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Study of the Mechanism of the Neuron Sensitization to the Repeated Glutamate Challenge

  • R. R. Sharipov
  • I. A. Krasilnikova
  • V. G. Pinelis
  • L. R. Gorbacheva
  • A. M. SurinEmail author
ARTICLES

Abstract

Exposure of cultured neurons to high concentrations of Glu leads to a strong depolarization of mitochondria, which develops synchronously with the secondary rise in the intracellular Ca2+ concentration (delayed calcium deregulation, DCD). In this study, using the primary culture of rat cerebellar neurons, we investigated the mechanism of neuronal sensitization, which manifests itself in the reduction of latent periods of DCD during repeated exposures to Glu. It was shown that the most likely cause of sensitization is the inability of mitochondria to maintain a high transmembrane potential (ΔΨm) as a result of an increase in the proton conductivity of the internal mitochondrial membrane, but not the opening of the mitochondrial permeability transition pore in the inner mitochondrial membrane. Mitochondrial dysfunction reduces the production of ATP, leading to the inability of neurons to quickly restore the concentration of Na+, ATP, and NADH in the intervals between successive Glu administrations. One of the reasons that aggravate the dysfunction of mitochondria and contribute to the sensitization of neurons to the repeated action of Glu is Ca2+ accumulated in the mitochondria during the first glutamate impact.

Keywords:

glutamate sensitization calcium sodium ATP mitochondria neuronal cultures 

Notes

ACKNOWLEDGMENTS

The authors thank Prof. H. Imamura who kindly provided plasmid Ateam1.03 encoding ATP sensor. The work was supported by the Russian Foundation for Basic Research (project nos. 16-04-00792 and 16-04-01869) and by the Russian Science Foundation (project no. 17-15-01487).

COMPLIANCE WITH ETHICAL STANDARDS

Conflict of interests. The authors declare that they have no conflict of interest.

Statement on the welfare of animals. Experiments involving animals were carried out in accordance with ethical principles and normative documents recommended by the European Convention on the protection of vertebrate animals used in experiments (Guide for the Care and Use of Laboratory Animals: Eighth Edition, 2010), as well as in compliance with the Regulations of the appropriate laboratory practice as approved by the order no. 199n of 01.04.2016 of the Russian Federation Ministry of Health.

