权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Investigation of signal transduction in cerebellar Purkinje cell by in vivo expression of pertusis toxin.

通过百日咳毒素的体内表达研究小脑浦肯野细胞的信号转导。

基本信息

批准号：
16300099
负责人：
AIBA Atsu
金额：
$ 9.6万
依托单位：
Kobe University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16300099/
关键词：
Purkinje cell Gi o pertusis toxin L7 transgenic mouse insulator RGS

项目摘要

In order to investigate the role of trimetric GTP binding proteins in cerebellar Purkinje cell, we generated two different kind of transgenic mice.First we generated transgenic mice which express pertusis toxin (PTX) in Purkinje cells to inhibit the signal via a Gi/o family. Secondly, we generated mutant mice which lack RGS8 that has a GTPase activating domein for Gi/o family and is expressed prominently in Purkinje cells. We generated L7-PTX transgenic mice by microinjection of PTX gene driven by Purkinje cell specific L7 promoter. One L7-PTX transgenic mouse expressed PTX mRNA in cerebellum and showed reduction in spontaneous movement. The transgenic mouse died at 7 weeks old. Other transgenic mice expressed little PTX mRNA in the cerebellum and transgene were not transmitted to the offspring. These results suggested that expression of PTX in the cerebellum results in reduced viability. To overcome this situation, we generated a transgenic vector in which PTX expression was suppressed by doxycycline. We generated RGS8 knockout mice (KO) by gene targeting. We introduced loxP sites and neomycin resistant gene flanked by two FRT sites. We generated RGS8 mutant carrying loxP sequence with intact exons as well as the RGS8 KO mice in which two exons were deleted. RGS8 KO showed normal motor coordination, spontaneous movement and body weight. However, body temperature of RGS8 KO was lower than that of wild-type littermate. Furthermore, hypothermia induced by GABAB receptor agonist baclofen was mimicked in RGS8 KO mice. These results suggested that RGS8 regulates body temperature by controlling GABAB receptor signaling.

为了研究三价GTP结合蛋白在小脑浦肯野细胞中的作用，我们制备了两种不同类型的转基因小鼠。首先，我们获得了在浦肯野细胞中表达PTX的转基因小鼠，通过Gi/o家族抑制该信号。其次，我们产生了缺乏RGS8的突变小鼠，RGS8具有激活Gi/o家族的Domein的GTP酶，并且在Purkinje细胞中显著表达。我们通过显微注射Purkinje细胞特异性L7启动子驱动的PTX基因获得了L7-PTX转基因小鼠。1只L7-PTX转基因小鼠在小脑表达PTX mRNA，并表现出自发活动减少。这只转基因小鼠在7周大时死亡。其他转基因小鼠在小脑中几乎没有表达PTX基因，转基因小鼠没有遗传给后代。这些结果表明，PTX在小脑中的表达导致存活率降低。为了克服这种情况，我们构建了一个转基因载体，其中PTX的表达被多西环素抑制。我们通过基因打靶获得了RGS8基因敲除小鼠(KO)。我们引入了loxP位点和新霉素抗性基因，并在其两侧引入了两个FRT位点。我们获得了带有完整外显子的携带loxP序列的突变体RGS8，以及缺失了两个外显子的RGS8 KO小鼠。RGS8KO显示运动协调、自发活动和体重正常。但RGS8KO的体温低于野生型。此外，还模拟了GABAB受体激动剂巴氯芬对RGS8KO小鼠的降温作用。这些结果表明，RGS8通过调控GABAB受体信号来调节体温。

项目成果

期刊论文数量（15）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Synaptically driven endocannabinoid release requires Ca2+-assisted metabotropic glutamate receptor subtype 1 to phospholipase Cbeta4 signaling cascade in the cerebellum.

突触驱动的内源性大麻素释放需要 Ca2 辅助的代谢型谷氨酸受体亚型 1 与小脑中磷脂酶 Cbeta4 信号级联反应。

DOI：
发表时间：
2005
期刊：
J. Neurosci 25
影响因子：
0
作者：
Maejima;T.;Oka;S.;Hashimotodani;Y.;Ohno-Shosaku;T.;Aiba;A.;Wu;D.;Waku;K.;Sugiura;T.;Kano;M.
通讯作者：
M.

Signaling complex formation of phospholipase Cbeta4 with metabotropic glutamate receptor type lalplia and 1,4,5-tris-phosphate receptor at the perisynapse and endoplasmic reticulum in the mouse brain.

磷脂酶 Cbeta4 与小鼠大脑突触周围和内质网的代谢型谷氨酸受体类型 lalplia 和 1,4,5-tris-磷酸受体形成信号复合物。

DOI：
发表时间：
2004
期刊：
Eur.J.Neurosci. 20(11)
影响因子：
0
作者：
Nakamura M;Sato K;Fukaya M;Araishi K;Aiba A;Kano M;Watanabe M.
通讯作者：
Watanabe M.

Farnesylation of retinal transducin underlies its translocation during light adaptation

DOI：
10.1016/j.neuron.2005.07.025
发表时间：
2005-08-18
期刊：
NEURON
影响因子：
16.2
作者：
Kassai, H;Aiba, A;Fukada, Y
通讯作者：
Fukada, Y

Signaling complex formation of phospholipase Cbeta4with metabotropic glutamate receptor type 1alpha and 1,4,5-tris-phosphate receptor at the perisynapse and endoplasmic reticulum in the mouse brain.

磷脂酶 Cbeta4 与小鼠大脑突触周围和内质网处的代谢型谷氨酸受体 1α 和 1,4,5-tris-磷酸受体形成信号复合物。

DOI：
发表时间：
2004
期刊：
Eur. J. Neurosci. 20・11
影响因子：
0
作者：
Tomemori Y;Ichiba M;Kusumoto A;Mizuno E;Sato D;Muroya S;Nakamura M;Kawaguchi H;Yoshida H;Ueno S;Nakao K;Nakamura K;Aiba_A;Katsuki M;Sano A.;Atsushi KIDO;M.J.K.Klein;大高洋司;Maejima et al.;Seishi ISAKA;入口敦志;S.Goto;Tomemori et al.;Kouichi MORIMOTO;相田満;J.H.Kupers;Takashi KURIHARA;Nakamura et al.
通讯作者：
Nakamura et al.

Subcellular and subsynaptic localization of group I metabotropic glutamate receptors in the monkey subthalamic nucleus