Abstract
To establish an assay system for evaluation of the uptake and reversed transport of glutamate, we examined the effects of Na+-concentration and pharmacological agents on the extracellular glutamate concentration ([Glu]o) in rat cortical synaptosomes in vitro. There was a decrease and increase of the [Glu]o at high and low Na+ concentrations, respectively, in a Ca2+-free medium. The changes in [Glu]o in both directions were temperature-sensitive, and reversed at around 30 mM of Na+. Dihydrokainate (DHK), a non-transportable inhibitor selective for glial glutamate transporter GLT-1, suppressed the decrease in [Glu]o, and the reversal of [Glu]o change was shifted to about 60 mM Na+. There was no change in the maximum [Glu]o at total Na+ substitution. Further pharmacological analysis revealed that D-aspartate and DL-threo-β-hydroxy-aspartate (THA), transportable substrates of glutamate transporters, increased the [Glu]o in standard media. In contrast, β-phenylglutamic acid, a structural analogue of glutamate, suppressed both the decrease in [Glu]o in standard medium and the increase in [Glu]o in low Na+ medium. It is, thus, concluded that both the direction and the amount of [Glu]o changes are determined by a balance of the uptake and reversed transport of glutamate, and that this assay system is suitable for evaluation of the effect of this on glutamate transporters.
