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目的观察钾通道阻断剂四乙铵(TEA)对缺氧缺血损伤整体、离体模型的保护作用,阐明Kv2.1电压依赖性钾通道在缺氧缺血细胞损伤中发挥的作用。方法应用大鼠暂时性大脑中动脉阻塞(tMCAO)脑缺血模型验证TEA对脑缺血损伤的保护作用。采用膜片钳技术观察氧糖剥夺(OGD)对稳定转染Kv2.1钾通道的人胚胎肾293(HEK293)细胞膜电位以及TEA对OGDKv2.1-HEK293细胞钾电流的影响。MTT法观察OGD对Kv2.1-HEK293的损伤及TEA的保护作用。结果TEA(5μg·kg-1)侧脑室注射可以显著减小tMCAO大鼠的脑梗死体积;应用转基因细胞的研究证实Kv2.1-HEK293对OGD损伤的敏感性明显提高;OGD可以降低Kv2.1-HEK293细胞膜电位;TEA(10mmol·L-1)能显著抑制OGDKv2.1-HEK293钾电流,同时使其所受的细胞损伤降低。结论钾通道阻断剂TEA对整体和离体缺氧缺血损伤模型均发挥细胞保护作用,这种保护作用与对Kv2.1的阻断密切相关;提示Kv2.1可能是抗脑缺血药物开发的潜在靶点。
Objective To observe the protective effect of tetraethylammonium (TEA), a blocker of potassium channel, on global and in vitro models of hypoxic-ischemic injury and to elucidate the role of Kv2.1-voltage-dependent potassium channel in hypoxic-ischemic injury. Methods The protective effect of TEA on cerebral ischemic injury was validated by transient middle cerebral artery occlusion (tMCAO) in rats. The effects of oxygen glucose deprivation (OGD) on membrane potential of human embryonic kidney 293 (HEK293) stably transfected with Kv2.1 potassium channel and the effect of TEA on the potassium current of OGDKv2.1-HEK293 cells were observed by patch clamp technique. MTT assay OGD on Kv2.1-HEK293 injury and TEA protective effect. Results Intracerebroventricular injection of TEA (5μg · kg -1) significantly reduced the infarction volume in tMCAO rats. The transgenic mice showed that the sensitivity of Kv2.1-HEK293 to OGD injury was significantly increased. OGD decreased the expression of Kv2.1 -HEK293 cell membrane potential; TEA (10mmol·L-1) can significantly inhibit the OGDKv2.1-HEK293 potassium current, while reducing cell damage suffered by them. Conclusion Potassium channel blocker TEA plays a protective role on both global and isolated hypoxic-ischemic injury models. This protective effect is closely related to the blockade of Kv2.1, suggesting that Kv2.1 may be an anti-cerebral ischemia drug Development of potential targets.