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Several model systems have been used to understand the cellular and molecular mechanisms of differentiation of mammalian nerve cells. Each model system has unique advantages and disadvantages and is suited for the study of only certain aspects of differentiation. In this book, the techniques of these models and the usefulness and limita tion of each model system are discussed. An awareness of the use and misuse of each model system is important for a rational interpretation of data and for a reasonable comparison of data obtained from different model systems. With the use of clonal lines of neuronal cells and hybrid neural cells (neural cells x nonneural cells), many new concepts have emerged con cerning the regulation of differentiated functions, the relationship between the expressions of individual differentiated functions, and the relationship between differentiation and malignancy. Some of these concepts have already proved to be relevant to regulation of differentiation in vivo. These new emerging concepts are discussed extensively in this book. Many new agents (physiological and nonphysiological) which induce or increase the expression of one or more differentiated functions have been identified. These agents will be useful biological tools for further studies of the regulation of differentiation in mamma lian nerve cells. This book describes the role of each agent in d- vii viii PREFACE ferentiation of nerve cells by focusing on different model systems and provides a rational basis for selecting the particular differentiating agents for specific problems of differentiation processes.
This volume is one in a series of monographs being issued under the general title of "Disorders of Human Communication". Each monograph deals in detail with a particular aspect of vocal communication and its disorders, and is written by internationally distinguished experts. Therefore, the series will provide an authoritative source of up-to-date scientific and clinical informa tion relating to the whole field of normal and abnormal speech communication, and as such will succeed the earlier monumental work "Handbuch der Stimm und Sprachheilkunde" by R. Luchsinger and G. E. Arnold (last issued in 1970). This series will prove invaluable for clinicians, teachers and research workers in phoniatrics and logopaedics, phonetics and linguistics, speech pathology, otolaryngology, neurology and neurosurgery, psychology and psychiatry, paediatrics and audiology. Several of the monographs will also be useful to voice and singing teachers, and to their pupils. G. E. Arnold, Jackson, Miss. F. Winckel, Berlin B. D. Wyke, London Preface Neurologists, neuropsychologists, speech pathologists and other clinicians who care for dysphasic patients have often complained that available books on dysphasia tend to be parochially theoretical, and insufficiently directed towards clinical reality. These books provide the categories, labels, and theoretical speculations of one school or another; but dysphasic patients as often as not do not fit neatly into a specific theoretical category. Clinical patterns of dysphasic syndromes of most patients with dysphasia rarely conform fully to the pictures painted in the textbooks.
A fundamental problem in neuroscience is the elucidation of the cellular and molecular mechanisms underlying the development and function of the nervous system. The complexity of organization, the heteroge neity of cell types and their interactions, and the difficulty of controlling experimental variables in intact organisms make this a formidable task. Because of the ability that it affords to analyze smaller components of the nervous system (even single cells in some cases) and to better control experimental variables, cell culture has become an increasingly valuable tool for neuroscientists. Many aspects of neural development, such as proliferation, differentiation, synaptogenesis, and myelination, occur in culture with time courses remarkably similar to those in vivo. Thus, in vitro methods often provide excellent model systems for investigating neurobiological questions. Ross Harrison described the first culture of neural tissue in 1907 and used morphological methods to analyze the cultures. Since that time the technique has been progressively modified and used to address an ever widening range of developmental questions. In recent years a con vergence of new or improved cell culture, biochemical, electrophysiol ogical, and immunological methods has occurred and been brought to bear on neurobiological questions. This volume is intended not to be comprehensive but rather to highlight some of the latest findings, with a review of previous important work as well, in which combinations of these methods are used.
This symposium entitled Calcium, Neuronal Function and Transmitter Re lease, was in the framework of the regional meeting of the International Union of Physiological Sciences, that took place in Jerusalem between August 26-31, 1984. The symposium dealt with the role of calcium ions in regulation of a large number of important processes in modern neurobiology, from molecular and cellular points of view. In this context, we consider heart and most cells as 'honorary neurons'. The meeting was comprised of lectures and quite intense discussions. We hope that the transcription of the discussions which follow the articles will give the reader a feeling of the intense, but pleasant atmosphere that per vaded during this symposium. It is our pleasure to thank Mrs. Miriam Silber, the assistant editor of this book, for her hard work in transcribing the discussions, retyping large portions of the book, and getting the approval of the authors. We express our thanks to Dr. Halina Meiri and Dr. Simona Ginsburg for their editorial assis tance. The unfailing help of Ms. Rachel Klein and Ms. Shoshana Wineberg is greatly appreciated. xxvii SECTION 1 CALCIUM CHANNELS, TRANSPORTERS & CALCIUM REGULATED CHANNELS EFFECTS OF TEMPERATURE ON SINGLE CHANNEL AND WHOLE CELL Ca CURRENTS A. M. BROWN, D. L. KUNZE, H.D. LUX AND D. L. WILSON INTRODUCTION Calcium tall currents have a large,fast component which Is not detectable when the currents are turned on or acti vated from the completely rested state (Brown et aI, 1983).
