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Screening for Developmental Neurotoxicity in Zebrafish Larvae: Assessment of Behavior and Malformations.
Citation:
PADILLA, S. J., A. Desai, S. Frady, B. Padnos, D. L. HUNTER, J. Harrill, W. R. MUNDY, AND R. C. MACPHAIL. Screening for Developmental Neurotoxicity in Zebrafish Larvae: Assessment of Behavior and Malformations. Presented at 7th European Zebrafish Meeting, Edinburgh, SCOTLAND, July 05 - 09, 2011.
Impact/Purpose:
We have developed a paradigm that objectively assesses both overt and neurotoxic endpoints
Description:
The U.S. Environmental Protection Agency is evaluating methods to screen and prioritize large numbers of chemicals for developmental toxicity. As part of this approach, it is important to be able to separate overt toxicity (Le., malformed larvae) from the more specific neurotoxic effect (Le., behavioral changes). We have developed a paradigm that objectively assesses both overt and neurotoxic endpoints. Zebrafish embryos/larvae are dosed from day 0 post fertilization (dpf) to day 4 pf with the compounds to be screened. After one day of depuration (Le. on day 6 pf) behavior is assessed by simultaneously testing individual larvae under both light and dark conditions in a 96-well plate using a video tracking system. By controlling the duration and intensity of light, we are able to assess visual threshold, changes in locomotion during light-dark transitions, and adaptation to both light and dark during approximately 1 hour of testing. The testing format allows evaluation of large numbers of larvae, chemicals across a wide dose range. Following the behavioral test, the larvae are euthanized, fixed, and assessed for dysmorphology with the Zebratox V3 BioApplication (Thermo Scientific) using the Cellornics" Array Scan® high content image analysis platform. This paradigm analyzes each individual larva for various features (e.g., total area, width, spine length, head-totail length, length-to-width ratio, perimeter-to-area ratio). Comparison of these automated measures to detailed visual inspection of the larvae showed excellent congruence. Using this paradigm we have tested a training set of chemicals that are either known or generally considered as positive or negative controls for producing developmental neurotoxicity in mammals. We have found that many developmentally neurotoxic compounds perturb behavior at doses that do not produce malformations, while many developmentally non-neurotoxic compounds either do not perturb behavior at any dose or only perturb behavior at doses which produce malformations. Therefore, results from the training set indicate that a combination of behavioral and morphological evaluation of zebrafish larvae may be able to identify accurately mammalian developmental neurotoxicants. This abstract may not necessarily reflect official Agency policy.