Effects of ionic strength and chloride ion on activities of the glucose-6-phosphatase system: regulation of the biosynthetic activity of glucose-6-phosphatase by chloride ion inhibition/deinhibition

We investigated the transport pathways available for the uptake of vitamin C in the human placental choriocarcinoma cell line, JAR. These cells were found to possess the capacity to accumulate the vitamin when presented either in the oxidized form (dehydroascorbic acid) or in the reduced form (ascorbate). Dithiothreitol and 5,5'-dithiobis(2-nitrobenzoic acid) were used to maintain vitamin C as ascorbate and dehydroascorbic acid, respectively. The uptake of these two forms of vitamin C in JAR cells was found to occur by different mechanisms. The uptake of the dehydroascorbic acid was Na(+)-independent and was mediated by facilitative glucose transporters as evidenced from the inhibition of the uptake process by glucose. On the other hand, the uptake of ascorbate was Na(+)-dependent and was not sensitive to inhibition by glucose. Substitution of Na+ with other monovalent cations abolished the uptake of ascorbate completely. The uptake process was, however, not influenced by anions. Kinetic analysis indicated the presence of a single saturable transport system for ascorbate with a Michaelis-Menten constant of 22 +/- 1 microM. The dependence of the uptake rare of ascorbate on Na+ concentration exhibited sigmoidal kinetics, suggesting interaction of more than one Na+ ion with the transporter. The Hill coefficient for the Na+ interaction was 2, indicating that the Na(+)-dependent ascorbate transport is electrogenic. The Na(+)-dependent stimulation of ascorbate uptake was primarily due to an increase in the affinity of the transporter for ascorbate in the presence of Na+. It is concluded that the JAR placental trophoblast cell line expresses two different transport systems for vitamin C: one for the reduced form of the vitamin ascorbate; and the other for the oxidized form of the vitamin dehydroascorbic acid.