大気圧走査電子顕微鏡を用いたシナプス構造と分子局在の解析  [in Japanese] Analysis of Molecular Localization and Synaptic Structure Using ASEM  [in Japanese]

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Abstract

<p>シナプス形成は複雑かつ精巧な脳神経ネットワーク構築の要のステップの一つである.近年,小脳におけるGluRδ2-Cbln1-NRXN複合体などの脳シナプス形成を担うシナプスオーガナイザーの存在が明らかになってきたが,シナプスの形成過程,分子基盤については不明な点が多い.SiN薄膜窓を底面にもつASEMディッシュ上に初代培養神経細胞を培養し,光学顕微鏡と電子顕微鏡による同一視野観察が可能な大気圧走査電子顕微鏡(ASEM)を用いてシナプスに局在する分子の観察を試みた.さらに,シナプス形成を誘導するタンパク質をコートした磁気ビーズを培養神経細胞に添加することで,ASEMディッシュ上の任意の場所にシナプス前部の構造を作り観察できることが示された.初代神経細胞培養とASEMによる解析は,シナプス形成の分子機構を解明する上で有力なツールになるものと期待される.</p>

<p>Synapse formation is one of the key steps in the development of neural networks. Precise synaptic connections between nerve cells are crucial for brain development and functions. Recently, several synapse organizers including cerebellar GluRδ2-Cbln1-neurexin triad have been identified. However, their intracellular downstream signals and process of synapse formation are still largely unknown. Atmospheric scanning electron microscope (ASEM) is an electron microscope which was developed recently to directly observe subcellular structures and organelles through a silicon nitride (SiN) film under atmospheric pressure. ASEM dish with the SiN film can be used for incubation of cultured primary neurons. Cultured primary neurons were analyzed by integrated fluoresence microscope and ASEM. Presynaptic differentiation by GluRδ2-NTD-Fc-coated magnetic beads was analyzed by fluoresence microscope and ASEM. Fluorescence signals for presynaptic proteins were accumulated on the beads particularly concentrated at the site of contact with axon. Subsequent ASEM analysis revealed that presynaptic protein, presumably of an indistinguishable neurite branch, surrounded each beads. Thus, ASEM is a potentially useful tool for analysis of molecular localization and fine structure of synapse.</p>

Journal

  • KENBIKYO

    KENBIKYO 49(1), 7-9, 2014

    The Japanese Society of Microscopy

Codes

  • NII Article ID (NAID)
    130007701701
  • NII NACSIS-CAT ID (NCID)
    AA11917781
  • Text Lang
    JPN
  • ISSN
    1349-0958
  • NDL Article ID
    025543022
  • NDL Call No.
    Z16-896
  • Data Source
    NDL  J-STAGE 
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