Trafficking of the water channel aquaporin-2 (AQP2) is classically regulated by the action of the antidiuretic hormone arginine-vasopressin (AVP). The AVP signal is followed by the phosphorylation and translocation of AQP2 to the apical plasma membrane of the collecting duct (CD) principal cell. So far the regulation of AQP2 has been studied in whole animals and different cell models. In this study we made use of the isolated perfused rat kidney (IPK). IPKs were perfused for a period of at least 90min and the urine was collected every 8–10min. Compared to kidneys perfused with control solution (without AVP) the osmolality significantly increased when AVP was added to the perfusate (25 or 250pM), thus the free water reabsorption was significantly enhanced under the influence of AVP. This demonstrates that the IPK responds to the AVP treatment. Analysis of AQP2 localization with an anti-AQP2 antibody (H27) by immuno-fluorescence showed after AVP (25pM) treatment an enrichment at the apical plasma membrane. Using an antibody directed against the protein kinase A phosphorylation site at position S256 showed that AQP2 phosphorylated at S256 after AVP treatment (25pM) was localized at the apical membrane. Interestingly the treatment with a higher concentration of AVP (250pM) leads in addition also to a basolateral staining of AQP2 with the H27 antibody. However under the same treatment S256 phsphorylated AQP2 is exclusively localized at the apical membrane. In conclusion we could show that the IPK is an adequate model to analyze the regulation of AQP2 in the collecting duct cells.