XE991 and Linopirdine Are State-Dependent Inhibitors for Kv7/KCNQ Channels that Favor Activated Single Subunits
XE991 and Linopirdine Are State-Dependent Inhibitors for Kv7/KCNQ Channels that Favor Activated Single Subunits
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DOI：
10.1124/jpet.117.241679
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发表时间：
2017-07-01
影响因子：
3.5
通讯作者：
Hoshi, Naoto
中科院分区：
文献类型：
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作者：
Greene, Derek L.;Kang, Seungwoo;Hoshi, Naoto
M-通道抑制剂，特别是XE 991，在动物实验中的使用越来越多;然而，XE 991的不充分表征有时会混淆使用该化合物时结果的解释。在这里，我们证明，XE 991和linopirdine是状态依赖性抑制剂，有利于激活亚基的神经元Kv 7/KCNQ通道。我们对中国仓鼠卵巢细胞中表达的同源Kv7.2或异源Kv7.2/3通道进行了膜片钳实验，以表征XE 991和利诺吡啶。这两种抑制剂在生理条件下细胞的静息膜电位周围都不有效。XE 991对Kv7.2和Kv7.2/3通道的抑制作用与通道激活密切相关。当激活的电压依赖性由瑞替加滨左移或由突变Kv7.2（R214 D）右移时，半激活电压的偏移与XE 991抑制的半有效电位的偏移成比例地一致。XE 991洗入过程中的抑制动力学在去极化电位下得到促进。XE 991的10分钟洗脱导致类似于30%的电流恢复，其中大部分归因于Kv7.2通道的表面转运。Linopirdine也表现出类似的抑制特性，除了在去极化电位洗脱后几乎完全的电流恢复。XE 991和利诺吡啶的抑制动力学对电压变化不像通过开放通道抑制预测的那样敏感。相反，它们被很好地解释为与单个活化亚基结合。当在实验中使用这些M通道抑制剂时，应考虑XE 991和利诺吡啶的特性。
M-channel inhibitors, especially XE991, are being used increasingly in animal experiments; however, insufficient characterization of XE991 at times confounds the interpretation of results when using this compound. Here, we demonstrate that XE991 and linopirdine are state-dependent inhibitors that favor the activated-subunit of neuronal Kv7/KCNQ channels. We performed patch-clamp experiments on homomeric Kv7.2 or heteromeric Kv7.2/3 channels expressed in Chinese hamster ovary cells to characterize XE991 and linopirdine. Neither inhibitor was efficacious around the resting membrane potential of cells in physiologic conditions. Inhibition of Kv7.2 and Kv7.2/3 channels by XE991 was closely related with channel activation. When the voltage dependence of activation was left-shifted by retigabine or right-shifted by the mutation, Kv7.2(R214D), the shift in half-activation voltage proportionally coincided with the shift in the half-effective potential for XE991 inhibition. Inhibition kinetics during XE991 wash-in was facilitated at depolarized potentials. Ten-minute washout of XE991 resulted in similar to 30% current recovery, most of which was attributed to surface transport of Kv7.2 channels. Linopirdine also exhibited similar inhibition characteristics, with the exception of near-complete current recovery after washout at depolarized potentials. Inhibition kinetics of both XE991 and linopirdine was not as sensitive to changes in voltage as would be predicted by open-channel inhibition. Instead, they were well explained by binding to a single activated subunit. The characteristics of XE991 and linopirdine should be taken into account when these M-channel inhibitors are used in experiments.