Kir 2.3过表达抑制原代海马神经元样放电

Kir 2.3 Overexpression Inhibits Epileptiform Discharges in Primary Cultured Hippocampus Neurons

目的探究内向整流钾(Kir) 2. 3通道过表达对环噻唑诱导的原代海马神经元样放电的影响。方法分离培养大鼠原代海马神经元,分别使用GFP质粒或Kir 2. 3-GFP质粒转染神经元。通过免疫荧光染色和电压钳技术检测转染效果。使用环噻唑诱导神经元样放电,电流钳记录膜电位钳制在-70 m V条件下的自发放电,膜片钳记录各组神经元的膜电位。结果转染Kir 2. 3-GFP质粒的神经元Kir 2. 3表达水平显著增高(P <0. 05)。环噻唑能够诱导未转染神经元(n=12)及GFP质粒转染神经元(n=6)出现样爆发放电,但是未能诱导Kir 2. 3-GFP质粒转染神经元(n=5)出现样爆发放电。Kir 2. 3-GFP质粒转染组出现样爆发放电神经元的比例显著低于未转染组(P <0. 01)及GFP质粒转染组(P <0. 05)。此外,Kir 2. 3-GFP质粒转染组(n=10)神经元膜电位较未转染组(n=6)及GFP质粒转染组(n=8)显著增高。结论 Kir 2. 3过表达能显著抑制环噻唑诱导的原代海马神经元样放电,其作用机制可能与静息膜电位增高相关。

Aim To investigate the effect of Kir 2. 3 overexpression on epileptiform discharges of primary cultured hippocampus neurons induced by cyclothiazide( CTZ). Methods Rat hippocampus neurons were primarily cultured and transfected with plasmid containing either GFP or Kir 2. 3-GFP.Immunofluorescence staining and voltage clamp technique were used to check the transfection effect.Epileptiform discharges were induced by CTZ,and the spontaneous discharge were recorded by current clamp with the membrane potential held around-70 m V. Membrane potential of neurons in different group were determined by patch clamp. Results Kir 2. 3 expression of neurons transfected with Kir 2. 3-GFP plasmid augmented significantly( P < 0. 05). Epileptiform burst discharges were induced by CTZ in non-transfected neurons( n = 12) or GFP plasmid transfected control neurons( n = 6) but failed in neurons with Kir 2. 3-GFP plasmid transfected( n = 5). The proportion of burst neurons in Kir 2. 3-GFP plasmid transfected neurons were significantly lower than that of non-transfected neurons( P < 0. 01) and GFP plasmid transfected neurons( P < 0. 05). Furthermore,the membrane potential of Kir 2. 3-GFP transfected neurons increased significantly compared with non-transfected neurons and GFP plasmid transfected control neurons.Conclusion Kir 2. 3 overexpression could significantly suppress the epileptiform discharges of primary cultured hippocampus neurons induced by CTZ,which might result from the augment of the membrane potential.