Bacterial LPS is a potent proinflammatory molecule. In the lungs, LPS induces alterations in surfactant pool sizes and phospholipid (PL) contents, although direct actions of LPS on the alveolar type II cells (AT II) are not yet clear. For this reason, we studied short- and long-term effects of LPS on basal and agonist-stimulated secretory responses of rat AT II by using Ca2+microfluorimetry, a microtiter plate-based exocytosis assay, by quantitating PL and3H-labeled choline released into cell supernatants and by using quantitative PCR and Western blot analysis. Long term, but not short term, exposures to LPS led to prolonged ATP-induced Ca2+signals and an increased rate in vesicle fusions with an augmented release of surfactant PL. Most notably, the stimulatory effect of LPS was ATP-dependent and may be mediated by the upregulation of the purinergic receptor subtype P2Y2. Western blot analysis confirmed higher levels of P2Y2, and suramin, a P2Y receptor antagonist, was more effective in LPS-treated cells. From these observations, we conclude that LPS, probably via Toll-like receptor-4, induces a time-dependent increase in P2Y2receptors, which, by yet unknown mechanisms, leads to prolonged agonist-induced Ca2+responses that trigger a higher activity in vesicle fusion and secretion. We further conclude that chronic exposure to endotoxin sensitizes AT II to increase the extracellular surfactant pool, which aids in the pulmonary host defense mechanisms.