Mitochondrial injury by disulfiram: two different mechanisms of the mitochondrial permeability transition

Disulfiram (Ds), a clinically employed alcohol deterrent of the thiuram, di sulfide (TD) class of compounds, is known to cause hepatitis and neuropathi es. Although this drug has been shown to inhibit different thiol-containing enzymes, the actual mechanism of Ds toxicity is not clear. We have previou sly demonstrated that Ds impairs the permeability of inner mitochondrial me mbrane (IMM) [Arch. Biochem. Biophys. 356 (1998) 46]. In this report, the e ffect of Ds and its structural analogue thiram (Th) on mitochondrial functi ons was studied in detail. We found that mitochondria metabolize TDs in a N AD(P)H- and GSH-dependent manner. At the concentration above characteristic threshold, TDs induced irreversible oxidation of NAD(P)H and glutathione ( GSH) pools, collapse of transmembrane potential, and inhibition of oxidativ e phosphorylation. The presence of Ca2+ and exhaustion of mitochondrial glu tathione (GSH+GSSG) decreased the threshold concentration of TDs. Swelling of the mitochondria and leakage of non-transported fluorescent dye BCECF fr om the matrix indicated that TDs induced the mitochondrial permeability tra nsition (MPT). Mitochondrial permeabilization by TDs involves two, apparent ly distinct mechanisms. In the presence of Ca2+, TDs produced cylosporin A- sensitive swelling of mitochondria, which was inhibited by ADP and accelera ted by carboxyatractyloside (CATR) and phosphate. In contrast, the swelling produced by TDs in the absence of Ca2+ was not sensitive to cyclosporin A (CsA), ADP and CATR but was inhibited by phosphate. Titration with N-ethylm aleimide revealed that these two mechanisms involve different SH-groups and probably different transport proteins on the IMM. Our findings indicate th at at pharmacologically relevant concentrations TDs may cause. In irreversi ble mitochondrial injury as a result of induction of the MPT. (C) 2001 Else vier Science Ireland Ltd. All rights reserved.