Enhanced-contrast two-photon optogenetic pH sensing and pH-resolved brain imaging.

Area of Science:

• Neuroscience
• Biophysics
• Optical Imaging

Background:

• Accurate pH monitoring is crucial for understanding brain function and disease.
• Existing pH sensing methods have limitations in resolution and applicability to deep brain tissues.

Purpose of the Study:

• To develop a novel two-photon optogenetic pH sensing technique for high-contrast imaging.
• To demonstrate the utility of this method in cell cultures and brain slices.

Main Methods:

• Utilized a genetically encoded yellow fluorescent protein sensor (SypHer3s).
• Employed a tailored laser driver with two spectrally isolated components for excitation.
• Optimized laser spectrum for maximum contrast in ratiometric two-photon fluorescence readout.

Main Results:

• Achieved high-contrast optogenetic pH sensing and pH-resolved two-photon excited fluorescence (2PEF) imaging.
• Demonstrated a broad dynamic range for pH measurements.
• Successfully applied the technique to cell cultures and brain slices, enabling deep-brain imaging.

Conclusions:

• The developed method provides a powerful tool for precise pH monitoring in biological systems.
• This technique enhances the capability for deep-brain pH sensing and imaging.
• Offers potential for advancing research in neuroscience and related fields.