The CFPAC-1 cell line was derived from a pancreas duct carcinoma from a patient suffering from ΔF 508 cystic fibrosis (CF). This cell therefore should be well suited to study the Cl– transport defect in CF. We used slow whole cell (SWC) and cell-attached nystatin (CAN) patch clamp techniques to characterize these cells and we confirm that cAMP fails to induce Cl– currents. Ca2+-mobilizing agents such as neurotensin (NT), ATP, and carbachol all induce a depolarization together with an increase in conductance. This was revealed in SWC and in apical membrane CAN experiments. The half-maximal conductance effect of NT was at 10–9 mol/l, that of ATP at 10–6 mol/l. For carbachol, only 10–4 mol/l was examined. In CAN experiments, no clear current events were detectable in apical membrane patches, suggesting that the Cl– channel responsible for the measured increase in conductance has a conductance too small to be resolved as a single channel event. Hypotonic cell swelling also induced a biphasic voltage reponse with a predominant increase in Cl– conductance. This effect was detectable in SWC and CAN recordings for 240 mosm/1 NaCl and was further enhanced at 160 mosm/1 NaCl. The conductance sequence for this and the ATP-induced pathway was F = Br– = Cl–. Both pathways were inhibited equally well by 10–3 mol/l DIDS, and they were not additive with respect to their conductance effects. These data indicate that cell swelling as well as Ca2+ mobilization activates a Cl– conductance which consists of the synchronous activity of nondetectable single Cl– currents. These currents appear to be the same as those recently described in HT29 cells.