An Ultrasensitive Genetically Encoded Voltage Indicator Uncovers the Electrical Activity of Non‐Excitable Cells
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Abstract
Most animal cell types are classified as non‐excitable because they do not generate action potentials observed in excitable cells, such as neurons and muscle cells. Thus, resolving voltage signals in non‐excitable cells demands sensors with exceptionally high voltage sensitivity. In this study, the ultrabright, ultrasensitive, and calibratable genetically encoded voltage sensor rEstus is developed using structure‐guided engineering. rEstus is most sensitive in the resting voltage range of non‐excitable cells and offers a 3.6‐fold improvement in brightness change for fast voltage spikes over its precursor ASAP3. Using rEstus, it is uncovered that the membrane voltage in several non‐excitable cell lines (A375, HEK293T, MCF7) undergoes spontaneous endogenous alterations on a second to millisecond timescale. Correlation analysis of these optically recorded voltage alterations provides a direct, real‐time readout of electrical cell–cell coupling, showing that visually connected A375 and HEK293T cells are also largely electrically connected, while MCF7 cells are only weakly coupled. The presented work provides enhanced tools and methods for non‐invasive voltage imaging in living cells and demonstrates that spontaneous endogenous membrane voltage alterations are not limited to excitable cells but also occur in a variety of non‐excitable cell types.
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Fig. 2
It would be interesting to know what is the limit of detection? i.e. the smallest voltage fluctuation that could be reliably detected?
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cell types
Another interesting cell type could be insulin-secreting beta cells. A similar ratiometric sensor was developed and tested in those cells in 2017: Schifferer, Martina, Dmytro A. Yushchenko, Frank Stein, Andrey Bolbat, and Carsten Schultz. “A Ratiometric Sensor for Imaging Insulin Secretion in Single β Cells.” Cell Chemical Biology 24, no. 4 (April 20, 2017): 525-531.e4. https://doi.org/10.1016/j.chembiol.2017.03.001.
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Fig. 5. Fluorescence–fluctuation correlation analysis uncovers strong electrical coupling among HEK293T and A375cells and weak coupling among MCF7 cells
This is a very cool result and it would be very interesting to see actual videos of the electrical coupling in action. e.g. would it be possible to see millisecond-scale membrane activity actually spreading via the gap junctions? Similar to what was done in Fig 3c of https://doi.org/10.1038/nmeth.3000?
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