Endothelial cells control the exchange on fluid and macromolecules between the intravascular- and the interstitial space by a transcellular and paracellular way. During inflammation a loss of paraendothelial barrier function increases the extravasation of macromolecules and water causing edemas and endothelila dysfunction. The vascular endothelial (VE) cadherin/b-,a-catenin complex is critical in regulation of paracellular barrier function. We show that the thrombin-induced decrease in paraendothelial barrier function is due to dynamin II dependent and caveolin-1-dependent endocytosis of the VE-cadherin/b-catenin complex. This process involves the epidermal-growth-factor-receptor-pathway-substrate 15 (Eps15) that interact via a specific binding sequence with caveolin-1 and the VE-cadherin/b-catenin complex followed by endocytosis. This mechanism is verified by using cavelin-1 deficient endothelioma cell lines and dominant negative mutant, dynamin II K44A, that are expressed in cultured endothelial cells via adenoviral and lentiviral vectors. The findings indicate a novel mechanism of regulation of endothelial barrier function by endocytosis.