STUDY OF THE PHENOTYPES AND FUNCTIONS OF TRP FAMILY MEMBERS EXPRESSED IN NEURONS

TRP家族成员神经元表达的表型和功能研究

基本信息

批准号：
15590059
负责人：
KANEKO Shuji
金额：
$ 2.3万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

项目摘要

(1) A brief exposure to hydrogen peroxide (H_2O_2) induces severe deterioration of primary cultured neurons in vitro. We have investigated a possible link between the H_2O_2-induced neuronal death and Ca^<2+>-permeable TRPM2 channels that are supposed to be regulated by ADP-ribose (ADPR) and/or H_2O_2. In most of cultured cerebral cortical neurons from fetal rat, TRPM2 proteins were detected at cell bodies immunocytochemically. Application of H_2O_2 to cultured neurons elicited an increase in intracellular Ca^<2+> concentration ([Ca^<2+>]_i) caused by Ca^<2+> influx and subsequent neuronal death in a similar concentration range. Molecular cloning of rat TRPM2 cDNA revealed several differences in amino acid sequences within Nudix box region as compared with those of human and mouse TRPM2. ADPR-induced current responses, H_2O_2-induced Ca^<2+> influx and H_2O_2-induced cell death were elicited in human embryonic kidney cells heterogeneously expressing rat TRPM2. Treatment of cultured neurons with small interfering RNA (siRNA) against rat TRPM2 caused decrease in immunoreactive TRPM2 content and the H_2O_2-induced Ca^<2+> influx. Moreover, H_2O_2-induced neuronal death was significantly inhibited by the siRNA treatment. These results suggest a critical role of TRPM2 in regulation of neuronal survival as well as the intracellular Ca^<2+> homeostasis.(2)In cultured cerebral cortical neurons from feral rat, TRPC1,3,4,5 and 6 proteins are detected by immunocytochemistry. In Fura-2 loaded neurons, Ca^<2+> influx was observed after P2Y receptor stimulation with ATP, which was inhibited by La^<3+>, Zn^<2+> and SKF96365. Both TRPC3 immunoreactivity and the receptor-activated Ca^<2+> influx were decreased in the neurons treated with anti-TRPC3 antisense oligodeoxynucleotide. There results suggest that TRPC3 channels are involved in the receptor-operated Ca^<2+> influx in immature cerebral cortical neurons.

（1）短暂暴露于过氧化氢（H_2O_2）会在体外诱导原发性培养的神经元严重恶化。我们已经研究了H_2O_2诱导的神经元死亡与Ca^<2+> - 可渗透的TRPM2通道之间的可能联系，该通道应由ADP-ribose（ADPR）和/或H_2O_2调节。在胎儿大鼠的大多数培养的脑皮质神经元中，通过免疫细胞化学的细胞体检测到TRPM2蛋白。 H_2O_2在培养的神经元中的应用引起了细胞内Ca^<2+>浓度的增加（[Ca^<2+>] _ I），由Ca^<2+>涌入和随后在类似浓度范围内的神经元死亡引起。与人和小鼠TRPM2相比，大鼠TRPM2 cDNA的分子克隆揭示了Nudix框区域内氨基酸序列的几个差异。 ADPR诱导的电流反应，H_2O_2诱导的Ca^<2+>涌入和H_2O_2诱导的细胞死亡在人类胚胎肾细胞中引起了异质表达大鼠TRPM2的人类胚胎肾细胞。用小干扰RNA（siRNA）对大鼠TRPM2的培养神经元的处理导致免疫反应性TRPM2含量降低，而H_2O_2诱导的Ca^<2+>涌入。此外，siRNA处理可显着抑制H_2O_2诱导的神经元死亡。这些结果表明，TRPM2在调节神经元存活以及细胞内Ca^<2+>稳态中的关键作用。（2）在野性大鼠的培养的脑皮质神经元中，TRPC1,3,4,5和6个蛋白质通过免疫细胞学检测。在Fura-2负载的神经元中，在用ATP刺激P2Y受体刺激后观察到Ca^<2+>流动，这被La^<3+>，Zn^<2+>和SKF96365抑制。在用抗TRPC3抗义寡脱氧核苷酸处理的神经元中，TRPC3免疫反应性和受体激活的CA^<2+>流入均降低。结果表明，TRPC3通道参与了未成熟脑皮质神经元中受体操的Ca^<2+>流入。

项目成果

期刊论文数量（41）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Serofendic acid prevents acute glutamate neurotoxicity in cultured cortical neurons.

Serofendic Acid 可防止培养的皮质神经元中的急性谷氨酸神经毒性。

DOI：
发表时间：
2003
期刊：
Eur.J.Pharmacol. 477
影响因子：
0
作者：
H.Toyohara et al.;M.Kinoshita-Kawada et al.;Haruhiko Toyohara et al.;Mariko Kinoshita-Kawada et al.;M.Kinoshita-Kawada et al.;H.Toyohara et al.;F.Osakada et al.;S.Fujimoto et al.;Shinji Fujimoto et al.;Feng Wang et al.;Fujimoto et al.;F.Osakada et al.;N.Sakka et al.;R.Taguchi et al.;Y.Takada et al.;Noriko Sakka et al.;Yuki Takada et al.;Haruki Shibata et al.;Ryota Taguchi et al.
通讯作者：
Ryota Taguchi et al.

Identification of scallop DMT as a metal transporter responsible for cadmium accumulation

鉴定扇贝 DMT 作为负责镉积累的金属转运蛋白

DOI：
发表时间：
2005
期刊：
FEBS Letters (In press)
影响因子：
0
作者：
H.Toyohara et al.;M.Kinoshita-Kawada et al.;Haruhiko Toyohara et al.;Mariko Kinoshita-Kawada et al.;M.Kinoshita-Kawada et al.;H.Toyohara et al.
通讯作者：
H.Toyohara et al.

α-Tocotrienol provides the most potent neuroprotection among vitamin E analogs on cultured striatal neurons

DOI：
10.1016/j.neuropharm.2004.06.029
发表时间：
2004-11-01
期刊：
NEUROPHARMACOLOGY
影响因子：
4.7
作者：
Osakada, F;Hashino, A;Akaike, A
通讯作者：
Akaike, A

「研究成果報告書概要(欧文)」より

摘自《研究结果报告摘要（欧洲）》

DOI：
发表时间：
2006
期刊：
Seibutsu Butsuri 46(1)
影响因子：
0
作者：
Yasushi Shigeri;Keiko Shimamoto
通讯作者：
Keiko Shimamoto

Mechanisms of oxygen glucose deprivation-induced goutamate release from cerebrocortical slice cultures

氧葡萄糖剥夺诱导脑皮质切片培养物释放古氨酸的机制

DOI：
发表时间：
2004
期刊：
Neuroscience Research 50
影响因子：
0
作者：
H.Toyohara et al.;M.Kinoshita-Kawada et al.;Haruhiko Toyohara et al.;Mariko Kinoshita-Kawada et al.;M.Kinoshita-Kawada et al.;H.Toyohara et al.;F.Osakada et al.;S.Fujimoto et al.
通讯作者：
S.Fujimoto et al.