Neurotoxicity of the Cu(OH)2 Nanopesticide through Perturbing Multiple Neurotransmitter Pathways in Developing Zebrafish.

The copper hydroxide [Cu(OH)2] nanopesticide is an emerging agricultural chemical that can negatively impact aquatic organisms. This study evaluated the behavioral changes of zebrafish larvae exposed to the Cu(OH)2 nanopesticide and assessed its potential to induce neurotoxicity. Metabolomic and transcriptomic profiling was also conducted to uncover the molecular mechanisms related to potential neurotoxicity. The Cu(OH)2 nanopesticide at 100 µg/L induced zebrafish hypoactivity, dark avoidance, and response to the light stimulus, suggestive of neurotoxic effects. Altered neurotransmitter-related pathways (serotoninergic, dopaminergic, glutamatergic, GABAergic) and reduction of serotonin (5-HT), dopamine (DA), glutamate (GLU), γ-aminobutyric acid (GABA), and several of their precursors and metabolites were noted following metabolomic and transcriptomic analyses. Differentially expressed genes (DEGs) were associated with the synthesis, transport, receptor binding, and metabolism of 5-HT, DA, GLU, and GABA. Transcripts (or protein levels) related to neurotransmitter receptors for 5-HT, DA, GLU, and GABA and enzymes for the synthesis of GLU and GABA were downregulated. Effects on both the glutamatergic and GABAergic pathways in zebrafish were specific to the nanopesticide and differed from those in fish exposed to copper ions. Taken together, the Cu(OH)2 nanopesticide induced developmental neurotoxicity in zebrafish by inhibiting several neurotransmitter-related pathways. This study presented a model for Cu(OH)2 nanopesticide-induced neurotoxicity in developing zebrafish that can inform ecological risk assessments.

Dopaminergic and serotoninergic neurotoxicity of lanthanide phosphate (TbPO4) in developing zebrafish.

Rare earth elements (REEs) are exploited for global use in manufacturing. Such activities result in their release into the environment and the transformation into more stable phosphate deposition. The objective of this study was to evaluate molecular and behavioral changes of zebrafish exposed to the synthesized terbium phosphate (TbPO4) at concentrations of 10, 20, and 50 mg/L and to determine its potential for neurotoxicity. Metabolomics related to neurotransmitters, and assessment of transcripts and proteins were conducted to uncover the molecular mechanisms underlying TbPO4 with emphasis on neurotransmitter systems. Exposure to 20 mg/L TbPO4 induced larval hyperactivity and perturbed the cholinergic system in zebrafish. Based on metabolomics related to neurotransmitters, dopamine (DA), serotonin (5-HT), and many of their precursors and metabolites were decreased in abundance by TbPO4. In addition, the expression levels of transcripts related to the synthesis, transport, receptor binding, and metabolism of DA and 5-HT were analyzed at the mRNA and protein levels. Transcript and protein levels for tyrosine hydroxylase (TH), the rate-limiting enzyme for DA synthesis, were down-regulated in larval fish. Monoamine oxidase (MAO), an enzyme that catabolizes monoamines DA and 5-HT, was also reduced in mRNA abundance. We hypothesize that DA synthesis and monoamine metabolism are associated with behavioral alterations. This study elucidates putative mechanisms and exposure risks to wildlife and humans by characterizing phosphatic REE-induced neurotoxicity in developing zebrafish.

Metabolomic responses in livers of female and male zebrafish (Danio rerio) following prolonged exposure to environmental levels of zinc oxide nanoparticles.

