1990 Fiscal Year Final Research Report Summary
Intracellular Mechanisms Controlling Neuritic Processes of Mouse Neuroblastoma Cells
| Project/Area Number |
01571207
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Biological pharmacy
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| Research Institution | Okayama University, faculty of Pharmaceutical Sciences |
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Principal Investigator |
TSUDA Masaaki Okayama Univ., Fac. of Pharmaceutical Sci., Associate Professor, 薬学部, 助教授 (80132736)
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| Co-Investigator(Kenkyū-buntansha) |
KANAZAWA Hiroshi Okayama Univ., Fac. of Technology, Professor, 工学部, 教授 (50116448)
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| Project Period (FY) |
1989 – 1990
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| Keywords | synaptic plasticity / neuroblastoma / neurite outgrowth / neurotransmitter / transcription / DNA-binding / gene-transfer / lipofectin |
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| Research Abstract |
To elucidate the mechanisms of the synaptic plasticity, we investigated the genetical responses of neurons to several stimuli including neurotransmitters.
(1) Protein kinase inhibitor H-7 [1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride] can stimulate the neurite outgrowth of mouse neuroblastoma N18TG2 cell and NG108-15 hybrid cell. Morphology of the neurites induced by H-7 was different from that by dibutyryl (dB)-cAMP. Stimulation of the neurite outgrowth by H-7 was also observed with cerebellar granule cells prepared from 1-week-old mice, the morphology of which was different from that by dB-cAMP.
(2) We found that the administration of carbachol, noradrenaline or brdykinin can induce the expression of egr mRNA in NG108-15 cells, which was maximally observed 1 hr after administration. The induction of egr-1 mRNA expression was abolished by the presence of specific antagonists, indicating that the induction is evoked through specific receptors to stimuli. As another appro
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ach, we investigated the genetical responses of cultured cerebellar granule cells to neurotransmitters. As the result, N-Methyl-D-Aspartate (NMDA) induced the TRE (TPA responsive element) binding activities, which were detected by a convenient gel-shift assay. This induction included the c-fos mRNA induction and abolished by the presence of specific antagonists. We have found that the induction of TRE binding activities depends upon the presence of extracellular Ca^<++>.
(3) We are now trying to clone the cDNA for the antigen which is detected mainly with the primary dendrite of the Purkinje cell of mouse cerebellum. Purification of the proteins corresponding to the antigen was accomplished, and the partial amino acid sequences were already determined.
(4) We tried to introduce foreign genes by direct injection of DNA with lipofectin (BRL) into several tissues of living animals. As the results, we have succeeded in introducing plasmid DNAs into mouse brain cells and muscle cells, although the efficiency of the introduction is still very low. Less
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[Publications] Tsuda, M., Ono, K., Katayama, N., Yamagata, Y., Kikuchi, K. and Tsuchiya, T.: "Neurite outgrowth from mouse neuroblastoma and cerebellar cells induced by the protein kinase inhibitor H-7" Neurosci. Lett.105. 241-245 (1989)
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[Publications] Katayama, N., Yamagata, Y., Hashimoto, H., Kanazawa, H., Tsuchiya, T. and Tsuda, M.: "Concanavalin A affects beta-tubulin mRNA expression during neuritic processes of mouse neuroblastoma N18TG2 cells in a different manner from colchicine" Biochem. Biophys. Res. Commun.167. 1348-1354 (1990)
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[Publications] Tsuda, M., Yuasa, S., Fujino, Y., Sekikawa, M., Ono, K., Tsuchiya, T. and Kawamura, K.: "Retrovirus-mediated gene transfer into mouse cerebellar primary culrure and its application to the neural transplantation" Brain Res. Brll.24. 787-792 (1990)
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