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Acidosis and 5-(N-ethyl-N-isopropyl)amiloride (EIPA) attenuate zinc/kainate toxicity in cultured cerebellar granule neurons

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Abstract

Cultured cerebellar granule neurons (CGNs) are resistant to the toxic effect of ZnCl2 (0.005 mM, 3 h) and slightly sensitive to the effect of kainate (0.1 mM, 3 h). Simultaneous treatment of CGNs with kainate and ZnCl2 caused intensive neuronal death, which was attenuated by external acidosis (pH 6.5) or 5-(N-ethyl-N-isopropyl)amiloride (EIPA, Na+/H+ exchange blocker, 0.03 mM). Intracellular zinc and calcium ion concentrations ([Zn2+]i and [Ca2+]i) were increased under the toxic action of kainate + ZnCl2, this effect being significantly decreased on external acidosis and increased in case of EIPA addition. Neuronal Zn2+ imaging demonstrated that EIPA increases the cytosolic concentration of free Zn2+ on incubation in Zn2+-containing solution. These data imply that acidosis reduces ZnCl2/kainate toxic effects by decreasing Zn2+ entry into neurons, and EIPA prevents zinc stores from being overloaded with zinc.

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References

  1. Toth, K. (2011) Zinc in neurotransmission, Annu. Rev. Nutr., 31, 139–153.

    Article  CAS  PubMed  Google Scholar 

  2. Shuttleworth, C. W., and Weiss, J. H. (2011) Zinc: new clues to diverse roles in brain ischemia, Trends Pharmacol. Sci., 32, 480–486.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  3. Pivovarova, N. B., Stanika, R. I., Kazanina, G., Villanueva, I., and Andrews, S. B. (2014) The interactive roles of zinc and calcium in mitochondrial dysfunction and neurodegeneration, J. Neurochem., 128, 592–602.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  4. Zhao, Y., Pan, R., Li, S., Luo, Y., Yan, F., Yin, J., Qi, Z., Yan, Y., Ji, X., and Liu, K. J. (2014) Chelating intracellularly accumulated zinc decreased ischemic brain injury through reducing neuronal apoptotic death, Stroke, 45, 1139–1147.

    Article  CAS  PubMed  Google Scholar 

  5. Dineley, K. E., Votyakova, T. V., and Reynolds, I. J. (2003) Zinc inhibition of cellular energy production: implications for mitochondria and neurodegeneration, J. Neurochem., 85, 563–570.

    Article  CAS  PubMed  Google Scholar 

  6. Sensi, S. L., Paoletti, P., Bush, A. I., and Sekler, I. (2009) Zinc in the physiology and pathology of the CNS, Nat. Rev. Neurosci., 10, 780–791.

    Article  CAS  PubMed  Google Scholar 

  7. Sheline, C. T., Zhu, J., Zhang, W., Shi, C., and Cai, A. L. (2013) Mitochondrial inhibitor models of Huntington’s disease and Parkinson’s disease induce zinc accumulation and are attenuated by inhibition of zinc neurotoxicity in vitro or in vivo, Neurodegener. Dis., 11, 49–58.

    Article  PubMed  Google Scholar 

  8. Weiss, J. H., Hartley, D. M., Koh, J. Y., and Choi, D. W. (1993) AMPA receptor activation potentiates zinc neurotoxicity, Neuron, 10, 43–49.

    Article  CAS  PubMed  Google Scholar 

  9. Colvin, R. A. (2002) pH dependence and compartmentalization of zinc transported across plasma membrane of rat cortical neurons, Am. J. Physiol. Cell Physiol., 282, 317–329.

    Article  Google Scholar 

  10. Isaev, N. K., Stelmashook, E. V., Lukin, S. V., Freyer, D., Mergenthaler, P., and Zorov, D. B. (2010) Acidosisinduced zinc-dependent death of cultured cerebellar granule neurons, Cell. Mol. Neurobiol., 30, 877–883.

    Article  CAS  PubMed  Google Scholar 

  11. Kiedrowski, L. (2014) Proton-dependent zinc release from intracellular ligands, J. Neurochem., 130, 87–96.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  12. Frazzini, V., Rapposelli, I. G., Corona, C., Rockabrand, E., Canzoniero, L. M., and Sensi, S. L. (2007) Mild acidosis enhances AMPA receptor-mediated intracellular zinc mobilization in cortical neurons, Mol. Med., 13, 356–361.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  13. Stelmashook, E. V., Isaev, N. K., Plotnikov, E. Y., Uzbekov, R. E., Alieva, I. B., Arbeille, B., and Zorov, D. B. (2009) Effect of transitory glucose deprivation on mitochondrial structure and functions in cultured cerebellar granule neurons, Neurosci. Lett., 461, 140–144.

    Article  CAS  PubMed  Google Scholar 

  14. Medvedeva, Y. V., Lin, B., Shuttleworth, C. W., and Weiss, J. H. (2009) Intracellular Zn2+ accumulation contributes to synaptic failure, mitochondrial depolarization, and cell death in an acute slice oxygen-glucose deprivation model of ischemia, J. Neurosci., 29, 1105–1114.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  15. Zhu, L., Tang, Y., Wang, H. D., Zhang, Z. Y., and Pan, H. (2012) Immersion autometallographic demonstration of pathological zinc accumulation in human acute neural diseases, Neurol. Sci., 33, 855–861.

