Table Of ContentMethods in Neurosciences
Edited by
P. Michael Conn
Department of Pharmacology
The University of Iowa
College of Medicine
Iowa City, Iowa
V o l u me 11
Receptors: Model Systems and
Specific Receptors
ACADEMIC PRESS, INC.
Harcourt Brace Jovanovich, Publishers
San Diego New York Boston London Sydney Tokyo Toronto
Front cover photograph (paperback edition only): Autoradiographic distribution of
the thymic ß2-adrenergic receptor. A Loats Image Analysis System (Amersham) was
used to analyze differences in receptor localization. Cold colors (including blue, violet,
and rose) indicate the absence or low concentration of receptor; green, orange, yellow,
and red represent increasing receptor concentrations. In the section shown from a
proestrous female rat thymus gland, a high concentration of receptor can be seen dis
tributed over the medullary regions. Courtesy of Drs. Bianca Marchetti and Maria C.
Morale, Department of Pharmacology, Medical School, University of Catania, Italy,
This book is printed on acid-free paper. @
Copyright © 1993 by ACADEMIC PRESS, INC.
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Contributors to Volume 11
Article numbers are in parentheses following the names of contributors. Affiliations listed are current.
ALAIN BEAUDET (20), Laboratory of Neuroanatomy, Montreal Neurobio-
logical Institute, McGill University, Montreal, Quebec H3A 2B4, Canada
ANN M. BENZ (15), Department of Psychiatry, Washington University
School of Medicine, St. Louis, Missouri 63110
ERIC L. BITTMAN (6), Department of Zoology and Program in Neuroscience
and Behavior, University of Massachusetts, Amherst, Massachusetts 01003
STEPHEN R. BLOOM (8), Department of Medicine, Royal Postgraduate Medi
cal School, Hammersmith Hospital, London W12 ONN, England
S. MARC BREEDLOVE (1), Department of Psychology and Graduate Group in
Neurobiology, University of California, Berkeley, Berkeley, California
94720
A. CADIEUX (4), Department of Pharmacology, University of Sherbrooke
Medical School, Sherbrooke, Quebec JIH 5N4, Canada
ANNICK CAUVIN (24), Laboratoire de Chimie Biologique et de la Nutrition,
Université Libre de Bruxelles, Â-1070 Brussels, Belgium
JoÉLLE CHABRY (20), Institut de Pharmacologic Moléculaire et Cellulaire,
Centre National de la Recherche Scientifique, Sophia Antipolis, 06560
Valbonne, France
FREDERIC CHECLER (20), Institut de Pharmacologic Moléculaire et CeUu-
laire, Centre National de la Recherche Scientifique, Sophia Antipolis, 06560
Valbonne, France
YI-ZHANG CHEN (2), Laboratory of Neuroscience, Department of
Physiology, The Second Military Medical University, Shanghai 200433, Peo
ple's Republic of China
JEAN CHRISTOPHE (24), Laboratoire de Chimie Biologique et de la Nutrition,
Université Libre de Bruxelles, B-1070 Brussels, Belgium
DANIELA CIRILLO (23), Department of Biomedical Science and Oncology,
University of Torino Medical School, Torino, Italy 10126
CS. COCKRAM (12), Department of Medicine, The Chinese University of
Hong Kong, Prince of Wales Hospital, Shatin N.T., Hong Kong
viii CONTRIBUTORS TO VOLUME 11
P. DORLÉANS-JUSTE (4), Department of Pharmacology, University of
Sherbrooke Medical School, Sherbrooke, Quebec JIH 5N4, Canada
Errol B. De Souza (3), CNS Diseases Research, The Du Pont Merck
Pharmaceutical Company, Experimental Station, Wilmington, Delaware
19880
M. B. EMERIT (22), INSERM U. 288, Neurobiologie Cellulaire et Fonction-
nelle, Faculté de Médecine Pitié-Salpétriére, 75634 Paris, France
LOUISE M. FREEMAN (1), Department of Psychology, University of Califor
nia, Berkeley, Berkeley, California 94720
HONG FU (2), Laboratory of Neuroscience, Department of Physiology, The
Second Military Medical University, Shanghai 200433, People's Republic of
China
PAOLA GALLO (7), Department of Neurology, Second Neurologic Clinic,
University of Padova School of Medicine, 35137 Padova, Italy
STEEN GAMMELTOFT (13), Department of Clinical Chemistry, Bispebjerg
