Three-dimensional (3D) temporal and spatial reconstructions of adult zebrafish swim paths rapidly expose overall affective phenotype.

After indicated experimental manipulations (Fig. 3), zebrafish novel tank behavior was manually observed and video-tracked using EthoVision XT7 program. Raw track and behavioral endpoints were processed, formatted, and visualized in a 3D scatter plot using RapidMiner 5.0 software; traditional computer-generated two-dimensional (2D) swim path traces were placed at t = 0.0 s (top left part of each panel) for reference. Representative 3D reconstructions were selected by comparing swim paths of all subjects within a cohort, ranking them from 1 to n based on similarity to each other (low/no to high activity) and choosing the middle for the illustrations. For better visuality and consistency, fish used for spatial 3D imaging were the same as those used for the respective temporal 3D reconstructions. For a more detailed analysis of 3D reconstructions, the average velocity (m/s) of each fish was reflected by changes in color from blue to green, yellow and red, as the velocity increases. Note that any other computer-generated behavioral indices (Table S1 of Supporting Information) may be expressed in color on 3D reconstructions of zebrafish locomotion paths. Overall, these 3D traces reveal striking differences between zebrafish high- and low-anxiety behaviors, thereby enabling a rapid visualization and interpretation of the observed phenotypes.