REFERENCES

  1. 1.
    Wang Y., Qin Z. 2010. Molecular and cellular mechanisms of excitotoxic neuronal death. Apoptosis. 15 (11), 1382–1402.CrossRefGoogle Scholar
  2. 2.
    Zhou Y., Danbolt N.C. 2014. Glutamate as a neurotransmitter in the healthy brain. J. Neural. Transm. 121 (8), 799–817.CrossRefGoogle Scholar
  3. 3.
    Gudiño-Cabrera G., Ureña-Guerrero M.E., Rivera-Cervantes M.C., Feria-Velasco A.I., Beas-Zárate C. 2014. Excitotoxicity triggered by neonatal monosodium glutamate treatment and blood–brain barrier function. Arch. Med. Res. 45 (8), 653–659.CrossRefGoogle Scholar
  4. 4.
    Plitman E., Nakajima S., de la Fuente-Sandoval C., Gerretsen P., Chakravarty M.M., Kobylianskii J., Chung J.K., Caravaggio F., Iwata Y., Remington G., Graff-Guerrero A. 2014. Glutamate-mediated excitotoxicity in schizophrenia: A review. Eur. Neuropsychopharmacol. 24 (10), 1591–1605.CrossRefGoogle Scholar
  5. 5.
    Kostic M., Zivkovic N., Stojanovic I. 2013. Multiple sclerosis and glutamate excitotoxicity. Rev. Neurosci. 24 (1), 71–88.CrossRefGoogle Scholar
  6. 6.
    Verkhratsky A., Kirchhoff F. 2007. NMDA receptors in glia. Neurosci. 13 (1), 28–37.Google Scholar
  7. 7.
    Gerkau N.J., Rakers C., Petzold G.C., Rose C.R. 2017. Differential effects of energy deprivation on intracellular sodium homeostasis in neurons and astrocytes. J. Neurosci. Res. 95 (11), 2275–2285.CrossRefGoogle Scholar
  8. 8.
    Olney J.W., Gubareff T. 1978. Glutamate neurotoxicity and Huntington’s chorea. Nature. 271 (5645), 557–559.CrossRefGoogle Scholar
  9. 9.
    Choi D.W. 1988. Glutamate neurotoxicity and diseases of the nervous system. Neuron. 1 (8), 623–634.CrossRefGoogle Scholar
  10. 10.
    Nicholls D.G., Budd S.L. 2000. Mitochondria and neuronal survival. Physiol. Rev. 80 (1), 315–360.CrossRefGoogle Scholar
  11. 11.
    Khodorov B. 2004. Glutamate-induced deregulation of calcium homeostasis and mitochondrial dysfunction in mammalian central neurones. Prog. Biophys. Mol. Biol. 86 (2), 279–351.CrossRefGoogle Scholar
  12. 12.
    Duchen M.R. 2012. Mitochondria, calcium-dependent neuronal death and neurodegenerative disease. Pflügers Arch. Eur. J. Physiol. 464 (1), 111–121.CrossRefGoogle Scholar
  13. 13.
    Kornberg H. 2000. Krebs and his trinity of cycles. Nat. Rev. Mol. Cell Biol. 1 (3), 225–228.CrossRefGoogle Scholar
  14. 14.
    Skulachev V.P., Bogachev A.V., Kasparinskii F.O. 2010. Membrannaya bioenergetika (Membrane bioenergetics). M., MSU.Google Scholar
  15. 15.
    Nicholls D.G., Ferguson S.J. 2013. Bioenergetics. London: Elsevier, Acad. Press. 4 ed.Google Scholar
  16. 16.
    Tymianski M., Charlton M.P., Carlen P.L., Tator C.H. 1993. Source specificity of early calcium neurotoxicity in cultured embryonic spinal neurons. J. Neurosci. 13 (5), 2085–2104.CrossRefGoogle Scholar
  17. 17.
    Nicholls D.G., Ward M.W. 2000. Mitochondrial membrane potential and neuronal glutamate excitotoxicity: Mortality and millivolts. Trends Neurosci. 23 (4), 166–174.CrossRefGoogle Scholar
  18. 18.
    Abramov A.Y., Duchen M.R. 2010. Impaired mitochondrial bioenergetics determines glutamate-induced delayed calcium deregulation in neurons. Biochim. Biophys. Acta. Gen. Subj. 1800 (3), 297–304.Google Scholar
  19. 19.
    Ames A. 2000. CNS energy metabolism as related to function. Brain Res. Rev. 34 (1–2), 42–68.CrossRefGoogle Scholar
  20. 20.
    Kiedrowski L., Brooker G., Costa E., Wroblewski J.T. 1994. Glutamate impairs neuronal calcium extrusion while reducing sodium gradient. Neuron. 12 (2), 295–300.CrossRefGoogle Scholar
  21. 21.
    Kato H., Araki T., Kogure K. 1992. Repeated focal cerebral ischemia in gerbils is associated with development of infarction. Brain Res. 596 (1–2), 315–319.CrossRefGoogle Scholar
  22. 22.
    Dambinova S.A., Shikuev A.V., Weissman J.D., Mullins J.D. 2013. AMPAR peptide values in blood of nonathletes and club sport athletes with concussions. Mil. Med. 178 (3), 285–290.CrossRefGoogle Scholar
  23. 23.
