A single cell model of myocardial reperfusion injury:: Changes in intracellular Na+ and Ca2+ concentrations in guinea pig ventricular myocytes

To investigate the contribution of the changes in intracellular Na+ and Ca2+ concentrations ([Na+](i) and [Ca2+](i)) to myocardial reperfusion injury, we made an ischemia/reperfusion model in intact guinea pig myocytes. Myocardial ischemia was simulated by the perfusion of metabolic inhibitors (3.3 mM amobarbital and 5 mu M carbonyl cyanide m-chlorophenylhydrazone) with pH 6.6 and reperfusion was achieved by the washout of them with pH 7.4. [Na+](i) increased from 7.9 +/- 2.0 to 14.0 +/- 3.4 mM (means +/- S.E., p < 0.01) during 7.5 min of simulated ischemia (SI) and increased further to 18.8 +/- 3.0 mM at 7.5 min after reperfusion. [Ca2+](i), expressed as the ratio of fluo 3 fluorescence intensity, increased to 133 +/- 8% (p < 0.01) during SI and gradually returned to the control level after reperfusion. Intracellular pH decreased from 7.53 +/- 0.04 to 6.31 +/- 0.04 (p < 0.01) and recovered quickly after reperfusion. Reperfusion with the acidic solution or the continuous perfusion of hexamethylene amiloride (2 mu M) prevented the reperfusion-induced increase in [Na+](i). When the duration of SI was prolonged to 15 min, the cell response after reperfusion varied, 16 of 37 cells kept quiescent, 21 cells showed spontaneous Ca2+ waves, and 4 cells out of these 21 cells became hypercontracted. In quiescent cells, both [Na+](i) and [Ca2+](i) decreased immediately after reperfusion. In cells with Ca2+ waves, [Na+](i) transiently increased further at the early phase of reperfusion, while [Ca+](i) declined. In hypercontracted cells, [Na+](i) increased as much as in 'Ca2+ wave' cells, but [Ca2+](i) increased extensively and both ion concentrations continued to increase. Reperfusion with the Ca2+-free solution prevented both the [Ca2+](i) increase and morphological change. In the presence of ryanodine (10 mu M), the increase in [Ca2+](i) after reperfusion was augmented and some cells became hypercontracted. We concluded that (1) Na+/H+ exchange is active both during SI and reperfusion, resulting in the additional [Na+](i) elevation on reperfusion, (2) the [Na+](i) level after reperfusion and the following Ca2+ influx via Na+/Ca2+ exchange are crucial for reperfusion cell injury, and (3) the Ca2+ buffering capacity of sarcoplasmic reticulum would also contribute to the Ca2+ regulation and cell injury after reperfusion.