Abstract
Purpose :
The inactivation of visual pigments by GRK1-mediated phosphorylation is essential for the termination of rod and cone photoresponses and the recovery of photoreceptor sensitivity following a bleach. Recently, the negative regulation of GRK1 activity via cAMP-dependent phosphorylation of its amino-terminal Ser-21 residue in the dark has been described in wild type mouse retinas. Here, we tested the hypothesis that this process contributes to timely dark adaptation of mammalian rods.
Methods :
We generated mice in which the amino acid Ser-21, the cAMP-dependent phosphorylation site in GRK1, has been substituted by Ala (GRK1-S21A). The expression of GRK1 was determined by western blotting, and retinal morphology was analyzed by H&E staining and light microscopy. We assessed the function of control and mutant rods by transretinal ERG recordings. The effect of the S21A mutation in GRK1 on rod photoreceptor dark adaptation was determined by in vivo ERG.
Results :
Mutating GRK1 to block its cAMP-dependent phosphorylation at Ser-21 site did not affect the total expression level of GRK1 in mouse rods and did not cause retinal degeneration by up to 5 months of age. Rod photoresponse amplitude and sensitivity were identical in dark-adapted control and GRK1-S21A animals. The lack of GRK1 phosphorylation at Ser-21 did not reduce the amplification of rod phototransduction. The response recovery following dim to saturating test flashes was also normal in GRK1-S21A rods. However, following >90% pigment bleach, the dark adaptation of GRK1-S21A rods was compromised in vivo, and the final relative level of response amplitude recovery 60 min after the bleach was ~12% lower in mutant mice. Similarly, the recovery of their rod photosensitivity was reduced compared to controls.
Conclusions :
The phosphorylation-dependent quenching of GRK1 activity in the dark does not affect the kinetics of rod phototransduction cascade. However, it modulates the dark adaptation of mouse rods in vivo, allowing more efficient resetting of rhodopsin’s ground state after photoactivation. As in the mouse GRK1 is found not only in rods but also in cones, experiments seeking to determine whether this modulation takes place in cones are currently underway.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.