Phosphatidylinositol-3-Kinase Regulates Mast Cell Ion Channel Activity

Abstract

Stimulation of the mast cell IgE-receptor (FcεRI) by antigen leads to stimulation of Ca²⁺ entry with subsequent mast cell degranulation and release of inflammatory mediators. Ca²⁺ further activates Ca²⁺-activated K⁺ channels, which in turn provide the electrical driving force for Ca²⁺ entry. Since phosphatidylinositol (PI)-3-kinase has previously been shown to be required for mast cell activation and degranulation, we explored, whether mast cell Ca²⁺ and Ca²⁺-activated K⁺ channels may be sensitive to PI3-kinase activity. Whole-cell patch clamp experiments and Fura-2 fluorescence measurements for determination of cytosolic Ca²⁺ concentration were performed in mouse bone marrow-derived mast cells either treated or untreated with the PI3-kinase inhibitors LY-294002 (10 µM) and wortmannin (100 nM). Antigen-stimulated Ca²⁺ entry but not Ca²⁺ release from the intracellular stores was dramatically reduced upon PI3-kinase inhibition. Ca²⁺ entry was further inhibited by TRPV blocker ruthenium red (10 µM). Ca²⁺ entry following readdition after Ca⁺-store depletion with thapsigargin was again decreased by LY-294002, pointing to inhibition of store-operated channels (SOCs). Moreover, inhibition of PI3-kinase abrogated IgE-stimulated, but not ionomycin-induced stimulation of Ca²⁺-activated K⁺ channels. These observations disclose PI3-kinase-dependent regulation of Ca²⁺ entry and Ca²⁺-activated K⁺-channels, which in turn participate in triggering mast cell degranulation.