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Copper acutely impairs behavioral function and muscle acetylcholinesterase activity in zebrafish (Danio rerio)
... In this sense, our results indicate that exposure to Cu and Cu+MPs induces hypoactivity and a disorganized swimming pattern in zebrafish larvae. These findings are corroborated by prior studies, which documented impairment of swimming behavior in fish exposed either to Cu (Haverroth et al., 2015;Puglis et al., 2019;Tilton et al., 2011;Vieira et al., 2009;Zhang et al., 2015) or MPs combined with other pollutants (Barboza et al., 2018b;Chen et al., 2017a;Le Bihanic et al., 2020). On the other hand, a subchronic exposure to MPs and Cu, alone or combined, during early life stages had no significant effect on the locomotor activity of 14 dpf zebrafish larvae, under alternating light-dark periods, nor in shoaling behavior. ...
... This was corroborated in the present study by the negative correlation observed between AChE activity and larvae mortality. Supporting our results, significant inhibition of AChE activity by MPs and Cu have also been previously reported in carp (Cyprinus carpio) (Banaee et al., 2019), European seabass (Dicentrarchus labrax) (Barboza et al., 2018a), streaked prochilod (Prochilodus lineatus) (Roda et al., 2020) and in zebrafish (Haverroth et al., 2015;Tilton et al., 2011). Furthermore, a few studies have correlated AChE inhibition and swimming pattern alterations in fish exposed to pesticides (Beauvais et al., 2000;Zhang et al., 2016), heavy metals (Haverroth et al., 2015;Vieira et al., 2009) and, more recently, nanoplastics (Chen et al., 2017a). ...
... Supporting our results, significant inhibition of AChE activity by MPs and Cu have also been previously reported in carp (Cyprinus carpio) (Banaee et al., 2019), European seabass (Dicentrarchus labrax) (Barboza et al., 2018a), streaked prochilod (Prochilodus lineatus) (Roda et al., 2020) and in zebrafish (Haverroth et al., 2015;Tilton et al., 2011). Furthermore, a few studies have correlated AChE inhibition and swimming pattern alterations in fish exposed to pesticides (Beauvais et al., 2000;Zhang et al., 2016), heavy metals (Haverroth et al., 2015;Vieira et al., 2009) and, more recently, nanoplastics (Chen et al., 2017a). How metals interact with and inhibit AChE is not completely understood. ...
Microplastics (MPs, <5 mm) have been frequently detected in aquatic ecosystems, representing both health and ecological concerns. However data about the combined effects of MPs and other contaminants is still limited. This study aimed to evaluate the impact of MPs and the heavy metal copper (Cu) on zebrafish (Danio rerio) larvae development and behavior. Zebrafish embryos were subchronically exposed to MPs (2 mg/L), two sub-lethal concentrations of Cu (60 and 125 µg/L) and binary mixtures of MPs and Cu using the same concentrations, from 2-h post fertilization until 14 days post fertilization. Lethal and sub-lethal responses (mortality, hatching, body length) were evaluated during the embryogenesis period, and locomotor, avoidance, anxiety and shoaling behaviors, and acetylcholinesterase (AChE) activity were measured at 14 dpf. The results showed that survival of larvae was reduced in groups exposed to MPs, Cu and Cu+MPs. Regarding the behavioral patterns, the higher Cu concentration and mixtures decreased significantly the mean speed, the total distance traveled and the absolute turn angle, demonstrating an adverse effect on swimming competence of zebrafish larvae. Exposure to MPs and Cu, alone or combined, also affected avoidance behavior of zebrafish, with larvae not reacting to the aversive stimulus. There was a significant inhibition of AChE activity in larvae exposed to all experimental groups, compared to the control group. Moreover, a higher inhibition of AChE was noticed in larvae exposed to MPs and both Cu+MPs groups, comparatively to the Cu alone groups. Our findings demonstrate the adverse effects of MPs, alone or co-exposed with Cu, on fish early life stages behavior. This study highlights that MPs and heavy metals may have significant impacts on fish population fitness by disrupting locomotor and avoidance behaviors.
... Copper (Cu), an essential micronutrient, but toxic to organisms at high levels (Sorensen, 1991), is commonly found in aquatic ecosystems at concentrations ranging from 0.04 to 560 mg/L (Oliveira et al., 2008;USEPA, 2007). Moreover, a wide variety of adverse effects, including biochemical, physiological and behavioral changes have been reported in fish exposed to Cu (Haverroth et al., 2015;Sonnack et al., 2015). Cu is ubiquitous in rivers (Couto et al., 2018;Milivojevic et al., 2016), estuaries (Couto et al., 2018) and coastal ecosystems (Rossi and Jamet, 2008), where MPs pollution also has a high occurrence (e.g. ...
... In PCA analysis, a positive correlation between AChE and hatching was observed, which suggests that a decrease in embryo motility derived from AChE inhibition may have contributed to the lower hatching observed in the exposed embryos. Previous studies have reported inhibition of AChE activity in fish exposed to Cu (Haverroth et al., 2015;Roda et al., 2020;Tilton et al., 2011). In fish exposed to the MPs alone, no significant differences were perceived in this parameter, although a slight decrease of AChE activity was noticed. ...
... In fact, an opposed tendency was revealed by PCA analysis between LDH activity, involved in the metabolic rate, and the total distance moved by larvae, emphasizing that assessing behavioral alterations may provide a linkage between physiological processes and interaction with the environment (Sandoval-Herrera et al., 2019). Similarly, locomotor activity disturbances have been reported in fish exposed to Cu or to MPs, alone or combined (Acosta et al., 2016;Chen et al., 2017;Haverroth et al., 2015;Qiang and Cheng, 2019). The locomotor response involves several neurochemical systems (Sharma, 2019), which when disrupted by exposure to xenobiotics can be translated in behavior impairment. ...
Data about the toxicological interactions of MPs and heavy metals in biota is limited, particularly in fish early life stages. This study aimed to evaluate the toxicological effects of MPs and copper (Cu), alone or combined, in zebrafish early life stages. Embryos were exposed from 2 until 96-h post-fertilization (hpf) to MPs (2 mg/L), three sub-lethal concentrations of Cu (15, 60 and 125 μg/L) and binary mixtures containing Cu and MPs (Cu15+MPs, Cu60+MPs, Cu125+MPs). Lethal and sub-lethal parameters, histopathological changes, biochemical biomarkers, gene expression and behavior were assessed. Our findings showed that Cu and Cu+MPs decreased embryos survival and hatching rate. Increased ROS levels were observed in larvae exposed to the two lowest Cu and Cu+MPs groups, suggesting an induction of oxidative stress. An increased CAT and GPx activities were observed in Cu and Cu+MPs, implying a response of the antioxidant defense system to overcome the metal and MPs stress. The sod1 expression was downregulated in all Cu groups and in the two highest Cu+MPs exposed groups. AChE was significantly inhibited in Cu and Cu+MPs groups, indicating neurotoxicity. A disruption of avoidance and social behaviors were also noticed in the Cu125 and Cu125+MPs exposed larvae. Evidences of Cu-toxicity modulation by MPs were observed in some endpoints. Overall, the findings of this study highlight that Cu alone or co-exposed with MPs lead to oxidative stress, neurotoxicity and ultimately behavioral alterations in early life stages of zebrafish, while MPs alone do not produce significant effects on zebrafish larvae.
... Tissue samples were digested with 2 mL of 65% HNO 3 at 120°C for 5 h, according to the methodology adapted from Haverroth et al. (2015). The concentration of heavy metals in brains, livers, and muscles was determined by atomic absorption spectroscopy (model 800 AAnalyst AAS-Perkin Elmer®), following the graphite furnace method. ...
... The novel tank apparatus consists of a 2.7-L tank (24 × 8 × 20 cm width × depth × height), virtually divided into three equal horizontal zones (bottom, middle, and top). The following parameters were analyzed: total distance traveled (m), mean speed (m/s), number of crossings among the three zones, turn angle (°), and time (s) and distance (m) in the bottom zone of the tank (Haverroth et al. 2015;Idalencio et al. 2015). ...
... Our results demonstrated a decrease in muscle AChE activity at higher concentrations of Fe and Mn, in accordance with El Haimeur et al. (2017) and Kumar et al. (2017), studies using aquatic organisms. Haverroth et al. (2015) described that the decrease in AChE activity may be related to the binding of the metal to functional groups such as sulfhydryl, imidazole, and carboxyl, compromising the catalytic activity of the enzyme. Sant' Anna et al. (2011) reported increased activity of this enzyme in acute exposure to Fe (15 mg/L), above the one tested here. ...
Iron (Fe) and manganese (Mn) are metals commonly found at high concentrations in underground water. These metals are essential for the good functioning of living organisms, but high concentrations lead to imbalance, potentiating the appearance of pathologies. This study aimed to evaluate the effect of exposure to naturally occurring metals in groundwater, using zebrafish (Danio rerio) as an experimental model. Thus, zebrafish were exposed to Fe (0.8 and 1.3 mg/L), Mn (0.2 and 0.4 mg/L), and groundwater collected from deep tube wells with Fe and Mn (Fe 0.8/Mn 0.2 mg/L and Fe 1.3/Mn 0.4 mg/L) for 30 days. Bioaccumulation of these metals has been demonstrated in the livers and muscles of zebrafish. Acetylcholinesterase activity changed only in zebrafish muscles in all groups. Sulfhydryl levels changed mainly in the group Mn 0.4. SOD/CAT ratio decreased in the groups Fe 0.8 and 1.3, Mn 0.4, and Fe 0.8/Mn 0.4. An increase in the frequency of micronucleus in all groups was shown as a consequence of these changes. Behavioral parameters (time and distance traveled, mean speed, turn angle, latency, and number of crossings between compartments) have also changed, mainly in the groups Fe 1.3, Mn 0.4, and Fe 1.3/Mn 0.4. Therefore, long-term exposure to Fe and Mn, even at not so high concentrations, may cause biochemical, genotoxic, and behavioral changes in zebrafish.
... This assay is particularly relevant to toxicants directly impacting activity of AChE such as organophosphate and carbamate pesticides and has been demonstrated in diverse aquatic species Hagstrom et al., 2018;Hagstrom et al., 2017;Gravato et al., 2021;Haverroth et al., 2015). It has been also reported that diverse toxicants such as metals, polyaromatic hydrocarbons (PAHs), perfluorooctane sulfonate, dioxins and even plastic additives such as bisphenol A and 1-hydroxycyclohexyl phenyl ketone (1-HCHPK) may also alter AChE activity following, e.g., degradation of cholinergic neurons (Walpitagama et al., 2019;Haverroth et al., 2015;Senger et al., 2011;Xu et al., 2013;Sant'Anna et al., 2011;Kang and Fang, 1997;Yuan et al., 2018;Miyagawa et al., 2007). ...
... This assay is particularly relevant to toxicants directly impacting activity of AChE such as organophosphate and carbamate pesticides and has been demonstrated in diverse aquatic species Hagstrom et al., 2018;Hagstrom et al., 2017;Gravato et al., 2021;Haverroth et al., 2015). It has been also reported that diverse toxicants such as metals, polyaromatic hydrocarbons (PAHs), perfluorooctane sulfonate, dioxins and even plastic additives such as bisphenol A and 1-hydroxycyclohexyl phenyl ketone (1-HCHPK) may also alter AChE activity following, e.g., degradation of cholinergic neurons (Walpitagama et al., 2019;Haverroth et al., 2015;Senger et al., 2011;Xu et al., 2013;Sant'Anna et al., 2011;Kang and Fang, 1997;Yuan et al., 2018;Miyagawa et al., 2007). Although AChE was reported to be a sensitive biomarker of neurotoxic alterations, it is only specific to AChE-inhibiting chemicals (i.e. ...
During the last decade, there has been an increase in awareness of how anthropogenic pollution can alter behavioural traits of diverse aquatic organisms. Apart from understanding profound ecological implications, alterations in neuro-behavioural indices have emerged as sensitive and physiologically integrative endpoints in chemical risk assessment. Accordingly, behavioural ecotoxicology and broader eco-neurotoxicology are becoming increasingly popular fields of research that span a plethora of fundamental laboratory experimentations as well as applied field-based studies.
