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Gene Expression Changes in Developing Zebrafish as Potential Markers for Rapid Developmental Neurotoxicity Screening
FAN, C., J. COWDEN, S. SIMMONS, S. J. PADILLA, AND T. V. RAMABHADRAN. Gene Expression Changes in Developing Zebrafish as Potential Markers for Rapid Developmental Neurotoxicity Screening. NEUROTOXICOLOGY AND TERATOLOGY. Elsevier Science Ltd, New York, NY, 32(1):91-98, (2010).
Alternative species based models such as zebrafish are being increasingly used as rapid screening of the toxic potential of environmental compounds including potential developmental neurotoxicants. This is because this and other similar systems provide economical and rapid screening and also reduce the use of rodents in toxicological testing. Advantages of fish also include high fecundity, a transparent embryo and ex vivo development, that latter negating the confounding maternal effects when tested in mammals. A key component of this screen development effort is the identification of biomarkers that permit rapid assessment of key toxic events, developmental neurotoxicity in this case. During the development of the embryo, a temporally-ordered transcriptional program unfolds. Measurement the emergence of transcripts of genes during development and therefore the measurement of these transcripts can provide accurate timers that track the normalcy or deviations of the developmental program. For this effort it is important to identify transcripts that could serve as appropriate biomarkers. In this paper, using an advanced technique of quantitative RT-PCR, we have examined the expression profile of ten gene transcripts, many of which are brain specific, for suitability as developmental biomarkers. As expected these show consistent but different temporal expression profile. Using a known developmental neurotoxicant, ethanol, we show that the expression profiles are altered in measurable fashion upon exposure to sublethal concentrations of the agent. These results establish that measurement of RNA expression profiles in zebrafish using quantitative RT-PCR as an efficient approach for screening developmental neurotoxicants as well as toxicants in general.
Sparse information exists on many chemicals to guide developmental neurotoxicity (DNT) risk assessments. As DNT testing using rodents is laborious and expensive, alternative species such as zebrafish are being adapted for toxicity screening. Assessing the DNT potential of chemicals requires identifying and characterizing zebrafish biomarkers. To this end, the developmental expression profiles of ten nervous system genes were characterized in 1 to 6 days post fertilization zebrafish embryos/larvae using real time quantitative PCR (RT-qPCR). Transcripts of synapsinII a (syn2a) and myelin basic protein (mbp) increased throughout development, while transcripts of gap43, neurogenin1 (ngn1), α1-tubulin, and glial fibrillary acidic protein (gfap) initially increased, but subsequently declined during development. Transcripts for nestin and sonic hedgehog a (shha) decreased during development. To test whether these potential DNT biomarkers are sensitive to DNT compounds, we investigated the gene expression profiles following exposure to ethanol, a known developmental neurotoxicant. Ethanol exposure depressed mbp expression, but increased the expression of gfap, nestin, neurogenin1, nkx2.2a, and α1-tubulin by 3 dpf (compared to untreated control). Levels of gfap and nestin transcripts remained elevated at 6 dpf. Collectively, these proof-ofconcept data indicate that transcript levels of genes are responsive to ethanol exposure and that the gene expression profiles maybe useful as an endpoint to evaluate the DNT potential of chemicals.
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Record Details:Record Type: DOCUMENT (JOURNAL/PEER REVIEWED JOURNAL)
Organization:U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
NATIONAL HEALTH AND ENVIRONMENTAL EFFECTS RESEARCH LABORATORY
CELLULAR AND MOLECULAR TOXICOLOGY BRANCH