Scanless two-photon voltage imaging

Area of Science:

• Neuroscience
• Biophysics
• Optical Imaging

Background:

• Two-photon voltage imaging offers transformative potential for neuroscience.
• Development of novel imaging approaches is needed for genetically encoded voltage indicators.

Purpose of the Study:

• To demonstrate high-SNR two-photon voltage imaging using parallel excitation.
• To characterize scanless two-photon voltage imaging with different illumination approaches and lasers.
• To enable simultaneous optogenetic control and voltage imaging.

Main Methods:

• Whole-cell patch-clamp electrophysiology.
• Characterization of scanless two-photon voltage imaging with three parallel illumination approaches.
• Utilizing lasers with varying repetition rates and wavelengths.
• Co-expression of JEDI-2P-Kv and ChroME-ST.

Main Results:

• High-SNR two-photon voltage imaging achieved with parallel excitation.
• Successful voltage recordings of high-frequency spike trains and sub-threshold depolarizations.
• Multi-cell recordings of up to fifteen neurons simultaneously using a low repetition-rate laser.
• Simultaneous optogenetic stimulation and voltage imaging of action potentials.
• In vivo imaging of multiple cells up to 250 µm deep in mouse barrel cortex.

Conclusions:

• Scanless two-photon voltage imaging with parallel excitation enhances SNR for genetically encoded voltage indicators.
• This method allows for high-resolution voltage recordings and simultaneous optogenetic manipulation.
• The technique is applicable for in vivo multi-cell recordings in neuroscience research.