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首页> 外文期刊>Journal of Neuroscience Methods >Identification of neuromuscular junctions by correlative confocal and transmission electron microscopy.
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Identification of neuromuscular junctions by correlative confocal and transmission electron microscopy.

机译：通过相关的共聚焦和透射电子显微镜鉴定神经肌肉接头。

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摘要

The physiological processes regulating neuromuscular transmission are highly dependent on the structural features of the motor neuron and motor endplate, and detailing the structure of neuromuscular junctions (NMJs) in muscle biopsies is a powerful method for research and diagnostics. The observation of NMJ ultrastructure, however, is complicated by the difficulty in locating NMJs for analysis by electron microscopy. Consequently, a correlative confocal-transmission electron microscopy method was developed. Fixed muscle samples were cryo-protected in sucrose, sectioned on a cryostat, and stained with fluorescent alpha-bungarotoxin for confocal microscopy. Sections containing junctions were mapped and then processed for transmission electron microscopy (TEM). Cryostat sections allowed large expanses of muscle tissue to be rapidly screened and enabled specific junctions to be targeted for TEM. The morphology of the junctions was well preserved with all essential features of the pre- and postsynaptic elements readily identifiable without freeze damage. Unlike NMJ correlative methods using histochemical stains and DAB photo-oxidation, no electron dense precipitate was deposited over the NMJ, enabling an unobstructed view of the pre- and postsynaptic structures.
机译：调节神经肌肉传递的生理过程高度依赖于运动神经元和运动终板的结构特征，在肌肉活检中详述神经肌肉接头（NMJ）的结构是研究和诊断的有力方法。然而，由于难以定位用于通过电子显微镜分析的NMJ，因此NMJ超微结构的观察变得复杂。因此，发展了一种相关的共聚焦透射电子显微镜方法。将固定的肌肉样品在蔗糖中冷冻保护，在低温恒温器上切片，并用荧光α-真菌毒素染色以进行共聚焦显微镜检查。绘制包含结的切片，然后进行透射电子显微镜（TEM）处理。低温恒温器切片可快速筛查大面积的肌肉组织，并使特定的接合处成为TEM的靶标。交界处的形态被很好地保留，突触前和突触后元件的所有基本特征都易于识别，而不会受到冻害。与使用组织化学染色剂和DAB光氧化的NMJ相关方法不同，在NMJ上没有电子致密的沉淀物沉积，因此可以清晰地观察突触前和突触后的结构。

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