Astrocytes are thought to control extracellular glutamate concentrations ([Glu]o) in the brain, thereby protecting neurons from excitotoxic injury. We investigated the effects of metabotropic glutamate receptor (mGluR) agonists on glutamate transport and [Glu]o in primary hippocampal astrocytic cultures. Acute or chronic exposure of astrocytes to the mGluR agonist trans-1-aminocyclopentane-1,3-dicarboxylic acid (trans-ACPD) or its active isomer 1S,3R-ACPD reduced [Glu]o in a time- and dose-dependent manner (44.5 +/- 3.6% reductions of [Glu]o in astrocytes from P0-P10 rats and 65.9 +/- 4.1% from rats P20 by 100 microM 1S,3R-ACPD, EC50 approximately 5 microM). 1S,3R-ACPD effects developed slowly (median effective at approximately 60 min) and persisted for several hours after agonist removal. ACPD-pretreated astrocytes established lower steady-state [Glu]o levels. ACPD effects persisted in the presence of the glutamate uptake inhibitors D,L-threo-beta-hydroxyaspartate (THA) and L-trans-pyrrolidine-2,4-dicarboxylate (PDC) but were impaired by disruption of the transmembrane Na+, K+, or H+ gradients. In addition, 1S,3R-ACPD had no effects on intracellular glutamate content and did not directly block glutamate transport. Furthermore, ACPD effects could be mimicked by glutamate per se and several other compounds presumed to be mGluR agonists, although (S)-3,5-dihydroxyphenylglycine (DHPG), (2S,2R,3R)-2-(2,3-dicarboxycyclopropyl)glycine (DCG-IV), and L-(+)-2-amino-4-phosphonobutyric acid (L-AP4) were without effect. These data suggest that glutamate and certain mGluR agonists may regulate [Glu]o by modulating the transmembrane equilibrium of glutamate transport, especially by attenuating glutamate release.