一种改进的适用于膜片钳技术的海马神经元分离方法

An improved method of hippocampal neuron separation suitable for patch-clamp technique

目的探讨获得适用于膜片钳技术的单个海马神经元的方法。方法取3～6日龄Sprague-Dawley新生大鼠10只,应用酶消化及机械分离法,急性分离出大鼠海马神经元,通过倒置显微镜直接观察和全细胞膜片钳技术分别对分离得到的海马细胞的形态学和电生理学特征进行研究。结果急性分离所得海马具有锥形胞体的顶树突特征,立体感强。在全细胞膜片钳记录模式下可记录到全细胞电流及钠通道电流,该电流可被1μmol·L^(-1)的河豚毒素完全阻断。结论应用该酶消化及机械分离法急性分离的新生大鼠海马神经元形态和生理特性良好,适用于海马神经元的膜片钳研究。

Objective To discuss the method of obtaining the single hippocampal neurons suitable for patch-clamp tech- nique. Methods Ten Sprague-Daweley rats （3 -6 days） were taken and the hippocampal neurons were dissociated by enzyme digestion and mechanical separation methods, and morphological and physiological characteristics of the hippocampal cells were studied through the inverted microscope direct observations and whole cell patchclamp technique respectively. Results Hippocampus had a top dendritic characteristics of cone cell bodies and a strong tereo sense. Whole-cell current and sodium channel currents were recorded by whole-cell patch-clamp modle and the current could be completely blocked by 1 μmol .L-1globefish toxins. Conclusion Morphological and physiological characteristics of hippocampal neuronal of newborn rat through enzyme digestion and mechanical separation methods were good and suitable for patch-clamp study of hippocampal neurons.