The role of depolarization in the survival and differentiation of cerebellar granule cells in culture

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Journal of Neuroscience, Vol 7, 2203-2213, Copyright © 1987 by Society for Neuroscience

ARTICLE

The role of depolarization in the survival and differentiation of cerebellar granule cells in culture

V Gallo, A Kingsbury, R Balazs and OS Jorgensen

Cultures greatly enriched in granule cells from early postnatal cerebellum (P8) were grown in a medium containing fetal calf serum. Under the conditions used, nerve cells died, usually within a week, unless the K+ concentration in the medium was greater than or equal to 20 mM. The requirement for elevated [K+]e was manifested by about 3 d in vitro, and after this time continuous exposure to high [K+]e was essential for the survival of the granule cells. The initial morphological and biochemical maturation of the granule cells was similar in the presence and the absence of elevated [K+]e, suggesting that the dependence on depolarizing conditions develops in parallel with the expression of the differentiated characteristics of the cells. The positive effect of elevated [K+]e on granule cell survival was not influenced by preventing bioelectric activity in the cultures with TTX and xylocaine. On the other hand, depolarization-induced transmembrane Ca2+ flux was essential in securing the maintenance of the granule cells. Depolarized nerve cells were compromised when Ca2+ entry was blocked by elevated Mg2+, EGTA, or organic Ca2+ antagonists, while dihydropyridine Ca2+ agonists [BAY K 8644, (+)-(S)-202 79 1 and CGP 28392] were potent agents preventing nerve cell loss in the presence of 15 mM [K+]e, which was ineffective on its own. Calmodulin inhibitors (1 microM trifluoperazine or calmidazolium) blocked the beneficial effect of K+-induced depolarization on granule cells. The comparison of the timing of the differentiation and innervation of the postmitotic granule cells in vivo with the development of the K+ dependence in vitro would indicate that depolarization of the granule neurons in culture mimics the influence of the physiological stimulation in vivo through excitatory amino acid receptors, including N-methyl-D-aspartate receptors, involving Ca2+ entry and the activation of a Ca2+/calmodulin- dependent protein kinase.

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F. X. Zhang, R. Rubin, and T. A. Rooney
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S. Tanaka, K. Suzuki, M. Watanabe, A. Matsuda, S. Tone, and T. Koike
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J. R. Mellor, D. Merlo, A. Jones, W. Wisden, and A. D. Randall
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R. M. Soler, J. Egea, G. M. Mintenig, C. Sanz-Rodriguez, M. Iglesias, and J. X. Comella
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Y. Du, K. R. Bales, R. C. Dodel, E. Hamilton-Byrd, J. W. Horn, D. L. Czilli, L. K. Simmons, B. Ni, and S. M. Paul
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T. M. Miller, K. L. Moulder, C. M. Knudson, D. J. Creedon, M. Deshmukh, S. J. Korsmeyer, and E. M. Johnson Jr.
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S. P. Yu, C. Yeh, S. L. Sensi, B. J. Gwag, L. M. Canzoniero, Z. S. Farhangrazi, H. S. Ying, M. Tian, L. L. Dugan, and D. W. Choi
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A. M Wood, P. Tiwari, and D. R Bristow
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M. J. Courtney, K. E.�O. Akerman, and E. T. Coffey
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M. Deshmukh and E. M. Johnson Jr.
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T. Ono, K. Inokuchi, A. Ogura, Y. Ikawa, Y. Kudo, and S. Kawashima
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T. M. Miller, M. G. Tansey, E. M. Johnson Jr., and D. J. Creedon
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L. M. Gault and R. E. Siegel
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R. Ishitani, K. Sunaga, M. Tanaka, H. Aishita, and D.-M. Chuang
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J. L. Hegarty, A. R. Kay, and S. H. Green
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S. R. D'Mello, K. Borodezt, and S. P. Soltoff
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T. M. Miller and E. M. Johnson Jr.
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E. H. Holt, A. E. Broadus, and M. L. Brines
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A. de Luca, M. Weller, and A. Fontana
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A Ghosh, J Carnahan, and M. Greenberg
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L. Li, D. Guerini, and E. Carafoli
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M. J. O'Hare, S. T. Hou, E. J. Morris, S. P. Cregan, Q. Xu, R. S. Slack, and D. S. Park
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J. Biol. Chem., August 11, 2000; 275(33): 25358 - 25364.
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