Studies during the previous funding period have established that cyclooxygenase-2 (COX-2), a rate-limiting enzyme for prostanoid synthesis, contributes to ischemic brain injury and glutamate excitotoxicity. However, the reaction products responsible for these deleterious effects, their receptor interactions, and the mechanisms of their action have not been established. Identifying the mediators of neurotoxicity provides the bases for treatment modalities targeting the deleterious effects of COX-2 without opposing the beneficial vascular actions of this enzyme. The long-term goals of this renewal application are to elucidate the specific COX-2 reaction products that mediate ischemic brain injury and to begin to define their mechanism of action. The proposed studies will test the following hypotheses: (1) Prostanoids rather than reactive oxygen species are the main COX-2 reaction product initiating the injury; (2) Prostaglandin E2, acting through neuronal EP1 receptors, is responsible for the deleterious effects of COX-2; (3) EP1 receptors play a role in ischemic brain injury by contributing to glutamate excitotoxicity; (4) EP1 receptors contribute to excitotoxicity by amplifying the Ca++ dysregulation produced by activation of glutamate receptors. Experiments will be conducted in mice in which cerebral ischemia is produced by transient occlusion of the middle cerebral artery. The role of COX-2 reaction products will be investigated using pharmacological inhibitors, transgenic mice overexpressing the antioxidant enzyme superoxide dismutase 1, or null mice lacking COX-2 or EP1 receptors. Ischemic brain injury will be assessed by histological and behavioral criteria. Cellular, molecular, biochemical and neuroanatomical techniques will be used to define the mechanisms by which COX-2 reaction products contribute to brain injury. The results of these studies will provide novel insights into the specific factors mediating COX-2-dependent neurotoxicity, and will offer the opportunity for new therapeutic approaches that selectively target the deleterious effects of COX-2. ? ?
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