The Conventional and Breakthrough Tool for the Study of L-Glutamate Transporters
Abstract
:1. Basic Background on EAAT2 and Other L-Glutamate Transporters in the Central Nervous System
2. EAAT2 Pharmacology
3. Reasons to Choose Xenopus Oocytes for the Study of EAAT2
3.1. Advantages of Xenopus Oocytes That Overexpress EAAT2
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- Direct injection enables to express the targeted proteins with a high success rate.
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- Approximately 200 or more oocytes at defolliculated stage V or VI that are suitable for cRNA injection can be collected from one Xenopus. Four to six operations with 1-week intervals are possible per one Xenopus. Additionally, three injections are possible for the oocytes collected in one operation. Therefore, ready-to-use oocytes can be obtained 12 times a month.
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- The transfection protocol is easy and feasible (Figure 1A(a1)). To overexpress the targeted protein in Xenopus oocytes, 50 nL of capped cRNA (10 ng) solution is directly injected into an oocyte by a nanoinjector installed with a glass microelectrode. An oocyte is a spherical cell with a diameter of 1–1.2 mm, large enough to confirm a successful injection by checking the swelling of the oocyte. Furthermore, the diameter of the microelectrode tip is 20–25 μm, which enables the minimization of membrane damage and the initiation of electrophysiological recording the next day.
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- By modulating the interval between the injection and the analysis, the expression level of the targeted proteins can be regulated. Conversely, experiments using oocytes expressing almost the same level of the targeted proteins can be performed.
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- The interactions of the targeted protein with the accessory subunits and/or modulatory proteins can be examined through the co-transfection of these proteins at the optimal ratio [e.g., protein interacting with C kinase 1 (PICK1), containing the postsynaptic density protein (PDZ) domain, with GLT1 (EAAT2 in rodents) [53]].
3.2. Measurement of the Activity of EAAT2 Expressed in Xenopus Oocytes
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- Because a Xenopus oocyte is a large single cell, TEVC methods can be applied. Among the various types of electrophysiological techniques, it is rather easy to become proficient in performing TEVC. Furthermore, the oocyte membrane is so strong and stable that obtaining recordings from 10–30 oocytes per day and for longer than 30 min per oocyte is possible [53,56,57].
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- Because the current values are so large (31.4 ± 21.7 nA, n = 27, holding potential = −50 mV) in Xenopus oocytes transfected with EAAT2 [58], accurate quantification of any modulation is possible.
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- It is possible to modify intracellular conditions in addition to the extracellular condition during recording. For example, it is possible to change intracellular conditions by filling glass microelectrodes with compounds such as H2O2 or DTT [57]. The extracellular conditions can be modulated by changing the pH, Cl− concentration, etc. Furthermore, the chronic effects of compounds can be examined [61] through incubation with the conditioned medium for ~2 days.
3.3. Application of Molecular Biological Techniques to Xenopus Oocytes
4. New Findings about the Interactions between PUFAs and EAAT2 Obtained with Xenopus Oocyte Experiments
5. New Insights Suggested by the Interactions between DHA and L-Glu Transporters
5.1. Some PUFAs Modulate EAATs as Allosteric Modulators
5.2. Physiological Significance: The Potential of DHA as a Neurotransmission Modulator
6. Summary
Author Contributions
Funding
