Signal transduction in rod photoreceptors begins with light activation of rhodopsin and ends with a decrease in the intracellular level of cyclic GMP. The decrease in cGMP is what the rod uses to produce an electrical signal. The recovery of cGMP level, and termination of the light-induced signal depends on the deactivation of a rod cell component called phosphodiesterase (PDE). A defect in PDE -- or in one of the components that regulate PDE activity -- will result both in abnormal levels of cGMP, and in poor vision. For example, retinitis pigmentosa (RP) is a group of disorders that causes retinal degeneration. An early symptom of RP is night blindness. One hypothesis for the underlying basis for sensitivity deficits in RP patients, is that cGMP metabolism in rods is defective. Testing this hypothesis requires an understanding of the role each cellular component plays in the light activation and subsequent deactivation of PDE. The long term objectives of this application are (i) to examine processes involved in the inactivation and adaptation of the photoresponse in normal vision, and (ii) to examine whether defects in these processes underlie visual dysfunctions that occur in diseases such as RP. One specific objective of this research is to examine the effect of light on the cellular localization and metabolic regulation of a retinal protein designated """"""""48K"""""""", """"""""S- antigen"""""""" or """"""""arrestin"""""""" that has been shown to quench light activation of PDE. A second specific objective is to examine whether reversible methylation of PDE is a mechanism for regulating PDE activity in the presence of steady background illumination. These studies will provide normative data about processes that are central to the regulation of PDE activity in rods. In view of (i) the role of PDE in the regulation of cGMP levels (ii) the central role of cGMP in phototransduction, and (iii) data indicating that abnormal cGMP metabolism can result in poor vision, proposed experiments will provide much needed information about processes that may underlie certain forms of retinal degenerative diseases such as RP. To achieve these goals, the following experiments are proposed: 1) To examine whether light induces a migration of 48K from the inner segment of rod photoreceptor to the outer segment. Experiments involve quantitation of 48K in dark- (DA) vs. light- adapted (LA) rod outer segments. 2) To measure the turnover rate of 48K in DA rod photoreceptors, and to examine whether light adaptation affects turnover rate of 48K. Experiments involve radioactive labeling of retinal proteins, and determination of specific activity of 48K in retinal subcellular fractions. 3) To examine whether reversible methylation of PDE is involved in adaptational processes in rods.
