title: Scavenging ROS dramatically increase NMDA receptor whole-cell currents in painted turtle cortical neurons.
abstract: Oxygen deprivation triggers excitotoxic cell death in mammal neurons through excessive calcium loading via over-activation of N-methyl-d-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. This does not occur in the western painted turtle, which overwinters for months without oxygen. Neurological damage is avoided through anoxia-mediated decreases in NMDA and AMPA receptor currents that are dependent upon a modest rise in intracellular Ca(2+) concentrations ([Ca(2+)]i) originating from mitochondria. Anoxia also blocks mitochondrial reactive oxygen species (ROS) generation, which is another potential signaling mechanism to regulate glutamate receptors. To assess the effects of decreased intracellular [ROS] on NMDA and AMPA receptor currents, we scavenged ROS with N-2-mercaptopropionylglycine (MPG) or N-acetylcysteine (NAC). Unlike anoxia, ROS scavengers increased NMDA receptor whole-cell currents by 100%, while hydrogen peroxide decreased currents. AMPA receptor currents and [Ca(2+)]i concentrations were unaffected by ROS manipulation. Because decreases in [ROS] increased NMDA receptor currents, we next asked whether mitochondrial Ca(2+) release prevents receptor potentiation during anoxia. Normoxic activation of mitochondrial ATP-sensitive potassium (mKATP) channels with diazoxide decreased NMDA receptor currents and was unaffected by subsequent ROS scavenging. Diazoxide application following ROS scavenging did not rescue scavenger-mediated increases in NMDA receptor currents. Fluorescent measurement of [Ca(2+)]i and ROS levels demonstrated that [Ca(2+)]i increases before ROS decreases. We conclude that decreases in ROS concentration are not linked to anoxia-mediated decreases in NMDA/AMPA receptor currents but are rather associated with an increase in NMDA receptor currents that is prevented during anoxia by mitochondrial Ca(2+) release.
authors: Department of Cell and Systems Biology and Department of Ecology and Evolutionary Biology, University of Toronto, Toronto, ON M5S 3G5, Canada.
authors: David James
authors: DJ
authors: Dukoff
authors: Department of Cell and Systems Biology and Department of Ecology and Evolutionary Biology, University of Toronto, Toronto, ON M5S 3G5, Canada.
authors: David William
authors: DW
authors: Hogg
authors: Department of Cell and Systems Biology and Department of Ecology and Evolutionary Biology, University of Toronto, Toronto, ON M5S 3G5, Canada.
authors: Peter John
authors: PJ
authors: Hawrysh
authors: Department of Cell and Systems Biology and Department of Ecology and Evolutionary Biology, University of Toronto, Toronto, ON M5S 3G5, Canada les.buck@utoronto.ca.
authors: Leslie Thomas
authors: LT
authors: Buck
mesh_terms: D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002540:Cerebral Cortex / Q000166:cytology* / Q000502:physiology; D005260:Female; D016166:Free Radical Scavengers; D006861:Hydrogen Peroxide; D008564:Membrane Potentials / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D010100:Oxygen; D018408:Patch-Clamp Techniques; D017382:Reactive Oxygen Species / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000502:physiology*; D012402:Rotenone; D008625:Tiopronin / Q000494:pharmacology; D014426:Turtles / Q000502:physiology*
chemical_list: D016166:Free Radical Scavengers; D017382:Reactive Oxygen Species; D016194:Receptors, N-Methyl-D-Aspartate; D012402:Rotenone; C487056:N-Acetylcysteinamide; D006861:Hydrogen Peroxide; D008625:Tiopronin; D010100:Oxygen; D002118:Calcium; D000111:Acetylcysteine
keywords: Anoxia tolerance; Calcium; Channel arrest; Fluorescence; Pyramidal neurons; Reactive oxidative species; Whole-cell patch-clamp