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Yan Cao, Johan Pahlberg, Ignacio Sarria, Naomi Kamasawa, Alapakkam P. Sampath, Kirill A. Martemyanov; RGS7 and RGS11 Provide the Main GAP for the Gαo and Set the Sensitivity and Time Scale of Rod ON Bipolar Light Responses. Invest. Ophthalmol. Vis. Sci. 2012;53(14):3154.
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The depolarizing light-evoked responses of ON bipolar cells (ON-BCs) are generated by a metabotropic signaling cascade involving the glutamate receptor, mGluR6, the heterotrimeric G protein, Gαo, and the cationic channel, TRPM1. The deactivation of Gαo is required to generate the rising phase of the light-evoked response in ON-BCs. Fast kinetics of the ON-BC response suggest that slow spontaneous deactivation of Gαo is catalyzed by GTPase Activating Proteins (GAP). However, the identity of the dominant GAP in the ON-BC that drives generation of light responses is not known. Here we studied the role of RGS7 and RGS11 proteins in ON-BC function.
We generated a line of knockout mice with complete elimination of RGS7. These mice were crossed with RGS11 knockouts to generate a line of double knockouts (DKO). Protein expression, localization, and synaptic morphology were analyzed by Western blotting, immunohistochemistry and electron microscopy, respectively. The light-evoked responses of ON-BCs were studied in intact retinas by ERG, and in single rods and rod bipolar cells from dark adapted retina using suction and patch electrodes, respectively.
We found that elimination of RGS7 alone did not alter the light-evoked responses of ON-BCs. However, simultaneous knockout of RGS7 and RGS11 resulted in severe disruption of ON-BC function with no effect on rod photoresponses. ERG recordings showed that threshold for the generation of b waves was much higher in the DKO mice. Furthermore, the onset of the b-wave was slowed by more than an order of magnitude. Single cell recordings revealed that flash responses were completely absent in DKO rod ON-BCs. However, bright steps of light in DKO rod ON-BCs elicited inward currents that were ~50-fold smaller and had ~20-fold slower onset as compared to wild-type rod ON-BCs. The synaptic morphology and retina cytoarchitecture in DKO was normal.
These results are consistent with persistently high G protein activity in the dendrites of DKO ON-BCs, suggesting that RGS7 and RGS11 together provide the dominant GAP in the mGluR6 signaling cascade that is essential for setting the sensitivity and temporal resolution of ON-BCs.
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