Role of the GLT-1 subtype of glutamate transporter in glutamate homeostasis: the GLT-1-preferring inhibitor WAY-855 produces marginal neurotoxicity in the rat hippocampus.
Glutamate is the major excitatory neurotransmitter in the central nervous system and is tightly regulated by cell surface transporters to avoid increases in concentration and associated neurotoxicity. Selective blockers of glutamate transporter subtypes are sparse and so knock-out animals and antisense techniques have been used to study their specific roles. Here we used WAY-855, a GLT-1-preferring blocker, to assess the role of GLT-1 in rat hippocampus. GLT-1 was the most abundant transporter in the hippocampus at the mRNA level. According to [(3)H]-l-glutamate uptake data, GLT-1 was responsible for approximately 80% of the GLAST-, GLT-1-, and EAAC1-mediated uptake that occurs within dissociated hippocampal tissue, yet when this transporter was preferentially blocked for 120 h with WAY-855 (100 microm), no significant neurotoxicity was observed in hippocampal slices. This is in stark contrast to results obtained with TBOA, a broad-spectrum transport blocker, which, at concentrations that caused a similar inhibition of glutamate uptake (10 and 30 microm), caused substantial neuronal death when exposed to the slices for 24 h or longer. Likewise, WAY-855, did not significantly exacerbate neurotoxicity associated with simulated ischemia, whereas TBOA did. Finally, intrahippocampal microinjection of WAY-855 (200 and 300 nmol) in vivo resulted in marginal damage compared with TBOA (20 and 200 nmol), which killed the majority of both CA1-4 pyramidal cells and dentate gyrus granule cells. These results indicate that selective inhibition of GLT-1 is insufficient to provoke glutamate build-up, leading to NMDA receptor-mediated neurotoxic effects, and suggest a prominent role of GLAST and/or EAAC1 in extracellular glutamate maintenance.
Selkirk
Julie V
JV
Neurosciences Department, Neurocrine Biosciences Inc., 12970 El Camino Real, San Diego, CA 92130, USA. jselkirk@neurocrine.com
Nottebaum
Lisa M
LM
Vana
Alicia M
AM
Verge
Gail M
GM
Mackay
Kenneth B
KB
Stiefel
Theodore H
TH
Naeve
Greg S
GS
Pomeroy
Jordan E
JE
Petroski
Robert E
RE
Moyer
John
J
Dunlop
John
J
Foster
Alan C
AC
D027322:Amino Acid Transport System X-AG / Q000502:physiology; D000818:Animals; D000831:Animals, Newborn; D001224:Aspartic Acid / Q000494:pharmacology; D015153:Blotting, Western / Q000379:methods; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000633:toxicity*; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D027342:Excitatory Amino Acid Transporter 2 / Q000235:genetics / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000502:physiology; D018698:Glutamic Acid / Q000378:metabolism*; D006536:Heptanes / Q000633:toxicity*; D006575:Heterocyclic Compounds, 3-Ring / Q000633:toxicity*; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects*; D006706:Homeostasis / Q000187:drug effects*; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D017403:In Situ Hybridization / Q000379:methods; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D016202:N-Methylaspartate / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D018408:Patch-Clamp Techniques / Q000379:methods; D051381:Rats; D013997:Time Factors; D014316:Tritium / Q000378:metabolism
C487627:3-aminotricyclo(2.2.1.02.6)heptane-1,3-dicarboxylic acid; D027322:Amino Acid Transport System X-AG; D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D018691:Excitatory Amino Acid Antagonists; D027342:Excitatory Amino Acid Transporter 2; D006536:Heptanes; D006575:Heterocyclic Compounds, 3-Ring; C120673:benzyloxyaspartate; D014316:Tritium; D001224:Aspartic Acid; D018698:Glutamic Acid; D016202:N-Methylaspartate; D016291:Dizocilpine Maleate