    Article  PubMed  Google Scholar 

  16. Suh, S. W., Garnier, P., Aoyama, K., Chen, Y., and Swanson, R. A. (2004) Zinc release contributes to hypoglycemiainduced neuronal death, Neurobiol. Dis., 16, 538–545.

    Article  CAS  PubMed  Google Scholar 

  17. Bitanihirwe, B. K., and Cunningham, M. G. (2009) Zinc: the brain’s dark horse, Synapse, 63, 1029–1049.

    Article  CAS  PubMed  Google Scholar 

  18. Sensi, S. L., Paoletti, P., Koh, J. Y., Aizenman, E., Bush, A. I., and Hershfinkel, M. (2011) The neurophysiology and pathology of brain zinc, J. Neurosci., 31, 16076–16085.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  19. Stelmashook, E. V., Isaev, N. K., Genrikhs, E. E., Amelkina, G. A., Khaspekov, L. G., Skrebitsky, V. G., and Illarioshkin, S. N. (2014) Role of Zn2+ and Cu2+ in pathogenic mechanisms of Alzheimer’s and Parkinson’s diseases, Biochemistry (Moscow), 79, 501–508.

    Google Scholar 

  20. Lozier, E. R., Stelmashook, E. V., Uzbekov, R. E., Novikova, S. V., Zorov, S. D., Alieva, I. B., Arbeille, B., Zorov, D. B., and Isaev, N. K. (2012) Stimulation of kainate toxicity by zinc in cultured cerebellar granule neurons and the role of mitochondria in this process, Toxicol. Lett., 208, 36–40.

    Article  CAS  PubMed  Google Scholar 

  21. Andreeva, N., Khodorov, B., Stelmashook, E., Sokolova, S., Gragoe, E., Jr., and Victorov, I. (1992) 5-(N-ethyl-Nisopropyl)amiloride and mild acidosis protect cultured cerebellar granule cells against glutamate-induced delayed neuronal death, Neuroscience, 49, 175–181.

    Article  CAS  PubMed  Google Scholar 

  22. Ou-Yang, Y., Kristian, T., Mellergerd, P., and Siesjo, B. K. (1994) The influence of pH on glutamateand depolarization-induced increases of intracellular calcium concentration in cortical neurons in primary culture, Brain Res., 646, 65–72.

    Article  CAS  PubMed  Google Scholar 

  23. Tombaugh, G. C., and Somjen, G. G. (1996) Effects of extracellular pH on voltage-gated Na+, K+ and Ca2+ currents in isolated rat CA1 neurons, J. Physiol., 493, 719–732.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  24. Schneider, D., Gerhardt, E., Bock, J., Muller, M. M., Wolburg, H., Lang, F., and Schulz, J. B. (2004) Intracellular acidification by inhibition of the Na+/H+- exchanger leads to caspase-independent death of cerebellar granule neurons resembling paraptosis, Cell Death Differ., 11, 760–770.

    Article  CAS  PubMed  Google Scholar 

  25. Luo, J., Chen, H., Kintner, D. B., Shull, G. E., and Sun, D. (2005) Decreased neuronal death in Na+/H+ exchanger isoform 1-null mice after in vitro and in vivo ischemia, J. Neurosci., 25, 11256–11268.

    Article  CAS  PubMed  Google Scholar 

  26. Rogers, T. B., Inesi, G., Wade, R., and Lederer, W. J. (1995) Use of thapsigargin to study Ca2+ homeostasis in cardiac cells, Biosci. Rep., 15, 341–349.

    Article  CAS  PubMed  Google Scholar 

  27. Park, K. S., Poburko, D., Wollheim, C. B., and Demaurex, N. (2009) Amiloride derivatives induce apoptosis by depleting ER Ca2+ stores in vascular endothelial cells, Br. J. Pharmacol., 156, 1296–1304.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  28. Stork, C. J., and Li, Y. V. (2010) Zinc release from thapsigargin/IP3-sensitive stores in cultured cortical neurons, J. Mol. Signal., 5, DOI: 10.1186/1750-2187-5-5.

  29. Toda, T., Kadono, T., Hoshiai, M., Eguchi, Y., Nakazawa, S., Nakazawa, H., Higashijima, N., and Ishida, H. (2007) Na+/H+ exchanger inhibitor cariporide attenuates the mitochondrial Ca2+ overload and PTP opening, Am. J. Physiol. Heart Circ. Physiol., 293, 3517–3523.

    Article  Google Scholar 

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Correspondence to E. V. Stelmashook or N. K. Isaev.

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Published in Russian in Biokhimiya, 2015, Vol. 80, No. 8, pp. 1282–1288.

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Stelmashook, E.V., Novikova, S.V., Amelkina, G.A. et al. Acidosis and 5-(N-ethyl-N-isopropyl)amiloride (EIPA) attenuate zinc/kainate toxicity in cultured cerebellar granule neurons. Biochemistry Moscow 80, 1065–1072 (2015). https://doi.org/10.1134/S000629791508012X

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  • DOI: https://doi.org/10.1134/S000629791508012X

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