Automated analysis of behavior in zebrafish larvae

Zebrafish larvae have become a popular model system to examine genetic and environmental factors that affect behavior. However, studying complex behavior in large numbers of fish larvae can be challenging. The present study describes a novel high-resolution imaging system that is unique in its ability to automatically analyze the location and orientation of zebrafish larvae in multiwell plates. The system revealed behaviors in zebrafish larvae that would have been missed by more manual approaches, including a preference to face a threatening stimulus from a distance and a clockwise orientation in a two-fish assay. The clockwise orientation of the larvae correlates with a clockwise orientation of molecular structures during early development. Larvae with reversed embryonic asymmetries display a counter-clockwise orientation in the two-fish assay, suggesting that embryonic asymmetry and chiral behavior are regulated by the same developmental mechanisms. The developed imaging techniques may be used in large-scale screens to identify genes, pharmaceuticals, and environmental toxicants that influence complex behaviors.