Regulation of aquaporin-2 trafficking by vasopressin in the renal collecting duct -: Roles of ryanodine-sensitive Ca2+ stores and calmodulin

In the renal collecting duct, vasopressin increases osmotic water permeability (P-f) by triggering trafficking of aquaporin-2 vesicles to the apical plasma membrane. We investigated the role of vasopressin-induced intracellular Ca2+ mobilization in this process. In isolated inner medullary collecting ducts (IMCDs), vasopressin (0.1 nM) and 8-(4-chlorophenylthio)-cAMP (0.1 mM) elicited marked increases in [Ca2+](i) (fluo-4), Vasopressin-induced Ca2+ mobilization was completely blocked by preloading with the Ca2+ chelator BAPTA, In parallel experiments, BAPTA completely blocked the vasopressin-induced increase in P-f without affecting adenosine 3',5'-cyclic monophosphate (cAMP) production. Previously, we demonstrated the lack of activation of the phosphoinositide-signaIing pathway by vasopressin in IMCD, suggesting an inositol 1,4,5-trisphosphate-independent mechanism of Ca2+ release. Evidence for expression of the type I ryanodine receptor (RyR1) in IMCD was obtained by immunofluorescence, immunoblotting, and reverse transcription-polymerase chain reaction, Ryanodine (100 muM), a ryanodine receptor antagonist, blocked the arginine vasopressin-mediated increase in P-f and blocked vasopressin-stimulated redistribution of aquaporin-2 to the plasma membrane domain in primary cultures of IMCD cells, as assessed by immunofluorescence immunocytochemistry. Calmodulin inhibitors (W7 and trifluoperazine) blocked the P-f response to vasopressin and the vasopressin-stimulated redistribution of aquaporin-2. The results suggest that Ca2+ release from ryanodine-sensitive stores plays an essential role in vasopressin-mediated aquaporin-2 trafficking via a calmodulin-dependent mechanism.