Fast whole-brain imaging of seizures in zebrafish larvae by two-photon light-sheet microscopy

TL;DR

This paper introduces a two-photon light-sheet microscopy technique that enables fast, non-invasive, whole-brain imaging of zebrafish larvae, allowing detailed study of seizure dynamics with minimal laser exposure.

Contribution

The authors developed a 2P LSFM system that significantly increases volumetric imaging speed while reducing laser intensity, facilitating unbiased, real-time brain activity imaging in zebrafish.

Findings

Achieved 5 Hz volumetric imaging rate of zebrafish brain

First observation of caudo-rostral ictal waves during seizures

Demonstrated non-invasive imaging with low laser intensity

Abstract

Light-sheet fluorescence microscopy (LSFM) enables real-time whole-brain functional imaging in zebrafish larvae. Conventional one photon LSFM can however induce undesirable visual stimulation due to the use of visible excitation light. The use of two-photon (2P) excitation, employing near-infrared invisible light, provides unbiased investigation of neuronal circuit dynamics. However, due to the low efficiency of the 2P absorption process, the imaging speed of this technique is typically limited by the signal-to-noise-ratio. Here, we describe a 2P LSFM setup designed for non-invasive imaging that enables quintuplicating state-of-the-art volumetric acquisition rate of the larval zebrafish brain (5 Hz) while keeping low the laser intensity on the specimen. We applied our system to the study of pharmacologically-induced acute seizures, characterizing the spatial-temporal dynamics of pathological activity and describing for the first time the appearance of caudo-rostral ictal waves (CRIWs).