Despite mounting interest in aquatic behavioural ecotoxicology studies, there is, however, a considerable paucity in deciphering the mechanistic foundations underlying behavioural alterations upon exposure to pollutants. The behavioural phenotype is indeed the highest-level integrative neurobiological phenomenon, but at its core lie myriads of intertwined biochemical, cellular, and physiological processes. Therefore, the mechanisms that underlie changes in behavioural phenotypes can stem among others from dysregulation of neurotransmitter pathways, electrical signalling, and cell death of discrete cell populations in the central and peripheral nervous systems. They can, however, also be a result of toxicity to sensory organs and even metabolic dysfunctions. In this critical review, we outline why behavioural phenotyping should be the starting point that leads to actual discovery of fundamental mechanisms underlying actions of neurotoxic and neuromodulating contaminants. We highlight potential applications of the currently existing and emerging neurobiology and neurophysiology analytical strategies that should be embraced and more broadly adopted in behavioural ecotoxicology. Such strategies can provide new mechanistic discoveries instead of only observing the end sum phenotypic effects.
... Consider, for example, effects on defensive behavior: contaminants that change the appropriate levels of antipredator defense and cautious exploratory behavior can lead to a decreased ability to escape or avoid actual or potential threats [4], causing either death or the loss of important resources, such as access to mates or food. Examples of contaminants that have been shown to change cautious exploratory behavior include atrazine [5], methylmercury [6,7], PCB126 [8], dimethyl sulfoxide [9], and copper [10]. Ammonia [11] and IPBC (3-iodo-2-propynyl-N-butyl carbamate) [12] have been shown to impact antipredatory and alarm responses. ...
... It has been observed that for standardized behavioral practices within the laboratory, such experimental conditions (test days and batches of fish) may have relatively few experimental effects on the outcomes of anxiety and locomotor activity [71]. Examples of toxicants that have been shown to affect anxiety-like behavior in the zebrafish light/dark and novel tank tests can be found on Table 3. Copper sulfate 60.0769 nM Acute LDT, NTT [10] Alterations in anxiety-like behavior are relevant not only to antipredator defenses, but also to foraging and resource finding: if an animal is "too cautious" (i.e., increased anxiety-like behavior) due to the effects of a toxicant, it can miss important opportunities to reproduce or to forage outside its home range. Conversely, decreased anxiety-like behavior can lead to "reckless" behavior that ends in being attacked by a predator, as in migrating salmon smolts [4]. ...
The industry is increasingly relying on fish for toxicity assessment. However, current guidelines for toxicity assessment focus on teratogenicity and mortality. From an ecotoxicological point of view, however, these endpoints may not reflect the “full picture” of possible deleterious effects that can nonetheless result in decreased fitness and/or inability to adapt to a changing environment, affecting whole populations. Therefore, assessing sublethal effects add relevant data covering different aspects of toxicity at different levels of analysis. The impacts of toxicants on neurobehavioral function have the potential to affect many different life-history traits, and are easier to assess in the laboratory than in the wild. We propose that carefully-controlled laboratory experiments on different behavioral domains—including anxiety, aggression, and exploration—can increase our understanding of the ecotoxicological impacts of contaminants, since these domains are related to traits such as defense, sociality, and reproduction, directly impacting life-history traits. The effects of selected contaminants on these tests are reviewed, focusing on larval and adult zebrafish, showing that these behavioral domains are highly sensitive to small concentrations of these substances. These strategies suggest a way forward on ecotoxicological research using fish.
... Some impairments may not be found using a certain neurotransmitter at a determined concentration, but some adjustment in exposures could lead to the identification of subtle effects. Adults zebrafish acetylcholinesterase activity (AChE) in the muscles but not in the brain was reduced within 24 h of exposure to 6 µg/L of Cu [31]. This low concentration of copper changes the locomotion and the tendency to avoid intense lighting [31]. ...
... Adults zebrafish acetylcholinesterase activity (AChE) in the muscles but not in the brain was reduced within 24 h of exposure to 6 µg/L of Cu [31]. This low concentration of copper changes the locomotion and the tendency to avoid intense lighting [31]. Exposure to high concentrations of lead during zebrafish development generates abnormalities in the region of the posterior brain and neurotoxicity due to alterations in the development of the neural vasculature and branchiomotor neurons [32]. ...
Objective
Because of pipelines corrosion, lead (Pb) and copper (Cu) ions could be generated at low levels in tap water leading to a chronic exposure with the potential to affect the neurochemical balance in the organism. Malfunction of dopamine (DA) and serotonin (5-HT) systems are significantly implicated on learning, working memory, attention problems among others health impairment in humans. The aim of this research was to determine the toxic effect of Pb and Cu ions on the DA and 5-HT of the model organism zebrafish (Danio rerio).Methods
Forty-eight adult zebrafish were exposed to the nominal concentrations of 12 µg/L of Pb2+, 39 µg/L of Cu2+ and to the co-exposure of both ions. Three subjects of each treatment were extracted every seven days of exposure. The fish brains were analyzed through liquid chromatography to quantify DA and 5-HT while the bodies were digested to measure the bioaccumulation with atomic absorption.ResultsThere were no differences attributed to the different periods of time through the 28 days of exposure at any treatment. Significant differences were found between the means of the treatment populations, meaning that there was an effect on neurotransmitters due to exposure to metals. By the post-hoc test results, it was almost general that there were significant differences between the treatments mean population of the fish exposed to Cu vs control group, and between those exposed to Cu with the population exposed to Pb. Copper exposures increased the levels of DA and 5-HT in the brain of the zebrafish, whereas Pb effects were the total opposite.Conclusions
As it has been observed, the effects on DA and 5-HT are different and mostly affected by Cu. In addition, co-exposure of the metals exacerbates the toxic effects on the zebrafish by lowering the lethal concentrations.
... Examples of contaminants that have been shown to change cautious exploratory behavior include atrazine [5], methylmercury [6,7], PCB126 [8], dimethyl sulfoxide [9], and copper [10]. Ammonia [11] and IPBC (3-iodo-2-propynyl-N-butyl carbamate) [12] have been shown to impact antipredatory and alarm responses. ...
... Examples of toxicants which have been shown to affect anxiety-like behavior in the zebrafish light/dark and novel tank tests can be found on Table 3. Copper sulfate 60.0769 nM Acute LDT, NTT [10] Alterations in anxiety-like behavior are relevant not only to antipredator defenses, but also to foraging and resource finding: if an animal is "too cautious" (i.e., increased anxiety-like behavior) due to the effects of a toxicant, it can miss important opportunities to reproduce or to forage outside its home range. Conversely, decreased anxiety-like behavior can lead to "reckless" behavior that ends in being attacked by a predator, as in migrating salmon smolts [4]. ...
The industry is increasingly relying on fish for toxicity assessment. However, current guidelines for toxicity assessment focus on teratogenicity and mortality. From an ecotoxicological point of view, however, these endpoints may not reflect the “full picture” of possible deleterious effects that can nonetheless result in decreased fitness and/or inability to adapt to a changing environment, affecting whole populations. Therefore, assessing sublethal effects add relevant data covering different aspects of toxicity at different levels of analysis. Impacts of toxicants on neurobehavioral function have the potential to affect many different life-history traits, and are easier to assess in the laboratory than in the wild. We propose that carefully-controlled laboratory experiments on different behavioral domains – including anxiety, aggression, and exploration – can increase our understanding of the ecotoxicological impacts of contaminants, since these domains are related to traits such as defense, sociality, and reproduction, directly impacting life-history traits. The effects of selected contaminants on these tests are reviewed, focusing on larval and adult zebrafish, showing that these behavioral domains are highly sensitive to small concentrations of these substances. These strategies suggest a way forward on ecotoxicological research using fish.
... Exposed larvae covered similar distances as control larvae and no significant difference was observed on traveled distance. The study of Haverroth et al. (2015) showed significant reduction on total distance traveled and restricted mobility on zebrafish (Danio rerio) exposed to 6 µg/L Cu after 24 h. This can suggest that locomotor patterns were weakened (Haverroth et al., 2015). ...
... The study of Haverroth et al. (2015) showed significant reduction on total distance traveled and restricted mobility on zebrafish (Danio rerio) exposed to 6 µg/L Cu after 24 h. This can suggest that locomotor patterns were weakened (Haverroth et al., 2015). Atchinson et al. (1980) described the different ways by which metals can alter the locomotion on fish, either by the influence on the attraction or the evasion to a contaminated area, the impaired sensory perception on hyper or hypoactivity, or reducing the swimming performance. ...
... This suggests that elevated environmental metal concentrations may act as inhibitors of AChE activity, potentially disrupting the transmission of nervous impulses in organisms. The inhibition of AChE by metals occurs because these compounds can bind to the functional groups of proteins, compromising catalytic activity and resulting in the loss of enzymatic function (Haverroth et al. 2015). AChE plays a key role in acetylcholine (ACh) hydrolysis at cholinergic synapses, ensuring the intermittent nature of nervous impulses responsible for neuronal communication (Lopes et al. 2019;Araújo et al. 2016). ...
Major tailings dam failures have occurred recently around the world and resulted in severe environmental impacts, such as metal contamination. Manganese is a metal highly associated with mining activities, largely detected in mining dam collapses. This metal is considered necessary for different organisms, but it can be toxic and cause oxidative stress and genetic damage in fishes. In this study, we investigated the toxic effects of manganese on Astyanax lacustris, by exposing the fish individually to different concentrations of this metal (2.11, 5.00, and 10.43 mg/L) for 96 h. To assess the effects of manganese, we used biochemical biomarkers (glutathione S-transferase, catalase, and acetylcholinesterase enzyme activity) and the manganese bioaccumulation in different tissues (liver and gills). The obtained data showed that only at concentrations of 5.00 mg/L and 10.43 mg/L the activity of glutathione S-transferase differed significantly. Additionally, the acetylcholinesterase activity in the brain tissue was inhibited. The highest level of manganese bioaccumulation was observed in the liver and branchial tissue. Overall, we concluded that high concentrations of manganese may cause physiological changes in Astyanax lacustris.
... Besides developmental delay, treatment with copper sulphate leads to a dysfunctional locomotor behaviour of zebrafish larvae [16,42,43]. We used the light-dark locomotion test to analyse the recovery from the swimming impairment after the inflammatory induction and the following treatment with M-PSs for 24 and 48 h. ...
The Euganean Thermal District, situated in NorthEast Italy, is one of Europe's largest and oldest thermal centres. The topical application of its therapeutic thermal muds is recognised by the Italian Health System as a beneficial treatment for patients suffering from arthro-rheumatic diseases. Polysaccharides produced by the mud microbiota have been recently identified as anti-inflammatory bioactive molecules. In this paper we analysed the efficacy of Microbial-Polysaccharides (M-PS) derived from mature muds obtained at different maturation temperatures, both within and outside the codified traditional mud maturation range. M-PSs were extracted from six mature muds produced by five spas of the Euganean Thermal District and investigated for their chemical properties, monosaccharide composition and in vivo anti-inflammatory potential, using the zebrafish model organism. Additionally, mature muds were characterized for their microbiota composition using Next-Generation Sequencing. The results showed that all M-PSs exhibit similar anti-inflammatory potential, referable to their comparable chemical composition. This consistency was observed despite changes in cyanobacteria populations, suggesting a possible role of the entire microbial community in shaping the properties of these biomolecules. These findings highlight the importance of scientific research in untangling the origins of the therapeutic efficacy of Euganean Thermal muds in the treatment of chronic inflammatory conditions.
... Moreover, negative effects of chronic exposure to Cu on oxidative status/profile have been described to be a key mechanism involved in Cu toxicity (Zebral et al. 2019;Malhotra et al. 2020;Tavares-Dias 2021). Other morphological, biochemical and physiological impairments associated with fish exposure to Cu are linked to negative effects on growth, reproduction and behavior (Haverroth et al. 2015; Adalto Bianchini adaltobianchini@furg.br 1 idea that H. luetkenii exposure to elevated concentrations of waterborne Cu in the field affects lipid metabolism at different levels. The induction of several genes implicated in lipid and steroid transformation suggests that lipid metabolism is one of the most affected process after acute and chronic exposure of H. luetkenii to the Cu mining impacted area (Abril et al. 2022). ...
In this study, we examine markers of oxidative stress in the tetra Hyphessobrycon luetkenii collected from two locations in the copper contaminated João Dias creek (southern Brazil). Also, specimens were translocated from a clean reference section of the creek to a polluted stretch and vice-versa. Fish were held at in submerged cages for 96 h and then sacrificed. Nuclear abnormalities in erythrocytes and total antioxidant capacity, lipid peroxidation and protein carbonylation in gills, brain, liver and muscle displayed similar trends in both groups. Lipid peroxidation increased in all tissues of individuals translocated to the polluted site but only in liver and muscle of those translocated to the reference site. Increased protein carbonylation was also observed in gills of individuals translocated to the reference location. These results suggest similar oxidative stress among fish from the reference and polluted locations and that long-term metals exposure may require adaptations toward oxidative stress responses.
