SH-EP1细胞株转染表达神经元N受体α4β2、α4β4和α7亚型激活态的电生理学特征

ELECTROPHYSIOLOGICAL CHARACTERISTICS OF THE OPEN STATE OF NEURONAL α4β2,α4β4 AND α7-NICOTINIE ACETYLCHOLINE RECEPTORS

目的:建立SH-EP1细胞株转染表达的神经元N受体α4β2、α4β4和α7亚型的膜片钳全细胞记录技术并研究其激活态的电生理学特征。方法:用高糖DMEM培养液培养细胞。传代3d后,细胞可用于膜片钳全细胞记录。通过多管给药电极喷射给予激动剂烟碱(α4β2,α4β4)或胆碱(α7)诱导受体由静息态转变为激活态。结果:传代培养的SH-EP1细胞功能状态良好,受体表达丰富。α4β2、α4β4和α7三种亚型均可被激动剂快速激活诱发内向电流,且该三种通道的内向电流均具有激动剂浓度依赖性、电压依赖性和内向整流特性。三种通道由静息态转变为激活态、以及激活态受体数目减少的速度各不相同。其中,α7亚型激活速度最快,失活速度也最快;α4β4亚型激活速度最慢,失活速度也最慢;α4β2亚型则均居中。结论:在激动剂作用下神经元N受体α4β2、α4β4和α7亚型激活态的形成和转变各具特点,但其介导的内向电流均具有激动剂浓度依赖性、电压依赖性和内向整流特性。

Aim： To establish the whole-cell recording techniques of the neuronal α4β2, α4β4 and α7-nicotinic acetylcholine receptors heterologously expressed in SH-EP1 cell line and discuss the electrophysiological characteristics of their open states. Methods： The cells were cultured with DEME medium（high glucose） and suitable for electrophysiological experiments three days after passage. The receptors were induced from resting states into open states by rapid application of nicotine（α4β2, α4β4） or choline（α7）. Results： The SH-EP1 cells cultured by this method were in good conditions and expressed plenty of receptors, α4β2, α4β4 and α7 inward currents could be induced by rapid application of agonists but had different dynamic processes against time. All the three types of currents were dose and voltage-dependent and had inward rectification property. Conclusion： The open states of neuronal α4β2, α4β4 and α7-uicotinic acetylchuline receptors and their transitions have distinct characteristics and the inward currents of all this three types of receptors are dose and voltage-dependent and have inward rectification property.