The connectome is the comprehensive map of the brain represented by wiring diagram of the full set of neuro-glia and synapses within entire brain of an organism. Some recent scientific efforts have successfully been made to visualize such map at neuro-glial networking level, however, capturing it as one unit of the entire brain have never been elucidated. Moreover, in order to derive structure-function relationship of different brain regions in response to a defined stimulus, there is a need to elucidate the connectome at single neuro-glial ensemble level after brain is challenged with the known memory function. This needs developing molecular approaches to tag neuro-glial activities in response to a conditioned brain function. Such approaches of using specific molecular tags have been tried to visualize independently neuron and glial specific events in response to a memory function, however, they could not tag the connectome together at single neuro-glia ensemble level. Therefore, there is a need to develop new methods for mapping entire connectome up to a single neuro-glial precision and resolution, with a purpose of tagging specific brain region accountable to execute a special memory formation process. The present hypothetical paper aims to propose a novel molecular method to generate the structural connectome at neuro-glial level in mice brain. Herein, we propose to tag the entire connectome at neuro-glia precision by generating a transgenic mice via transposing and recombining engineered novel "Neuro-Glia specific Vectors" (NGVs: specific to excitatory neurons, inhibitory neurons and glial cells) vis a vis "Transcriptional/ Translational Messenger (TMs: specific to metalloproteinases, MMP-9) coupled with different color protein tags, followed by the Clarity. Herein, the NGVs will be translated via Neuro-glia specific promoters, while TMs will be translated via endogenous MMP-9 promoter in all neuro-glial cells. The viability of all constructs will be verified in cortical/ hippocampal culture by inducing them to undergo chemically induced long term potentionation (cLTP) following visualization of different colored pattern. This will be further confirmed by Immunostaning, Western Blot and RT-PCR analysis. Additionally, in this approach, one can decipher the dynamics of molecular and cellular events associated with MMP-9 seretome by monitoring the trafficking of tagged endogenous MMP-9 protein after neuronal stimulation by cLTP in vitro. However, for visualizing complete connectome, the adult transgenic mice will be challenged with fear consolidation (Fear context and contextual cue) tests followed by Clarity coupled Light Sheet Microscopy to analyze neuro-glia ensemble following whole brain imaging.

中文翻译：

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映射哺乳动物大脑中的连接组：生物工程神经胶质细胞特异性载体的一种新方法。

连接组是大脑的综合图，由生物体整个大脑内全套神经胶质和突触的接线图表示。最近的一些科学努力已经成功地在神经胶质网络级别可视化这种地图，但是，从未阐明将其作为整个大脑的一个单位来捕获。此外，为了获得不同大脑区域响应特定刺激的结构-功能关系，需要在大脑受到已知记忆功能挑战后在单个神经胶质集合水平上阐明连接组。这需要开发分子方法来标记神经胶质活动以响应条件性大脑功能。这种使用特定分子标签的方法已尝试独立可视化神经元和神经胶质特定事件以响应记忆功能，但是，它们无法在单个神经胶质集合水平上将连接组标记在一起。因此，需要开发将整个连接组映射到单个神经胶质精度和分辨率的新方法，目的是标记负责执行特殊记忆形成过程的特定大脑区域。本假设论文旨在提出一种新的分子方法，以在小鼠大脑的神经胶质水平上生成结构连接组。在此，我们建议通过转座和重组工程化的新型“神经胶质细胞特异性载体”（NGV：人们可以通过在体外 cLTP 神经元刺激后监测标记的内源性 MMP-9 蛋白的运输来破译与 MMP-9 血清组相关的分子和细胞事件的动态。然而，为了可视化完整的连接组，成年转基因小鼠将接受恐惧巩固（恐惧上下文和上下文提示）测试，然后是 Clarity 耦合光片显微镜，以在全脑成像后分析神经胶质集合。