Na+ transport by human placental brush border membranes: are there several mechanisms?

Na+ transport was evaluated in brush border membrane vesicles isolated from the human placental villous tissue. Na+ uptake was assayed by the rapid filtration technique in the presence and the absence of an uphill pH gradient. Amiloride strongly decreased Na+ uptake whether a pH gradient was present or not. In pH gradient conditions (pH 7.5 in and 9.0 out), 1 mM amiloride decreased the 10 mM Na+ uptake by 84%. In the absence of pH gradient (pH 7.5 in and out), Na+ uptake was lower but still sensitive to amiloride. The Lineweaver-Burk plot of Na+ uptake consistently showed a single kinetics. Increasing the pH gradient decreased Km values of the amiloride-sensitive Na+ uptake, leaving the Vmax unchanged. In the absence of a pH gradient, the amiloride sensitive Na+ transport was maximal at pH 7.5. Here again, a single kinetics was observed, and pH influenced exclusively the Km of Na+. Since ethylisopropylamiloride, the specific Na/H exchanger inhibitor mimicked the effects of amiloride, decreasing by 98% the 10 mM Na+ uptake, whereas benzamil, the Na+ channel blocker, had no effect, it was concluded that the amiloride sensitive Na+ uptake was predominantly or exclusively due to a Na+-H+ exchanger activity. K+ in trans-position significantly decreased the amiloride sensitive uptake. In contrast, the presence of the cation in cis-position had no effect. The amiloride resistant Na+ transport was neither influenced by pH, nor saturable. Incubation of the placental tissue with 100 microM or 1 mM dibutyryl cAMP, 0.1 or 1 microM phorbol myristate acetate, 10(-7) M insulin, 10(-10) M angiotensin II, or 10(-8) M human parathyroid hormone (PTH) did not influence Na+ transport by subsequently prepared brush border membranes. Finally, we failed to demonstrate any Na+-H+ exchange activity in the basal plasma membrane. These results indicate that (1) in the absence of cosubstrates such as phosphate and aminoacids, the Na+-H+ exchange is probably the unique mechanism of Na+ transport by the placental brush border membrane, (2) the placental isoform of the exchanger is not regulated by PTH, angiotensin, nor insulin and, therefore, is different from the isoform present in the renal brush border membrane, and (3) there is no exchanger activity in the basal plasma membrane.