Yamane T, Furukawa T, Hiraoka M
Department of Cardiovascular Diseases and Autonomic Physiology, Tokyo Medical and Dental University, Japan.
Am J Physiol. 1995 Aug;269(2 Pt 2):H556-64. doi: 10.1152/ajpheart.1995.269.2.H556.
The blocking action of 4-aminopyridine (4-AP) on the cloned K+ channel Kv1.5 expressed in Xenopus oocytes was studied using the two-microelectrode voltage-clamp method. Application of 4-AP to the bath solution reversibly suppressed the expressed current in a voltage- and concentration-dependent manner decreasing with membrane depolarization and with a half-maximal inhibitory concentration of 0.14 mM (at +40 mV). Both block and unblock occurred mainly during a depolarization when channels were activated. With successive depolarizations, 4-AP decreased not only the peak amplitudes of the current in successive pulses, but also the current during a depolarization. Upon washout of 4-AP, the current recovered with successive depolarizations, whereas no recovery of the current was noted in the absence of depolarizations. The extent of block markedly increased with alkalization of the external solution and decreased with acidification. External application of 4-amino-pyridine methiodide, a charged form of a quaternary 4-AP derivative, did not affect the current, but internal application markedly suppressed the current, indicating the drug gained access to the channel from the cytoplasmic side. These data suggest that 4-AP crosses the membrane in its uncharged form and acts from inside of the cell in its charged form, resulting in block of the channels with higher affinity to the open state.
采用双微电极电压钳法研究了4-氨基吡啶（4-AP）对非洲爪蟾卵母细胞中表达的克隆钾通道Kv1.5的阻断作用。将4-AP加入浴液中，以电压和浓度依赖性方式可逆地抑制表达的电流，该电流随膜去极化而降低，半最大抑制浓度为0.14 mM（在+40 mV时）。阻断和解除阻断主要发生在通道激活时的去极化过程中。随着连续去极化，4-AP不仅降低了连续脉冲中电流的峰值幅度，还降低了去极化期间的电流。在洗脱4-AP后，电流随着连续去极化而恢复，而在没有去极化的情况下未观察到电流恢复。阻断程度随着外部溶液碱化而显著增加，随着酸化而降低。外部应用4-氨基吡啶甲碘化物（一种季铵化4-AP衍生物的带电形式）不影响电流，但内部应用显著抑制电流，表明药物从细胞质侧进入通道。这些数据表明，4-AP以不带电形式穿过膜，并以带电形式从细胞内部起作用，导致对开放状态具有更高亲和力的通道被阻断。
