Bradykinin (BK) plays a key role in collecting duct functions. Using an established line of principal cells of the rabbit collecting duct (R.C. SV3), we examined the characteristics of the BK receptors in these cells. [<sup>3</sup>H]-BK bound specifically to R.C. SV3. Saturation binding analyses allowed K<sub>D</sub> (968 ± 232 p M) and B<sub>max</sub> values (356 ± 43 fmol/mg protein) to be calculated. Competitive displacement of [<sup>3</sup>H]-BK was observed with Hoe-140, a specific type 2 BK receptor (BKR-2) antagonist, but not with des arg<sup>9</sup>-BK, a BKR-1 agonist. The presence of BKR-2 was confirmed by the reverse-transcription polymerase chain reaction technique. BK stimulated cytosolic calcium and inositol phosphate formation in a dose-dependent manner (from 1 n M to 1 µ M). BK also inhibited the arginine vasopressin dependent increase of cyclic adenosine monophosphate. This effect could not be related to the production of prostaglandin E<sub>2</sub>. These results demonstrate the presence of high-affinity BKR-2 in the principal cells of the rabbit collecting duct that are linked to phospholipase C activity and are involved in arginine vasopressin related regulatory loops.