Zebrafish Pharmacological Assays

MTZ (MilliporeSigma, M3761-25G) was dissolved in system water at a concentration of 5 mM and 0.5% DMSO. Adult zebrafish were exposed to MTZ for 16 hours at ambient temperature in the dark. APAP (MilliporeSigma, A7085-100G) was dissolved in system water at a concentration of 15 mM. EtOH (MilliporeSigma, EX0276-3) was dissolved in system water at a concentration of 1%. Adult zebrafish were exposed to APAP or EtOH for 24 hours at ambient temperature in the dark. Stock solutions for AG1478 (Selleck Chemicals, S2728), U0126 (Cayman Chemical Company, 70970), LY290002 (Selleck Chemicals, S1105), and Rapamycin (Selleck Chemicals, AY-22989) were made by dissolution in DMSO at a concentration of 10 μM, 20 μM, 10 μM, and 10 μM, respectively. These chemicals were diluted 1:1,000 in system water to make a working solution. Adult zebrafish were immersed in the working solution. Working solution was changed daily. 4-OHT (Hello Bio, HB2508) was dissolved in DMSO at a concentration of 10 mM, vortexed continuously for 15 minutes, and stored at –20°C. Prior to use, an aliquot of 4-OHT was incubated at 65°C for 10 minutes. Triple-transgenic larvae were exposed to 4-OHT between 2 and 3 days after fertilization to induce recombination.