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 microM 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 microM) 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 [Ca2+]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 microM), 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.
Nakamura
T
T
The Third Department of Internal Medicine, Hamamatsu University School of Medicine, Japan.
Hayashi
H
H
Satoh
H
H
Katoh
H
H
Kaneko
M
M
Terada
H
H
D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D005260:Female; D006168:Guinea Pigs; D006352:Heart Ventricles / Q000187:drug effects / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008954:Models, Biological; D009200:Myocardial Contraction; D015428:Myocardial Reperfusion Injury / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D012433:Ryanodine / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism*
C057360:5-(N,N-hexamethylene)amiloride; D012433:Ryanodine; D000584:Amiloride; D012964:Sodium; D002118:Calcium