Automated Tracking of Zebrafish Shoals and the Analysis of Shoaling Behavior

Zebrafish spend the majority of their time in groups, called shoals. Shoaling behavior is complex and dynamic: fish leave and rejoin the shoal, distances between shoal-members oscillate, and the speed and polarization of the shoal change on timescales of seconds to minutes. All these features of shoals can be modified by various pharmacological and environmental—and possibly also genetic—manipulations and a thorough characterization of shoaling behavior can therefore be used as an effective assay for complex aspects of vertebrate social behavior. We present methods for acquiring and analyzing detailed trajectory data from shoals of zebrafish and demonstrate how these methods can be used to distinguish episodes of shoaling under different conditions. These methods could be further developed to create a standardized assay of shoaling behavior that will allow for an in-depth exploration of social behaviors in zebrafish.