CAREER: Voltage-sensitive Dyes based on Mesoscopic Electron Transfer

本文提出的研究是创建一类新的电压敏感染料来测量细胞膜电位和跟踪神经元活动。含有由间隔物隔开的电子供体-受体对，染料基于电子转移的电场依赖性，其与荧光发射竞争并将电场的变化转化为荧光强度的变化。这种类型的染料是独特的，因为它们的荧光强度具有与波长无关的电压响应，因此它们所有发射的光子都可以被收集用于电压检测。这与现有的电压敏感染料形成鲜明对比，在现有的电压敏感染料中，只有一小部分发射的光子是有用的，因此可能导致光子计数大数百倍，并将测量中的信噪比提高超过一个数量级。在理论计算和光学表征的指导下，目标是合成具有亚微秒时间响应、高电压灵敏度和大荧光量子产率的染料，以便它们能够促进具有亚毫秒时间分辨率和亚微米空间分辨率的神经元活动的多位点光学跟踪。此外，基于这种一般的电压传感机制，可以开发出更新颖的光学电压探针，例如双光子吸收电压敏感荧光染料，这将允许三维脑成像和基于荧光蛋白的探针。

The research proposed herein is to create a new class of voltage sensitive dyes to measure cell membrane potentials and to track neuronal activity. Containing an electron donor-acceptor pair separated by a spacer, the dyes are based on the electric-field dependence of electron transfer, which competes with fluorescence emission and converts changes in electric fields to changes in fluorescence intensity. This type of dyes is unique in that their fluorescence intensity has wavelength-independent voltage response, and therefore all their emitted photons may be collected for voltage detection. This is in sharp contrast to existing voltage-sensitive dyes in which only a small portion of emitted photons are useful, and may therefore lead to photon counts hundreds of times larger and improve signal-to-noise ratio in measurements by more than one order of magnitude. Guided by both theoretical calculations and optical characterization, the aim is to synthesize dyes with sub-microsecond time response, high voltage sensitivity, and large fluorescence quantum yield, so that they can facilitate multi-site optical tracking of neuronal activity with sub-millisecond time resolution and sub-micrometer spatial resolution. Furthermore, based on this general voltage-sensing mechanism, more novel optical voltage probes can be developed, such as two-photon-absorbing voltage-sensitive fluorescent dyes that would allow for three-dimensional brain imaging and probes based on fluorescent proteins.