Function of the visual pigment in situ

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DESCRIPTION (provided by applicant): The sensitivity and response kinetics of a vertebrate photoreceptor depend on the intrinsic properties of the visual pigment molecule, such as its spectral sensitivity, as well as environmental factors that affect visual pigment function, such as the lateral mobility of rhodopsin in the disk membrane. The goal of this application is to explore the impact of two environmental factors on the photoresponse: rhodopsin packing density and type of transducin present. The first three components involved in the activation of phototransduction: rhodopsin, transducin and phosphodiesterase, interact on the disk membrane surface. Rhodopsin kinase and RGS9, two key proteins in the shutoff of phototransduction are also attached to the membrane. In normal rods, the rate-limiting steps in the activation and recovery of the photoresponse are limited by the rates of molecular collisions on the disk membrane surface. These rates are slowed by the prohibitive effect of the high rhodopsin packing density on membrane protein mobility, because a 50% reduction in the rhodopsin packing density accelerated the rising and falling phases of the photoresponse. The effect of a further decrease in rhodopsin packing density on the photoresponse cannot be predicted reliably, because the identity of the rate-limiting step could change. The magnitude of the effect of an increase in packing density cannot be predicted either, because the change in protein mobility with packing density is not known in vivo. These conditions will be tested in rods of transgenic mice that express sub- and supranormal amounts of rhodopsin in their disk membranes. Phototransduction will be evaluated from suction electrode recordings of the electrical responses of the rods to flashes and steps of light. Rods and cones differ in their sensitivity and response kinetics. They appear to have similar pigment packing densities but in general, the identities of their pigments and transducins are different. The effects of pigment and transducin type on photoresponse sensitivity and kinetics will be tested in rods and cones of salamander, where green rods and blue-sensitive cones couple the same visual pigment to different transducins and UV-sensitive cones contain multiple pigments coupled to the same transducin.