... The same profile was repeated in the April collection, but without significant differences. It is known that the reduced activity of AChE can lead to the accumulation of acetylcholine in central cholinergic synapses and neuromuscular junctions ( Haverroth et al., 2015 ), resulting in locomotor behavioral impairment and death ( Tilton et al., 2011 ). This response profile was observed in fish exposed to metals, especially due to the characteristic of these pollutants being linked to AChE functional groups ( Najimi et al., 1997 ;Lima et al., 2012 ). ...
The Ilha River is one of the main tributaries of the Sinos River, consequently contributing to its ecological impact. The present study aimed to associate the use of different biomarkers in vitro and in vivo to assess the water quality of three points along the Ilha River during 2019. For this purpose, water samples were collected at the source and mouth of the river, and in an intermediate section. In the laboratory, samples were used for in vitro toxicity tests with the V79 cell line, in which we evaluated the protein content and mitochondrial activity through the MTT assay. The results indicated a possible disruption in cellular mitochondrial activity. Acute toxicity tests were also carried out in Danio rerio and Daphnia magna, and a chronic toxicity test was performed in C. elegans. No significant differences were observed in the antioxidant defense enzymes (SOD, CAT and GPx); however, the point of greatest agricultural impact inhibited the activity of muscle AChE in zebrafish. All samples delayed the development of C. elegans, characterizing toxicity. Phosphorus was above the limit established by Brazilian legislation at the three sampling points in two of the three collections, indicating the probable presence of agricultural components in the water. The results alert to the need for the association of different bioindicators and, whenever possible, the use of chronic toxicity tests for the best environmental diagnosis.
... Heavy metals have shown to cause changes in circadian behavior and inverse relationship between exposure time and activity in addition to decreased AChE activity. Decreased activity by heavy metals exposure were reported by Cunha et al. [65] in marine gastropods for cadmium and copper, [66] for mercury and lead in brain of zebra fish, Haverroth et al. [67] in Zebra fish for Copper, Leitemperge et al. [68] in silver catfish Rhamdia quelen for copper, Kim and Kang et al. [22] for cadmium in brain and muscles of juvenile rock fish Sebastes schgeleii, Pan et al. [69], Zhang et al. [70] in fish Danio rerio for cadmium chloride and deltamethrin, Lee and Freeman [71] documented lead toxicity in Zebra fish, Green et al. [72]. ...
... An acute exposure to Cu with concentrations higher than 20 μg·L −1 disturbs the circadian rhythm in the liver and brain, interfering with the correct synchronization between the organism and environment (behavior) (Doria et al., 2018). These behavioral changes may be due to the effects of Cu in specific neurochemical circuits, such as cholinergicmediated signaling (De Lima et al., 2013;Haverroth et al., 2015). The characteristic hypermobility associated with anxiety-like behavior may also be the indicative of pain (Kalueff et al., 2013); this possible painrelated hypermobility reinforces the pain-induced stress hypothesis. ...
Viticulture plays an important role in generating income for small farms globally. Historically, vineyards use large quantities of phytosanitary products, such as Bordeaux mixture [Ca(OH)2 + CuSO4], to control plant diseases. These products result in the accumulation of copper (Cu) in the soil and increases the risk of transfer to water bodies. Thus, it is important to evaluate whether the presence of Cu-bearing particles in water is toxic to aquatic fauna. This study conducted chemical, mineralogical, and particle size evaluations on water samples and sediments collected from a watershed predominantly cultivated with old vineyards. The proportion of Cu-rich nanoparticles (<10 nm) in the sediment was ~27%. We exposed zebrafish to different dilutions of water and sediment samples that collected directly from the study site (downstream river) under laboratory conditions. Then, we evaluated their exploratory behavior and the stress-related endocrine parameter, whole-body cortisol. We also carried out two experiments in which zebrafish were exposed to Cu. First, we determined the median lethal concentration (LC50–96 h) of Cu and then assessed whether Cu exposure results in effects similar to those associated with exposure to the water and sediment samples collected from the study site. The water and sediment samples directly impacted the exploratory behavior of zebrafish, showing clear anxiety-like behavioral phenotype and stress in terms of cortisol increase (during the second rain event). The Cu exposure did not mimic the same behavioral changes triggered by the water and sediment samples, although it had caused similar stress in the fish. Our results highlight that even at low concentrations, the water and sediment samples from vineyard watershed runoff were able to induce behavioral and endocrine changes that may harm the ecological balance of an aquatic environment.
... In the present study, no correlation between AChE and swimming behavior was found. Also, previous studies reported rather poor correlations between swimming behavior alterations and AChE inhibition in zebrafish (Haverroth et al. 2015;Tilton et al. 2011), which might indicate that AChE is not a very sensitive biomarker for the detection of effects from PAHs. ...
In situ burning (ISB) is discussed to be one of the most suitable response strategies to combat oil spills in extreme conditions. After burning, a highly viscous and sticky residue is left and may over time pose a risk of exposing aquatic biota to toxic oil compounds. Scientific information about the impact of burn residues on the environment is scarce. In this context, a comprehensive ISB field experiment with approx. 1000L IFO 180 was conducted in a fjord in Greenland. The present study investigated the toxicity of collected ISB residues to early life stages of zebrafish (Danio rerio) as a model for potentially exposed pelagic organisms. The toxicity of ISB residues on zebrafish embryos was compared with the toxicity of the initial (unweathered) IFO 180 and chemically dispersed IFO 180. Morphological malformations, hatching success, swimming behavior, and biomarkers for exposure (CYP1A activity, AChE inhibition) were evaluated in order to cover the toxic response on different biological organization levels. Across all endpoints, ISB residues did not induce greater toxicity in zebrafish embryos compared with the initial oil. The application of a chemical dispersant increased the acute toxicity most likely due to a higher bioavailability of dissolved and particulate oil components. The results provide insight into the adverse effects of ISB residues on sensitive life stages of fish in comparison with chemical dispersant application.
... These include high synteny with the human genome, optical clarity of embryos, highly developed immune system, rapid life cycle, and ease of genetic modification, among others. 64 In Brazil, the zebrafish has been used to characterize the mechanism of action and toxicity of several chemicals, such as metals, [65][66][67] pesticides, [68][69][70] pharmaceutical compounds, 71-73 and nanomaterial. [74][75][76] Pharmacology & Toxicology is the second largest area researched in Brazil, and accounts for 16.9% of the articles published up to 2019, contrary to what is observed worldwide, where more developmental biology, cellular biology, molecular biology, biochemistry, and genetics studies have been published. ...
The zebrafish (Danio rerio) is an emerging model system in several research areas worldwide, especially in the Global South. In this context, the present study revised the historical use and trends of zebrafish as experimental models in Brazil. The data concerning the bibliometric parameters, research areas, geographic distribution, experimental design, zebrafish strain, and reporter lines, as well as recent advances were revised. In addition, the comparative trends of Brazilian and global research were discussed. Revised data showed the rapid growth of Brazilian scientific production using zebrafish as a model, especially in three main research areas (Neuroscience &and Behavior, Pharmacology and Toxicology, and Environment/Ecology). Studies were conducted in 19 Brazilian states (70.37%), confirming the wide geographic distribution and importance of zebrafish research. Results indicated that research related to toxicological approaches are widespread in Global South countries such as Brazil. Studies were performed mainly using in vivo tests (89.58%) with adult fish (59.75%) and embryos (30.67%). Moreover, significant research gaps and recommendations for future research are presented. The present study shows that the zebrafish is a suitable vertebrate model system in the Global South.
... Cognitive deficits and behavioral functions can be easily modulated by cholinergic signaling and this has been shown to be altered by Cu 2+ . [66][67][68] Thus, DPA may modulate locomotor activity and cognitive deficits in D. melanogaster. ...
D-penicillamine (DPA) is an amino-thiol that has been established as a copper chelating agent for the treatment of Wilson's disease. DPA reacts with metals to form complexes and/or chelates. Here, we investigated the survival rate extension capacity and modulatory role of DPA on Cu2+-induced toxicity in Drosophila melanogaster. Adult Wild type (Harwich strain) flies were exposed to Cu2+ (1 mM) and/or DPA (50 μM) in the diet for 7 days. Additionally, flies were exposed to acute Cu2+ (10 mM) for 24 h, followed by DPA (50 μM) treatment for 4 days. Thereafter, the antioxidant status [total thiol (T-SH) and glutathione (GSH) levels and glutathione S-transferase and catalase activities] as well as hydrogen peroxide (H2O2) level and acetylcholinesterase activity were evaluated. The results showed that DPA treatment prolongs the survival rate of D. melanogaster by protecting against Cu2+-induced lethality. Further, DPA restored Cu2+-induced depletion of T-SH level compared to the control (P < 0.05). DPA also protected against Cu2+ (1 mM)-induced inhibition of catalase activity. In addition, DPA ameliorated Cu2+-induced elevation of acetylcholinesterase activity in the flies. The study may therefore have health implications in neurodegenerative diseases involving oxidative stress such as Alzheimer's disease.
... Furthermore, systemic AChE inhibition may also affect the levels of acetylcholine in the peripheral nervous system (Colović et al., 2013), which, in turn, may impact zebrafish swimming and cortisol levels. For instance, exposure to copper reduces zebrafish locomotor activity, paralleled by lower AChE activity in the muscle tissue (Haverroth et al., 2015). Thus, future studies may examine the activity of AChE in the periphery (e.g. in zebrafish bodies) as well. ...
Background
A potent acetylcholinesterase inhibitor, donepezil is a cognitive enhancer clinically used to treat neurodegenerative diseases. However, its complete pharmacological profile beyond cognition remains unclear. The zebrafish ( Danio rerio) is rapidly becoming a powerful novel model organism in neuroscience and central nervous system drug screening.
Aim
Here, we characterize the effects of 24-h donepezil administration on anxiety-like behavioral and endocrine responses in adult zebrafish.
Methods
We evaluated zebrafish anxiety-like behaviors in the novel tank, the light-dark and the shoaling tests, paralleled by assessing brain acetylcholinesterase activity and whole-body cortisol levels.
Results
Overall, donepezil dose-dependently decreased zebrafish locomotor activity in the novel tank test and reduced time in light in the light-dark test, likely representing hypolocomotion and anxiety-like behaviors. Donepezil predictably decreased brain acetylcholinesterase activity, also increasing whole-body cortisol levels, thus further linking acetylcholinesterase inhibition to anxiety-like behavioral and endocrine responses.
Conclusion
Collectively, these findings suggest negative modulation of zebrafish affective behavior by donepezil, support the key role of cholinergic mechanisms in behavioral regulation in zebrafish, and reinforce the growing utility of zebrafish models for studying complex behavioral processess and their neuroendocrine and neurochemical regulation.
... Furthermore, AChE inhibition may also affect the levels of acetylcholine in the peripheral nervous system (Colović et al., 2013), which, in turn, may impact zebrafish swimming and cortisol levels. For instance, exposure to cooper reduces zebrafish locomotor activity, paralleled by lower AChE activity in the muscle tissue (Haverroth et al., 2015). Thus, future studies may examine the activity of AChE in the periphery (e.g., in zebrafish bodies) as well. ...
Background: An acetylcholinesterase (AChE) inhibitor, donepezil is a cognitive enhancer
... Behavioural damages could find their origin in different biochemical and physiological impairments such as sensory system disruption, endocrine disruption, neurotoxicity or energy disturbances (Amiard-Triquet and Amiard, 2012) at a lower level and population effects at a higher level of organisation. In the laboratory, numerous studies showed that decreased in AChE activity induced behavioural disturbances in different species exposed to several contaminants (Sismeiro-Vivas et al., 2007;Ren et al., 2015;Haverroth et al., 2015;Geraudie et al., 2016). However, in the field, it is necessary to conduct studies able to demonstrate a causal relationship between behavioural responses and effects at higher levels of biological organisation such as population effects (Amiard-Triquet et al., 2015). ...
The main objective of the present study was to explore the potential link between acetylcholinesterase (AChE) activity and burrowing behaviour of the ragworm Hediste diversicolor, which may have consequences at higher levels of biological organisation. Two complementary studies were conducted. AChE activity, at the sub-individual level, and behavioural responses, at the individual level, were evaluated in worms from the Loire estuary (France), whereas density and biomass of H. diversicolor were determined at the population level. A Spearman positive correlation between both biomarkers (AChE and burrowing) suggested that inhibition of AChE activity was linked to behaviour impairments. At the population level, lower AChE and behaviour activities were detected in worms corresponding to lower population density and biomass. These results provide direct empirical field evidence demonstrating the sensitivity of behaviour of H. diversicolor as a biomonitor of estuarine health status assessment.
