This proposal would continue a long-term investigation of phototransduction mechanisms in Limulus photoreceptors. CaM-kinase has been implicated in the light-dependent phosphorylation of arrestin, but its function is unknown. The possible role of CaM-kinase in regulating equivalent light and other transduction processes will be tested by intracellular injection of active kinase or kinase inhibitors. A second set of experiments will attempt to elucidate the late stages of transduction which remain unknown. These experiments are predicated on the hypothesis that the Ca2+ elevation produced by the phospholipase C (PLC)-IP3 pathway triggers a cyclase; the resulting elevation of cyclic nucleotide could activate the cyclic nucleotide-gated channels observed in earlier experiments on excised patches. If confirmed, this hypothesis would provide a complete pathway from rhodopsin to channel and a framework for understanding the degeneration produced by light in invertebrates. A final set of experiments builds upon the earlier observation that CaM may also be involved in an early step of transduction. Physiological and biochemical experiments are proposed to localize this step. Preliminary evidence suggests that it may be at PLC. These results, if confirmed, may reveal altogether new regulatory mechanisms for phospholipase and Ca2+, insight that may be important in understanding the role of this enzyme in vertebrate cones, where its role remains completely unclear.
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