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

Background: Extracellular adenosine 5′-triphosphate (ATP) increases human eosinophil intracellular Ca2+ 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). Objective: This study is aimed at identifying the functional purinergic receptors in human eosinophils. Methods: The relative potency of ATP, uridine (UTP), cytidine (CTP), and inosine (ITP) 5′-triphosphates (P2Y agonists); 2-methylthio-ATP (P2Y1 agonist); and 2 P2X agonists, α,β-methylene-ATP and β,γ-methylene-ATP on intracellular Ca2+ concentration was examined in Ca2+-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. Results: The nucleotide potency order was UTP ≥ ATP > ITP >>> 2-methylthio-ATP > α,β-methylene-ATP = β,γ-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 μmol/L) partially inhibited the effect of ATP in eosinophils but totally blocked it in neutrophils. Both cells constitutively express mRNA for P2X1, P2X4, P2X5, P2Y1, and P2Y2, but not P2X7, with much weaker expressions of P2X4 and P2X5 in neutrophils. Eosinophils cultured with the TH1 cytokine, IFN-γ, expressed mRNA for P2X7, a receptor linked to apoptosis. Conclusions: 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. (J Allergy Clin Immunol 2001;107:849-55.)