原代培养神经细胞膜经N-甲基-D-天冬氨酸和MK-801作用后原子力显微镜观察

Atomic force microscopic observation of the neurons membrane in primary culture after treatment with N-Methyl-D-Asph

目的：对比观察正常培养皮层神经元在N－甲基－D－天冬氨酸（NMDA）及其受体拮抗剂－MK－801作用下膜表面三维构像形态的改变。方法：利用分辨率为0．1－0．01nm的原子力显微镜（AFM）对原代培养大鼠皮层神经细胞膜表面进行纳米尺度的扫描观测。结果：正常神经元膜表面光滑，起伏均匀，隆起的颗粒状蛋白密集，间隔规律。NMDA损伤后神经元破碎，崩解，膜失去连续性，NMDA＋MK－801作用下神经元膜皱折增加，边缘粗糙，起伏程度介于前两者之间，结论：（1）AFM具有分辨率高，制样简单特点，（2）AFM能细微地分辨损伤保护作用后引起的细胞膜表面三维形态改变。（3）NMDA作用后膜结构开始解体，膜蛋白颗粒聚集增大，脂质凹陷加深，间距增宽，表面粗糙度增加。

Objective To observe the changes in three-dimensional morpholog y of the membrane of the cortex neurons in pri-mary culture in response to treatm ent with N-Methyl-D-Asp (NMDA) and MK-801 using atomic force microscope (AFM). Methods Following NMDA treatment, the changes in the membrane of the neurons fix ed by glutaraldehyhyde were examined, and the protective effect of MK-801 on th e neurons was also observed using an ATM at the resolution of 0.01-0.1 nm. Resul ts Normal neurons presented smooth membrane surface with regular undulation and densely but well arrayed protein granules. In response to NMDA treatment, the n eurons were disrupted, falling into small pieces, and their membrane appeared di scontinu-ous. MK-801 treatment increased the folds of the membrane that showed r ough edges, with membrane undulation only sec-ondary to normal condition. Conclu sions AFM, with the merits of high resolution and easy sample preparation, can f inely dis-play the three-dimensional morphology of the surface of the neurons, w hich, after NMDA treatment, becomes disintegrated with increased bulk of the pr otein granules, deepened caving of the lipids, and rougher membrane surface.