α-synuclein in pathology, 303 PBN, see Parabrachial complex PD, see Parkinson's disease Periaqueductal gray (PAG), autonomic function, 10–11 Peripheral neuropathy, diagnostic workup with dysautonomia, 473–474 PET, see Positron emission tomography Phenylephrine, pupil response, 240

701

Index

Phenylethanolamine-N-methyltransferase (PNMT), functional overview, 38 Pheochromocytoma diagnosis, 421–424 differential diagnosis, 422 gene mutations and phenotypes, 421–422 malignant, 424 treatment, 424 Phox2 genes, autonomic nervous system development and specification role, 5 PHOX2B function, 445 genotype–phenotype correlations cardiac asystole, 446 continuous ventilatory dependence, 446 facial dysmorphology, 447 frameshift mutations, 447 Hirschsprung disease, 446 later-onset disease, 447–448 tumors of neural crest origin, 446 locus, 445 Physical activity, see Exercise Physical examination autonomic dysfunction, 379 peripheral neuropathy with dysautonomia, 473 Physiome project autonomic nervous system modeling, 675–677 overview, 673 physiome standards, 673–675 prospects, 677–678 Physostigmine, pupil response, 240 Pilocarpine, pupil response, 239–240 Pituitary adenylate cyclase-activating polypeptide (PACAP), migraine role, 545 PNMT, see Phenylethanolamine-Nmethyltransferase PNS, see Parasympathetic nervous system POA, see Preoptic area Porphyria autonomic dysfunction, 497 clinical manifestations, 497 gene mutations, 496–497 treatment, 497 Positron emission tomography (PET) dysautonomia sympathetic imaging, 401–402 Lewy body disease findings, 402 multiple system atrophy findings, 456 pure autonomic failure findings, 468 tracers for sympathetic imaging, 399–401 Post-traumatic stress disorder (PTSD), autonomic function, 292 Postprandial hypotension (PPH), 356, 639–641 Postural orthostatic tachycardia syndrome (POTS) chronic fatigue syndrome, 533 clinical features, 517 epidemiology, 517 exercise therapy, 278 follow-up, 518 head-up tilt table testing, 384, 386 joint hypermobility association, 536–537 management, 518 octreotide management, 646–647 pathophysiology hyperadrenergic POTS

central hyperadrenergic POTS, 521 mast cell activation disorder, 522 norepinephrine transporter deficiency, 521–522 hypovolemia and blood volume regulation, 523 neuropathic POTS, 522–523 stroke volume reduction, 523 phenotypes deconditioning-associated POTS, 518 hyperadrenergic POTS, 517 neuropathic POTS, 517 sex differences, 263, 517 symptoms, 378 tachycardia origins, 525–526 reflex tachycardia classification high flow POTS, 527 low flow POTS, 526–527 normal flow POTS, 528 water drinking therapy, 615 Posture, see Orthostatic hypertension; Orthostatic hypotension; Orthostatic stress POTS, see Postural orthostatic tachycardia syndrome PP2A, see Protein phosphatase 2A PPH, see Postprandial hypotension Preeclampsia, 267 Pregnancy baroreceptor reflex impairment, 265–267 preeclampsia, 267 sympathetic nervous system activation, 265 Preoptic area (POA), thermoregulation, 244– 246, 287 Protein phosphatase 2A (PP2A), norepinephrine transporter interactions, 49–50 Psychological stress, see Stress PTSD, see Post-traumatic stress disorder Puborectalis, fecal continence, 601 Pupil combined sympathetic and parasympathetic defects, 242 diabetic autonomic dysfunction findings, 476 epilepsy changes, 550 instilled drug response adrenergic blockers, 240 adrenergic drugs, 240 anticholinergic drugs, 239 cholinergic drugs, 239 iris pigment effects, 241 miscellaneous drugs, 240–241 parasympathetic defects, 242 sympathetic defects, 241–242 Pure autonomic failure (PAF) catecholamine studies, 467–468 clinical features, 467 differential diagnosis, 467 imaging studies, 468 management, 468 neuroendocrine studies, 468 neuropathology, 468 Purinergic receptors, see also Adenosine receptor classification, 89–91 P2X receptors, 90 P2Y receptors, 90, 92

