Abstract

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.

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