| Project/Area Number |
04670035
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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 |
神経解剖学
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| Research Institution | Chiba University |
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Principal Investigator |
KADOTA Tomoko Chiba University, Medical School Associate Professor, 医学部, 助教授 (00089864)
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| Co-Investigator(Kenkyū-buntansha) |
TOYAMA Yoshiro Chiba University, Medical School Assistant Professor, 医学部, 講師 (70009637)
MIZOTE Muneaki Teikyo University of Technology, Information Engineering Professor, 情報工学, 教授 (70009645)
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| Project Period (FY) |
1992 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1994: ¥400,000 (Direct Cost: ¥400,000)
Fiscal Year 1993: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1992: ¥600,000 (Direct Cost: ¥600,000)
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| Keywords | neurotransmitter transporter / oligopeptide / dopamine transporter / PC12 cells / monoclonal antibody / immunohistochemistry / シナプス / シナプス小胞 / シナプトファイシン / 細胞骨格 / 免疫細胞化学 / シナプス小胞膜蛋白 / シナプス後部濃物質 / 免疫電顕 |
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| Research Abstract |
1.Synthesis of oligopeptides of the neurotransmitter transporter and production of the anti-peptide antibodies
The neurotransmitter transporters presenting in the presynaptic membrane work to reuptake the neurotransmitter released into the synaptic cleft and thereby to terminate the postsynaptic response. cDNA of many neurotransmitter transporters have been cloned. In the present study dopamine transporter, serotonin transporter and glutamate transporter were examined. The specific parts of their amino acid sequences were designed and oligopeptides were chemically synthesized. Then, the antipeptide antibodies were produced with in vitro immunization method using these oligopeptides as antigens. These antibodies were characterized the specificity to the original transporter proteins by Western blotting.
2.Expression of dopamine transporter in the PC12 cells during differentiation.
The expression and subcellular localization of dopamine transporter(DAT)was examined in the PC12 cells (h-clone) treated with NGF.DAT was immunohistochemically detected on the surface of the PC12 cells. When the cells were treated with NGF,the expression of DAT was significantly emphasized first in the Golgi area and then on the surface of the cell. The surface area, then, extended outwards forming neurites. The tips of the neurites, growth cones, were strongly labeled with the antibody. Many small spots positively responsing to the antibody were detected in the cytoplasm of the cell body and in the neurites.
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