Novel vasocontractile role of the <scp>P2Y₁₄</scp> receptor: characterization of its signalling in porcine isolated pancreatic arteries

<jats:sec><jats:title>Background and Purpose</jats:title><jats:p>The <jats:styled-content style="fixed-case">P2Y<jats:sub>14</jats:sub></jats:styled-content> receptor is the newest member of the <jats:styled-content style="fixed-case">P2Y</jats:styled-content> receptor family; it is <jats:styled-content style="fixed-case">G</jats:styled-content><jats:sub>i/o</jats:sub> protein‐coupled and is activated by <jats:styled-content style="fixed-case">UDP</jats:styled-content> and selectively by <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose and <jats:styled-content style="fixed-case">MRS2690</jats:styled-content> (2‐thiouridine‐5′‐diphosphoglucose) (7–10‐fold more potent than <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose). This study investigated whether <jats:styled-content style="fixed-case">P2Y<jats:sub>14</jats:sub></jats:styled-content> receptors were functionally expressed in porcine isolated pancreatic arteries.</jats:p></jats:sec><jats:sec><jats:title>Experimental Approach</jats:title><jats:p>Pancreatic arteries were prepared for isometric tension recording and <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose, <jats:styled-content style="fixed-case">UDP</jats:styled-content> and <jats:styled-content style="fixed-case">MRS2690</jats:styled-content> were applied cumulatively after preconstriction with <jats:styled-content style="fixed-case">U46619</jats:styled-content>, a <jats:styled-content style="fixed-case">TxA<jats:sub>2</jats:sub></jats:styled-content> mimetic. Levels of phosphorylated myosin light chain 2 (<jats:styled-content style="fixed-case">MLC2</jats:styled-content>) were assessed with <jats:styled-content style="fixed-case">W</jats:styled-content>estern blotting. <jats:styled-content style="fixed-case">cAMP</jats:styled-content> concentrations were assessed using a competitive enzyme immunoassay kit.</jats:p></jats:sec><jats:sec><jats:title>Key Results</jats:title><jats:p>Concentration‐dependent contractions with a rank order of potency of <jats:styled-content style="fixed-case">MRS2690</jats:styled-content> (10‐fold) > <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose ≥ <jats:styled-content style="fixed-case">UDP</jats:styled-content> were recorded. These contractions were reduced by <jats:styled-content style="fixed-case">PPTN</jats:styled-content> {4‐[4‐(piperidin‐4‐yl)phenyl]‐7‐[4‐(trifluoromethyl)phenyl]‐2‐naphthoic acid}, a selective antagonist of <jats:styled-content style="fixed-case">P2Y<jats:sub>14</jats:sub></jats:styled-content> receptors, which did not affect responses to UTP. Contraction to <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose was not affected by <jats:styled-content style="fixed-case">MRS2578</jats:styled-content>, a <jats:styled-content style="fixed-case">P2Y<jats:sub>6</jats:sub></jats:styled-content> receptor selective antagonist. Raising <jats:styled-content style="fixed-case">cAMP</jats:styled-content> levels and forskolin, in the presence of U46619, enhanced contractions to <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose. In addition, <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose and <jats:styled-content style="fixed-case">MRS2690</jats:styled-content> inhibited forskolin‐stimulated <jats:styled-content style="fixed-case">cAMP</jats:styled-content> levels. Removal of the endothelium and inhibition of endothelium‐derived contractile agents (<jats:styled-content style="fixed-case">TxA<jats:sub>2</jats:sub></jats:styled-content>, <jats:styled-content style="fixed-case">PGF<jats:sub>2α</jats:sub></jats:styled-content> and endothelin‐1) inhibited contractions to UDP glucose. <jats:styled-content style="fixed-case">Y</jats:styled-content>‐27632, nifedipine and thapsigargin also reduced contractions to the agonists. <jats:styled-content style="fixed-case">UDP</jats:styled-content>‐glucose and <jats:styled-content style="fixed-case">MRS2690</jats:styled-content> increased <jats:styled-content style="fixed-case">MLC2</jats:styled-content> phosphorylation, which was blocked by <jats:styled-content style="fixed-case">PPTN</jats:styled-content>.</jats:p></jats:sec><jats:sec><jats:title>Conclusions and Implications</jats:title><jats:p><jats:styled-content style="fixed-case">P2Y<jats:sub>14</jats:sub></jats:styled-content> receptors play a novel vasocontractile role in porcine pancreatic arteries, mediating contraction via <jats:styled-content style="fixed-case">cAMP</jats:styled-content>‐dependent mechanisms, elevation of intracellular <jats:styled-content style="fixed-case">Ca<jats:sup>2</jats:sup></jats:styled-content><jats:sup>+</jats:sup> levels, activation of <jats:styled-content style="fixed-case">RhoA</jats:styled-content>/<jats:styled-content style="fixed-case">ROCK</jats:styled-content> signalling and <jats:styled-content style="fixed-case">MLC2</jats:styled-content>, along with release of <jats:styled-content style="fixed-case">TxA<jats:sub>2</jats:sub></jats:styled-content>, <jats:styled-content style="fixed-case">PGF<jats:sub>2α</jats:sub></jats:styled-content> and endothelin‐1.</jats:p></jats:sec>