Carbenoxolone induces permeability transition pore opening in rat mitochondria via the translocator protein TSPO and connexin43

Ca2+-induced permeability transition pore (mPTP) opening in isolated rat brain mitochondria is promoted through targeting of connexin43. After a threshold Ca2+ load, mitochondrial membrane potential drops and efflux of accumulated Ca2+ from the mitochondrial matrix occurs, indicating the mPTP opening. Specific antibodies were used to assess the role of the translocator protein (18 kDa; TSPO) and connexin43 in swelling of isolated rat liver and brain mitochondria induced by carbenoxolone and the endogenous TSPO ligand protoporphyrin IX. Mitochondrial membrane potential, Ca2+ transport and oxygen consumption were determined using selective electrodes. All the parameters were detected simultaneously in a chamber with the selective electrodes. The phosphorylation state of mitochondrial protein targets was assessed. We report that Ca2+-induced mitochondrial swelling was strengthened in the presence of both carbenoxolone and protoporphyrin IX. The carbenoxolone- and protoporphyrin IX-accelerated mPTP induction in brain mitochondria was completely prevented by antibodies specific for the mitochondrial translocator protein (TSPO). The anti-TSPO antibodies were more effective than anti-сonnexin43 antibodies. Moreover, carbenoxolone-stimulated phosphorylation of mitochondrial proteins was inhibited by anti-TSPO antibodies. Taken together, the data suggests that, in addition to acting via connexion43, carbenoxolone may exert its effect on mPTP via mitochondrial outer membrane TSPO.