Extracellular ATP stimulates exocytosis via localized Ca2+ release from acidic stores in rat pancreatic β cells

Three different methods, membrane capacitance (C-m) measurement, amperometry and FM dye labeling were used to investigate the role of extracellular ATP in insulin secretion from rat pancreatic beta cells. We found that extracellular application of ATP mobilized intracellular Ca2+ stores and synchronously triggered vigorous exocytosis. No influence of ATP on the readily releasable pool of vesicles was observed, which argues against a direct modulation of the secretory machinery at a level downstream of Ca2+ elevation. The stimulatory effects of ATP were greatly reduced by intracellular perfusion of BAPTA but not EGTA, suggesting a close spatial association of fusion sites with intracellular Ca2+ releasing sites. ATP-induced Ca2+ transients and exocytosis were not blocked by thapsigargin (TG), by a ryanodine receptor antagonist or by dissipation of pH in acidic stores by monensin alone, but they were greatly attenuated by IP3 receptor inhibition as well as ionomycin plus monensin, suggesting involvement of IP3-sensitive acidic Ca2+ stores. Taken together, our data suggest that extracellular ATP triggers exocytosis by mobilizing spatially limited acidic Ca2+ stores through IP3 receptors. This mechanism may explain how insulin secretion from the pancreas is coordinated through diffusible ATP that is co-released with insulin.