An Autoradiographic Study of Neuroglia
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This dissertation has been microfilmed exactly as received 6 6 —10 ,0 3 6 SCHWYN, Robert Conrad, 1938- AN AUTORADIOGRAPHIC STUDY OF NEUROGLIA. The Ohio State University, Ph.D., 1966 Anatomy University Microfilms, Inc., Ann Arbor, Michigan ( t ) Copyright by Robert Conrad Schwyn AN AUTORADIOGRAPHIC STUD! OF NEUROGLIA DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Robert Conrad Schwyn, B .S ., M.Sc. * * * * * * * The Ohio State University 1966 Approved by Adviser Department of Anatomy ACKNOWLEDGMENTS The author wishes to express his sincere gratitude to Dr. James L. H all, Department of Anatomy, School of Medicine, The Ohio State University, for his assistance and encouragement during this p ro je c t. A special word of thanks is extended to my wife, Jane, whose patience and devotion have made this endeavor possible, and to our daughter, Christine, who did her best. i i VITA June 8, 1938 . Born - Toledo, Ohio 1960 ..... B .S ., The Ohio S tate U niversity, Columbus, Ohio 1960-1962 . United States Aimed Forces, U.S. Array, Medical Service Corps, Landstuhl, Germany. 1962-196^ • • Graduate A ssistan t, Department o f Anatomy, The Ohio S tate U niversity, Columbus, Ohio 1963 ................. M.Sc., The Ohio S tate U niversity, Columbus, Ohio 1965 ................. Teaching A ssistan t, Department o f Anatomy, The Ohio S tate U niversity, Columbus, Ohio 1965-1966 . Teaching A ssociate, Department o f Anatomy, The Ohio S tate U niversity, Columbus, Ohio PUBLICATIONS "Studies of Neuroglial Activity in Autonomic Ganglia during Electrical Stimulation and Drug Administration." Anat. Rec., 151: March 1965. (Abstract) "A Modified Tri-basic Eye Technique for Neuroglia in Autonomic Gang lia." Ohio J. Sci., 6j>: 20^208. 1965. FIELDS OF STUD! Major F ield: Anatomy Studies in Neuroanatomy. Professor James L. Hall Studies in Gross Anatomy. P rofessors Linden F. Edwards and George R. L. Gaughran Studies in Comparative Anatomy. Professor W. James Leach Studies in Histology. Professor G. Adolph Ackerman Studies in Etabryology. Professor John C. Weston i i i CONTENTS Page ACKNOWLEDGMENTS i i VITA i i i INTRODUCTION 1 HISTORICAL REVIEW 5 METHODS 18 A. General operative procedures B. Autoradiographic techniques C. Autoradiographic development technique D. Staining procedure E. H istochem istry F. Standard histological technique OBSERVATIONS............................................. 27 I A* Observations of control ganglia removed from unstimulated animals B. Observations of stimulated superior cervical ganglia C. Observations of stimulated ganglia removed from drug-injected animals D. Histochemical observations of superior cervical ganglia DISCUSSION.............................................................. 41 CONCLUSIONS . .................... 56 SUMMARY . .................... .......................................... 58 APPENDIX ............................ 60 BIBLIOGRAPHY............................................. 67 iv INTRODUCTION The neuron doctrine, formulated by Waldeyer in 1891 and championed by Ramon y Cajal during the years to follow (Rasmussen, 19^7)» described the neuron as the anatomical, functional, embryo- logical, and trophic unit of the nervous system. Under the guidance of these fundamental aspects of neurology, our knowledge of nerve structure and function surged forward very rapidly, gathering momen tum as it progressed. In comparison, information related to the functional significance of the non-nervous cellular components of the nervous system remained relatively stagnant. These cells consist of a small nucleus, while their cytoplasm is drawn out into thin pro cesses. The original descriptive term applied to this mass of supporting cells was translated into the English language as "nerve glue" and was appropriate at that time. This trend of thought per sisted during the great boom of neurological investigation. Although there were earlier suggestions that neuroglia cells were involved in processes other than their own maintenance and neuron suspension, there was insufficient experimental data to lend much support to the concept. Within the last twenty years, our understanding of neuro glia elements has increased; but it has remained in the shadow of investigations conducted on neuronal function. At present, sufficient 1 data has been compiled to implicate these cells in the metabolic support of neurons. The supporting cellular elements located in peripheral ganglia were also designated as neuroglia because they demonstrated anatomical and physiological relationships similar to oligodendrocytes identified within the central nervous system. The functional capacities of these cells are even more obscure