... It has been observed that standardized behavioral practices within the laboratory such environmental conditions (test days and batches of fish) may have relatively few experimental effects on the outcomes of anxiety and locomotor activity [54]. Examples of toxicants which have been shown to affect anxiety-like behavior in the zebrafish light/dark and novel tank tests can be found on Table 4. [58] Dimethyl sulfoxide 0.05% Acute NTT [59] Copper 0.006 mg/L Acute LDT, NTT [60] Alterations in anxiety-like behavior are relevant not only to antipredator defenses, but also to foraging and resource finding: if an animal is "too cautious" (i.e., increased anxiety-like behavior) due to the effects of a toxicant, it can miss important opportunities to reproduce or to forage outside its home range. Conversely, decreased anxiety-like behavior can lead to "reckless" behavior that ends in being attacked by a predator. ...
The industry is increasingly relying on fish for toxicity assessment. However, current guidelines for toxicity assessment focus on teratogenicity and mortality. From an ecotoxicological point of view, however, these endpoints are not sensitive enough, as they are not able to detect sub-lethal or non-teratogenic effects that can nonetheless result in decreased fitness and/or inability to adapt to a changing environment, affecting whole populations. Impacts of toxicants on neurobehavioral function have the potential to affect many different life-history traits, and are easier to assess in the laboratory than in the wild. We propose that carefully-controlled laboratory experiments on different behavioral domains – including anxiety, aggression, and exploration – can increase our understanding of the ecotoxicological impacts of contaminants, since these domains are related to traits such as defense, sociality, and reproduction, directly impacting life-history traits. We review the effects of selected contaminants on these tests, focusing on larval and adult zebrafish, showing that these behavioral domains are highly sensitive to small concentrations of these substances. These strategies suggest a way forward on ecotoxicological research using fish.
... Among the contaminants of environmental concern, copper deserves special attention as it is frequently used in mining, agricultural, industrial, commercial, and homestead activities. In addition, copper is known to increase the reactive oxygen species, affect extracellular enzymes of the brain such as nucleotidases, decreasing their expression and activities (da Acosta et al., 2016), to inhibit cholinesterase activity and to impair several physiological activities and neurochemical functions (Falfushynska et al., 2013;Haverroth et al., 2015). Due to the neurochemical effects caused by copper, when a forced and continuous exposure is used in ecotoxicity tests, the effects on behavior might be exacerbated (Kimball and Levin, 1985); because the possibility to spatially avoid copper and move towards more favorable area is prevented (Lopes et al., 2004;Gutierrez et al., 2012;Araújo et al., 2014a). ...
Aquatic ecosystems receive run-off and discharges from different sources that lead to the accumulation of contaminants such as copper. Besides producing lethal and sub-lethal effects, copper has shown to be aversive to zebrafish (Danio rerio) by triggering avoidance response. The primary aim of the present study was to evaluate how a copper gradient could affect the spatial distribution of D. rerio by triggering avoidance, preventing recolonization and isolating populations. Secondly, to what extent the food availability in a previously avoided environment could make it a less aversive environment was assessed. A non-forced, multi-compartmented exposure system with a copper gradient (0–300 μg·L⁻¹), through which fish could move, was used for the avoidance and recolonization assays. To test the effect of copper on population isolation, two uncontaminated connected zones were separated by a chemical barrier with a copper concentration of 90 μg·L⁻¹ (a concentration producing an avoidance of 50% - AC50). Zebrafish avoided copper and the 2 h-AC50 was 90.8 μg·L⁻¹. The recolonization was in accordance with avoidance and the relationship ACx/RC100-x (RC: recolonization concentration) was around 2.5. When food was provided in the highest copper concentration, the recolonization pattern was altered, although the distribution of the fish was not statistically different from the scenario without food. The chemical barrier formed by copper (90 μg·L⁻¹) impaired the migratory potential of the fish population by 41.3%; when food was provided in the last compartment, no statistically significant trend of fish moving towards that concentration was observed. Copper might act as an environmental disruptor by triggering spatial avoidance, preventing recolonization and isolating populations in zebrafish. The present study allows simultaneously including three ecological concepts to ecotoxicological studies that have received little attention: habitat selection, recolonization and habitat chemical fragmentation.
... In the current study, we used transgenic zebrafish with confocal imaging and phenotypic anchoring of olfactory behavior to better understand cellular mechanisms of Cu olfactory injury and recovery of specific neuron populations. In this regard, previous studies of toxic effects of metals on the olfactory sensory epithelium offish generally relied on histology, immunostaining, scanning electron microscopy (SEM), and behavioral studies (Dew et al., 2014;Haverroth et al., 2015;Heffern et al., 2018;Lazzari et al., 2017;Williams et al., 2016) to distinguish between the various OSN populations. However, imaging of histological and/or immunostained tissue sections can yield low cellular-level resolution and difficulty in distinguishing different cell types that are interspersed within the olfactory sensory epithelium (Lazzari et al., 2017). ...
Fish rely heavily on their sense of smell to maintain behaviors essential for survival, such as predator detection and avoidance, prey selection, social behavior, imprinting, and homing to natal streams and spawning sites. Due to its direct contact with the outside environment, the peripheral olfactory system of fish is particularly susceptible to dissolved contaminants. In particular, environmental exposures to copper (Cu) can cause a rapid loss of olfactory function. In this study, confocal imaging of double-transgenic zebrafish larvae with differentially labeled ciliated and microvillous olfactory sensory neurons (OSNs) were used to examine cell death and regeneration following Cu exposure. Changes in cell morphologies were observed at varying degrees within both ciliated and microvillous OSNs, including the presence of round dense cell bodies, cell loss and fragmentation, retraction or loss of axons, disorganized cell arrangements, and loss of cells and fluorescence signal intensity, which are all indicators of cell death after Cu exposure. A marked loss of ciliated OSNs relative to microvillous OSNs occurred after exposure to low Cu concentrations for 3 hours, with some regeneration observed after 72 hours. At higher Cu concentrations and 24-h exposures, ciliated and microvillous OSNs were damaged with increased severity of injury with longer Cu exposures. Interestingly, microvillous, but not ciliated OSNs, regenerated rapidly within the 72-h time period of recovery after death from Cu exposure, suggesting that microvillous OSNs may be replaced in lieu of ciliated OSNs. An increase in bromodeoxyuridine labeling was observed 24 h after Cu-induced OSN death, suggesting that increased proliferation of the olfactory stem cells replaced the damaged OSNs. Olfactory behavioral analyses supported our imaging studies and revealed both initial loss and restoration of olfactory function after Cu exposures. In summary, our studies indicate that following zebrafish OSN damage by Cu, regeneration of microvillous OSNs may occur exceeding ciliated OSNs, likely via increased proliferation of the cellular reservoir of neuronal OSC precursors. Transgenic zebrafish are a valuable tool to study metal olfactory injury and recovery and to characterize sensitive olfactory neuron populations in fish exposed to environmental pollutants.
... Previous work has demonstrated oxidative stress responses from Cu NP or CuO NP exposure, but it is often unclear whether the observed toxicity is derived from released ionic species or nanomaterial exposure. 9,18,[37][38][39] Others have suggested a Cu-based NP-specific effect based on the Trojan horse mechanism, in which whole NPs are taken up by the organism and endocytotic transport to acidic compartments (i.e. lysosomes) causes release of toxic ions directly into the cytoplasm. ...
Determining the specific nanomaterial features that elicit adverse biological responses is important to inform risk assessments, develop targeted applications, and rationally design future nanomaterials. Embryonic zebrafish are often employed to study nanomaterial-biological interactions, but few studies address the role of the chorion in nanomaterial exposure and toxicity. Here, we used chorion-intact (CI) or dechorionated (DC) embryonic zebrafish to investigate the influence of the chorion on copper-based nanoparticle toxicity. We found that despite higher dissolution and uptake, CuO NPs were less toxic than Cu NPs regardless of chorion status and did not cause 100% mortality at even the highest exposure concentration. The presence of the chorion inhibited Cu toxicity: DC exposures to Cu NPs had an LC50 of 2.5 ± 0.3 mg/L compared to a CI LC50 of 13.7 ± 0.8 mg/L. This highlights the importance of considering zebrafish chorion status during nanotoxicological investigations, as embryo sensitivity increased by one order of magnitude or more when chorions were removed. Agglomerate size, zeta potential, and dissolved Cu did not sufficiently explain the differences in toxicity between Cu NPs and CuO NPs; however, reactive oxygen species (ROS) generation did. Cu NPs generated ROS in a concentration-dependent manner, while CuO did not and generated less than Cu NPs. We believe that the differences between the toxicities of Cu NPs and CuO NPs are due in part to their ability to generate ROS which could and should be a hazard consideration for risk assessments.
Food standards control copper in water and several foods by setting ADI values. The metal, an essential element influencing biochemical functions, is involved in oxidative stress resistance. However, high metal concentrations can be toxic, especially in nerve functions. Copper at 1 and 2 mM could toxic generations of Drosophila melanogaster flies by assessing the development, learning, memory, locomotion, activities of catalase and acetylcholinesterase, and the neutralizing DPPH-free radicals. Flies cultured in a copper-containing medium lengthened their maturation period and reduced hatching numbers. The learning ability in larvae and the climbing ability of adult flies also decreased between generations. The antioxidative capacity showed the differences between fly groups. Catalase activity gradually decreased with each generation, while acetylcholinesterase did not indicate profound differences between generations. In conclusion, a daily intake of 1–2 mM copper could affect fruit flies’ behaviors, nervous system functions, and the benchmark dose of copper in food.
AD disease (AD) is a multifaceted and intricate neurodegenerative disorder characterized by intracellular neurofibrillary tangle (NFT) formation and the excessive production and deposition of Aβ senile plaques. While transgenic AD models have been found instrumental in unravelling AD pathogenesis, they involve cost and time constraints during the preclinical phase. Zebrafish, owing to their simplicity, well-defined behavioural patterns, and relevance to neurodegenerative research, have emerged as a promising complementary model. Zebrafish possess glutaminergic and cholinergic pathways implicated in learning and memory, actively contributing to our understanding of neural transmission processes. This review sheds light on the molecular mechanisms by which various neurotoxic agents, including okadaic acid (OKA), cigarette smoke extract, metals, and transgenic zebrafish models with genetic similarities to AD patients, induce cognitive impairments and neuronal degeneration in mammalian systems. These insights may facilitate the identification of effective neurotoxic agents for replicating AD pathogenesis in the zebrafish brain. In this comprehensive review, the pivotal role of zebrafish models in advancing our comprehension of AD is emphasized. These models hold immense potential for shaping future research directions and clinical interventions, ultimately contributing to the development of novel AD therapies.
Copper is a significant environmental contaminant resulting from electronic waste and pesticide use. It is also a micronutrient that is essential for life because it is an enzyme cofactor. However, excess exposure to this heavy metal can lead to adverse physiologic effects. Prior studies have demonstrated neurotoxic effects of copper exposure on arthropods, mollusks, amphibians, and mammals, via mechanisms ranging from oxidative stress to changes in neurotransmitter levels. We focused on toxic effects of copper exposure on the nervous system of the Hirudo verbana leech, hypothesizing that copper exposure leads to neurobehavioral changes associated with structural changes to and proteomic changes within nerve ganglia. After exposing leeches to graded concentrations of copper sulfate, we performed neurobehavioral testing to measure their ability to ingest liver and record their movements in a 3-D printed pool. We dissected out nerve ganglia and examined them with light microscopy following hematoxylin and eosin staining. We also extracted, identified, and quantified proteins from the nerve ganglia using high performance liquid chromatography-mass spectrometry. We found that copper exposure was associated with neurobehavioral changes including food avoidance, diminished dark-seeking behavior, and decreased motility. Furthermore, we found histopathologic changes and proteomic alterations within nerve ganglia of copper-exposed leeches. These findings contribute to our understanding of potential mechanisms for nervous system damage with copper exposure. The results are concerning as they indicate that copper accumulation in the environment has tangible neurological consequences for exposed organisms.
