title: Glutathione modulation influences methyl mercury induced neurotoxicity in primary cell cultures of neurons and astrocytes.
abstract: Methyl mercury (MeHg) is highly neurotoxic and may lead to numerous neurodegenerative disorders. In this study, we investigated the role of glutathione (GSH) and reactive oxygen species (ROS) in MeHg-induced neurotoxicity, using primary cell cultures of cerebellar neurons and astrocytes. To evaluate the effect of GSH on MeHg-induced cytotoxicity, ROS and GSH were measured using the fluorescent indicators chloro methyl derivative of di-chloro di-hydro fluorescein diacetate (CMH(2)DCFDA) and monochlorobimane (MCB). Cell-associated MeHg was measured with (14)C-radiolabeled MeHg. Mitochondrial dehydrogenase activity was detected by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]. MTT timeline study was also performed to evaluate the effects of both the concentration and duration of MeHg exposure. The intracellular GSH content was modified by pretreatment with N-acetyl cysteine (NAC) or di-ethyl maleate (DEM) for 12 h. Treatment with 5 microM MeHg for 30 min led to significant (p<0.05) increase in ROS and reduction (p<0.001) in GSH content. Depletion of intracellular GSH by DEM further increased the generation of MeHg-induced ROS in both cell cultures. Conversely, NAC supplementation increased intracellular GSH and provided protection against MeHg-induced oxidative stress in both cell cultures. MTT studies also confirmed the efficacy of NAC supplementation in attenuating MeHg-induced cytotoxicity. The cell-associated MeHg was significantly (p<0.02) increased after DEM treatment. In summary, depletion of GSH increases MeHg accumulation and enhances MeHg-induced oxidative stress, and conversely, supplementation with GSH precursor protects against MeHg exposure in vitro.
authors.affiliation: Department of Neuroscience, Norwegian University of Science and Technology, N-7489 Trondheim, Norway.
authors.forename: Parvinder
authors.initials: P
authors.lastname: Kaur
authors.forename: Michael
authors.initials: M
authors.lastname: Aschner
authors.forename: Tore
authors.initials: T
authors.lastname: Syversen
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000187:drug effects*; D002452:Cell Count; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D005978:Glutathione / Q000502:physiology*; D020262:Mercury Poisoning, Nervous System / Q000209:etiology / Q000378:metabolism; D008767:Methylmercury Compounds / Q000378:metabolism / Q000633:toxicity*; D051379:Mice; D008928:Mitochondria / Q000187:drug effects; D009474:Neurons / Q000187:drug effects*; D010088:Oxidoreductases / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D013778:Tetrazolium Salts; D013844:Thiazoles; D013997:Time Factors
chemical_list: D004791:Enzyme Inhibitors; D008767:Methylmercury Compounds; D017382:Reactive Oxygen Species; D013778:Tetrazolium Salts; D013844:Thiazoles; D010088:Oxidoreductases; C022616:thiazolyl blue; D005978:Glutathione; D005033:Ethylmaleimide; D000111:Acetylcysteine