The effect of the spider venom argiopine on L-glutamate-activated membrane channels of crayfish muscle was investigated using the patch-clamp technique. 2. When 10(-2) M-glutamate and 10(-9) M-argiopine were contained in the pipette solution of a cell-attached patch, bursts of openings of excitatory channels appeared after formation of the patch. These bursts ceased abruptly after variable periods of time in the range of 5 min. Higher concentrations of argiopine (up to 10(-6) M) blocked more rapidly, approximately in proportion to concentration. 3. The block of excitatory channels could be partially or completely reversed by hyperpolarizing the membrane by up to -190 mV from the resting potential. The time constant of the recovery of channel opening decreased with increasing hyperpolarization and was 2 ms with -160 mV hyperpolarization. Switching back from the hyperpolarized level to the resting potential, the time constant for the resulting block was about 3 s (10(-7) M-argiopine). Potential-dependent block by argiopine with similar characteristics was also observed in outside-out patches. 4. Up to argiopine concentrations of 10(-7) M the kinetics of channel openings and of bursts measured in pre-block periods or during reversal of the block by hyperpolarization were indistinguishable from controls. 5. When the potential-dependent block observed in the presence of 10(-6) M-argiopine and 10(-2) M-glutamate was reversed by hyperpolarization, additional short closings occurred during bursts. This 'flickering block' did not change burst length appreciably, but an additional open time component (tau = 0.1 ms) appeared and the average open time per burst was reduced. 6. At least two reaction steps seem necessary to model the behaviour of the potential-dependent block. The flickering block may be described as intermittent blocking of the channel which does not interfere with the reactions between glutamate and the channel.
摘要
采用膜片钳技术研究了蜘蛛毒液精氨酸对小龙虾肌肉中L-谷氨酸激活的膜通道的影响。2. 当在细胞贴附式膜片的微管溶液中含有10⁻² M -谷氨酸和10⁻⁹ M -精氨酸时，在膜片形成后出现兴奋性通道的开放爆发。这些爆发在5分钟范围内的不同时间段后突然停止。更高浓度的精氨酸（高达10⁻⁶ M）阻断得更快，大致与浓度成正比。3. 通过将膜电位从静息电位超极化至-190 mV，兴奋性通道的阻断可部分或完全逆转。通道开放恢复的时间常数随超极化程度增加而减小，在超极化-160 mV时为2毫秒。从超极化水平切换回静息电位时，产生阻断的时间常数约为3秒（10⁻⁷ M -精氨酸）。在外侧向外式膜片中也观察到了具有相似特征的精氨酸电位依赖性阻断。4. 直到精氨酸浓度达到10⁻⁷ M，在阻断前阶段或超极化逆转阻断期间测量的通道开放动力学和爆发动力学与对照无差异。5. 当在10⁻⁶ M -精氨酸和10⁻² M -谷氨酸存在下观察到的电位依赖性阻断通过超极化逆转时，爆发期间会出现额外的短暂关闭。这种“闪烁阻断”并未明显改变爆发长度，但出现了一个额外的开放时间成分（τ = 0.1毫秒），且每次爆发的平均开放时间缩短。6. 至少需要两个反应步骤来模拟电位依赖性阻断的行为。闪烁阻断可描述为通道的间歇性阻断，不干扰谷氨酸与通道之间的反应。
