Kehl Steven J
Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver, British Columbia, Canada
J Pharmacol Exp Ther. 2017 Nov;363(2):184-195. doi: 10.1124/jpet.117.243097. Epub 2017 Sep 1.
Voltage clamp recordings of macroscopic currents were made from rat potassium-gated potassium 4.2(Kv4.2) channels expressed in human embryonic kidney (HEK293) cells with the main goals of quantifying the concentration, time, and voltage dependence of the block and to generate a state model that replicates the features of the block. When applied either externally or internally, the block of Kv4.2 currents by 4-aminopyridine (4AP) occurs at the holding potential (-80 mV), is affected by the stimulus frequency, and is relieved by membrane depolarization. The for the tonic block at -80 mV was 0.9 ± 0.07 mM and was consistent with 1:1 binding. Relief of block during a step to 50 mV was well fitted by a single exponential with a time constant of ∼40 milliseconds. At -80 mV, the association rate constant was 0.08 mM s, and the off-rate was 0.08 s The state model replicates the features of the experimental data reasonably well by assuming that 4AP binds only to closed states, that 4AP binding and inactivation are mutually exclusive processes, and that the activation of closed-bound channels is the same as for closed channels. Since the open channel has a very low or no affinity for 4AP, channel opening promotes the unbinding of 4AP, which accounts for the reverse use dependence of the block.
在人胚肾（HEK293）细胞中表达的大鼠钾门控钾通道4.2（Kv4.2）上进行了宏观电流的电压钳记录，主要目的是量化阻断的浓度、时间和电压依赖性，并生成一个复制阻断特征的状态模型。当从外部或内部施加时，4-氨基吡啶（4AP）对Kv4.2电流的阻断发生在保持电位（-80 mV），受刺激频率影响，并通过膜去极化解除。在-80 mV时，强直阻断的Kd为0.9±0.07 mM，与1:1结合一致。在阶跃到50 mV期间阻断的解除由一个时间常数约为40毫秒的单指数很好地拟合。在-80 mV时，结合速率常数为0.08 mM-1 s-1，解离速率为0.08 s-1。该状态模型通过假设4AP仅与关闭状态结合、4AP结合和失活是相互排斥的过程以及关闭结合通道的激活与关闭通道相同，相当好地复制了实验数据的特征。由于开放通道对4AP的亲和力非常低或没有亲和力，通道开放促进了4AP的解离，这解释了阻断的反向使用依赖性。
