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Glutamate transporters

Glutamate neurotransmitter transporters of CNS GLAST-1 (EAAT-1), GLT-1 (EAAT2) and EAAC-1 (EAAT3) in excitatory neurotransmission and neurotoxicity.

Discovery of glutamate neurotransmitter transporters of CNS GLAST-1 (EAAT-1)
L-Glutamate is the neurotransmitter of excitatory synapses in CNS. The glu concentration in the synaptic cleft is fine tuned by the orchestra of neurotransmitter glutamate release, glutamate receptors and Na+-dependent, high affinity (Km < 10 µM) glutamate uptake systems to prevent neuronal excitotoxicity. We have discovered the first CNS L-glutamate transporter GLAST1 (Glu-Asp-Transporter). Subsequently four more completed the family of Glu transporters: GLT1 (Pines et al.), EAAC1 (Kanai et al.), EAAT4 and EAAT5 (Arizza et al.). GLAST1 and GLT1 are expressed only in astrocytes, EAAC1 in neurons, kidney and jejunum. Different from the 12-TMD topology of the other neurotransmitter transporter topology, the EAAT- family is characterized by a six TMD N-terminus followed by a four ß-barrel C-terminal domain, the transport active site of GLAST1 and presumably the other members of the glutamate transporter family. We derived this topology by "N-glycosylation scanning" (figure 2). Despite extensive biochemical and electrophysiological studies of these most important uptake systems, their individual role in vivo remains elusive. Therefore we have generated and characterized so far the GLAST1 and EAAC1 single and GLAST1-EAAC1 deficient mouse models. Surprisingly these null mutants lacked the expected epileptic and neurotoxic phenotype. We have now focused on establishing the GLT1 deficient mouse model, because electrophysiological studies provided evidence that GLT1 is the "work horse", but GLAST1 and EAAC1 are the modulators of the glutamatergic synapses. The conventional and conditional glt1-/- mouse models will be of great importance in unravelling the basis of the molecular processes leading to excitotoxicity under experimental conditions and in human neurological diseases, particularly in epilepsy.

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Figure 1: a) Affinity purification of Glast-1. b) Derived sequence of Glast-1 and related bacterial dicaboxylic acid transporters


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Figure 2: The proposed topology of Glast-1 in plasmamembrane


References
  1. Pietro Peghini, Julia Janzen and Wilhelm Stoffel 1997. Glutamate Transporter EAAC-1-Deficient Mice Develop Dicarboxylic Aminoaciduria and Behavioral Abnormalities but no Neurodegeneration EMBO J. 16, 3822-3832
  2. Conradt, M. and Stoffel, W. 1997. Inhibition of High Affinity Brain Glutamate Transporter GLAST-1 via direct Phosphorylation J. Neurochem. 68, 1244-1251
  3. Wahle, S. and Stoffel, W. 1996. Membrane Topology of the High Affinity L-Glutamate Transporter (GLAST-1) of the Central Nervous System. J.Cell Biol. 135, 1867-1877.
  4. Stoffel, W., Sasse, J., Düker, M., Müller, R., Hofmann, K., Fink, Th., Lichter, P. (1996) Human High Affinity, Na+-Dependent L-Glutamate/L-Aspartate Transporter GLAST-1 (EAAT-1): Gene Structure and Localization to Chromosome 5p11-p12 FEBS Letters 386, 189-193
  5. M. Conradt, W. Stoffel (1995) Functional analysis of the high-affinity, Na+-dependent glutamate transporter GLAST-1 by site-directed mutagenesis J. Biol. Chemistry, 270, No. 42, 25207-25212
  6. W. Stoffel, R. Blau Rat and Human Glutamate Transporter GLAST1. Stable Heterologous Expression, Biochemical and Functional Characterization Biol. Chem. Hoppe-Seyler 376, 511-514 (1995)
  7. M. Conradt, T. Storck, W. Stoffel Localization of N-glycosylation sites and functional role of carbohydrate units of GLAST-1, a cloned rat brain L-glutamate/L-aspartate transporter Eur. J. Biochem. 229, 682-687 (1995)
  8. U. Klöckner, T. Storck, M. Conradt, W. Stoffel Functional properties and substrate specificity of the cloned L-glutamate/L-aspartate transporter GLAST-1 from rat brain expressed in Xenopus oocytes J. Neurosc. 14, 5759-5765 (1994)
  9. K. Hofmann, M. Düker, Th. Fink, P. Lichter, W. Stoffel Human neutral amino acid transporter ASCT1: structure of the gene (SLC1A4) and localization to chromosome 2p13-p15 Genomics 24, 20-26 (1994)
  10. U. Klöckner, T. Storck, M. Conradt, W. Stoffel Electrogenic L-glutamate uptake in Xenopus laevis oocytes expressing a cloned rat brain L-glutamate/L-aspartate transporter (GLAST-1) J. Biol. Chem., 268, 14594-14596 (1993)


July 12, 2011
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