than those in the central nervous system. Although the term neuroglia, in reference to these cells, appears in most textbooks and periodicals, many prominent neuroanatomists today hesitate to identify them as neuroglia, perhaps avoiding terminology that may not'correctly reflect their functional or anatomical signi ficance. For want of better descriptive terminology, the supporting cellular elements located in the superior cervical sympathetic gang lion will be referred to collectively as neuroglial cells. These neuroglial cells will also be considered as interstitial or peri- neuronal satellite cells because of their anatomical location in relation to the neurons. Furthermore, perineuronal satellite cells will be referred to as either primary or secondary satellite cells, again determined by their location. The group of satellite cells excludes Schwann cells, endothelial cells, fibroblasts, or other connective tissue cells that may be present in the superior cervical ganglion. The premise of this investigation was the possibility that neuroglia within autonomic ganglia are involved in maintenance of environmental conditions which are conducive to efficient transmission 3 of an impulse. Many processes are involved during the transmission of a nervous impulse from preganglionic fibers to end organs receiving postganglionic fiber terminations. Synaptic transmission has been the subject of many histological, physiological, and pharmacological investigations. Most of the evidence, however, has been concerned with neuronal function, rarely implicating the presence of neuroglia or their functional activities. With the aid of previous data on the anatomical and biochemical properties of neuroglia cells and of autoradio graphic and histochemical techniques employed in this in vestigation, an attempt is made here to implicate neuroglial activity in relation to repetitive neuronal transmission. Autoradiographic techniques, although relatively new and in early stages of application, have been utilized very successfully during the past decade in determining the dynamics of cellular activity. Almost any biochemical compound or precursor can be labeled with various radioisotopes. Autoradiographic identification of a cell component, then, depends upon the availability of the labeled compound' and the time involved for its cellular absorption. Histochemical procedures were undertaken in order that the labeling of a cell component could be correlated with enzymatic and chemical activities occurring simultaneously within the ganglion. By apply ing electrical stimulation to preganglionic fibers terminating in the superior cervical ganglion, and in some instances introducing certain drugs into the organism, an attempt was made to alter and control the activity of gLial cells under specific known conditions. It is hoped that the results of this investigation will shed additional light upon the possibility of neuroglial involvement in synaptic activity and neuronal metabolism in autonomic ganglia. The following information may be of value to those interested in the functional relationship existing between neurons and neuroglia. Since facets of various disciplines are encountered during this study, it is presumed that individuals involved in anatomical, physiological, biochemical, and pathological research may find some aspect of this paper applicable to their own work. HISTORICAL REVIEW The earliest accounts of neuroglia investigation, according to Glees (1955)» date back to 18^6 -when Virchow first described these structures as independent cellular elements of the nervous system. This marked the beginning of an era concerned primarily with the morphological characteristics and identification of these newly identified elements. Descriptions of neuroglia cells and their processes were more numerous after the turn of the century when many new staining techniques were developed. Two of the most widely known staining methods are those of Golgi and Weigert. Glees (1955) stated that Held (1909) developed a technique by which he was able to show that glia cellular fibers form a syncytial net in which the primary nervous structures are suspended. In addition to advancing the theory that the neuroglia cells form a mechanical supporting network, Held suggested that they are involved in the metabolic activity of neurons. It was his identification of terminal end feet on glia processes attached to blood vessels which led him to suggest that glia elements may serve as a transport for metabolic products. According to Glees (1955)» Held also observed granular inclusions in glia processes which he thought to be secretion; further substantiating a function other than mechanical support. Held's observations, however, were highly advanced