ROLE OF CYCLIC GMP IN ROD PHOTORECEPTOR FUNCTION

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The objective of this research is to elucidate the biochemical mechanism by which cGMP influences visual transduction in vertebrate rod photoreceptors. Both biochemical studies of the cGMP pathway (rhodopsin to GTP-bining protein to phosphodiesterase) and electrophysiological effects of cGMP on whole cell recordings and isolated membrane patches demonstrate the importance of cGMP in modulating the light-dependent permeability mechanism in the plasma membrane. The molecular mechanism by which cGMP acts has not, however, been determined. The aims of the proposed research are as follows: (1) To evaluate whether small light-induced decreases in total cGMP concentration reflect larger changes in cGMP concentration in the cytoplasm. The observed decrease in total cGMP concentration is too small to exert a significant effect on the permeability mechanism by a simple binding model. However, subcellular partitioning of cGMP into free and bound pools may occur, and will be examined by fractionating total cellular cGMP to determine whether a significant portion of total cGMP is bound and hence not sensitive to rapid light-induced hydrolysis by phosphodiesterse. (2) To identify possible sites of action of cGMP in the rod. cGMP is known to have effects on ion permeability of the plasma membrane, Ca2+ release from disc membranes, protein kinase activity, and GTP-binding protein. Identification and characterization of cGMP binding sites will lead to purification of protein components that may regulate, via changes in cGMP concentration and/or binding, the ionic permeability of the rod. These experiments should permit more definitive tests of various models for cGMP action in visual transduction, and lead to increased understanding of why defects in cGMP meatabolism lead to photoreceptor cell degeneration and death.