Potassium channels are required for the absorption and secretion of fluids and electrolytes in epithelia. Calu-3 cells possess a secretory phenotype, and are a model human airway submucosal gland serous cell. Short-circuit current (I_sc) recordings from Calu-3 cells indicated that basal anion secretion was reduced by apical application of the K⁺ channel inhibitors bupivicaine, lidocaine, clofilium, and quinidine. Application of riluzole resulted in a large increase in I_sc, inhibited by apical application of either bupivicane or the cystic fibrosis transmembrane conductance regulator (CFTR) Cl^– channel blocker DPC. These results suggested that one or more members of the two-pore-domain K⁺ (K_2P) channel family could influence anion secretion. Using RT-PCR, we found that Calu-3 cells express mRNA transcripts for TASK-2 (KCNK5), TWIK-1 (KCNK1), TWIK-2 (KCNK6) and TREK-1 (KCNK2). TASK-2, TWIK-2 and TREK-1 protein were detected by Western blotting, while immunolocalization of polarized cells confirmed protein expression of TREK-1 and TWIK-2 at the plasma cell membrane. TASK-2 protein staining was localized to intracellular vesicles, located beneath the apical membrane. While the pro-secretory role of basolateral K⁺ channels is well established, we suggest that apically located K_2Pchannels, not previously described in airway epithelial cells, also play an important role in controlling the rate of transepithelial anion secretion.