Characterisation of P2Y2 receptors in human vascular endothelial cells using AR-C118925XX, a competitive and selective P2Y2 antagonist

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Abstract

Background and Purpose: There is a lack of potent, selective antagonists at most subtypes of P2Y receptor. The aims of this study were to characterise the pharmacological properties of the proposed P2Y 2 receptor antagonist, AR-C118925XX, and then to use it to determine the role of P2Y 2 receptors in the action of the P2Y 2 agonist, UTP, in human vascular endothelial cells. Experimental Approach: Cell lines expressing native or recombinant P2Y receptors were superfused constantly, and agonist-induced changes in intracellular Ca 2+ levels monitored using the Ca 2+-sensitive fluorescent indicator, Cal-520. This set-up enabled full agonist concentration–response curves to be constructed on a single population of cells. Key Results: UTP evoked a concentration-dependent rise in intracellular Ca 2+ in 1321N1-hP2Y 2 cells. AR-C118925XX (10 nM to 1 μM) had no effect per se on intracellular Ca 2+ but shifted the UTP concentration–response curve progressively rightwards, with no change in maximum. The inhibition was fully reversible on washout. AR-C118925XX (1 μM) had no effect at native or recombinant hP2Y 1, hP2Y 4, rP2Y 6, or hP2Y 11 receptors. Finally, in EAhy926 immortalised human vascular endothelial cells, AR-C118925XX (30 nM) shifted the UTP concentration–response curve rightwards, with no decrease in maximum. Conclusions and Implications: AR-C118925XX is a potent, selective and reversible, competitive P2Y 2 receptor antagonist, which inhibited responses mediated by endogenous P2Y 2 receptors in human vascular endothelial cells. As the only P2Y 2-selective antagonist currently available, it will greatly enhance our ability to identify the functions of native P2Y 2 receptors and their contribution to disease and dysfunction.

LanguageEnglish
Pages2894-2904
Number of pages11
JournalBritish Journal of Pharmacology
Volume176
Issue number16
Early online date22 May 2019
DOIs
Publication statusPublished - 1 Aug 2019

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Purinergic P2Y2 Receptors
Uridine Triphosphate
Endothelial Cells
Pharmacology
Cell Line
Population

Keywords

  • AR-C118925XX
  • P2Y2 receptor
  • competitive antagonist
  • endothelial cells

Cite this

@article{59d36cfb19464d5cb70e9f7140426c94,
title = "Characterisation of P2Y2 receptors in human vascular endothelial cells using AR-C118925XX, a competitive and selective P2Y2 antagonist",
abstract = "Background and Purpose: There is a lack of potent, selective antagonists at most subtypes of P2Y receptor. The aims of this study were to characterise the pharmacological properties of the proposed P2Y 2 receptor antagonist, AR-C118925XX, and then to use it to determine the role of P2Y 2 receptors in the action of the P2Y 2 agonist, UTP, in human vascular endothelial cells. Experimental Approach: Cell lines expressing native or recombinant P2Y receptors were superfused constantly, and agonist-induced changes in intracellular Ca 2+ levels monitored using the Ca 2+-sensitive fluorescent indicator, Cal-520. This set-up enabled full agonist concentration–response curves to be constructed on a single population of cells. Key Results: UTP evoked a concentration-dependent rise in intracellular Ca 2+ in 1321N1-hP2Y 2 cells. AR-C118925XX (10 nM to 1 μM) had no effect per se on intracellular Ca 2+ but shifted the UTP concentration–response curve progressively rightwards, with no change in maximum. The inhibition was fully reversible on washout. AR-C118925XX (1 μM) had no effect at native or recombinant hP2Y 1, hP2Y 4, rP2Y 6, or hP2Y 11 receptors. Finally, in EAhy926 immortalised human vascular endothelial cells, AR-C118925XX (30 nM) shifted the UTP concentration–response curve rightwards, with no decrease in maximum. Conclusions and Implications: AR-C118925XX is a potent, selective and reversible, competitive P2Y 2 receptor antagonist, which inhibited responses mediated by endogenous P2Y 2 receptors in human vascular endothelial cells. As the only P2Y 2-selective antagonist currently available, it will greatly enhance our ability to identify the functions of native P2Y 2 receptors and their contribution to disease and dysfunction.",
keywords = "AR-C118925XX, P2Y2 receptor, competitive antagonist, endothelial cells",
author = "Muoboghare, {Markie O.} and Drummond, {Robert M.} and Charles Kennedy",
year = "2019",
month = "8",
day = "1",
doi = "10.1111/bph.14715",
language = "English",
volume = "176",
pages = "2894--2904",
journal = "British Journal of Pharmacology",
issn = "0007-1188",
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TY - JOUR

T1 - Characterisation of P2Y2 receptors in human vascular endothelial cells using AR-C118925XX, a competitive and selective P2Y2 antagonist

AU - Muoboghare, Markie O.