Increasing evidence has shown that gut microbes play an important role in the reproductive endocrine system and the development of polycystic ovary syndrome (PCOS). However, whether environmental factors are involved in these gut microbiota alterations has seldom been studied. In this study, we aimed to explore the crucial role of an imbalanced gut microbiota on abnormal ovarian follicle development induced by Cu. A 1:1 matched case-control study with 181 PCOS patients and 181 controls was conducted using a propensity score matching protocol. Information regarding dietary Cu intake was obtained from a face-to-face dietary intake interview. Alterations in the gut microbiota were detected by high-throughput 16 S rDNA sequencing. The results showed that dietary Cu intake was positively correlated with the risk of PCOS, and the risk threshold was approximately 1.992 mg/d. Compared with those with dietary Cu intakes lower than 1.992 mg/d, those who had a higher dietary Cu intake had a 1.813-fold increased risk of PCOS (OR=1.813, 95% CI: 1.150-2.857). PCOS patients had a lower relative abundance of Bacteroides than controls (P = 0.003), and Bacteroides played a partial mediating role between dietary Cu exposure and PCOS (Pindirect effect=0.026, 95% CI: 0.002-0.072). In addition, an animal model of Cu exposure through the diet showed that Cu can induce gut microbiota disorder; increase serum levels of LPS, MDA, and IL-6; and alter host ovarian steroidogenesis to affect ovarian follicle development. Staphylococcus played a partial mediating role between Cu exposure and CYP17A1 (Pg_Staphylococcus=0.083, 95% CI: 0.001-0.228). Overall, this study shows that long-term exposure to high dietary Cu levels can affect the composition of the gut microbiota, cause inflammation and oxidative stress, and then interfere with hormone signaling, ultimately affecting ovarian follicle development.
Background
Several activities can build up soil copper (Cu) concentrations leading to incorporation into the food chain and adversely affect natural and managed ecosystems. This study provides a synthesis of Cu sources, ecological risks, and mitigation strategies of Cu-contaminated soils in Brazil.
Methods
A systematic data search was conducted based on publications from three databases. Data on Cu contamination in Brazilian soils were collected from selected publications; descriptive statistics, regressions, and geo-accumulation indices were obtained. Ecological risks and remediation methods were identified.
Results
Based on total soil Cu concentrations and the geo-accumulation index (Igeo), it was evident that some Cu sources are more problematic for soils. These sources include antifouling paint particles (Igeo = 9.5), tailings from Cu mining (Igeo = 2.8), and Cu-based fungicides applied to vineyards/orchards (Igeo = 1.8). Other Cu sources such as organic waste, solid waste, dust fallout, and metal scraps have generally resulted in moderate to low levels of soil contamination. Soils contaminated by Cu can also impact water since soil erosion carries elements into water bodies. Contaminated environments may lead to increased Cu absorption by organisms, which pose ecological risks. Reference values of total Cu concentrations used for assessing soil quality have been proposed for almost half of Brazilian states. The most prescribed Cu mitigation actions for soil amelioration are phytomanagement with non-accumulator cover crops or native plant species that can accumulate up to 1000 mg Cu kg−1 in roots/stems combined with liming/fertilization and microbial inoculation.
Conclusion
The most Cu contaminated sites in Brazil occur on soils affected by Cu mine tailings and vineyards where Cu-based fungicides have been applied for several decades, which results in environmental, social, economic, and health issues. Strategies have been proposed for the mitigation of Cu contaminated soils, mainly phytomanagement options. However, there is still a strong need for joint efforts by public and private sectors to implement remediation practices on contaminated soils and to prevent Cu contamination of new areas.
Nicotine is a highly addictive substance that can cause teratogenic impacts in the embryo through redox-dependent pathways. As antioxidants, naturally occurring chemicals can protect cells from redox imbalance. The purpose of this study was to evaluate the effectiveness of 24-epibrassinolide (24-EPI), a natural brassinosteroid with well-known antioxidant properties, in protecting zebrafish embryos against nicotine's teratogenic effects. For 96 h, embryos (2 h post-fertilization - hpf) were exposed to 100 μM nicotine, co-exposed with 24-EPI (0.01, 0.1, and 1 μM), and 24-EPI alone (1 μM). Lethal and sublethal developmental characteristics were evaluated during exposure. Biochemical tests were performed at the conclusion of the exposure, and distinct behavioural paradigms were analysed 24 h later. Nicotine exposure resulted in a higher proportion of larvae with deformities, which were decreased following co-exposure to 24-EPI. Nicotine exposure also caused an increase in oxidative stress as observed by the increased activity of superoxide dismutase and catalase accompanied by an increase in the malondialdehyde levels. Besides, metabolic changes were noticed as observed by the increased lactate dehydrogenase activity that were hypothesised to be associated to nicotine-induced hypoxia which may be responsible for the increased oxidative damage. In addition, locomotor deficits were observed as well as a decrease in the acetylcholinesterase activity denoting nicotine-induced cognitive dysfunction. However, co-exposure to 24-EPI alleviated behavioural deficits and improved nicotine-induced emotional states. Overall, and although further studies are required to clarify these effects, 24-EPI showed promising ameliorative properties against the teratogenic effects induced by nicotine.
Currently, antibiotics and heavy metal contaminants have posed a great threat for ecological security and human health. Herein, the lanthanide functionalized ZIF (named ZIF-90-PABA-Eu) is constructed by coordinating with Eu3+ via p-aminobenzoic acid intermediate. Due to the excellent fluorescence properties, the novel fluorescent probe can selectively monitor flumequine based on "turn on" mode. Furthermore, the obtained new material (named ZIF-90-PABA-Eu-Flu) can be used as "turn off" sensor for selective detection of both radioactive and nonradioactive heavy metal ions (UO22+, Ni2+ and Cu2+) which are the main component of nuclear industrial wastewater. ZIF-90-PABA-Eu-Flu shows ultra-short fluorescence response time (3 s) and ultra-low limit of detection (9.0 × 10-3, 1.3 × 10-2 and 6.1 × 10-4 ppm) for three metal ions, which may be attributed to its good affinity with UO22+, Ni2+ and Cu2+. Moreover, principal component analysis (PCA) is applied to distinguish the three metal ions. Additionally, the possible sensing mechanism is investigated by the UV-vis spectra, luminescence lifetimes and theoretical calculation analysis. Based on these results, ZIF-90-PABA-Eu possesses promising potential in practical application and provides insight for the design of novel probes to continuously monitor flumequine, radioactive and nonradioactive heavy metal ions.
Aquatic organisms are concomitantly exposed to multiple noxious chemicals that can be discharged into water bodies. We aimed to investigate the single and simultaneous sub-acute exposure to copper and methomyl on juvenile Oreochromis niloticus. Compared to the controls, the outcomes revealed that brain of methomyl-exposed fish displayed significant declines in the activities of SOD, CAT, and GST in addition to higher MDA and lower GSH levels. Methomyl induced notable declines in levels of GABA and acetylcholine esterase in brain and muscle of exposed fish. Noteworthy downregulated gene expression levels of TNF-α, HSP-70 together with upregulated c-fos were evident in brain of fish expose to either of tested compounds. Marked apoptotic changes were observed in fish brain exposed to copper and methomyl indicated by augmented immune expression of caspase-3. Conclusively, the results indicated the possible interaction between both compounds with subsequent toxic effects that differ from their single exposure.
Therapeutic thermal mud produced by spas of the Euganean Thermal District (Italy) is used as a treatment for arthro-rheumatic diseases. Its production involves the growth of a specific microbiota embedded in a polysaccharidic matrix. Polysaccharides (Microbial-PolySaccharides, M-PS) released in the mud by the resident microorganisms were extracted and analyzed. The monosaccharidic composition analysis showed the presence of galacturonic acid, mannose, xylose, ribose and glucose and a high percentage of sulfated groups in the polymers. To assess their involvement in the therapeutic efficacy of the mud, the M-PS were tested using the model organism zebrafish (Danio rerio). The anti-inflammatory and antioxidant activities were evaluated after confirming the lack of toxic effects during development. Inflammatory state was induced chemically with copper sulfate, or through tail fin amputation procedure and UVB exposure. Recovery from inflammatory condition after exposure to M-PS was always observed with specific morphometric analyses, and further supported by qPCR. Genes linked with the inflammatory and oxidative stress response were investigated confirming the M-PS treatment's efficacy.
Copper leached from recreational vessel antifouling paints can pose a threat to aquatic organisms. To date, leaching rates have mainly been studied in seawater and brackish water. The aim of this study was to investigate the copper input from antifouling paints to freshwater using field and laboratory studies. Therefore, a large sailing area in Berlin was sampled. According to a risk assessment, the use of copper containing antifouling paints in Berlins is predicted to have no significant impact on aquatic organisms. An exception was found in sport boat marinas where, in dependence on the boat number, the sediment copper concentrations were up to 2 times higher than the German environmental quality standard for sediments. The most important result revealed by the laboratory batch tests is that the copper leaching rate in freshwater is much lower compared to seawater and brackish water. The dissolution of copper from antifouling paints into freshwater is overestimated up to now. Nevertheless, the leached copper from antifouling paints is the second largest anthropogenic copper source in the urban area of Berlin and the third largest anthropogenic copper source in German freshwaters.
The Bordeaux mixture, fungicides composed of copper sulfate, is commonly used in organic production. This use has been questioned owing to the impact of copper accumulation in the environment, which can affect living organisms negatively. Here, was analyzed the presence of copper levels in soils and in orange and grape juices that received Bordeaux mixture fungicide treatments. Furthermore, we evaluated copper toxicity in Caenorhabditis elegans nematodes by assessing some of their biochemical and behavioral parameters after acute and chronic copper exposure. The metal exposure in vivo was performed at concentrations of 0.05, 0.1, 0.3, and 0.7 mg/L, compared with a control group that received ultrapure water. The results showed that copper was present in the sampled soils at higher levels than in the control area soil. Copper was detected in an average value of 0.25 mg/L in orange juice and 0.69 mg/L in grape juice. The acute and chronic exposure conditions showed behavioral changes and an activation of the acetylcholinesterase enzyme. However, chronic exposure to low copper concentrations led to inhibition of this enzyme. Thus, the copper concentrations detected in the fruit juices acted as modulators of the cholinergic system and modified the behavioral parameters in C. elegans.
L’utilisation excessive de pesticides engendre des pollutions et des dégradations importantes de l’environnement y compris sur les écosystèmes aquatiques. L’activité viticole ne fait pas exception à cette règle, et c’est pour cette raison que ce travail de thèse s’est intéressé aux réponses adaptatives ou délétères aux premiers stades de vie de poissons exposés à trois produits phytosanitaires utilisés en viticulture ainsi qu’à leur mélange et à des échantillons environnementaux. La thèse s’intéresse aux stades précoces de développement de la truite arc-en-ciel, mais également à la lignée cellulaire de foie de truite RTL-W1. Les embryons de truite ont été exposés au cuivre, au glyphosate et au chlorpyrifos seuls ou en mélange. Une autre partie de ce travail a consisté à étudier les effets toxiques des échantillons environnementaux d’eau et de sédiment provenant d’un cours d’eau, La Livenne, à proximité de parcelles viticoles. Toutes les expériences ont été faites en conditions contrôlées de laboratoire. Les réponses induites par ces expositions ont été mesurées à différents niveaux d’organisation biologique : au niveau moléculaire, phénotypique et comportemental pour l’étude in vivo (larves de la truite arc-en-ciel) ; et l’analyse des effets cytotoxiques de l’induction des espèces réactives de l’oxygène (ROS) et des dommages à l’ADN pour l’étude in vitro (lignée RTL-W1). Les résultats de ce travail ont montré que l’exposition aux pesticides individuels, ou en mélange, chez les larves de truites arc-en-ciel n’a pas produit d’effets létaux aux concentrations testés. En revanche, ces substances ont provoqués différents effets sub-létaux, selon le composé et les concentrations étudiés, dont des effets tératogènes, des perturbations du comportement natatoire, des effets génotoxiques et l’expression différentielle de gènes cibles. Le cuivre s’est avéré le plus toxique provoquant un échec d’éclosion important. Des effets cytotoxiques et une production d’espèces réactives de l’oxygène (ERO) ont été observés sur cellules de truite exposées à des extraits d’eau de rivière. Par ailleurs, des effets comportementaux ont été observés sur les larves de truites exposées pendant 48 h à des sédiments et de l’eau de la Livenne. En conclusion, ces travaux de thèse ont mis en évidence des effets sub-létaux sur les cellules et les stades précoces de développement de truite arc-en-ciel exposés à des concentrations environnementales de pesticides utilisés en viticulture.
Drug discovery and development for Alzheimer’s disease (AD) are complex and challenging due to the higher failure rate in the drug development process. The overproduction and deposition of Aβ senile plaque and intracellular neurofibrillary tangle (NFT) formation are well-recognized diagnostic hallmarks of AD. Numerous transgenic models of Alzheimer’s disease have restrictions on cost-effectiveness and time in the preclinical setup. Zebrafish has emerged as an excellent complementary model for neurodegenerative research due to simpler organisms with robust, clearly visible behavior forms. Glutaminergic and cholinergic pathways responsible for learning and memory are present in zebrafish and actively participate in the transmission process. Therefore, it is imperative to study neurotoxic agents’ mechanisms that induce dysfunction of memory, learning, and neurons in the zebrafish. This review illustrates the in-depth molecular mechanism of several neurotoxic agents such as okadaic acid, cigarette smoke extract, and metals to produce cognitive deficits or neurodegeneration similar to mammals. These updates would determine an ideal and effective neurotoxic agent for producing AD pathophysiology in the zebrafish brain for preclinical screening.
