Skip to main content
SpringerLink
Log in

Comparison of Effects of DL-Threo-β-Benzyloxyaspartate (DL-TBOA) and L-Trans-Pyrrolidine-2,4-Dicarboxylate (t-2,4-PDC) on Uptake and Release of [3H]D-Aspartate in Astrocytes and Glutamatergic Neurons

  • Published:
Neurochemical Research Aims and scope Submit manuscript

Abstract

Uptake and release processes in cerebellar astrocytes and granule neurons (glutamatergic) for glutamate were investigated by the use of [3H]D-aspartate, a non-metabolizable glutamate analog. The effects of DL-threo-β-benzyloxyaspartate (DL-TBOA) and L-trans-pyrrolidine-2,4-dicarboxylate (t-2,4-PDC) on uptake and release of [3H]D-aspartate were studied. Both compounds inhibited potently uptake of [3H]D-aspartate in neurons and astrocytes (IC50 values 10-100 μM), DL-TBOA being slightly more potent than t-2,4-PDC. Release of preloaded [3H]D-aspartate from neurons or astrocytes could be stimulated by addition of excess t-2,4-PDC whereas addition of DL-TBOA had no effect on [3H]D-aspartate efflux. Moreover, DL-TBOA inhibited significantly the depolarization-induced (55 mM KCl) release of preloaded [3H]D-aspartate in the neurons. The results reflect the fact that DL-TBOA is not transported by the glutamate carriers while t-2,4-PDC is a substrate which may heteroexchange with [3H]D-aspartate. It is suggested that DL-TBOA may be used to selectively inhibit depolarization coupled glutamate release mediated by reversal of the carriers.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

REFERENCES

  1. Fonnum, F. 1984. Glutamate: A neurotransmitter in mammalian brain. J. Neurochem. 42:1–11.

    Google Scholar 

  2. Schousboe, A. 1981. Transport and metabolism of glutamate and GABA in neurons and glial cells. Int. Rev. Neurobiol. 22:1–45.

    Google Scholar 

  3. Gegelashvili, G. and Schousboe, A. 1997. High-affinity glutamate transporters: Regulation of expression and activity. Mol. Pharmacol. 52:6–15.

    Google Scholar 

  4. Danbolt, N. C. 1994. The high affinity uptake system for excitatory amino acids in the brain. Prog. Neurobiol. 44:377–396.

    Google Scholar 

  5. Schousboe, A., Svenneby, G., and Hertz, L. 1977. Uptake and metabolism of glutamate in astrocytes cultured from dissociated mouse brain hemispheres. J. Neurochem. 29:999–1005.

    Google Scholar 

  6. Hertz, L., Schousboe, A., Boechler, N., Mukerji, S., and Fedoroff, S. 1978. Kinetic characteristics of the glutamate uptake into normal astrocytes in cultures. Neurochem. Res. 3:1–14.

    Google Scholar 

  7. Drejer, J., Larsson, O. M., and Schousboe, A. 1982. Characterization of glutamate uptake into and release from astrocytes and neurons cultured from different brain regions. Exp. Brain Res. 47:259–269.

    Google Scholar 

  8. Lehre, K. P., Levy, L. M., Ottersen, O. P., Storm-Mathisen, J., and Danbolt, N. C. 1995. Differential expression of two glial glutamate transporters in the rat brain: Quantitative and immunocytochemical observations. J. Neurosci. 15:1835–1853.

    Google Scholar 

  9. Lehre, K. P. and Danbolt, N. C. 1998. The number of glutamate transporter subtype molecules at glutamatergic synapses: Chemical and stereological quantification in young adult rat brain. J. Neurosci. 18:8751–8757.

    Google Scholar 

  10. Frandsen, A. and Schousboe, A. 1997. Glutamate receptors and neurotoxicity. Pages 239–251, in Stone, T. W. (ed.), CNS Neurotransmitters and Neuromodulators: Glutamate. CRC Press, Boca Raton, FL.

    Google Scholar 

  11. Rothstein, J. D., Dykes-Hoberg, M., Pardo, C. A., Bristol, L. A., Jin, L., Kuncl, R. W., Kanai, Y., Hediger, M. A., Wang, Y., Schielke, J. P., and Welty, D. F. 1996. Knockout of glutamate transporters reveals a major role for astroglial transport in excitotoxicity and clearance of glutamate. Neuron 16:675–686.

