Dendritic cells (DCs) play a central role in immune regulation, ranging from tolerance induction to the induction of specific immune responses. DC functions are under regulation of cytosolic Ca²⁺. Ca²⁺ entry is sensitive to membrane potential and may depend on Cl^- channel activity. The present study employed whole-cell patch clamp experiments on mouse bone marrow-derived DCs to identify expression and regulation of Cl^- channels. Outwardly rectifying Cl^- currents were activated by elevation of cytosolic Ca²⁺, triggered either by ionomycin in the presence of extracellular Ca²⁺ or by IP₃-induced Ca²⁺ release from intracellular stores. The Ca²⁺-activated Cl^- channels (CaCCs) were activated by the chemokine CCL21 (75 ng/ml). The currents showed sensitivity to CaCC blockers niflumic acid (300 mM) and AO1 (20 mM). According to Fura2 fluorescence, the CaCC blockers blunted the store operated Ca²⁺ entry following depletion of intracellular Ca²⁺ stores with thapsigargin (1 mM) or upon stimulation with CCL21. The CaCC blockers further inhibited spontaneous, CXCL12 (50 ng/ml)- and CCL21 (25 ng/ml)-induced migration in transwell experiments and secretion of interleukin-2 (IL-2) and IL-6. According to RT-PCR and Western blot data, Anoctamin 6 is expressed in DCs and can contribute to CaCCs in DCs. Our data identify CaCCs in DCs as major players in Ca²⁺ homeostasis and Ca²⁺-regulated DC functions.