Endothelial cell junctional integrity modulation by serotonin: an ultrastructural analysis

We have reported previously that exogenous serotonin (5-hydroxytryptamine, 5-HT) alters cultured bovine aortic endothelial cell (BAEC) structural integrity by modulating the assembly of stress fibers. In the present study a 5-HT stimulus-coupled change in BAEC junctional integrity was quantitated by determining the width and percentage of intercellular openings in a monolayer. BAEC treated with 5-HT at concentrations of 10(-9) M to 10(-3) M caused a significant dose-dependent decrease in interendothelial cell junctional openings compared to controls, with the greatest reduction induced at 10(-6) M (92% from control). Treatment of BAEC with histamine (10(-4) M) increased the junctional openings by 82% when compared to controls. This change could be prevented by either pretreatment of the monolayers with 5-HT or by adding 5-HT in conjunction with the histamine. To assess a direct interaction of 5-HT with actin filaments, cultured BAEC monolayers were extracted, treated with 5-HT, and processed for immunocytochemical localization of 5-HT using the Avidin-Biotin method. Electron microscopy revealed 5-HT antibody bound to actin filaments and dense in areas of filament intersection, which implies a role for internalized 5-HT in stimulating the assembly of an actin filament network. Collectively, these results suggest that 5-HT helps to regulate the endothelial junctional barrier by promoting actin filament formation and stability, which may in turn increase the junctional apposition between endothelial cells.