Related Experiment Video
Updated: May 31, 2025
High Throughput Danio Rerio Energy Expenditure Assay
Published on: January 27, 2016
Methylene blue at recommended concentrations alters metabolism in early zebrafish development
Niepukolie Nipu1, Lai Wei1, Lauren Hamilton2
1Department of Biology, University of Ottawa, Ottawa, ON, K1N 6N5, Canada.
Abstract:
Methylene blue (MB) is an antifungal agent widely used during critical stages of zebrafish development. Most guidelines recommend 0.00005% or 0.0001% of MB for embryo/larval rearing. The Organisation for Economic Co-operation and Development zebrafish embryo toxicity test guideline omits MB recommendations, leading to inconsistent MB use in zebrafish research. Because MB affects oxidative energy metabolism in vitro and in vivo, we investigate possible metabolic effects of recommended MB concentrations in developing zebrafish (1-5 days post-fertilization (dpf)). MB increases O2 consumption rate at 1 dpf, followed by an overall reduction in oxidative energy metabolism in post-hatch eleutheroembryos (4-5 dpf). Concomitantly, mitochondrial transcripts decrease in 1 and 4 dpf zebrafish. Our findings show that MB, at recommended husbandry concentrations, affects oxidative metabolism and can thus confound experiments. Since the zebrafish embryo/larval model is gaining traction as a high-throughput New Approach Methodology (NAM) for toxicity assessment, researchers should reconsider MB use.
Insights
Methylene blue, used in zebrafish studies, alters their energy metabolism and mitochondrial gene expression. Researchers should reconsider its use due to potential experimental interference.
Area of Science:
- Developmental Biology
- Toxicology
- Metabolomics
Background:
- Methylene blue (MB) is commonly used in zebrafish husbandry.
- Recommended concentrations vary, and guidelines lack standardization.
- MB is known to affect cellular metabolism.
Purpose of the Study:
- To investigate the metabolic effects of recommended Methylene blue concentrations in developing zebrafish.
- To assess the impact of MB on oxidative energy metabolism and mitochondrial gene expression.
Main Methods:
- Zebrafish embryos and larvae (1-5 days post-fertilization) were exposed to recommended MB concentrations.
- Oxygen consumption rates were measured.
- Mitochondrial gene expression was analyzed.
Main Results:
- Methylene blue increased oxygen consumption at 1 day post-fertilization.
- A reduction in oxidative energy metabolism was observed in later stages (4-5 dpf).
- Mitochondrial transcript levels decreased at 1 and 4 dpf.
Conclusions:
- Recommended Methylene blue concentrations impact zebrafish oxidative metabolism and mitochondrial activity.
- MB can confound experimental results in zebrafish models.
- Reconsideration of MB use in zebrafish research, especially in high-throughput toxicity testing, is warranted.
