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Justin D. Berry, Rikard Frederiksen, Soile Nymark, M. Carter Cornwall; Phosphorylation State Of Opsin Affects The Rate Of Pigment Regeneration In Mouse Rod Photoreceptor Outer Segments. Invest. Ophthalmol. Vis. Sci. 2012;53(14):748.
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Following exposure to bright bleaching light, the sensitivity of rod photoreceptors in the intact eye recovers slowly, with full dark-adapted sensitivity reestablished in 30-40 min. We tested the hypothesis that the slow rate of pigment regeneration which is generally thought to underlie sensitivity recovery is limited by the slow rate of dephosphorylation at rhodopsin’s c-terminus.
Microspectrophotometric measurements of rhodopsin concentration were made from outer segments of rods in patches of retinae from wild-type and GRK1-/- mice. Spectra were measured in darkness and after exposure to a bright light that bleached ~ 50% of the visual pigment. These bleached retinae were then left in darkness for sufficient time to allow complete decay of bleaching photoproducts, Meta II and Meta III. Bleached retinae were then treated exogenously with 11-cis retinal to regenerate fully the visual pigment. We measured and compared the time course of visual pigment regeneration in wild-type and GRK1 -/- outer segments under these conditions to determine the effect that opsin phosphorylation has on the intrinsic rate of visual pigment regeneration in intact rod outer segments.
Visual pigment regenerated in outer segments of wild-type mice with a time constant of ~ 9 min; complete recovery occurred over about 30 min. In contrast, pigment regeneration in GRK1-/- rod outer segments treated identically, regenerated with a time constant of about 3 min, more than 3-fold faster than wild-type. Under these conditions over 95% of the visual pigment was regenerated within about 10-12 min after retinoid exposure.
Visual pigment regeneration in rods in which opsin phosphorylation is not possible occurs very rapidly, on a time course that is more typical of cone photoreceptors. We propose that the slowness of visual pigment regeneration in wild-type rods occurs for one of two reasons: (1) Visual pigment regeneration is slowed by a slow rate of dephosphorylation of bleached opsin, or (2) Visual pigment regeneration is slowed by the binding of arrestin to phosphorylated opsin, which must unbind before rapid pigment regeneration can take place. Experiments with arrestin-/- mice are underway to differentiate between these two possibilities.
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