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EFFECT OF METHYLENE BLUE ON DEVELOPING ZEBRAFISH EMBRYOS Danio rerio
Citation:
Hedge, J., E. Sanders, K. Jarema, D. Hunter, AND S. Padilla. EFFECT OF METHYLENE BLUE ON DEVELOPING ZEBRAFISH EMBRYOS Danio rerio. Aquaculture America 2017, San Antonio, TX, February 19 - 22, 2017.
Impact/Purpose:
Our laboratory routinely conducts zebrafish (Danio rerio) developmental studies on 100s of chemicals, it is therefore imperative that the chemicals we use for animal husbandry are safe. Therefore we did this developmental screen using methylene blue, a common antifungal used in zebrafish research.
Description:
EFFECT OF METHYLENE BLUE ON DEVELOPING ZEBRAFISH EMBRYOS Danio rerioJoan M. Hedge*, Erik Sanders, Kimberly A. Jarema, Deborah Hunter, and Stephanie PadillaIntegrated Systems Toxicology Division, NHEERL, US EPA, Research Triangle Park, NC 27709hedge.joan@epa.govOur laboratory routinely conducts zebrafish (Danio rerio) developmental studies on 100s of chemicals, it istherefore imperative that the chemicals we use for animal husbandry are safe. It was brought to our attentionthat methylene blue, an antifungal agent commonly used in zebrafish embryo rearing, has been reported toaffect swim bladder inflation (angelfish), increased fetal death (rats and humans), and producehyperbilirubinemia, hemolytic anemia, and intestinal atresia (humans). Therefore, we conducted adevelopmental toxicity and neurotoxicity assessment of methylene blue in zebrafish using concentrationscommonly employed by zebrafish researchers.A 6 day developmental/neurodevelopmental assay was conducted using zebrafish embryos/larvae. At 0days post fertilization (dpf), methylene blue exposure began in 96 well plates, which were kept in a 26°Cincubator on a 14:10 (light:dark) light cycle. Embryos were dosed once daily on 0-4 dpf with 0.6, 1.6, 5.0 or10.0 DM methylene blue (MEB); 1 DM chlorpyrifos (CPF) was used as a positive control (n=24 perchemical per dose). At 5 dpf embryos were removed from the chemical, and placed in water, and thentested on 6 dpf.Behavioral testing consisted of assessing the locomotor activity of individual larval zebrafish in a 96-wellplate, under a light:dark test paradigm: 20 minute acclimation in the dark, followed by 40 minutes of light (5lux) and then 40 minutes of dark. Embryos were examined daily on 0-5 dpf and after locomotor testing on6 dpf for death, hatching, swim bladder inflation, and morphological appearance.Although the CPF treated larvae showed the expected decrement in locomotor activity (Figure 1 inset), nodifferences were seen between MEB exposed and control larval locomotor activity (Figure 1) or any of thedevelopmental aspects (death, hatching rate, swim bladder inflation or dysmorphology). These dataindicate that MEB is not developmentally or neurodevelopmentally toxic to larval zebrafish at commonlyused concentrations. This abstract does not necessarily reflect U.S. EPA policy.