Department of Biomedical Engineering, Vanderbilt University, Nashville, Tennessee.
Department of Biomedical Engineering, Vanderbilt University, Nashville, Tennessee; Department of Neurosurgery, Vanderbilt University Medical Center, Nashville, Tennessee.
Biophys J. 2022 Apr 19;121(8):1525-1540. doi: 10.1016/j.bpj.2022.03.006. Epub 2022 Mar 8.
Infrared neural stimulation (INS) uses pulsed infrared light to yield label-free neural stimulation with broad experimental and translational utility. Despite its robust demonstration, INS's mechanistic and biophysical underpinnings have been the subject of debate for more than a decade. The role of lipid membrane thermodynamics appears to play an important role in how fast IR-mediated heating nonspecifically drives action potential generation. Direct observation of lipid membrane dynamics during INS remains to be shown in a live neural model system. We used hyperspectral stimulated Raman scattering microscopy to study biochemical signatures of high-speed vibrational dynamics underlying INS in a live neural cell culture model. The findings suggest that lipid bilayer structural changes occur during INS in vitro in NG108-15 neuroglioma cells. Lipid-specific signatures of cell stimulated Raman scattering spectra varied with stimulation energy and radiation exposure. The spectroscopic observations agree with high-speed ratiometric fluorescence imaging of a conventional lipophilic membrane structure reporter, 4-(2-(6-(dibutylamino)-2-naphthalenyl)ethenyl)-1-(3-sulfopropyl)pyridinium hydroxide. The findings support the hypothesis that INS causes changes in the lipid membrane of neural cells by changing the lipid membrane packing order. This work highlights the potential of hyperspectral stimulated Raman scattering as a method to safely study biophysical and biochemical dynamics in live cells.
红外神经刺激 (INS) 使用脉冲红外光实现无标记神经刺激，具有广泛的实验和转化应用。尽管已经得到了强有力的证明，但 INS 的机制和生物物理基础已经成为十多年来争论的主题。脂质膜热力学的作用似乎在 IR 介导的加热如何快速非特异性地驱动动作电位产生方面起着重要作用。在活体神经模型系统中，仍需要直接观察 INS 期间脂质膜动力学。我们使用高光谱受激拉曼散射显微镜研究了活体神经细胞培养模型中 INS 下高速振动动力学的生化特征。研究结果表明，在体外 NG108-15 神经胶质瘤细胞中进行 INS 时会发生脂质双层结构变化。细胞受激拉曼散射光谱的脂质特异性特征随刺激能量和辐射暴露而变化。这些光谱观察结果与传统亲脂性膜结构报告器 4-(2-(6-(二丁基氨基)-2-萘基)乙烯基)-1-(3-磺丙基)吡啶𬭩氢氧化物的高速比率荧光成像一致。这些发现支持了 INS 通过改变脂质膜堆积顺序引起神经细胞脂质膜变化的假设。这项工作强调了高光谱受激拉曼散射作为一种安全研究活细胞生物物理和生化动力学的方法的潜力。
