CA2+ DEPLETION PREVENTS ANOXIC DEATH OF HEPATOCYTES BY INHIBITING MITOCHONDRIAL PERMEABILITY TRANSITION

Removal of Ca2+ from the culture medium or treatment with the intracel lular Ca2+ chelator ,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA-AM) prevented the killing of rat hepa tocytes by anoxia and rotenone, but not by cyanide. Neither manipulati on prevented the loss of the mitochondrial membrane potential or the d epletion of ATP. A mitochondrial permeability transition (MPT) was dem onstrated in digitonin-permeabilized hepatocytes as an increased [H-3] sucrose-accessible space sensitive to cyclosporin A (CyA). Ca2+ deplet ion by either means prevented the MPT measured in intact cells made an oxic or treated with rotenone. In isolated mitochondria deenergized by rotenone, BAPTA-AM prevented the MPT induced by palmitoyl CoA. By con trast, in isolated mitochondria deenergized by cyanide, BAPTA-AM alone did not prevent the MPT. Rather, BAPTA-AM plus CyA were required. Sim ilarly, the killing of cultured hepatocytes by cyanide was prevented b y BAPTA-AM plus CyA, but not by either agent alone. The MPT in intact cells treated with cyanide was also prevented by BAPTA-AM plus CyA The se data define a specific requirement for Ca2+ in the killing of hepat ocytes that follows the inhibition of electron transport. A model is p resented in which the MPT depends on factors that modulate the sensiti vity of the permeability transition to the matrix concentration of Ca2 +.