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Selective inhibition of NAALADase, which converts NAAG to glutamate, reduces ischemic brain injury

Nature Medicine volume 5, pages 1396–1402 (1999)Cite this article

Abstract

We describe here a new strategy for the treatment of stroke, through the inhibition of NAALADase (N-acetylated-α-linked-acidic dipeptidase), an enzyme responsible for the hydrolysis of the neuropeptide NAAG (N-acetyl-aspartyl-glutamate) to N-acetyl-aspartate and glutamate. We demonstrate that the newly described NAALADase inhibitor 2-PMPA (2-(phosphonomethyl)pentanedioic acid) robustly protects against ischemic injury in a neuronal culture model of stroke and in rats after transient middle cerebral artery occlusion. Consistent with inhibition of NAALADase, we show that 2-PMPA increases NAAG and attenuates the ischemia-induced rise in glutamate. Both effects could contribute to neuroprotection. These data indicate that NAALADase inhibition may have use in neurological disorders in which excessive excitatory amino acid transmission is pathogenic.

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Figure 1: The NAALADase inhibitor 2-PMPA and a less-active analog 2-PMSA.
Figure 2: Protective effects of NAALADase inhibition in a cell culture model of cerebral ischemia.
Figure 3: Protective effect of NAALADase inhibition in a rat MCAO model of focal ischemia.
Figure 4: NAALADase inhibition alters glutamate and NAAG levels after MCAO.
Figure 5: Neurotoxicity of high affinity glutamate receptor blockade compared with NAALADase inhibition.

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References

  1. Benveniste, H., Drejer, J., Schousboe, A. & Diemer, N.H. Elevation of the extracellular concentrations of glutamate and aspartate in rat hippocampus during transient cerebral ischemia monitored by intracerebral microdialysis. J. Neurochem. 43, 1369–1374 (1984).

    Article  CAS  Google Scholar 

  2. Butcher, S.P., Bullock, R., Graham, D.I. & McCulloch, J. Correlation between amino acid release and neuropathologic outcome in rat brain following middle cerebral artery occlusion. Stroke 21, 1727–1733 (1990).

    Article  CAS  Google Scholar 

  3. Fagg, G.E. & Foster, A.C. Excitatory amino acid synaptic mechanisms and neurological function. Trends Pharmacol. Sci. 7, 357–363 (1986).

    Article  CAS  Google Scholar 

  4. Rothman, S.M. & Olney, J.W. Glutamate and the pathophysiology of hypoxic-ischemic brain death. Ann. Neurol. 19, 105–111 (1986).

    Article  CAS  Google Scholar 

  5. Choi, D.W. & Rothman, S.M. The role of glutamate neurotoxicity in hypoxic-ischemic neuronal death. Annu. Rev. Neurosci. 13, 171–182 (1990).

    Article  CAS  Google Scholar 

  6. Meldrum, B.S. Protection against ischaemic neuronal damage by drugs acting on excitatory neurotransmission. Cerebrovasc. Brain Metab. Rev. 2, 27–57 (1990).

    CAS  PubMed  Google Scholar 

  7. Wahlgren, N.G. in International Review of Neurobiology: Neuroprotective Agents and Cerebral Ischemia (eds. Green, A.R. & Cross, A.J.) 337–363 (Academic, San Diego, 1997).

    Google Scholar 

  8. Coyle, J.T. et al. in Excitatory Amino Acids (eds. Meldrum, B.S., Moroni, F., Simon, R.P. & Woods, J.H.) 69–77 (Raven, New York, 1991).

    Google Scholar 

  9. Coyle, J.T. The nagging question of the function of N-acetylaspartylglutamate. Neurobiol. Dis. 4, 231–238 (1997).

    Article  CAS  Google Scholar 

  10. Tsai, G., Stauch, B.L., Vornov, J.J., Deshpande, J.K. & Coyle, J.T. Selective release of N- acetylaspartylglutamate from rat optic nerve terminals in vivo. Brain Res. 518, 313–316 (1991).

    Article  Google Scholar 

  11. Wrobleska, B. et al. NAAG selectively activates mGluR3 receptors in transfected cells. J. Neurochem. 69, 174–181 (1997).

    Article  Google Scholar 

  12. Vallivullah, H.M., Lancaster, J., Sweetnam, P.M. & Neale, J.H. High concentrations of N- acetylaspartylglutamate and AMPA, kainate, and NMDA binding sites. J. Neurochem. 63, 1714–1719 (1994).