Graphical Abstract
The frequency of wildfire is expected to increase with time as a function of climate change. Recent studies in our laboratory have demonstrated that pyrogenic polycyclic aromatic hydrocarbons (PAHs) can cause greater‐than‐additive effects in Hyalella azteca in the presence of low concentrations of copper (Cu). We hypothesized that freshwater animals inhabiting Cu‐contaminated sites, such as those in the vicinity of Cu mines, may be vulnerable to non‐additive toxicity from contaminants released by wildfires. To investigate the interaction between Cu and water conditioned by burnt wood ash (fire extract; FE) we exposed H. azteca for 14 days to binary mixtures of 225 mg/kg Cu‐enriched artificial sediment (225 mg Cu/kg) and a FE dilution series (12.5%, 25%, 50%, 75%, and 100%). All binary mixtures of Cu‐enriched sediment and FE resulted in complete mortality with the exception of Cu‐enriched sediment + 12.5% FE. The combination of Cu‐enriched sediment with 12.5% FE had a more‐than‐additive effect on survival and tissue Cu concentration but there was no reduction in growth or acetylcholinesterase activity compared to the 225 mg/kg Cu‐contaminated sediment or FE control, respectively. Acetylcholinesterase activity was significantly reduced in amphipods exposed to FE but the presence of Cu did not exacerbate this effect. Results from this study suggest that Cu‐contaminated water bodies that receive runoff from wildfires are at risk of enhanced toxicity. This article is protected by copyright. All rights reserved.
We have evaluated the interactive toxicity of Cu(II) and Cd(II) in water with different hardness levels using adult zebrafish (Danio rerio). Zebrafish were exposed to Cd(II) (0.2-22 μM) or Cu(II) (0.1-8 μM) in single or binary exposures in very soft, moderately hard or very hard water. The whole body burdens of Cd(II) and Cu(II) reflect the net effect of biouptake and elimination, mortality was the indicator of toxicity, and whole body major ion content was measured to assess ion regulatory functions. Cu(II) was found to be more toxic than Cd(II) for zebrafish, and Cu(II) and Cd(II) exhibited a significant synergistic effect. The toxicity of metal ions increased upon decreasing the ionic strength of the exposure medium, probably due to elevated competition between metal ions with other cations in hard water and increased activity of Ca2+ pathways in soft water treatments. Whole body metal accumulation and the accumulation rate of both Cu and Cd increased as the metal ion concentration in the exposure medium increased. Nevertheless, neither parameter explained the observed synergistic effect on mortality. Finally, we observed a significant loss of whole body Na+ in fish which died during the metal exposure compared to surviving fish, irrespective of exposure conditions. Such an effect was not observed for other major cations (K+, Ca2+ and Mg2+). This observation suggests that, under the applied exposure conditions, survival was correlated to the capacity of the organism to maintain Na+ homeostasis.
Graphene oxide quantum dots (GOQDs) have broad applications such as bioimaging and drug delivery, among others, even expanding into the aquatic environment. However, reports on the adverse effects of GOQDs on fish development are limited. In this study, we exposed zebrafish embryos to GOQDs for 7 days after fertilization and found that GOQDs exposure at low concentrations (12.5, 25, 50 or 100 μg/L) decreased the total distance and the mean velocity of larvae movement. Additionally, the GOQDs significantly reduced the enzyme activity related to energy supply and locomotor capacity, including Ca2+‐ATPase in the 12.5, 25, 50 and 100 μg/L GOQDs groups and Na+/K+‐ATPase in the 25 and 50 μg/L GOQDs groups. Moreover, GOQD exposure altered the mRNA expression of genes involved in energy supply and calcium transport. The levels of the atp2a2b, atp2a1, and cacna1sb genes were significantly downregulated in the 25, 50 and 100 μg/L GOQDs groups, and ryr3 expression was significantly reduced in the 25 and 50 μg/L GOQDs groups. The expression level of cacna1c was significantly upregulated in the 50 and 100 μg/L GOQDs groups. In summary, GOQD exposure induced a decrease in locomotor capacity in zebrafish, which may be due to the reduction of Ca2+‐ATPase and Na+/K+‐ATPase activity levels, and dysregulated expression of the genes involved in energy metabolism and calcium transport. Our study provides novel insight into the effects of GOQDs on the embryonic development of fish, which will be useful for the development of environment‐friendly GOQDs that reduce the potential hazard to aquatic species. Zebrafish embryos exposure to GOQDs (12.5, 25, 50, and 100 μg/L) for 7 days impaired the locomotor capacity in zebrafish larvae, which may be due to the dysregulated expression of genes involved in energy metabolism and calcium transport, and the reduction of Ca2+‐ATPase and Na+/K+‐ATPase activity levels.
Environmental contaminants such as metal ions can have detrimental effects on aquatic organisms at the molecular, organismal and population levels. In the present work, we examined the interactive effect of Cu(II) and conspecific alarm substance on zebrafish behavioural responses utilizing the novel tank diving assay. To this end, 3 novel tank diving tests (on day 0, 3 and 10 of the experimental phase) were conducted on zebrafish in 4 experimental groups: (1) control: no Cu(II) and no alarm substance, (2) Cu(II) only: exposed to 0.78 µM Cu(II) (25% of the 240 hr LC50) in the home tank for 10 days, (3) alarm substance only: exposed to alarm substance for 6 min concomitant with behavioural testing, and (4) Cu(II) + alarm substance: exposed to 0.78 µM Cu(II) in the home tank for 10 days and treated with alarm substance for 6 min during the behavioural testing. Results showed robust habituation response of zebrafish. Exposure to Cu(II) did not affect the behavioural phenotypes of zebrafish in the novel tank diving test or habituation responses. Alarm substance treatment evoked strong anxiety-like behaviour. Finally, zebrafish in the Cu(II) + alarm substance group lost their sensitivity to alarm substance in repeated novel tank assays throughout the concomitant Cu(II) exposure; this observation is tentatively ascribed to Cu(II)-induced olfactory impairment.
Triclosan is widely used in consumer products as an antimicrobial agent. Epidemiological studies have reported the association of triclosan with adverse birth outcomes. The toxic effects of triclosan on the developing stages of zebrafish are reported, however, its role as behavioral modifier is limited. In the present study, adult zebrafish were exposed to triclosan (0.3 and 0.6 mg/L) for 48 h and the exploratory behavior was analyzed using ZebraTrack. Triclosan exposed group showed significantly reduced locomotion concomitant with increased freezing duration. They also showed erratic movements suggesting that triclosan induced anxiety-like behavior in adult zebrafish. Next, we tested the hypothesis that the anxiety-like behavior is linked to altered acetylcholinesterase activity. We found that the triclosan exposure decreased acetylcholinesterase activity in the brain and skeletal muscle but acetylcholinesterase (ache) gene was significantly down-regulated only in the skeletal muscle of the adult zebrafish exposed to triclosan. In addition, we also observed a down-regulation of myelin basic protein (mbp) gene in the skeletal muscle of adult zebrafish treated with triclosan. Thus, our data indicates that even short exposure of triclosan is potent enough to induce behavioral anomalies in adult zebrafish that appear to involve acetylcholinesterase and other structural proteins especially in the skeletal muscle.
Copper is an essential element in many biological processes, but may exert toxic effects at levels surplus to metabolic requirements. Herein we assess the effect of copper on zebrafish behaviour using two assays, namely the novel tank diving test and a T-maze test with food reward. Novel tank diving tests were conducted on days 0, 4, and 10 of a 10 day Cu exposure (at concentrations of 0.77 μM (25% of the 240 h LC50) and 1.52 μM (50% of the 240 h LC50) to assess the alterations of behavioural responses in repeating novel tank diving assays and the effect of Cu on these patterns. Results demonstrate habituation to novelty, which is an indicator of spatial memory. Copper exposure had no effect on the latency of entry into the upper zones of the tank, nor on the total time spent therein, but did cause a greater number of freezing bouts in comparison to the control group. Additionally, Cu exposure had no effect on the habituation responses of zebrafish. Using the T-maze assay, we tested the effect of prior exposure to Cu for 10 days on subsequent behavioural trainings. The T-maze protocol was based on associative learning, where a visual stimulus (colour) was linked with a natural stimulus (food). Results of the control group showed that zebrafish are able to perform associative learning tasks. Moreover, Cu was found to negatively affect the associative learning capabilities. Specifically, while zebrafish in the control group achieved a significant number of correct choices (leading to food reward) throughout the T-maze training, such a trend was not observed for Cu exposed fish. Thus at the exposure concentrations and exposure times considered herein, Cu has no determinative impact on instinctual behavioural responses of zebrafish in repeated novel tank diving assays but does limit the associative learning capabilities.
The effects of dietary zinc, copper or mercury (9.8, 9.5 and 0.66 mg/kg wet weight of forage, respectively) on different aspects of fish feeding behavior in common carp were evaluated. The latency to leave the starting chamber after its front wall was raised (t1) and latency to feed (latent time of feeding, t2) as well as the amount of food consumed or ration (the quantity of eaten chironomid larvae for 3 min of observation) were registered. By the end of the experiment under the impact of Hg, the t1 value increased by 20% and t2 value increased 5.6 times. The amount of food consumed decreased by 38%. The latency to leave the starting chamber under the impact of metal significantly increased during 20–30 days, followed by decline. The dynamics of the feeding reaction rate under the impact of metal differed: the maximum changes in t2 value are noted under the impact of Cu: they were increased in the 2nd and 5th decade (3.7 and 6.1 times, respectively). The amount of food consumed in the presence of Zn either increased or remained at the control group level, but under the action of Cu and Hg was in most cases lower (by 37% at the most).
Two experiments were performed to determine whether oral administration of copper oxide capsules controlled helminthic infections in Lacaune sheep without acute collateral effects on animal health. In experiment 1, 48 multiparous lactating sheep (60.1 ± 8.5 kg) were stratified according to initial number of eggs (Haemonchus contortus) per gram of feces (EPG) and were assigned randomly to 1 of two treatments (24 sheep/treatment): no oral administration (control) or oral administration of two copper capsules (treated; approximately 58 mg copper/kg body weight). Blood and fecal samples were collected on days 0, 15 and 45. Animals treated with copper capsules showed lower of EPG, eosinophils, acetylcholinesterase (AChE) in whole blood, and lower butyrylcholinesterase (BChE) activity in serum. Treated sheep had higher erythrocyte numbers, hemoglobin concentrations, hematocrit, and lymphocyte numbers. In experiment 2, 12 male lambs negative for helminths and coccidia were assigned randomly to one of two treatments (six lambs/treatment): control or treated (one copper capsule; approximately 58 mg copper/kg body weight); the experiment was designed to determine whether the results of experiment 1 were due to treatment or parasitism. Blood samples were collected on days 0, 5, 10 and 15 and fecal samples were collected on days 0, 7 and 15. Treated animals showed greater concentrations of lymphocytes; however, treatment had no effect on other hemogram variables, AChE and BChE activities, or levels of alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transferase, creatinine, urea, albumin, total protein, and reactive oxygen species. These data suggest that copper capsules in dairy sheep efficiently controlled H. contortus infections. Treatment was not harmful to lambs during the first 15 days, i.e. there were no signs of acute toxicity.
Lines of evidence used in ecological risk assessment (ERA) are essentially three: chemistry, biology and ecotoxicology. Until now, the fundamental assumption made when measuring ecological risks is that organisms are forcedly exposed to stressors. However, when organisms can avoid disturbed habitats by escaping to less stressful areas, the assumption that exposure is mandatory to pose risk may not match field disturbance scenarios. A non-forced exposure approach using a linear free-choice multi-compartmented system has been proposed previously as a complementary tool to assess the effects on organisms' spatial avoidance/preference responses. Yet, the linearity of the latter system limits avoidance measurements to one spatial dimension. A novel, heterogeneous multi-habitat assay system (HeMHAS) consisting of 18 connected circular compartments (3 compartments on a vertical axis in each one of 6 zones on a longitudinal axis; a 2D system) is put forward here to be used in heterogeneous-habitat selection studies, as it makes it possible to assess the ability of organisms to detect contamination and other stressors and select more favorable habitats. In the present study, the avoidance to copper by zebrafish (Danio rerio) was tested after exposing organisms to a copper gradient in the HeMHAS and compared with that in the linear system. Avoidance occurred for all copper concentrations: 43% in the lowest (21 μg·L-1) to 72% in the highest (221 μg·L-1). Results obtained within the HeMHAS (AC50: 60 μg·L-1) were statistically (p = 0.72) similar to avoidance of copper by D. rerio in the linear non-forced system (AC50: 89 μg·L-1). In summary, the simulation of a copper gradient in the HeMHAS (2D system) allowed to assess the potential repellency of copper to zebrafish and to corroborate the ability of organisms to detect and avoid potentially toxic concentrations.
