GPCR-dependent biasing of GIRK channel signaling dynamics by RGS6 in mouse sinoatrial nodal cells

How G protein -coupled receptors (GPCRs) evoke specific biological outcomes while utilizing a limited array of G proteins and effectors is poorly understood, particularly in native cell systems. Here, we examined signaling evoked by muscarinic (M2R) and adenosine (AIR) receptor activation in the mouse sinoatrial node (SAN), the cardiac pacemaker. M2R and AIR activate a shared pool of cardiac G protein -gated inwardly rectifying K+ (GIRK) channels in SAN cells from adult mice, but A(1)R-GIRK responses are smaller and slower than M2R-GIRK responses. Recordings from mice lacking Regulator of G protein Signaling 6 (RGS6) revealed that RGS6 exerts a GPCR-dependent influence on GIRK-dependent signaling in SAN cells, suppressing M2R-GIRK coupling efficiency and kinetics and A(1)R-GIRK signaling amplitude. Fast kinetic bioluminescence resonance energy transfer assays in transfected HEK cells showed that RGS6 prefers Ga. over Ga, as a substrate for its catalytic activity and that M2R signals preferentially via Ga,, while A(1)R does not discriminate between inhibitory G protein isoforms. The impact of atrial/SAN-selective ablation of Ga. or Gait was consistent with these findings. Gait ablation had minimal impact on M2R-GIRK and A(1)R-GIRK signaling in SAN cells. In contrast, Ga. ablation decreased the amplitude and slowed the kinetics of M2R-GIRK responses, while enhancing the sensitivity and prolonging the deactivation rate of A(1)R-GIRK signaling. Collectively, our data show that differences in GPCR-G protein coupling preferences, and the Ga. substrate preference of RGS6, shape A(1)R- and M2R-GIRK signaling dynamics in mouse SAN cells.