A central goal in systems neuroscience is to understand how distributed neural circuitry dynamics drive animal behaviors. In recent years, larval zebrafish (Danio rerio) have become an increasingly attractive model system to investigate the neural correlates of behaviors owing to their small brain size, optical transparency, and rich behavioral repertoire.

Applications of advanced optical imaging methods makes it possible to record whole brain neural activity of a movement-restrained larval zebrafish at high spatial-temporal resolution in the virtual reality paradigm. The behavioral repertoire, however, may be further expanded in freely swimming zebrafish whose behavioral states can be directly inferred and when sensory feedback loops are mostly intact and active.

Whole brain imaging in freely behaving animals has been previously reported in a roundworm Caenorhabditis elegans, by integrating spinning-disk confocal microscopy with a 2D tracking system. This method in freely swimming zebrafish may allow optical interrogation of brain circuits underlying a range of less explored behaviors.

A new research carried out by the Optical Neuroimaging Group led by Dr. WANG Kai at the Institute of Neuroscience of Chinese Academy of Sciences collaborating with the Biophysical and Computational Neuroscience group led by Dr. WEN Quan at University of Science and Technology of China reported a novel volume imaging and 3D tracking technique that monitors whole brain neural activity in freely swimming larval zebrafish. This study was published online in eLife.

In this study, Dr. WANG’s group developed two key techniques, i.e. a high-speed closed-loop system to retain the entire fish head within the field of view of a high numerical aperture (25×, NA = 1.05) objective in 3D, and a new imaging technique called eXtended field-of-view Light Field Microscopy (XLFM) implemented with flashed excitation laser exposure that can image sparse neural activity over the larval zebrafish brain simultaneously and at high speed.

By integrating this imaging technique with a set of high-speed three-dimensional tracking system, they successfully carried out rapid whole brain neural activity recording in freely behaving larval zebrafish. For the first time, they captured whole brain neural activity at high spatiotemporal resolution during the prey capture behavior of zebrafish. The team demonstrated the ability of the system during visually evoked and prey capture behaviors in larval zebrafish.

This study extends traditional experimental paradigms, which can only be performed on head-fixed zebrafish, to freely behaving animals. It opens new window into whole brain dynamics of larval zebrafish in its native and natural environment at unprecedented resolution.

This study was supported by the CAS Strategic Priority Research Program of the Chinese Academy of Sciences, National Science Foundation of China, China Youth Thousand Talents Program, and CAS Hundreds of Talents Program of China.