As the most abundant natural polymer, cellulose presents a unique advantage for large‐scale applications. To fully unlock its potential, the introduction of desired functional groups onto the cellulose backbone is required, which can be realized by either chemical bonding or physical surface interactions. This review gives an overview of the chemistry behind the state‐of‐the‐art functionalization methods (e.g., oxidation, esterification, grafting) for cellulose in its various forms, from nanocrystals to bacterial cellulose. The existing and foreseeable applications of the obtained products are presented in detail, spanning from water purification and antibacterial action, to sensing, energy harvesting, and catalysis. A special emphasis is put on the interactions of functionalized cellulose with heavy metals, focusing on copper as a prime example. For the latter, its toxicity can either have a harmful influence on aquatic life, or it can be conveniently employed for microbial disinfection. The reader is further introduced to recent sensing technologies based on functionalized cellulose, which are becoming crucial for the near future especially with the emergence of the internet of things. By revealing the potential of water filters and conductive clothing for mass implementation, the near future of cellulose‐based technologies is also discussed. An overview on the synthesis and applications of functionalized cellulose reveals the role of functionalization methods (e.g., oxidation; esterification; grafting; conductive nanoparticle, carbon, or polymer composite formation) on its physical properties. Existing and foreseeable applications are presented in detail, spanning from heavy metal water purification and antibacterial action, to sensing, energy harvesting, and catalysis.
Dysfunction of copper homeostasis can lead to a host of disorders, which might be toxic sometimes. 4-Methoxy-5-hydroxy-canthin-6-one (CAN) is one of the major constituents from Picrasma quassioides and responsible for its therapeutic effects. In this work, we evaluated the toxic effect of CAN (7.5μM) on zebrafish embryos. CAN treatment decreased survival, delayed hatching time and induced malformations (loss of pigmentation, pericardial edema, as well as hematologic and neurologic abnormalities). Besides, exogenous copper supplementation rescued the pigmentation and cardiovascular defects in CAN-treated embryos. Further spectroscopic studies revealed a copper-chelating activity of CAN. Then its regulation on the expressions of copper homeostasis related genes also be analyzed. In addition, CAN lowered the total activity of SOD, elevated the ROS production and altered the oxidative related genes transcriptions, which led to oxidative stress. In conclusion, we demonstrated that CAN (7.5μM) might exert its toxic effects in zebrafish embryos by causing copper dyshomeostasis and oxidative stress. It will give insight into the risk assessment and prevention of CAN-mediated toxicity.
The rate of autoxidation of epinephrine and the sensitivity of this autoxidation to inhibition by superoxide dismutase were both augmented, as the pH was raised from 7.8 → 10.2. O2⁻, generated by the xanthine oxidase reaction, caused the oxidation of epinephrine to adrenochrome and the yield of adrenochrome produced per O2⁻ introduced, increased with increasing pH in the range 7.8 → 10.2 and also increased with increasing concentration of epinephrine. These results, in conjunction with complexities in the kinetics of adrenochrome accumulation, lead to the proposal that the autoxidation of epinephrine proceeds by at least two distinct pathways, only one of which is a free radical chain reaction involving O2⁻ and hence inhibitable by superoxide dismutase. This chain reaction accounted for a progressively greater fraction of the total oxidation as the pH was raised. The ability of superoxide dismutase to inhibit the autoxidation of epinephrine at pH 10.2 has been used as the basis of a convenient and sensitive assay for this enzyme.
Iron, copper, zinc, and eight other minerals are classified as essential trace elements because they present in minute in vivo quantities and are essential for life. Because either excess or insufficient levels of trace elements can be detrimental to life (causing human diseases such as iron-deficiency anemia, hemochromatosis, Menkes syndrome and Wilson's disease), the endogenous levels of trace minerals must be tightly regulated. Many studies have demonstrated the existence of systems that maintain trace element homeostasis, and these systems are highly conserved in multiple species ranging from yeast to mice. As a model for studying trace mineral metabolism, the zebrafish is indispensable to researchers. Several large-scale mutagenesis screens have been performed in zebrafish, and these screens led to the identification of a series of metal transporters and the generation of several mutagenesis lines, providing an in-depth functional analysis at the system level. Moreover, because of their developmental advantages, zebrafish have also been used in mineral metabolism-related chemical screens and toxicology studies. Here, we systematically review the major findings of trace element homeostasis studies using the zebrafish model, with a focus on iron, zinc, copper, selenium, manganese, and iodine. We also provide a homology analysis of trace mineral transporters in fish, mice and humans. Finally, we discuss the evidence that zebrafish is an ideal experimental tool for uncovering novel mechanisms of trace mineral metabolism and for improving approaches to treat mineral imbalance-related diseases.
The utility of early effect endpoints as biomarkers of ecotoxicity of natural sediments in water-sediment static system was investigated. The particular goal was to evaluate the ecotoxicity of the sediment samples from La Choza stream, located in upper basin of the Reconquista river, the second most polluted river of Argentina. Native juveniles Hyalella curvispina were used as test organisms evaluating survival, growth, oxidative stress parameters (SOD; CAT, TBARS) and the electron transport system (ETS) activity as early toxic effect. This study used methodologies and techniques that allow the assessment of sediment pollution with a native species as test organism and provided data to discuss the viability of sublethal endpoints as tools for freshwater sediment assessment. In spring and in summer two ten-day series of whole-sediment assays were conducted simultaneously: (a) standard assays and (b) biomarkers assays. A control sediment was ran simultaneously in which no--effect on survival was measured. In summer there was a significant increase in length and biomass in both exposed and control groups. In spring an inhibitory effect on growth and an increase in oxidative damage with a concomitant rise in antioxidant defenses, was observed in animals exposed to La Choza sediment. ETS measurement indicated a significant depression of metabolic activity of amphipods exposed to contaminated sediments. The measured biomarkers represent the first record for juvenile H. curvispina exposed to polluted natural sediments under standardized laboratory conditions. The used bioanalytical tools demonstrated higher sensitivity and a more accurate assessment of the effects than those obtained by the standard tests of survival and growth. We propose their adoption in biomonitoring of freshwater sediment toxicity.
The sublethal effects of three different pesticides (a metal, organophosphate, and pyrethroid) on juvenile coho salmon (Oncorhynchus kisutch) were evaluated using paired electrophysiological recordings from the olfactory epithelium and the olfactory bulb. Animals were exposed to copper (5-20 μg·L-1), chlorpyrifos (0.625-2.5 μg·L -1), or esfenvalerate (0.05-0.20 μg·L-1) for 7 days. Sublethal neurotoxicity was examined by recording odor-evoked field potentials from the sensory epithelium and olfactory forebrain using two natural odorants (taurocholic acid or L-serine). Copper and chlorpyrifos decreased the amplitudes of the epithelial and bulbar responses to both odorants in a concentration-dependent manner. Benchmark concentrations for a 20% loss of sensory function were 4.4 μg·L-1 for copper and 0.72 μg·L-1 for chlorpyrifos. Esfenvalerate did not affect the amplitude of odor-evoked field potentials. However, in the olfactory bulbs of coho exposed to 0.2 μg esfenvalerate·L-1, L-serine evoked distinct and irregular bursts of postsynaptic activity in the olfactory bulb, possibly indicating sublethal excitotoxicity to central networks. Collectively, these data indicate that periodic, non-point source contamination of salmon habitats with current-use pesticides could interfere with olfactory function and, by extension, olfactory-mediated behaviors that are important for the survival and migration of salmonids.
Zebrafish have been used predominantly in developmental biology and molecular genetics, but their value in toxicology as well as drug discovery has been recognized. To evaluate the toxicity of a chemical, it is essential to identify the endpoints of toxicity and their dose-response relationships, elucidate the mechanisms of toxicity, and determine the toxicodynamics of the chemical. In addition to detailed toxicological investigations of a single chemical, there also is a need for high-throughput large- scale screening for toxicity of several hundreds of chemicals at a time. In both cases, the zebrafish has numerous attributes. More is probably known about ''what is normal'' in the zebrafish than any other fish species. This includes morpho- logical, biochemical, and physiological information at all stages of early development and in juveniles and adults of both sexes. This makes using the zebrafish ideal for toxicology research where the objective is to identify adverse effects of chemical exposure.
Juvenile rainbow trottt (Oncorhynchus mykiss) werc exposed to I I (control), 300 (medium), and 1000 pg Cug-r (high) (as CuSOa.5H2O) in the diet for 28 days at a daily ration of 4%o wet body weight, with a background waterborne Cu concentration of3 pg.L-|. There was no effect ofdietary Cu on growth, condition factor, or food con- version efficiency. Whole-body Cu content increased continuously over the exposure period in all groups and was two- fold and fourfold higher than controls at day 28 for the medium- and high-Cu diets, respectively. Copper accumulated mailly in liver and gut tissue, with the latter stabilizing by day 14. Accumulation also occurred in gill, kidney, and carcass. Plasma Cu concentration was not different from the controls whereas Cu ir bile was greatly elevated an indi- cation of increased hepatobiliary excretion. Dietary Cu pre-exposure decreased the uptake of waterborne Cu across the gills, providing the fust evidence of homeostatic interaction between the two routes of uptake. Electron microscopic ob- sewations of the midintestine revealed numerous mitochondria, lysosomes, lamellated bodies, and extensive lamellar processes ir the enterocytes. Apoptosis, mitosis, and eosinophilic granule cells were more apparent in Cu-exposed fish. R6sum6 : Des jeunes Truites arc-en-ciel (Oncfutrhynchus mykiss) ont 6t€ exposdes pendant 28 jours i des concentra- tions de 11 (temoin), de 300 (moyenne) ou de 1000 pg Cug-t t6lev6e), sous forme de CuSOo'5HrO, dans des rations alimentaires 6quivalant d 4o/o de ieur masse corporelle humide et dans une eau contenant des concentrations d'arridre plan de Cu de 3 pg'L-I. Le Cu alimentaire est rest6 sans effet sur la croissance, le coefficient d'embonpoint et
Zebrafish have become a popular organism for the study of vertebrate gene function. The virtually transparent embryos of this species, and the ability to accelerate genetic studies by gene knockdown or overexpression, have led to the widespread use of zebrafish in the detailed investigation of vertebrate gene function and increasingly, the study of human genetic disease. However, for effective modelling of human genetic disease it is important to understand the extent to which zebrafish genes and gene structures are related to orthologous human genes. To examine this, we generated a high-quality sequence assembly of the zebrafish genome, made up of an overlapping set of completely sequenced large-insert clones that were ordered and oriented using a high-resolution high-density meiotic map. Detailed automatic and manual annotation provides evidence of more than 26,000 protein-coding genes, the largest gene set of any vertebrate so far sequenced. Comparison to the human reference genome shows that approximately 70% of human genes have at least one obvious zebrafish orthologue. In addition, the high quality of this genome assembly provides a clearer understanding of key genomic features such as a unique repeat content, a scarcity of pseudogenes, an enrichment of zebrafish-specific genes on chromosome 4 and chromosomal regions that influence sex determination.
Zebrafish (Danio rerio) are rapidly gaining popularity in translational neuroscience and behavioral research. Physiological similarity to mammals, ease of genetic manipulations, sensitivity to pharmacological and genetic factors, robust behavior, low cost, and potential for high-throughput screening contribute to the growing utility of zebrafish models in this field. Understanding zebrafish behavioral phenotypes provides important insights into neural pathways, physiological biomarkers, and genetic underpinnings of normal and pathological brain function. Novel zebrafish paradigms continue to appear with an encouraging pace, thus necessitating a consistent terminology and improved understanding of the behavioral repertoire. What can zebrafish ‘do’, and how does their altered brain function translate into behavioral actions? To help address these questions, we have developed a detailed catalog of zebrafish behaviors (Zebrafish Behavior Catalog, ZBC) that covers both larval and adult models. Representing a beginning of creating a more comprehensive ethogram of zebrafish behavior, this effort will improve interpretation of published findings, foster cross-species behavioral modeling, and encourage new groups to apply zebrafish neurobehavioral paradigms in their research. In addition, this glossary creates a framework for developing a
zebrafish neurobehavioral ontology, ultimately to become part of a unified animal neurobehavioral ontology, which
collectively will contribute to better integration of biological data within and across species.
