ATP-sensitive K+ channel activation by nitric oxide and protein kinase G in rabbit ventricular myocytes

The present investigation tested the hypothesis that nitric oxide (NO) potentiates ATP-sensitive K+ (K-ATP) channels by protein kinase G (PKG)-dependent phosphorylation in rabbit ventricular myocytes with the use of patch-clamp techniques. Sodium nitroprusside (SNP; 1 mM) potentiated K-ATP channel activity in cell-attached patches but failed to enhance the channel activity in either inside-out or outside-out patches. The 8-(4- chlorophenylthio)-cGMP Rp isomer (Rp-CPT-cGMP, 100 muM) suppressed the potentiating effect of SNP. 8-( 4-Chlorophenylthio)-cGMP (8-pCPT-cGMP, 100 muM) increased KATP channel activity in cell-attached patches. PKG (5 U/mul) added together with ATP and cGMP (100 muM each) directly to the intracellular surface increased the channel activity. Activation of K-ATP channels was abolished by the replacement of ATP with ATPgammaS. Rp-pCPT-cGMP (100 muM) inhibited the effect of PKG. The heat-inactivated PKG had little effect on the K-ATP channels. Protein phosphatase 2A (PP2A, 1 U/ml) reversed the PKG-mediated K-ATP channel activation. With the use of 5 nM okadaic acid (a PP2A inhibitor), PP2A had no effect on the channel activity. These results suggest that the NO-cGMP-PKG pathway contributes to phosphorylation of K-ATP channels in rabbit ventricular myocytes.