Neuronal cell cultures from aplysia for high-resolution imaging of growth cones.
Neuronal cell cultures from aplysia for high-resolution imaging of growth cones.
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DOI：
10.3791/662
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发表时间：
2008-02-20
期刊：
影响因子：
--
通讯作者：
Suter, Daniel
中科院分区：
文献类型：
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作者：
Lee, Aih Cheun;Decourt, Boris;Suter, Daniel
神经元生长锥是位于轴突顶端的高度运动的结构，其可以检测环境中的引导线索并将该信息转化为朝向适当的靶细胞的定向运动。为了充分了解指导信息是如何从细胞表面传输到底层的动态细胞骨架网络，需要一个模型系统，适用于活细胞成像的蛋白质动力学在高的时间和空间分辨率。典型的脊椎动物生长锥太小，无法定量分析F-肌动蛋白和微管动力学。来自加利福尼亚海兔的神经元比脊椎动物神经元大5-10倍，可以容易地保存在室温下，并且是用于显微操作和生物物理测量的非常稳健的细胞。它们的生长锥有非常明确的细胞质区域和一个描述良好的细胞骨架系统。神经元细胞体可以显微注射各种探针用于研究生长锥运动和指导。在本协议中，我们演示了一个程序，用于解剖的腹神经节，培养袋细胞神经元和建立一个成像室的生长锥活细胞成像。
Neuronal growth cones are the highly motile structures at the tip of axons that can detect guidance cues in the environment and transduce this information into directional movement towards the appropriate target cell. To fully understand how guidance information is transmitted from the cell surface to the underlying dynamic cytoskeletal networks, one needs a model system suitable for live cell imaging of protein dynamics at high temporal and spatial resolution. Typical vertebrate growth cones are too small to quantitatively analyze F-actin and microtubule dynamics. Neurons from the sea hare Aplysia californica are 5-10 times larger than vertebrate neurons, can easily be kept at room temperature and are very robust cells for micromanipulation and biophysical measurements. Their growth cones have very defined cytoplasmic regions and a well-described cytoskeletal system. The neuronal cell bodies can be microinjected with a variety of probes for studying growth cone motility and guidance. In the present protocol we demonstrate a procedure for dissection of the abdominal ganglion, culture of bag cell neurons and setting up an imaging chamber for live cell imaging of growth cones.