Properties of Cl- channels from surface membranes of acutely dissociated rat cerebral cortical neurons were studied with the patch clamp technique. These channels were present in the majority of excised inside-out membrane patches. 2. Cl- channels were rarely observed in cell-attached membrane patches, and usually several minutes elapsed following excision of the patch before Cl- channels became active. 3. Under asymmetric ionic conditions (1000 mM-KCli, 140 mM-KClo), neuronal Cl- channels are fairly selective for Cl- over K+ and Na+, with permeability ratios, determined by reversal potential shifts of 4.8 for both PCl/PK and PCl/PNa. 4. Neuronal Cl- channel kinetic activity remained stable over periods of time long enough to collect up to 500,000 open and closed intervals. Occasionally, the channels entered altered modes of activity. In the 'buzz mode' the open and closed interval durations became much shorter than normal for several hundreds of intervals. In the 'subconductance mode' the channel opened to a current level about two-thirds of the normal level. 5. Using the method of maximum likelihood, sums of exponentials were fitted to the distributions of open and closed interval durations. Open interval distributions required at least two exponential components with time constants of less than 1 ms. At least six or seven exponential components were required to fit the closed interval distributions with time constants ranging from 30 microseconds to several hundreds of milliseconds. This suggests that neuronal Cl- channels enter at least two open and six or seven closed kinetic states during normal activity. 6. Cl- channels often entered long-duration closed states of several minutes which could not be accounted for by the sums of exponentials fitted to the distribution of closed interval durations. 7. Neuronal Cl- channels exhibit a marked voltage dependence with the percentage of time the channels are open increasing with depolarization. Most of the observed voltage dependence can be accounted for by a decrease in the mean closed interval duration with depolarization. The mean open interval was relatively independent of voltage. 8. These results suggest a high degree of similarity in kinetic behaviour and conductance properties between the fast Cl- channels of tissue-cultured rat skeletal muscle and fast Cl- channels in acutely dissociated rat cerebral cortical neurons.
摘要
采用膜片钳技术研究了急性分离的大鼠大脑皮层神经元表面膜氯离子通道的特性。这些通道存在于大多数切除的内面向外膜片中。2. 在细胞贴附膜片中很少观察到氯离子通道，通常在膜片切除后几分钟，氯离子通道才开始活跃。3. 在不对称离子条件下（1000 mM - KCli，140 mM - KClo），神经元氯离子通道对氯离子的选择性远高于钾离子和钠离子，通过反转电位偏移确定的通透率比值，PCl/PK和PCl/PNa均为4.8。4. 神经元氯离子通道的动力学活性在足够长的时间内保持稳定，足以收集多达500,000个开放和关闭间隔。偶尔，通道会进入改变的活动模式。在“嗡嗡模式”下，开放和关闭间隔持续时间会在数百个间隔内变得比正常情况短得多。在“亚电导模式”下，通道开放到约为正常水平三分之二的电流水平。5. 使用最大似然法，将指数和拟合到开放和关闭间隔持续时间的分布上。开放间隔分布至少需要两个时间常数小于1毫秒的指数成分。至少需要六到七个指数成分来拟合关闭间隔分布，其时间常数范围从30微秒到数百毫秒。这表明在正常活动期间，神经元氯离子通道至少进入两种开放和六到七种关闭动力学状态。6. 氯离子通道经常进入长达几分钟的长时间关闭状态，这无法用拟合到关闭间隔持续时间分布的指数和来解释。7. 神经元氯离子通道表现出明显的电压依赖性，通道开放时间的百分比随去极化增加。观察到的大部分电压依赖性可以通过去极化时平均关闭间隔持续时间的减少来解释。平均开放间隔相对与电压无关。8. 这些结果表明，组织培养的大鼠骨骼肌快速氯离子通道与急性分离的大鼠大脑皮层神经元快速氯离子通道在动力学行为和电导特性上具有高度相似性。
