Investigation of the functional expression of receptors for ATP in porcine isolated pancreatic arteries

Mouhamed Alsaqati, Susan L.F. Chan, Vera Ralevic. University of Nottingham, Nottingham, UK

Background: Adenosine-5’-triphosphate (ATP) and other nucleotides may play an important role in regulating blood flow in tissues, the effects of which are dependent on P2 receptors expressed in the vasculature. ATP has been shown to induce smooth muscle vasoconstriction and vasorelaxation via endothelial cells which release endothelial derived relaxing factors (1). However, little is known about the effects of ATP in porcine pancreatic blood vessels. The aim of the current study was to examine the effects of ATP on vascular tone and its mechanism of action in porcine isolated pancreatic arteries.

Methods: Pancreases from pigs were obtained on ice from a local abattoir. Segments of pancreatic arteries were prepared for isometric tension recording in warmed oxygenated Krebs-Henseleit buffer. The effects of ATP were examined at concentrations from 1µM-10mM after preconstriction with U46619 (10-100nM), a thromboxane A2-mimetic. Subsequently a sub-maximal concentration of ATP (1mM) was investigated in the absence and presence of NF449 (10μM), a highly selective P2X1 antagonist, suramin (100μM) and pyridoxalphosphate-6-azophenyl-2′,4′-disulphonate (PPADS) (10μM)(non-selective P2 receptor antagonists), XAC (10μM), a non-selective adenosine receptor antagonist, and αβ-meATP (1μM), the desensitising agent. In some preparations, the endothelium was removed by gently rubbing the lumen of the artery with forceps. All the drugs were dissolved in distilled water. Results were compared by one-way ANOVA with Bonferroni’s post-hoc test or unpaired Student’s t-test. Data are presented as mean ± S.E.M. Contractions to ATP were expressed in g, while the relaxations were expressed as a percentage of the U46619-induced contraction.

Results: ATP (1µM-10mM) generated a non-cumulative vasoconstriction EC50 value (mean EC50 value was 0.5 mM (95% confidence interval (CI): 0.1-1.8 mM); Emax value was 9.5g, followed by a vasorelaxation EC50 value (mean EC50 value was 60 µM (95% confidence interval (CI): 5µM-0.8mM); Emax value was -6.5g; n=7, in pancreatic arteries. The contraction to ATP (1mM) was blocked by each of NF449, suramin and PPADS (P < 0.01, one-way ANOVA n=6-13). ATP-induced contraction was inhibited in the presence of αβ-meATP (P < 0.01, one-way ANOVA n=6-9). The relaxation to ATP was not affected by the non-selective P2 receptor antagonists or by the desensitising agent (P > 0.05, one-way ANOVA n=7), but it was inhibited significantly by XAC (P < 0.001, unpaired Student’s t-test, n=8-10). Removal of the endothelium had no effect on the contraction or the relaxation to ATP (P > 0.05, one-way ANOVA n=9-11).

Conclusions: These results suggest that, in porcine isolated pancreatic arteries, ATP induces a vasoconstriction mediated by P2X1 receptors followed by a vasorelaxation evoked by adenosine receptors. Data obtained in endothelium-denuded vessels showed that the P2X1 receptors and the adenosine receptors were present on the vascular smooth muscles of the porcine pancreatic arteries.

(1) Erlinge D & Burnstock G, Purinergic Signal 4: 1-20, 2008