title: The toxicity of N-methyl-alpha-methyldopamine to freshly isolated rat hepatocytes is prevented by ascorbic acid and N-acetylcysteine.
abstract: In the past decade, clinical evidence has increasingly shown that the liver is a target organ for 3,4-methylenedioxymethamphetamine (MDMA, "ecstasy") toxicity. The aims of the present in vitro study were: (1) to evaluate and compare the hepatotoxic effects of MDMA and one of its main metabolites, N-methyl-alpha-methyldopamine (N-Me-alpha-MeDA) and (2) to investigate the ability of antioxidants, namely ascorbic acid and N-acetyl-L-cysteine (NAC), to prevent N-Me-alpha-MeDA-induced toxic injury, using freshly isolated rat hepatocytes. Cell suspensions were incubated with MDMA or N-Me-alpha-MeDA in the final concentrations of 0.1, 0.2, 0.4, 0.8, and 1.6 mM for 3 h. To evaluate the potential protective effects of antioxidants, cells were preincubated with ascorbic acid in the final concentrations of 0.1 and 0.5 mM, or NAC in the final concentrations of 0.1 and 1 mM for 15 min before treatment with 1.6 mM N-Me-alpha-MeDA for 3 h (throughout this incubation period the cells were exposed to both compounds). The toxic effects were evaluated by measuring the cell viability, glutathione (GSH) and glutathione disulfide (GSSG), ATP, and the cellular activities of GSH peroxidase (GPX), GSSG reductase (GR), and GSH S-transferase (GST). MDMA induced a concentration- and time-dependent GSH depletion, but had a negligible effect on cell viability, ATP levels, or on the activities of GR, GPX, and GST. In contrast, N-Me-alpha-MeDA was shown to induce not only a concentration- and time-dependent depletion of GSH, but also a depletion of ATP levels accompanied by a loss in cell viability, and decreases in the antioxidant enzyme activities. For both compounds, GSH depletion was not accompanied by increases in GSSG levels, which seems to indicate GSH depletion by adduct formation. Importantly, the presence of ascorbic acid (0.5 mM) or NAC (1 mM) prevented cell death and GSH depletion induced by N-Me-alpha-MeDA. The results provide evidence that MDMA and its metabolite N-Me-alpha-MeDA induce toxicity to freshly isolated rat hepatocytes. Oxidative stress may play a major role in N-Me-alpha-MeDA-induced hepatic toxicity since antioxidant defense systems are impaired and administration of antioxidants prevented N-Me-alpha-MeDA toxicity.
authors.affiliation: REQUIMTE, Serviço de Toxicologia, Faculdade de Farmácia, Universidade do Porto, Rua Aníbal Cunha 164, 4099/030, Portugal. marciacarvalho@ff.up.pt
authors.forename: Márcia
authors.initials: M
authors.lastname: Carvalho
authors.forename: Fernando
authors.initials: F
authors.lastname: Remião
authors.forename: Nuno
authors.initials: N
authors.lastname: Milhazes
authors.forename: Fernanda
authors.initials: F
authors.lastname: Borges
authors.forename: Eduarda
authors.initials: E
authors.lastname: Fernandes
authors.forename: Félix
authors.initials: F
authors.lastname: Carvalho
authors.forename: Maria Lourdes
authors.initials: ML
authors.lastname: Bastos
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001205:Ascorbic Acid / Q000494:pharmacology*; D002469:Cell Separation; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D003846:Deoxyepinephrine / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors* / Q000633:toxicity*; D016166:Free Radical Scavengers / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism; D005980:Glutathione Reductase / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006213:Hallucinogens / Q000037:antagonists & inhibitors / Q000633:toxicity; D022781:Hepatocytes / Q000187:drug effects* / Q000201:enzymology; D066298:In Vitro Techniques; D008297:Male; D018817:N-Methyl-3,4-methylenedioxyamphetamine / Q000037:antagonists & inhibitors / Q000633:toxicity; D051381:Rats; D013056:Spectrophotometry, Ultraviolet
chemical_list: D000975:Antioxidants; D016166:Free Radical Scavengers; D006213:Hallucinogens; C023492:alpha-methyldopamine; D000255:Adenosine Triphosphate; D005979:Glutathione Peroxidase; D005980:Glutathione Reductase; D005982:Glutathione Transferase; D005978:Glutathione; D018817:N-Methyl-3,4-methylenedioxyamphetamine; D001205:Ascorbic Acid; D003846:Deoxyepinephrine; D000111:Acetylcysteine