Institutional Review Board Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
ALS | Amyotrophic lateral sclerosis |
DHA | Docosahexaenoic acid |
EAAT | Excitatory amino acid transporter |
HP | Helical hairpin |
IFS | Inward facing state |
iPLA2 | Ca2+-independent phospholipase A2 |
IV relation | Current–voltage relation |
L-Asp | Aspartate |
L-Glu | Glutamate |
OFS | Outward facing state |
PUFA | Polyunsaturated fatty acid |
SCAM | Substituted cysteine accessibility method |
TEVC | Two-electrode whole cell voltage clamp |
TM | Transmembrane |
References
- Arriza, J.L.; Fairman, W.A.; Wadiche, J.I.; Murdoch, G.H.; Kavanaugh, M.P.; Amara, S.G. Functional comparisons of three glutamate transporter subtypes cloned from human motor cortex. J. Neurosci. 1994, 14, 5559–5569. [Google Scholar] [CrossRef]
- Kanai, Y.; Hediger, M.A. Primary structure and functional characterization of a high-affinity glutamate transporter. Nature 1992, 360, 467–471. [Google Scholar] [CrossRef]
- Fairman, W.A.; Vandenberg, R.J.; Arriza, J.L.; Kavanaugh, M.P.; Amara, S.G. An excitatory amino-acid transporter with properties of a ligand-gated chloride channel. Nature 1995, 375, 599–603. [Google Scholar] [CrossRef] [PubMed]
- Arriza, J.L.; Eliasof, S.; Kavanaugh, M.P.; Amara, S.G. Excitatory amino acid transporter 5, a retinal glutamate transporter coupled to a chloride conductance. Proc. Natl. Acad. Sci. USA 1997, 94, 4155–4160. [Google Scholar] [CrossRef] [PubMed]
- Tanaka, K.; Watase, K.; Manabe, T.; Yamada, K.; Watanabe, M.; Takahashi, K.; Iwama, H.; Nishikawa, T.; Ichihara, N.; Kikuchi, T.; et al. Epilepsy and exacerbation of brain injury in mice lacking the glutamate transporter GLT-1. Science 1997, 276, 1699–1702. [Google Scholar] [CrossRef] [PubMed]
- Michaelis, E.K. Molecular biology of glutamate receptors in the central nervous system and their role in excitotoxicity, oxidative stress and aging. Prog. Neurobiol. 1998, 54, 369–415. [Google Scholar] [CrossRef] [PubMed]
- Mennerick, S.; Dhond, R.P.; Benz, A.; Xu, W.; Rothstein, J.D.; Danbolt, N.C.; Isenberg, K.E.; Zorumski, C.F. Neuronal expression of the glutamate transporter GLT-1 in hippocampal microcultures. J. Neurosci. 1998, 18, 4490–4499. [Google Scholar] [CrossRef] [PubMed]
- Zerangue, N.; Kavanaugh, M.P. Flux coupling in a neuronal glutamate transporter. Nature 1996, 383, 634–637. [Google Scholar] [CrossRef] [PubMed]
- Herman, M.A.; Jahr, C.E. Extracellular glutamate concentration in hippocampal slice. J. Neurosci. 2007, 27, 9736–9741. [Google Scholar] [CrossRef] [PubMed]
- Rose, E.M.; Koo, J.C.; Antflick, J.E.; Ahmed, S.M.; Angers, S.; Hampson, D.R. Glutamate transporter coupling to Na,K-ATPase. J. Neurosci. 2009, 29, 8143–8155. [Google Scholar] [CrossRef] [PubMed]
- Voutsinos-Porche, B.; Bonvento, G.; Tanaka, K.; Steiner, P.; Welker, E.; Chatton, J.Y.; Magistretti, P.J.; Pellerin, L. Glial glutamate transporters mediate a functional metabolic crosstalk between neurons and astrocytes in the mouse developing cortex. Neuron 2003, 37, 275–286. [Google Scholar] [CrossRef] [PubMed]
- Magistretti, P.J.; Allaman, I. A cellular perspective on brain energy metabolism and functional imaging. Neuron 2015, 86, 883–901. [Google Scholar] [CrossRef] [PubMed]
- Héja, L.; Nyitrai, G.; Kékesi, O.; Dobolyi, A.; Szabó, P.; Fiáth, R.; Ulbert, I.; Pál-Szenthe, B.; Palkovits, M.; Kardos, J. Astrocytes convert network excitation to tonic inhibition of neurons. BMC Biol. 2012, 10, 26. [Google Scholar] [CrossRef] [PubMed]
- Wadiche, J.I.; Amara, S.G.; Kavanaugh, M.P. Ion fluxes associated with excitatory amino acid transport. Neuron 1995, 15, 721–728. [Google Scholar] [CrossRef] [PubMed]
- Kato, T.; Kusakizako, T.; Jin, C.; Zhou, X.; Ohgaki, R.; Quan, L.; Xu, M.; Okuda, S.; Kobayashi, K.; Yamashita, K.; et al. Structural insights into inhibitory mechanism of human excitatory amino acid transporter EAAT2. Nat. Commun. 2022, 13, 4714. [Google Scholar] [CrossRef] [PubMed]
