Investigation of the functional expression of purine and pyrimidine receptors in porcine isolated pancreatic arteries.

Receptors for purines and pyrimidines are expressed throughout the cardiovascular system. This study investigated their functional expression in porcine isolated pancreatic arteries. Pancreatic arteries (endothelium intact or denuded) were prepared for isometric tension recording and preconstricted with U46619, a thromboxane A(2) mimetic; adenosine-5'-diphosphate (ADP), uridine-5'-triphosphate (UTP) and MRS2768, a selective P2Y(2) agonist, were applied cumulatively, while adenosine-5'-triphosphate (ATP) and αß-methylene-ATP (αß-meATP) response curves were generated from single concentrations per tissue segment. Antagonists/enzyme inhibitors were applied prior to U46619 addition. ATP, αß-meATP, UTP and MRS2768 induced vasoconstriction, with a potency order of αß-meATP > MRS2768 > ATP ≥ UTP. Contractions to ATP and αß-meATP were blocked by NF449, a selective P2X(1) receptor antagonist. The contraction induced by ATP, but not UTP, was followed by vasorelaxation. Endothelium removal and DUP 697, a cyclooxygenase-2 inhibitor, had no significant effect on contraction to ATP but attenuated that to UTP, indicating actions at distinct receptors. MRS2578, a selective P2Y(6) receptor antagonist, had no effect on contractions to UTP. ADP induced endothelium-dependent vasorelaxation which was inhibited by MRS2179, a selective P2Y(1) receptor antagonist, or SCH58261, a selective adenosine A(2A) receptor antagonist. The contractions to ATP and αß-meATP were attributed to actions at P2X(1) receptors on the vascular smooth muscle, whereas it was shown for the first time that UTP induced an endothelium-dependent vasoconstriction which may involve P2Y(2) and/or P2Y(4) receptors. The relaxation induced by ADP is mediated by P2Y(1) and A(2A) adenosine receptors. Porcine pancreatic arteries appear to lack vasorelaxant P2Y(2) and P2Y(4) receptors.

Novel vasocontractile role of the P2Y14 receptor: characterization of its signalling in porcine isolated pancreatic arteries.

BACKGROUND AND PURPOSE: The P2Y14 receptor is the newest member of the P2Y receptor family; it is G(i/o) protein-coupled and is activated by UDP and selectively by UDP-glucose and MRS2690 (2-thiouridine-5'-diphosphoglucose) (7-10-fold more potent than UDP-glucose). This study investigated whether P2Y14 receptors were functionally expressed in porcine isolated pancreatic arteries. EXPERIMENTAL APPROACH: Pancreatic arteries were prepared for isometric tension recording and UDP-glucose, UDP and MRS2690 were applied cumulatively after preconstriction with U46619, a TxA2 mimetic. Levels of phosphorylated myosin light chain 2 (MLC2) were assessed with Western blotting. cAMP concentrations were assessed using a competitive enzyme immunoassay kit. KEY RESULTS: Concentration-dependent contractions with a rank order of potency of MRS2690 (10-fold) > UDP-glucose ≥ UDP were recorded. These contractions were reduced by PPTN {4-[4-(piperidin-4-yl)phenyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthoic acid}, a selective antagonist of P2Y14 receptors, which did not affect responses to UTP. Contraction to UDP-glucose was not affected by MRS2578, a P2Y6 receptor selective antagonist. Raising cAMP levels and forskolin, in the presence of U46619, enhanced contractions to UDP-glucose. In addition, UDP-glucose and MRS2690 inhibited forskolin-stimulated cAMP levels. Removal of the endothelium and inhibition of endothelium-derived contractile agents (TxA2, PGF(2α) and endothelin-1) inhibited contractions to UDP glucose. Y-27632, nifedipine and thapsigargin also reduced contractions to the agonists. UDP-glucose and MRS2690 increased MLC2 phosphorylation, which was blocked by PPTN. CONCLUSIONS AND IMPLICATIONS: P2Y14 receptors play a novel vasocontractile role in porcine pancreatic arteries, mediating contraction via cAMP-dependent mechanisms, elevation of intracellular Ca²âº levels, activation of RhoA/ROCK signalling and MLC2, along with release of TxA2, PGF(2α) and endothelin-1.