Dynamics of Neuron-Specific Gene Expression During Development and in Response to Selective Lesions of the Rat Central Nervous System: A Dissertation

Abstract

Synapse development and injury-induced reorganization in the nervous system have been extensively characterized morphologically, although, relatively little is known regarding the molecular and biochemical events that underlie these processes. In an attempt to better understand, at the molecular level, the role of the expression of synaptic proteins during synapse establishment and regeneration, this dissertation examines the dynamics of expression of the neuron-specific gene synapsin I during development and in response to selective lesions of the rat central nervous system. Synapsin I is the best characterized member of a family of nerve-terminal specific phosphoproteins implicated in the regulation of neurotransmitter release. During development, the expression of synapsin I correlates temporally and topographically with synapse formation, and recent physiological studies by Lu et al., (1992) have suggested that synapsin I may participate in the functional maturation of synapses. To better understand the temporal relationship between synapsin I gene expression and particular cellular events during development, we have used in situhybridization histochemistry to localize synapsin I mRNA in the rat central and peripheral nervous systems throughout embryonic and postnatal development, and into the adult period. During development, from the earliest embryonic time point examined (E12), the expression of the synapsin I gene was detectable in both the rat central and peripheral nervous systems. While, in general, levels of synapsin I mRNAs were high in utero, synapsin I cDNA probes revealed specific patterns of hybridization in different regions of the embryonic nervous system. To precisely determine the temporal onset of expression of the synapsin I gene during neuronal development, we examined in detail the appearance of synapsin I mRNA during the well characterized postnatal development of the cerebellum and hippocampus. In both regions, the onset of synapsin I gene expression correlated with the period of stem cell commitment to terminal differentiation. In a second phase, in accord with prior analyses, synapsin I gene expression increases to a maximum for a given neuronal population during synapse formation. In the adult rat brain, our data demonstrates a widespread yet regionally variable pattern of expression of synapsin I mRNA similar to that seen at earlier time points, with noteworthy exceptions. The greatest abundance of synapsin I mRNA was found in the pyramidal neurons of the CA3 and CA4 fields of the hippocampus, and in the mitral and internal granular cell layers of the olfactory bulb. Other areas abundant in synapsin I mRNA were the layer n neurons of the piriform and entorhinal cortices, the granule cell neurons of the dentate gyrus, the pyramidal neurons of hippocampal fields CA1 and CA2, and the cells of the parasubiculum. In general, the pattern of expression of synapsin I mRNA paralleled those encoding other synaptic terminal-specific proteins, such as synaptophysin, VAMP-2, and SNAP-25. Then, to determine specifically how synapsin I mRNA levels are related to levels of synapsin I protein in the adult rat brain, we employed in situhybridization histochemistry and immunohistochemistry to examine in detail the local distribution of both synapsin I mRNA and protein in the hippocampus. In short, these data revealed differential levels of expression of synapsin I mRNA and protein within defined synaptic circuits of the rat hippocampus. Based on these data we hypothesized that locally high levels of synapsin I mRNA in neuronal somata may reflect the ability of the nervous system to respond to select enviromental stimuli and/or injury by producing longterm changes in synaptic circuitry. To test this hypothesis and to better understand the regulation and putative role of synapsin I gene expression in the development of functional synapses in the central nervous system, we first examined the developmental pattern of expression of the synapsin I gene; in dentate granule neurons of the dentate gyrus and their accompaning mossy fibers during the main period of synaptogenic differentiation in the rat hippocampus. The results of these studies indicate a significant difference between the temporal expression of synapsin I mRNA in dentate granule cell somata and the appearence of protein in their mossy fiber terminals during the posmatal development of these neurons. Next, to investigate the regulation and putative role of synapsin I gene expression during the restoration of synaptic contacts in the central nervous system, we examined the expression of the synapsin I mRNA and protein following lesions of hippocampal circuitry. These studies show marked changes in the pattern and intensity of synapsin I immunoreactivity in the dendritic fields of dentate granule cell neurons following perforant pathway transection. In contrast, changes in synapsin I mRNA expression in target neurons, and in those neurons responsible for the reinnervation of this region of the hippocampus, were not found to accompany new synapse formation. On a molecular level, both developmental and lesion data suggest that the expression of the synapsin I gene is tightly regulated in the central nervous system, and that considerable changes in synapsin I protein may occur in neurons without concommitant changes in the levels of its mRNA. From a functional standpoint, our results suggest that the appearance of detectable levels of synapsin I protein in developing and sprouting synapses does not reflect simply synaptogenesis, but coincides with the acquisition of function by those central synapses.