Intracellular Ca(2+) regulates responsiveness of cardiac L-type Ca(2+) current to protein kinase A: role of calmodulin.
The goal of this study was to determine whether the protein kinase A (PKA) responsiveness of the cardiac L-type Ca(2+) current (ICa) is affected during transient increases in intracellular Ca(2+) concentration. Ventricular myocytes were isolated from 3- to 4-day-old neonatal rats and cultured on aligned collagen thin gels. When measured in 1 or 2 mM Ca(2+) external solution, the aligned myocytes displayed a large ICa that was weakly regulated (20% increase) during stimulation of PKA by 2 microM forskolin. In contrast, application of forskolin caused a 100% increase in ICa when the external Ca(2+) concentration was reduced to 0.5 mM or replaced with Ba(2+). This Ca(2+)-dependent inhibition was also observed when the cells were treated with 1 microM isoproterenol, 100 microM 3-isobutyl-1-methylxanthine, or 500 microM 8-bromo-cAMP. The responsiveness of ICa to PKA was restored during intracellular dialysis with a calmodulin (CaM) inhibitory peptide but not during treatment with inhibitors of protein kinase C, Ca(2+)/CaM-dependent protein kinase, or calcineurin. Adenoviral-mediated expression of a CaM molecule with mutations in all four Ca(2+)-binding sites also increased the PKA sensitivity of ICa. Finally, adult mouse ventricular myocytes displayed a greater response to forskolin and cAMP in external Ba(2+). Thus Ca(2+) entering the myocyte through the voltage-gated Ca(2+) channel regulates the PKA responsiveness of ICa.
Walsh
Kenneth B
KB
Department of Pharmacology, Physiology, and Neuroscience, School of Medicine, University of South Carolina, Columbia, SC 29208, USA. walsh@med.sc.edu
Cheng
Qi
Q
D000818:Animals; D000831:Animals, Newborn; D001464:Barium / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000502:physiology*; D002147:Calmodulin / Q000502:physiology*; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology*; D004553:Electric Conductivity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism*; D051379:Mice; D009206:Myocardium / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism; D051381:Rats
D020746:Calcium Channels, L-Type; D002147:Calmodulin; D004791:Enzyme Inhibitors; D005576:Colforsin; D001464:Barium; D017868:Cyclic AMP-Dependent Protein Kinases; D002118:Calcium