PKC regulation of cardiac CFTR Cl- channel function in guinea pig ventricular myocytes.
The role of protein kinase C (PKC) in regulating the protein kinase A (PKA)-activated Cl- current conducted by the cardiac isoform of the cystic fibrosis transmembrane conductance regulator (cCFTR) was studied in guinea pig ventricular myocytes using the whole cell patch-clamp technique. Although stimulation of endogenous PKC with phorbol 12,13-dibutyrate (PDBu) alone did not activate this Cl- current, even when intracellular dialysis was limited with the perforated patch-clamp technique, activation of PKC did elicit a significant response in the presence of PKA-dependent activation of the current by the beta-adrenergic receptor agonist isoproterenol. PDBu increased the magnitude of the Cl- conductance activated by a supramaximally stimulating concentration of isoproterenol by 21 +/- 3.3% (n = 9) when added after isoproterenol and by 36 +/- 16% (n = 14) when introduced before isoproterenol. 4alpha-Phorbol 12, 13-didecanoate, a phorbol ester that does not activate PKC, did not mimic these effects. Preexposure to chelerythrine or bisindolylmaleimide, two highly selective inhibitors of PKC, significantly reduced the magnitude of the isoproterenol-activated Cl- current by 79 +/- 7.7% (n = 11) and 52 +/- 10% (n = 8), respectively. Our results suggest that although acute activation of endogenous PKC alone does not significantly regulate cCFTR Cl- channel activity in native myocytes, it does potentiate PKA-dependent responses, perhaps most dramatically demonstrated by basal PKC activity, which may play a pivotal role in modulating the function of these channels.
Middleton
L M
LM
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106-4970, USA.
Harvey
R D
RD
D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000187:drug effects / Q000502:physiology*; D004553:Electric Conductivity; D004789:Enzyme Activation; D005260:Female; D006168:Guinea Pigs; D006321:Heart / Q000502:physiology*; D006352:Heart Ventricles; D007545:Isoproterenol / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D018408:Patch-Clamp Techniques; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010703:Phorbol Esters / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*
D000318:Adrenergic beta-Agonists; D010703:Phorbol Esters; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; C008876:phorbol-12,13-didecanoate; D015240:Phorbol 12,13-Dibutyrate; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D007545:Isoproterenol