Inhibition of glutamate uptake induces progressive accumulation of extracellular glutamate and neuronal damage in rat cortical cultures.
It is known that neurons exposed to high concentrations of glutamate degenerate and die. The clearance of this amino acid from the extracellular space depends on their active transport by Na(+)-dependent high-affinity carriers. In the present study we tested whether inhibition of glutamate transport in mixed glial/neuronal cortical cultures induces accumulation of extracellular glutamate and whether such increase results in cell damage. Three inhibitors of glutamate transport were used: L-trans-pyrrolidine-2,4-dicarboxylate (PDC), DL-threo-beta-hydroxyaspartate (THA), and dihydrokainate (DHK). Cell damage was assessed by light microscopy observations, reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, and leakage of lactate dehydrogenase. PDC induced a significant concentration- and time-dependent neuronal damage, whereas pure glial cultures were not affected. A good correlation was found between this damage and elevations of glutamate concentration in the medium. These effects of PDC were similar in glutamine-free medium and in medium supplemented with glutamine. THA induced identical cell damage and elevations of extracellular glutamate to those produced by PDC, while DHK did not affect at all any of these parameters. PDC- and THA-induced toxicity was protected by the N-methyl-D-aspartate receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo-(a,d)cyclohepten-5,10-imine maleate but not by the non-N-methyl-D-aspartate receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)quinoxaline.
Velasco
I
I
Departamento de Neurociencias, Universidad Nacional Autónoma de México, México.
Tapia
R
R
Massieu
L
L
D000818:Animals; D000900:Anti-Bacterial Agents / Q000633:toxicity; D001224:Aspartic Acid / Q000031:analogs & derivatives / Q000633:toxicity; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D003998:Dicarboxylic Acids / Q000633:toxicity; D018691:Excitatory Amino Acid Antagonists / Q000633:toxicity*; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D007608:Kainic Acid / Q000031:analogs & derivatives / Q000633:toxicity; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009410:Nerve Degeneration / Q000187:drug effects; D009474:Neurons / Q000187:drug effects* / Q000648:ultrastructure; D014179:Neurotransmitter Uptake Inhibitors / Q000633:toxicity; D011759:Pyrrolidines / Q000633:toxicity; D051381:Rats; D017208:Rats, Wistar; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors
D000900:Anti-Bacterial Agents; D003998:Dicarboxylic Acids; D018691:Excitatory Amino Acid Antagonists; D014179:Neurotransmitter Uptake Inhibitors; D011759:Pyrrolidines; D016194:Receptors, N-Methyl-D-Aspartate; C011274:3-hydroxyaspartic acid; D001224:Aspartic Acid; D018698:Glutamic Acid; C020889:dihydrokainic acid; C083486:pyrrolidine-2,4-dicarboxylic acid; D007770:L-Lactate Dehydrogenase; D007608:Kainic Acid