Background: 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). 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 (P2Y 1 agonist); and 2 P2X agonists, α,β-methylene-ATP and β,γ-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. 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 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-γ, expressed mRNA for P2X 7 , 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.)