    Google Scholar 

  12. Nicholls, D. and Attwell, D. 1990. The release and uptake of excitatory amino acids. TIPS 11:462–468.

    Google Scholar 

  13. Jensen, J. B. Pickering, D. S., and Schousboe, A. 2000. Depolarization-induced release of [3H]D-aspartate from GABAergic neurons caused by reversal of glutamate transporters. Int. J. Devl. Neurosci. 18:309–315.

    Google Scholar 

  14. Belhage, B., Hansen, G. H., and Schousboe, A. 1993. Depolarization by K+ and glutamate activates different neurotransmitter release mechanisms in GABAergic neurons: Vesicular versus non-vesicular release of GABA. Neuroscience 54:1019–1034.

    Google Scholar 

  15. Gegelashvili, G. and Schousboe, A. 1998. Cellular distribution and kinetic properties of high-affinity glutamate transporters. Brain Res. Bull. 45:233–238.

    Google Scholar 

  16. Dunlop, J., Grieve, A., Damgaard, I., Schousboe, A., and Griffiths, R. 1992. Sulphur-containing excitatory amino acid evoked Ca2+-independent release of D-[3H]aspartate from cultured cerebellar granule cells: The role of glutamate receptor activation coupled to reversal of the acidic amino acid plasma membrane carrier. Neuroscience 50:107–115.

    Google Scholar 

  17. Lebrun, B., Sakaitani, M., Shimamoto, K., Yasuda-Kamatani, Y., and Nakajima, T. 1997. New beta-hydroxyaspartate derivatives are competitive blockers for the bovine glutamate/aspartate transporter. J. Biol. Chem. 272:20336–20339.

    Google Scholar 

  18. Shimamoto, K., Lebrun, B., Yasuda-Kamatani, Y., Sakaitani, M., Shigeri, Y., Yumoto, N., and Nakajima, T. 1998. DL-threobeta-benzyloxyaspartate, a potent blocker of excitatory amino acid transporters. Mol. Pharmacol. 53:195–201.

    Google Scholar 

  19. Drejer, J., Larsson, O. M., and Schousboe, A. 1983. Characterization of uptake and release processes for D-and L-aspartate in primary cultures of astrocytes and cerebellar granule cells. Neurochem. Res. 8:231–243.

    Google Scholar 

  20. Hertz, L., Juurlink, B. H. J., Hertz, E., Fosmark H., and Schousboe A. 1989. Preparation of primary cultures of mouse (rat) astrocytes. Pages 105–108, in Shahar, A., De Vellis, J., Vernadakis, A., and Haber, B. (eds.), A Dissection and Tissue Culture Manual for the Nervous System. Alan R Liss, NY.

    Google Scholar 

  21. Hertz, L., Juurlink, B. H. J., Fosmark, H., and Schousboe A. 1982. Astrocytes in primary cultures. Pages 175–186, in Pfeiffer, S. E., (ed.), Neuroscience Approached through Cell Culture, Vol 1. CRC Press, Boca Raton, FL.

    Google Scholar 

  22. Schousboe, A., Meier, E., Drejer, J., and Hertz L. 1989. Preparation of primary cultures of mouse (rat) cerebellar granule cells. Pages 203–206, in Shahar, A., De Vellis, J., Vernadakis, A., and Haber, B. (eds.), A Dissection and Tissue Culture Manual for the Nervous System. Alan R Liss, NY.

    Google Scholar 

  23. Palaiologos, G., Hertz, L., and Schousboe, A. 1989. Role of aspartate aminotransferase and mitochondrial dicarboxylate transport for release of endogenously and exogenously supplied neurotransmitter in glutamatergic neurons. Neurochem. Res. 14:359–366.

    Google Scholar 

  24. Belhage, B., Rehder, V., Hansen, G. H., Kater, S. B., and Schousboe, A. 1992. [3H]D-aspartate release from cerebellar granule neurons is differentially regulated by glutamate-and K(+)-stimulation. J. Neurosci. Res. 33:436–444.

    Google Scholar 

  25. Cousin, M. A. and Nicholls, D. G. 1997. Synaptic vesicle recycling in cultured cerebellar granule cells: Role of vesicular acidification and refilling. J. Neurochem. 69:1927–1935.

    Google Scholar 

  26. Drejer, J., Honore, T., and Schousboe, A. 1987. Excitatory amino acid-induced release of 3H-GABA from cultured mouse cerebral cortex interneurons. J. Neurosci. 7:2910–2916.

    Google Scholar 

  27. Gegelashvili, G., Danbolt, N. C., and Schousboe, A. 1997. Neuronal soluble factors differentially regulate the expression of the GLT1 and GLAST glutamate transporters in cultured astroglia. J. Neurochem. 69:2612–2615.