AU - Drummond, Robert M.

AU - Kennedy, Charles

PY - 2019/8/1

Y1 - 2019/8/1

N2 - Background and Purpose: There is a lack of potent, selective antagonists at most subtypes of P2Y receptor. The aims of this study were to characterise the pharmacological properties of the proposed P2Y 2 receptor antagonist, AR-C118925XX, and then to use it to determine the role of P2Y 2 receptors in the action of the P2Y 2 agonist, UTP, in human vascular endothelial cells. Experimental Approach: Cell lines expressing native or recombinant P2Y receptors were superfused constantly, and agonist-induced changes in intracellular Ca 2+ levels monitored using the Ca 2+-sensitive fluorescent indicator, Cal-520. This set-up enabled full agonist concentration–response curves to be constructed on a single population of cells. Key Results: UTP evoked a concentration-dependent rise in intracellular Ca 2+ in 1321N1-hP2Y 2 cells. AR-C118925XX (10 nM to 1 μM) had no effect per se on intracellular Ca 2+ but shifted the UTP concentration–response curve progressively rightwards, with no change in maximum. The inhibition was fully reversible on washout. AR-C118925XX (1 μM) had no effect at native or recombinant hP2Y 1, hP2Y 4, rP2Y 6, or hP2Y 11 receptors. Finally, in EAhy926 immortalised human vascular endothelial cells, AR-C118925XX (30 nM) shifted the UTP concentration–response curve rightwards, with no decrease in maximum. Conclusions and Implications: AR-C118925XX is a potent, selective and reversible, competitive P2Y 2 receptor antagonist, which inhibited responses mediated by endogenous P2Y 2 receptors in human vascular endothelial cells. As the only P2Y 2-selective antagonist currently available, it will greatly enhance our ability to identify the functions of native P2Y 2 receptors and their contribution to disease and dysfunction.

AB - Background and Purpose: There is a lack of potent, selective antagonists at most subtypes of P2Y receptor. The aims of this study were to characterise the pharmacological properties of the proposed P2Y 2 receptor antagonist, AR-C118925XX, and then to use it to determine the role of P2Y 2 receptors in the action of the P2Y 2 agonist, UTP, in human vascular endothelial cells. Experimental Approach: Cell lines expressing native or recombinant P2Y receptors were superfused constantly, and agonist-induced changes in intracellular Ca 2+ levels monitored using the Ca 2+-sensitive fluorescent indicator, Cal-520. This set-up enabled full agonist concentration–response curves to be constructed on a single population of cells. Key Results: UTP evoked a concentration-dependent rise in intracellular Ca 2+ in 1321N1-hP2Y 2 cells. AR-C118925XX (10 nM to 1 μM) had no effect per se on intracellular Ca 2+ but shifted the UTP concentration–response curve progressively rightwards, with no change in maximum. The inhibition was fully reversible on washout. AR-C118925XX (1 μM) had no effect at native or recombinant hP2Y 1, hP2Y 4, rP2Y 6, or hP2Y 11 receptors. Finally, in EAhy926 immortalised human vascular endothelial cells, AR-C118925XX (30 nM) shifted the UTP concentration–response curve rightwards, with no decrease in maximum. Conclusions and Implications: AR-C118925XX is a potent, selective and reversible, competitive P2Y 2 receptor antagonist, which inhibited responses mediated by endogenous P2Y 2 receptors in human vascular endothelial cells. As the only P2Y 2-selective antagonist currently available, it will greatly enhance our ability to identify the functions of native P2Y 2 receptors and their contribution to disease and dysfunction.

KW - AR-C118925XX

KW - P2Y2 receptor

KW - competitive antagonist

KW - endothelial cells

UR - https://www.bps.ac.uk/publishing/our-journals/british-journal-of-pharmacology

U2 - 10.1111/bph.14715

DO - 10.1111/bph.14715

M3 - Article

VL - 176

SP - 2894

EP - 2904

JO - British Journal of Pharmacology

T2 - British Journal of Pharmacology

JF - British Journal of Pharmacology

SN - 0007-1188

IS - 16

ER -