Regulation of Mitochondrial Permeability Transition by ADP

Abstract

Mitochondrial ADP serves at least two important functions: provides a substrate for ATP generation and inhibits the mitochondrial permeability transition pore (mPTP). The mechanism for the inhibitory effect of ADP on mPTP remains unclear. Moreover, the contribution of this ADP-dependent regulation of mPTP in protection from mitochondrial injury has yet to be elucidated. We investigated the inhibitory effects of ADP on mPTP in mitochondria isolated from mouse hearts by measuring mitochondrial Ca2+ uptake capacity. Mitochondria isolated from control mice were able to sequester Ca2+ from up to 10 pulses of 40 μM Ca2+ (389 ± 45 μM, n=7) before the occurrence of a sudden and irreversible rise of Ca2+ in the incubating solution, indicative of mPTP opening. The presence of 500 μM ADP shifted the Ca2+ uptake capacity to 867 ± 89 μM (n=6), similar to that of cyclosporine A treatment (900 ± 139 μM, n=4) or cyclophilin D knockout mice (1300 ± 212 μM, n=4). Interestingly, ADP further increased the Ca2+ uptake capacity in cyclophilin D knockout mice (2075 ± 284 μM, n=4). 1 mM ATP and 10 mM creatine mimicked the effects of ADP (800 ± 124 μM, n=4). Also, oligomycin augmented ADP efficacy. Addition of 10 μM of atractyloside abolished the ADP-mediated increase in Ca2+ uptake capacity. The effect of Ca2+ on mPTP opening may be related to the production of ROS. The increase in ROS caused by Ca2+ addition is reversed by ADP, cyclosporine A, and in cyclophilin D knockout mice. In summary, we show that ADP is a powerful inhibitor of mPTP opening and that its regulatory mechanism could be different from that of cyclophilin D.Supported by HL033333, HL093671 and in part by ETF 8041 and ESF DoRa program activity 6.