From The Institute of Academic Anaesthesia, Division of Neuroscience, School of Medicine, Ninewells Hospital, University of Dundee, Dundee, United Kingdom.
Anesth Analg. 2018 Sep;127(3):650-660. doi: 10.1213/ANE.0000000000003597.
Cardiotoxic effects of local anesthetics (LAs) involve inhibition of NaV1.5 voltage-gated Na channels. Metastatic breast and colon cancer cells also express NaV1.5, predominantly the neonatal splice variant (nNaV1.5) and their inhibition by LAs reduces invasion and migration. It may be advantageous to target cancer cells while sparing cardiac function through selective blockade of nNaV1.5 and/or by preferentially affecting inactivated NaV1.5, which predominate in cancer cells. We tested the hypotheses that lidocaine and levobupivacaine differentially affect (1) adult (aNaV1.5) and nNaV1.5 and (2) the resting and inactivated states of NaV1.5.
The whole-cell voltage-clamp technique was used to evaluate the actions of lidocaine and levobupivacaine on recombinant NaV1.5 channels expressed in HEK-293 cells. Cells were transiently transfected with cDNAs encoding either aNaV1.5 or nNaV1.5. Voltage protocols were applied to determine depolarizing potentials that either activated or inactivated 50% of maximum conductance (V½ activation and V½ inactivation, respectively).
Lidocaine and levobupivacaine potently inhibited aNaV1.5 (IC50 mean [SD]: 20 [22] and 1 [0.6] μM, respectively) and nNaV1.5 (IC50 mean [SD]: 17 [10] and 3 [1.6] μM, respectively) at a holding potential of -80 mV. IC50s differed significantly between lidocaine and levobupivacaine with no influence of splice variant. Levobupivacaine induced a statistically significant depolarizing shift in the V½ activation for aNaV1.5 (mean [SD] from -32 [4.6] mV to -26 [8.1] mV) but had no effect on the voltage dependence of activation of nNaV1.5. Lidocaine had no effect on V½ activation of either variant but caused a significantly greater depression of maximum current mediated by nNaV1.5 compared to aNaV1.5. Similar statistically significant shifts in the V½ inactivation (approximately -10 mV) occurred for both LAs and NaV1.5 variants. Levobupivacaine (1 μM) caused a significantly greater slowing of recovery from inactivation of both variants than did lidocaine (10 μM). Both LAs caused approximately 50% tonic inhibition of aNaV1.5 or nNaV1.5 when holding at -80 mV. Neither LA caused tonic block at a holding potential of either -90 or -120 mV, voltages at which there was little steady-state inactivation. Higher concentrations of either lidocaine (300 μM) or levobupivacaine (100 μM) caused significantly more tonic block at -120 mV.
These data demonstrate that low concentrations of the LAs exhibit inactivation-dependent block of NaV1.5, which may provide a rationale for their use to safely inhibit migration and invasion by metastatic cancer cells without cardiotoxicity.
局部麻醉剂（LA）的心脏毒性作用涉及抑制 NaV1.5 电压门控 Na 通道。转移性乳腺癌和结肠癌细胞也表达 NaV1.5，主要是新生剪接变异体（nNaV1.5），LA 对其的抑制作用可降低侵袭和迁移。通过选择性阻断 nNaV1.5 和/或通过优先影响在癌细胞中占优势的失活 NaV1.5 来靶向癌细胞，同时保留心脏功能可能是有利的。我们测试了以下假设：利多卡因和左布比卡因（levobupivacaine）（1）对成人（aNaV1.5）和 nNaV1.5 有不同的影响；（2）对 NaV1.5 的静息和失活状态有不同的影响。
使用全细胞膜片钳技术评估利多卡因和左布比卡因对表达于 HEK-293 细胞的重组 NaV1.5 通道的作用。细胞瞬时转染编码 aNaV1.5 或 nNaV1.5 的 cDNA。应用电压程序来确定去极化电位，这些电位要么激活，要么失活 50%的最大电导（分别为 V½激活和 V½失活）。
在 -80 mV 的保持电位下，利多卡因和左布比卡因均能有效抑制 aNaV1.5（IC50 平均值 [标准差]：分别为 20 [22] 和 1 [0.6] μM）和 nNaV1.5（IC50 平均值 [标准差]：分别为 17 [10] 和 3 [1.6] μM）。IC50 显著不同利多卡因和左布比卡因之间无剪接变异体的影响。左布比卡因引起 aNaV1.5 的 V½激活发生统计学上显著的去极化偏移（从 -32 [4.6] mV 到 -26 [8.1] mV），但对 nNaV1.5 的激活电压依赖性没有影响。利多卡因对任一变体的 V½激活均无影响，但与 aNaV1.5 相比，nNaV1.5 介导的最大电流的抑制作用明显更大。两种 LA 和 NaV1.5 变体的 V½失活（约 -10 mV）也发生类似的统计学显著偏移。与利多卡因（10 μM）相比，左布比卡因（1 μM）导致两种变体的失活恢复明显减慢。两种 LA 在 -80 mV 时均导致 aNaV1.5 或 nNaV1.5 产生约 50%的持续抑制。LA 在 -80 mV 或 -90 mV 或 -120 mV 的保持电位下均不引起持续阻断，这些电压下基本没有稳态失活。较高浓度的利多卡因（300 μM）或左布比卡因（100 μM）在 -120 mV 时引起的持续阻断明显更多。
这些数据表明，低浓度的 LA 表现出失活依赖性的 NaV1.5 阻断，这可能为其安全抑制转移性癌细胞的迁移和侵袭而不产生心脏毒性提供了一个合理的依据。