    Nakata N., Kato H., Kogure K. 1993. Effects of repeated cerebral ischemia on extracellular amino acid concentrations measured with intracerebral microdialysis in the gerbil hippocampus. Stroke. 24 (3), 458–464.CrossRefGoogle Scholar
  24. 24.
    Ueda Y., Obrenovitch T.P., Lok S.Y., Sarna G.S., Symon L. 1992. Changes in extracellular glutamate concentration produced in the rat striatum by repeated ischemia. Stroke. 23 (8), 1125–1130.CrossRefGoogle Scholar
  25. 25.
    Khodorov B.I., Mikhailova M.M., Bolshakov A.P., Surin A.M., Sorokina E.G., Rozhnev S.A., Pinelis V.G. 2012. Dramatic effect of glycolysis inhibition on the cerebellar granule cells bioenergetics. Biochem. (Moscow) Suppl. Ser. A Membr. Cell Biol. 6 (2), 186–197.Google Scholar
  26. 26.
    Surin A.M., Gorbacheva L.R., Savinkova I.G., Sharipov R.R., Khodorov B.I., Pinelis V.G. 2014. Study on ATP concentration changes in cytosol of individual cultured neurons during glutamate-induced deregulation of calcium homeostasis. Biochemistry (Moscow). 79 (2), 146–157.CrossRefGoogle Scholar
  27. 27.
    Imamura H., Huynh Nhat K.P., Togawa H., Saito K., Iino R., Kato-Yamada Y., Nagai T., Noji H. 2009. Visualization of ATP levels inside single living cells with fluorescence resonance energy transfer-based genetically encoded indicators. Proc. Natl. Acad. Sci. USA. 106 (37), 15651–15656.CrossRefGoogle Scholar
  28. 28.
    Schindelin J., Arganda-Carreras I., Frise E., Kaynig V., Longair M., Pietzsch T., Preibisch S., Rueden C., Saalfeld S., Schmid B., Tinevez J.-Y.J.-Y., White D.J., Hartenstein V., Eliceiri K., Tomancak P., Cardona A., Liceiri K., Tomancak P., A. C. 2012. Fiji: An open source platform for biological image analysis. Nat. Methods. 9 (7), 676–682.CrossRefGoogle Scholar
  29. 29.
    Keelan J., Vergun O., Duchen M.R. 1999. Excitotoxic mitochondrial depolarisation requires both calcium and nitric oxide in rat hippocampal neurons. J. Physiol. 520 (3), 797–813.CrossRefGoogle Scholar
  30. 30.
    Vergun O., Keelan J., Khodorov B.I., Duchen M.R. 1999. Glutamate-induced mitochondrial depolarisation and perturbation of calcium homeostasis in cultured rat hippocampal neurones. J. Physiol. 519 (2), 451–466.CrossRefGoogle Scholar
  31. 31.
    Rueda C.B., Llorente-Folch I., Traba J., Amigo I., Gonzalez-Sanchez P., Contreras L., Juaristi I., Martinez-Valero P., Pardo B., Del Arco A., Satrustegui J. 2016. Glutamate excitotoxicity and Ca2+-regulation of respiration: Role of the Ca2+ activated mitochondrial transporters (CaMCs). Biochim. Biophys. Acta. 1857 (8), 1158–1166.Google Scholar
  32. 32.
    Nicholls D.G. 2008. Oxidative stress and energy crises in neuronal dysfunction. Ann. N.Y. Acad. Sci. 1147 (1), 53–60.CrossRefGoogle Scholar
  33. 33.
    Bernardi P. 1999. Mitochondrial transport of cations: Channels, exchangers, and permeability transition. Physiol. Rev. 79 (4), 1127–1155.CrossRefGoogle Scholar
  34. 34.
    Wabnitz A.V., Storozhevykh T.P., Pinelis V.G., Khodorov B.I. 2005. The permeability transition pore is not a prerequisite for glutamate-induced calcium deregulation and mitochondrial depolarization in brain neurons. Biol. Membr. (Rus.). 22 (4), 378–382.Google Scholar
  35. 35.
    Bolshakov A.P., Mikhailova M.M., Szabadkai G., Pinelis V.G., Brustovetsky N., Rizzuto R., Khodorov B.I. 2008. Measurements of mitochondrial pH in cultured cortical neurons clarify contribution of mitochondrial pore to the mechanism of glutamate-induced delayed Ca2+ deregulation. Cell Calcium. 43 (6), 602–614.CrossRefGoogle Scholar
  36. 36.
    Lazarewicz J.W., Wroblewski J.T., Costa E. 1990. N‑methyl-D-aspartate-sensitive glutamate receptors induce calcium-mediated arachidonic acid release in primary cultures of cerebellar granule cells. J. Neurochem. 55 (6), 1875–1881.CrossRefGoogle Scholar
  37. 37.
