FM1-43造影观察PC12神经元与原代皮层神经元间的突触形成

Neuronal PC12 cells develop functional synapses with neurons from neonatal cortex (a FM1-43 imaging investigation)

目的 观察种植到原代皮层神经元中的PC12细胞,在神经生长因子(nervegrowthfactorNGF)作用下分化成的神经元样细胞与原代皮层神经元之间功能性突触的形成,研究细胞移植环境中宿主细胞和移植细胞形成神经连接的可能及过程。方法 将绿色荧光蛋白(enhencedgreenflourescentproteinEGFP)标记的PC12细胞种植到原代神经元中,建立共培养系统。NGF诱导其中的PC12细胞分化为神经元样细胞,可能与共培养的原代神经元形成突触连接。高钾离子的去极化刺激使突触末端囊泡被FM1 43染色,激光共聚焦扫描显微镜下观察,电子图像显示出活性的突触末端。结果 和对照组相比共培养系统中PC12神经元更成熟,FM1 43造影显示这些新的神经元与原代神经元之间产生了功能性突触囊泡活动。结论 与原代的神经细胞共培养有利于PC12细胞在NGF作用下分化为神经元样细胞及形成神经突起,新形成的神经元可以和宿主神经元形成突触连接。

Objective To investigate whether neuronal PC12 cells, when cultured in a transplantation simulating co cultured system, could build functional synapses with primary neurons from neonatal cortex. Methods PC12 cells were transfected to stably express enhanced green flourescent protein (EGFP), and then these cells were transplanted into primary neural cells from neonatal cortex. 25ng/ml NGF was added into the medium to induce the differentiation of PC12 cells. Synaptic terminals were dyed with FM1 43 using a depolarizing stimulus to induce exocytosis. Results Compared with the control group(PC12 cells cultured alone), PC12 cells co cultured with primary neural cells differentiated into more mature neuron like cells when induced by NGF. FM1 43 imaging demonstrated that these neuron like cells had synaptic exocytosis with primary neurons. Conclusions Co cultured with primary neural cells is helpful for PC12 cells′ differentiation and their neurite growth induced by NGF. These PC12 derived neuron like cells could successfully form functional synapses with primary neurons.