Zinc oxide nanoparticles (ZnONPs) are widespread pollutants that are present in diverse environmental samples. Here, we determined metabolomic and bioenergetic responses in the liver of female and male zebrafish exposed to a prolonged environmentally relevant concentration of ZnONPs. Metabolome analysis revealed that exposure to 500 µg/L ZnONPs reduced the abundance of metabolites in the tricarboxylic acid (TCA) cycle by modulating the activities of rate-limiting enzymes α-ketoglutarate dehydrogenase and isocitrate dehydrogenase. Moreover, oxidative phosphorylation (OXPHOS) was negatively impacted in the liver based upon decreased activities of mitochondrial Complex I and V in both female and male livers. Our results revealed that bioenergetic responses were not attributed to dissolved Zn2+ and were not sex-specific. However, the metabolic responses in liver following exposure to ZnONPs did show sex-specific responses. Females exposed to ZnONPs compensated for the energetic stress via increasing fatty acids and amino acids metabolism, while males compensated to ZnONPs exposure by adjusting amino acids metabolism, based upon transcript profiles. This study demonstrates that zebrafish adjust the transcription of metabolic enzymes in the liver to compensate for metabolic disruption following ZnONPs exposure. Taken together, this study contributes to a comprehensive understanding of risks related to ZnONPs exposure in relation to metabolic activity in the liver. Environmental implication Zinc oxide nanoparticles (ZnONPs) are widely used in industry and are subsequently released into environments. However, biological responses between female and male following ZnONPs exposure has never been compared. Our data revealed for the first time that female and male zebrafish showed comparable bioenergetic responses, but different metabolic responses to ZnONPs at an environmentally relevant dose. Females compensated for the energetic stress via increasing fatty acids and amino acids metabolism, while males compensated to ZnONPs exposure by adjusting amino acids metabolism in livers. This study reveals that sex may be an important variable to consider in risk assessments of nanoparticles released into environments.

Comparative toxicity of [C8mim]Br and [C8py]Br in early developmental stages of zebrafish (Danio rerio) with focus on oxidative stress, apoptosis, and neurotoxicity.

The increasing production and usage of ionic liquids (ILs) have raised global ecotoxicological concerns regarding their release into the environment. While the effects of side chains on the IL-induced toxicity in various aquatic organisms have been well-recognized, the role of cationic cores in determining their ecotoxicity remains to be elucidated. Herein, the comparative bioavailability and toxicity of two ILs with different cationic cores but the same anion and side chain in zebrafish embryos were determined. 1-octyl-3-methylimidazolium bromide ([C8mim]Br) has higher accumulation in zebrafish, and triggered developmental toxicity by inducing oxidative stress and apoptosis. Meanwhile, 1-octyl-1-methylpyridium bromide ([C8py]Br) enhanced SOD activity and upregulated anti-apoptotic bcl-2 gene expression, contributing to its much lower neurodevelopmental toxicity. Our study demonstrates the vital role of cationic core in determining the developmental toxicity of ILs and highlights the need for further investigations into the toxicity of imidazolium and pyridinium based ILs in aquatic ecosystems.

Developmental toxicity of fenbuconazole in zebrafish: Effects on mitochondrial respiration and locomotor behavior.

Triazole fungicides are used to control the disease of cereal crops but may also cause adverse effects on non-target organisms. There is a lack of toxicity data for some triazoles such as fenbuconazole in aquatic organisms. This research was conducted to evaluate the toxicity of fenbuconazole at environmentally relevant concentrations with attention on the mitochondria, antioxidant system, and locomotor activity in zebrafish. Zebrafish were exposed to one concentration of 5, 50, 200 or 500 ng/L fenbuconazole for 96 h. There was no effect on survival nor percentage of fish hatched, but exposure to 200 and 500 ng/L fenbuconazole resulted in malformation and hypoactivity in zebrafish. Oxygen consumption rates (OCR) of embryos were measured to determine if the fungicide impaired mitochondrial respiration. Exposure to 500 ng/L fenbuconazole reduced basal OCR and oligomycin-induced ATP linked respiration in exposed fish. Fenbuconazole reduced mitochondrial membrane potential and reduced the activities of mitochondrial Complex II and III. Transcript levels of both sdhc and cyc1, each related to Complex II and III, were also altered in expression by fenbuconazole exposure, consistent with mitochondrial dysfunction in embryos. Fenbuconazole activated the antioxidant system, based upon both transcriptional and enzymatic data in zebrafish. Consistent with mitochondrial impairment, molecular docking confirmed a strong binding capacity of the fungicide at the Qi site of Complex III, revealing this complex is susceptible to fenbuconazole. This study reveals potential toxicity pathways related to fenbuconazole exposure in aquatic organisms; such data can improve risk assessments for triazole fungicides.

