Functions of neuronal membrane-associated proteins for neuronal connection and synapase formation.

神经元膜相关蛋白对神经元连接和突触形成的功能。

• 批准号: 12680728
• 负责人: INOUE Akihiro
• 金额: $ 2.24万
• 依托单位: Tokyo Medical & Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12680728/
• 关键词: Retino-tectal projection; Organotypic culture; Cell adhesion molecule; Synapse formation; GFP; Cadherin; Adenovirus vector; Laminar specificity; 網膜-視蓋投射; アデノウィルスベクター

1. Many parts of brain have laminated structures, and each lamina receives afferents from specific sets of neuron groups, then make physiologically relevant synapses. In the chick tectum, sixteen layers can be distinguished, and retinal inputs make arborizations and synapses only in the three of those lamina (retinorecipient laminae; RRL) in its superficial region. N-cadherin is concentrated specifically in RRL, and remained in adult, while SC1/JC7/DM-GRASP/BEN, an immunoglobulin superfamily cell adhesion molecule appeared to be expressed in deeper two RRL with its peak at embryonic day 14-16, then gradually decreased. A coculture system was developed to analyze the lamina-specificity and retinal neuron terminal behavior. When antibodies against N-cadherin were applied, more overshooting of neurites beyond RRL border, and reduced terminal arborization were observed. SC1 antibodies had similar but less effects on the overshooting to N-cadherin, however arborization was not affected. Tho … More se results suggest that some of cell-surface molecules with spatially-restricted expression patterns may regulate the development of lamina-specific retinotectal neuronal connectivity in the chick, and may also be applicable to other laminated brain portions and/or other species.2. We developed an hippocampal slice culture system from neonatal mice of which organotypic structures are well maintained for at least three weeks. In order to obtain neuron-specific expression of exogenous genes, we prepared the slice cultures from a transgenic line with Cre recombinase under control of CaMKII gene promoter, the the cultures are infected with adenoviruses bearing a promoter and EGFP with loxP-STOP-loxP cassette in between. Adenoviruses are chosen, because of their low cytotoxicity to mammalian cultures. High resolutional observation of synaptic structures in living neurons can be achieved using a two-photon laser microscope. We also analyzed dynamics of PSD-95-GFP and GFP-Zip45, showing that they behave in similar way to that of dissociation cultures which we previously reported. To study the function of N-cadherin in synapse formation and plastic change, we are now preparing several systems for suppressing N-cadherin gene expression with anti-sense oligonucleotides as well as direct inhibition of its adhesion activity with inhibitory peptides or proteins. Less

1。大脑的许多部分具有层压结构，每个层层都会从特定的神经元组中接收传入，然后制作出与物理相关的突触。在雏鸡底部，可以区分16层，视网膜输入仅在其表面区域中的三个薄片（视网膜层层层）中才能使一克和突触。 N-钙粘着蛋白特别浓缩在RRL中，并保留在成年中，而SC1/JC7/DM-GRASP/BEN是一种免疫球蛋白超家族细胞粘附分子，似乎在更深的两个RRL中表达，其峰值在14-16的峰值峰值，然后逐渐改善。开发了一种共培养系统来分析椎板特异性和残留神经元终末行为。当应用针对N-钙黏着蛋白的抗体时，RRL边界以外的神经运动的过度旋转，并观察到减少的末端树皮化。 SC1抗体对与N-钙黏着蛋白的过度施加的影响相似但较少，但是一克化不受影响。 ……更多的SE结果表明，一些具有空间限制性表达模式的细胞表面分子可能调节雏鸡中层状特异性视网膜直直直透明神经元连接的发展，并且也可能适用于其他层压的脑部部分和/或其他物种。2。我们从新生小鼠开发了海马切片培养系统，其中有机结构至少维持了至少三周。为了获得外源基因的神经特异性表达，我们从CAMKII基因启动子控制的转基因系中从转基因系中制备了切片培养物，这些培养物被腺病毒感染带有启动子和EGFP的腺病毒，并带有LOXP-Stop-Stop-Stop-Stop-loxp-loxp-loxp cassette。选择腺病毒，因为它们对哺乳动物培养物的细胞毒性低。使用两光子激光显微镜可以实现生物神经元突触结构的高分辨率观察。我们还分析了PSD-95-GFP和GFP-ZIP45的动力学，表明它们的行为与我们先前报道的分离培养物相似。为了研究N-钙粘着蛋白在突触形成和塑性变化中的功能，我们现在正在准备几种用于抑制N-钙粘蛋白基因表达的系统，并用抗义寡核苷酸以及用抑制性辣椒或蛋白质直接抑制其粘合剂活性。较少的

项目成果

Okabe, S., Miwa, A., H.Okado: "Spine formation and correlated assembly of presynaptic and postsynaptic moelcules"Journal of Neuroscience. 21,1. 6105-6114 (2001)

Okabe, S.、Miwa, A.、H.Okado：“脊柱形成和突触前和突触后分子的相关组装”神经科学杂志。

Umeda, T., S.Okabe: "Visualizing synapse formation and remodeling : recent advanced in real-time imageing of CNS synapses"Neuroscience Research. 40. 291-300 (2001)

Umeda, T., S.Okabe：“可视化突触形成和重塑：中枢神经系统突触实时成像的最新进展”神经科学研究。

Okabe, S., Miwa, A., and H. Okado: "Spine formation and correlated assembly of presynaptic and postsynaptic moelcules."Journal of Neuroscience. Vol.21. 6105-6114 (2001)

Okabe, S.、Miwa, A. 和 H. Okado：“脊柱形成以及突触前和突触后分子的相关组装。”神经科学杂志。

Okabe, S., Umshido, T., Konno, D., Okado, H., K.Sobue: "Rapid redistribution of the postsynaptic density protein PSD-Zip45(Homer 1c)and its differential regulation by NMDA receptors and calcium channels"Journal of Neuroscience. 21,2. 9561-9571 (2001)

Okabe, S.、Umshido, T.、Konno, D.、Okado, H.、K.Sobue：“突触后密度蛋白 PSD-Zip45 (Homer 1c) 的快速重新分布及其 NMDA 受体和钙通道的差异调节”

Okabe, S., Urushido, T., Konno, D., Okado, H., and K. Sobue: "Rapid redistribution of the postsynaptic density protein PSD-Zip45 (Homer 1c) and its differential regulation by NMDA receptors and calcium channels."Journal of Neuroscience. Vol.21. 9561-9571

Okabe, S.、Urushido, T.、Konno, D.、Okado, H. 和 K. Sobue：“突触后密度蛋白 PSD-Zip45 (Homer 1c) 的快速重新分布及其 NMDA 受体和钙通道的差异调节