Liver function was examined in group-housed mice given saline or 5, 10 or 20 mg/kg MDMA i.p and maintained either at 20 C or 30 C, with body temperature measured via implanted chip rather than rectal probe. Biochemical analysis was used to measure liver function, and histological analyses were performed on liver tissue removed from animals killed 24 h post-drug. Liver functions analyzed included assessment of hepatic glutathione (GSH) via HPLC, amount of transaminases and alkaline phosphatase, extent of lipid peroxidation, function of the antioxidant enzymes catalase, selenium-dependent glutathione peroxidase (GPx), copper/zinc superoxide dismutase (CuZnSOD), and manganese superoxide dismutase (MnSOD). Light microscopy and staining was used to detect blood clots and hepatocyte vacuolization in tissue. As was expected, MDMA at all doses increased body temperature, with elevated body temperature lasting for 10 h post-drug. High ambient temperature (30 vs 20 C) increased mortality from 10% to 40% (all fatalities replaced to ensure that all conditions contained 5 mice per group). GSH depletion occurred in MDMA-treated mice at normal and high ambient temperatures. Lipid peroxidation increased only in mice given 20 mg/kg MDMA and housed in 30 C environment. Mice given 10 or 20 mg/kg MDMA and kept at 30 C showed decreased catalase activity, but GPx activity decreased in saline-treated mice kept at 30 C. All other liver functions remained similar across treatments (both MDMA and temperature). MDMA dose-dependently produced hepatocyte vacuolization and blood clots radiating from the central vein, with vacuolation appearing at 5 mg/kg, and clotting appearing at 10 mg/kg, and high (versus low) ambient temperature aggravated MDMA-associated apparent liver damage. Both MDMA and high ambient temperature appear to affect liver function, independently and in an additive manner, with the function of some antioxidant enzymes more strongly affected than others. Study findings suggest a role for high ambient temperature in increasing the likelihood of MDMA-associated hepatotoxicity in mammals. However, since there is some evidence that different mechanisms are responsible for some toxicological effects of MDMA in mice versus rats, it is possible that hepatic effects in rats may differ from the reported findings.