Ex vivo imaging of motor axon dynamics in murine triangularis sterni explants
Ex vivo imaging of motor axon dynamics in murine triangularis sterni explants
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DOI：
10.1038/nprot.2008.160
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发表时间：
2008-01-01
期刊：
影响因子：
14.8
通讯作者：
Misgeld, Thomas
中科院分区：
文献类型：
--
作者：
Kerschensteiner, Martin;Reuter, Miriam S.;Misgeld, Thomas
我们提供了一个协议，描述了一个外植体系统，允许运动轴突的动态成像。这种方法是基于从小鼠的胸骨三角肌（一种覆盖胸腔内部的薄肌肉）制备的神经肌肉外植体。这些外植体可以存活数小时，含有长延伸的外周运动轴突，包括其末端分支和神经肌肉接头。可以从在神经元或神经胶质细胞中表达荧光蛋白的转基因小鼠系制备外植体，这使得能够通过荧光显微镜直接观察它们的细胞和亚细胞形态。延时成像提供了一种方便和可靠的方法来跟踪运动轴突的动态行为，其周围的神经胶质细胞和细胞内细胞器具有高的时间和空间分辨率。胸骨三角肌外植体可在15分钟内制备，离体成像数小时，并在约2小时内进行免疫组织化学处理。
We provide a protocol that describes an explant system that allows the dynamics of motor axons to be imaged. This method is based on nerve-muscle explants prepared from the triangularis sterni muscle of mice, a thin muscle that covers the inside of the thorax. These explants, which can be maintained alive for several hours, contain long stretches of peripheral motor axons including their terminal arborizations and neuromuscular junctions. Explants can be prepared from transgenic mouse lines that express fluorescent proteins in neurons or glial cells, which enables direct visualization of their cellular and subcellular morphology by fluorescence microscopy. Time-lapse imaging then provides a convenient and reliable approach to follow the dynamic behavior of motor axons, their surrounding glial cells and their intracellular organelles with high temporal and spatial resolution. Triangularis sterni explants can be prepared in 15 min, imaged ex vivo for several hours and processed for immunohistochemistry in about 2 h.