title: Methylmercury-induced reactive oxygen species formation in neonatal cerebral astrocytic cultures is attenuated by antioxidants.
abstract: Excessive generation of reactive oxygen species (ROS) has been suggested as a causal factor in various neurodegenerative disorders, such as Parkinson's disease and Alzheimer's disease [Brain Res. 830 (1999) 10-15; Biochem. J. 310 (1995) 83-90; Free Radic. Biol. Med. 27 (1999) 612-616]. The present work examined the role of ROS in the neurotoxicity of methylmercury (MeHg). ROS formation in primary astrocytic cultures of neonatal rat cerebral cortex was monitored by 2',7'-dichlorodihydrofluorescein diacetate (H(2)DCF-DA) fluorescence. MeHg, at 10 and 20 microM caused a significant increase in ROS formation (10 microM, P<0.01; 20 microM, P<0.001). Additional studies established the effectiveness of antioxidants/free radical scavengers in attenuating the MeHg-stimulated ROS formation in the following rank-order: (1) Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid), a non-thiol containing antioxidant, (2) n-propyl gallate (PG), a free radical scavenger, (3) superoxide dismutase (SOD), an antioxidant enzyme that dismutates superoxide anion radical, (4) alpha-phenyl-tert-butyl nitrone (PBN), a lipophilic hydroxyl radical spin trapping agent. A significant inhibition of MeHg-induced ROS generation was also noted in astrocytes preincubated (3 h) with arachidonyl trifluoromethyl ketone (AACOCF(3,) 20 microM, P<0.05), a specific inhibitor of cytosolic phospholipase A(2) (cPLA(2)). Conversely, pretreatment (24 h) with 100 microM buthionine-L-sulfoxamine [BSO, a glutathione (GSH) synthesis inhibitor], significantly increased (P<0.05) ROS formation in MeHg treated astrocytes compared to controls. Combined, these studies invoke ROS as potent mediators of MeHg cytotoxicity and support the hypothesis that excessive ROS generation, at least in part, plays an important role in MeHg-induced neurotoxicity.
authors.affiliation: Department of Physiology and Pharmacology, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1083, USA.
authors.forename: Gouri
authors.initials: G
authors.lastname: Shanker
authors.forename: Michael
authors.initials: M
authors.lastname: Aschner
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D000975:Antioxidants / Q000494:pharmacology*; D001095:Arachidonic Acids / Q000494:pharmacology; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D002478:Cells, Cultured; D002839:Chromans / Q000494:pharmacology*; D003497:Cyclic N-Oxides; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D008767:Methylmercury Compounds / Q000633:toxicity*; D018696:Neuroprotective Agents / Q000494:pharmacology; D009589:Nitrogen Oxides / Q000494:pharmacology; D011435:Propyl Gallate / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013482:Superoxide Dismutase / Q000494:pharmacology
chemical_list: D000975:Antioxidants; D001095:Arachidonic Acids; D002839:Chromans; D003497:Cyclic N-Oxides; D004791:Enzyme Inhibitors; D008767:Methylmercury Compounds; D018696:Neuroprotective Agents; D009589:Nitrogen Oxides; D017382:Reactive Oxygen Species; C081565:arachidonyltrifluoromethane; C029217:phenyl-N-tert-butylnitrone; D011435:Propyl Gallate; D013482:Superoxide Dismutase; D005978:Glutathione; C004925:methylmercuric chloride; C010643:6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid