Live Cell Light Sheet Imaging with Low- and High-Spatial-Coherence Detection Approaches Reveals Spatiotemporal Aspects of Neuronal Signaling.
Live Cell Light Sheet Imaging with Low- and High-Spatial-Coherence Detection Approaches Reveals Spatiotemporal Aspects of Neuronal Signaling.
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DOI：
10.3390/jimaging9060121
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发表时间：
2023-06-16
影响因子：
3.2
通讯作者：
中科院分区：
文献类型：
--
作者：
活细胞中的光片显微镜需要最小的激发强度，并快速解析三维（3D）信息。晶格光片显微镜（LLSM）的工作原理类似，但使用贝塞尔光束的晶格配置来生成更平坦的衍射限制z轴片，适用于研究亚细胞区室，具有更好的组织穿透性。我们开发了一种LLSM方法，用于原位研究组织的细胞特性。神经结构提供了一个重要的目标。神经元是复杂的3D结构，细胞和亚细胞结构之间的信号传导需要高分辨率成像。我们开发了一个LLSM配置的基础上Janelia研究校园的设计或原位记录，允许同时电生理记录。我们给出了使用LLSM原位评估突触功能的例子。在突触前，诱发的Ca2+内流引起囊泡融合和神经递质释放。我们证明了使用LLSM来测量刺激诱发的局部突触前Ca2+进入和跟踪突触囊泡回收。我们还证明了在单个突触中突触后Ca2+信号的分辨率。3D成像中的一个挑战是需要移动发射物镜以保持聚焦。我们发展了一种非相干全息点阵光片（IHLLS）技术，用双衍射透镜代替LLS管透镜，获得了物体衍射的空间非相干光的三维图像。在扫描体积内再现3D结构而不移动发射物镜。这消除了机械伪影并提高了时间分辨率。我们专注于LLS和IHLLS应用和神经科学中获得的数据，并强调使用这些方法提高时间和空间分辨率。
Light sheet microscopy in live cells requires minimal excitation intensity and resolves three-dimensional (3D) information rapidly. Lattice light sheet microscopy (LLSM) works similarly but uses a lattice configuration of Bessel beams to generate a flatter, diffraction-limited z-axis sheet suitable for investigating subcellular compartments, with better tissue penetration. We developed a LLSM method for investigating cellular properties of tissue in situ. Neural structures provide an important target. Neurons are complex 3D structures, and signaling between cells and subcellular structures requires high resolution imaging. We developed an LLSM configuration based on the Janelia Research Campus design or in situ recording that allows simultaneous electrophysiological recording. We give examples of using LLSM to assess synaptic function in situ. In presynapses, evoked Ca2+ entry causes vesicle fusion and neurotransmitter release. We demonstrate the use of LLSM to measure stimulus-evoked localized presynaptic Ca2+ entry and track synaptic vesicle recycling. We also demonstrate the resolution of postsynaptic Ca2+ signaling in single synapses. A challenge in 3D imaging is the need to move the emission objective to maintain focus. We have developed an incoherent holographic lattice light-sheet (IHLLS) technique to replace the LLS tube lens with a dual diffractive lens to obtain 3D images of spatially incoherent light diffracted from an object as incoherent holograms. The 3D structure is reproduced within the scanned volume without moving the emission objective. This eliminates mechanical artifacts and improves temporal resolution. We focus on LLS and IHLLS applications and data obtained in neuroscience and emphasize increases in temporal and spatial resolution using these approaches.
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影响因子：
2.9
作者：
SPENCER, HJ;GRIBKOFF, VK;LYNCH, GS
通讯作者：
LYNCH, GS
DOI：
10.1073/pnas.1315858110
发表时间：
2013-12-24
影响因子：
11.1
作者：
Schulz, Olaf;Pieper, Christoph;Enderlein, Joerg
通讯作者：
Enderlein, Joerg
影响因子：
3.8
作者：
Ga, Liang;Tang, Wei-Chun;Chen, Bi-Chang
通讯作者：
Chen, Bi-Chang
影响因子：
2.9
作者：
PETSAS, KI;COATES, AB;GRYNBERG, G
通讯作者：
GRYNBERG, G
影响因子：
3.8
作者：
Potcoava, Mariana;Mann, Christopher;Alford, Simon
通讯作者：
Alford, Simon