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哺乳动物中耳蜗内毛细胞带状突触(ribbon synapse)是外界声音信号向大脑听中枢传递路径上的第一个感觉神经突触结构,它在声音的编码和神经递质的释放过程中发挥着重要作用.然而由于带状突触的数量相对比较少,而且空间位置深在,所以仅应用电子显微镜观察的方法定量分析其数量始终在技术上存在一定的困难.本研究通过对C57BL/6J小鼠耳蜗基底膜标本带状突触中的特异突触前结构RIBEYE和非特异突触后结构GluR 2&3同时进行免疫荧光标记,采用激光共聚焦显微镜对标本进行光学连续切片,利用3DS MAX软件对连续切片标本进行三维建模,结果中每个荧光色对代表一个突触的存在.在此基础上对内毛细胞带状突触的数量进行计数,结果显示,耳蜗内毛细胞带状突触的空间分布及数量均可以清晰完整的重现出来.在本实验中,每个内毛细胞带状突触数量平均为(16.10±1.03)个.本实验结果表明,利用免疫荧光对小鼠耳蜗带状突触的前后膜结构进行双重标记,采用激光共聚焦显微镜对标本进行光学连续切片,应用3DS MAX软件对连续切片进行三维建模后可以准确计数带状突触的数量.实验表明,本方法准确可行,对今后深入探讨感觉神经性聋的机制具有重要意义。
The ribbon synapse of mammalian inner ear cochlea is the first sensory synapse structure on the path of the external auditory signal to the auditory nerve of the brain. It plays a role in the process of sound coding and neurotransmitter release However, because of the relatively small number of ribbon synapses, and the spatial location of the deep, so only the application of electron microscopy quantitative analysis of its quantitative methods there are always technically difficult.This study by C57BL / 6J The specific pre-synaptic structure RIBEYE and the non-specific post-synaptic structure GluR 2 & 3 in the striated synapse of mouse cochlea basal lamina were immunofluorescently labeled at the same time, and the specimens were optically cut by laser confocal microscopy. Using 3DS MAX software, Serial sections of specimens for three-dimensional modeling, the results of each fluorescent color on behalf of the existence of a synapse.On the basis of the number of ribbons in the inner hair cells counted, the results show that the cochlear hair cell ribbon synapses The number of spatial distribution and the number can be clearly and completely reproduced in this experiment, the average number of ribbons per inner hair cell (16.10 ± 1.03) The experimental results show that immunofluorescence double labeling of the anterior and posterior membrane structure of cochlear strip synapse in mice, using laser confocal microscopy of optical serial sectioning of the specimen, the use of 3DS MAX software for continuous slice three-dimensional modeling can accurately count The number of banded synapses shows that this method is accurate and feasible, which is of great significance for further exploration of the mechanism of sensory nerve deafness in the future.