    Surin A.M., Bolshakov A.P., Mikhailova M.M., Sorokina E.G., Senilova Ya.E., Pinelis V.G., Khodorov B.I. 2006. Arachidonic acid enhances intracellular [Ca2+]i increase and mitochondrial depolarization induced by glutamate in cerebellar granule cells. Biochemistry (Moscow). 71 (8), 864–870.CrossRefGoogle Scholar
  38. 38.
    Mironova G.D., Saris N.-E.L., Belosludtseva N.V., Agafonov A.V., Elantsev A.B., Belosludtsev K.N. 2015. Involvement of palmitate/Ca2+(Sr2+)-induced pore in the cycling of ions across the mitochondrial membrane. Biochim. Biophys. Acta. Biomembr. 1848 (2), 488–495.Google Scholar
  39. 39.
    Belosludtsev K.N., Belosludtseva N. V., Agafonov A. V., Astashev M.E., Kazakov A.S., Saris N.-E.L., Miro-nova G.D. 2014. Ca2+-dependent permeabilization of mitochondria and liposomes by palmitic and oleic acids: A comparative study. Biochim. Biophys. Acta. Biomembr. 1838 (10), 2600–2606.Google Scholar
  40. 40.
    Abramov A.Y., Duchen M.R. 2008. Mechanisms underlying the loss of mitochondrial membrane potential in glutamate excitotoxicity. Biochim. Biophys. Acta. 1777 (7–8), 953–964.Google Scholar
  41. 41.
    Surin A.M., Zobova S.N., Tukhbatova G.R., Senilova Ya.E., Pinelis V.G., Khodorov B.I. 2010. Changes in mitochondrial NAD(P)H and glutamate-induced delayed calcium deregulation in cultured rat cerebellar neurons. Biochem. (Moscow) Suppl. Series A: Membr. Cell Biol. 4 (1), 32–37.Google Scholar
  42. 42.
    Surin A.M., Krasilnikova I.A., Pinelis V.G., Khodorov B.I. 2014. Study of the relationships between glutamate-induced delayed calcium deregulation, mitochondrial depolarization, and subsequent neuronal death. Patogenez (Rus.). 12 (4), 40–46.Google Scholar
  43. 43.
    Yoshida T., Alfaqaan S., Sasaoka N., Imamura H. 2017. Application of FRET-based biosensor “ATeam” for visualization of ATP levels in the mitochondrial matrix of living mammalian cells. Methods Mol. Biol. 1567, 231–243.CrossRefGoogle Scholar
  44. 44.
    Liemburg-Apers D.C., Imamura H., Forkink M., Nooteboom M., Swarts H.G., Brock R., Smeitink J.A.M., Willems P.H.G.M., Koopman W.J.H. 2011. Quantitative glucose and ATP sensing in mammalian cells. Pharm. Res. 28 (11), 2745–2757.CrossRefGoogle Scholar
  45. 45.
    Brittain M.K., Brustovetsky T., Sheets P.L., Brittain J.M., Khanna R., Cummins T.R., Brustovetsky N. 2012. Delayed calcium dysregulation in neurons requires both the NMDA receptor and the reverse Na+/Ca2+ exchanger. Neurobiol. Dis. 46 (1), 109–117.CrossRefGoogle Scholar
  46. 46.
    Kiedrowski L. 1999. N-methyl-D-aspartate excitotoxicity: Relationships among plasma membrane potential, Na+/Ca2+ exchange, mitochondrial Ca2+ overload, and cytoplasmic concentrations of Ca2+, H+, and K+. Mol. Pharmacol. 56 (3), 619–632.CrossRefGoogle Scholar
  47. 47.
    Kiedrowski L. 2007. NCX and NCKX operation in ischemic neurons. Ann. N.Y. Acad. Sci. 1099 (1), 383–395.CrossRefGoogle Scholar
  48. 48.
    Pinelis V.G., Segal M., Greenberger V., Khodorov B.I. 1994. Changes in cytosolic sodium caused by a toxic glutamate treatment of cultured hippocampal neurons. Biochem. Mol. Biol. Int. 32 (3), 475–482.Google Scholar
  49. 49.
    Nicholls D.G., Vesce S., Kirk L., Chalmers S. 2003. Interactions between mitochondrial bioenergetics and cytoplasmic calcium in cultured cerebellar granule cells. Cell Calcium. 34 (0143–4160 (Print)), 407–424.Google Scholar
  50. 50.
    Brocard J.B., Tassetto M., Reynolds I.J. 2001. Quantitative evaluation of mitochondrial calcium content in rat cortical neurones following a glutamate stimulus. J. Physiol. 531 (Pt 3), 793–805.CrossRefGoogle Scholar
  51. 51.
    Chernyak B.V. 1999. Induction of the non-selective mitochondrial pore in lymphoid cells. 2. Intact rat thymocytes. Biochemistry (Moscow). 64 (8), 922–928.Google Scholar
  52. 52.
    Xu-Friedman M.A., Regehr W.G. 1999. Presynaptic strontium dynamics and synaptic transmission. Biophys. J. 76 (4), 2029–2042.CrossRefGoogle Scholar
  53. 53.
