Winegar B D, Haws C M, Lansman J B
Department of Cellular and Molecular Pharmacology, University of California, San Francisco 94143-0450, USA.
J Gen Physiol. 1996 Mar;107(3):433-43. doi: 10.1085/jgp.107.3.433.
The activity of single mechanosensitive channels was recorded from cell-attached patches on acutely isolated skeletal muscle fibers from the mouse. The experiments were designed to investigate the mechanism of channel block produced by externally applied aminoglycoside antibiotics. Neomycin and other aminoglycosides reduced the amplitude of the single-channel current at negative membrane potentials. The block was concentration-dependent, with a half-maximal concentration of approximately 200 microM. At high drug concentrations, however, block was incomplete with roughly one third of the current remaining unblocked. Neomycin also caused the channel to fluctuate between the open state and a subconductance level that was also roughly one third the amplitude of the fully open level. An analysis of the kinetics of the subconductance fluctuations was consistent with a bimolecular reaction between an aminoglycoside molecule and the open channel (kon = approximately 1 x 10(6) M-1s-1 and koff = approximately 400 s-1 at -60 mV). Increasing the external pH reduced both the rapid block of the open channel and the frequency of the subconductance fluctuations, as if both blocking actions were produced by a single active drug species with a pKa = approximately 7.5. The results are interpreted in terms of a mechanism in which an aminoglycoside molecule partially occludes ion flow through the channel pore.
从小鼠急性分离的骨骼肌纤维的细胞贴附膜片上记录单个机械敏感通道的活性。这些实验旨在研究外部应用氨基糖苷类抗生素产生通道阻断的机制。新霉素和其他氨基糖苷类药物在负膜电位下降低了单通道电流的幅度。这种阻断呈浓度依赖性，半数最大浓度约为200微摩尔。然而，在高药物浓度下，阻断并不完全，约有三分之一的电流仍未被阻断。新霉素还导致通道在开放状态和一个亚电导水平之间波动，该亚电导水平的幅度也大致是完全开放水平的三分之一。对亚电导波动动力学的分析与氨基糖苷类分子与开放通道之间的双分子反应一致（在-60 mV时，结合速率常数约为1×10⁶ M⁻¹s⁻¹，解离速率常数约为400 s⁻¹）。提高外部pH值可降低开放通道的快速阻断以及亚电导波动的频率，似乎这两种阻断作用都是由一种pKa约为7.5的单一活性药物分子产生的。这些结果是根据一种机制来解释的，即氨基糖苷类分子部分阻塞了通过通道孔的离子流。
