In polycystic kidney disease (PKD), cyst growth results from both renal epithelial cell proliferation and fluid accumulation within the cyst lumen. Cystic fibrosis transmembrane conductance regulator (CFTR) chloride channels located in the apical membrane of renal epithelial cells play an important role in fluid accumulation within cysts (Sullivan et al. 1998). However, the role of other types of epithelial Cl^–channels is unclear.In this study, we investigated the role of calcium-activated Cl^–channels (CaCCs) in the growth of renal cysts using MDCK cysts as a model system. We used the Ussing chamber technique to study transepithelial ion transport and grew cysts as previously described (Li & Sheppard, 2002). To activate CaCCs, we used ionomycin (1 µM) and UTP (100 µM). As a control, we studied the CFTR Cl^–channel.Addition of ionomycin to the apical membrane activated a transient increase in short-circuit current (I_sc) of similar magnitude and time-course to that stimulated by forskolin (10 µM) (ionomycin: 11.9±1.7 µA/cm²; mean±SEM; n = 8; forskolin: 12.1±1.2 µA/cm²;n = 18; P > 0.05; Student′s unpaired t-test). However, the currents activated by ionomycin and forskolin had distinct pharmacology: pretreatment of epithelia with DIDS (200 µM) or niflumic acid (NFA, 100 µM) abolished the ionomycin-activated I_sc (DIDS: 1.5±0.9 µA/cm²;NFA:0.4±0.5 µA/cm²;n = 6; P < 0.05 vs. ionomycin data), but not the forskolin-stimulated I_sc (DIDS: 16.7±1.7 µA/cm²;n = 6; NFA: 10.0±2.2 µA/cm²;n = 8; P > 0.05 vs. forskolin data). These data suggest that MDCK epithelia likely possess both CFTR and CaCC.Consistent with previous results, when cultured in the presence of media containing forskolin (10 µM), MDCK cells formed cysts in collagen gels. Similarly, ionomycin (1 µM) and UTP (100 µM) stimulated cyst formation. However, after six days, the volume and number of cysts observed in the presence of either ionomycin or UTP were reduced compared to those observed in the presence of forskolin (ionomycin: volume, 0.0003±0.00008 mm³;n = 20; number, 24±5, n = 4; forskolin: volume, 0.002±0.0003 mm³;n = 186; number, 44±4; n = 23; P < 0.05 for forskolin vs. ionomycin data). Interestingly, when cysts grown in the presence of forskolin for six days were treated with ionomycin in the continuous presence of forskolin for a further six days, cyst growth was inhibited dramatically (n = 9). Moreover, this effect of ionomycin was not relieved by DIDS (200 µM; n = 10). We interpret our data to suggest that CaCC only weakly stimulate cyst formation and growth. However, the data also suggest that increasing the intracellular calcium concentration might inhibit cAMP-stimulated cyst growth.