Hospital, DK-2400 Copenhagen NV, Denmark
J. A. GINGRICH (22), INSERM U. 288, Neurobiologie Cellulaire et Fonction-
nelle. Faculté de Médecine Pidé-Salpétriére, 75634 Paris, France
BRUNO GIOMETTO (7), Department of Neurology, Second Neurologie
Clinic, University of Padova School of Medicine, 35137 Padova, Italy
H. GOZLAN (22), INSERM U. 288, Neurobiologie Cellulaire et Foncdon-
nelle, Faculté de Médecine Pidé-Salpétriére, 75634 Paris, France
Zuo Guo (2), Laboratory of Neuroscience, Department of Physiology, The
Second Military Medical University, Shanghai 200433, People's Republic of
China
M. HAMON (22), INSERM U. 288, Neurobiologie CeUulaire et Foncdon-
nelle. Faculté de Médecine Pidé-Salpétriére, 75634 Paris, France
JANICE Ě. HICKOK (15), Department of Psychiatry, Washington University
School of Medicine, St. Louis, Missouri 63110
ROBERT T. JENSEN (17), Digesdve Disorders Branch, National Institute of
Diabetes and Digestive and Kidney Diseases, National Institutes of Health,
Bethesda, Maryland 20892
RICHARD KRIS (23), Department of Pharmacology, New York University
Medical Center, New York, New York 10016
CONTRIBUTORS TO VOLUME 11 ix
ELLEN E. LADENHEIM (17), Department of Psychiatry and Behavioral Sci
ences, The Johns Hopkins University School of Medicine, Baltimore, Mary
land 21205
MANFRED P. LUTZ (10), Center for Basic Research in Digestive Diseases,
Mayo Clinic, Rochester, Minnesota 55905
PAUL G. LYSKO (11), Department of Cardiovascular Pharmacology,
SmithKhne Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406
JEAN MAZELLA (20), Institut de Pharmacologic Moléculaire et Cellulaire,
Centre National de la Recherche Scientiñque, Sophia Antipolis, 06560
Valbonne, France
LAURENCE J. MILLER (10), Center for Basic Research in Digestive Diseases,
Mayo Medical School, Clinic and Foundation, Rochester, Minnesota 55905
M.-C. MIQUEL (22), INSERM U. 288, Neurobiologie Cellulaire et Fonction-
nelle. Faculté de Médecine Pitié-Salpétriére, 75634 Paris, France
TERRY W. MOODY (23), Department of Biochemistry and Molecular Biol
ogy, The George Washington University Medical Center, Washington, D.C.
20037
TIMOTHY H. MORAN (17), Department of Psychiatry and Behavioral Sci
ences, The Johns Hopkins University School of Medicine, BaUimore, Mary
land 21205
LuiGi NALDINI (23), Department of Biomedical Science and Oncology, Uni
versity of Torino Medical School, Torino, Italy 10126
PoNNAL NAMBÍ (11), Department of Renal Pharmacology, SmithKhne
Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406
M. G. NICHOLLS (12), Department of Medicine, Christchurch Hospital,
Christchurch 1, New Zealand
D. REGOLI (4), Department of Pharmacology, University of Sherbrooke
Medical School, Sherbrooke, Quebec JIH 5N4, Canada
PATRICK ROBBERECHT (24), Laboratoire de Chimie Biologique et de la Nutri
tion, Université Libre de Bruxelles, B-1070 Brussels, Belgium
STEVEN A. ROSENZW^EIG (18), Department of CeU and Molecular Pharma
cology and Experimental Therapeutics, Medical University of South Caro
lina, Charleston, South Carolina 29425
ENRIQUE ROZENGURT (16), Imperial Cancer Research Fund, Laboratory of
Growth Regulation, London WC2A 3PX, England
CONTRIBUTORS TO VOLUME 11
JAMES SINNETT-SMITH (16), Imperial Cancer Research Fund, Laboratory of
Growth Reguladon, London WC2A 3PX, England
DAVID M. SMITH (8), Department of Medicine, Royal Postgraduate Medical
School, Hammersmith Hospital, London W12 ONN, England
BARBARA C. SW^ARZENSKI (15), Department of Psychiatry, Washington Uni
versity School of Medicine, St. Louis, Missouri 63110
TosHiHiRO TAKAO (3), CNS Diseases Research, The Du Pont Merck
Pharmaceudcal Company, Experimental Station, Wilmington, Delaware
19880
JEFFREY B. TATRO (5), Division of Endocrinology, Diabetes, Metabolism,
and Molecular Medicine, New England Medical Center Hospitals; and De
partment of Medicine, Tufts University School of Medicine, Boston, Massa
chusetts 02111
BRUNO TAVOLATO (7), Department of Neurology, Second Neurologic