    Severin F.F., Severina I.I., Antonenko Y.N., Rokitskaya T.I., Cherepanov D.A., Mokhova E.N., Vyssokikh M.Y., Pustovidko A.V., Markova O.V., Yaguzhinsky L.S., Korshunova G.A., Sumbatyan N.V., Skulachev M.V., Skulachev V.P. 2010. Penetrating cation/fatty acid anion pair as a mitochondria-targeted protonophore. Proc. Natl. Acad. Sci. USA. 107 (2), 663–668.CrossRefGoogle Scholar
  54. 54.
    Surin A.M., Sharipov R.R., Krasilnikova I.A., Boyarkin D.P., Lisina O.Yu., Gorbacheva L.R., Avetisian A.V., Pinelis V.G. 2017. Disruption of functional activity of mitochondria during the MTT assay of viability of cultured neurons. Biochamistry (Moscow). 82 (6), 737–749.CrossRefGoogle Scholar
  55. 55.
    Jekabsons M.B., Nicholls D.G. 2004. In situ respiration and bioenergetic status of mitochondria in primary cerebellar granule neuronal cultures exposed continuously to glutamate. J. Biol. Chem. 279 (31), 32989–33000.CrossRefGoogle Scholar
  56. 56.
    Jekabsons M.B., Nicholls D.G. 2006. Bioenergetic analysis of cerebellar granule neurons undergoing apoptosis by potassium/serum deprivation. Cell Death Differ. 13 (9), 1595–1610.CrossRefGoogle Scholar
  57. 57.
    Liu D., Gharavi R., Pitta M., Gleichmann M., Mattson M.P. 2009. Nicotinamide prevents NAD+ depletion and protects neurons against excitotoxicity and cerebral ischemia: NAD+ consumption by SIRT1 may endanger energetically compromised neurons. Neuromolec. Med. 11 (1), 28–42.CrossRefGoogle Scholar
  58. 58.
    Pinelis V.G., Bykova L.P., Bogachev A.P., Isaev N.K., Viktorov I. V, Khodorov B.I. 1997. Toxic effect of glutamate on cultured cerebellar granular cells reduces the intracellular level of ATP. The role of Ca2+ ions. Bull. Eksp. Biol. Med. (Rus.). 123 (2), 162–164.Google Scholar
  59. 59.
    Kovacs-Bogdan E., Sancak Y., Kamer K.J., Plovanich M., Jambhekar A., Huber R.J., Myre M.A., Blower M.D., Mootha V.K. 2014. Reconstitution of the mitochondrial calcium uniporter in yeast. Proc. Natl. Acad. Sci. USA. 111 (24), 8985–8990.CrossRefGoogle Scholar
  60. 60.
    McIntosh D.B., Woolley D.G., Vilsen B., Andersen J.P. 1996. Mutagenesis of segment 487Phe-Ser-Arg-Asp-Arg-Lys492 of sarcoplasmic reticulum Ca2+-ATPase produces pumps defective in ATP binding. J. Biol. Chem. 271 (42), 25778–25789.CrossRefGoogle Scholar
  61. 61.
    Saris N.-E.L., Carafoli E. 2005. A historical review of cellular calcium handling, with emphasis on mitochondria. Biochemistry (Moscow). 70 (2), 187–194.CrossRefGoogle Scholar
  62. 62.
    Vyklicky V., Korinek M., Smejkalova T., Balik A., Krausova B., Kaniakova M., Lichnerova K., Cerny J., Krusek J., Dittert I., Horak M., Vyklicky L. 2014. Structure, function, and pharmacology of NMDA receptor channels. Physiol. Res. 63 (Suppl. 1), 191–203.Google Scholar
  63. 63.
    Novelli A., Reilly J.A., Lysko P.G., Henneberry R.C. 1988. Glutamate becomes neurotoxic via the N-methyl-D-aspartate receptor when intracellular energy levels are reduced. Brain Res. 451 (1–2), 205–212.CrossRefGoogle Scholar
  64. 64.
    Rose C.R., Konnerth A. 2001. NMDA receptor-mediated Na+ signals in spines and dendrites. J. Neurosci. 21 (12), 4207–4214.CrossRefGoogle Scholar
  65. 65.
    Cortassa S., Aon M.A., Marbán E., Winslow R.L., O’Rourke B. 2003. An integrated model of cardiac mitochondrial energy metabolism and calcium dynamics. Biophys. J. 84 (4), 2734–2755.CrossRefGoogle Scholar
  66. 66.
    Cortassa S., Aon M.A. 2012. Computational modeling of mitochondrial function. Methods Mol. Biol. 810, 311–326.CrossRefGoogle Scholar

Copyright information

© Pleiades Publishing, Ltd. 2018

Authors and Affiliations

  • R. R. Sharipov
    • 1
  • I. A. Krasilnikova
    • 2
  • V. G. Pinelis
    • 2
  • L. R. Gorbacheva
    • 3
    • 4
  • A. M. Surin
    • 1
    • 2
    • 3
    Email author
  1. 1.Scientific Center of Children’s HealthMoscowRussia
  2. 2.Institute of General Pathology and PathophysiologyMoscowRussia
  3. 3.Pirogov Russian National Research Medical UniversityMoscowRussia
  4. 4.Biology Department, Moscow Lomonosov State UniversityMoscowRussia

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