Protein kinase C inhibits Ca2+ accumulation in cardiac sarcoplasmic reticulum.
It is now recognized that phorbol esters are negative inotropic agents in mammalian heart which presumably act via stimulation of Ca2(+)-activated phospholipid-dependent protein kinase (PKC). The goal in the present study was to identify the underlying cellular processes. Digitonin-permeabilized cultured neonatal rat ventricular myocytes were used to study biochemical and functional effects of phorbol esters on cardiac sarcoplasmic reticulum (SR). These cells contracted spontaneously at 3 microM Ca2+. Beating was inhibited by 10 microM ryanodine and was insensitive to 1 microM nifedipine. Thus, beating behavior results from the phasic oscillation of Ca2+ transport by SR in this preparation. Phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), decreased frequency by 30%, suggesting that Ca2+ transport by SR had been reduced. Whereas cAMP stimulated the rate of oxalate-supported 45Ca2+ uptake 2-fold, phorbol esters, TPA, and phorbol 12,13-dibutyrate inhibited this process by about 45%. The effects of phorbols were specific: (a) the alpha-analogues of TPA and phorbol 12,13-dibutyrate were inactive; and (b) the phorbol esters had no effect on Ca2+ transport in cells that had been depleted of PKC. TPA decreased oxalate-stimulated Ca2+ uptake over the entire range of Ca2+ concentrations, from 0.1 to 10 microM, by at least 70% without shifting the half-maximal effective Ca2+ concentration. Taken together these results indicate that the effects of phorbol ester on cardiac contraction are due to decreased Ca2+ transport by the SR and that these responses are mediated by PKC. These studies support the interpretation that the negative inotropic effects of phorbol esters are due, in part, to decreased SR function.
Rogers
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TB
Department of Biological Chemistry, University of Maryland School of Medicine, Baltimore 21201.
Gaa
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Massey
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C
Dösemeci
A
A
D000818:Animals; D000831:Animals, Newborn; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002132:Calcium Radioisotopes; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000494:pharmacology; D009200:Myocardial Contraction / Q000187:drug effects; D009206:Myocardium / Q000378:metabolism*; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D012433:Ryanodine / Q000494:pharmacology; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002132:Calcium Radioisotopes; D012433:Ryanodine; D015240:Phorbol 12,13-Dibutyrate; D000242:Cyclic AMP; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium