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Change in Gene Expression in Zebrafish as an Endpoint for Developmental Neurotoxicity Screening
Citation:
FAN, C., J. COWDEN, S. J. PADILLA, AND T. V. RAMABHADRAN. Change in Gene Expression in Zebrafish as an Endpoint for Developmental Neurotoxicity Screening. Presented at TestSmart DNT 2 Developmental Neurotoxicity Meeting, Reston, VA, November 12 - 14, 2008.
Impact/Purpose:
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Description:
Chemicals that adversely affect the developing nervous system may have long-term consequences on human health. Little information exists on a large number of environmental chemicals to guide the risk assessments for developmental neurotoxicity (DNT). As traditional developmental neurotoxicity screening using rodents is costly, time consuming and labor intensive, alternative species such as zebrafish are being adapted for high-throughput toxicity screening. Hence, it is necessary to identify and characterize rapid, yet specific biomarkers for assessing the DNT potential of chemicals. Our approach is to use gene expression of developmental and/or nervous-system genes as biomarkers of appropriate development in zebrafish embryos/larvae. First, we described the time course of developmental expression profiles of neural genes expressed in untreated zebrafish embryos and larvae from 1 to 6 days post fertilization (dpf) determined using real time quantitative PCR (RT-qPCR). Among the genes expressed in nervous system, synapsin II and myelin basic protein (mbp) showed the largest increase in transcript levels throughout the early development phase, while transcripts of nestin and sonic hedgehog (shh) decreased relative to levels at 1dpf. Interestingly, huC, gap43, neurogenin 1 (ngn1), t1α tubulin, nkx2.2a, and gfap showed an early increase in gene expression followed by down-regulation. We next investigated the changes of gene expression following exposure to known DNT chemicals ethanol and valproate. Our results showed that both ethanol and valproate significantly affected gene expression. When embryos were exposed to 1% ethanol, transcripts of gfap, nestin and ngn1 were increased to 1.5, 1.7, 2.7-fold respectively compared to untreated controls, while mbp expression declined to 30% of control at 3 dpf. At 6 dpf, gfap and nestin transcription was still higher that control samples (1.8, 1.7-fold respectively). Following valproate exposure, transcripts of gap43, mbp, synapsin II declined in a dose-dependent manner in 3 dpf zebrafish embryos, and valproate upregulated transcripts of nestin and ngn1. At 6 dpf, most of gene expression examined returned to control levels, with the exception of mbp, gfap and ngn1. Collectively, these data indicate that gene transcription is responsive to developmental neurotoxicant exposure. It is possible to utilize the gene expression profiling as an endpoint to evaluate the development neurotoxicity. (This abstract does not necessarily reflect USEPA policy)