| Project/Area Number |
14572164
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
応用薬理学・医療系薬学
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| Research Institution | Showa Pharmaceutical University |
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Principal Investigator |
UTSUNOMIYA Iku Showa Pharmaceutical University, Department of Neuroscience, Assistant Professor, 講師 (70168722)
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| Co-Investigator(Kenkyū-buntansha) |
MIYATAKE Tadashi Showa Pharmaceutical University, Department of Neuroscience, Professor, 教授 (50048998)
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| Project Period (FY) |
2002 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2004: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2003: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2002: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Keywords | motor neuron / potassium channel / western blot / RT-PCR / patch clamp / mRNA / neurotrophic factor / ALS / CNTF / ツボクサ / イチョウ / ハイブリドーマ / アセチルコリンレセプター |
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| Research Abstract |
To develop pharmacotherapy for motor neuron disease, we examined whether ciliary neurotrophic factor (CNTF) can alter serum-free cell survival of immortalized motor neuron-like cells, which were established by fusing mouse neuroblastoma N18TG2 with mouse motor neurons. One of the cell lines, NSC-34 exhibited cell survival in the presence of CNTF. NSC-34 preserves the most characteristics of motor neurons, such as the formation of neuromuscular junctions on co-cultured myotube. We looked for CNTF-like activity in some natural low molecular compounds and found Madecassoside in Cenfella asiatica L. and Ginkgol acid in Ginkgo biloba L. to be candidates. These extracts promote NSC-34 cell growth in the presence of serum. Autoimmunity as well as cell death could underlie the pathogenesis of motor neuron disease. Autoantibody may attack some ion channels which mediate neuronal transmission. Therefore the expression of potassium ion channel on NSC-34 cells was examined. Western blot analysis using polyclonal antibodies to shaker-related potassium channel (Kv 1.1-Kv1.6) revealed all Kv1 channel proteins tested were contained in NSC-34 cell lysate. In immunocytochemistry these Kv1 channels were observed on the cell surface. All Kv 1 mRNAs were also detected in NSC-34 cells by RT PCR using specific primers. Potassium ion current was detected on NSC-34 cells by patch-clamp method. As all Kv 1 channels were detected biochemically, histochemically and electrophysiologically in NSC-34 cells we measured the content of each channel to determine what type of Kv1 channel is abundant. Quantitative western blot analysis revealed that Kv 1.4 is most abundant in NSC-34 cells.
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