Abstract. Chloride (Cl) conductances were studied in primary cultures of the bright part of rabbit distal convoluted tubule (DCTb) by the whole cell patch clamp technique. The bath solution (33C) contained (in mm): 140 NaCl, 1 CaCl2, 10 N-2-hydroxy-ethylpiperazine-N-2-ethanesulfonic acid (HEPES), pH 7.4 and the pipette solution 140 N-methyl-d-glucamine (NMDG)-Cl, 5 MgATP, 1 ethylene-glycol-bis(b-aminoethyl ether)-N,N,N,N-tetraacetic acid (EGTA), 10 HEPES, pH 7.4. We identified a Cl current activated by 105m forskolin, 103m 8-bromo adenosine 3,5-cyclic monophophosphate (8 Br-cAMP), 106m phorbol 12-myristate 13-acetate (PMA), 103m intracellular adenosine 3,5-cyclic monophophosphate (cAMP) and 107m calcitonin. The current-voltage relationship was linear and the relative ion selectivity was Br Cl I glutamate. This current was inhibited by 103m diphenylamine-2-carboxylate (DPC) and 104m 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) and was insensitive to 103m 4,4-diisothiocyanostilbene-2,2-disulfonic acid (DIDS). These characteristics are similar to those described for the cystic fibrosis transmembrane conductance regulator (CFTR) Cl conductance. In a few cases, forskolin and calcitonin induced an outwardly rectifying Cl current blocked by DIDS. To determine the exact location of the Cl conductance 6-methoxy-1-(3-sulfonatopropyl) quinolinium (SPQ) fluorescence experiments were carried out. Cultures seeded on collagen-coated permeable filters were loaded overnight with 5 mm SPQ and the emitted fluorescence analyzed by laser-scan cytometry. Cl removal from the apical solution induced a Cl efflux which was stimulated by 105m forskolin, 107 calcitonin and inhibited by 105m NPPB. In 140 mm NaBr, forskolin stimulated an apical Br influx through the Cl pathway. Forskolin and calcitonin had no effect on the basolateral Cl permeability. Thus in DCTb cultured cells, exposure to calcitonin activates a Cl conductance in the apical membrane through a cAMP-dependent mechanism.