Pyridostigmine efficacy, 629 mechanism of action, 629

Q QDIRT, see Quantitative direct and indirect test of pseudomotor function QSART, see Quantitative sudomotor axon reflex test QST, see Quantitative sensory testing Quantitative direct and indirect test of pseudomotor function (QDIRT), 410 Quantitative sensory testing (QST), female sexual dysfunction assessment, 237 Quantitative sudomotor axon reflex test (QSART), 410–411, 474, 519

R RAS, see Renin-angiotensin system Rasagiline, multiple system atrophy management, 456–457 Reactive amyloidosis, see Amyloidosis Reactive oxygen species, see Oxidative stress Recto-anal reflexes, fecal continence, 601–602 Renal norepinephrine spillover (RNES), heart failure studies, 367 Renin-angiotensin system (RAS) angiotensin II cardiovascular effects in nucleus of the solitary tract, 142–143 angiotensin receptors, 113–114, 168 angiotensin-converting enzyme, 113–116 brain system autonomic regulation, 115–116 components, 114–115 classical system autonomic regulation, 113–114 components and features, 113 low-renin hypertension, 118–119 orthostatic stress response, 195 renal dopaminergic system interactions, 222–223–224 salt-sensitivity of blood pressure, 314, 316 Renorenal reflex, see Kidney Respiratory-cardiovascular coupling cardiovascular modulation of respiratory activity, 154 high frequency oscillation cardiovascular disease response, 154 functions, 153 overview, 151 prospects for study, 154 respiratory pattern generator coupling to cardiovascular autonomic activity, 151–152 respiratory sinus arrhythmia cardiovascular disease response, 153–154 functions, 152–153 Respiratory sinus arrhythmia (RSA), respiratory-cardiovascular coupling, 151–154 Resting metabolic rate (RMR), sympathetic stimulation contributions, 253 Rifampicin, multiple system atrophy management, 457 Riley–Day syndrome, see Familial dysautonomia Rivastigmine, 632 RMR, see Resting metabolic rate

702

Index

RNES, see Renal norepinephrine spillover Rostral raphe pallidus area (rRPa), thermoregulation, 246 Rostral ventrolateral medulla (RVLM) adenosine receptors, 96 autonomic function, 11, 13 baroreceptor reflex impairment in pregnancy, 266–267 functional magnetic resonance imaging, 15 rRPa, see Rostral raphe pallidus area RSA, see Respiratory sinus arrhythmia RVLM, see Rostral ventrolateral medulla

S SA node, see Sinoatrial node SAH, see Subarachnoid hemorrhage Salivary gland, autonomic control of secretion, 205 Salt-sensitivity of blood pressure (SSBP) clinical significance, 317–318 defects arachidonic acid metabolites, 317 endothelin, 316 nitric oxide, 316 renin-angiotensin-aldosterone system, 314, 316 sympathetic nervous system neurotransmitters, 317 gene–environment interactions, 314 genetics, 313–316 SCG, see Superior cervical ganglia SCM, see Stress cardiomyopathy SCN, see Suprachiasmatic nuclei Scopolamine, pupil response, 239 SCR, see Skin conductance response Serotonin autonomic function, 85–86 false cotransmitter function, 28, 32 localization, 83 metabolism, 590 panic disorder and brain release, 605–606 sympathetic rhythms, 148 synthesis and metabolism, 83 transporters, 83–84 Serotonin receptor pharmacology, 85 signaling, 84–85 Sex differences, autonomic function hypertension, 263 normal autonomic function, 261 orthostatic hypotension, 262–263 overview, 261 Sexual function, see Female sexual function SIADH, see Syndrome of inappropriate antidiuretic hormone Sildenafil, erectile dysfunction management, 563 Single-photon emission computed tomography (SPECT) dysautonomia sympathetic imaging, 401–402 Lewy body disease findings, 402 multiple system atrophy findings, 456 tracers for sympathetic imaging, 399–401 Sinoatrial (SA) node, 178–179, 526 Skeleton, see Bone Skin biopsy applications, 414–416, 474 neuroanatomy arrector pilorum muscles, 414

