Genetic analysis for learning and instinctive behaviors by gene targeting mice.

通过基因靶向小鼠对学习和本能行为进行遗传分析。

• 批准号: 06454674
• 负责人: YAGI Takeshi
• 金额: $ 4.48万
• 依托单位: (National Institute of Physiological Sciences) Okazaki National Research Institutes
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06454674/
• 关键词: Gene targeting; Fyn; NMDA; Emotion; RNP; GABA; Seizure; behavior; ピューロマイシン; 行動; ターゲティング; チロシンキナーゼ

The mammalian nervous system constitutes an enormous neural network and regulates various behaviors such as locomotion, sense, recognition, emotion, memory and learning. The question is how these neural networks are systematically formed during development and how they regulate behaviors. Fyn is a member of the Src family kinases, a non-receptor-type tyrosine kinase and its deficiencies causes impairments of spatial learning in mice. In this reserch project, firstly we have confirmed that gene disruption of Fyn causes several other behavioral abnormalities. They have suckling problem and higher fear-response scores in the novelty preferance and passive avoidance tests, exhibit stronger light aversion in the light-dark choice test, and are hyperresponsive to fear-inducing environments and to acoustically primed audiogenic seizures. Anatomical defects in the neural cell layr of the hippocampal formation and in the glomeruli of the olfactory bulb were also observed in the mutants. Thus, t … More he Fyn signaling pathway should offer insights into the morphological, electrophysiological, and behavioral events in the mammalian nervous system. Second we focused to molecular function of Fyn during brain formation and on several behavior formation. We examined the susceptibility to seizures induced by various convulsive drugs, including, pentylenetetrazol, picrotoxin, bicuculline, kainic acid, N-methyl-D-aspartate (NMDA) and strychine. Fyn-deficient mice were significantly more likely to show myoclonic convulsions, when pentylenetetrazol, picrotoxin, bicuculline, kainic acid, or NMDA were administered. On the other hand, no difference in seizure susceptibility was found in Fyn-deficient mice, when strychinine was administered. These results provide evidence of abnormal susceptibility to seizures induced by anti-GABAergic agents and agonists for central excitatory amino acids in Fyn-deficient mice. And we isolated five cDNA clones that directly associate with Fyn in neonatal mouse brain by using a two hybrid yeast system. Sequence analyzes revealed that three of them are previously reported molecules, SON,tctex-1 and hnRNP K,and two clones encode novel sequences. The hnRNP K has been reported to asssociate with Fyn. A full-length cDNA of novel clone 82 was obtained and its deduced amino acid sequence was homologous to the RNA-binding proteins. Isolation of many Fyn-binding molecules suggest that, in the mouse brain, Fyn mediates multiple signaling patheways by binding to multiple molecules and that some of these pathways play critical roles in determing a certain type of behavior. Third we newly producted gene-knocked out mice for NMDA receptor subtypes ; epsilon1, epsilon2, epsilon4, glutamate receptor ; delta2, and GAD65. These gene-dificient mice have impaired of spatial learning, defects of suckling behavior, reduced spontaneous activity, malformation of synapse and different susceptibility to seizure-induced drug, respectivelity. We think to be successful in Less

哺乳动物的神经系统构成了一个庞大的神经网络，调节着运动、感觉、识别、情感、记忆和学习等多种行为。问题是这些神经网络在发育过程中是如何系统形成的，以及它们是如何调节行为的。Fyn是Src家族激酶的一员，是一种非受体型酪氨酸激酶，它的缺乏会导致小鼠空间学习障碍。在这个研究项目中，我们首先确认了Fyn基因的破坏会导致其他几种行为异常。他们在新奇偏好和被动回避测试中有哺乳问题和更高的恐惧反应得分，在光暗选择测试中表现出更强的光厌恶，并且对引起恐惧的环境和声启动的听源性癫痫反应过度。在突变体中还观察到海马神经细胞层和嗅球肾小球的解剖缺陷。因此，Fyn信号通路应该为哺乳动物神经系统的形态学、电生理和行为事件提供更多的见解。其次，我们重点研究了Fyn在脑形成过程中的分子功能和几种行为形成。我们检测了各种惊厥药物对癫痫发作的敏感性，包括戊四唑、微旋毒素、双青碱、kainic酸、n -甲基- d -天冬氨酸（NMDA）和士的宁。当给药戊四氮、微毒素、双曲碱、甜菜酸或NMDA时，fny缺陷小鼠更容易出现肌阵挛性惊厥。另一方面，当给药士的宁时，fyn缺陷小鼠的癫痫易感性没有差异。这些结果为fyn缺陷小鼠抗gaba能药物和中枢兴奋性氨基酸激动剂诱导癫痫的异常易感性提供了证据。利用双杂交酵母系统分离出5个与新生小鼠大脑Fyn直接相关的cDNA克隆。序列分析显示，其中3个是先前报道的分子，SON， text1和hnRNP K，两个克隆编码新序列。据报道，hnRNP - K与Fyn有关联。获得了新克隆82的全长cDNA，其氨基酸序列与rna结合蛋白同源。许多Fyn结合分子的分离表明，在小鼠大脑中，Fyn通过结合多个分子介导多种信号通路，其中一些通路在决定某种类型的行为中起着关键作用。第三，我们新建立了NMDA受体亚型基因敲除小鼠；Epsilon1, epsilon2, epsilon4，谷氨酸受体；δ 2和GAD65。这些基因缺陷小鼠分别表现为空间学习障碍、哺乳行为缺陷、自发性活动减少、突触畸形和对癫痫药物的不同易感性。我们认为在Less是成功的

项目成果

Yagi,T.: "Gene targeting of fyn tyrosine kinase in murine nervous system.Genes for development,Cell growh and infectious diseases" Genes for development,Cell growth and infectious diseases,John Libbey Eurotext in Paris. 20. 17-27 (1995)

Yagi,T.：“小鼠神经系统中 fyn 酪氨酸激酶的基因靶向。发育、细胞生长和传染病的基因”发育、细胞生长和传染病的基因，John Libbey Eurotext，巴黎。

Miyakawa,T.,Yagi,T.,Kagiyama,A.and Niki,H.: "Radial maze performance,open-field and elevated plus-maze behaviors in Fyn-kinase deficient mice ; Further evidence for increased fearfulness" Molecular Brain Research. 37. 145-150 (1996)

Miyakawa,T.、Yagi,T.、Kagiyama,A. 和 Niki,H.：“Fyn 激酶缺陷小鼠的径向迷宫表现、开放场和升高的十字迷宫行为；恐惧增加的进一步证据”分子脑研究

Asada,H.,et al: "Mice lacking the 65 kDa isoform of glutamic acid decarboxylase (GAD65) maintain normal levels of GAD67 and GABA in their brains but are susceptible to seizures" Biochemical and Biophysical Research Communications. 229. 891-895 (1996)

Asada, H. 等人：“缺乏 65 kDa 谷氨酸脱羧酶 (GAD65) 亚型的小鼠大脑中 GAD67 和 GABA 保持正常水平，但容易癫痫发作”《生物化学和生物物理研究通讯》。

Umemori,H.,et al: "Initial events of myelination involve Fyn tyrosine kinase signalling" Nature. 367. 572-576 (1994)

Umemori, H., et al：“髓鞘形成的初始事件涉及 Fyn 酪氨酸激酶信号传导”Nature。

八木健 等: "遺伝子欠損マウスにおける行動異常解析" 実験医学. 12. 2456-2460 (1994)

Ken Yagi 等人：“基因缺陷小鼠的行为异常分析”《实验医学》12. 2456-2460 (1994)。