Protein kinase C and preconditioning: role of the sarcoplasmic reticulum.
Activation of protein kinase C (PKC) is cardioprotective, but the mechanism(s) by which PKC mediates protection is not fully understood. Inasmuch as PKC has been well documented to modulate sarcoplasmic reticulum (SR) Ca2+ and because altered SR Ca2+ handling during ischemia is involved in cardioprotection, we examined the role of PKC-mediated alterations of SR Ca2+ in cardioprotection. Using isolated adult rat ventricular myocytes, we found that addition of 1,2-dioctanoyl-sn-glycerol (DOG), to activate PKC under conditions that reduced myocyte death associated with simulated ischemia and reperfusion, also reduced SR Ca2+. Cell death was 57.9 +/- 2.9% and 47.3 +/- 1.8% in untreated and DOG-treated myocytes, respectively (P < 0.05). Using fura 2 fluorescence to monitor Ca2+ transients and caffeine-releasable SR Ca2+, we examined the effect of DOG on SR Ca2+. Caffeine-releasable SR Ca2+ was significantly reduced (by approximately 65%) after 10 min of DOG treatment compared with untreated myocytes (P < 0.05). From our examination of the mechanism by which PKC alters SR Ca2+, we present the novel finding that DOG treatment reduced the phosphorylation of phospholamban (PLB) at Ser16. This effect is mediated by PKC-epsilon, because a PKC-epsilon-selective inhibitory peptide blocked the DOG-mediated decrease in phosphorylation of PLB and abolished the DOG-induced reduction in caffeine-releasable SR Ca2+. Using immunoprecipitation, we further demonstrated that DOG increased the association between protein phosphatase 1 and PLB. These data suggest that activated PKC-epsilon reduces SR Ca2+ content through PLB dephosphorylation and that reduced SR Ca2+ may be important in cardioprotection.
Yamamura
Ken
K
Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, National Institutes of Health, 111 T. W. Alexander Dr., Bldg. 101, MD F2-07, Research Triangle Park, NC 27709, USA.
Steenbergen
Charles
C
Murphy
Elizabeth
E
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015687:Cell Hypoxia / Q000502:physiology; D007511:Ischemia / Q000378:metabolism*; D019157:Ischemic Preconditioning, Myocardial / Q000379:methods*; D008297:Male; D032383:Myocytes, Cardiac / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D054645:Protein Phosphatase 1; D051381:Rats; D017207:Rats, Sprague-Dawley; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D002118:Calcium