- Takaki, J.; Fujimori, K.; Miura, M.; Suzuki, T.; Sekino, Y.; Sato, K. L-glutamate released from activated microglia downregulates astrocytic L-glutamate transporter expression in neuroinflammation: The ‘collusion’ hypothesis for increased extracellular L-glutamate concentration in neuroinflammation. J. Neuroinflamm. 2012, 9, 275. [Google Scholar] [CrossRef] [PubMed]
- Rothstein, J.D.; Van Kammen, M.; Levey, A.I.; Martin, L.J.; Kuncl, R.W. Selective loss of glial glutamate transporter GLT-1 in amyotrophic lateral sclerosis. Ann. Neurol. 1995, 38, 73–84. [Google Scholar] [CrossRef] [PubMed]
- Heider, J.; Vogel, S.; Volkmer, H.; Breitmeyer, R. Human iPSC-Derived Glia as a Tool for Neuropsychiatric Research and Drug Development. Int. J. Mol. Sci. 2021, 22, 10254. [Google Scholar] [CrossRef] [PubMed]
- Wilton, D.K.; Stevens, B. The contribution of glial cells to Huntington’s disease pathogenesis. Neurobiol. Dis. 2020, 143, 104963. [Google Scholar] [CrossRef]
- Garcia, V.J.; Rushton, D.J.; Tom, C.M.; Allen, N.D.; Kemp, P.J.; Svendsen, C.N.; Mattis, V.B. Huntington’s Disease Patient-Derived Astrocytes Display Electrophysiological Impairments and Reduced Neuronal Support. Front. Neurosci. 2019, 13, 669. [Google Scholar] [CrossRef] [PubMed]
- Tyzack, G.; Lakatos, A.; Patani, R. Human Stem Cell-Derived Astrocytes: Specification and Relevance for Neurological Disorders. Curr. Stem Cell Rep. 2016, 2, 236–247. [Google Scholar] [CrossRef] [PubMed]
- Hinoi, E.; Takarada, T.; Tsuchihashi, Y.; Yoneda, Y. Glutamate transporters as drug targets. Curr. Drug Targets CNS Neurol. Disord. 2005, 4, 211–220. [Google Scholar] [CrossRef] [PubMed]
- Lin, L.; Yee, S.W.; Kim, R.B.; Giacomini, K.M. SLC transporters as therapeutic targets: Emerging opportunities. Nat. Rev. Drug Discov. 2015, 14, 543–560. [Google Scholar] [CrossRef] [PubMed]
- Wang, W.W.; Gallo, L.; Jadhav, A.; Hawkins, R.; Parker, C.G. The Druggability of Solute Carriers. J. Med. Chem. 2020, 63, 3834–3867. [Google Scholar] [CrossRef] [PubMed]
- Bunch, L.; Erichsen, M.N.; Jensen, A.A. Excitatory amino acid transporters as potential drug targets. Expert Opin. Ther. Targets 2009, 13, 719–731. [Google Scholar] [CrossRef] [PubMed]
- Bridges, R.J.; Esslinger, C.S. The excitatory amino acid transporters: Pharmacological insights on substrate and inhibitor specificity of the EAAT subtypes. Pharmacol. Ther. 2005, 107, 271–285. [Google Scholar] [CrossRef] [PubMed]
- Vandenberg, R.J.; Ryan, R.M. Mechanisms of glutamate transport. Physiol. Rev. 2013, 93, 1621–1657. [Google Scholar] [CrossRef] [PubMed]
- Koch, H.P.; Kavanaugh, M.P.; Esslinger, C.S.; Zerangue, N.; Humphrey, J.M.; Amara, S.G.; Chamberlin, A.R.; Bridges, R.J. Differentiation of substrate and nonsubstrate inhibitors of the high-affinity, sodium-dependent glutamate transporters. Mol. Pharmacol. 1999, 56, 1095–1104. [Google Scholar] [CrossRef] [PubMed]
- Willis, C.L.; Humphrey, J.M.; Koch, H.P.; Hart, J.A.; Blakely, T.; Ralston, L.; Baker, C.A.; Shim, S.; Kadri, M.; Chamberlin, A.R.; et al. L-trans-2,3-pyrrolidine dicarboxylate: Characterization of a novel excitotoxin. Neuropharmacology 1996, 35, 531–539. [Google Scholar] [CrossRef] [PubMed]
- Bridges, R.J.; Kavanaugh, M.P.; Chamberlin, A.R. A pharmacological review of competitive inhibitors and substrates of high-affinity, sodium-dependent glutamate transport in the central nervous system. Curr. Pharm. Des. 1999, 5, 363–379. [Google Scholar] [CrossRef] [PubMed]
- Vandenberg, R.J.; Mitrovic, A.D.; Chebib, M.; Balcar, V.J.; Johnston, G.A. Contrasting modes of action of methylglutamate derivatives on the excitatory amino acid transporters, EAAT1 and EAAT2. Mol. Pharmacol. 1997, 51, 809–815. [Google Scholar] [CrossRef] [PubMed]
- Donevan, S.D.; Beg, A.; Gunther, J.M.; Twyman, R.E. The methylglutamate, SYM 2081, is a potent and highly selective agonist at kainate receptors. J. Pharmacol. Exp. Ther. 1998, 285, 539–545. [Google Scholar] [PubMed]
