Transducin Mutation Affects Both Activation and Recovery of Photoresponses in Transgenic Mouse Rods

Abstract

Abstract: : Purpose: The mutation responsible for the Nougaret form of dominant stationary night blindness in humans is a G38D missense mutation in the rod transducin alpha subunit (Gtα). The purpose of this study was to assess transducin function in a transgenic mouse model of this disease. Methods: Transgenic mouse lines expressing mutant GtαG38D on Gtα +/– and –/– backgrounds were generated. Transgenic and wildtype mice were sacrificed after overnight dark–adaptation and the retinas were removed. Using suction electrodes, responses of individual rods to brief flashes of light were recorded. Results: Rods expressing GtαG38D on a Gtα –/– background (G38D –/–) were significantly less sensitive than wildtype rods and displayed greatly delayed response recoveries. Both the flash strength required to generate a half–maximal response (Io) and the flash sensitivity of the dim flash response (Sf) indicated that the G38D –/– rods were more than an order of magnitude less sensitive than wildtype rods. Rods from GtαG38D mice on a Gtα +/– background (G38D +/–) were slightly less sensitive than wildtype rods as measured by both Io and Sf. The rate of rise of the light–activated phosphodiesterase (PDE) activity in G38D –/– rods was much smaller than wildtype rods. In G38D +/– rods, there was a slight decrease in the rate of rise of PDE activity. The recovery of photoresponses in G38D –/– rods was significantly slower than in wildtype rods. The response to dim flashes recovered with a time constant of 1–5 seconds, as compared to wildtype recovery time constant of approximately 200 ms. As flash strength increased, the time constant of recovery slowed further. Recovery of G38D +/– rods to dim flashes of light displayed normal kinetics. However, at the brightest flash strengths, response recovery was greatly delayed, often prolonging saturation for 10s of seconds. Conclusions: Mouse rods expressing GtαG38D were less sensitive and displayed impaired activation and recovery of the phototransduction cascade. The reduction in sensitivity and cascade activation in transgenic rods likely reflect both the lower expression levels of GtαG38D and the lowered ability of the activated form of GtαG38D to activate PDE. The delayed recovery of the photoresponse after bright flashes of light in G38D +/– rods may explain the dominant nature of the mutation in humans with the Nougaret form of night blindness.

Keywords: photoreceptors • protein structure/function • signal transduction