Analysis of local neural circuit in the central nervous system

中枢神经系统局部神经回路分析

• 批准号: 16200025
• 负责人: KANEKO Takeshi
• 金额: $ 32.12万
• 依托单位: Kyoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16200025/
• 关键词: Neuroscience; Anatomy; Signal Transduction; Cell and Tissue; Physiology; 中枢神経系; 局所神経回路; 投射ニューロン; インターニューロン; トランスジェニックマウス; ウィルスベクター; ゴルジ染色様標識; 細胞内染色

To reveal the local circuit in the central nervous system, we examine the circuit under the strategy of 'From one to group'.1) Single GABAergic neurons were labeled by intracellular staining after whole cell clamp in the cortical slices of adult VGAT/GFP transgenic rats. We analyzed the contacts of these GABAergic axons onto the dendrites of corticospinal neurons, which were retrogradely labeled before the slice preparation.2) To visualize the whole dendrites of a specific group of neurons, we developed a dendritic membrane-targeted reporter protein (myrGFP-LDLRCT) using a test system of lentiviral vectors. We further produced three kinds of transgenic mice with Thy1 promoter, GAD67 short promoter, and parvalbumin BAC promoter. In all the mice, we found that the dendrites and cell bodies of a specific neuron group were completely labeled with GFP fluorescence or immunofluorescence. Thus, we can now use these mice in the 'From one to group' study combining with an intracellular staining technique.3) To visualize the whole dendrites of a specific group of neurons, we also developed a viral vectors for retrograde labeling. We have tried pseudorabies virus, rabies virus glycoprotein (RVG)-pseudotyped Sindbis virus, and RVG-pseudotyped lentivirus, and got a partial success in Sindbis virus and lentivirus. In addition, when myrGFP-LDLRCT-expressing adenovirus was injected into the thalamus with 0.6 M NaCl, the dendrites of corticothalamic neurons were efficiently labeled. Thus, we have started the analysis on the local connection of a pyramidal cell to corticothalamic neurons.4) In the striatum, single medium-sized spiny neurons were labeled with Sindbis virus exressing membrane-targeted GFP. Cortical and thalamic inputs to the single neurons were examined with VGluT1 and VGluT2 immunoreactivities. This may be a 'From group to one' analysis of neural circuit.

为了揭示中枢神经系统的局部回路，我们按照“从一个到一组”的策略来检查回路。1）成年VGAT/GFP转基因大鼠皮层切片中全细胞钳夹后，通过细胞内染色标记单个GABA能神经元。我们分析了这些 GABA 能轴突与皮质脊髓神经元树突的接触，这些轴突在切片制备之前被逆行标记。2）为了可视化特定神经元组的整个树突，我们使用慢病毒载体测试系统开发了树突膜靶向报告蛋白（myrGFP-LDLRCT）。我们进一步制备了三种带有Thy1启动子、GAD67短启动子和小清蛋白BAC启动子的转基因小鼠。在所有小鼠中，我们发现特定神经元组的树突和细胞体完全被 GFP 荧光或免疫荧光标记。因此，我们现在可以在“从一个到一组”研究中结合细胞内染色技术使用这些小鼠。3）为了可视化特定神经元组的整个树突，我们还开发了用于逆行标记的病毒载体。我们尝试过伪狂犬病病毒、狂犬病病毒糖蛋白（RVG）-伪型辛德比斯病毒、RVG-伪型慢病毒，在辛德比斯病毒和慢病毒方面均取得了部分成功。此外，当将表达myrGFP-LDLRCT的腺病毒注射到含有0.6 M NaCl的丘脑时，皮质丘脑神经元的树突被有效标记。因此，我们开始分析锥体细胞与皮质丘脑神经元的局部连接。4) 在纹状体中，单个中等大小的多刺神经元被表达膜靶向 GFP 的辛德比斯病毒标记。使用 VGluT1 和 VGluT2 免疫反应性检查单个神经元的皮质和丘脑输入。这可能是对神经回路的“从一组到一个”分析。

项目成果

Efferent and afferent connections of GABAergic neurons in the supratrigeminal and the intertrigeminal regions An immunohistochemical tract-tracing study in the GAD67-GFP knock-in mouse

• DOI: 10.1016/j.neures.2004.10.005
• 发表时间: 2005-01
• 期刊: Neuroscience Research
• 影响因子: 2.9
• 作者: Jinlian Li;Sheng-Xi Wu;Ryohei Tomioka;K. Okamoto;KouichiC . Nakamura;T. Kaneko;N. Mizuno

Two medullospinal sympathetic premotor pathways mediated by different vesicular glutamate transporters.

由不同囊泡谷氨酸转运蛋白介导的两种髓脊髓交感前运动通路。

• 发表时间: 2004
• 作者: Nakamura;S-X Wu;K Matsumura;T Huebschle;Y Nakamura;F Fujiyama;H Hioki;R Gerstberger;S Kobayashi;T Kaneko
• 通讯作者: T Kaneko

Presynaptic localization of AMPA-type glutamate receptor in corticostriatal and thalamostriatal axon terminals.

AMPA 型谷氨酸受体在皮质纹状体和丘脑纹状体轴突末端的突触前定位。

• 发表时间: 2004
• 作者: F Fujiyama;E Kuramoto;K Okamoto;H Hioki;T Furuta;L Zhou;S Nomura;T Kaneko
• 通讯作者: T Kaneko

Association of group 3 metabotropic glutamate receptor-equipped axon terminals with the dendrites of cholinergic neurons in ratneostriatum.

大鼠新纹状体中第 3 组代谢型谷氨酸受体装备的轴突末端与胆碱能神经元树突的关联。

• 发表时间: 2004
• 作者: E Kuramoto;F Fujiyama;R Sigemoto;T Kaneko
• 通讯作者: T Kaneko

Shemically Specific Circuit Composed of VGLUT3-and PPTB-Producing Interneurons in Rat Neocortex

大鼠新皮质中由产生 VGLUT3 和 PPTB 的中间神经元组成的化学特异性电路

• 发表时间: 2004
• 作者: Hiroyki;Hioki;Fumino;Fujiyama;Kazuhiro;Nakamura;Wakoto;Matsuda;Takeshi;Kaneko
• 通讯作者: Kaneko