Behavioral tests of hearing in fish are relatively rare and are generally based upon aversive conditioning, with little data available for the positive reinforcement methods common in other vertebrates. Despite its increasing importance as an auditory model, no behavioral hearing measures have been conducted on zebrafish (Danio rerio), with only physiological hearing estimates available. In the current study, a new behavioral testing paradigm is developed to assess sound detection abilities of zebrafish and the effect of training frequency on hearing sensitivity. Zebrafish were trained to respond to either a 400 Hz or a 1000 Hz tone, and behavioral thresholds were then measured to tones from 200 to 1000 Hz. Significant threshold differences existed between the behavioral audiograms, with fish from each set most sensitive to their conditioned frequency. Furthermore, fish acoustically conditioned to 1000 Hz were most sensitive to the upper range of test frequencies (600-1000 Hz). This appears to be the first study utilizing a positive reinforcement behavioral assay for testing hearing in zebrafish and provides further evidence of fine-scale auditory filtering in fish.