Visualization of the dendritic arbor of neurons in intact 500 μm thick brain slices
Visualization of the dendritic arbor of neurons in intact 500 μm thick brain slices
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DOI：
10.1016/s0165-0270(02)00341-2
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发表时间：
2003-02-15
影响因子：
3
通讯作者：
Kennedy, TE
中科院分区：
文献类型：
--
作者：
Hamam, BN;Kennedy, TE
表征单个神经元的结构和电生理特性对于理解单个细胞如何促进神经元网络的功能至关重要。在对急性脑切片中的神经元进行细胞内记录后，通常通过组织切片的连续切片然后重建感兴趣的神经元来检查记录的细胞的结构；这是一个劳动密集型且耗时的过程。在这里，我们采用了一种整体免疫组织化学技术，并用它来可视化厚达 500 微米的成人中枢神经系统组织切片中单个神经元的树突状乔木。切片的透化和广泛的清洗允许组织化学试剂渗透并从切片上清洗，产生有限的背景染色。使用这种方法，可以对切片内的细胞进行光学切片并使用光学切片进行重建。我们展示了 500 微米厚的成年大鼠内嗅皮层和海马切片中神经元树突树的图像，这些图像通过免疫组织化学标记，或在全细胞膜片钳或尖锐电极记录后注射生物胞素标记。获得的分辨率足以可视化切片深处的树突棘。该方法没有与切割连续切片相关的伪影，并且广泛适用于需要可视化中枢神经系统组织厚切片中单个细胞精细结构的任务。 (C) 2002 Elsevier Science B.V. 保留所有权利。
Characterizing the structure and electrophysiological properties of single neurons is essential for understanding how individual cells contribute to the function of neuronal networks. Following intra-cellular recording from neurons in acute brain slices, the structure of the recorded cell has typically been examined by serial sectioning of the tissue slice and then reconstructing the neuron of interest; a labor-intensive and time-consuming process. Here, we have adapted a whole-mount inummohistochemical technique and used it to visualize the dendritic arbor of individual neurons in sections of adult CNS tissue up to 500 mum thick. Permeabilization of the slice and extensive washing allow histochemical reagents to penetrate and be washed from the section, producing limited background staining. Using this method, the cell within the slice can be sectioned optically and reconstructed using the optical sections. We present images of the dendritic trees of neurons in 500 mum thick slices of adult rat entorhinal cortex and hippocampus, labeled either immunohistochemically, or by biocytin injection following whole-cell patch clamp or sharp electrode recordings. The resolution obtained is sufficient to visualize dendritic spines deep within the section. The method is free from artifacts associated with cutting serial sections and is broadly applicable to tasks that require visualization of the fine structure of individual cells in thick slices of CNS tissue. (C) 2002 Elsevier Science B.V. All rights reserved.