Clinic, University of Padova School of Medicine, 35137 Padova, Italy
IAN L. TAYLOR (19), Departments of Medicine and CeU Biology, Division of
Gastroenterology, Duke University Medical Center, Durham, North Caro-
hna 27710; and Durham Veterans Administration Medical Center, Durham,
North Carolina 27705
RICHARD D. TODD (15), Departments of Psychiatry and Genetics, Washing
ton University School of Medicine, St. Louis, Missouri 63110
ANDRE VANDERMEERS (24), Laboratoire de Chimie Biologique et de la Nu
trition, Université Libre de Bruxelles, B-1070 Brussels, Belgium
MARIE-CLAIRE VANDERMEERS-PIRET (24), Laboratoire de Chimie Biologi
que et de la Nutrition, Université Libre de Bruxelles, B-1070 Brussels,
Belgium
STEVEN R. VIGNA (19), Departments of Cell Biology and Medicine, Division
of Gastroenterology, Duke University Medical Center, Durham, North Car
olina 27710; and Durham Veterans Administradon Medical Center, Durham,
North Carolina 27705
JEAN-PIERRE VINCENT (20), Institut de Pharmacologic Moléculaire et Cellu
laire, Centre National de la Recherche Scientifique, Sophia Antipolis, 06560
Valbonne, France
ANDREW^ A. WELCHER (21), Amgen Inc., Amgen Center, Thousand Oaks,
California 91320
CONTRIBUTORS TO VOLUME 11 XI
PATRICIA M. WHITAKER-AZMITIA (14), Department of Psychiatry, Health
Sciences Center, State University of New York at Stony Brook, Stony
Brook, New York 11794
DAVID C. WHITCOMB (19), Departments of Medicine and Physiology, Divi
sion of Gastroenterology and Hepatology, University of Pittsburgh Medical
Center, Pittsburgh, Pennsylvania 15213
ZSUZSANNA WIESENFELD-HALLIN (9), Section of Chnical Neurophysiology,
Department of Clinical Physiology, Karolinska Institute, Huddinge Univer
sity Hospital, S-141 86 Huddinge, Sweden
XIAO-JUN XU (9), Section of Chnical Neurophysiology, Department of Clin
ical Physiology, Karolinska Institute, Huddinge University Hospital, S-141
86 Huddinge, Sweden
V. T. F. YEUNG (12), Department of Medicine, The Chinese University of
Hong Kong, Prince of Wales Hospital, Shatin N.T., Hong Kong
IAN ZACHARY (16), Imperial Cancer Research Fund, Laboratory of Growth
Regulation, London WC2A 3PX, England
Preface
Receptors initiate the means by which ceUular regulators exert their actions
on target cells. Because of the central role of cell-cell communication and
signal transduction, receptors are of intrinsic interest to neuroscientists.
Receptor studies utilize both traditional methods of analysis and modern
molecular techniques. Volumes 11 and 12 of this series are divided into
sections describing, in a pragmatic way, ''model" receptor techniques,
molecular techniques, and techniques for the determination of receptor sub
classes and for localization and consideration in ligand design. The chapters
are written in a way that will ahow readers to "export" the technology
described to the study of other receptor systems in their own areas of inter
est. Techniques include PCR protocols, methods for the assessment of gene
expression, transfection, cloning, autoradiography, in situ hybridization, ra
dioligand binding, receptor solubilization and purification, and coupling to
effector systems.
The goal of these volumes—and of the others in this series—is to provide
in one source a view of the contemporary techniques significant to a particu
lar branch of the neurosciences, information which will prove invaluable not
only to the experienced researcher but to the student as well. Of necessity
some archival material has been included, but the authors have been encour
aged to present information that has not yet been published, to compare (in a
way not found in other publications) different approaches to similar prob
lems, and to provide tables that direct the reader, in a systematic fashion, to
earlier literature as an efficient means to summarize data. Flow diagrams and
summary charts will guide the reader through the processes described.