- Shimamoto, K.; Lebrun, B.; Yasuda-Kamatani, Y.; Sakaitani, M.; Shigeri, Y.; Yumoto, N.; Nakajima, T. DL-threo-beta-benzyloxyaspartate, a potent blocker of excitatory amino acid transporters. Mol. Pharmacol. 1998, 53, 195–201. [Google Scholar] [CrossRef] [PubMed]
- Shigeri, Y.; Shimamoto, K.; Yasuda-Kamatani, Y.; Seal, R.P.; Yumoto, N.; Nakajima, T.; Amara, S.G. Effects of threo-beta-hydroxyaspartate derivatives on excitatory amino acid transporters (EAAT4 and EAAT5). J. Neurochem. 2001, 79, 297–302. [Google Scholar] [CrossRef] [PubMed]
- Jabaudon, D.; Shimamoto, K.; Yasuda-Kamatani, Y.; Scanziani, M.; Gähwiler, B.H.; Gerber, U. Inhibition of uptake unmasks rapid extracellular turnover of glutamate of nonvesicular origin. Proc. Natl. Acad. Sci. USA 1999, 96, 8733–8738. [Google Scholar] [CrossRef] [PubMed]
- Shimamoto, K.; Sakai, R.; Takaoka, K.; Yumoto, N.; Nakajima, T.; Amara, S.G.; Shigeri, Y. Characterization of novel L-threo-beta-benzyloxyaspartate derivatives, potent blockers of the glutamate transporters. Mol. Pharmacol. 2004, 65, 1008–1015. [Google Scholar] [CrossRef] [PubMed]
- Dunlop, J.; McIlvain, H.B.; Carrick, T.A.; Jow, B.; Lu, Q.; Kowal, D.; Lin, S.; Greenfield, A.; Grosanu, C.; Fan, K.; et al. Characterization of novel aryl-ether, biaryl, and fluorene aspartic acid and diaminopropionic acid analogs as potent inhibitors of the high-affinity glutamate transporter EAAT2. Mol. Pharmacol. 2005, 68, 974–982. [Google Scholar] [CrossRef]
- Fontana, A.C.; de Oliveira Beleboni, R.; Wojewodzic, M.W.; Ferreira Dos Santos, W.; Coutinho-Netto, J.; Grutle, N.J.; Watts, S.D.; Danbolt, N.C.; Amara, S.G. Enhancing glutamate transport: Mechanism of action of Parawixin1, a neuroprotective compound from Parawixia bistriata spider venom. Mol. Pharmacol. 2007, 72, 1228–1237. [Google Scholar] [CrossRef]
- Fontana, A.C.; Guizzo, R.; de Oliveira Beleboni, R.; Meirelles, E.S.A.R.; Coimbra, N.C.; Amara, S.G.; dos Santos, W.F.; Coutinho-Netto, J. Purification of a neuroprotective component of Parawixia bistriata spider venom that enhances glutamate uptake. Br. J. Pharmacol. 2003, 139, 1297–1309. [Google Scholar] [CrossRef] [PubMed]
- Kortagere, S.; Mortensen, O.V.; Xia, J.; Lester, W.; Fang, Y.; Srikanth, Y.; Salvino, J.M.; Fontana, A.C.K. Identification of Novel Allosteric Modulators of Glutamate Transporter EAAT2. ACS Chem. Neurosci. 2018, 9, 522–534. [Google Scholar] [CrossRef]
- Rothstein, J.D.; Patel, S.; Regan, M.R.; Haenggeli, C.; Huang, Y.H.; Bergles, D.E.; Jin, L.; Dykes Hoberg, M.; Vidensky, S.; Chung, D.S.; et al. Beta-lactam antibiotics offer neuroprotection by increasing glutamate transporter expression. Nature 2005, 433, 73–77. [Google Scholar] [CrossRef] [PubMed]
- Cudkowicz, M.E.; Titus, S.; Kearney, M.; Yu, H.; Sherman, A.; Schoenfeld, D.; Hayden, D.; Shui, A.; Brooks, B.; Conwit, R.; et al. Safety and efficacy of ceftriaxone for amyotrophic lateral sclerosis: A multi-stage, randomised, double-blind, placebo-controlled trial. Lancet Neurol. 2014, 13, 1083–1091. [Google Scholar] [CrossRef] [PubMed]
- Lehre, K.P.; Levy, L.M.; Ottersen, O.P.; Storm-Mathisen, J.; Danbolt, N.C. Differential expression of two glial glutamate transporters in the rat brain: Quantitative and immunocytochemical observations. J. Neurosci. 1995, 15, 1835–1853. [Google Scholar] [CrossRef] [PubMed]
- Haugeto, O.; Ullensvang, K.; Levy, L.M.; Chaudhry, F.A.; Honoré, T.; Nielsen, M.; Lehre, K.P.; Danbolt, N.C. Brain glutamate transporter proteins form homomultimers. J. Biol. Chem. 1996, 271, 27715–27722. [Google Scholar] [CrossRef] [PubMed]
- Sato, K.; Matsuki, N.; Ohno, Y.; Nakazawa, K. Estrogens inhibit l-glutamate uptake activity of astrocytes via membrane estrogen receptor alpha. J. Neurochem. 2003, 86, 1498–1505. [Google Scholar] [CrossRef] [PubMed]
