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Screening of drugs for neurodegenerative disorders using neurotransmitter release machinery expressed in Xenopus oocytes

利用非洲爪蟾卵母细胞表达的神经递质释放机制筛选治疗神经退行性疾病的药物

基本信息

批准号：
05557010
负责人：
SHUNTOH Hisato
金额：
$ 3.58万
依托单位：
Kobe University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Developmental Scientific Research (B)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1995
项目状态：
已结题

项目摘要

1. Complementary DNA clones encoding neural visinin-like calcium-binding proteins (NVP-1, NVP-2 and NVP-3) were isolated from rat brain cDNA library. NVP-1, NVP-2 and NVP-3 mRNAs are most highly expressed in brain and are differentially expressed in various regions of rat brain. Furthermore, alterations in NVP-1 may be involvedin the pathology of Phecyclidine-induced psychosis. 2. The effect of phorbol ester on the Na+-dependent uptake of GABA by rat cortical synaptosomes and Xenopus oocytes expressing GAT1 (neuronal GABA transporter) was examined. Protein kinase C may modulate the GABA uptake into presynaptic terminals through the inhibition of GAT1 activity. 3. The serotonin release from RBL-2H3 cells is subject to tonic, Ca2+-independent, inhibition by protein phosphatase 1.4. In the Schaffer collateral-CA1 pathway in rat hippocampal slices, calyculin A induced a long-lasting potentiation of the extracellularfield potentials with a transient increase in glutamate release. 5. Heat shock transiently suppresses the inositol trisphosphate-mediated intracellular calcium signal transduction system and that HSP72 is involved in the recovery of receptor-stimulated calcium rise in the neuronal cells. 6. By screening the mutagen-treated fission yeast cells, immunosuppressant- and temperature- sensitive (its1-6) mutant strains were isolated. Gene products of its1+ and its3+ were beta-adaptin homolog, which is a subunit of adapter protein AP-2, and Pl (4) P5-kinase homolog, which is required for exocytosis, respectively. These results suggest that calcineurin plays an important role in vesicular recycling. In conclusion, our several results provide important clues about the molecular basis of neurotransmitter release and secretory vesicle recycling or endocytosis.

1.从大鼠脑cDNA文库中克隆了编码神经视素样钙结合蛋白（NVP-1、NVP-2和NVP-3）的互补DNA克隆。NVP-1、NVP-2和NVP-3 mRNA在大脑中表达最高，并且在大鼠大脑的各个区域中表达差异。此外，NVP-1的改变可能参与Phosphoclidine诱导的精神病的病理过程。2.研究了佛波酯对大鼠皮层突触体和表达GAT 1（神经元GABA转运蛋白）的爪蟾卵母细胞Na+依赖性摄取GABA的影响。蛋白激酶C可能通过抑制GAT 1的活性来调节GABA进入突触前末梢。3.从RBL-2 H3细胞释放的5-羟色胺受到紧张性的、不依赖于Ca 2+的蛋白磷酸酶1.4的抑制。在大鼠海马脑片的Schaffer侧支-CA 1通路中，calyculin A诱导了细胞外场电位的持久增强，并短暂增加了谷氨酸的释放。5.热休克可短暂抑制三磷酸肌醇介导的细胞内钙信号转导系统，热休克蛋白72参与了受体刺激的神经细胞内钙升高的恢复。6.通过对诱变处理的裂殖酵母细胞进行筛选，分离到了免疫抑制剂和温度敏感（its 1 -6）突变株。its 1+和its 3+的基因产物分别是衔接蛋白AP-2的一个亚基β-衔接素同源物和胞吐作用所需的P1（4）P5-激酶同源物。这些结果表明，钙调神经磷酸酶在囊泡循环中起着重要作用。总之，我们的几个结果提供了重要的线索，神经递质释放和分泌囊泡回收或内吞的分子基础。