Abstract
Purpose :
The Cngb1 gene encodes the β-subunit of the rod photoreceptor cyclic nucleotide-gated (CNG) cation channel. GARP2, a splice variant of the Cngb1 gene, is exclusively expressed in rods and suggested to function as a structural protein, a calcium-binding protein, and a modulator of the basal activity of cGMP phosphodiesterase (PDE). We set out to further assess the structural and functional role of GARP2 within the retina.
Methods :
GARP2 knockout mice were generated using ZFN-mediated gene editing. Morphological features were assessed by optical coherence tomography and immunohistochemistry techniques at P180. Functional properties were then assessed by in-vivo electroretinogram (ERG) and whole-cell patch-clamp recordings from retinal slices.
Results :
The retina in KO mice exhibited no major alterations, except for occasional longer and bent rod outer segments. The maximum amplitude of the ERG a-waves (460 ± 150 µV vs. 306 ± 70 µV) and b-waves (1108 ± 333 µV vs. 589 ± 103 µV) were reduced in KO mice, with KO mice showing lower than predicted b-wave amplitudes for the measured a-wave. However,single-cell recordings showed no significant differences in the physiological properties of rods and RBCs between the genotypes. Surprisingly, KO rod photoreceptors showed a significant decrease in dark noise levels compared to WT (0.57 ± 0.07 ΔpA2/Hz vs. 0.3 ± 0.04 ΔpA2/Hz), p< 0.01).
Conclusions :
Our morphological analysis suggests that the retina in KO mice develops correctly, with a possible minor role for GARP2 in the structural stability in rod photoreceptors. Importantly, we were able to confirm the modulatory role of GARP2 on the basal activity of PDE, its important role in controlling dark noise levels and implications for regulating visual phototransduction and single-photon sensitivity.
This is a 2021 ARVO Annual Meeting abstract.