The nature of this series permits the presentation of methods in fine detail,
reveahng "tricks" and short cuts that frequendy do not appear in the litera
ture owing to space limitations. Lengthy operating instructions for common
equipment will not be included except in cases of unusual application. The
contributors have been given wide latitude in nomenclature and usage since
they are best able to make judgments consistent with current changes.
I wish to express my appreciation to Mrs. Sue Birely for assisdng in the
organization and maintenance of records, and to the staff of Academic Press
for their efficient coordination of production. Appreciation is also expressed
to the contributors, particularly for meeting their deadlines for the prompt
and dmely publicadon of this volume.
P. MICHAEL CONN
xni
Methods in Neurosciences
Volume 1 Gene Probes
Edited by P. Michael Conn
Volume 2 Cell Culture
Edited by P. Michael Conn
Volume 3 Quantitative and Qualitative Microscopy
Edited by P. Michael Conn
Volume 4 Electrophysiology and Microinjection
Edited by P. Michael Conn
Volume 5 Neuropeptide Technology: Gene Expression and Neuropeptide Receptors
Edited by P. Michael Conn
Volume 6 Neuropeptide Technology: Synthesis, Assay, Purification,
and Processing
Edited by P. Michael Conn
Volume 7 Lesions and Transplantation
Edited by P. Michael Conn
Volume 8 Neurotoxins
Edited by P. Michael Conn
Volume 9 Gene Expression in Neural Tissues
Edited by P. Michael Conn
Volume 10 Computers and Computations in the Neurosciences
Edited by P. Michael Conn
Volume 11 Receptors: Model Systems and Specific Receptors
Edited by P. Michael Conn
Volume 12 Receptors: Molecular Biology, Receptor Subclasses, Localization, and Ligand Design
(in preparation)
Edited by P. Michael Conn
Volume 13 Neuropeptide Analogs, Conjugates, and Fragments (in preparation)
Edited by P. Michael Conn
Volume 14 Paradigms for the Study of Behavior (in preparation)
Edited by P. Michael Conn
Volume 15 Photoreceptor Cells (in preparation)
Edited by Paul A. Hargrave
Volume 16 Neurobiology of Cytokines (Part A) (in preparation)
Edited by Errol B. De Souza
Volume 17 Neurobiology of Cytokines (Part B) (in preparation)
Edited by Errol B. De Souza
XV
[1] Steroid Receptors in the Central
Nervous System
Louise Μ. Freeman and S. Marc Breedlove
Introduction
Steroid hormones exert powerful influences over the structure and function
of both the developing and the mature nervous system. In fact, the proposi
tion that steroid hormones could alter the developing and mature nervous
system was first indicated on the basis of behavior. Castrated males of
most species will eventually stop copulating and testosterone treatment can
reinstate the behavior as long as the regimen is maintained. Similarly, sexual
receptivity in females is abolished by ovariectomy and, in rats, for example,
sequential treatment with estrogen and progesterone will result, a few hours
later, in 6-8 hr of receptivity. Phoenix et al. (I) proposed that, in addition to
such "activational" effects, steroids could also 'Organize" the developing
nervous system, permanently rendering it masculine or feminine. The even
tual discovery of sexual dimorphism in the vertebrate central nervous system
(CNS) and the discovery that early exposure to testicular steroids was respon
sible for the development of masculine morphology amply confirmed this
organizational hypothesis (2). Although the organizational-activational dis
tinction has been useful to behavioral endocrinologists, the basic cellular
action of steroids, change in gene expression, is similar throughout develop
ment (3). This change can often manifest itself as an alteration in the behavior
of the organism. As such, steroids provide an opportunity to perturb normal
neural processes and monitor the resulting changes in behavior, thereby
illuminating the structural and/or physiological bases of neural development
and function. We will review the methods of monitoring steroid influence,
which center, as we will see, on monitoring the receptors for these hormones.
Steroids are lipophiHc molecules consisting of four interconnected carbon
rings, the so-called cyclopentanoperhydrophenanthrene rings, also known
as sterane (Fig. 1). Cholesterol serves as the primary precursor to steroid
synthesis, and the capacity and proclivity with which an organ will produce
a particular steroid depend on the activity of various enzymes required for
synthesis of the steroid. There are several competing systems of nomencla
ture for the various steroids, but we will rely on the names most commonly
associated with each hormone in the neuroscience literature. Steroids are
Methods in Neurosciences, Volume 11
Copyright © 1993 by Academic Press, Inc. All rights of reproduction in any form reserved.
