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A systematic method of noise analysis for channels with single-channel conductance of fA level

单通道电导fA级通道噪声分析的系统方法

基本信息

批准号：
17590184
负责人：
OIKI Shigetoshi
金额：
$ 2.3万
依托单位：
University of Fukui
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

项目摘要

Single channel conductance of the voltage-gated proton channel was expected to be below pS. The noise analysis method for evaluating such small conductance channels was examined in this study. Whole-cell recordings of the voltage-gated proton channel from microglial cells were performed. The time course of activation at depolarization potential was very slow for the proton channel and long recording time was necessary to collect enough numbers of current traces for depolarizing pulses. For the analysis we found that the most critical step was detrend upon the selection of current traces. In the long time of recordings current traces change their shape. Current traces showing these systematic changes, rather than stochastic changes, must be eliminated from the analysis. Several criteria for selecting current traces were imposed. Ensemble average was performed from those selected traces. Fluctuating components were extracted and power-spectrum and non-stationary noise analysis were performed at various experimental conditions. Double-Lorentzian fit was done for the power spectral density. The cut-off frequency of 5 kHz was set for the non-stationary noise analysis. The variance-mean plot was drawn and we found that this plot gave valuable criteria for the selection of traces. The process from selecting traces with different criteria to the variance-mean plot was repeated until the fitted parameters were converged. From variance-mean plots amplitudes of single channel current, numbers of active channels and maximal open probability were estimated. The estimated single channel current for the voltage-gated proton channel was 12 fA. Underlying mechanisms for improvements of the data analysis using selected traces were discussed.

预期电压门控质子通道的单通道电导低于pS。在这项研究中，评估这种小电导通道的噪声分析方法进行了检查。进行小胶质细胞的电压门控质子通道的全细胞记录。质子通道在去极化电位激活的时间过程非常缓慢，需要长的记录时间来收集足够数量的去极化脉冲的电流轨迹。对于分析，我们发现最关键的步骤是在选择电流迹线时的去趋势。在长时间的记录中，电流迹线的形状发生了变化。显示这些系统性变化而不是随机变化的电流迹线必须从分析中消除。选择电流轨迹的几个标准被强加。从这些选择的迹线进行包络平均。在不同的实验条件下，波动成分被提取和功率谱和非平稳噪声分析进行。对功率谱密度进行双洛伦兹拟合。对于非平稳噪声分析，将截止频率设定为5 kHz。绘制了方差-均值图，我们发现该图为迹线的选择提供了有价值的标准。重复从选择具有不同标准的迹线到方差-平均值图的过程，直到拟合参数收敛。从方差-平均值图的单通道电流的幅度，活跃通道的数量和最大开放概率估计。电压门控质子通道的估计单通道电流为12 fA。讨论了使用选定轨迹改进数据分析的基本机制。