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Updated: Feb 15, 2026

A Rhodopsin Transport Assay by High-Content Imaging Analysis
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Two-photon voltage imaging with rhodopsin-based sensors.

Christiane Grimm1, Ruth R Sims1, Dimitrii Tanese1

  • 1Institut de la Vision, Sorbonne Université, INSERM, CNRS, 75012 Paris, France.

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|February 13, 2026
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Summary

Researchers engineered rhodopsin-based genetically encoded voltage indicators (GEVIs) for two-photon (2P) imaging. This breakthrough enables high-speed, high-signal-to-noise ratio action potential detection in vivo and in vitro.

Keywords:
action potential detectionfast functional imagingholographic illuminationmicrobial rhodopsinscanless imagingtwo-photon microscopyvoltage indicators

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Area of Science:

  • Neuroscience
  • Optical Imaging
  • Molecular Biology

Background:

  • Two-photon (2P) voltage imaging offers in-depth, high-resolution neuronal activity monitoring.
  • Current 2P voltage imaging primarily uses ASAP-type sensors, while rhodopsin-based sensors are mainly used with one-photon (1P) illumination.

Purpose of the Study:

  • To demonstrate the compatibility of rhodopsin-based genetically encoded voltage indicators (GEVIs) with 2P illumination.
  • To engineer novel rhodopsin-based GEVIs for enhanced 2P voltage imaging.
  • To assess the performance of these sensors in various biological preparations and conditions.

Main Methods:

  • Rational engineering of a novel rhodopsin-based GEVI, named Jarvis.
  • Testing Jarvis and existing FRET-opsin GEVIs (pAce, Voltron2) under 1P and 2P illumination.
  • Comparing 2P scanless and fast 2P scanning illumination methods.
  • Evaluating action potential detection in mouse hippocampal slices, zebrafish larvae, and awake mouse cortex.

Main Results:

  • Demonstrated successful 2P illumination compatibility for rhodopsin-based GEVIs, including Jarvis, pAce, and Voltron2.
  • Identified improved signal-to-noise ratio (SNR) for FRET-opsin GEVIs under low irradiance using scanless 2P illumination.
  • Achieved high-SNR action potential detection at kilohertz rates using Jarvis and pAce in vitro and in vivo.

Conclusions:

  • Rhodopsin-based GEVIs are effective tools for high-performance 2P voltage imaging.
  • Scanless 2P illumination enhances SNR for FRET-opsin GEVIs.
  • This work expands the toolkit for in-depth, high-speed neuronal activity monitoring in complex biological systems.