Isomerically Pure Tetramethylrhodamine Voltage Reporters

• Published in: Journal of the American Chemical Society 2016-07, Vol.138 (29), p.9085-9088
• Main Authors: Deal, Parker E, Kulkarni, Rishikesh U, Al-Abdullatif, Sarah H, Miller, Evan W
• Format: Article
• Language: English
• Subjects: Electron Transport; Fluorescent Dyes - chemistry; Fluorescent Dyes - metabolism; Isomerism; Membrane Potentials; Photochemical Processes; Rhodamines - chemistry; Rhodamines - metabolism
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/jacs.6b05672

We present the design, synthesis, and application of a new family of fluorescent voltage indicators based on isomerically pure tetramethylrhodamines. These new Rhodamine Voltage Reporters, or RhoVRs, use photoinduced electron transfer (PeT) as a trigger for voltage sensing, display excitation and emission profiles in the green to orange region of the visible spectrum, demonstrate high sensitivity to membrane potential changes (up to 47% ΔF/F per 100 mV), and employ a tertiary amide derived from sarcosine, which aids in membrane localization and simultaneously simplifies the synthetic route to the voltage sensors. The most sensitive of the RhoVR dyes, RhoVR 1, features a methoxy-substituted diethylaniline donor and phenylenevinylene molecular wire at the 5′-position of the rhodamine aryl ring, exhibits the highest voltage sensitivity to date for red-shifted PeT-based voltage sensors, and is compatible with simultaneous imaging alongside green fluorescent protein-based indicators. The discoveries that sarcosine-based tertiary amides in the context of molecular-wire voltage indicators prevent dye internalization and 5′-substituted voltage indicators exhibit improved voltage sensitivity should be broadly applicable to other types of PeT-based voltage-sensitive fluorophores.