title: Extracellular reduced glutathione increases neuronal vulnerability to combined chemical hypoxia and glucose deprivation.
abstract: In addition to its intracellular antioxidant role, reduced glutathione (GSH) is released by CNS cells and may mediate or modulate excitatory neurotransmission. Although extracellular GSH levels rise in the ischemic cortex, its effect on the viability of energy-compromised neurons has not been defined. In this study, we tested the hypothesis that exogenous GSH would increase the vulnerability of cultured cortical neurons to azide-induced chemical hypoxia combined with glucose deprivation. Thirty minutes azide exposure in a glucose-free buffer was tolerated by most neurons, with release of less than 10% of neuronal LDH over the subsequent 21-25 h. Concomitant treatment with 10-100 microM GSH increased cell death in a concentration-dependent fashion, to 71.6+/-5.1% of neurons at 100 microM; GSH alone was nontoxic. Injury was blocked by the selective N-methyl-d-aspartate (NMDA) antagonist MK-801 but not by the AMPA/kainate antagonist NBQX. The sulfhydryl reducing agent mercaptoethanol (10-100 microM) mimicked the action of GSH; however, the zinc chelator ethylenediaminetetraacetic acid (EDTA) was ineffective. Two GSH analogues that lack a sulfhydryl group, S-hexylglutathione (SHG) and oxidized glutathione (GSSG), were inactive per se but attenuated the effect of both GSH and mercaptoethanol. These results suggest that micromolar concentrations of GSH enhance neuronal loss due to energy depletion by altering the extracellular redox state, resulting in increased NMDA receptor activation.
authors.affiliation: Division of Emergency Medicine, Thomas Jefferson University, 1020 Sansom Street, 239 Thompson Building, Philadelphia, PA 19107, USA. raymond.f.regan@mail.tju.edu
authors.forename: R F
authors.initials: RF
authors.lastname: Regan
authors.forename: Y
authors.initials: Y
authors.lastname: Guo
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005947:Glucose / Q000172:deficiency*; D005978:Glutathione / Q000494:pharmacology*; D019803:Glutathione Disulfide / Q000494:pharmacology; D002534:Hypoxia, Brain / Q000139:chemically induced / Q000188:drug therapy* / Q000473:pathology; D008623:Mercaptoethanol / Q000494:pharmacology; D051379:Mice; D009474:Neurons / Q000187:drug effects*; D010084:Oxidation-Reduction; D011810:Quinoxalines / Q000494:pharmacology; D009435:Synaptic Transmission / Q000187:drug effects*
chemical_list: D018691:Excitatory Amino Acid Antagonists; D011810:Quinoxalines; C062865:2,3-dioxo-6-nitro-7-sulfamoylbenzo(f)quinoxaline; D008623:Mercaptoethanol; D016291:Dizocilpine Maleate; D004492:Edetic Acid; D005978:Glutathione; D005947:Glucose; D019803:Glutathione Disulfide