ATP DEPLETION RATHER THAN MITOCHONDRIAL DEPOLARIZATION MEDIATES HEPATOCYTE KILLING AFTER METABOLIC INHIBITION

The importance of ATP depletion and mitochondrial depolarization in th e toxicity of cyanide, oligomycin, and carbonyl cyanide m-choloropheny lhydrazone (CCCP), an uncoupler, was evaluated in rat hepatocytes. Oli gomycin, an inhibitor of the reversible mitochondrial ATP synthase (F1 F0-adenosinetriphosphatase), caused dose-dependent cell killing with 0 .1 mu g/ml being the minimum concentration causing the maximum cell ki lling. Oligomycin also caused rapid ATP depletion without causing mito chondrial depolarization. Fructose (20 mM), a potent glycolytic substr ate in liver, protected completely against oligomycin toxicity. CCCP ( 5 mu M) also caused rapid killing of hepatocytes. Fructose retarded ce ll death caused by CCCP but failed to prevent lethal cell injury. Alth ough oligomycin (1.0 mu g/ml) was lethally toxic by itself, in the pre sence of fructose it protected completely against CCCP induced cell ki lling. Cyanide (2.5 mM), an inhibitor of mitochondrial respiration, ca used rapid cell killing that was reversed by fructose. CCCP completely blocked fructose protection against cyanide, causing mitochondrial de polarization and rapid ATP depletion. In the presence of fructose and cyanide, oligomycin protected cells against CCCP-induced ATP depletion and cell death but did not prevent mitochondrial depolarization. In e very instance, cell killing was associated with ATP depletion, whereas protection against lethal cell injury was associated with preservatio n of ATP. In conclusion, protection by fructose against toxicity of cy anide, oligomycin, and CCCP was mediated by glycolytic ATP formation r ather than by preservation of the mitochondrial membrane potential. Th ese findings support the hypothesis that inhibition of cellular ATP fo rmation is a crucial event in the progression of irreversible cell inj ury.