In Vivo Two-Photon Voltage Imaging with Sulfonated Rhodamine Dyes

Area of Science:

• Neuroscience
• Biophotonics
• Molecular Imaging

Background:

• Optical methods using fluorescence are crucial for understanding neuronal activity in awake animals.
• Existing fluorescent indicators often lack the required brightness, sensitivity, or compatibility for in vivo applications.
• There is a need for advanced voltage-sensitive dyes for high-resolution neural circuit interrogation.

Purpose of the Study:

• To develop a new class of fluorescent, voltage-sensitive dyes for mapping neuronal membrane potential changes.
• To characterize the performance of these dyes in terms of brightness, sensitivity, and two-photon imaging capabilities.
• To evaluate the utility of these dyes for in vivo neural activity recording in intact brains.

Main Methods:

• Synthesis of sulfonated rhodamine voltage reporters (sRhoVR) based on rhodamine fluorophores and molecular wires.
• Characterization of dye photophysical properties, including voltage sensitivity and two-photon absorption.
• Application of sRhoVR dyes to cultured neurons and in vivo mouse brain preparations for optical imaging of action potentials and local field potentials.

Main Results:

• The novel sRhoVR dyes exhibit high voltage sensitivity and excellent two-photon performance.
• sRhoVR 1 demonstrates a 44% fluorescence increase per 100 mV depolarization and rapid detection of action potentials.
• In vivo imaging in mouse brains shows sRhoVR dyes effectively record neuronal transmembrane potentials, complementing electrophysiology.

Conclusions:

• Sulfonated rhodamine voltage reporters (sRhoVR) represent a promising new class of voltage-sensitive dyes.
• sRhoVR dyes offer superior performance for in vivo optical imaging of neural activity in awake, behaving animals.
• These dyes facilitate high-resolution interrogation of neural computations and brain circuit dynamics.