Cyclooxygenase-2 (COX-2) is the inducible form of cyclooxygenase and catalyzes the rate-limiting step in the formation of prostaglandins from arachidonic acid. COX-2 expression and prostaglandin production increase markedly in neurons following a variety of stimuli including inflammation and excitotoxicity. Previous studies in rodent models of ischemia have demonstrated that COX-2 activity potentiates neuronal injury. The mechanism by which COX-2 promotes neuronal injury has not been determined, but likely involves cellular changes resulting from activation of specific prostaglandin receptors, all of which are present in neurons. From preliminary studies, we hypothesize that distinct prostaglandin products may act to promote toxicity through activation of specific prostaglandin receptors and second messenger systems. In preliminary studies, we have determined that prostaglandin receptors can mediate either toxic or protective effects on neuronal survival in culture. Using in vitro culture techniques as well as in vivo studies of PG receptor knockout mice, we propose to define specific COX-2 prostaglandin metabolites and receptor mediated signaling pathways that function in promoting the toxic effects of COX-2 enzymatic activity in stroke.
In Aim 1, using receptor specific agonists and antagonists in neuronal culture, we will determine the effects of activation of individual PG receptor subtypes on neuronal survival. We will then correlate the toxic or protective effects of individual PG receptors with downstream signaling pathways focusing on cAMP production and changes in intracellular Ca2+ levels in Aim 2.
In Aim 3, we will quantify the prostaglandin response following ischemia at various time points during reperfusion and determine the level of expression and anatomic localization of PG receptors. We will test whether toxic prostaglandins and prostaglandin receptors are dominantly induced in this paradigm.
In Aim 4, we will confirm our findings in vivo, and test the function of selected PG receptors using prostaglandin receptor knockout mice that have been made available to us for study. From these studies, we hope to define specific PG receptor mediated pathways that impact on neuronal survival in stroke, and that can be targeted in future studies for therapeutic intervention.
