The mast cell lines rat basophilic leukemia (RBL) and mouse C57 cells respond to IgE/antigen complexes by degranulation. Treatment of these cells with 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), (10-100 nM) for 24-48 h enhanced IgE/antigen-induced exocytosis as monitored by release of hexosaminidase. A short term incubation with the hormone did not affect exocytosis, ruling out a rapid non genomic mechanism. The presence of vitamin D receptors, demonstrated by immunoblotting and the lack of effect of 24,25(OH)2D3 suggest a role for these receptors in the enhancing effect. 1,25(OH)2D3 also enhanced exocytosis induced by the calcium ionophore A23187 in the presence or absence of phorbol ester indicating modulation of events distal to signal transduction. 1,25(OH)2D3 enhanced exocytosis in the presence of cytochalasin D, indicating that the action of the hormone is not due to effects on microfilament structure. The results of this study suggest that 1,25(OH)2D3 may affect the allergic or pro-inflammatory potential of mast cells.