Effects of purine and pyrimidine nucleotides on intracellular Ca2+ in human eosinophils: activation of purinergic P2Y receptors.

Extracellular adenosine 5'-triphosphate (ATP) increases human eosinophil intracellular Ca(2+) concentration; the mechanism of action is not fully known. ATP, a physiologic regulator, acts through 2 purinergic receptor types: cation channels (P2X) and G protein-coupled receptors (P2Y). This study is aimed at identifying the functional purinergic receptors in human eosinophils. The relative potency of ATP, uridine (UTP), cytidine (CTP), and inosine (ITP) 5'-triphosphates (P2Y agonists); 2-methylthio-ATP (P2Y(1) agonist); and 2 P2X agonists, alpha,beta-methylene-ATP and beta,gamma-methylene-ATP on intracellular Ca(2+) concentration was examined in Ca(2+)-sensitive Fura-2-labeled human eosinophils. For comparison, ATP effects were similarly studied in human neutrophils. P2X/P2Y mRNA expression in cells was examined by reverse transcription and PCR. The nucleotide potency order was UTP > or = ATP > ITP >>> 2-methylthio-ATP > alpha,beta-methylene-ATP = beta,gamma-methylene-ATP = CTP = 0 in eosinophils. Pertussis toxin (500 ng/mL) pretreatment abolished the effect of lower (10(-6) mol/L) but not higher (10(-5) mol/L) concentrations of ATP in eosinophils, whereas it attenuated the effects of 10(-4) mol/L ATP in neutrophils. The phospholipase C inhibitor U73122 (2 micromol/L) partially inhibited the effect of ATP in eosinophils but totally blocked it in neutrophils. Both cells constitutively express mRNA for P2X(1), P2X(4), P2X(5), P2Y(1), and P2Y(2), but not P2X(7), with much weaker expressions of P2X(4) and P2X(5) in neutrophils. Eosinophils cultured with the T(H)1 cytokine, IFN-gamma, expressed mRNA for P2X(7), a receptor linked to apoptosis. These results suggest that the P2 purinergic receptor signal transduction pathways in eosinophils and neutrophils are different and are mediated by more than 1 subtype of functional P2Y receptors.