Alloxan-induced Mitochondrial Permeability Transition Triggered by Calcium, Thiol Oxidation, and Matrix ATP

Abstract

In addition to their critical function in energy metabolism, mitochondria contain a permeability transition pore, which is regulated by adenine nucleotides. We investigated conditions required for ATP to induce a permeability transition in mammalian mitochondria. Mitochondrial swelling associated with mitochondria permeability transition (MPT) was initiated by adding succinate to a rat liver mitochondrial suspension containing alloxan, a diabetogenic agent. If alloxan was added immediately with or 5 min after adding succinate, MPT was strikingly decreased. MPT induced by alloxan was inhibited by EGTA and several agents causing thiol oxidation, suggesting that alloxan leads to permeability transition through a mechanism dependent on Ca(2+) uptake and sulfhydryl oxidation. Antimycin A and cyanide, inhibitors of electron transfer, carbonyl cyanide m-chlorophenylhydrazone, and oligomycin all inhibited MPT. During incubation with succinate, alloxan depleted ATP in mitochondria after an initial transient increase. However, in a mitochondrial suspension containing EGTA, ATP significantly increased in the presence of alloxan to a level greater than that of the control. These results suggest the involvement of energized transport of Ca(2+) in the MPT initiation. Addition of exogenous ATP, however, did not trigger MPT in the presence of alloxan and had no effect on MPT induced by alloxan. We conclude that alloxan-induced MPT requires mitochondrial energization, oxidation of protein thiols, and matrix ATP to promote energized uptake of Ca(2+).