Millonas M M, Hanck D A
Department of Pharmacological and Physiological Sciences, The University of Chicago, Illinois 60637, USA.
Biophys J. 1998 Jan;74(1):210-29. doi: 10.1016/S0006-3495(98)77781-1.
We describe a new electrophysiological technique called nonequilibrium response spectroscopy, which involves application of rapidly fluctuating (as high as 14 kHz) large-amplitude voltage clamp waveforms to ion channels. As a consequence of the irreversible (in the sense of Carnot) exchange of energy between the fluctuating field and the channel protein, the gating response is exquisitely sensitive to features of the kinetics that are difficult or impossible to adequately resolve by means of traditional stepped potential protocols. Here we focus on the application of dichotomous (telegraph) noise voltage fluctuations, a broadband Markovian colored noise that fluctuates between two values. Because Markov kinetic models of channel gating can be embedded within higher-dimensional Markov models that take into account the effects of the voltage fluctuations, many features of the response of the channels can be calculated algebraically. This makes dichotomous noise and its generalizations uniquely suitable for model selection and kinetic analysis. Although we describe its application to macroscopic ionic current measurements, the nonequilibrium response method can also be applied to gating and single channel current recording techniques. We show how data from the human cardiac isoform (hH1a) of the Na+ channel expressed in mammalian cells can be acquired and analyzed, and how these data reveal hidden aspects of the molecular kinetics that are not revealed by conventional methods.
我们描述了一种名为非平衡响应光谱学的新电生理技术，该技术涉及向离子通道施加快速波动（高达14kHz）的大幅度电压钳波形。由于波动场与通道蛋白之间存在不可逆（从卡诺意义上讲）的能量交换，门控响应对动力学特征极为敏感，而这些特征通过传统的阶梯电位协议很难或无法充分解析。在这里，我们重点关注二分（电报）噪声电压波动的应用，这是一种在两个值之间波动的宽带马尔可夫有色噪声。由于通道门控的马尔可夫动力学模型可以嵌入考虑电压波动影响的高维马尔可夫模型中，通道响应的许多特征可以通过代数方法计算。这使得二分噪声及其推广形式特别适用于模型选择和动力学分析。尽管我们描述了其在宏观离子电流测量中的应用，但非平衡响应方法也可应用于门控和单通道电流记录技术。我们展示了如何获取和分析在哺乳动物细胞中表达的钠通道人类心脏亚型（hH1a）的数据，以及这些数据如何揭示传统方法未揭示的分子动力学的隐藏方面。
