ABSTRACT
Ascorbic acid is frequently added in the incubation medium to prevent oxidation of dopamine (DA) during uptake assays. However, a preliminary study showed that the presence of ascorbic acid induced a decrease of DA uptake after prolonged incubation. The purpose of this study was to determine the mechanism underlying ascorbic acid-induced alterations of DA uptake in rat striatal synaptosomes. In this context, the effects of physiological concentrations of ascorbic acid (100-500 microM) on DA uptake and Na+/K+ ATPase activity (which is essential for DA transporter function) were assessed in synaptosomes before and after incubation at 37 degrees C. The capacity of synaptosomes to take up DA was significantly decreased after incubation owing to a reduction in DA transporters (but with no modification of their affinity for DA). This partial inhibition was associated with a decrease of Na+/K+ ATPase activity, a production of thiobarbituric acid reactive substances (TBARS) and malonaldehyde (MDA), and a loss of sulfhydryl group content. Addition of Trolox C to the medium prevented the reduction of DA uptake, the inhibition of Na+/K+ ATPase activity, the decrease in sulfhydryl group content and the production of TBARS and MDA. These results suggest that ascorbic acid in the presence of contaminant ferrous ions induced a decrease in functional DA transporters, probably through a lipid peroxidation process involving oxidation of sulfhydryl groups and at least in part through a decrease of Na+/K+ ATPase activity.
