CAMP AND G-PROTEIN REGULATION OF CARDIAC SODIUM CHANNELS

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DESCRIPTION (adapted from the applicant's abstract): This proposal is an extension of observations to pursue the basic mechanisms by which cAMP- dependent phosphorylation and the direct action of Gsa modulate INa in rabbit cardiac myocytes. First, the mechanisms by which cAMP- dependent phosphorylation modulates INa will be determined. Studying INa in cell- attached and inside-out membrane patches, the effects of forskolin, cell permeable cAMP analogs, and the catalytic subunit of PKA on the number of active channels (N), the probability of channel opening (Po), and the unitary current amplitude (i) of the sodium channel will be determined. The first latency and channel reopening, m(t), distributions and the analysis of runs will also be performed. The effect of protein phosphatases on cardiac INa will also be studied. Second, the mechanisms by which the direct effects of Gsa on INa will be determined. The cAMP-dependent pathways will be investigated using the PKA inhibitor peptide (PKI) and the voltage-dependence of the Gsa effects on INa will be studied. The effect of Gsa on INa activation and inactivation on N, Po, and i of single sodium channels will also be examined. The applicant also plans to delineate the INa-activating domain(s) of Gsa by studying the effects of synthetic peptides with sequences homologous to various segments of native Gsa. Finally, the applicant proposes that there are important interactions between the direct Gsa and the cAMP-mediated pathways in the modulation of INa. He will study the effects of sodium channel phosphorylation and dephosphorylation on the modulation of INa by the direct effects of Gsa.