Differential Ca mobilization and mast cell degranulation by FcεRI- and GPCR-mediated signaling.

Mast cells play a primary role in allergic diseases. During an allergic reaction, mast cell activation is initiated by cross-linking IgE-FcεRI complex by multivalent antigen resulting in degranulation. Additionally, G protein-coupled receptors also induce degranulation upon activation. However, the spatio-temporal relationship between Ca mobilization and mast cell degranulation is not well understood. We investigated the relationship between oscillations in Ca level and mast cell degranulation upon stimulation in rat RBL-2H3 cells. Nile red and Fluo-4 were used as probes for monitoring histamine and intracellular Ca levels, respectively. Histamine release and Ca oscillations in real-time were monitored using total internal reflection fluorescence microscopy (TIRFM). Mast cell degranulation followed immediately after FcεRI and GPCR-mediated Ca increase. FcεRI-induced Ca increase was higher and more sustained than that induced by GPCRs. However, no significant difference in mast cell degranulation rates was observed. Although intracellular Ca release was both necessary and sufficient for mast cell degranulation, extracellular Ca influx enhanced the process. Furthermore, cytosolic Ca levels and mast cell degranulation were significantly decreased by downregulation of store-operated Ca entry (SOCE) via Orai1 knockdown, 2-aminoethyl diphenylborinate (2-APB) or tubastatin A (TSA) treatment. Collectively, this study has demonstrated the role of Ca signaling in regulating histamine degranulation.