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Identifying Negative Control Chemicals for use in Larval Zebrafish Behavior Assays
Citation:
Knapp, B., B. Hill, D. Hunter, M. Lowery, AND S. Padilla. Identifying Negative Control Chemicals for use in Larval Zebrafish Behavior Assays. Carolinas Society of Environmental Toxicology & Chemistry (SETAC), Charleston, SC, April 19 - 21, 2023. https://doi.org/10.23645/epacomptox.22798736
Impact/Purpose:
Poster presented to the Carolinas Society of Environmental Toxicology & Chemistry (SETAC) Meeting April 2023. Although prior research has identified chemicals that alter larval zebrafish behavior after developmental exposure, there has been little attention given to identifying negative controls, or chemicals that do not alter behavior. Reliable positive and negative control test compounds allow researchers to measure the sensitivity and specificity of their assays. Martin and coworkers (2022) identified a group of 9 candidate negative control chemicals for developmental neurotoxicity testing (e.g., glycerol, L-ascorbic acid, ibuprofen, D-mannitol, selegiline hydrochloride, omeprazole, metformin hydrochloride, sodium benzoate, and saccharin) given that they had been shown in the literature not to induce developmental neurotoxicity in mammals. So far, we have found that saccharin, L-ascorbic acid, D-mannitol, glycerol, omeprazole, and selegiline hydrochloride do not produce any behavioral alterations, indicating that they would be reasonable negative controls for the larval zebrafish locomotor assay.
Description:
Our laboratory assesses the developmental neurotoxicity potential of chemicals using a medium-throughput larval zebrafish screening assay which measures larval zebrafish locomotor behavior in response to visual stimuli. Although prior research has identified chemicals that alter larval zebrafish behavior after developmental exposure, there has been little attention given to identifying negative controls, or chemicals that do not alter behavior. Reliable positive and negative control test compounds allow researchers to measure the sensitivity and specificity of their assays. Martin and coworkers (2022) identified a group of 9 candidate negative control chemicals for developmental neurotoxicity testing (e.g., glycerol, L-ascorbic acid, ibuprofen, D-mannitol, selegiline hydrochloride, omeprazole, metformin hydrochloride, sodium benzoate, and saccharin) given that they had been shown in the literature not to induce developmental neurotoxicity in mammals. To see if these chemicals change zebrafish behavior in our assay, we first performed a dose range-finding developmental toxicity study at ≤ 100 µM, to assess the lethality and teratogenic potential of each chemical. At 100 µM, only ibuprofen was developmentally toxic. Subsequently, after exposing the zebrafish to non-developmentally toxic concentrations of each chemical during development, we assessed their locomotor activity at 6 days post fertilization. This method uses a light/dark transition test consisting of 40 minutes of light (3,500 lux illumination) followed abruptly by 40 minutes of dark (12 lux illumination). So far, we have found that saccharin, L-ascorbic acid, D-mannitol, glycerol, omeprazole, and selegiline hydrochloride do not produce any behavioral alterations, indicating that they would be reasonable negative controls for the larval zebrafish locomotor assay. Standardizing a set of negative controls will allow researchers to compare data from them to the results obtained from unknown chemicals as well as assist with validating new testing methods. This abstract does not reflect the official policy of the US EPA.
URLs/Downloads:
DOI: Identifying Negative Control Chemicals for use in Larval Zebrafish Behavior Assays
BK 2023 CSETAC POSTER FINAL VERSION.PDF (PDF, NA pp, 635.563 KB, about PDF)