P276	Characterization of murine endothelial P2Y receptors.
1P-J.Guns, 2T.Van Assche, 2P.Fransen, 2B.Robaye, 3J-M.Boeynaems, 1H.Bult
1University of Antwerp, Antwerp, BE; 2Gosselies, BE; 3Brussels, BE & 2Antwerp, BE.

Objectives: Nucleotides regulate various biological functions including vascular tone. This study was aimed to characterize P2Y receptors on endothelial cells of the murine aorta. In view of the lack of selective purinergic agonists and antagonists, we used P2Y2- and P2Y4-deficient mice. Methods: Five adjacent thoracic aorta segments (width 2mm) of wild type (C57BL6) or P2Y2- or P2Y4-deficient (CD1/129SV) mice were mounted in organ baths to measure isometric force development. Results: Nucleotides evoked complete (ATP, UTP, UDP; >90%) or partial (ADP) relaxation of phenylephrine pre-contracted thoracic aortic rings of wild type mice. The rank order of potency was: UDP~UTP~ADP>ATPγS>ATP. These results are compatible with the presence of P2Y1 (ADP>ATP), P2Y2 or P2Y4 (ATP and UTP) and P2Y6 (UDP) receptors. P2Y4-deficient compared to wild type mice displayed unaltered responses to both ATP as well as UTP. Therefore, P2Y4 receptors are not involved in relaxation. Furthermore, mRNA of P2Y1, P2Y2 and P2Y6 subtypes was present in whole vessel homogenates, whereas P2Y4 mRNA could not be detected. Finally, P2Y2-deficient mice showed impaired ATP-evoked relaxation, suggesting that in wild type mice ATP activates predominantly the P2Y2-subtype. Moreover, in P2Y2-deficient mice ATP-evoked relaxation was rescued by P2Y1, since it was sensitive to MRS2179, a P2Y1 selective antagonist. In contrast, UTP-evoked relaxation was normal in P2Y2-deficient mice. Therefore, the action of UTP is probably mediated by a P2Y6(like) receptor subtype. Conclusion: Taken together, these results point to the presence of functional P2Y1 (ADP>ATP), P2Y2 (ATP) and P2Y6 (UDP) receptor subtypes on murine aortic endothelial cells. UTP probably acts via a P2Y6(like) subtype, since P2Y4 and P2Y2 subtypes were not involved in the UTP-evoked relaxation.

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