- Gegelashvili, G.; Danbolt, N.C.; Schousboe, A. Neuronal soluble factors differentially regulate the expression of the GLT1 and GLAST glutamate transporters in cultured astroglia. J. Neurochem. 1997, 69, 2612–2615. [Google Scholar] [CrossRef] [PubMed]
- Schlag, B.D.; Vondrasek, J.R.; Munir, M.; Kalandadze, A.; Zelenaia, O.A.; Rothstein, J.D.; Robinson, M.B. Regulation of the glial Na+-dependent glutamate transporters by cyclic AMP analogs and neurons. Mol. Pharmacol. 1998, 53, 355–369. [Google Scholar] [CrossRef] [PubMed]
- Swanson, R.A.; Liu, J.; Miller, J.W.; Rothstein, J.D.; Farrell, K.; Stein, B.A.; Longuemare, M.C. Neuronal regulation of glutamate transporter subtype expression in astrocytes. J. Neurosci. 1997, 17, 932–940. [Google Scholar] [CrossRef] [PubMed]
- Yang, Y.; Gozen, O.; Watkins, A.; Lorenzini, I.; Lepore, A.; Gao, Y.; Vidensky, S.; Brennan, J.; Poulsen, D.; Won Park, J.; et al. Presynaptic regulation of astroglial excitatory neurotransmitter transporter GLT1. Neuron 2009, 61, 880–894. [Google Scholar] [CrossRef] [PubMed]
- Berry, C.B.; Hayes, D.; Murphy, A.; Wiessner, M.; Rauen, T.; McBean, G.J. Differential modulation of the glutamate transporters GLT1, GLAST and EAAC1 by docosahexaenoic acid. Brain Res. 2005, 1037, 123–133. [Google Scholar] [CrossRef] [PubMed]
- Dunlop, J.; Lou, Z.; Zhang, Y.; McIlvain, H.B. Inducible expression and pharmacology of the human excitatory amino acid transporter 2 subtype of L-glutamate transporter. Br. J. Pharmacol. 1999, 128, 1485–1490. [Google Scholar] [CrossRef] [PubMed]
- Duffield, M.; Patel, A.; Mortensen, O.V.; Schnur, D.; Gonzalez-Suarez, A.D.; Torres-Salazar, D.; Fontana, A.C.K. Transport rate of EAAT2 is regulated by amino acid located at the interface between the scaffolding and substrate transport domains. Neurochem. Int. 2020, 139, 104792. [Google Scholar] [CrossRef] [PubMed]
- Sogaard, R.; Borre, L.; Braunstein, T.H.; Madsen, K.L.; MacAulay, N. Functional modulation of the glutamate transporter variant GLT1b by the PDZ domain protein PICK1. J. Biol. Chem. 2013, 288, 20195–20207. [Google Scholar] [CrossRef] [PubMed]
- Dvorak, V.; Wiedmer, T.; Ingles-Prieto, A.; Altermatt, P.; Batoulis, H.; Bärenz, F.; Bender, E.; Digles, D.; Dürrenberger, F.; Heitman, L.H.; et al. An Overview of Cell-Based Assay Platforms for the Solute Carrier Family of Transporters. Front. Pharmacol. 2021, 12, 722889. [Google Scholar] [CrossRef] [PubMed]
- Kolen, B.; Kortzak, D.; Franzen, A.; Fahlke, C. An amino-terminal point mutation increases EAAT2 anion currents without affecting glutamate transport rates. J. Biol. Chem. 2020, 295, 14936–14947. [Google Scholar] [CrossRef] [PubMed]
- Trotti, D.; Danbolt, N.C.; Volterra, A. Glutamate transporters are oxidant-vulnerable: A molecular link between oxidative and excitotoxic neurodegeneration? Trends Pharmacol. Sci. 1998, 19, 328–334. [Google Scholar] [CrossRef] [PubMed]
- Trotti, D.; Rolfs, A.; Danbolt, N.C.; Brown, R.H., Jr.; Hediger, M.A. SOD1 mutants linked to amyotrophic lateral sclerosis selectively inactivate a glial glutamate transporter. Nat. Neurosci. 1999, 2, 427–433. [Google Scholar] [CrossRef]
- Takahashi, K.; Chen, L.; Sayama, M.; Wu, M.; Hayashi, M.K.; Irie, T.; Ohwada, T.; Sato, K. Leucine 434 is essential for docosahexaenoic acid-induced augmentation of L-glutamate transporter current. J. Biol. Chem. 2022, 299, 102793. [Google Scholar] [CrossRef] [PubMed]
- Otis, T.S.; Kavanaugh, M.P. Isolation of current components and partial reaction cycles in the glial glutamate transporter EAAT2. J. Neurosci. 2000, 20, 2749–2757. [Google Scholar] [CrossRef] [PubMed]
- Wadiche, J.I.; Kavanaugh, M.P. Macroscopic and microscopic properties of a cloned glutamate transporter/chloride channel. J. Neurosci. 1998, 18, 7650–7661. [Google Scholar] [CrossRef]
- Fairman, W.A.; Sonders, M.S.; Murdoch, G.H.; Amara, S.G. Arachidonic acid elicits a substrate-gated proton current associated with the glutamate transporter EAAT4. Nat. Neurosci. 1998, 1, 105–113. [Google Scholar] [CrossRef] [PubMed]