A major advance in the biological sciences in the past decade has been the biochemical identification of cell membrane receptors. The existence of re ceptor substances on the surface of cells that recognize and bind to extracellular molecules was proposed at the beginning of the century by the pharmacologist and immunologist Paul Ehrlich and the physiologist J. N. Langley. Since then, receptors have been found to play an important role in numerous physiological and pathological processes. Over the years many attempts have been made to physically isolate and chemically characterize receptors, but because of the receptors' extremely low concentration and membrane localization, these ef forts have met with limited success. Yet, despite the failure to characterize receptor substances, the concept of the presence of such molecules has had considerable heuristic value. Using pharmacological, physiological, and im munologic approaches, researchers have identified several specific receptors, e. g. , a-and ~-adrenergic, nicotinic and muscarinic cholinergic, and histami nergic. With the characterization of various types of receptors on cell mem branes, many drugs were developed that proved to be experimentally and ther apeutically useful. It was only in the early 1970s that methods for the specific measurement, chemical characterization, and physical isolation of cell membrane receptors were developed. These advances were made possible by the availability of ligands with high specific radioactivity that retained their biological activity and of experimental procedures that differentiated between specific and non specific binding of ligands.
During the past decade, great strides have been made in our un derstanding of the biochemistry and pharmacology of the alpha-l adrenergic receptor. The alpha-l adrenergic receptor plays a key role in biological function. This is evidenced by the fact that the alpha-l adrenergic receptor plays a prominent functional role in most organs of the body and in the key systems responsible for survival of the organism and maintenance of optimum biological activity. This is most apparent in the cardiovascular system, in which alpha-l adrenergic receptors are the single most important receptor involved in the maintenance of blood pressure and circu latory function. It is appropriate, therefore, that recent findings related to the pharmacology and biochemistry of the alpha-l adrenergic receptor be compiled, since this subject has not been reviewed in detail in recent years. It is the purpose of this book to present a series of reviews of key experimental findings that shed new light on the alpha-l adrenergic receptor and the manner in which it functions. Classically, most receptors have been characterized based on structure-activity relationships obtained for selective agonists and antagonists interacting with the receptor. Although there are many newer and more sophisticated approaches to receptor char acterization, structure-activity relationships still provide impor tant information regarding the chemical requirements made by the receptor for its occupation by ligands and its subsequent acti vation by those ligands possessing intrinsic efficacy and, there fore, agonist activity.
Work in the field of neuroprosthetics requires multidisciplinary teams, but these collaborators must meet on common ground to develop an understanding of the capabilities and limitations of each part of a bioengineering project. The Handbook of Neuroprosthetic Methods provides a comprehensive resource for the techniques, methodologies, and options available to properly design and undertake experiments within the field of neuroprosthetics. It combines the most commonly employed concepts, applications, and knowledge from the many disciplines associated with neuroprosthetic research to foster more effective, profitable, and productive collaborations.

From basic neurophysiology to emerging technologies, this book provides a clear introduction to the entire range of neuroprosthetic systems. Each chapter includes background information, methodology, illustrative figures that clarify experimental methods, and tables that outline and compare experimental choices. The last part of each chapter provides practical applications and examples that relate the topic to the actual design and implementation of a neuroprosthetic system or device.

Through its exploration of a variety of developmental processes, the book provides guidance on issues that have yet to be solved, strategies for solving such problems, and the pitfalls often encountered when developing neural prostheses. Whether you are new to or a veteran of the field, whether you work directly or indirectly with neuroprosthesis projects, the Handbook of Neuroprosthetic Methods provides an accessible common ground for all involved in neuroprosthetic design and research.
