脑细胞内NADH的多光子、无标记光声和光学成像

IF 23.4 Q1 OPTICS
Tatsuya Osaki, W. David Lee, Xiang Zhang, Rebecca E. Zubajlo, Mercedes Balcells-Camps, Elazer R. Edelman, Brian W. Anthony, Mriganka Sur, Peter T. C. So
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引用次数: 0

摘要

在大脑中以单细胞分辨率对生物事件进行无标记检测,可以无创地捕获大脑状态,用于医学诊断和基础神经科学研究。NADH是一种通用的辅酶,不仅在细胞代谢中起着核心作用,而且还可以作为生物标志物来捕捉脑细胞和结构中的代谢过程。利用近红外飞秒激光,研制了一种新型无标记多光子光声显微镜(LF-MP-PAM),用于观察活细胞内源性NAD(P)H。NAD(P)H对近紫外荧光的强吸收使其在脑组织中的成像深度限制在~100 μm。在这里,低量子产率荧光团NAD(P)H的多光子(三光子)激发的热特征的声学检测允许在前所未有的深度进行检测,而聚焦激发确保了高空间分辨率。我们通过监测在NADH溶液中培养的HEK293T细胞和HepG2细胞内NAD(P)H的增加,验证了光声检测NAD(P)H。我们还证明了在700 μm深度的脑切片和1100 μm深度的脑类器官中检测到内源性NAD(P)H光声信号。最后,我们通过实时图像采集和处理管道开发并演示了脑细胞中NAD(P)H的同时光声和光学成像。这种方法可以在人类和动物大脑深处的单细胞水平上,为监测大脑发育和疾病期间的代谢变化,以及神经元活动引起的变化打开一扇新的大门。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Multi-photon, label-free photoacoustic and optical imaging of NADH in brain cells

Multi-photon, label-free photoacoustic and optical imaging of NADH in brain cells

Label-free detection of biological events at single-cell resolution in the brain can non-invasively capture brain status for medical diagnosis and basic neuroscience research. NADH is an universal coenzyme that not only plays a central role in cellular metabolism but may also be used as a biomarker to capture metabolic processes in brain cells and structures. We have developed a new label-free, multiphoton photoacoustic microscope (LF-MP-PAM) with a near-infrared femtosecond laser to observe endogenous NAD(P)H in living cells. The imaging depth of NAD(P)H in tissues with all-optical methods is limited to ~100 μm in brain tissue by the strong absorption of the near-ultraviolet fluorescence. Here, acoustic detection of the thermal signature of multi-photon (three-photon) excitation of NAD(P)H, a low quantum yield fluorophore, allows detection at an unprecedented depth while the focused excitation ensures high spatial resolution. We validated the photoacoustic detection of NAD(P)H by monitoring an increase in intracellular NAD(P)H in HEK293T cells and HepG2 cells incubated in NADH solution. We also demonstrated the detection of endogenous NAD(P)H photoacoustic signals in brain slices to 700 μm depth and in cerebral organoids to 1100 μm depth. Finally, we developed and demonstrated simultaneous photoacoustic and optical imaging of NAD(P)H in brain cells with a real-time image acquisition and processing pipeline. This approach could open a new door to monitor brain metabolic changes during development and disease, and changes due to neuronal activity, at single-cell level deep in the brains of both humans and animals.

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来源期刊
Light-Science & Applications
Light-Science & Applications 数理科学, 物理学I, 光学, 凝聚态物性 II :电子结构、电学、磁学和光学性质, 无机非金属材料, 无机非金属类光电信息与功能材料, 工程与材料, 信息科学, 光学和光电子学, 光学和光电子材料, 非线性光学与量子光学
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发文量
803
审稿时长
2.1 months
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