- Akabas, M.H. Cysteine Modification: Probing Channel Structure, Function and Conformational Change. Adv. Exp. Med. Biol. 2015, 869, 25–54. [Google Scholar] [CrossRef]
- Akabas, M.H.; Stauffer, D.A.; Xu, M.; Karlin, A. Acetylcholine receptor channel structure probed in cysteine-substitution mutants. Science 1992, 258, 307–310. [Google Scholar] [CrossRef] [PubMed]
- Grunewald, M.; Bendahan, A.; Kanner, B.I. Biotinylation of single cysteine mutants of the glutamate transporter GLT-1 from rat brain reveals its unusual topology. Neuron 1998, 21, 623–632. [Google Scholar] [CrossRef]
- Grunewald, M.; Menaker, D.; Kanner, B.I. Cysteine-scanning mutagenesis reveals a conformationally sensitive reentrant pore-loop in the glutamate transporter GLT-1. J. Biol. Chem. 2002, 277, 26074–26080. [Google Scholar] [CrossRef] [PubMed]
- Rong, X.; Tan, F.; Wu, X.; Zhang, X.; Lu, L.; Zou, X.; Qu, S. TM4 of the glutamate transporter GLT-1 experiences substrate-induced motion during the transport cycle. Sci. Rep. 2016, 6, 34522. [Google Scholar] [CrossRef]
- Zhang, Y.; Kanner, B.I. Two serine residues of the glutamate transporter GLT-1 are crucial for coupling the fluxes of sodium and the neurotransmitter. Proc. Natl. Acad. Sci. USA 1999, 96, 1710–1715. [Google Scholar] [CrossRef] [PubMed]
- Brocke, L.; Bendahan, A.; Grunewald, M.; Kanner, B.I. Proximity of two oppositely oriented reentrant loops in the glutamate transporter GLT-1 identified by paired cysteine mutagenesis. J. Biol. Chem. 2002, 277, 3985–3992. [Google Scholar] [CrossRef] [PubMed]
- Vandenberg, R.J.; Arriza, J.L.; Amara, S.G.; Kavanaugh, M.P. Constitutive ion fluxes and substrate binding domains of human glutamate transporters. J. Biol. Chem. 1995, 270, 17668–17671. [Google Scholar] [CrossRef] [PubMed]
- Mitrovic, A.D.; Amara, S.G.; Johnston, G.A.; Vandenberg, R.J. Identification of functional domains of the human glutamate transporters EAAT1 and EAAT2. J. Biol. Chem. 1998, 273, 14698–14706. [Google Scholar] [CrossRef]
- Storck, T.; Schulte, S.; Hofmann, K.; Stoffel, W. Structure, expression, and functional analysis of a Na(+)-dependent glutamate/aspartate transporter from rat brain. Proc. Natl. Acad. Sci. USA 1992, 89, 10955–10959. [Google Scholar] [CrossRef] [PubMed]
- Yernool, D.; Boudker, O.; Jin, Y.; Gouaux, E. Structure of a glutamate transporter homologue from Pyrococcus horikoshii. Nature 2004, 431, 811–818. [Google Scholar] [CrossRef]
- Zhang, Z.; Chen, H.; Geng, Z.; Yu, Z.; Li, H.; Dong, Y.; Zhang, H.; Huang, Z.; Jiang, J.; Zhao, Y. Structural basis of ligand binding modes of human EAAT2. Nat. Commun. 2022, 13, 3329. [Google Scholar] [CrossRef] [PubMed]
- Jardetzky, O. Simple allosteric model for membrane pumps. Nature 1966, 211, 969–970. [Google Scholar] [CrossRef] [PubMed]
- Boudker, O.; Ryan, R.M.; Yernool, D.; Shimamoto, K.; Gouaux, E. Coupling substrate and ion binding to extracellular gate of a sodium-dependent aspartate transporter. Nature 2007, 445, 387–393. [Google Scholar] [CrossRef] [PubMed]
- Verdon, G.; Boudker, O. Crystal structure of an asymmetric trimer of a bacterial glutamate transporter homolog. Nat. Struct. Mol. Biol. 2012, 19, 355–357. [Google Scholar] [CrossRef] [PubMed]
- Akyuz, N.; Georgieva, E.R.; Zhou, Z.; Stolzenberg, S.; Cuendet, M.A.; Khelashvili, G.; Altman, R.B.; Terry, D.S.; Freed, J.H.; Weinstein, H.; et al. Transport domain unlocking sets the uptake rate of an aspartate transporter. Nature 2015, 518, 68–73. [Google Scholar] [CrossRef] [PubMed]
- Reyes, N.; Ginter, C.; Boudker, O. Transport mechanism of a bacterial homologue of glutamate transporters. Nature 2009, 462, 880–885. [Google Scholar] [CrossRef] [PubMed]
- Canul-Tec, J.C.; Assal, R.; Cirri, E.; Legrand, P.; Brier, S.; Chamot-Rooke, J.; Reyes, N. Structure and allosteric inhibition of excitatory amino acid transporter 1. Nature 2017, 544, 446–451. [Google Scholar] [CrossRef] [PubMed]
