Cell Signal 2014 Jun 01;266:1182-1192. doi: 10.1016/j.cellsig.2014.02.007.

Differential effects of genetically-encoded Gβγ scavengers on receptor-activated and basal Kir3.1/Kir3.4 channel current in rat atrial myocytes.

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Opening of G-protein-activated inward-rectifying K(+) (GIRK, Kir3) channels is regulated by interaction with βγ-subunits of Pertussis-toxin-sensitive G proteins upon activation of appropriate GPCRs. In atrial and neuronal cells agonist-independent activity (Ibasal) contributes to the background K(+) conductance, important for stabilizing resting potential. Data obtained from the Kir3 signaling pathway reconstituted in Xenopus oocytes suggest that Ibasal requires free Gβγ. In cells with intrinsic expression of Kir3 channels this issue has been scarcely addressed experimentally. Two Gβγ-binding proteins (myristoylated phosducin - mPhos - and Gαi1) were expressed in atrial myocytes using adenoviral gene transfer, to interrupt Gβγ-signaling. Agonist-induced and basal currents were recorded using whole cell voltage-clamp. Expression of mPhos and Gαi1 reduced activation of Kir3 current via muscarinic M2 receptors (IK(ACh)). Inhibition of IK(ACh) by mPhos consisted of an irreversible component and an agonist-dependent reversible component. Reduction in density of IK(ACh) by overexpressed Gαi1, in contrast to mPhos, was paralleled by substantial slowing of activation, suggesting a reduction in density of functional M2 receptors, rather than Gβγ-scavenging as underlying mechanism. In line with this notion, current density and activation kinetics were rescued by fusing the αi1-subunit to an Adenosine A1 receptor. Neither mPhos nor Gαi1 had a significant effect on Ibasal, defined by the inhibitory peptide tertiapin-Q. These data demonstrate that basal Kir3 current in a native environment is unrelated to G-protein signaling or agonist-independent free Gβγ. Moreover, our results illustrate the importance of physiological expression levels of the signaling components in shaping key parameters of the response to an agonist.

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Species referenced: Xenopus
Genes referenced: kcnj3 kcnj5