The ultimate goal of this grant proposal is to describe at the molecular level developmental changes during neuromuscular junction formation. Toward this goal we will continue to study in detail alterations occurring during nerve-muscle interaction in cultures of the African toad, Xenopus laevis. The excellent visibility and the ease in experimental manipulations are two major advantages of this system. We will correlate the synaptic current with properties of acetylcholine (ACh) receptor channels at the initial stage of interaction. As a continuation of the previous work on Na+ currents in the pre-synaptic neuron, we will examine further CA2+ and K+ currents. We will analyse these currents at pre-synaptic varicosities, enlarged portions of neurites, which are amenable to patch clamping. We also have evidence bearing on the induction process for ACh receptor accumulation. We will further characterize this new developmental process as it may lead us to the molecular mechanism of induction. The developmental phenomena are ultimately controlled by genes. Multiple genes must be contributing to each process and phenotypic expression of those genes are orchestrated to produce the neuromuscular junction. By studying each contributing gene we can further dissect the process into finer elementary steps. The fruit fly, Drosophila malanogaster, with its short life cycle and genetic accessibility, is ideal for this kind of dissection. We will first describe electrophysiologically early developmental changes during neuromuscular junction formation. Then using various mutants which have a defect in synaptic transmission we will define and describe finer steps during this process. Neuromuscular diseases are often the final complex products of many intermediate reactions in various participating cell types. The primary causes of these events could be simple defective steps during early cell interactions. The results of this research proposal may provide information as to the primary defective step and possible cure for some neuromuscular diseases.
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