title: Measurement of glutamate uptake and reversed transport by rat synaptosome transporters.
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 Ca(2+)-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-beta-hydroxy-aspartate (THA), transportable substrates of glutamate transporters, increased the [Glu](o) in standard media. In contrast, beta-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.
authors.affiliation: Discovery and Pharmacology Research Laboratories, Tanabe Seiyaku Co., Ltd., Toda, Saitama, Japan. bon@tanabe.co.jp
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nishida
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Iwata
authors.forename: Yukitsuka
authors.initials: Y
authors.lastname: Kudo
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Yuzo
authors.initials: Y
authors.lastname: Matsuoka
authors.forename: Yoshikatsu
authors.initials: Y
authors.lastname: Kanai
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Endou
mesh_terms: D000818:Animals; D001418:Baclofen / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D027342:Excitatory Amino Acid Transporter 2 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism*; D008297:Male; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism*
chemical_list: D027342:Excitatory Amino Acid Transporter 2; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins; D018698:Glutamic Acid; D001418:Baclofen