Protein kinase G reverses all isoproterenol induced changes of cardiac single L-type calcium channel gating

Objective: cGMP reduces the effect of beta -adrenoceptor agonists on cardia c L-type calcium current by protein kinase G activation. Stimulation of bet a -adrenoceptors increases protein kinase A dependent phosphorylation of L- type calcium channels via cAMP. At the single channel level, protein kinase A dependent phosphorylation increases bath availability and open probabili ty. The present study investigates how cGMP antagonises protein kinase A in duced changes of single L-type calcium channel gating. Methods: Single L-ty pe calcium channels were recorded in the cell attached configuration of the patch clamp technique in isolated mouse ventricular myocytes. Results: The beta -adrenoceptor agonist isoproterenol (10(-6) M) enhanced single channe l peak average current by increasing availability and open probability and decreasing the time constant of long close times. 8-Br-cGMP (10(-3) M) comp letely reversed these effects. The phosphatase inhibitor okadaic acid (10(- 6) M) did not influence the effect of 8-Br-cGMP. The protein kinase G inhib itor Rp-8Br-PET-cGMPS (10(-7) M) abated the effect of 8-Br-cGMP. Activation of protein kinase A by the hydrolysis-resistant cAMP derivative 8-Br-cAMP (10(-3) M) enhanced L-type calcium channel activity like isoproterenol and its effect was also reversed by 8-Br-cGMP. Conclusion: 8-Br cGMP diminishes beta -adrenoceptor activation of L-type calcium channels via protein kinas e G. It interacts with the beta -adrenoceptor signaling pathway distal of a denylyl cyclase. Our observations suggest that protein kinase G interacts e ither with protein kinase A or directly with the L-type calcium channel. (C ) 2000 Elsevier Science B.V. All rights reserved.