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Abstract
The primary objectives stemming from this project were to characterize fully both the equineadenosine A2A (eA2A-R) and A3 (eA3-R) receptors to determine if they would be suitable pharmacological targets for the treatment of equine endotoxemia. To that end, heterologous expression systems were established to express the receptors in human embryonic kidney cells (HEK293) for their elucidation and study. Initially, pharmacological characterization was determined via equilibrium radioligand binding and adenylate cyclase assays. To further characterize the receptors, reporter gene assays were performed in order to determine what effect
receptor activation played on intracellular signaling and inhibition of specific components of the inflammatory cascade. Pharmacological characterization of the eA2A-R and eA3-R revealed that that the heterologously expressed receptors had a pharmacologic profile that was in accordance with that of other mammalian A2A and A3 receptors. Furthermore, it was demonstrated in adenylate cyclase assays that the expressed receptors functionally coupled to the intracellular Gprotein complex as evidenced by the generation of [3H]cAMP for the eA2A-R, and an inhibition of [3H]cAMP for the eA3-R. To characterize these receptors further, reporter gene assays were then performed to ascertain what effect receptor activation would have on intracellular signal transduction; specifically, the NF-