The neuromuscular junction (NMJ) serves as a model in the analysis of aglng of synapses. We propose to evaluate the mechanisms of two improtant age changes at the NMJ - altered transmitter release and increased outgrowth and retraction of motor nerve terminals. Indirect data indicate that the alteration in synaptic transmission in aging NMJ is due to altered nerve terminal depolarization or calcium influx. Because mamalian motor nerve terminals are too small for intracellular recording, these two key parameters of synaptic efficacy will be evaluated with new voltage- or calcium-sensitive fluorescent dyes. With additional electrophysiologic techniques, the ion channel properties of synaptic terminals will be determined as a function of age and directly related ion channel properties of synaptic terminals will be determined as a function of age and directly related to altered transmitter release in individual identified junctions. By comparing muscles in which there is age related increase, decrease, or unchanged transmitter release in individual identified junctions. By comparing muscles in which there is age-related increase, decrease, or unchanged transmitter release, physiologically relevant age changes can be separated from those that do not correlate with altered transmitter release. Also age changes in propogation or wave-form of depolarization and those involving calcium channels can be distinguished. This new approach will also shed light on synaptic function in young animals. Two hypotheses will be tested which account for the morphologic remodelling of NMJ in aging mice. The first is that adhesion of nerve terminal to synaptic matrix is reduced, allowing nerve terminal outgrowth. The following predictions will be tested with immunocytochemical and tissue culture techniques; 1. During regeneration in vivo or outgrowth of ciliary ganglion cells on cryostat sections in vitro, preferential adhesion to synaptic sites will be diminished in old muscle; 2. Intrinsic neuronal or extrinsic matrix components related to adhesion will be reduced with age; and 3. Outgrowth sites of nerve terminals will be associated with reduced density of adhesive components. A second hdypotheseis is that nerve terminal remodelling in old mice results form diminution of supply or detabolism and removal of membrane components. Microimmunocuytochemical methods have been devised to compare amounts of transported material individual motor nerves. These studies will elucidate mechanisms of morphologic instabilty of aging synapses.
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