| Project/Area Number |
01570982
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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 |
Ophthalmology
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| Research Institution | Kitasato University |
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Principal Investigator |
WAKAKURA Masato Kitasato Univ. Ophthalmology, Associate Professor, 医学部, 助教授 (50137931)
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| Co-Investigator(Kenkyū-buntansha) |
FUKUSHIMA Kazuya Kitasato Univ. Ophthalmology Assistant, 医学部, 助手 (60199205)
YAMAMOTO Noboru Kitasato Univ. Functional Morph. Professor, 看護学部, 教授 (10050543)
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| Project Period (FY) |
1989 – 1991
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| Project Status |
Completed (Fiscal Year 1991)
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| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1991: ¥400,000 (Direct Cost: ¥400,000)
Fiscal Year 1990: ¥400,000 (Direct Cost: ¥400,000)
Fiscal Year 1989: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Keywords | Muller cells / retinal neurons / axonal transport / neurotransmitters / aminoadipic acid / 4-aminopyridine / culture / glial fibrillary acidic protein / aminoadipic acid / intracellular calcium ion / stereospecificity / neurotoxicity / gliotoxcity / 網膜ニュ-ロン / ミュ-ラ-細胞 / 培養 / 細胞内カルシウム濃度 / 細胞内カルシウム動員 / 4アミノピリジン / カルシウム拮抗剤 / アミノアジピン酸 / 神経伝達物質 / 細胞内カルシウムイオン濃度 / カルシウム動員 / NCAM / 軸索輸送 |
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| Research Abstract |
A number of neurotransmitters elicit postsynaptic release through influx of extracellular calcium ion. Retinal neurons cultured from neonatal rat eyes, in which pure Muller cell culture was used as a feeder layer, expressed not only neurofilament peptide also various neurotransmitters. Using the cultured retinal neurons, cellular response to 4-aminopyridine(4-AP)was investigated. Free calcium in the neurons was labelled with Fura-2 AM and the rapid change in intracellular calcium ion[Ca^<2+>]_i was analyzed by a computerassisted fluorospectrometry. Rapid increase of [Ca^<2+>]_i was seen significantly following treatment beyond 4mM of 4-AP in all neurons. Since the response was not completely blocked during incubation with calcium-free EGTA containing medium, intracellular calcium mobilization system appeared also to be exerted.
Administration of excitatory amino acids may be toxic for retinal cells and block axonal transport in retinal neurons. Effects of aminoadipic acid(AAA)on culture
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d retinal cells were investigated by the above method of [Ca^<2+>]_i measurement. The D, L- and D-forms of AAA activated neurons at low concentrations and activated the Muller cells at higher concentrations. The D-form may act selectively on retinal neurons, suggesting that it may be an agonist of an excitatory amino acid receptor. The results indicate that the ability of AAA to elevate cytosolic [Ca^<2+>]_i depends upon the spectrospecificity of the AAA and on cell type. The prolonged effect of D, L-AAA on cultured retinal cells was also studied using morphological observation and immunofluorescence of glial fibrillary acidic, protein(GFAP). Vacuolations in Muller cells were found after 8.0 mM treatment. Reduction of neurons was evident after treatment beyond 1.6 mM. Administration of AAA was followed by accumulation of GFAP in Muller cells but never resulted in reduction of the cells or reduction of DNA content in the culture, suggesting strong. cellualr resistance. These results indicate AAA not to be highly toxic for Muller cells and to be rather toxic for retinal neurons. The results support that the cellular response to AAA depends upon Cellular constituent. Less
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