Tonic regulation of excitation-contraction coupling by basal protein kinase C activity in isolated cardiac myocytes.
A high-speed imaging technique was used to investigate the effects of inhibitors and activators of protein kinase C (PKC) on the [Ca2+]i transients and contraction of fura-2 loaded rat ventricular cardiac myocytes. The amplitude of the [Ca2+]i transient was reduced following treatment with 100 nM phorbol 12,13-dibutyrate (PDBu), whereas the PKC inhibitors staurosporine (0.5 microM) and calphostin C (10 microM) increased [Ca2+]i transient amplitude, elevated basal [Ca2+]i and slowed the decay of the [Ca2+]i transient. These changes were paralleled by similar alterations in the rate and extent of cell shortening. The activity of nitrendipine-sensitive Ca2+ channels was monitored indirectly as the rate of Mn2+ quench of cytosolic fura-2 in electrically-paced cells. PDBu reduced Mn2+ influx by six-fold, whereas staurosporine and calphostin C increased the influx rate by eight-fold and seven-fold over basal quench, respectively. The caffeine releasable Ca2+ pool was reduced in the presence of PDBu and increased transiently in presence of staurosporine. The effects of PKC activation and inhibition on sarcoplasmic reticulum Ca2+ content may be secondary to alterations of sarcolemmal Ca2+ influx. However, the PKC inhibitors also decreased the rate of sarcoplasmic reticulum Ca2+ uptake in permeabilized myocytes, suggesting that a direct effect of PKC on the sarcoplasmic reticulum may contribute to the prolongation of the [Ca2+]i transient under these conditions. The present work demonstrates that basal PKC activity has a potent depressant effect, mediated primarily through inhibition of sarcolemmal Ca2+ influx, which may play a key role in setting the basal tone of cardiac muscle.
Nicolás
J M
JM
Department of Anatomy, Pathology and Cell Biology, Thomas Jefferson University, Philadelphia, PA 19107, USA.
Renard-Rooney
D C
DC
Thomas
A P
AP
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D000252:Calcium-Transporting ATPases; D003285:Contractile Proteins / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006321:Heart / Q000502:physiology*; D066298:In Vitro Techniques; D007700:Kinetics; D008274:Magnesium / Q000502:physiology; D008297:Male; D008856:Microscopy, Fluorescence; D009200:Myocardial Contraction; D009206:Myocardium / Q000201:enzymology; D009281:Naphthalenes / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012508:Sarcolemma / Q000502:physiology; D012519:Sarcoplasmic Reticulum / Q000502:physiology; D019311:Staurosporine / Q000494:pharmacology; D013997:Time Factors