SYNAPTIC SPECIFICITY IN THE SPINAL CORD

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The long term objective of our studies is the identification of nerve terminal components involved in the formation and maintenance of specific patterns of synaptic connections in the spinal cord. Our fundamental hypothesis is that distinctive molecules are present in different populations of presynaptic nerve terminals which mediate selective interaction with appropriate target neurons to stabilize and maintain specific synaptic linkages. Our investigations have employed a monoclonal antibody to localize and partially characterize a novel nerve terminal antigen, termed NT75, which has several properties expected of a protein involved in selective synaptic development. NT75 is a neuron-specific, acidic 75KDa, membrane protein which makes a conspicuous appearance in selected populations of nerve terminals during synaptogenesis and is maintained in a subpopulation of nerve terminals in adulthood. Additional study of the NT75 protein is needed, however, before it can be considered a candidate for mediating selective synapse formation and maintenance. One set of studies will examine the localization and regulation of NT75 in developing, regenerating and adult nerve terminals. Electron microscopy will identify NT75 immunoreactivity in developing spinal nerve terminals to correlate the appearance of the protein with specific stages of synaptic maturation and with levels of NT75 during development. In dorsal root ganglion neurons maintained in vitro, studies will determine how NT75 is regulated in conjunction with neurite outgrowth, contact with appropriate vs. inappropriate target cells, synapse formation and synaptic activity. Additional studies will determine whether NT75 is present in developing motor nerve terminals and will examine the regulation of NT75 during peripheral nerve regeneration and abortive regeneration in the CNS. In situ hybridization combined with complementary immunocytochemical methods will also be used to determine precisely those neurons in spinal cord and brain which express NT75 in adulthood. Another set of studies will purify NT75, develop additional antibodies to the protein, and analyze the membrane orientation of NT75, especially with regard to whether the protein is exposed on the surface of the nerve terminal membrane. Finally, a cDNA library will be screened to obtain probes for the in situ hybridization studies and for future analyses of the amino acid sequence of the NT75 protein. The results of these studies are expected to have significant implications for understanding specific molecular mechanisms involved in the formation and maintenance of selective synaptic connections in spinal cord and brain.