The original premise of the Editors in initiating this series was that there existed a readership ofneurochemists with considerable biochemical back ground who would make use of a series dedicated to both new develop ments and specialized reviews in neurochemistry. Having selected our authors, we have offered them virtually complete freedom to reflect and speculate in a field in which they have achieved prominence. The response to the first two volumes has been rewarding. The present one continues in this tradition. While we have not attempted to publish specialized volumes, the present volume contains two somewhat related chapters (Chapters 4 and 5, on the role of amino acid neurotransmitters). The first three chapters examine three diverse approaches, each of current interest, in neurochemi cal approaches to the molecular bases of neuronal and glial structure. B. W. Agranoff M. H. Aprison vii CONTENTS CHAPTER 1 2',3'-CYCUC NUCLEOTIDE 3'-PHOSPHODIESTERASE NEIL RAYMOND SIMS AND PATRICK ROBERT CARNEGIE 1. Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 2. Assay of CNPase . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 2. 1. Need for Activation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 2. 2. Comparison of Assays . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 3. Association of CNPase with Myelin . . . . . . . . . . . . . . . . . . . . . . . 7 3. 1. Historical. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 3. 2. Subcellular Fractionation. . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 3. 3. Development. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 3. 4. Mutant Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9 3. 5. Use as a Myelin Marker . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 3. 6. Myelin-Related Fractions and Peripheral Nerve Myelin. 11 4. CNPase in Nonmyelin Fractions. . . . . . . . . . . . . . . . . . . . . . . . . . 13 5. Activation and Isolation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13 5. 1. Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13 5. 2. Activation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14 5. 3. Solubilization and Fractionation. . . . . . . . . . . . . . . . . . . . . . 15 6. Properties. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19 6. 1. Substrate Specificity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
One of the most active and productive areas of biological science in the past decade has been the study of the biochemical and biophysical prop erties of cell membranes. There is little doubt that membranes are essen tial components of all cellular systems and that each type of membrane manifests specific and characteristic cellular functions. In the nervous system, important events such as neurotransmission, receptor binding, ion transport, axonal transport, and cell uptake are all known to take place within the neural cell membrane. Phospholipids, one of the major components of membranes, not only provide the membrane with its structural integrity and physical proper ties, but also play an important role in regulating membrane function. Attention has recently been focused on the asymmetric localization of these molecules, the identification of discrete metabolic pools of phospholipids within the membrane matrix, and their involvement in sig nal transmission. Although synaptic membranes generally lack an active mechanism for the de novo biosynthesis of phospholipids, a number of enzymic routes are present for their interconversions and for facilitating metabolic turnover. Metabolites generated during the interconversion reactions may also exert a great influence in modulating membrane func tions. The phosphogylcerides of neural membranes are especially enriched in polyunsaturated fatty acids. However, only very small amounts of these fatty acids are present in the free form, and they are maintained in dynamic equilibrium with the membrane phospholipids.
This book is a result of our combined major interests in oral and facial function. Since most of our research efforts have been concentrated on fundamental neural mechanisms, the book emphasizes basic research in this area. However, our back grounds in clinical dentistry have always made us acutely aware of the relevance of these findings to clinical problems in dentistry and medicine, and such correlations are emphasized throughout the text. The term, "oral and facial function," will here include the sensory and motor neural mechanisms of the face, mouth, pharynx, and larynx. Detailed discussions of nasal function, olfaction, and speech mechanisms have been omitted; these areas would encompass a book in themselves. A chapter on the subject of taste presents a brief overview in relation to other chapters in the book and clinical significance. We have not intended each chapter to be a review of the literature in a given area but have chosen to emphasize significant findings for total function of the area. References are limited to review articles whenever possible and the reader is invited to search such reviews for original articles of interest. Where such reviews are not available, original articles are usually referenced so that the book provides a path to source material for those so inclined. Some of the chapters on special areas of interest such as teeth, periodontium, and jaw reflexes, however, are extensively referenced because of their unique relationship to the subject matter of the book.
Historically, the major emphasis on the study of purinergic systems has been predominantly in the areas of physiology and gross pharmacology. The last decade has seen an exponential in crease in the number of publications related to the role of both adenosine and A TP in mammalian tissue function, a level of interest that has evolved from a more molecular focus on the identity of adenosine and A TP receptor subtypes and the search for selective ligands and development of radioligand binding assays by Fred Bruns and colleagues (especially that for A receptors) that played z a highly significant role in advancing research in the area. In the 60 years since adenosine was first shown to be a potent hypotensive agent, a considerable investment has been made by several pharmaceutical companies-including Abbott, Byk Gulden, Takeda, Warner-LambertlParke Davis, Boehringer Mann heim, Boehringer Ingelheim, Nelson/Whitby Research and CffiA Geigy-as well as John Daly's laboratory at the National Institutes of Health, to design new adenosine receptor ligands, and both agon ists and antagonists with the aim of developing new therapeutic entiities. Numerous research tools have derived from these efforts including: 2-chloroadenosine, R-PIA (~-phenylisopropyladeno sine; NECA (5' N-ethylcarboxamidoadenosine); CV1808; CI936; PD 125,944; ~-benzyladenosine; PACPX; CPX; CPT; XAC; CGS 15943 and CGS 21680. Yet in the realm of therapeutics it was only in 1989 that adenosine itself was approved for human use in the treatment of supraventricular arrythmias.
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