    Article  Google Scholar 

  13. Sekiguchi, M., Okamoto, K. & Sakai, Y. Low concentration N-acetylaspartylglutamate suppresses the climbing fiber response of Purkinje cells in Guinea pig cerebellar slices and the responses to excitatory amino acids of Xenopus laevis oocytes injected with cerebellar mRNA. Brain Res. 482, 87–96 (1989).

    Article  CAS  Google Scholar 

  14. Puttfarcken, P.S., Montgomery, D., Coyle, J.T. & Werling, L.L. N-acetyl-L-aspartyl-L-glutamate (NAAG) modulation of NMDA-stimulated [3H]norepinephrine release from rat hippocampal slices. Pharmacol. Exp. Ther. 266, 796–803 (1993).

    CAS  Google Scholar 

  15. Slusher, B.S. et al. Rat brain N-acetylated-α-linked acidic dipeptidase activity. Purification and immunologic characterization. J. Biol. Chem. 265, 21297–21301 (1990).

    CAS  PubMed  Google Scholar 

  16. Robinson, M.B., Blakely, R.D., Couto, R. & Coyle, J.T. Hydrolysis of the brain dipeptide N-acetyl- L-aspartyl-L-glutamate. Identification and characterization of a novel N-acetylated alpha-linked acidic dipeptidase activity from rat brain. J. Biol. Chem. 262, 14498–14506 (1987).

    CAS  PubMed  Google Scholar 

  17. Stauch-Slusher, B., Robinson, M.B., Forloni, G., Tsai, G. & Coyle, J.T. The effects of N-acetylated-α-linked acidic dipeptidase (NAALADase) inhibitors on [3H]NAAG catabolism in vivo. Neuro. Sci. Lett. 100, 295–300 (1989).

    Article  Google Scholar 

  18. Fuhrman, S., Palkovits, M., Cassidy, M. & Neale, J.H. The regional distribution of N- acetylaspartylglutamate (NAAG) and peptidase activity against NAAG in the rat nervous system. J. Neurochem. 62, 275–281 (1994).

    Article  CAS  Google Scholar 

  19. Meyerhoff, J.L., Carter, R.E., Yourick, D.L., Stauch-Slusher, B. & Coyle, J.T. Genetically epilepsy- prone rats have increased brain regional activity of an enzyme which liberates glutamate from N-acetylaspartylglutamate. Brain Res. 593, 140–143 (1992).

    Article  CAS  Google Scholar 

  20. Bruno, V. et al. Activation of class II or III metabotropic glutamate receptors protects cultured cortical neurons against excitotoxic degeneration. Eur. J. Pharm. 7, 1906–1913 (1995).

    CAS  Google Scholar 

  21. Serval, V. et al. Competitive inhibition of N-acetylated-α-linked acidic dipeptidase activity by N-acetyl-L-aspartyl-α-linked-L-glutamate. J. Neurochem. 55, 39–46 (1990).

    Article  CAS  Google Scholar 

  22. Jackson, P.F. et al. Design, synthesis, and biological activity of a potent inhibitor of the neuropeptidase N-acetylated-α-linked acidic dipeptidase. J. Med. Chem. 39, 619–622 (1996).

    Article  CAS  Google Scholar 

  23. Vornov, J.J. Toxic NMDA-receptor activation occurs during recovery in a tissue culture model of ischemia. J. Neurochem. 65, 1681–1691 (1995).

    Article  CAS  Google Scholar 

  24. Vornov, J.J., Thomas, A.G. & Jo, D. Protective effects of extracellular acidosis and blockade of sodium/hydrogen ion exchange during recovery from metabolic inhibition in neuronal tissue culture. J. Neurochem. 67, 2379–2389 (1996).

    Article  CAS  Google Scholar 

  25. Takahashi, H. et al. PPBP [4- phenyl-1-(4-phenylbutyl)piperidine] decreases brain injury after transient focal ischemic in rats. Stroke 27, 2120–2123 (1996).

    Article  CAS  Google Scholar 

  26. Britton, P., Lu, M.X.C., Laskosky, M.S. & Tortella, F.C. Dextromethorphan protects against cerebral injury following transient, but not permanent, focal ischemia in rats. Life Sci. 60 (20), 1729–1740 (1997).

    Article  CAS  Google Scholar 

  27. Longa, E.Z., Weinstein, P.R., Carlson, S. & Cummins, R. Reversible middle cerebral artery occlusion without craniectomy in rats. Stroke 20: 84–91 (1989).

