Calcium handling and purinoceptor subtypes involved in ATP-induced contraction in rat small mesenteric arteries

1. The relationship between the stimulation of ATP receptors, the increase in intracellular free calcium concentration ([Ca2+](i); measured using the fluorescent indicator fura-2), contraction and the subtypes of purinoceptors involved were investigated in the small mesenteric artery of the rat. 2. In normal physiological solution, ATP (0.001-3 mM) caused concentration-dependent increases in both [Ca2+](i) and contraction. Both responses produced by ATP (1 mM) were inhibited by 50 % in the presence of nitrendipine (1 mu M) and were abolished in the presence of nitrendipine plus SK&F 96385 (30 mu M). 3.In Ca2+-free medium, ATP (3 mM) elicited a transient increase in both [Ca2+](i) and tension which were abolished by caffeine and decreased by 65% by thapsigargin (1 mu M). Moreover, ATP (1 and 3 mM) produced increases in the [H-3]D-myo-inositol 1,4,5-trisphosphate ([H-3]IP3) content of vessels in a concentration-dependent manner. 4. Treatment of the vessels with Bordetella pertussis toxin (PTX) inhibited contractions: to ATP linked to the influx of calcium through nitrendipine-sensitive mechanisms, but not those linked to the release of Ca2+ from intracellular stores nor the capacity of ATP in increasing IP3 content of the vessels. 5. The order of potency of ATP and its analogues in eliciting contraction was alpha,beta-methylene-ATP (alpha,beta-MeATP)>28-methylthio-BTP (2-MeSATP)>ATP=ADP. The response to ATP was inhibited by suramin. Reactive Blue 2 (up to 100 mu M) did not affect the contractile response to ATP. Pyridoxal-phosphate-8-azophenyl-2',4'-disulphonic acid 4-sodium (PPADS) and alpha,beta-MeATP abolished the response to low concentrations of ATP and reduced contractions elicited by high concentrations of ATP. 6. After blockade of P-2X-purinoceptors with PPADS, the order of potency of ATP and its analogues was 2-MeSATP>ATP=ADP. UTP produced concentration-dependent contractions which were not affected by suramin, Reactive Blue 2, PPADS or alpha,beta-MeATP, suggesting the presence of P-2U-purinoceptors. 7. The results suggest that low concentrations of ATP activate P-2X-purinoceptors and produce an influx of calcium through both voltage-dependent calcium channels sensitive to nitrendipine and through receptor-operated calcium channels sensitive to SK&F 96365. High concentrations of ATP activate P-2Y-purinoceptors which promote firstly a nitrendipine-sensitive calcium influx via a PTX-sensitive G protein and secondly a release of Ca2+ from an internal source via the production of IP3.