Background: Zebrafish are a popular animal model for investigations of the neurobiological mechanisms of learning and memory. However, there remains a need for a standardized set of behavioral methods for use with this species. New method: The goldfish conditioned withdrawal preparation was recently upgraded, and its efficacy for use with zebrafish tested. These upgrades consisted of, 1) automating the presentation of stimulus events, 2) adding a third conditioned stimulus (CS) to the previous two, 3) using a larger conditioning chamber to make it easier to track a subject's position, and 4) employing a special floor insert to prevent the occurrence of defensive shoaling behavior, directed towards the subject's reflection, which can interfere with testing performance. Results: Conditioning was successfully demonstrated with all three CSs. Importantly, however, these demonstrations required the use of more stringent criteria for the scoring of a conditioned withdrawal response than those used previously with goldfish. This was necessary to accommodate their comparatively high rates of locomotor activity. Comparison with existing methods: Several classical conditioning preparations are available for use with zebrafish. However, a potential hindrance to their being standardized is the apparent difficulty of their being accurately reproduced. This is due to such factors as insufficient construction information, the high level of technical expertise required, and/or prohibitively high costs. In contrast, the preparation tested in this report can be easily and inexpensively constructed by virtually any student or professional researcher. Conclusions: The goldfish conditioned withdrawal preparation is a highly effective, efficient, and easily-obtainable tool for the study of classical fear conditioning in zebrafish.