blood vessels, 414 hair follicle, 413 sweat gland, 413 Skin conductance response (SCR) decision-making studies, 295 somatic marker hypothesis, 295 Skin sympathetic nerve activity (SSNA), see also Sympathetic microneurography applications radiculoneuropathies, 393 sweat disturbances, 394 cocaine effects, 580 Sleep apnea autonomic function during sleep, 565 central sleep apnea, 569 muscle sympathetic nerve activity, 394–395 obstructive sleep apnea acute changes during episodes, 565, 567 atrial fibrillation association, 567–568 chronic changes during episodes, 567 epidemiology, 565 heart rate variability, 568 sudden cardiac death association, 568 Small intestine, see Intestine SMP, see Sympathetically maintained pain SNS, see Sympathetic nervous system Somatostatin agonists, see also Octreotide adverse effects, 647–648 hot flash management, 647 postural tachycardia syndrome management, 646–647 autonomic neuropathy role, 645–646 receptors, 645 SP, see Substance P Space physiology management of disorders, 285 motion sickness, 284–285 orthostatic intolerance following flight, 284–285 overview, 284 SPECT, see Single-photon emission computed tomography Spinal cord injury bionic baroreflex system application, 662 cardiovascular system effects, 505–507 cutaneous circulation, 507 gastrointestinal effects, 608 reproductive system effects, 508 spinal shock, 505 sudomotor function, 508 thermoregulation, 507–508 urinary system effects, 508 Splanchic circulation lidocaine and hepatic blood flow, 665 local regulation, 211 overall circulatory function impact, 211 sympathetic control disease, 212–213 overview, 211, 212 special circumstances, 212 Spleen circulation, see Splanchic circulation Squatting, autonomic dysfunction management, 609–610 SSBP, see Salt-sensitivity of blood pressure SSNA, see Skin sympathetic nerve activity SSR, see Sympathetic skin response Stomach autonomic control of secretion, 205

circulation, see Splanchic circulation diabetic autonomic dysfunction and gastric emptying, 476–477 gut motility control, 206, 208 normal motor function, 208–209 Stress anxiety disorder, 292 autonomic dysfunction, 25 cardiac disease, 292 gastrointestinal effects, 292 noradrenergic versus adrenergic activation in stress, 42–43 panic disorder, 292 post-traumatic stress disorder, 292 psychosomatic disorders, 292 sympathetic nerve responses, 291–293 Stress cardiomyopathy (SCM) causes, 371–373 coronary artery spasm, 371–373 diagnosis, 371, 373 sympathetic activation, 373–374 Stress urinary incontinence (SUI), 232 Stroke volume, postural orthostatic tachycardia syndrome, 523 Subarachnoid hemorrhage (SAH), endothelin role, 138 Substance P (SP) airway control, 203 cotransmission, 29 Sudden cardiac death, obstructive sleep apnea association, 568 Sudden unexpected death in epilepsy patients (SUDEP), 551 SUDEP, see Sudden unexpected death in epilepsy patients Sudomotor function axon reflex testing, 409–411 diabetic autonomic dysfunction, 479 overview of tests, 409 skin potentials, 411 spinal cord injury, 508 sweating disorders and testing, 556, 558 thermoregulatory sweat test, 411 SUI, see Stress urinary incontinence Superior cervical ganglia (SCG), sympathetic nervous system, 17–20, 22–23 Suprachiasmatic nuclei (SCN) circadian and sleep control of autonomic nervous system, 158–159 output and autonomic control, 157–158 pacemaker cells, 157 Sweat glands denervation effects, 250–251 density and distribution, 249 factors affecting sweat response, 250 functions, 250, 289 innervation, 250 innervation, 413 normal factors affecting sweating, 553–554 physiology, 249–250 skin sympathetic nerve activity in disturbances, 394 thermoregulatory sweat test, 411 types, 249 Sweating disorders, see Anhidrosis; Hyperhidrosis; Hypohidrosis Sympathectomy, hyperhidrosis management, 649

