Tang Qiang, Jin Man-Wen, Xiang Ji-Zhou, Dong Min-Qing, Sun Hai-Ying, Lau Chu-Pak, Li Gui-Rong
Department of Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong, SAR, China.
Biochem Pharmacol. 2007 Dec 3;74(11):1596-607. doi: 10.1016/j.bcp.2007.07.042. Epub 2007 Aug 3.
BAPTA-AM is a well-known membrane permeable Ca(2+) chelator. The present study found that BAPTA-AM rapidly and reversibly suppressed human ether a-go-go-related gene (hERG or Kv11.1) K(+) current, human Kv1.3 and human Kv1.5 channel currents stably expressed in HEK 293 cells, and the effects were not related to Ca(2+) chelation. The externally applied BAPTA-AM inhibited hERG channels in a concentration-dependent manner (IC(50): 1.3 microM). Blockade of hERG channels was dependent on channel opening, and tonic block was minimal. Steady-state activation V(0.5) of hERG channels was negatively shifted by 8.5 mV (from -3.7+/-2.8 of control to -12.2+/-3.1 mV, P<0.01), while inactivation V(0.5) was negatively shifted by 6.1 mV (from -37.9+/-2.0 mV of control to -44.0+/-1.6 mV, P<0.05) with application of 3 microM BAPTA-AM. The S6 mutant Y652A and the pore helix mutant S631A significantly attenuated blockade by BAPTA-AM at 10 microM causing profound blockade of wild-type hERG channels. In addition, BAPTA-AM inhibited hKv1.3 and hKv1.5 channels in a concentration-dependent manner (IC(50): 1.45 and 1.23 microM, respectively), and the blockade of these two types of channels was also dependent on channel opening. Moreover, EGTA-AM was found to be an open channel blocker of hERG, hKv1.3, hKv1.5 channels, though its efficacy is weaker than that of BAPTA-AM. These results indicate that the membrane permeable Ca(2+) chelator BAPTA-AM (also EGTA-AM) exerts an open channel blocking effect on hERG, hKv1.3 and hKv1.5 channels.
1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸-乙酰甲酯（BAPTA-AM）是一种著名的可透过细胞膜的钙离子螯合剂。本研究发现，BAPTA-AM能快速且可逆地抑制人醚-去极化相关基因（hERG或Kv11.1）钾电流、在人胚肾293（HEK 293）细胞中稳定表达的人Kv1.3和人Kv1.5通道电流，且这些效应与钙离子螯合无关。细胞外施加的BAPTA-AM以浓度依赖性方式抑制hERG通道（半数抑制浓度[IC50]：1.3微摩尔）。hERG通道的阻断依赖于通道开放，持续性阻断作用极小。应用3微摩尔BAPTA-AM时，hERG通道的稳态激活半值电压（V0.5）负向偏移8.5毫伏（从对照的-3.7±2.8毫伏变为-12.2±3.1毫伏，P<0.01），而失活半值电压负向偏移6.1毫伏（从对照的-37.9±2.0毫伏变为-44.0±1.6毫伏，P<0.05）。S6突变体Y652A和孔螺旋突变体S631A显著减弱了10微摩尔BAPTA-AM对野生型hERG通道的深度阻断作用。此外，BAPTA-AM以浓度依赖性方式抑制hKv1.3和hKv1.5通道（IC50分别为1.45和1.23微摩尔），对这两种通道的阻断也依赖于通道开放。此外，虽然乙二醇双（2-氨基乙基醚）四乙酸-乙酰甲酯（EGTA-AM）的效力比BAPTA-AM弱，但它也是hERG、hKv1.3、hKv1.5通道的开放通道阻断剂。这些结果表明，可透过细胞膜的钙离子螯合剂BAPTA-AM（以及EGTA-AM）对hERG、hKv1.3和hKv1.5通道发挥开放通道阻断作用。
