title: Cyclooxygenase-2 contributes to N-methyl-D-aspartate-mediated neuronal cell death in primary cortical cell culture.
abstract: Cyclooxygenase isozymes (COX-1 and COX-2) are found to be constitutively expressed in brain, with neuronal expression of COX-2 being rapidly induced after numerous insults, including cerebral ischemia. Because overactivation of N-methyl-D-aspartate (NMDA) receptors has been implicated in the cell loss associated with ischemia, we characterized the expression of the COX isozymes in murine mixed cortical cell cultures and used isozyme-selective inhibitors to determine their relative contribution to NMDA receptor-stimulated prostaglandin (PG) production and excitotoxic neuronal cell death. Immunocytochemical analysis of mixed cortical cell cultures revealed that COX-2 expression was restricted to neurons, whereas COX-1 was expressed in both neurons and astrocytes. Brief exposure to NMDA (5 min; 100 microM) elicited a time-dependent accumulation of PGs in the culture medium that preceded neuronal cell death and correlated with the induction of COX-2 mRNA. COX-1 expression remained unchanged. Flurbiprofen, a nonselective COX-1/COX-2 inhibitor, blocked NMDA-stimulated PG production and attenuated neuronal death in a concentration-dependent manner. Similar results were obtained with the specific COX-2 inhibitor NS-398 (10-30 microM) but not with the selective COX-1 inhibitor valeryl salicylate (10-300 microM). Inhibition of total constitutive COX activity with aspirin (100 microM, 1.5 h) before NMDA exposure did not prevent subsequent NMDA-mediated neuronal cell death. However, neuronal injury in aspirin-pretreated cultures was attenuated by flurbiprofen administration after NMDA exposure. Finally, the protection afforded by COX-2 inhibition was specific for NMDA because neither flurbiprofen nor NS-398 protected neurons against kainate-mediated neurotoxicity. Together, these results support the conclusion that newly synthesized COX-2 protein contributes to NMDA-induced neuronal injury.
authors.affiliation: Department of Pharmacology, Program in Neuroscience, University of Connecticut Health Center, Farmington, Connecticut 06030-6125, USA. shewett@neuron.uchc.edu
authors.forename: S J
authors.initials: SJ
authors.lastname: Hewett
authors.forename: T F
authors.initials: TF
authors.lastname: Uliasz
authors.forename: A S
authors.initials: AS
authors.lastname: Vidwans
authors.forename: J A
authors.initials: JA
authors.lastname: Hewett
mesh_terms: D000818:Animals; D015153:Blotting, Western; D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D018690:Excitatory Amino Acid Agonists / Q000633:toxicity*; D005480:Flurbiprofen / Q000494:pharmacology; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D007608:Kainic Acid / Q000633:toxicity; D008565:Membrane Proteins; D051379:Mice; D016202:N-Methylaspartate / Q000633:toxicity*; D009474:Neurons / Q000187:drug effects* / Q000201:enzymology; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D011453:Prostaglandins / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D018690:Excitatory Amino Acid Agonists; D007527:Isoenzymes; D008565:Membrane Proteins; D011453:Prostaglandins; D012333:RNA, Messenger; D005480:Flurbiprofen; D016202:N-Methylaspartate; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; C496512:Ptgs1 protein, mouse; D007608:Kainic Acid