College of Life Sciences, Nankai University, Tianjin 300071, China.
Toxicol Appl Pharmacol. 2011 Dec 15;257(3):377-87. doi: 10.1016/j.taap.2011.09.021. Epub 2011 Sep 29.
We expressed rat Na(v)1.6 sodium channels in combination with the rat β1 and β2 auxiliary subunits in human embryonic kidney (HEK293) cells and evaluated the effects of the pyrethroid insecticides tefluthrin and deltamethrin on expressed sodium currents using the whole-cell patch clamp technique. Both pyrethroids produced concentration-dependent, resting modification of Na(v)1.6 channels, prolonging the kinetics of channel inactivation and deactivation to produce persistent "late" currents during depolarization and tail currents following repolarization. Both pyrethroids also produced concentration dependent hyperpolarizing shifts in the voltage dependence of channel activation and steady-state inactivation. Maximal shifts in activation, determined from the voltage dependence of the pyrethroid-induced late and tail currents, were ~25mV for tefluthrin and ~20mV for deltamethrin. The highest attainable concentrations of these compounds also caused shifts of ~5-10mV in the voltage dependence of steady-state inactivation. In addition to their effects on the voltage dependence of inactivation, both compounds caused concentration-dependent increases in the fraction of sodium current that was resistant to inactivation following strong depolarizing prepulses. We assessed the use-dependent effects of tefluthrin and deltamethrin on Na(v)1.6 channels by determining the effect of trains of 1 to 100 5-ms depolarizing prepulses at frequencies of 20 or 66.7Hz on the extent of channel modification. Repetitive depolarization at either frequency increased modification by deltamethrin by ~2.3-fold but had no effect on modification by tefluthrin. Tefluthrin and deltamethrin were equally potent as modifiers of Na(v)1.6 channels in HEK293 cells using the conditions producing maximal modification as the basis for comparison. These findings show that the actions of tefluthrin and deltamethrin of Na(v)1.6 channels in HEK293 cells differ from the effects of these compounds on Na(v)1.6 channels in Xenopus oocytes and more closely reflect the actions of pyrethroids on channels in their native neuronal environment.
我们在人胚肾(HEK293)细胞中表达大鼠 Na(v)1.6 钠通道,并与大鼠 β1 和 β2 辅助亚基结合,然后使用全细胞膜片钳技术评估拟除虫菊酯杀虫剂四氟甲醚菊酯和溴氰菊酯对表达的钠电流的影响。两种拟除虫菊酯均产生浓度依赖性的静息修饰作用,使 Na(v)1.6 通道的失活和去激活动力学延长,从而在去极化期间产生持久的“晚期”电流,并在复极化后产生尾电流。两种拟除虫菊酯还使通道激活和稳态失活的电压依赖性产生浓度依赖性超极化偏移。最大激活偏移量是通过拟除虫菊酯诱导的晚期和尾电流的电压依赖性来确定的,四氟甲醚菊酯约为 25mV,溴氰菊酯约为 20mV。这些化合物的最高可达浓度也使稳态失活的电压依赖性偏移约 5-10mV。除了对失活的电压依赖性的影响外,两种化合物还导致在强去极化预脉冲后对钠电流的失活抗性比例产生浓度依赖性增加。我们通过确定在 20 或 66.7Hz 的频率下用 1 到 100 个 5ms 的去极化预脉冲进行的重复去极化对通道修饰程度的影响来评估四氟甲醚菊酯和溴氰菊酯对 Na(v)1.6 通道的使用依赖性作用。在这两种频率下的重复去极化使溴氰菊酯引起的修饰增加了约 2.3 倍,但对四氟甲醚菊酯引起的修饰没有影响。四氟甲醚菊酯和溴氰菊酯在 HEK293 细胞中作为 Na(v)1.6 通道的调节剂的效力相等,以产生最大修饰的条件作为比较的基础。这些发现表明,四氟甲醚菊酯和溴氰菊酯对 HEK293 细胞中的 Na(v)1.6 通道的作用与这些化合物对 Xenopus oocytes 中的 Na(v)1.6 通道的作用不同,并且更接近拟除虫菊酯在其天然神经元环境中的通道的作用。