703

Index

Sympathetic microneurography, see also Muscle sympathetic nerve activity; Skin sympathetic nerve activity analysis multi-unit activity, 391 single unit activity, 391 applications bone loss, 396–397 chronic kidney disease, 395–396 heart failure, 395 hypertension, 395 hypotensive attacks, 393 metabolic syndrome, 396 multiple system atrophy, 393 obstructive sleep apnea, 394–395 radiculoneuropathies, 393 sweat disturbances, 394 challenges, 392 electrode site, 392 equipment, 389 technique, 389–391 Sympathetic nervous system (SNS) cotransmission, 27–28 functions, 24–25 neurotransmitter phenotypes, 22–23 overview, 17–20 sympathoadrenal axis, 20–21 Sympathetic rhythm cardiac and respiratory rhythms entrainment, 147–148 mechanisms, 147 phasic inputs, 147 central oscillators, 148 definition, 147 functional significance, 148 spinal cord and sympathetic rhythms, 148 Sympathetic skin response (SSR) spinal cord injury, 508 sudomotor function testing, 411 Sympathetically maintained pain (SMP), complex regional pain syndrome, 586 Syncope, see Neurally mediated syncope Syndrome of inappropriate antidiuretic hormone (SIADH) autonomic dysfunction, 125 euvolemia, 123 Syntaxin 1A, norepinephrine transporter interactions, 49–50 α-Synuclein pathology Alzheimer's disease, 303 multiple system atrophy, 303 Parkinson's disease, 303 transgenic animal studies, 304 synucleinopathies, see Dementia with Lewy bodies; Multiple system atrophy; Parkinson's disease; Pure autonomic failure therapeutic targeting, 304 toxic species, 303–304

T Takotsubo syndrome, 371 TCD, see Transcranial Doppler TEF, see Thermogenic effect of feeding Tetrahydrobiopterin (BH4) deficiency clinical presentation, 427–428 diagnosis, 428–429 treatment, 429 functions, 427 TH, see Tyrosine hydroxylase Thermogenic effect of feeding (TEF), 254 Thermoregulation central control overview, 287 cocaine effects, 579 cutaneous thermal receptor afferent pathway, 243–244 dorsomedial hypothalamus, 245–246 hyperthermia response, 288–289 hypothermia response, 288 rostral raphe pallidus area, 246 sensorimotor integration in preoptic area, 244–245 spinal cord injury, 507–508 spinal sympathetic mechanisms, 246 sweat glands, 250, 289 thermoneutral environments, 287–288 Thermoregulatory sweat test, sudomotor function, 411 Tilt table testing, see Head-up tilt table testing Traditional Chinese medicine, see Acupuncture Transcranial Doppler (TCD), head-up tilt table testing monitoring, 384 Transient receptor potential channel (Trp), Trpv4 in osmoreception, 614 Transthyretin amyloidosis, see Amyloidosis Tropicamide, pupil response, 239 Trp, see Transient receptor potential channel Tyramine, pupil response, 240 Tyrosine hydroxylase (TH) activation, 38 deficiency clinical presentation, 428 diagnosis, 429 treatment, 429 functional overview, 38, 45, 427 gene mutations in deficiency, 46–47 splice variants, 45–46 structure, 45 gene therapy, 46–47

U Urethra, see Lower urinary tract Urination, see Lower urinary tract

V Vagal nerve stimulation (VNS), effects on cardiovascular function in epilepsy, 552

Valsalva maneuver, autonomic dysfunction evaluation, 380–381 Vasculature, see Blood vessel Vasoactive intestinal peptide (VIP) cotransmission, 27–31 vasomotor control, 187, 190 Vasomotor control, see also Endothelial function; Endothelin adrenergic vasoconstriction modulation, 191 aging effects, 273 differential control, 190 overview, 187–188 parasympathetic control, 189–190 sympathetic control neuroeffector junction, 188 neurotransmitters, 188–189 Vasopressin, see Arginine vasopressin Vesicular monoamine transporter (VMAT) functional overview, 38–39 serotonin transport, 83 VIP, see Vasoactive intestinal peptide Virtual Physiological Human (VPH) project autonomic nervous system modeling, 675–677 overview, 673 physiome standards, 673–675 prospects, 677–678 Visceral afferents anatomy, 171 autonomic reflex responses to activation, 173–174 fiber types, 171–172 functional overview, 23–24, 171 ischemia response, 171–173 pathological alterations, 174–175 stimuli, 171 VMAT, see Vesicular monoamine transporter VNS, see Vagal nerve stimulation VPH project, see Virtual Physiological Human project

W Water drinking, therapeutic utility, 614–615 WCH, see White coat hypertension White coat hypertension (WCH), 356 Wnt, autonomic neuron development role, 4

X Xanthine oxidase, reactive oxygen species production, 336

Y Yohimbine adverse effects, 624 dosing, 624 efficacy, 628 mechanism of action, 624, 628 sources, 627