The mechanisms by which neuromuscular junctions are established either during the course of embryonic development or following denervation are little understood. The specific objective of this proposal is to test the hypothesis that an interaction of the developing motor nerve fibers with muscle cells triggers the development of postsynaptic specializations on the embryonic muscle cells. I will monitor the development of aneural superior oblique muscle of the eye to determine if the postsynaptic specializations develop in the absence of motor nerve fibers. Muscle will be made aneural by permanent destruction of the trochlear motor neurons well in advance of their normal projection to the muscle. I will monitor the development of synaptic folds, postsynaptic densities, acetylcholinesterase and basal lamina in normal and aneural muscles at the light and electron microscopic levels. In addition, I propose to determine if the number of neuromuscular junctions is regulated by the number of motor neurons innervating the muscle. Immunoglobulin G from patients with myasthenia gravis is known to prevent the death of trochlear motor neurons which occurs normally during the course of embryonic development. Myasthenia gravis immunoglobulin G will be used to hyperinnervate the muscle. I will monitor the total number of neuromuscular junctions in normal and hyperinnervated muscles at the light microscopic level. This proposal fits in with my long-term research objective which is to understand the nature of mechanisms and signals between the nerve and muscle, both during the course of normal development, and in diseases affecting motor neurons, neuromuscular junctions and muscle. The investigation proposed here will rigorously test whether the motor nerve is necessary for the differentiation of the postsynaptic elements of the neuromuscular junction or whether the differentiation of the postsynaptic specializations is preprogrammed in the developing muscle cell. Further, it will reveal whether the total number of neuromuscular synapses in the muscle is controlled by the nerve or whether it is preprogrammed in the muscle. The results of this investigation will also increase our basic understanding of the mechanisms by which synapses are formed during embryogenesis.
