Abstract
Fatty acids are the preferred substrate of ischemic, reperfused myocardium and may account for the decreased cardiac efficiency during aerobic recovery. Neonatal cardiac myocytes in culture respond to hypoxia/serum- and glucose-free medium by a slow decline in ATP which reverses upon oxygenation. This model was employed to examine whether carnitine palmitoyltransferase I (CPT-I) modulates high rates of β-oxidation following oxygen deprivation. After 5 h of hypoxia, ATP levels decline to 30% control values and CPT- I activity is significantly stimulated in hypoxic myocytes with no alteration in cellular carnitine content or in the release of the mitochondrial matrix marker, citrate synthase. This stimulation was attributed to an increase in the affinity of hypoxic CPT-I for carnitine, suggesting that the liver CPT-I isoform is more dominant following hypoxia. However, there was no alteration in hypoxic CPT-I inhibition by malonyl-CoA. DNP-etomoxiryl-CoA, a specific inhibitor of the liver CPT-I isoform, uncovered identical Michaelis kinetics of the muscle isoform in control and hypoxic myocytes with activation of the liver isoform. Northern blotting did not reveal any change in the relative abundance of mRNA for the liver vs. the muscle CPT-I isoforms. The tyrosine phosphatase inhibitor, pervanadate, reversed the hypoxia-induced activation of CPT-I and returned the affinity of cardiac CPT-I for carnitine to control. Reoxygenation was also associated with a return of CPT-I activity to control levels. The data demonstrate that CPT-I is activated upon ATP depletion. Lower enzyme activities are present in control and reoxygenated cells where ATP is abundant or when phosphatases are inhibited. This is the first suggestion that phosphorylation may modulate the activity of the liver CPT-I isoform in heart.
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Renstrom B, Nellis SH, Liedtke AJ: Metabolic oxidation of glucose during early myocardial reperfusion. Circ Res 65: 1094–1101, 1989
Liedtke AJ, Nellis SH, Mjøs OD: Effects of reducing fatty acid metabolism on mechanical function in regionally ischemic hearts. Am J Physiol 247: H387–H394, 1984
Lopaschuk GD, Spafford MA, Davies NJ, Wall SR: Glucose and palmitate oxidation in isolated working hearts reperfused after a period of transient global ischemia. Circ Res 66: 546–553, 1990
Kudo N, Barr AJ, Barr RL, Desai S, Lopaschuk GD: High rates of fatty acid oxidation during reperfusion of ischemic hearts are associated with a decrease in malonyl-CoA levels due to an increase in 5′-AM-Pactivated protein kinase inhibition of acetyl-CoA carboxylase. J Biol Chem 270: 17513–17520, 1995
Hudson EK, Liu M-H, Buja LM, McMillin JB: Insulin-associated changes in carnitine palmitoyltransferase in cultured neonatal rat cardiac myocytes. J Mol Cell Cardiol 27: 599–613, 1995
Weis BC, Esser V, Foster DW, McGarry JD: Rat heart expresses two forms of mitochondrial carnitine palmitoyltransferase I. J Biol Chem 269: 18712–18715, 1994
Weis BC, Cowan AY, Brown N, Foster DW, McGarry JD: Use of a selective inhibitor of liver carnitine palmitoyltransferase I (CPT-I) allows quantification of its contribution to total CPT-I activity in rat heart. J Biol Chem 269: 35–43, 1994
Brown NF, Weis BC, Husti JE, Foster DW, McGarry JD: Mitochondrial carnitine palmitoyltransferase I isoform switching in the developing rat heart. J Biol Chem 270: 8952–8957, 1995
McMillin JB, Hudson EK, Buja LM: Long chain acyl-CoA metabolism by mitochondrial carnitine palmitoyltransferase: A cell model for pathological studies. Meth Toxicol II: 301–309, 1993
Rogers TB, Gaa ST, Massey C, Dosemeci A: Protein kinase C inhibits Ca2+ accumulation in cardiac sarcoplasmic reticulum. J Biol Chem 265: 4302–4308, 1990
McMillin JB, Wang D, Witters LA, Buja LM: Kinetic properties of carnitine palmitoyltransferase I in cultured neonatal rat cardiac myocytes. Arch Biochem Biophys 312: 375–384, 1994
Teshima R, Ikebuchi H, Nakanishi M, Sawada J: Stimulatory effect of pervanadate on calcium signals and histamine secretion of RBL-2H3 cells. Biochem J 302: 867–874, 1994
