Activation of two sites by adenosine receptor agonists to cause relaxation in rat isolated mesenteric artery

Abstract

1. In this study we have characterized the receptor(s) in the rat mesenteric artery mediating relaxant responses to adenosine and a number of adenosine analogues, N⁶ -R-phenylisopropyladenosine (R-PIA), N⁶-cyclopentyladenosine (CPA), N⁶-(3-iodo-benzyl)-adenosine-5′-N-methyluronamide (IB-MECA) and 5′-N-ethylcarboxamidoadenosine (NECA), by use of the non-selective antagonist 8-sulphophenyltheophylline (8-SPT) and the A_2A selective ligands 2-[p-(2-carbonylethyl)-phenylethylamino]-5′-N-ethylcarboxamidoadenosine (CGS 21680) and 4-(2-[7-amino-2-(2-furyl)[1,2,4]-triazolo[2,3-a][1,3,5]-triazin-5-ylamino]ethyl) phenol (ZM 241385). We have also studied the effects of endothelial removal and uptake inhibition by nitrobenzylthioinosine (NBTI) and the effects of the A₃ receptor antagonist 1,3-dipropyl-8-(4-acrylate)phenylxanthine (BWA1433).

2. Adenosine, NECA, CPA and R-PIA all elicited relaxant responses in tissues precontracted with phenylephrine (1 μM) with the following potency order: NECA>R-PIA>adenosine=CPA. However, E/[A] curves to NECA were biphasic. CGS 21680 was inactive at concentrations up to 30 μM and IB-MECA elicited relaxant responses which were resistant to blockade by 8-SPT and BWA1433 (100 μM).

3. Removal of the endothelium produced a small but significant decrease in the asymptote of the high potency phase of E/[A] curves to NECA with no change in p[A]₅₀. E/[A] curves to adenosine were not altered by removal of the endothelium. However, there were small rightward shifts of E/[A] curves to CPA and R-PIA in the absence of endothelium.

4. Inhibition of uptake by NBTI (1 μM) had no effect on E/[A] curves to NECA, CPA or R-PIA, but E/[A] curves to adenosine were significantly left-shifted in the presence of NBTI.

5. 8-SPT (10–100 μM) caused significant rightward shifts of the high potency phase of the E/[A] curves to NECA (pA₂=5.63±0.26). The second phase of the concentration-response curve to NECA appeared to be resistant to blockade by 8-SPT, as were E/[A] curves for adenosine, CPA or R-PIA. However, in the presence of NBTI (1 μM), 8-SPT (100 μM) gave significant rightward shifts of E/[A] curves to adenosine.

6. ZM 241385 (0.1–1 μM) produced significant rightward shifts of the high potency phase of NECA E/[A] curves (pA₂=7.65±0.25 in the presence and 7.20±0.12 in the absence of endothelium), while curves to R-PIA were not significantly shifted by 1 μM ZM 241385. In the presence of NBTI E/[A] curves to adenosine were significantly rightward shifted by ZM 241385 (0.1 μM, pA₂=7.50±0.16).

7. In conclusion, the results suggest activation of A_2B receptors located primarily on the smooth muscle by low concentrations of NECA and by adenosine under conditions of uptake blockade, and of another, as yet undefined site which may be intracellular, by higher concentrations of NECA, by CPA, R-PIA and adenosine under conditions where uptake is operational.

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