MYOCARDIAL PROTEIN KINASE C & B-ADRENOCEPTORS IN CHF

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Protein kinase C, a Ca2+- and phospholipid-dependent protein phosphorylation enzyme present in myocardium, has been shown recently to induce cardiac beta-adrenergic desensitization by phosphorylation of beta-adrenoceptors. To study whether activation of protein kinase C is responsible for cardiac beta-adrenergic desensitization in congestive heart failure, we propose 1) to compare the number and/or activity of cardiac sarcolemmal protein kinase C in heart failure and sham-operated dogs, 2) to characterize the protein kinase C-catalyzed phosphorylation of endogenous substrate proteins in the myocardium of the heart failure and sham-operated dogs, 3) to study the effect of congestive heart failure on protein kinase C-mediated cellular functions in vitro, using isolated myocyte preparations, 4) to determine whether changes in protein kinase C occur only in the pressure-overloaded right ventricle (chamber-specific) or they are also found in the left ventricle and circulating lymphocytes, and 5) to study whether inhibition of protein kinase C in vivo normalizes myocardial beta-receptor density, adenylate cyclase activity and beta-adrenergic responsiveness in heart failure dogs. Congestive right heart failure will be produced by tricuspid avulsion and progressive pulmonary artery constriction in dogs. Hemodynamic measurements will be taken to characterize the degree of heart failure before the animals are sacrificed. Intrinsic contractile function and inotropic responsiveness of the right ventricle to beta-agonists will be studied, using an isolated right ventricular papillary muscle preparation. Samples will be taken from the right and left ventricles for measuring (3H)-phorbol- 12,13-dibutyrate binding, protein kinase C activity, protein kinase C-catalyze phosphorylation of endogenous substrate proteins, beta- receptor density and adenylate cyclase activity. Isolated myocyte preparations will be used to study the effects of protein kinase C as influenced by congestive heart failure on intracellular hydrogen and calcium concentrations, using fluorescent indicators and spectrofluorimetry. The study incorporates physiological, biochemical and cellular measurements in a conscious animal model. This integrative study will not only further our understanding of the fundamental cellular mechanisms responsible for the decreased sensitivity of the heart to sympathetic stimulation in congestive heart failure, but also may provide us a new modality for the treatment of congestive heart failure.