Active pixel power control for crosstalk-free all-optical neural interrogation.

Area of Science:

• Neuroscience
• Biophysics
• Optical Engineering

Background:

• All-optical interrogation using two-photon microscopy enables simultaneous neuronal manipulation and monitoring.
• Unintended channelrhodopsin (ChR) excitation during two-photon calcium imaging with genetically encoded calcium indicators (GECIs) causes artifacts.
• Artifacts contaminate neural activity measurements, limiting the precision of all-optical techniques.

Purpose of the Study:

• To develop and validate an active pixel power control (APPC) method.
• To minimize cross-talk between optogenetic manipulation and calcium imaging.
• To enable simultaneous all-optical interrogation with a single laser system.

Main Methods:

• Implemented an active pixel power control (APPC) system.
• Dynamically adjusted imaging laser power per pixel.
• Applied the APPC approach to in vivo larval zebrafish brain preparations.

Main Results:

• APPC significantly suppressed optogenetic artifacts.
• GECI signal quality was preserved during simultaneous imaging and manipulation.
• Demonstrated high-fidelity neural circuit probing in vivo.

Conclusions:

• APPC enhances the accuracy of neural circuit dissection.
• This method advances the precision of all-optical interrogation.
• The technology offers a robust framework for studying neural dynamics and causality across model organisms and integrates with existing two-photon microscopes.