MITOCHONDRIAL PERMEABILITY TRANSITION IN PH-DEPENDENT REPERFUSION INJURY TO RAT HEPATOCYTES

To simulate ischemia and reperfusion, cultured rat hepatocytes were in cubated in anoxic buffer at pH 6.2 for 4 h and reoxygenated at pH 7.4. During anoxia, intracellular pH (pH(i)) decreased to 6.3, mitochondri a depolarized, and ATP decreased to <1% of basal values, but the mitoc hondrial permeability transition (MPT) did not occur as assessed by co nfocal microscopy from the redistribution of cytosolic calcein into mi tochondria. Moreover, cell viability remained >90%. After reperfusion at pH 7.4, pH(i) returned to pH 7.2, the MPT occurred, and most hepato cytes lost viability. In contrast, after reperfusion at pH 6.2 or with Na+-free buffer at pH 7.4, pH(i) did not rise and cell viability rema ined >80%. After acidotic reperfusion, the MPT did not occur. When hep atocytes were reperfused with cyclosporin A (0.5-1 mu M) at pH 7.4, th e MPT was prevented and cell viability remained >80%, although pH(i) i ncreased to 7.2. Reperfusion with glycine (5 mM) also prevented cell k illing but did not block recovery of pH(i) or the MPT. Retention of ce ll viability was associated with recovery of 30-40% of ATP. In conclus ion, preventing the rise of pH(i) after reperfusion blocked the MPT, i mproved ATP recovery, and prevented cell death. Cyclosporin A also pre vented cell killing by blocking the MPT without blocking recovery of p H(i). Glycine prevented cell killing but did not inhibit recovery of p H(i) or the MPT.