Modulation of rat cardiac sodium channel by the stimulatory G protein α subunit

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Abstract
1 Modulation of cardiac sodium currents (INa) by the G protein stimulatory α subunit (Gsα) was studied using patch-clamp techniques on freshly dissociated rat ventricular myocytes. 2 Whole-cell recordings showed that stimulation of β-adrenergic receptors with 10 μM isoprenaline (isoproterenol, ISO) enhanced INa by 68·4 ± 9·6 % (mean ±s.e.m.; n= 7, P < 0·05vs. baseline). With the addition of 22 μg ml−1 protein kinase A inhibitor (PKI) to the pipette solution, 10 μM ISO enhanced INa by 30·5 ± 7·0 % (n= 7, P < 0·05vs. baseline). With the pipette solution containing both PKI and 20 μg ml−1 anti-Gsα IgG or 20 μg ml−1 anti-Gsα IgG alone, 10 μM ISO produced no change in INa. 3 The effect of Gsα on INa was not due to changes in the steady-state activation or inactivation curves, the time course of current decay, the development of inactivation, or the recovery from inactivation. 4 Whole-cell INa was increased by 45·2 ± 5·3 % (n= 13, P < 0·05vs. control) with pipette solution containing 1 μM Gsα27-42 peptide (amino acids 27-42 of rat brain Gsα) without altering the properties of Na+ channel kinetics. Furthermore, application of 1 nM Gsα27-42 to Na+ channels in inside-out macropatches increased the ensemble-averaged INa by 32·5 ± 6·8 % (n= 8, P < 0·05vs. baseline). The increase in INa was reversible upon Gsα27-42 peptide washout. Single channel experiments showed that the Gsα27-42 peptide did not alter the Na+ single channel current amplitude, the mean open time or the mean closed time, but increased the number of functional channels (N) in the patch. 5 Application of selected short amino acid segments (Gsα27-36, Gsα33-42 and Gsα30-39) of the 16 amino acid Gsα peptide (Gsα27-42 peptide) showed that only the C-terminal segment of this peptide (Gsα33-42) significantly increased INa in a dose-dependent fashion. These results show that cardiac INa is regulated by Gsα via a mechanism independent of PKA that results in an increase in the number of functional Na+ channels. In addition, a 10 residue domain (amino acids 33-42) near the N-terminus of Gsα is important in modulating cardiac Na+ channels.