Coupling of vasopressin‐induced intracellular Ca²⁺ mobilization and apical exocytosis in perfused rat kidney collecting duct

<jats:p>Arginine vasopressin (AVP) regulates the osmotic water permeability of the kidney collecting duct by inducing exocytotic insertion of aquaporin‐2 into apical membrane. The coupling between AVP‐induced intracellular Ca<jats:sup>2+</jats:sup> mobilization and apical exocytosis was investigated in isolated perfused rat inner medullary collecting duct (IMCD) segments using confocal fluorescence microscopy. Changes of [Ca<jats:sup>2+</jats:sup>]<jats:sub>i</jats:sub> in IMCD cells were measured with fluo‐4. A novel confocal imaging technique using a styryl dye, FM1‐43, was developed to monitor real‐time exocytosis induced by arginine vasopressin. AVP (0.1 n<jats:sc>m</jats:sc>) triggered a rapid increase of [Ca<jats:sup>2+</jats:sup>]<jats:sub>i</jats:sub> in IMCD cells, followed by sustained oscillations. Ratiometric measurement of [Ca<jats:sup>2+</jats:sup>]<jats:sub>i</jats:sub> confirmed that the observed [Ca<jats:sup>2+</jats:sup>]<jats:sub>i</jats:sub> oscillation was a primary event and was not secondary to changes in cell volume. The frequencies of [Ca<jats:sup>2+</jats:sup>]<jats:sub>i</jats:sub> oscillations in each IMCD cell were independent and time variant. 1‐Deamino‐8‐<jats:sc>d</jats:sc>‐arginine vasopressin (a V<jats:sub>2</jats:sub> receptor agonist, 0.1 n<jats:sc>m</jats:sc>) simulated the effects of AVP by triggering [Ca<jats:sup>2+</jats:sup>]<jats:sub>i</jats:sub> oscillations. In the absence of extracellular Ca<jats:sup>2+</jats:sup>, ryanodine (0.1 m<jats:sc>m</jats:sc>) inhibited AVP‐induced Ca<jats:sup>2+</jats:sup> mobilization. AVP (0.1 n<jats:sc>m</jats:sc>) triggered accumulative apical exocytosis in IMCD cells within 20 s after application. Pre‐incubating the IMCD with an intracellular Ca<jats:sup>2+</jats:sup> chelator, BAPTA, prevented AVP‐induced intracellular Ca<jats:sup>2+</jats:sup> mobilization, apical exocytosis, and increase of osmotic water permeability. These results indicate that AVP, via the V<jats:sub>2</jats:sub> receptor, triggers a calcium signalling cascade observed as [Ca<jats:sup>2+</jats:sup>]<jats:sub>i</jats:sub> oscillations in the IMCD and that intracellular Ca<jats:sup>2+</jats:sup> mobilization is required for exocytotic insertion of aquaporin‐2.</jats:p>