Comprehensive Interrogation of Metabolic and Bioenergetic Responses of Early-Staged Zebrafish (Danio rerio) to a Commercial Copper Hydroxide Nanopesticide.

The use of copper hydroxide nanopesticide can pose exposure risks to aquatic organisms. In this study, the toxicity of a copper hydroxide nanopesticide, compared to conventional copper sulfate at environmentally relevant doses, was evaluated using metabolomics and bioenergetic assays in embryonic zebrafish. At a copper concentration of 100 µg/L, the nanopesticide caused higher mortality and deformity compared to copper ions alone; despite higher copper accumulation, increased metallothionein and elevated ATP-binding cassette (ABC) transporter activity in zebrafish exposed to copper ions were observed. Both nanopesticide and copper ions reduced the abundance of metabolites of glycolysis and induced energetic stress in zebrafish. The nanopesticide also increased concentrations of several organic acids involved in the tricarboxylic acid (TCA) cycle and elevated the activity of isocitrate dehydrogenase and α-ketoglutarate dehydrogenase, suggesting enhanced TCA cycle activity. Nanopesticide exposure depleted both glutamate and glutamine parallel to the upregulation of the TCA cycle. In addition, zebrafish exposed to the nanopesticide appeared to shift metabolism toward amino acid catabolism and lipid accumulation based upon altered expression profiles of glutaminase, glutamate dehydrogenase, fatty acid synthase, and acetyl-CoA carboxylase. Lastly, the ability of the ions to increase oxidative phosphorylation to alleviate energetic stress was reduced in the case of the nanopesticide. We hypothesize that, unlike copper ions alone, the nanopesticide induces higher toxicity to zebrafish because of increased protein catabolism. This study provides a comprehensive understanding of the risks of copper hydroxide nanopesticide exposure in relation to metabolic activity and mitochondrial function.

A comprehensive review of strobilurin fungicide toxicity in aquatic species: Emphasis on mode of action from the zebrafish model.

Strobilurins are popular fungicides used in agriculture on a global scale. Due to their widespread use as agrochemicals, they can enter aquatic environments at concentrations that can elicit adverse effects in organisms. This review synthesizes the current state of knowledge regarding the toxic effects of strobilurin fungicides on aquatic species, including algal species, Daphnia magna, and fish species, to determine risk to aquatic organisms and ecosystems. Data show that the toxicities of strobilurins vary widely across aquatic species. Strobilurins bind cytochrome bc1 in mitochondrial complex III in fungi, and as such, research in aquatic species has focused on mitochondria-related endpoints following exposures to strobilurins. In fish, studies into the activities of mitochondrial complexes and the expression of genes involved in the electron transfer chain have been conducted, converging on the theme that mitochondrial complexes and their enzymes are impaired by strobilurins. In general, the order of toxicity of strobilurins for fish species are pyraoxystrobin > pyraclostrobin ≈ trifloxystrobin > picoxystrobin > kresoxim-methyl > fluoxastrobin > azoxystrobin. In addition to mitochondrial toxicity, studies also report genotoxicity, immunotoxicity, cardiotoxicity, neurotoxicity, and endocrine disruption, and each of these events can potentially impact whole organism-level processes such as development, reproduction, and behavior. Screening data from the US Environmental Protection Agency ToxCast database supports the hypothesis that these fungicides may act as endocrine disruptors, and high throughput data suggest estrogen receptor alpha and thyroid hormone receptor beta can be activated by some strobilurins. It is recommended that studies investigate the potential for endocrine disruption by strobilurins more thoroughly in aquatic species. Based on molecular, physiological, and developmental outcomes, a proposed adverse outcome pathway is presented with complex III inhibition in the electron transfer chain as a molecular initiating event. This review comprehensively addresses sub-lethal toxicity mechanisms of strobilurin fungicides, important as the detection of strobilurins in aquatic environments suggests exposure risks in wildlife.