The pyrimidine nucleotides UTP and UDP, are ligands for the P2Y receptors P2Y₂, P2Y₄ and P2Y₆ (Ralevic & Burnstock, 1998). Pharmacologically, it may be possible to distinguish these receptors based on the relative potency of the pyrimidine nucleotides and the sensitivity to the P2 antagonists suramin and PPADS (Charlton et al., 1996; Communi et al., 1996). We set out to examine the expression of these pyrimidine nucleotide-sensitive receptors in porcine coronary artery (PCA) and ear artery (PEA) smooth muscle, by examining contractile responses to these ligands and RT-PCR to confirm the expression of mRNA coding for these receptors.

Non-cumulative concentration contraction curves were constructed in 5-8 separate preparations of endothelium-denuded PCA and PEA segments and initially expressed as a proportion of the response to 60 mM KCl. RNA extracted from endothelium-denuded tissue segments was reverse transcribed and subjected to PCR with pig-specific primers for hypoxanthine phosphoribosyltransferase (housekeeping gene), p2y2r, p2y4r and p2y6r. Annealing temperatures for PCR were 58, 57 and 55°C, with 30, 40 and 28 cycles for p2y2r, p2y4r and p2y6r, respectively. PCR products were visualised after separation by electrophoresis through 1.5% agarose. Results were assessed for statistical significance by 1-way ANOVA with Dunnett’s post-hoc test (*p<0.05, **p<0.01).

Table 1: Contractile responses to pyrimidine nucleotides

UTP and UDP induced concentration-dependent contractions in both PCA and PEA. In the presence of suramin, responses to 3 mM UTP were significantly inhibited in both PCA and PEA, while PPADS evoked a significant inhibition of UTP responses only in the PEA (Table 1). Responses to 1 mM UDP were unaffected in either tissue in the presence of suramin or PPADS (Table 1). RT-PCR analysis confirmed the presence of p2y2r, p2y4r and p2y6r in both PCA and PEA.

Taken together, these results provide evidence for functional P2Y 2 receptors activated by UTP in the PCA, since these receptors are sensitive to suramin and not PPADS (Charlton et al., 1996). It is possible that the PEA expresses functional P2Y₂ and P2Y₄ receptors, as responses to UTP were sensitive to both PPADS and suramin (Charlton et al., 1996; Communi et al., 1996). The modest response to UDP in the PEA and the lack of sensitivity of UDP responses to the antagonists in the PCA suggests the presence of functional P2Y₆ receptors in the latter tissue only.

Charlton SJ et al. (1996) Br. J. Pharmacol. 118, 704-710
Communi D et al. (1996) Eur. J. Pharmacol. 317, 383-389
Ralevic V & Burnstock G (1998) Pharmacol.Rev. 50, 413-492