Effect of Coenzyme A derivatives on P2Y1 purine receptor-mediated vasorelaxations in the rat isolated thoracic aorta

Stephen Alexander, E elEfaishat, Vera Ralevic

University of Nottingham, Nottingham, UK

Coenzyme A (CoA) and its acyl derivatives have been shown to be antagonists at human recombinant P2Y1 purine receptors (Coddou et al., 2003). These compounds have also been shown to inhibit platelet function by blocking P2Y1 and P2Y12 receptor-mediated responses (Manolopoulos et al., 2008). The present study investigated the effects of these compounds on vasorelaxation mediated by P2Y receptors in the rat isolated thoracic aorta. The rat thoracic aorta is known to coexpress vasorelaxant P2Y1 and P2Y2 receptors on the endothelium (Bultmann et al., 1998).

Segments of thoracic aorta from male Wistar rats (200-250 g) were mounted for isometric tension recordings, as described previously (Bultmann et al., 1998). Tissue viability was assessed by eliciting contractions with 60 mM KCl. The rings were preconstricted with methoxamine (0.3-2.0 μM) to a tension of 55-85% of the KCl contraction. Concentration-relaxation response curves were then constructed in the presence of increasing cumulative concentrations of ADP or UTP, P2Y1- and P2Y2-selective agonists respectively. In separate experiments CoA, acetyl CoA or palmitoyl CoA were added 10 min before methoxamine addition. Data reported are means ± s.e.m of results from at least 4 experiments and were compared for statistical analysis by ANOVA. P < 0.05 was taken as significant.

ADP evoked concentration-dependent relaxations of the pre-contracted aorta with a calculated maximal response of 110 ± 8 % relaxation and pEC50 value of 5.43 ± 0.41 (n = 21). In contrast, the relaxation evoked by UTP was bell-shaped with reduced relaxatory responses observed at concentrations above 3 μM. Curve fitting of relaxations to lower concentrations of UTP allowed calculation of a maximal response of 94 ± 5 % and pEC50 value of 6.64 ± 0.04 (n = 8). In the presence of CoA (10 μM), acetyl CoA (10 μM) and palmitoyl CoA (10 μM), 5-, 6-, and 692-fold rightward shifts in the ADP-mediated relaxation were observed. Extending the range of concentrations of palmitoyl CoA examined for Schild analysis allowed calculation of a pA2 value of 6.34 ± 0.08 with a slope of 2.09 ± 0.35. CoA and acetyl CoA had no significant effect on vasorelaxations to UTP, and there was only a small rightward 3-fold rightward shift in the presence of palmitoyl CoA (10 μM).

The preferential inhibition of ADP-evoked relaxations compared to UTP-elicited relaxations indicates that palmitoyl CoA can act as a potent and selective antagonist for the vascular P2Y1 receptor, with little effect at the P2Y2 receptor.

We gratefully acknowledge and appreciate the Jordanian government, who sponsored this research.

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Manolopoulos P, Glenn JR, Fox SC, May JA, Dovlatova NL, Tang S-W, Thomas NR, Ralevic V, Heptinstall S (2008). Platelets 19: 134-145