Semi-loose seal Neurobiotin electroporation for combined structural and functional analysis of neurons
Semi-loose seal Neurobiotin electroporation for combined structural and functional analysis of neurons
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DOI：
10.1007/s00424-008-0539-9
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发表时间：
2008-11-01
影响因子：
4.5
通讯作者：
Vaney, David I.
中科院分区：
文献类型：
--
作者：
Kanjhan, Refik;Vaney, David I.
细胞内锐电极、全细胞膜片钳和细胞旁标记方法已被开发用于神经元结构和功能的综合分析。我们描述了一种新的电穿孔技术，用于用神经生物素标记神经元，使用半松散密封结构（R=100-300 MO）的贴片电极，具有非常小的振幅脉冲（50 mV）。在贴片电极的细胞内溶液中加入2%的神经生物素可使介电膜击穿电压阈值降低约三倍。由此产生的孔形成允许(1)自发和光诱发的突触后电位的稳定记录，而没有明显的细胞质冲洗；(2)染料的通过而没有溢出。该方法的效率和可靠性使其特别适合于小区域组织中多个神经元的连续记录和标记。
Intracellular sharp-electrode, whole-cell patch clamp and juxtacellular labeling methods have previously been developed for combined analysis of neuronal structure and function. We describe a novel electroporation technique for labeling neurons with Neurobiotin, using patch electrodes in a semi-loose seal configuration (R=100-300 MO) with very small amplitude pulses (50 mV). The addition of 2% Neurobiotin to the intracellular solution in the patch electrode reduces the dielectric membrane breakdown voltage threshold by about threefold. The resulting pore formation allows for (1) the stable recording of spontaneous and light-evoked postsynaptic potentials without significant cytoplasmic washout and (2) the passage of dye without spillover. The efficiency and reliability of the method makes it particularly suitable for the serial recording and labeling of multiple neurons in a small area of tissue.