Copyright © 2005, European Society of Cardiology
Role of mitochondrial re-energization and Ca2+ influx in reperfusion injury of metabolically inhibited cardiac myocytes
Department of Cell Physiology and Pharmacology, University of Leicester, PO Box 138, Leicester LE1 9HN, UK
* Corresponding author. Current address: Department of Cardiovascular Sciences, University of Leicester, Clinical Sciences Wing, Glenfield General Hospital, Leicester LE3 9QP, UK. Tel.: +44 116 256 3028; fax: +44 116 287 5792. Email address: gcr4{at}le.ac.uk
Objective: We used isolated myocytes to investigate the role of mitochondrial re-energization and Ca2+ influx during reperfusion on hypercontracture, loss of Ca2+ homeostasis and contractile function.
Methods: Isolated adult rat ventricular myocytes were exposed to metabolic inhibition (NaCN and iodoacetate) and reperfusion injury was assessed from hypercontracture, loss of Ca2+ homeostasis ([Ca2+]i measured with fura-2) and failure of contraction in response to electrical stimulation. Mitochondrial membrane potential was followed using the potentiometric dye tetramethylrhodamine ethyl ester.
Results: Metabolic inhibition led to contractile failure and rigor accompanied by a sustained increase in [Ca2+]i. Reperfusion after 10 min metabolic inhibition led to an abrupt repolarization of the mitochondrial membrane potential (after 25.5 ± 1.2 s), a transient fall in [Ca2+]i followed by an abrupt hypercontracture (37.1 ± 1.8 s) in 84% of myocytes. Ca2+ homeostasis (diastolic [Ca2+]i<250 nM) recovered in only 23.3 ± 5.1% of cells and contractions recovered in 15.3 ± 2.2%. Oligomycin abolished the hypercontracture on reperfusion, but mitochondrial repolarization was unaffected. Preventing Ca2+ influx during reperfusion with Ca2+-free Tyrode or with an inhibitor of Na+/Ca2+ exchange did not prevent the hypercontracture, but increased the percentage of cells recovering Ca2+ homeostasis and contractile function. The presence of 0.5 µM cyclosporin A did not prevent hypercontracture but increased the percentage of cells recovering Ca2+ homeostasis to 56.2 ± 3.6% and contractile function to 52 ± 4.3%.
Conclusions: Reperfusion-induced hypercontracture, and loss of Ca2+ homeostasis and contractile function are initiated following mitochondrial re-energization. The hypercontracture requires the production of oxidative ATP but not Ca2+ influx during reperfusion. Loss of Ca2+ homeostasis and contractile function are linked to Ca2+ influx during reperfusion, probably via opening of mitochondrial permeability transition pores.
KEYWORDS Reperfusion; Myocytes; Mitochondria; Na+/Ca2+ exchanger; Calcium; Cardiac muscle; Ischaemia
Time for primary review 22 days
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