The stable pyrimidines UDPbetaS and UTPgammaS discriminate between the P2 receptors that mediate vascular contraction and relaxation of the rat mesenteric artery

Abstract

The contractile and relaxant effects of the different P2 receptors were characterized in the rat isolated mesenteric artery by use of extracellular nucleotides, including the stable pyrimidines uridine 5'-O-thiodiphosphate (UDPbetaS) and uridine 5'-O-3-thiotriphosphate (UTPgammaS). The selective P2X receptor agonist, alphabeta-methylene-adenosine triphosphate (alphabeta-MeATP) stimulated a potent (pEC(50)=6.0) but relatively weak contraction (E:(max)=57% of 60 mM K(+)). The contractile concentration-response curve of adenosine triphosphate (ATP) was biphasic when added in single concentrations. The first part of the response could be desensitized by alphabeta-MeATP, indicating involvement of P2X receptors, while the second part might be mediated by P2Y receptors. The contractile P2Y receptors were further characterized after P2X receptor desensitization with 10 microM alphabeta-MeATP. Uridine diphosphate (UDP), uridine triphosphate (UTP) and ATP stimulated contraction only in high concentrations (1 - 10 mM). The selective P2Y(6) agonist, UDPbetaS, and the P2Y(2)/P2Y(4)-receptor agonists UTPgammaS and adenosine 5'-O-3-thiotriphosphate (ATPgammaS) were considerably more potent and efficacious (E:(max) approximately 250% of 60 mM K(+)). Adenosine 5'-O-thiodiphosphate (ADPbetaS) was inactive, excluding contractile P2Y(1) receptors. After precontraction with 1 microM noradrenaline, UTP, ADP and ATP induced relaxations with similar potencies (pEC(50) approximately 5.0). UTPgammaS, ADPbetaS and ATPgammaS were approximately one log unit more potent indicating the presence of endothelial P2Y(1) and P2Y(2)/P2Y(4) receptors. The P2Y(6) receptor agonist, UDPbetaS, had no effect. UDPbetaS and UTPgammaS are useful tools when studying P2 receptors in tissue preparations with ectonucleotidase activity. Contractile responses can be elicited by stimulation of P2Y(6) and, slightly less potently, P2Y(2)/P2Y(4) receptors. The P2X response was relatively weak, and there was no P2Y(1) response. Stimulation of P2Y(1) and P2Y(2)/P2Y(4) receptors elicited relaxation, while P2Y(6) did not contribute.

Figure 1

Concentration-dependent contractions to αβ-MeATP, UDPβS, UDP, UTPγS, UTP, ADPβS and ATPγS in the rat mesenteric artery. All nucleotides, except αβ-MeATP, were added after P2X receptor desensitization with 10 μM αβ-MeATP. Contractions are expressed as percentage of an initial contraction induced by 60 mM K⁺. Data are shown as means±s.e.mean of 6–10 experiments.

Figure 2

Concentration-dependent contractions to αβ-MeATP and ATP added in single-concentration, ATP added cumulatively and ATP after P2X receptor desensitization with 10 μM αβ-MeATP. Contractions are expressed as percentage of an initial contraction induced by 60 mM K⁺. Data are shown as means±s.e.mean of 6–10 experiments.

Figure 3

Concentration-dependent relaxations to UDPβS, UDP, UTPγS, UTP, ADPβS, ATPγS and in the rat mesenteric artery. All nucleotides were added after P2X receptor desensitization with 10 μM αβ-MeATP. Relaxations are expressed as percentage of an initial contraction induced by 1 μM NA. Data are shown as means ±s.e.mean of 6–10 experiments.