Modulation of the Mitochondrial Cyclosporin A - sensitive Permeability Transition Pore

This paper reports an investigation on the minimal requirements for induction of the mitochondrial cyclosporin A-sensitive permeability transition pore (MTP). Energized rat liver mitochondria incubated in sucrose medium take up a Ca2+ pulse but do not show activation of the MTP upon addition of fully uncoupling concentrations of carbonyl cyanide-p-trifluoromethoxyphenyl hydrazone (FCCP). Yet, addition of ruthenium red immediately prior to FCCP or inclusion of acetate o r low concentrations of phosphate in the incubation medium restore the ability of FCCP to trigger MTP opening. Since ruthenium red, phosphate, or acetate do not induce the MTPper se, they must act upon some additional factor(s) that counteract the increase of por “open” probability induced by membrane depolarization (Bernardi, P. (1992) J. Biol. Chem. 267, 88348839). We show that two key factors determining the overall probability of pore opening are matrix pH and matrix [Ca2+] when the membrane is depolarized. Indeed, in the absence of ruthenium red, phosphate, or acetate the addition of FCCP is followed by fast Ca2+ efflux on the uniporter mirrored by intense matrix acidification following compensatory H+ influx via FCCP. Ruthenium red prevents both Ca2+ efflux and matrix acidification, acetate mainly prevents matrix acidification, and phosphate both slows down Ca2+ efflux and prevents matrix acidification. Taken together, our data indicate that membrane depolarization is both necessary and sufficient to trigger MTP opening when matrix acidification and rapid Ca2+ efflux are prevented. Thus, the wide variety of MTP “inducing agents” may largely reflect the wide variety of the possible combinatorial effects on the minimal requirements for MTP operation.