Department of Applied Physics and Science for Life Laboratory, KTH Royal Institute of Technology, 100 44 Stockholm, Sweden.
Department of Applied Physics and Science for Life Laboratory, KTH Royal Institute of Technology, 100 44 Stockholm, Sweden.
Neurobiol Dis. 2021 Jul;155:105361. doi: 10.1016/j.nbd.2021.105361. Epub 2021 Apr 12.
The classic view of organelle cell biology is undergoing a constant revision fueled by the new insights unraveled by fluorescence nanoscopy, which enable sensitive, faster and gentler observation of specific proteins in situ. The endoplasmic reticulum (ER) is one of the most challenging structure to capture due the rapid and constant restructuring of fine sheets and tubules across the full 3D cell volume. Here we apply STED and parallelized 2D and 3D RESOLFT live imaging to uncover the tubular ER organization in the fine processes of neuronal cells with focus on mitochondria-ER contacts, which recently gained medical attention due to their role in neurodegeneration. Multi-color STED nanoscopy enables the simultaneous visualization of small transversal ER tubules crossing and constricting mitochondria all along axons and dendrites. Parallelized RESOLFT allows for dynamic studies of multiple contact sites within seconds and minutes with prolonged time-lapse imaging at ~50 nm spatial resolution. When operated in 3D super resolution mode it enables a new isotropic visualization of such contacts extending our understanding of the three-dimensional architecture of these packed structures in axons and dendrites.
细胞器细胞生物学的经典观点正在不断修正，这得益于荧光纳米技术所揭示的新见解，该技术能够在体内对特定蛋白质进行敏感、快速和温和的观察。内质网（ER）是最难捕捉的结构之一，因为细片和小管在整个 3D 细胞体积中快速且不断地重构。在这里，我们应用 STED 和并行的 2D 和 3D RESOLFT 活细胞成像，揭示神经元细胞的精细过程中的管状 ER 组织，重点关注线粒体-ER 接触，由于它们在神经退行性变中的作用，最近引起了医学界的关注。多色 STED 纳米技术能够同时可视化穿过并收缩线粒体的小横向 ER 小管，这些小管遍布轴突和树突。并行的 RESOLFT 允许在几秒钟和几分钟内对多个接触点进行动态研究，并通过延长的延时成像以 ~50nm 的空间分辨率进行长时间延时成像。当在 3D 超分辨率模式下操作时，它能够对这些接触点进行新的各向同性可视化，从而扩展我们对这些在轴突和树突中紧密堆积结构的三维结构的理解。
