| Project/Area Number |
12680750
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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 |
Neurochemistry/Neuropharmacology
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| Research Institution | Kanazawa University |
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Principal Investigator |
KATO Satoru Kanazawa University Grad. Sch. Of Med. Professor, 大学院・医学系研究科, 教授 (10019614)
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| Co-Investigator(Kenkyū-buntansha) |
MATSUKAWA Toru Kanazawa University Grad. Sch. Of Med. Instructor, 大学院・医学系研究科, 助手 (30219414)
TANII Hideji Kanazawa University Grad. Sch. Of Med. Asociate Professor, 大学院・医学系研究科, 助教授 (90110618)
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| Project Period (FY) |
2000 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2001: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 2000: ¥1,900,000 (Direct Cost: ¥1,900,000)
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| Keywords | goldfish retina / optic nerve regeneration / gene expression / transglutaminase / ganglion sells / neurite outgrowth / transglutaminase / goldfish / nerve section / regeneration / visual system / リコンビナント蛋白 |
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| Research Abstract |
The goldfish optic nerve can regenerate after injury. Regrowth of optic nerve fibers starts to extend 7-10 days after optic nerve transection. They arrive at the optic tectum 3-5 weeks after axtfomy. Finally, visual function is restored 4-6 months after optic nerve transection. To understand the molecular mechanism of optic nerve regeneration, we identified genes whose expression is specifically upregulated during the early stage of optic nerve regeneration. A cDNA library constructed from goldfish retina 5 days after transection was screened Two clones were identified as the Na, K-ATPase catalytic subunitalpha 3 isoform and the neural type of transglutaminase (TG) by sequence analysis. In northern hybridization, the expression level of the both mRNAs was significantly increased at 2 days and peaked at 5-10 days, and then gradually decreased and returned to control level by 45-60 days after optic nerve transection. In situ hybridization have revealed the location of this transient retinal change after axotomy. The increased expression was observed only in the ganglion cell layer at 10-20 days after optic nerve transection, The cDNA of TG was inserted into the plasmid DNA and thus a fused protein of TG (ca. 70kDa) was obtained. Antiserum against the recombinant TG was made by immunization. Immunohistochemical staining further confirmed the data of in situ hybridization. In an explant culture system, neurite outgrowth fkm the retina 7 days after optic nerve transection was spontaneously promoted. A low concentration of ouabain (an inhibitor of Na,K ATPase alpha 3 subunit) or neutral antibody against TG, completely blocked the spontaneous neurite outgrowth from the lesioned retina. Together, these data indicate that upregulation of the Na, K-ATPase alpha 3 subunit or TG is involved in the regrowth of ganglion cell axons after axotomy. Such molecular techniques will become useful tool for future genetic studies of nerve regeneration.
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