Project description:Domoic acid (DA) is a naturally produced neurotoxin synthesized by the marine diatom genus Pseudo-nitzschia. DA accumulates in filter-feeders such as shellfish, and can produce severe neurotoxicity when contaminated seafood is ingested. DA poisoning is a significant public health concern, and though seafood regulations have effectively minimized the human risk of severe acute poisoning, the effects of exposure at asymptomatic levels are poorly understood. The objective of this study was to determine the effects of exposure to symptomatic and asymptomatic doses of DA on gene expression patterns in the zebrafish brain. We exposed adult zebrafish to either a symptomatic (1.1 ± 0.2 μg /g) or an asymptomatic (0.31 ± 0.03 µg DA/g fish) dose of DA by intracelomic injection and sampled at 24, 48 and 168 h post-injection. Transcriptional profiling was done using Agilent and Affymetrix microarrays. Our analysis revealed distinct, non-overlapping changes in gene expression between the two doses. We found that the majority of transcriptional changes were observed at 24 hours post-injection with both doses. In response to symptomatic dose exposure, we observed 328, 5, and 1 genes were differentially expressed at 24, 48 and 168 hours post-injection, respectively. In contrast, 136, 12, and 28 genes were differentially expressed in the asymptomatic dose at the same time points. Pathway analysis revealed symptomatic DA exposure affected genes associated with pathways including cell adhesion, inflammation, and amyloid proteins. Among the pathways enriched among genes differentially expressed with asymptomatic exposure were circadian rhythms and endocrine signaling. Overall, these results suggest that transcriptional responses are specific to the DA dose and that asymptomatic exposure can cause long-term changes. Many of the differentially expressed genes are important players in neuronal, neuroimmune and neuroendocrine function. Further studies are needed to characterize the potential downstream neurobehavioral impacts of DA exposure.

Project description:Domoic acid (DA) is a neuroexcitatory amino acid that is naturally produced by some marine diatom species of the genus Pseudo-nitzschia. Ingestion of DA-contaminated seafood by humans results in a severe neurotoxic disease known as amnesic shellfish poisoning (ASP). Clinical signs of ASP include seizures and neuronal damage from activation of AMPA and kainate receptors. However, the impacts of DA exposure at levels below those known to induce outward signs of neurobehavioral exicitotoxicity have not been well characterized. To further understand the mechanisms of neurotoxic injury associated with DA exposure, we examined the transcriptome of whole brains from zebrafish (Danio rerio) receiving intracoelomic (IC) DA at both symptomatic and asymptomatic doses. A majority of zebrafish exposed to high-dose DA (1.2 µg DA/g) exhibited clinical signs of neuroexcitotoxicity (EC50 of 0.86 µg DA/g) within 5 to 20 minutes of IC injection. All zebrafish receiving low-dose DA (0.47 µg DA/g) or vehicle only maintained normal behavior. Microarray analysis of symptomatic and asymptomatic exposures collectively yielded 306 differentially expressed genes (1.5-fold, p = 0.05) predominately represented by signal transduction, ion transport, and transcription factor functional categories. Transcriptional profiles were suggestive of neuronal apoptosis following an overwhelming of protective adaptive pathways. Further, potential molecular biomarkers of neuropathic injury, including Nrdg4, were identified and may be relevant to DA exposure levels below that causing neurobehavioral injury. Our results validate zebrafish as a vertebrate model to study mechanisms of DA neurotoxicity and provide a basis for identifying pathways of DA-induced injury as well as biomarkers of asymptomatic and symptomatic DA exposure levels. Keywords: neurotoxic disease To further understand the mechanisms of neurotoxic injury associated with DA exposure, we examined the transcriptome of whole brains from zebrafish (Danio rerio) receiving intracoelomic (IC) DA at both symptomatic and asymptomatic doses. A majority of zebrafish exposed to high-dose DA (1.2 µg DA/g) exhibited clinical signs of neuroexcitotoxicity (EC50 of 0.86 µg DA/g) within 5 to 20 minutes of IC injection. All zebrafish receiving low-dose DA (0.47 µg DA/g) or vehicle only maintained normal behavior. Microarray analysis of symptomatic and asymptomatic exposures collectively yielded 306 differentially expressed genes (1.5-fold, p = 0.05) predominately represented by signal transduction, ion transport, and transcription factor functional categories. All animal studies were carried out under approved IACUC protocols at the University of Washington.

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Danio rerio

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