UC Berkeley

Voltage-sensitive dyes have provided recent promise for affording new methods to interrogate the electrical activity of biological circuits. In particular, small-molecule based approaches have proven useful for designing sensitive molecules that can observe neuronal activity in a noninvasive, highly-parallel manner. However, currently-available voltage-sensitive dyes are marred by low sensitivity, brightness, and/or poor solubility. In order to overcome these challenges, we designed and synthesized voltage-sensitive dyes based on a new family of photoinduced-electron transfer-based voltage sensors, or VoltageFluors. Using the VoltageFluor scaffold, we designed two new families of rhodol- and rhodamine-based voltage-sensitive dyes optimized for two-photon voltage imaging in thick biological tissues, such as brain slices and intact brains. Using these dyes, we characterized the neuronal phenotype associated with the epileptic disorder tuberous sclerosis and monitored spiking activity in awake, behaving mice. We then developed a synergistic computational and experimental approach to guide the rational design of new VoltageFluor dyes. Through this approach, we design and test the most voltage-sensitive VoltageFluor dye to date and also the brightest, highest signal-to-noise fluorescein-based VoltageFluor dye. This work lays the foundation for the diverse array of biological applications of VoltageFluor dyes and develops the guiding principles for future dye design.