Near-infrared voltage-sensitive dyes based on chromene donor

Abstract

Voltage-sensitive dyes (VSDs) are used to image electrical activity in cells and tissues with submillisecond time resolution. Most of these fast sensors are constructed from push–pull chromophores whose fluorescence spectra are modulated by the electric field across the cell membrane. It was found that the substitution of naphthalene with chromene produces a 60 to 80 nm red-shift in absorption and emission spectra while maintaining fluorescence quantum efficiency and voltage sensitivity. One dye was applied to ex vivo murine heart with excitation at 730 nm, by far the longest wavelength reported in voltage imaging. This VSD resolves cardiac action potentials in single trials with 12% ΔF/F per action potential. The well-separated excitation spectra between these long-wavelength VSDs and channelrhodopsin (ChR2) enabled monitoring of action potential propagation in ChR2 hearts without any perturbation of electrical dynamics. Importantly, by employing spatially localized optogenetic manipulation, action potential dynamics can be assessed in an all-optical fashion with no artifact related to optical cross-talk between the reporter and actuator. These new environmentally sensitive chromene-based chromophores are also likely to have applications outside voltage imaging.

Recent advances and current limitations of available technology to optically manipulate and observe cardiac electrophysiology

“…Dyes operating in this window offer even further benefits as compared to red-shifted dyes due to the low light scattering and low absorption of (oxy)haemoglobin and water in the 650–900 nm range [ 55 ]. Recently, Yan and collaborators synthetised and characterised the novel VSD ElectroFluor 730p, which is excitable in the NIR region of the spectrum [ 57 ]. Since the excitation spectrum of this VSD is far removed from the ChR2 activation spectrum, this would allow novel combinations with other indicators operating in the window in between.…”

“…Hence, the excitation light used for the Ca i 2+ indicator X-Rhod-1 does not induce fluctuations in the emission of the VSD ElectroFluor 730p. This is due to the proximity of the 590-nm excitation light to the isosbestic point of the ElectroFluor 730p excitation spectrum [ 57 ], resulting in minimal differences in emission upon changes in transmembrane potential. Therefore, the excitation wavelengths selected for each dye do not affect emission of the other, minimising optical crosstalk.…”