Endogenous RGS proteins enhance acute desensitization of GABAB receptor-activated GIRK currents in HEK-293T cells

The coupling of GABA(B) receptors to G-protein-gated inwardly rectifying potassium (GIRK) channels constitutes an important inhibitory pathway in the brain. Here, we examined the mechanism underlying desensitization of agonist-evoked currents carried by homomeric GIRK2 channels expressed in HEK-293T cells. The canonical GABA(B) receptor agonist baclofen produced GIRK2 currents that decayed by 57.3 +/- 1.4% after 60 s of stimulation, and then deactivated rapidly ( time constant of 3.90 +/- 0.21 s) upon removal of agonist. Surface labeling studies revealed that GABA(B) receptors, in contrast to l opioid receptors (MOR), did not internalize with a sustained stimulation for 10 min, excluding receptor redistribution as the primary mechanism for desensitization. Furthermore, heterologous desensitization was observed between GABA(B) receptors and MOR, implicating downstream proteins, such G-proteins or the GIRK channel. To investigate the G-protein turnover cycle, the non-hydrolyzable GTP analogue (GTP gamma S) was included in the intracellular solution and found to attenuate desensitization to 38.3 +/- 2.0%. The extent of desensitization was also reduced ( 45.3 +/- 1.3%) by coexpressing a mutant form of the G alpha q G-protein subunit that has been designed to sequester endogenous RGS proteins. Finally, reconstitution of GABA(B) receptors with G alpha o G-proteins rendered insensitive to RGS resulted in significantly less desensitization ( 28.5 +/- 3.2%). Taken together, our results demonstrate that endogenous levels of RGS proteins effectively enhance GABA(B) receptor-dependent desensitization of GIRK currents.