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ: Protein measurement with the folin phenol reagent. J Biol Chem 193: 265–275, 1951
Xia Y, Buja LM, McMillin JB: Change in expression of heart carnitine palmitoyltransferase I isoforms with electrical stimulation of cultured rat neonatal cardiac myocytes. J Biol Chem 271: 12082–12087, 1996
Wang D, Buja LM, McMillin JB: Acetyl coenzyme A carboxylase activity in neonatal rat cardiac myocytes in culture: Citrate dependence and effects of hypoxia. Arch Biochem Biophys 325: 249–255, 1996
Henry PD, Shuchleib R, Davis J, Weiss ES, Sobel BE: Myocardial contracture and accumulation of mitochondrial calcium in ischemic rabbit heart. Am J Physiol 233: H677–H684, 1977
Kwast KE, Hand SC: Acute depression of mitochondrial protein synthesis during anoxia. J Biol Chem 271: 7313–7319, 1996
Chrzanowska-Lightowlers ZMA, Preiss T, Lightowlers RN: Inhibition of mitochondrial protein synthesis promotes increased stability of nuclear-encoded respiratory gene transcripts. J Biol Chem 269: 27322–27328, 1995
Harano Y, Kashiwagi A, Kojima H, Suzuki M, Hashimoto T, Shigeta Y: Phosphorylation of carnitine palmitoyltransferase and activation by glucagon in isolated rat hepatocytes. FEBS Lett 188: 267–272, 1985
Zhao Z, Tan Z, Diltz CD, You M, Fischer EH: Activation of mitogen-activated protein (MAP) kinase pathway by pervanadate, a potent inhibitor of tyrosine phosphatases. J Biol Chem 271: 22251–22255, 1996
Esser V, Brown NF, Cowan AT, Foster DW, McGarry JD: Expression of a cDNA isolated from rat brown adipose tissue and heart identifies the product as the muscle isoform of carnitine palmitoyltransferase I (MCPT-I). J Biol Chem 271: 6972–6977, 1996
Bird ML, Saggerson ED: Interacting effects of L-carnitine and malonyl-CoA on rat liver carnitine palmitoyltransferase. Biochem J 230: 161–167, 1985
Paulson DJ, Shug AL: Tissue specific depletion of L-carnitine in rat heart and skeletal muscle by D-carnitine. Life Sci 28: 2931–2938, 1981
Cook GA, Gamble MS: Regulation of carnitine palmitoyltransferase by insulin results in decreased activity and decreased apparent KI values for malonyl-CoA. J Biol Chem 262: 2050–2055, 1987
Guzmán M, Geelan MJH: Short-term regulation of carnitine palmitoyltransferase activity in isolated rat hepatocytes. Biochem Biophys Res Commun 181: 781–787, 1988
Guzmán M, Castro J: Okadaic acid stimulates carnitine palmitoyltransferase I activity and palmitate oxidation in isolated rat hepatocytes. FEBS Lett 291: 105–108, 1991
Cohen P, Holmes CFB, Tsukitani Y: Okadaic acid: A new probe for the study of cellular regulation. Trends Biochem Sci 15: 98–102, 1990
Winz R, Hess D, Aebersold R, Brownsey RW: Unique structural features and differential phosphorylation of the 280-kDDa component (isozyme) of rat liver acetyl-CoA carboxylase. J Biol Chem 269: 14438–14444, 1994
Superti-Furga G: Regulation of the src protein tyrosine kinase. FEBS Lett 369: 62–66, 1995
Morioka M, Fukunaga K, Yasugawa S, Nagahiro S, Ushio Y, Miyamoto E: Regional and temporal alterations in Ca2+/calmodulin-dependent protein kinase II and calcineurin in the hippocampus of rat brain after transient forebrain ischemia. J Neurochem 58: 1798–17809, 1992
Ricciutti MA: Myocardial lysosome stability in the early stages of acute ischemic injury. Am J Cardiol 30: 492–497, 1972
Takano S, Fukuyama H, Fukumoto M, Hirashimizu K, Higuchi T, Takenawa J, Nakayama H, Kimura J, Fujita J: Induction of CL100 protein tyrosine phosphatase following transient forebrain ischemia in the rat brain. J Cereb Blood Flow Metab 15: 33–41, 1995
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Wang, D., Xia, Y., Buja, L.M. et al. The liver isoform of carnitine palmitoyltransferase I is activated in neonatal rat cardiac myocytes by hypoxia. Mol Cell Biochem 180, 163–170 (1998). https://doi.org/10.1023/A:1006815814283
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DOI: https://doi.org/10.1023/A:1006815814283