Exploiting our expert anatomical knowledge of the fly's optic lobe, we will examine the expression and function of genes in Drosophila photoreceptor terminals and their target internourons in the first neuropile, the lamina. Our long-term objective, for which fly photorecepto tetrad synapses Are a genetically manipulable model, is to understand the functional organization of multiple-contact synapses (dyads, triads, etc.), their formation and the role of neural activity in this process. Current objectives are to study synaptic organization in photoreceptor terminals and their interneurons, and the functional role of genes that code for synaptic proteins, especially as defined by phenotypic analyses of, and patterns of gene expression in, the lamina. Using advanced confocal, quantitative-EM, serial-em, immuno-EM and E-D reconstruction methods, and skilled personnel to implement these, we will precisely assay the phenotypes of control and synaptic mutant flies. We will examine: A. The functional role of genes for synaptic proteins in photoreceptor terminals, from EM phenotypes of mutants such as milton, noC, amphiphysin, cspa and D-huntingtin, and of new mutants from screens being conducted in collaborating laboratories for whole-eye mosaic flies with ERG defects that are attributable to impaired transmission in the lamina. The in vivo and in vitro expression of gene products in photoreceptor terminals, using probes directed to the pre-synaptic site at the photoreceptor terminal and to its lamina post- synaptic target cells, visualized with deconvolved confocal images and localized by immuno-EM. C. Dynamic aspects of structural organization and gene expression at visual synapses, and the role of neural activity in both the development assembly of afferent photoreceptor tetrads and their feedback partners, and short-term synaptogenesis in adult flies after light-evoked activity. D. The micro-circuits of distal straita in the second neuropile, or medulla, innervated by long photoreceptor axon and lamina cell terminals, using targeted expression of EM markers in medulla cells, to provide a basis upon which to interpret mutant structure in this complex neuropile. The studies proposed here, because they aim to produce a basic model of synaptogenesis applicable to multiple-contact synapses, such as the dyads and triads found widely in visual systems, will contribute to knowledge of the perturbations in disease states to which visual synapses are susceptible during their growth, development and function, and to the genetic bases of these.
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