- Sherman, W.; Beard, H.S.; Farid, R. Use of an induced fit receptor structure in virtual screening. Chem. Biol. Drug Des. 2006, 67, 83–84. [Google Scholar] [CrossRef] [PubMed]
- Sherman, W.; Day, T.; Jacobson, M.P.; Friesner, R.A.; Farid, R. Novel procedure for modeling ligand/receptor induced fit effects. J. Med. Chem. 2006, 49, 534–553. [Google Scholar] [CrossRef] [PubMed]
- Friesner, R.A.; Banks, J.L.; Murphy, R.B.; Halgren, T.A.; Klicic, J.J.; Mainz, D.T.; Repasky, M.P.; Knoll, E.H.; Shelley, M.; Perry, J.K.; et al. Glide: A new approach for rapid, accurate docking and scoring. 1. Method and assessment of docking accuracy. J. Med. Chem. 2004, 47, 1739–1749. [Google Scholar] [CrossRef] [PubMed]
- Abrahamsen, B.; Schneider, N.; Erichsen, M.N.; Huynh, T.H.; Fahlke, C.; Bunch, L.; Jensen, A.A. Allosteric modulation of an excitatory amino acid transporter: The subtype-selective inhibitor UCPH-101 exerts sustained inhibition of EAAT1 through an intramonomeric site in the trimerization domain. J. Neurosci. 2013, 33, 1068–1087. [Google Scholar] [CrossRef] [PubMed]
- Chen, I.; Pant, S.; Wu, Q.; Cater, R.J.; Sobti, M.; Vandenberg, R.J.; Stewart, A.G.; Tajkhorshid, E.; Font, J.; Ryan, R.M. Glutamate transporters have a chloride channel with two hydrophobic gates. Nature 2021, 591, 327–331. [Google Scholar] [CrossRef]
- Attwell, D.; Miller, B.; Sarantis, M. Arachidonic acid as a messenger in the central nervous system. Semin. Neurosci. 1993, 5, 159–169. [Google Scholar] [CrossRef]
- Piomelli, D. Arachidonic acid in cell signaling. Curr. Opin. Cell Biol. 1993, 5, 274–280. [Google Scholar] [CrossRef] [PubMed]
- Piomelli, D.; Greengard, P. Lipoxygenase metabolites of arachidonic acid in neuronal transmembrane signalling. Trends Pharmacol. Sci. 1990, 11, 367–373. [Google Scholar] [CrossRef] [PubMed]
- Aid, S.; Vancassel, S.; Poumes-Ballihaut, C.; Chalon, S.; Guesnet, P.; Lavialle, M. Effect of a diet-induced n-3 PUFA depletion on cholinergic parameters in the rat hippocampus. J. Lipid Res. 2003, 44, 1545–1551. [Google Scholar] [CrossRef]
- Novak, E.M.; Dyer, R.A.; Innis, S.M. High dietary omega-6 fatty acids contribute to reduced docosahexaenoic acid in the developing brain and inhibit secondary neurite growth. Brain Res. 2008, 1237, 136–145. [Google Scholar] [CrossRef]
- Rapoport, S.I. Translational studies on regulation of brain docosahexaenoic acid (DHA) metabolism in vivo. Prostaglandins Leukot. Essent. Fat. Acids 2013, 88, 79–85. [Google Scholar] [CrossRef] [PubMed]
- Salem, N., Jr.; Litman, B.; Kim, H.Y.; Gawrisch, K. Mechanisms of action of docosahexaenoic acid in the nervous system. Lipids 2001, 36, 945–959. [Google Scholar] [CrossRef] [PubMed]
- Moore, S.A. Cerebral endothelium and astrocytes cooperate in supplying docosahexaenoic acid to neurons. Adv. Exp. Med. Biol. 1993, 331, 229–233. [Google Scholar] [PubMed]
- Moore, S.A. Polyunsaturated fatty acid synthesis and release by brain-derived cells in vitro. J. Mol. Neurosci. 2001, 16, 195–200; discussion 215–121. [Google Scholar] [CrossRef] [PubMed]
- Williard, D.E.; Harmon, S.D.; Kaduce, T.L.; Preuss, M.; Moore, S.A.; Robbins, M.E.; Spector, A.A. Docosahexaenoic acid synthesis from n-3 polyunsaturated fatty acids in differentiated rat brain astrocytes. J. Lipid Res. 2001, 42, 1368–1376. [Google Scholar] [CrossRef] [PubMed]
- Bazan, N.G. Synaptic lipid signaling: Significance of polyunsaturated fatty acids and platelet-activating factor. J. Lipid Res. 2003, 44, 2221–2233. [Google Scholar] [CrossRef]
- Green, J.T.; Orr, S.K.; Bazinet, R.P. The emerging role of group VI calcium-independent phospholipase A2 in releasing docosahexaenoic acid from brain phospholipids. J. Lipid Res. 2008, 49, 939–944. [Google Scholar] [CrossRef] [PubMed]
