The use of zebrafish in behavioral neuroscience is rapidly growing. Zebrafish can be assessed for alterations in multiple behavioral endpoints, creating opportunities to use this powerful model to identify chemicals that alter behavioral phenotypes. To evaluate the utility of zebrafish for neurotoxicity research, we designed custom instrumentation to evaluate numerous embryonic and adult zebrafish behaviors. PRAT or Photomotor Response Analysis Tool was used to analyze the embryonic photomotor response (EPR) behavior in embryonic zebrafish (24 hours post fertilization). Shuttleboxes were used to evaluate learning and active avoidance conditioning and a zebrafish Visual Imaging System (zVIS) was used to measure fear responses. Social behavior was observed using Viewpoint tracking software. Startle responses were also analyzed using taps and Noldus Ethiovision XT tracking software. EPR results showed differential movement activities throughout development of larval zebrafish. Highest movement peaks were seen in 35-37 hours post fertilization fish. Using these custom analysis tools, we also evaluated the impact of Vitamin E deficiency and developmental Benzo[a]pyrene exposure on complex adult behaviors. Generational effects of BaP exposure were also tested. Zebrafish were fed defined-diets that either had sufficient or deficient levels of Vitamin E. The vitamin E deficient zebrafish had a ~30% decrease in learning rate relative to the fish with sufficient levels of Vitamin E. Startle response data showed that vitamin E deficient fish do not get desensitized to tap stimulus. Three exposure groups and generations were reared and spawned for the BaP study (0.1% DMSO controls, 1.25 ppm BaP, 2.5 ppm BaP). The zVIS system consists of an array of 8 tanks with only single side views of video projections on LCD monitors. This allows individual fish to visualize either a group of swimming zebrafish or single predator fish. For the socialization assay zebrafish were tracked using Viewpoint tracking software. Distances apart from each other were measured and analyzed in BaP exposed fish. For the predator test, zebrafish were expected to move away from the screen. The proximity of the zebrafish is tracked relative to the LCD screen projections. The preliminary results from BaP exposed zebrafish and 0.1% DMSO controls showed the percent of time spent away from the screen during the predator test or fear response assay was in the high 70% range for all fish. The 2.5 ppm BaP fish had on average the highest percentage (65% vs 50%) time spent away from the screen. Although it is uncertain as of now if there are any generational effects because further analysis is needed. Preliminary shoaling data shows that shoaling speed may be affected by DMSO exposure. The use of DMSO controls may not be optimal for this study. Disassociation is seen in both 1.25 ppm BaP and 2.5 ppm BaP exposure groups in the F2 generation. Collectively, these data demonstrate that custom behavioral systems are able to measure complex behavioral phenotypes and suggests that there are enormous opportunities for translation neurotoxicity research using zebrafish.