    Article  CAS  Google Scholar 

  28. Small, D.L. & Buchan, A.M. in International Review of Neurobiology: Neuroprotective Agents and Cerebral Ischemia (eds. Green, A.R. & Cross, A.J.) 137–171 (Academic, San Diego, 1997).

    Google Scholar 

  29. Britton, P., Whitton, P.S., Fowler, L.J. & Bowery, N.G. Tetanus toxin-induced effects on extracellular amino acid levels in rat hippocampus: an in vivo microdialysis study. J. Neurochem. 67, 324–329 (1996).

    Article  CAS  Google Scholar 

  30. Robinson, M.B., Djali, S. & Buchhalter, J.R. Inhibition of glutamate uptake with L-trans- pyrrolidine-2,4-dicarboxylate potentiates glutamate toxicity in primary hippocampal cultures. J. Neurochem. 61, 2099–2103 (1993).

    Article  CAS  Google Scholar 

  31. Zollinger, M., Amsler, U. & Brauchli, J. Quantification of N-acetylaspartylglutamate, an N- terminal blocked dipeptide neurotransmitter candidate, in brain slices superfusates by gas chromatography-mass spectrometry. J. Chromatogr. 532 (1), 27–36 (1990).

    Article  CAS  Google Scholar 

  32. Goda, H. et al. Modulation of ischemia-evoked release of excitatory and inhibitory amino acids by adenosine A1 receptor agonists. Eur. J. Pharm. 357, 149–155 (1998).

    Article  CAS  Google Scholar 

  33. Muir, K.W. & Lees, K.R. A randomized, double-blind, placebo controlled ascending dose tolerance study of 619C89 in acute stroke. Stroke 26, 503–513 (1995).

    Article  CAS  Google Scholar 

  34. Wozniak, D.F. et al. MK-801 neurotoxicity in male mice: histologic effects and chronic impairment in spatial learning. Brain Res. 707, 165–179 (1996).

    Article  CAS  Google Scholar 

  35. Fix, A.S., Ross, J.F., Stitzel, S.R. & Switzer, R.C. Integrated evaluation of central nervous system lesions: stains for neurons, astrocytes and microglia reveal the spatial and temporal features of MK-801-induced neuronal necrosis in the rat cerebral cortex. Toxicol. Pathol. 24(3), 291–304 (1996).

    Article  CAS  Google Scholar 

  36. Chen, J., Graham, S.H. & Simon, R.P. A comparison of the effects of a sodium channel blocker and an NMDA antagonist upon extracellular glutamate in rat focal ischemia. Brain Res. 699, 121–124 (1995).

    Article  CAS  Google Scholar 

  37. Johansen, P.A. et al. Type 4A metabotropic glutamate receptor: identification of new potent agonists and differentiation from the L-(+)-2-amino-4-phosphonobutanoic acid-sensitive receptor in the lateral perforant pathway in rats. Mol. Pharm. 48, 140–149 (1995).

    CAS  Google Scholar 

  38. Thomsen, C., Boel, E. & Suzdak, P.D. Actions of phenylglycine analogs at subtypes of the metabotropic glutamate receptor family. Eur. J. Pharm. 267, 77–84 (1994).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors thank S. Snyder for his insight and discussions in the preparation of this manuscript and E. Lea and E. Lisska for their assistance in the amino-acid analyses. All experiments were supported by Guilford Pharmaceuticals.

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Authors and Affiliations

  1. Department of Research, Guilford Pharmaceuticals, Baltimore, 21224, Maryland, USA

    Barbara S. Slusher, James J. Vornov, Ajit G. Thomas, X.-C. May Lu, Krystyna M. Wozniak, Beth M. Potter & Paul F. Jackson

  2. Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, 21205, Maryland, USA

    James J. Vornov, Ajit G. Thomas & Anish Bhardwaj

  3. Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, 21205, Maryland, USA

    Patricia D. Hurn, Izumi Harukuni & Richard J. Traystman

  4. Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, 19104-4318, Pennsylvania, USA

    Michael B. Robinson & Marc Yudkoff

  5. Division of Neurosciences, Walter Reed Army Institute of Research, Washington, DC, 20307-5100, USA

    Paul Britton & Frank C. Tortella

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  1. Barbara S. Slusher
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Correspondence to Barbara S. Slusher.

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Slusher, B., Vornov, J., Thomas, A. et al. Selective inhibition of NAALADase, which converts NAAG to glutamate, reduces ischemic brain injury. Nat Med 5, 1396–1402 (1999). https://doi.org/10.1038/70971

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