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Abstract Harmful algal blooms (HABs) produce neurotoxins that affect human health. Developmental exposure of zebrafish embryos to the HAB toxin domoic acid (DomA) causes myelin defects, loss of reticulospinal neurons, and behavioral deficits. However, it is unclear whether DomA primarily targets myelin sheaths, leading to the loss of reticulospinal neurons, or reticulospinal neurons, causing myelin defects. Here, we show that while exposure to DomA at 2 dpf did not reduce the number of oligodendrocyte precursors prior to myelination, it led to fewer myelinating oligodendrocytes that produced shorter myelin sheaths and aberrantly wrapped neuron cell bodies. DomA-exposed larvae lacked Mauthner neurons prior to the onset of myelination, suggesting that axonal loss is not secondary to myelin defects. The loss of the axonal targets may have led oligodendrocytes to inappropriately myelinate neuronal cell bodies. Consistent with this, GANT61, a GLI1/2 inhibitor that reduces oligodendrocyte number, caused a reduction in aberrantly myelinated neuron cell bodies in DomA-exposed fish. Together, these results suggest that DomA initially alters reticulospinal neurons and the loss of axons causes aberrant myelination of nearby cell bodies. The identification of initial targets and perturbed cellular processes provides a mechanistic understanding of how DomA alters neurodevelopment, leading to structural and behavioral phenotypes. 

Exposure to environmental toxicants during preconception has been shown to affect offspring health and epigenetic mechanisms such as DNA methylation are hypothesized to be involved in adverse outcomes. However, studies addressing the effects of exposure to environmental toxicants during preconception on epigenetic changes in gametes are limited. The objective of this study is to determine the effect of preconceptional exposure to a dioxin-like polychlorinated biphenyl (3,3′,4,4′,5-pentachlorobiphenyl [PCB126]) on DNA methylation and gene expression in testis. Adult zebrafish were exposed to 3 and 10 nM PCB126 for 24 h and testis tissue was sampled at 7 days postexposure for histology, DNA methylation, and gene expression profiling. Reduced representation bisulfite sequencing revealed 37 and 92 differentially methylated regions (DMRs) in response to 3 and 10 nM PCB126 exposures, respectively. Among them, 19 DMRs were found to be common between both PCB126 treatment groups. Gene ontology (GO) analysis of DMRs revealed that enrichment of terms such as RNA processing, iron-sulfur cluster assembly, and gluconeogenesis. Gene expression profiling showed differential expression of 40 and 1621 genes in response to 3 and 10 nM PCB126 exposures, respectively. GO analysis of differentially expressed genes revealed enrichment of terms related to xenobiotic metabolism, oxidative stress, and immune function. There is no overlap in the GO terms or individual genes between DNA methylation and RNA sequencing results, but functionally many of the altered pathways have been shown to cause spermatogenic defects.

 

Abstract Harmful algal blooms produce potent neurotoxins that accumulate in seafood and are hazardous to human health. Developmental exposure to the harmful algal bloom toxin, domoic acid (DomA), has behavioral consequences well into adulthood, but the cellular and molecular mechanisms of DomA developmental neurotoxicity are largely unknown. To assess these, we exposed zebrafish embryos to DomA during the previously identified window of susceptibility and used the well-known startle response circuit as a tool to identify specific neuronal components that are targeted by exposure to DomA. Exposure to DomA reduced startle responsiveness to both auditory/vibrational and electrical stimuli, and even at the highest stimulus intensities tested, led to a dramatic reduction of one type of startle (short latency c-starts). Furthermore, DomA-exposed larvae had altered kinematics for both types of startle responses tested, exhibiting shallower bend angles and slower maximal angular velocities. Using vital dye staining, immunolabelling, and live imaging of transgenic lines, we determined that while the sensory inputs were intact, the reticulospinal neurons required for short latency c-starts were absent in most DomA-exposed larvae. Furthermore, axon tracing revealed that DomA-treated larvae also showed significantly reduced primary motor neuron axon collaterals. Overall, these results show that developmental exposure to DomA targets large reticulospinal neurons and motor neuron axon collaterals, resulting in measurable deficits in startle behavior. They further provide a framework for using the startle response circuit to identify specific neural populations disrupted by toxins or toxicants and to link these disruptions to functional consequences for neural circuit function and behavior. 

Abstract Chemical modifications of proteins, DNA, and RNA moieties play critical roles in regulating gene expression. Emerging evidence suggests the RNA modifications (epitranscriptomics) have substantive roles in basic biological processes. One of the most common modifications in mRNA and noncoding RNAs is N6-methyladenosine (m6A). In a subset of mRNAs, m6A sites are preferentially enriched near stop codons, in 3′ UTRs, and within exons, suggesting an important role in the regulation of mRNA processing and function including alternative splicing and gene expression. Very little is known about the effect of environmental chemical exposure on m6A modifications. As many of the commonly occurring environmental contaminants alter gene expression profiles and have detrimental effects on physiological processes, it is important to understand the effects of exposure on this important layer of gene regulation. Hence, the objective of this study was to characterize the acute effects of developmental exposure to PCB126, an environmentally relevant dioxin-like PCB, on m6A methylation patterns. We exposed zebrafish embryos to PCB126 for 6 h starting from 72 h post fertilization and profiled m6A RNA using methylated RNA immunoprecipitation followed by sequencing (MeRIP-seq). Our analysis revealed 117 and 217 m6A peaks in the DMSO and PCB126 samples (false discovery rate 5%), respectively. The majority of the peaks were preferentially located around the 3′ UTR and stop codons. Statistical analysis revealed 15 m6A marked transcripts to be differentially methylated by PCB126 exposure. These include transcripts that are known to be activated by AHR agonists (eg, ahrra, tiparp, nfe2l2b) as well as others that are important for normal development (vgf, cebpd, sned1). These results suggest that environmental chemicals such as dioxin-like PCBs could affect developmental gene expression patterns by altering m6A levels. Further studies are necessary to understand the functional consequences of exposure-associated alterations in m6A levels. 

Polychlorinated biphenyls (PCBs) are highly persistent and ubiquitously distributed environmental pollutants. Based on their chemical structure, PCBs are classified into non-ortho-substituted and ortho-substituted congeners. Non-ortho-substituted PCBs are structurally similar to dioxin and their toxic effects and mode of action are well-established. In contrast, very little is known about the effects of ortho-substituted PCBs, particularly, during early development. The objective of this study is to investigate the effects of exposure to an environmentally prominent ortho-substituted PCB (2,2’,4,4’,5,5’-hexachlorobiphenyl; PCB153) on zebrafish embryos. We exposed zebrafish embryos to 3 different concentrations of PCB153 starting from 4 to 120 hours post-fertilization (hpf). We quantified gross morphological changes, behavioral phenotypes, gene expression changes, and circadian behavior in the larvae. There were no developmental defects during the exposure period, but starting at 7 dpf, we observed spinal deformity in the 10 μM PCB153 treated group. A total of 633, 2227, and 3378 differentially expressed genes were observed in 0.1 μM (0.036 μg/ml), 1 μM (0.36 μg/ml), and 10 μM (3.6 μg/ml) PCB153-treated embryos, respectively. Of these, 301 genes were common to all treatment groups. KEGG pathway analysis revealed enrichment of genes related to circadian rhythm, FoxO signaling, and insulin resistance pathways. Behavioral analysis revealed that PCB153 exposure significantly alters circadian behavior. Disruption of circadian rhythms has been associated with the development of metabolic and neurological diseases. Thus, understanding the mechanisms of action of environmental chemicals in disrupting metabolism and other physiological processes is essential.