    Google Scholar 

  28. Jaboudon, D., Shimamoto, K., Yasuda-Kamatani, Y., Scanziani, M., Gähwiler, B. H., and Gerber, U. 1999. Inhibition of uptake unmasks rapid extracellular turnover of glutamate of nonvesicular origin. Proc. Natl. Acad. Sci. USA 96:8733–8738.

    Google Scholar 

  29. Bridges, R. J., Kavanaugh, M. P., and Chamberlin, A. R. 1999. A pharmacological review of competitive inhibitors and substrates of high-affinity, sodium-dependent glutamate transport in the central nervous system. Curr. Pharm. Des. 5:363–379.

    Google Scholar 

  30. Volterra, A., Bezzi, P., Rizzini, B. L., Trotti, D., Ullensvang, K., Danbolt, N. C., and Racagni, G. 1996. The competitive transport inhibitor L-trans-pyrrolidine-2,4-dicarboxylate triggers excitotoxicity in rat cortical neuron-astrocyte co-cultures via glutamate release rather than uptake inhibition. Eur. J. Neurosci. 8:2019–2028.

    Google Scholar 

  31. Blitzblau, R., Gupta, S., Djali, S., Robinson, M. B., and Rosenberg, P. A. 1996. The glutamate transport inhibitor L-transpyrrolidine-2,4-dicarboxylate indirectly evokes NMDA receptor mediated neurotoxicity in rat cortical cultures. Eur. J. Neurosci. 8:1840–1852.

    Google Scholar 

  32. Levy, L. M., Warr, O., and Attwell, D. 1998. Stoichiometry of the glial glutamate transporter GLT-expressed inducibly in a Chinese hamster ovary cell line selected for low endogenous Na+-dependent glutamate uptake. J. Neurosci. 18:9620–9628.

    Google Scholar 

  33. Meier, E., Drejer, J., and Schousboe, A. 1984. GABA induces functionally active low-affinity GABA receptors on cultured cerebellar granule cells. J. Neurochem. 43:1737–1744.

    Google Scholar 

  34. Palaiologos, G., Hertz, L., and Schousboe, A. 1988. Evidence that aspartate amino transferase activity and ketodicarboxylate carrier function are essential for biosynthesis of transmitter glutamate. J. Neurochem. 51:317–320.

    Google Scholar 

  35. Phillis, J. W., Ren, J., and O'Regan, M. H. 2000. Transporter reversal as a mechanism of glutamate release from the ischemic rat cerebral cortex: Studies with DL-threo-beta-benzyloxyaspartate. Brain. Res. 868:105–112.

    Google Scholar 

  36. Phillis, J. W., Smith-Barbour, M., Perkins, L. M., and O'Regan, M. H. 1994. Characterization of glutamate, aspartate, and GABA release from ischemic rat cerebral cortex. Brain Res. Bull. 34:457–466.

    Google Scholar 

  37. Seki, Y., Feustel, P. J., Keller, R. W., Tranmer, B. I., and Kimelberg, H. K. 1999. Inhibition of ischemia-induced glutamate release in rat striatum by dihydrokinate and an anion channel blocker. Stroke 30:433–440.

    Google Scholar 

  38. Waagepetersen, H. S., Sonnewald, U., Gegelashvili, G., Larsson, O. M., and Schousboe, A. 2001. Metabolic distinction between vesicular and cytosolic GABA in cultured GABAergic neurons using 13C MRS. J. Neurosci. Res. 63:347–355.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. NeuroScience PharmaBiotec Res. Center, Dept. of Pharmacology, The Royal Danish School of Pharmacy, DK-2100, Copenhagen, Denmark

    Helle S. Waagepetersen & Arne Schousboe

  2. Suntory Institute for Bioorganic Research, 1-1-1 Wakayamadai, Shimamoto-cho, Mshima-gun, Osaka, 618-8503, Japan

    Keiko Shimamoto

Authors
  1. Helle S. Waagepetersen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Keiko Shimamoto
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Arne Schousboe
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Waagepetersen, H.S., Shimamoto, K. & Schousboe, A. Comparison of Effects of DL-Threo-β-Benzyloxyaspartate (DL-TBOA) and L-Trans-Pyrrolidine-2,4-Dicarboxylate (t-2,4-PDC) on Uptake and Release of [3H]D-Aspartate in Astrocytes and Glutamatergic Neurons. Neurochem Res 26, 661–666 (2001). https://doi.org/10.1023/A:1010939304104

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1010939304104

Search

Navigation