Nau C, Wang S Y, Strichartz G R, Wang G K
Department of Anesthesia Research Laboratories, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA.
Anesthesiology. 2000 Oct;93(4):1022-33. doi: 10.1097/00000542-200010000-00026.
S(-)-bupivacaine reportedly exhibits lower cardiotoxicity but similar local anesthetic potency compared with R(+)-bupivacaine. The bupivacaine binding site in human heart (hH1) Na+ channels has not been studied to date. The authors investigated the interaction of bupivacaine enantiomers with hH1 Na+ channels, assessed the contribution of putatively relevant residues to binding, and compared the intrinsic affinities to another isoform, the rat skeletal muscle (mu1) Na+ channel.
Human heart and mu1 Na+ channel alpha subunits were transiently expressed in HEK293t cells and investigated during whole cell voltage-clamp conditions. Using site-directed mutagenesis, the authors created point mutations at positions hH1-F1760, hH1-N1765, hH1-Y1767, and hH1-N406 by introducing the positively charged lysine (K) or the negatively charged aspartic acid (D) and studied their influence on state-dependent block by bupivacaine enantiomers.
Inactivated hH1 Na+ channels displayed a weak stereoselectivity with a stereopotency ratio (+/-) of 1.5. In mutations hH1-F1760K and hH1-N1765K, bupivacaine affinity of inactivated channels was reduced by approximately 20- to 40-fold, in mutation hH1-N406K by approximately sevenfold, and in mutations hH1-Y1767K and hH1-Y1767D by approximately twofold to threefold. Changes in recovery of inactivated mutant channels from block paralleled those of inactivated channel affinity. Inactivated hH1 Na+ channels exhibited a slightly higher intrinsic affinity than mu1 Na+ channels.
Differences in bupivacaine stereoselectivity and intrinsic affinity between hH1 and mu1 Na+ channels are small and most likely of minor clinical relevance. Amino acid residues in positions hH1-F1760, hH1-N1765, and hH1-N406 may contribute to binding of bupivacaine enantiomers in hH1 Na+ channels, whereas the role of hH1-Y1767 remains unclear.
据报道，与R(+)-布比卡因相比，S(-)-布比卡因的心脏毒性较低，但局部麻醉效能相似。迄今为止，尚未对人心肌（hH1）钠通道中的布比卡因结合位点进行研究。作者研究了布比卡因对映体与hH1钠通道的相互作用，评估了可能相关残基对结合的贡献，并比较了其与另一种亚型大鼠骨骼肌（mu1）钠通道的内在亲和力。
人心脏和mu1钠通道α亚基在HEK293t细胞中瞬时表达，并在全细胞电压钳条件下进行研究。作者通过引入带正电荷的赖氨酸（K）或带负电荷的天冬氨酸（D），在hH1-F1760、hH1-N1765、hH1-Y1767和hH1-N406位点进行定点突变，并研究了它们对布比卡因对映体状态依赖性阻滞的影响。
失活的hH1钠通道表现出较弱的立体选择性，立体效价比（+/-）为1.5。在hH1-F1760K和hH1-N1765K突变中，失活通道的布比卡因亲和力降低了约20至40倍，在hH1-N406K突变中降低了约7倍，在hH1-Y1767K和hH1-Y1767D突变中降低了约2至3倍。失活突变通道从阻滞中恢复的变化与失活通道亲和力的变化相似。失活的hH1钠通道表现出比mu1钠通道略高的内在亲和力。
hH1和mu1钠通道之间布比卡因立体选择性和内在亲和力的差异很小，很可能在临床上意义不大。hH1-F1760、hH1-N1765和hH1-N406位点的氨基酸残基可能有助于布比卡因对映体与hH1钠通道的结合，而hH1-Y1767的作用仍不清楚。
