We propose to study the factors controlling the formation of synapses and gap junctions by photoreceptors in the first neuropile, the lamina, of the fly's optic lobe. In particular, our long-term objective is to understand the developmental assembly of multiple-contact synapses (dyads, triads, etc.), for which the fly receptor tetrad synapses are a model. This entails aspects that are both qualitative (how individual tetrads assemble) and quantitative (their number and location). The modules of the lamina, called cartridges, comprise small, fixed numbers of identified neurons with which we are intimately familiar. We will continue our analysis of these cells with: 1) EM counts of synapse numbers by the disector method; 2) image analysis measurements of synaptic junction sizes and cell dimensions; 3) serial-EM based computer 3-D reconstructions of immature lamina cells; 4) cryo-ultrathin and immuno-EM methods. We will examine: A. Synaptic plasticity from rapid synaptic changes occurring during light adaptation, by studying the location of new synaptic sites relative to old ones, to examine the influence of an existing site on the formation of a new synaptic site nearby. We will also examine the activity-dependence of synapse formation in mutants with impaired transmission to postsynaptic cells. We will relate synapse formation to the pattern of dendrite growth in immature lamina cells reconstructed in 3-D, from the spacing and composition of immature and surviving synapses. We will seek the foreign targets and their synaptic contacts from the photoreceptor axons of genetically induced ectopic eyes. B. Expression of proteins that localize at synapses or gap junctions, or act during synaptogenesis in photoreceptors and lamina cells. Proteins for transmitter systems, gap junctions and cell adhesion will be expored with antibody probes. C. Drosophila mutants that introduce genetically defined perturbations to lamina cells. We will examine the structure of mutants that mis-express the cadherin and fas II genes, and ERG mutants that lack normal transmission to the lamina, and the dye-coupling between photoreceptors in a gap junction mutant. D. Finally, we will exploit a tissue-culture system we have developed, to examine selective fasciculation amongst photoreceptor axons and their interactions with their lamina cell targets. 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 a general knowledge of the perturbations in disease states to which visual synapses are susceptible during their growth and-development.
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