title: Methylmercury alters the in vitro uptake of glutamate in GLAST- and GLT-1-transfected mutant CHO-K1 cells.
abstract: In order to maintain normal functioning of the brain, glutamate homeostasis and extracellular levels of excitotoxic amino acids (EAA) must be tightly controlled. This is accomplished, in large measure, by the astroglial high-affinity Na+-dependent EAA transporters glutamate/ aspartate transporter (GLAST) and glutamate transporter-1 (GLT-1). Methylmercury (MeHg) is a potent neurotoxicant. Astrocytes are known targets for MeHg toxicity, representing a site for mercury localization. MeHg is known to cause astrocytic swelling, EAA release, and uptake inhibition in astrocytes, leading to increased extracellular glutamate levels and ensuing neuronal excitotoxicity and degeneration. However, the mechanisms and contribution of specific glutamate transporters to MeHg-induced glutamate dyshomeostasis remain unknown. Accordingly, the present study was carried out to investigate the effects of MeHg on the transport of [d-2, 3-3H]-d-aspartate, a nonmetabolizable glutamate analog in Chinese hamster ovary cells (CHO) transfected with the glutamate transporter subtypes GLAST or GLT-1. Additional studies examined the effects of MeHg on mRNA and protein levels of these transporters. Our results indicate the following (1) MeHg selectively affects glutamate transporter mRNA expression. MeHg treatment (6 h) led to no discernible changes in GLAST mRNA expression; however, GLT-1 mRNA expression significantly (p < 0.001) increased following treatments with 5 or 10 microM MeHg. (2) Selective changes in the expression of glutamate transporter protein levels were also noted. GLAST transporter protein levels significantly (p < 0.001, both at 5 and 10 microM MeHg) increased and GLT-1 transporter protein levels significantly (p < 0.001) decreased following MeHg exposure (5 microM). (3) MeHg exposure led to significant inhibition (p < 0.05) of glutamate uptake by GLAST (both 5 and 10 microM MeHg), whereas GLT-1 transporter activity was significantly (p < 0.01) increased following exposure to 5 and 10 microM MeHg. These studies suggest that MeHg contributes to the dysregulation of glutamate homeostasis and that its effects are distinct for GLAST and GLT-1.
authors.affiliation: Department of Physiology and Pharmacology, Wake Forest University School of Medicine, Winston-Salem, NC 27157-1083, USA.
authors.forename: Lysette
authors.initials: L
authors.lastname: Mutkus
authors.forename: Judy L
authors.initials: JL
authors.lastname: Aschner
authors.forename: Tore
authors.initials: T
authors.lastname: Syversen
authors.forename: Michael
authors.initials: M
authors.lastname: Aschner
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism; D001692:Biological Transport; D015152:Blotting, Northern; D015153:Blotting, Western; D016466:CHO Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D027341:Excitatory Amino Acid Transporter 1 / Q000235:genetics* / Q000378:metabolism; D027342:Excitatory Amino Acid Transporter 2 / Q000235:genetics* / Q000378:metabolism; D005971:Glutamates / Q000493:pharmacokinetics*; D008767:Methylmercury Compounds / Q000494:pharmacology*; D009154:Mutation; D009474:Neurons / Q000378:metabolism; D009498:Neurotoxins / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D012964:Sodium / Q000737:chemistry; D013997:Time Factors; D014131:Trace Elements; D014162:Transfection
chemical_list: D027341:Excitatory Amino Acid Transporter 1; D027342:Excitatory Amino Acid Transporter 2; D005971:Glutamates; D008767:Methylmercury Compounds; D009498:Neurotoxins; D012333:RNA, Messenger; D014131:Trace Elements; D012964:Sodium; C004925:methylmercuric chloride