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通过噪声分析对牛蛙毛细胞转导通道进行表征。

The transduction channel of hair cells from the bull-frog characterized by noise analysis.

作者信息

Holton T, Hudspeth A J

出版信息

J Physiol. 1986 Jun;375:195-227. doi: 10.1113/jphysiol.1986.sp016113.

Abstract

Receptor currents in response to mechanical stimuli were recorded from hair cells in the excised epithelium of the bull-frog sacculus by the whole-cell, gigohm-seal voltage-clamp technique. The stimulus-dependent transduction current was separated from the cell's stimulus-independent K+ and Ca2+ currents; the K+ currents were blocked with an internal solution containing Cs+ while the Ca2+ current was reduced by holding the membrane potential below -70 mV. The temperature of the preparation was maintained at about 10 degrees C to slow the kinetics of the cells' transduction channels. Calibrated displacements of hair bundles of individual hair cells were made with a probe coupled by suction to the kinociliary bulb and moved with a piezoelectricbimorph stimulator. The root mean square noise of probe motion was less than 2 nm. The mean, I, and the variance, sigma 2, of the receptor current were measured from the response to saturating (+/- 0.5 micron) displacements of the hair bundle. I was corrected for current offsets and sigma 2 for the transduction-independent background variance. The relation between sigma 2 and I is consistent with the predictions of a two-conductance-state model of the transduction channel, a model having only one non-zero conductance state. The relation between sigma 2 and I was fitted by the equation sigma 2 = Ii-I2/N, where N is the number of transduction channels in the cell and i is the current through a single open channel. The conductance of the transduction channel is approximately ohmic with a reversal potential near 0 mV. The estimated conductance of a single transduction channel, gamma, is 12.7 +/- 2.7 pS (mean +/- S.D.; n = 18) at 10 degrees C. gamma is independent of the maximum transduction conductance of the cell, Gmax. The number of transduction channels, N, is proportional to Gmax. N ranges from 7 to 280 in cells with Gmax ranging from 0.08 to 2.48 nS. The largest values of N correspond to a few, perhaps four, active transduction channels per stereocilium. Control experiments show that transduction by the hair cell of two artifactual sources of hair-bundle stimulation, noisy or discontinuous motion of the probe, do not contribute substantially to the measured variance, sigma 2. Displacement-response curves are generally sigmoidal and symmetrical; they reasonably fit the predictions of a two-kinetic-state model, comprising one open state and one closed state. The estimated displacement-sensitive free energy, Z, is 5.7 +/- 1.1 kcal/mol micron (mean +/- S.D., n = 18).(ABSTRACT TRUNCATED AT 400 WORDS)

摘要

采用全细胞千兆欧姆封接电压钳技术,从牛蛙球囊切除上皮中的毛细胞记录对机械刺激的感受器电流。将依赖刺激的转导电流与细胞的非刺激依赖性钾离子和钙离子电流分离;钾离子电流用含铯离子的内部溶液阻断,而钙离子电流通过将膜电位保持在-70mV以下来降低。标本温度维持在约10℃,以减缓细胞转导通道的动力学。用通过吸力与动纤毛球相连并由压电双压电晶片刺激器移动的探针,对单个毛细胞的毛束进行校准位移。探针运动的均方根噪声小于2nm。从对毛束饱和(±0.5微米)位移的反应中测量感受器电流的均值I和方差σ²。I校正了电流偏移,σ²校正了与转导无关的背景方差。σ²与I的关系与转导通道的双电导状态模型的预测一致,该模型只有一个非零电导状态。σ²与I的关系用方程σ² =(Ii - I²)/N拟合,其中N是细胞中转导通道的数量,i是通过单个开放通道的电流。转导通道的电导近似为欧姆性,反转电位接近0mV。在10℃时,单个转导通道的估计电导γ为12.7±2.7pS(均值±标准差;n = 18)。γ与细胞的最大转导电导Gmax无关。转导通道的数量N与Gmax成正比。在Gmax范围为0.08至2.48nS的细胞中,N范围为7至280。N的最大值对应于每个静纤毛有几个,可能是四个,活跃的转导通道。对照实验表明,毛细胞对毛束刺激的两个人为来源(探针的噪声或不连续运动)的转导,对测量的方差σ²贡献不大。位移-反应曲线通常为S形且对称;它们合理地符合由一个开放状态和一个关闭状态组成的双动力学状态模型的预测。估计的位移敏感自由能Z为5.7±1.1kcal/mol微米(均值±标准差,n = 18)。(摘要截断于400字)

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