Preparation of plasma membrane vesicles from the rat placenta at term and measurement of Na+ uptake.

Cytochemistry revealed that alkaline phosphatase was localized predominantly to the maternal facing plasma membrane of syncytiotrophoblast layer II in the haemotrichorial rat placenta at term. Plasma membrane vesicles prepared from term rat placenta by homogenization, treatment with MgCl2, and differential centrifugation were enriched 14-fold in alkaline phosphatase activity as compared to homogenate. These vesicles were mainly oriented right side out, as shown by a lack of effect of saponin treatment on alkaline phosphatase activity. Na+ uptake into the vesicles under equilibrium exchange conditions was significantly stimulated (P less than 0.01) nearly threefold in the presence of an outwardly directed H+ gradient as compared to no gradient. The stimulation by a H+ gradient was abolished in the presence of 0.5 mM amiloride. Measurements with the pH-sensitive dye BCECF in the absence of Na+ showed that 95.2 +/- 0.6 per cent of a 1.2 pH unit H+ gradient was dissipated from the vesicles in 2 min, but the remaining gradient was maintained for up to 15 min. These experiments therefore provide evidence that vesicles derived mainly from the maternal facing plasma membrane of syncytiotrophoblast layer II of the rat placenta possess a Na+/H+ exchanger.