title: Phosphoinositide 3-kinase couples NMDA receptors to superoxide release in excitotoxic neuronal death.
abstract: Sustained activation of neuronal N-methly D-aspartate (NMDA)-type glutamate receptors leads to excitotoxic cell death in stroke, trauma, and neurodegenerative disorders. Excitotoxic neuronal death results in part from superoxide produced by neuronal NADPH oxidase (NOX2), but how NMDA receptors are coupled to neuronal NOX2 activation is not well understood. Here, we identify a signaling pathway coupling NMDA receptor activation to NOX2 activation in primary neuron cultures. Calcium influx through the NR2B subunit of NMDA receptors leads to the activation of phosphoinositide 3-kinase (PI3K). Formation of phosphatidylinositol (3,4,5)-triphosphate (PI(3,4,5)P3) by PI3K activates the atypical protein kinase C, PKC zeta (PKCζ), which in turn phosphorylates the p47(phox) organizing subunit of neuronal NOX2. Calcium influx through NR2B-containing NMDA receptors triggered mitochondrial depolarization, NOX2 activation, superoxide formation, and cell death. However, equivalent magnitude calcium elevations induced by ionomycin did not induce NOX2 activation or neuronal death, despite causing mitochondrial depolarization. The PI3K inhibitor wortmannin prevented NMDA-induced NOX2 activation and cell death, without preventing cell swelling, calcium elevation, or mitochondrial depolarization. The effects of wortmannin were circumvented by exogenous supply of the PI3K product, PI(3,4,5)P3, and by transfection with protein kinase M, a constitutively active form of PKCζ. These findings demonstrate that superoxide formation and excitotoxic neuronal death can be dissociated from mitochondrial depolarization, and identify a novel role for PI3K in this cell death pathway. Perturbations in this pathway may either increase or decrease superoxide production in response to NMDA receptor activation, and may thereby impact neurological disorders, in which excitotoxicity is a contributing factor.
authors.affiliation: Department of Neurology, San Francisco Veterans Affairs Medical Center, University of California, San Francisco, CA 94121, USA. angela.brennan@ucsf.edu
authors.forename: A M
authors.initials: AM
authors.lastname: Brennan-Minnella
authors.forename: Y
authors.initials: Y
authors.lastname: Shen
authors.forename: J
authors.initials: J
authors.lastname: El-Benna
authors.forename: R A
authors.initials: RA
authors.lastname: Swanson
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004622:Embryo, Mammalian; D005786:Gene Expression Regulation / Q000187:drug effects; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D051379:Mice; D018345:Mice, Knockout; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics* / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D061251:Primary Cell Culture; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D016194:Receptors, N-Methyl-D-Aspartate / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013481:Superoxides / Q000378:metabolism; D000077191:Wortmannin; D000099221:Protein Kinase C zeta
chemical_list: D000730:Androstadienes; D002118:Calcium; D008562:Membrane Glycoproteins; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D019869:Phosphatidylinositol 3-Kinases; D010716:Phosphatidylinositols; D011493:Protein Kinase C; D047428:Protein Kinase Inhibitors; D021122:Protein Subunits; D016194:Receptors, N-Methyl-D-Aspartate; D013481:Superoxides; D000077191:Wortmannin; D000099221:Protein Kinase C zeta; C118303:phosphoinositide-3,4,5-triphosphate; C488842:Cybb protein, mouse
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