果蝇三龄幼虫中枢神经元细胞培养

Dissociated neurons from Drosophila larval central nervous system in cell culture

目的 :建立果蝇幼虫中枢神经元细胞培养方法 ,为研究吸入麻醉药相关的离子通道建立实验模型。方法 :取晚期三龄幼虫的脑腹神经节经解剖、酶解、冲洗、吹打、接种、培养步骤进行分散细胞的原代培养 ,并用膜片钳技术记录全细胞电流以检测培养细胞的功能。结果 :培养的神经元细胞生长活跃 ,状态良好 ,具有神经元的特征 -轴突生长及形成网络连接。以“全细胞方式”记录到的电压依赖性K +电流在不同细胞之间电流动力学和幅度变化较大 ,呈多样性。结论 :果蝇中枢神经元可在体外进行培养 ,方法稳定可靠 ;培养的神经元细胞生长活跃 ,状态良好 ,为进行吸入麻醉药的电生理机制提供良好实验模型。

Objective: To establish a method of primary dissociated cell culture of Drosophila larval central nervous system.Methods:The brain and ventral ganglion of late third instar larvae were dissected, incubated with degestive enxymes, flushed, dispersed into single cells and cultured 1 to 7days. We have used whole-cell patch clamp technique to characterize ion channels in cultured larval Drosophila CNS neurons in order to determine the function of cells. Results: Divisions of neuroblasts and vigorous outgrowth of neurites could be observed in culture.Within 24 hrs cultured cells exhibited characteristic neuronal morphology-axon like processes and forming contacts with other cells. Whole-cell currents from different cells varied in current kinetics and magnitude. Conclusion: The central nervous system cells could be cultured in vitro. Divisions of neuroblasts and vigorous outgrowth of neurites could be observed in culture.So this model provided the possibility to study the electrical mechanism of inhaled anesthetics.