Department of Chemistry, University of California, Berkeley, California 94720-1460, United States.
Department of Molecular & Cell Biology, University of California, Berkeley, California 94720-1460, United States.
ACS Chem Biol. 2024 Oct 18;19(10):2214-2219. doi: 10.1021/acschembio.4c00442. Epub 2024 Oct 2.
Voltage imaging is an important complement to traditional methods for probing cellular physiology, such as electrode-based patch clamp techniques. Unlike the related Ca imaging, voltage imaging provides a direct visualization of bioelectricity changes. We have been exploring the use of sulfonated silicon rhodamine dyes (Berkeley Red Sensor of Transmembrane potential, BeRST) for voltage imaging. In this study, we explore the effect of converting BeRST to diEt BeRST, by replacing the dimethyl aniline of BeRST with a diethyl aniline group. The new dye, diEt BeRST, has a voltage sensitivity of 40% Δ/ per 100 mV, a 33% increase compared to the original BeRST dye, which has a sensitivity of 30% Δ/ per 100 mV. In neurons, the cellular brightness of diEt BeRST is about 20% as bright as that of BeRST, which may be due to the lower solubility of diEt BeRST (300 μM) compared to that of BeRST (800 μM). Despite this lower cellular brightness, diEt BeRST is able to record spontaneous and evoked action potentials from multiple neurons simultaneously and in single trials. Far-red excitation and emission profiles enable diEt BeRST to be used alongside existing fluorescent indicators of cellular physiology, like Ca-sensitive Oregon Green BAPTA. In hippocampal neurons, simultaneous voltage and Ca imaging reveals neuronal spiking patterns and frequencies that cannot be resolved with traditional Ca imaging methods. This study represents a first step toward describing the structural features that define voltage sensitivity and brightness in silicon rhodamine-based BeRST indicators.
电压成像技术是探测细胞生理学的传统方法的重要补充，例如基于电极的膜片钳技术。与相关的 Ca 成像不同，电压成像提供了对生物电变化的直接可视化。我们一直在探索使用磺化硅罗丹明染料（Berkeley Red 跨膜电位传感器，BeRST）进行电压成像。在这项研究中，我们通过用二乙苯胺基团取代 BeRST 的二甲苯胺来探索将 BeRST 转化为二乙 BeRST 的效果。新染料二乙 BeRST 的电压灵敏度为 40%Δ/每 100 mV，比原始 BeRST 染料的 30%Δ/每 100 mV 增加了 33%。在神经元中，二乙 BeRST 的细胞亮度约为 BeRST 的 20%，这可能是由于二乙 BeRST 的溶解度（300 μM）低于 BeRST（800 μM）。尽管细胞亮度较低，但二乙 BeRST 仍能够在单个试验中同时记录来自多个神经元的自发和诱发动作电位。远红色激发和发射谱使二乙 BeRST 能够与细胞生理学的现有荧光指示剂（如 Ca 敏感的 Oregon Green BAPTA）一起使用。在海马神经元中，同时进行电压和 Ca 成像揭示了传统 Ca 成像方法无法分辨的神经元尖峰模式和频率。这项研究代表了描述硅罗丹明基 BeRST 指示剂的电压灵敏度和亮度的结构特征的第一步。
