Effects of purine nucleotides and nucleoside on cytosolic calcium levels in rat tracheal smooth muscle cells.

Abstract

Extracellular adenosine triphosphate (ATP) has a range of effects on a wide variety of cells through the activation of specific purinoceptors. The aim of this study was to establish whether P2 purinoceptors are present on airway smooth muscle cells. Experiments were conducted on cultured rat tracheal smooth-muscle cells (first through third passage). Intracellular Ca2+ ([Ca2+]i) was measured using Fura-2 and dual-excitation wavelength microfluorometry. The effects of ATP, adenosine diphosphate (ADP), uridine triphosphate (UTP), and adenosine (ADO) were measured in concentrations from 10(-6) to 10(-3) M. At a concentration of 10(-4) M, the peak [Ca2+]i was 502 +/- 92 nM for ATP and 543 +/- 76 nM for UTP (mean +/- standard error of the mean). ADO had no significant effect on Ca2+ release. Peak [Ca2+]i induced by ATP was not dependent on extracellular Ca2+ but was blocked by U-73122, an inhibitor of phospholipase C. Pretreatment with adenosine deaminase and desensitization with alphabeta-MeATP had no effect on ATP-induced Ca2+ release. The effects of ATP (10(-4) M) on peak [Ca2+]i were potentiated by the presence of ADO 10(-5) M (969 +/- 257 nM; P < 0.05). The presence of XAC, a blocker of A1 and A2 ADO receptors did not prevent this effect. In the presence of XAC, ADO 10(-6) M potentiated the effects of ATP (peak [Ca2+]i: 1,300 +/- 229 nM). The addition of 1433U83, a blocker of A3 ADO receptors, blocked the synergistic effect of ADO 10(-6) M on ATP. These data show that P2 purinoceptors, most likely of the P2U subtype, are present on airway smooth muscle cells and that the newly discovered A3 ADO receptor appears to be also present.