Voltage Imaging with a NIR-Absorbing Phosphine Oxide Rhodamine Voltage Reporter

24 October 2019, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


Near infrared (NIR) fluorophores may hold the key for non-invasive optical imaging of deep structures in intact organisms with high spatial and temporal resolution. Yet, developing fluorescent dyes that emit and absorb light at wavelengths greater than 700 nm and that respond to biochemical and biophysical events in living systems remains an outstanding challenge. Here, we report the design, synthesis, and application of NIR-absorbing and -emitting, sulfonated, phosphine-oxide (po) rhodamines for voltage imaging in thick tissue from the central nervous system. We find po-rhodamine based voltage reporters, or poRhoVRs, display NIR excitation and emission profiles at greater than 700 nm, show best-in class voltage sensitivity (up to 43% ΔF/F per 100 mV in HEK cells), and can be combined with existing optical sensors, like Ca2+-sensitive fluorescent proteins (GCaMP), and actuators, like light-activated opsins ChannelRhodopsin-2 (ChR2). Simultaneous voltage and Ca2+ imaging reveals differences in activity dynamics in rat hippocampal neurons, and pairing poRhoVR with blue-light based ChR2 affords all-optical electrophysiology. In ex vivo retinas isolated from a mouse model of retinal degeneration, poRhoVR, together with GCaMP-based Ca2+ imaging and traditional multi-electrode array (MEA) recording, can provide a comprehensive physiological activity profile of neuronal activity. Taken together, these experiments establish that poRhoVR will open new horizons in optical interrogation of cellular and neuronal physiology in intact systems.


organic synthesis
voltage imaging
xanthene dyes


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