- Strokin, M.; Sergeeva, M.; Reiser, G. Docosahexaenoic acid and arachidonic acid release in rat brain astrocytes is mediated by two separate isoforms of phospholipase A2 and is differently regulated by cyclic AMP and Ca2+. Br. J. Pharmacol. 2003, 139, 1014–1022. [Google Scholar] [CrossRef]
- Strokin, M.; Sergeeva, M.; Reiser, G. Prostaglandin synthesis in rat brain astrocytes is under the control of the n-3 docosahexaenoic acid, released by group VIB calcium-independent phospholipase A2. J. Neurochem. 2007, 102, 1771–1782. [Google Scholar] [CrossRef] [PubMed]
- Kuratko, C.N.; Barrett, E.C.; Nelson, E.B.; Salem, N., Jr. The relationship of docosahexaenoic acid (DHA) with learning and behavior in healthy children: A review. Nutrients 2013, 5, 2777–2810. [Google Scholar] [CrossRef] [PubMed]
- Valtcheva, S.; Venance, L. Control of Long-Term Plasticity by Glutamate Transporters. Front. Synaptic Neurosci. 2019, 11, 10. [Google Scholar] [CrossRef] [PubMed]
- Tzingounis, A.V.; Wadiche, J.I. Glutamate transporters: Confining runaway excitation by shaping synaptic transmission. Nat. Rev. Neurosci. 2007, 8, 935–947. [Google Scholar] [CrossRef] [PubMed]
- Conti, F.; Weinberg, R.J. Shaping excitation at glutamatergic synapses. Trends Neurosci. 1999, 22, 451–458. [Google Scholar] [CrossRef]
- Rose, C.R.; Felix, L.; Zeug, A.; Dietrich, D.; Reiner, A.; Henneberger, C. Astroglial Glutamate Signaling and Uptake in the Hippocampus. Front. Mol. Neurosci. 2017, 10, 451. [Google Scholar] [CrossRef] [PubMed]
- Fujita, S.; Ikegaya, Y.; Nishikawa, M.; Nishiyama, N.; Matsuki, N. Docosahexaenoic acid improves long-term potentiation attenuated by phospholipase A(2) inhibitor in rat hippocampal slices. Br. J. Pharmacol. 2001, 132, 1417–1422. [Google Scholar] [CrossRef] [PubMed]
- Mazzocchi-Jones, D. Impaired corticostriatal LTP and depotentiation following iPLA2 inhibition is restored following acute application of DHA. Brain Res. Bull. 2015, 111, 69–75. [Google Scholar] [CrossRef] [PubMed]
Compounds | Selectivity | Type for EAAT2 | Effect on EAAT2 Current | Extra Info | Refs | |
---|---|---|---|---|---|---|
Inhibitors | THA DL-threo-β-hydroxyaspartate |
EAAT2 = 19 μM EAAT3 = 25 μM
| Substrate | - |
| [1,3,4] |
L-trans-2.4-PDC L-trans-2.4-pyrrolidine dicarboxylate |
EAAT2 = 8 μM EAAT3 = 61 μM EAAT4 = 2.6 μM
| Substrate | - |
| [1,3,4] | |
L-trans-2.3-PDC L-trans-2.3-pyrrolidine dicarboxylate |
| Blocker | ↓ |
| [28,29,30] | |
SYM2081 (2S,4R)-4-methylglutamate |
| Blocker | ↓ |
| [31,32] | |
TBOA DL-threo-β-benzyloxyaspartate |
EAAT2 = 6 μM EAAT3 = 6 μM EAAT4 = 4.4 μM EAAT5 = 3.2 μM | Blocker | ↓ |
| [33,34,35] | |
TFB-TBOA (2S,3S)-3-{3-[4-(trifluoromethyl)benzoylamino]benzyloxy}aspartate |
EAAT2 = 17 nM EAAT3 = 300 nM | Blocker | ↓ |
| [36] | |
DHK Dihydrokainic acid |
EAAT2 = 23 μM EAAT3 > 3 mM | Blocker | ↓ |
| [1,33] | |
WAY213613 N(4)-[4-(2-bromo-4,5-difluorophenoxy)phenyl]-L-asparagine |
EAAT2 = 85 nM EAAT3 = 3.8 μM | Blocker | ↓ |
| [37] | |
Enhancers | Parawixin1 |
| Allosteric | ↑ |
| [38,39] |
GT949 3-((4-cyclohexylpiperazin-1-yl)(1-phenethyl-1H-tetrazol-5-yl)methyl)-6-methoxyquinolin-2(1H)-one |
| Allosteric | ↑ | [40] |
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Takahashi, K.; Sato, K. The Conventional and Breakthrough Tool for the Study of L-Glutamate Transporters. Membranes 2024, 14, 77. https://doi.org/10.3390/membranes14040077
Takahashi K, Sato K. The Conventional and Breakthrough Tool for the Study of L-Glutamate Transporters. Membranes. 2024; 14(4):77. https://doi.org/10.3390/membranes14040077
Chicago/Turabian StyleTakahashi, Kanako, and Kaoru Sato. 2024. "The Conventional and Breakthrough Tool for the Study of L-Glutamate Transporters" Membranes 14, no. 4: 77. https://doi.org/10.3390/membranes14040077
APA StyleTakahashi, K., & Sato, K. (2024). The Conventional and Breakthrough Tool for the Study of L-Glutamate Transporters. Membranes, 14(4), 77. https://doi.org/10.3390/membranes14040077