Aggression is a psychological construct that is commonly used to classify zebrafish behavior. Aggression is a complex trait that can be difficult to accurately measure. The literature on fish behavior describes many different methodologies to examine aggression, which, we believe, have not been compared in a formal manner. In this study we observed 19 individual zebrafish (Danio rerio) and quantified bites, lateral displays, charges, darts, and time near the stimulus in six common assays used to measure aggression. The methodologies included an inclined mirror assay, two flat mirror assays with different acclimation periods, a live conspecific assay, a clay model stimulus assay, and a video recording assay. Our results indicate high repeatability in most aggressive behaviors over time, which confirms the value of each assay to measure personality. However, our results also indicate significant differences between the assays. Specifically, assays using a flat mirror or live conspecific as a stimulus for aggression elicited more attempted bites than an inclined mirror, a clay model stimulus, or a video recording stimulus. Furthermore, the inclined mirror stimulus provoked more darts than any other assay. The results suggest the need for researchers to consider specific research goals when selecting the appropriate stimulus to provoke aggression in zebrafish.
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