title: Combination effects of amphetamines under hyperthermia - the role played by oxidative stress.
abstract: Rise in body temperature is a life-threatening consequence of 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) abuse. We evaluated the impact of hyperthermia on the cytotoxicity of combinations of MDMA and three other amphetamines, often co-ingested. For this, Hep G2 cells were exposed to MDMA, d-amphetamine, methamphetamine and 4-methylthioamphetamine, individually or combined, at 40.5 °C. The results were compared with normothermia data (37.0 °C). Mixture additivity expectations were calculated by independent action and concentration addition (CA) models. To delineate the mechanism(s) underlying the elicited effects, a range of stress endpoints was evaluated, including quantification of reactive oxygen/nitrogen species (ROS/RNS), lipid peroxidation, reduced/oxidized glutathione (GSH/GSSG), ATP and mitochondrial membrane potential (Δψm) changes. Our data show that, in hyperthermia, amphetamines acted additively and mixture effects were accurately predicted by CA. At 40.5 °C, even slight increases in the concentrations of each drug/mixture promoted significant rises in cytotoxicity, which quickly shifted from roughly undetectable to maximal mortality. Additionally, the increase of RNS/ROS production, decrease of GSH, ATP depletion and mitochondrial impairment were exacerbated under hyperthermia. Importantly, when equieffective cytotoxic concentrations of the mixture and individual amphetamines were compared for all tested stress endpoints, mixture effects did not deviate from those elicited by individual treatments, suggesting that these amphetamines have a similar mode of action, which is not altered in combination. Concluding, our data indicate that amphetamine mixtures produce deleterious effects, even when individual drugs are combined at negligible concentrations. These effects are strongly exacerbated in hyperthermia, emphasizing the potential increased risks of ecstasy intake, especially when hyperthermia occurs concurrently with polydrug abuse.
authors.affiliation: Faculdade de Medicina, Universidade do Porto, 4200-319, Porto, Portugal; Institute for the Environment, Brunel University, Uxbridge, Middlesex, UB8 3PH, UK; REQUIMTE (Rede de Química e Tecnologia), Laboratório de Toxicologia, Departamento de Ciências Biológicas, Faculdade de Farmácia, Universidade do Porto, 4050-313, Porto, Portugal.
authors.forename: Diana Dias
authors.initials: DD
authors.lastname: da Silva
authors.forename: Elisabete
authors.initials: E
authors.lastname: Silva
authors.forename: Helena
authors.initials: H
authors.lastname: Carmo
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000662:Amphetamines / Q000633:toxicity*; D002470:Cell Survival / Q000187:drug effects; D003913:Dextroamphetamine / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D004734:Energy Metabolism / Q000187:drug effects; D005978:Glutathione / Q000378:metabolism; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D006979:Hyperthermia, Induced; D015227:Lipid Peroxidation / Q000187:drug effects; D008694:Methamphetamine / Q000633:toxicity; D008930:Mitochondria, Liver / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D051336:Mitochondrial Membranes / Q000187:drug effects; D018817:N-Methyl-3,4-methylenedioxyamphetamine / Q000633:toxicity; D018384:Oxidative Stress / Q000187:drug effects*; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D018570:Risk Assessment
chemical_list: D000662:Amphetamines; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D008694:Methamphetamine; C078956:4-methylthioamphetamine; D000255:Adenosine Triphosphate; D005978:Glutathione; D018817:N-Methyl-3,4-methylenedioxyamphetamine; D003913:Dextroamphetamine
keywords: amphetamines; concentration addition (CA); hepatocytes; hyperthermia; mixtures; oxidative stress