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M. L. Woodruff, E. V. Olshevskaya, M. C. Cilluffo, M. C. Cornwall, P. A. Sieving, G. L. Fain, A. M. Dizhoor; Congenital Night Blindness and the Mechanism of Constitutive Signaling of G90D Rhodopsin. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5829.
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Rhodopsin G90D produces autosomal dominant stationary night blindness. Rods are desensitized by an equivalent background light, but the origin of the equivalent background remains unclear.
We made suction-electrode recordings and Ca measurements from rods of G90D mice (Woodruff et al., 2007) produced in Rpe65+ or Rpe65 knockout backgrounds to evaluate G90D activity in vivo and the effect of regeneration with 11-cis retinal.
G90D+/-;Rh+/- (D+) retinas showed no evidence of degeneration, as previously reported (Sieving et al., 2001); however G90D+/+;Rh-/- (D+/+) retinas showed a slow loss of nuclei, consistent with greater constitutive stimulation of the transduction cascade in these retinas. D+ and D+/+ rods were less sensitive than WT; comparison to background light in WT rods indicated that the mutations produced an equivalent background of 130 and 890 Rh* s-1, similar to values previously reported. Response kinetics resembled light-adapted WT rods, with decreases in both tauREC and tauD. Free Ca was also decreased, from 240 nM in WT to 143 + 31 nM in D+ and 119 + 15 nM in D+/+. We investigated a role for opsin in producing the desensitization by perfusing D+ or D+/+ rods with lipid vesicles containing 11-cis retinal. We saw little change in sensitivity or response waveform, though 11-cis retinal produced nearly complete recovery of both sensitivity and tauREC in Rpe65 knockout rods. As a control for poor regeneration of G90D pigment, we added 11-cis retinal to D+ (Rh-/-) rods lacking Rpe65. The 11-cis retinal now produced a significant increase in sensitivity near to that for D+ rods with functional Rpe65. The equivalent background seems therefore to be produced by some form of rhodopsin rather than by opsin but is not generated by transitions from Rh to Rh*, since D+ rods do not have excess noise in darkness equal to that generated in a WT rod by real light producing an equivalent desensitization.
Our results indicate that desensitization produced by G90D in a mammalian rod is not generated by unregenerated opsin, as previously believed, but rather by some form of the mutant rhodopsin, which stimulates the cascade with low gain. There is however so much G90D rhodopsin in a dark-adapted rod that this stimulation is sufficient to produce night blindness.
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