Serotonin transport and fluidity in plasma membrane vesicles: effect of hyperoxia

Plasma membrane vesicles were prepared from porcine pulmonary artery endothelial cells by a dextran-polyethylene glycol two-phase system. Specific carrier-mediated transport of 5-hydroxytryptamine (5-HT) into the vesicles was examined. Transport required a Na+ gradient (out greater than in) across the membrane, and accumulated 5-HT rapidly effluxed out of the vesicles when the ionophore gramicidin was added. Transport was inhibited by the antidepressant imipramine. 5-HT transport into plasma membrane vesicles appeared saturable and exhibited Michaelis-Menten kinetics (Km 7.4 microM, maximal velocity 217 pmol.min-1.mg membrane protein-1). A 24-h exposure to 95% O2 at 1 atmosphere absolute resulted in a 21% decrease (P less than 0.05) in specific 5-HT transport by plasma membrane vesicles. Hyperoxia also caused a significant (P less than 0.01) decrease in plasma membrane fluidity, as measured with the fluorescence probe 1,6-diphenyl-1,3,5-hexatriene. These results indicate that pulmonary artery endothelial cell plasma membrane vesicles provide a good model for studying 5-HT transport activity in vitro. Hyperoxia affects plasma membrane fluidity and 5-HT transport in pulmonary artery endothelial cells, suggesting a possible cause-and-effect relationship between the two.