Kinetics of 9-aminoacridine block of single Na channels.
Kinetics of 9-aminoacridine block of single Na channels.
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DOI：
10.1085/jgp.84.3.361
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发表时间：
1984-09
影响因子：
3.8
通讯作者：
Yeh, J Z
中科院分区：
文献类型：
--
作者：
Yamamoto, D;Yeh, J Z
在用N-溴乙酰胺（NBA）消除钠通道失活的条件下，用膜片钳技术研究了9-氨基吖啶（9-AA）阻断神经母细胞瘤N1E-115细胞单钠通道的动力学。NBA治疗后，通过单个Na通道流动的电流表现为持续数毫秒至数十毫秒的方波开放事件。当9-AA以30至100 μ M的浓度应用于Na通道的细胞质面时，其在Na通道的单个开口内引起开放和阻断状态之间的重复快速转换（闪烁），而不影响单通道电流的幅度。阻塞状态持续时间的直方图和开放状态持续时间的直方图可以用单指数函数拟合。平均开放时间（τ o）随药物浓度增加而变短，而平均阻断时间（τ B）与药物浓度无关。从平均开放时间倒数与9-AA浓度相关的曲线斜率计算的缔合（阻断）速率常数kappa显示出很小的电压依赖性，速率常数约为1 × 10（7）M-1s-1。解离（解阻断）速率常数，l，从平均阻断时间计算，是强烈的电压依赖性，平均速率常数为214 s-1在0 mV和变得更大的膜被超极化。电压依赖性表明，一阶阻断位点位于至少63%的方式通过膜场从细胞质表面。9-AA阻断Na通道的平衡解离常数（由1/kappa的关系定义）计算为在0 mV时为21 μ M。tau-1o和tau-1b的Q10均为1.3，表明结合反应受扩散控制。在9-AA存在下的破裂时间（其为开放时间和阻断时间的总和）长于在没有药物的情况下开放通道的寿命。9-AA阻断单个钠通道的所有特征与9-AA分子阻断开放钠通道的顺序模型一致，被阻断的通道只有在9-AA分子离开通道中的阻断位点后才能关闭。
The kinetics of 9-aminoacridine (9-AA) block of single Na channels in neuroblastoma N1E-115 cells were studied using the gigohm seal, patch clamp technique, under the condition in which the Na current inactivation had been eliminated by treatment with N-bromoacetamide (NBA). Following NBA treatment, the current flowing through individual Na channels was manifested by square-wave open events lasting from several to tens of milliseconds. When 9-AA was applied to the cytoplasmic face of Na channels at concentrations ranging from 30 to 100 microM, it caused repetitive rapid transitions (flickering) between open and blocked states within single openings of Na channels, without affecting the amplitude of the single channel current. The histograms for the duration of blocked states and the histograms for the duration of open states could be fitted with a single-exponential function. The mean open time (tau o) became shorter as the drug concentration was increased, while the mean blocked time (tau b) was concentration independent. The association (blocking) rate constant, kappa, calculated from the slope of the curve relating the reciprocal mean open time to 9-AA concentration, showed little voltage dependence, the rate constant being on the order of 1 X 10(7) M-1s-1. The dissociation (unblocking) rate constant, l, calculated from the mean blocked time, was strongly voltage dependent, the mean rate constant being 214 s-1 at 0 mV and becoming larger as the membrane being hyperpolarized. The voltage dependence suggests that a first-order blocking site is located at least 63% of the way through the membrane field from the cytoplasmic surface. The equilibrium dissociation constant for 9-AA to block the Na channel, defined by the relation of l/kappa, was calculated to be 21 microM at 0 mV. Both tau -1o and tau -1b had a Q10 of 1.3, which suggests that binding reaction was diffusion controlled. The burst time in the presence of 9-AA, which is the sum of open times and blocked times, was longer than the lifetime of open channels in the absence of drug. All of the features of 9-AA block of single Na channels are compatible with the sequential model in which 9-AA molecules block open Na channels, and the blocked channels could not close until 9-AA molecules had left the blocking site in the channels.