Importance of glycolytically derived ATP for Na+ loading via Na+/H+ exchange during metabolic inhibition in guinea pig ventricular myocytes

The increase in the intracellular Na⁺ concentration ([Na⁺]_i) during myocardial ischaemia is crucial for ischaemia/reperfusion cell injury, and the cardiac subtype of the Na⁺/H⁺ exchanger (NHE-1) has been shown to be a major pathway for Na⁺ loading. While the importance of glycolytically derived ATP for the optimal functioning of membrane transporters and channels has been suggested, whether NHE-1 is actually activated during myocardial ischaemia remains controversial. Here we examined whether the activity of NHE-1 is predominantly dependent on intracellular ATP generated by glycolysis, and whether the additional inhibition of glycolysis can affect the increase in [Na⁺]_i during the inhibition of oxidative phosphorylation in intact guinea pig ventricular myocytes. The selective inhibition of glycolysis by 2-deoxyglucose prevented the recovery of intracellular pH and the transient increase in [Na⁺]_i following intracellular acidosis induced by a NH₄Cl pre-pulse. During severe metabolic inhibition (SMI; induced by amobarbital and carbonyl cyanide m-chlorophenylhydrazone in a glucose-free perfusate), most myocytes changed from rod-shaped to contracted forms by ~ 15 min. [Na⁺]_i increased linearly until rigor contracture occurred, but after rigor contracture the rate of increase was blunted. The increase in [Na⁺]_i during SMI was suppressed significantly by an inhibitor of NHE-1, hexamethylene amiloride. The increase in the intracellular Mg²⁺ concentration, which can reciprocally indicate depletion of intracellular ATP, was small during the initial 10 min of SMI, but became larger from just a few minutes before rigor contracture. In the presence of 2-deoxyglucose, the time to rigor during SMI was shortened, but the increase in [Na⁺]_i before rigor contracture was not significant, and was much less than that in the absence of 2-deoxyglucose. It is concluded that ATP generated by glycolysis is essential to activate NHE-1, and that the dependence of NHE-1 on glycolysis might affect the increase in [Na⁺]_i observed during myocardial ischaemia.