sq-23377 and taurolithocholic-acid-3-sulfate

Bile acids are known to induce Ca(2+) signals in pancreatic acinar cells. We have recently shown that phosphatidylinositol 3-kinase (PI3K) regulates changes in free cytosolic Ca(2+) concentration ([Ca(2+)](i)) elicited by CCK by inhibiting sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA). The present study sought to determine whether PI3K regulates bile acid-induced [Ca(2+)](i) responses. In pancreatic acinar cells, pharmacological inhibition of PI3K with LY-294002 or wortmannin inhibited [Ca(2+)](i) responses to taurolithocholic acid 3-sulfate (TLC-S) and taurochenodeoxycholate (TCDC). Furthermore, genetic deletion of the PI3K gamma-isoform also decreased [Ca(2+)](i) responses to bile acids. Depletion of CCK-sensitive intracellular Ca(2+) pools or application of caffeine inhibited bile acid-induced [Ca(2+)](i) signals, indicating that bile acids release Ca(2+) from agonist-sensitive endoplasmic reticulum (ER) stores via an inositol (1,4,5)-trisphosphate-dependent mechanism. PI3K inhibitors increased the amount of Ca(2+) in intracellular stores during the exposure of acinar cells to bile acids, suggesting that PI3K negatively regulates SERCA-dependent Ca(2+) reloading into the ER. Bile acids inhibited Ca(2+) reloading into ER in permeabilized acinar cells. This effect was augmented by phosphatidylinositol (3,4,5)-trisphosphate (PIP(3)), suggesting that both bile acids and PI3K act synergistically to inhibit SERCA. Furthermore, inhibition of PI3K by LY-294002 completely inhibited trypsinogen activation caused by the bile acid TLC-S. Our results indicate that PI3K and its product, PIP(3), facilitate bile acid-induced [Ca(2+)](i) responses in pancreatic acinar cells through inhibition of SERCA-dependent Ca(2+) reloading into the ER and that bile acid-induced trypsinogen activation is mediated by PI3K. The findings have important implications for the mechanism of acute pancreatitis since [Ca(2+)](i) increases and trypsinogen activation mediate key pathological processes in this disorder.

Topics: Androstadienes; Animals; Bile Acids and Salts; Calcium; Cells, Cultured; Cholecystokinin; Chromones; Enzyme Activation; Enzyme Inhibitors; Inositol 1,4,5-Trisphosphate Receptors; Ionomycin; Mice; Mice, Inbred C57BL; Mice, Knockout; Morpholines; Pancreas, Exocrine; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Taurochenodeoxycholic Acid; Taurolithocholic Acid; Thapsigargin; Wortmannin

In single liver cells, the D-myo-inositol 1,4,5-triphosphate (InsP3)-dependent agonists such as noradrenaline and angiotensin II evoke oscillations in intracellular calcium [Ca2+]i resulting mostly from the periodic release and reuptake of calcium from intracellular stores. In the present work, we have reexamined the effects of these agonists and investigated whether the natural bile acid taurolithocholic acid 3-sulfate (TLC-S), which permeabilizes the endoplasmic reticulum, could initiate oscillations of [Ca2+]i. Oscillations of [Ca2+]i were monitored with the Ca2(+)-dependent K+ permeability in whole-cell voltage-clamped guinea pig liver cells. Our results confirm the presence of two types of oscillations induced by hormones. They could be distinguished by their frequency periods. The fast (type I) had periods ranging from 5 to 12 s and the slow (type II) from 60 to 240 s. They have been respectively attributed to second messenger- and receptor-controlled oscillations, respectively. Our results also show that TLC-S, as noradrenaline and angiotensin II, induced the activation of this Ca(+)-dependent K+ current and was able to reproduce both types of oscillations. The bile acid effect was not blocked by intracellular perfusion of heparin known to inhibit both InsP3 binding and InsP3-evoked Ca2+ release in several tissues. In these conditions, TLC-S only evoked type I oscillations, suggesting that these fluctuations could originate from a mechanism that is independent of InsP3 and is an intrinsic property of internal Ca2+ stores.

Topics: Angiotensin II; Animals; Calcium; Cells, Cultured; Guinea Pigs; Heparin; Inositol 1,4,5-Trisphosphate; Ionomycin; Kinetics; Liver; Male; Norepinephrine; Potassium; Potassium Channels; Saponins; Taurolithocholic Acid; Time Factors