INSULIN-ASSOCIATED CHANGES IN CARNITINE PALMITOYLTRANSFERASE IN CULTURED NEONATAL RAT CARDIAC MYOCYTES

Insulin increases the synthesis of mitochondrial proteins in the isolated perfused heart and total cell protein synthesis in neonatal cardiac myocytes. Since carnitine-dependent fatty acid oxidation is modulated by insulin in a variety of tissues, the effects of 1.7 mu M insulin on the mitochondrial enzyme(s), carnitine palmitoyltransferase (malonyl-CoA-senstive CPT-I and the matrix-facing CPT-II), were studied in neonatal rat cardiac myocytes cultured in the absence of serum. Following incubation in serum-free medium, there is a four-fold increase in the I-50 of CPT-I for malonyl-CoA (3.8 mu M) compared to cells cultured in serum-free medium to which insulin has been added (I-50 = 0.8 mu M). CPT I activity in the insulin-supplemented, serum-free cultures is 57% higher (P < 0.002) than CPT-I activity in cells cultured in the absence of insulin; CPT-II activity is also significantly increased (P < 0.01) in the presence of insulin, Since CPT-II is an inner membrane protein, the CPT response to insulin may be coordinately regulated with other mitochondrial activities, Similar to CPT, cytochrome oxidase activity of cardiac myocytes in serum-free medium is increased 33% by insulin. Consistent with this finding, both CPT-II and cytochrome oxidase mRNA expression is elevated over control in the presence of insulin. CPT-II activity increases significantly only at very high insulin concentrations (1.7 mu M), suggesting a role for insulin-like growth factor pathway. When myocytes are cultured in the presence of 1.7 mu M insulin and then transferred to an insulin-free medium, subsequent addition of insulin does not stimulate uptake of deoxyglucose. These results suggest that the response of CPT to insulin is mediated by insulin-like growth factor activity and not by cellular glucose availability. The response of CPT to insulin does not appear to be mediated by the protein kinase C pathway since CPT-IT activity is not reduced by the protein kinase C inhibitor, chelerythrine. Insulin significantly increases protein synthesis in the neonatal cardiac myocyte and in isolated mitochondria by increasing incorporation of labelled amino acid into total myocyte and/or mitochondrial protein. The degradation rate of radiolabelled protein in cardiac myocytes cultured in the presence of insulin is not different from that of insulin-deprived cells. The data suggest that insulin can affect the activity and expression of mitochondrial proteins, e.g., CPT, through the insulin-like growth factor-I pathway in neonatal cardiac myocytes.