The accumulation of cadmium, copper and zinc and the induction of metallothioneins (MT) in liver of three freshwater fish species was studied. Gudgeon (Gobio gobio), roach (Rutilus rutilus) and perch (Perca fluviatilis) were captured at 6 sampling sites along a cadmium and zinc gradient and one reference site in a tributary of the Scheldt River in Flanders (Belgium).
At each site up to 10 individuals per species were collected and analyzed on their general condition factor (K), hepatosomatic index (HSI) and gonadosomatic index (GSI). From each individual fish the liver was dissected and analyzed on Cd, Cu and Zn and MT-content. Although not all species were present at each site, hepatic Cd and Zn levels generally followed the pollution gradient and highest levels were measured in perch, followed by roach and gudgeon. Nevertheless also an effect of site was observed on this order. MT-levels appeared to be the highest in gudgeon although differences with the other species were not very pronounced and depended on the site. Significant relationships were found between hepatic zinc accumulation and MT levels. For each species the ratio MTtheoretical/ MTmeasured was calculated, which gives an indication of the relative capacity to induce MTs and thus immobilize the metals. Perch had the lowest capacity in inducing MTs (highest ratio). Relationships between hepatic metal levels and fish condition indices were absent or very weak.
Widely used behavioral assays need re-evaluation and validation against their intended use. We focus here on measures of chronic anxiety in mouse models and posit that widely used assays such as the open-field test are performed at the wrong time, for inadequate durations and using inappropriate mouse strains. We propose that behavioral assays be screened for usefulness on the basis of their replicability across laboratories.
The recent introduction of tasks to assess the behavior of zebrafish in novel and/or aversive envi-ronments has spurred great interest, prompting attempts to determine which constructs are modeled by these tasks (e.g., fear, anxiety, or some other construct). A review of the pharmacological and behavioral experiments indicates that not all behavioral testing models are equivalent. A more pre-cise understanding of the parameters that influence task performance affords a wider selection of experimental procedures for investigating a particular construct, and also provides tools for dif-ferentiating the various constructs that may ultimately be of interest. In this review we will more closely examine two behavioral assays commonly used to measure the construct of 'anxiety' in adult zebrafish, with the conclusion that they do not both appear to be measuring a single underlying state.
We assessed whether fish that tolerate higher levels of Cu exposure have a higher capacity to induce metallothionein (MT) synthesis than other, more sensitive, fish species. Furthermore, we examined if a correlation could be found between tissue Cu accumulation and MT levels. Cu accumulation and MT concentrations in gill, liver, kidney and muscle of rainbow trout (Oncorhynchus mykiss), common carp (Cyprinus carpio) and gibel carp (Carassius auratus gibelio) were measured during a 1 week exposure to a sublethal Cu (1 μM).
Fish behavior can be altered by contaminants. There is an extensive literature on laboratory behavioral assays, with many chemicals impairing feeding or predator avoidance. However, there is not extensive work on fishes that live in contaminated environments. Therefore, we then review our recent research on feeding and trophic relations of populations from contaminated estuaries compared with relatively unpolluted sites. The mummichog Fundulus heteroclitus, is a non-migratory fish; those from more contaminated areas are poor predators and slower to capture active prey (grass shrimp, Palaemonetes pugio). In the field, they consume much detritus and sediment, which is not nutritious. They are less active than fish from cleaner sites and more vulnerable to predation. They have altered thyroid glands and neurotransmitter levels, which may underlie altered behaviors. Fish from the reference site kept in tanks with sediment and food from the polluted site showed bioaccumulation and reduced prey capture after two months, although fish from the polluted site did not show significant improvement when maintained in a clean environment. Poor nutrition and predator avoidance may be responsible for their being smaller and having a shorter life span than reference fish. Bluefish Pomatomus saltatrix, are a marine species in which the young-of-the-year spend their first summer in estuaries. We found bioaccumulation of contaminants and reduced activity, schooling, and feeding in young-of-the-year bluefish from a relatively unpolluted site that were fed prey fish from a contaminated site. They also had altered thyroid glands and neurotransmitter levels. Many field-caught specimens had empty stomachs, which is rare in this species. In the fall, when they migrate back out to the ocean, they are smaller, slower, and more likely to starve or to be eaten than those that spent their summer in cleaner estuaries.
Water quality issues are a major challenge that humanity is facing in the twenty-first century. Here, we review the main groups of aquatic contaminants, their effects on human health, and approaches to mitigate pollution of freshwater resources. Emphasis is placed on chemical pollution, particularly on inorganic and organic micropollutants including toxic metals and metalloids as well as a large variety of synthetic organic chemicals. Some aspects of waterborne diseases and the urgent need for improved sanitation in developing countries are also discussed. The review addresses current scientific advances to cope with the great diversity of pollutants. It is organized along the different temporal and spatial scales of global water pollution. Persistent organic pollutants (POPs) have affected water systems on a global scale for more than five decades; during that time geogenic pollutants, mining operations, and hazardous waste sites have been the most relevant sources of long-term regional and local water pollution. Agricultural chemicals and waste-water sources exert shorter-term effects on regional to local scales.
Ultrastructural biomarkers in gill, liver, andkidney of brown trout (Salmo truttaf. fario) and stone loach (Barbatulabarbatula) were investigated over afive year period. The cellular damage of theorgans was assessed semi-quantitatively basedon a three-step classification ofultrastructural responses. Data obtained forfish exposed under semi-field conditions in twodifferently polluted test streams and in thelaboratory demonstrated that the ultrastructureof the organs can be correlated with differentpollutant exposure conditions. Cellular damagewas generally more severe in fish exposed to acomplexly polluted stream than in those exposedto a moderately polluted stream or to tap waterin the laboratory. Histopathological effects inliver and gill of trout were more pronounced inferal fish than in transplanted fish, whereasresponses in the kidney in both species, and inliver and gill of loach, were similar forintroduced and feral fish. In a laboratoryexperiment where trout were exposed todifferent mixtures of pollutants includingpesticides, PAH, and ammonia, only theultrastructure of kidney and liver showedsignificant differences between the threeexperimental set-ups. In a recovery experiment,where trout were transferred from thesemi-field condition back to the laboratory,ultrastructural investigations showed adifferential capacity of the respective organsto recover from stress under field conditions.Kidney and liver fully recovered after threemonths under control condition whereas gillsdemonstrated only partial recovery.
The effects of wall color stimuli on diving, and the effects of depth stimuli on scototaxis, were assessed in zebrafish. Three groups of fish were confined to a black, a white, or a transparent tank, and tested for depth preference. Two groups of fish were confined to a deep or a shallow tank, and tested for black-white preference. As predicted, fish preferred the deep half of a split-tank over the shallow half, and preferred the black half of a black/white tank over the white half. Results indicated that the tank wall color significantly affected depth preference, with the transparent tank producing the strongest depth preference and the black tank producing the weakest preference. Tank depth, however, did not significantly affect color preference. Additionally, wall color significantly affected shuttling and immobility, while depth significantly affected shuttling and thigmotaxis. These results are consistent with previous indications that the diving response and scototaxis may reflect dissociable mechanisms of behavior. We conclude that the two tests are complementary rather than interchangeable, and that further research on the motivational systems underlying behavior in each of the two tests is needed.
Many chemical pollutants have become ubiquitous in the environment, including some that interfere with hormones and other physiological mechanisms. These ‘endocrine-disrupting chemicals’ (EDCs) have harmful effects on development and physiology. We reviewed published evidence and found that EDCs also have adverse effects on a wide range of behaviours, including sexual and other reproductive behaviours, activity, motivation, communication, aggression, dominance and other social behaviours, and learning and other cognitive abilities. We also reviewed recent findings that challenge common assumptions in toxicology. For example, EDCs have several unanticipated properties, such as nonmonotonic dose effects and synergy. Furthermore, harmful effects of EDCs sometimes become apparent only when tested in natural ecological conditions, such as social stress and infection. These findings raise questions about the practicality and feasibility of testing chemical pollutants adequately. Finally, we discuss the implications of EDC research for species conservation and human health, and how toxicology and behavioural research might be better integrated. Behaviour might provide a useful indicator or biomarker for detecting harmful chemical contaminants; however, more integration between toxicology and behavioural ecology is needed to determine whether and how EDCs affect humans and other vertebrates outside of the laboratory.
Effect of quinalphos on the freshwater fish, Cyprinus carpio to
elucidate inhibitory effect of quinalphos on acetyl cholinesterase activity associated
behavioral changes. Fishes were exposed to one fifth (0.15μg/l) and one tenth (0.75
μg/l) of the lethal concentration (7.5 μg/l) of quinalphos for a period of 1, 7 and 14
days, and were allowed to recover for 7 days. Maximum decrement in acetyl
cholinesterase activity of the exposed fish was recorded in brain followed by
muscle, gill and liver. Recovery tenures witnessed increment in acetyl
cholinesterase activity but significantly differed in comparison with control group.
Depression of acetyl cholinesterase activity suggests decreased cholinergic
transmission and consequent accumulation of acetylcholine in the tissues leading to
cessation of nerve impulses. This has lead to behavioral and morphological
changes due to impaired neurophysiology of the fish.
Several components of branchial copper uptake were identified in juvenile freshwater rainbow trout (Oncorhynchus mykiss) using 64Cu. On the basis of competitive interactions between sodium and copper uptake,inhibition of copper uptake by a proton pump inhibitor (bafilomycin A1, 2μmol l-1) and a Na+ channel blocker (phenamil, 100μmol l-1), it appears that a proportion of the branchial copper uptake occurs via an apical Na+ channel. This sodium-sensitive copper uptake demonstrates saturation kinetics, with a Km of 7.1 nmol l-1 and a Jmax of 21.2 pmol g-1 h-1, and is characterized by an IC50 of 104 μmol l-1 sodium. On the basis of residual copper uptake in the presence of high sodium concentrations (20 mmol l-1) and differential inhibition of sodium and copper uptake by phenamil (100 μmol l-1), a sodium-insensitive component of copper uptake is also present in trout gills. It demonstrates saturation kinetics with a comparably low Km (9.6 nmol l-1) but a lower maximum transport capacity (Jmax=3.5 pmol g-1 h-1)than the sodium-insensitive system. Sodium uptake exhibits saturation kinetics with a Km of 69 μmol l-1. Copper reduced branchial sodium transport affinity but increased the maximal sodium transport capacity.
Aquatic toxicology is facing the challenge to assess the impact of complex mixtures of compounds on diverse biological endpoints. So far, ecotoxicology focuses mainly on apical endpoints such as growth, lethality and reproduction, but does not consider sublethal toxic effects that may indirectly cause ecological effects. One such sublethal effect is toxicant-induced impairment of neurosensory functions which will affect important behavioural traits of exposed organisms. Here, we critically review the mechanosensory lateral line (LL) system of zebrafish as a model to screen for chemical effects on neurosensory function of fish in particular and vertebrates in general. The LL system consists of so-called neuromasts, composed of centrally located sensory hair cells, and surrounding supporting cells. The function of neuromasts is the detection of water movements that is essential for the fish's ability to detect prey, to escape predator, to socially interact or to show rheotactic behaviour. Recent advances in the study of these organs provided researchers with a broad area of molecular tools for easy and rapid detection of neuromasts dysfunction and/or disturbed development. Further, genes involved in neuromasts differentiation have been identified using auditory/mechanosensory mutants and morphants. A number of environmental toxicants including metals and pharmaceuticals have been shown to affect neuromasts development and/or function. The use of the LL organ for toxicological studies offers the advantage to integrate the available profound knowledge on developmental biology of the neuromasts with the study of chemical toxicity. This combination may provide a powerful tool in environmental risk assessment.
The main threats to human health from heavy metals are associated with exposure to lead, cadmium, mercury and arsenic. These metals have been extensively studied and their effects on human health regularly reviewed by international bodies such as the WHO. Heavy metals have been used by humans for thousands of years. Although several adverse health effects of heavy metals have been known for a long time, exposure to heavy metals continues, and is even increasing in some parts of the world, in particular in less developed countries, though emissions have declined in most developed countries over the last 100 years. Cadmium compounds are currently mainly used in re-chargeable nickel-cadmium batteries. Cadmium emissions have increased dramatically during the 20th century, one reason being that cadmium-containing products are rarely re-cycled, but often dumped together with household waste. Cigarette smoking is a major source of cadmium exposure. In non-smokers, food is the most important source of cadmium exposure. Recent data indicate that adverse health effects of cadmium exposure may occur at lower exposure levels than previously anticipated, primarily in the form of kidney damage but possibly also bone effects and fractures. Many individuals in Europe already exceed these exposure levels and the margin is very narrow for large groups. Therefore, measures should be taken to reduce cadmium exposure in the general population in order to minimize the risk of adverse health effects. The general population is primarily exposed to mercury via food, fish being a major source of methyl mercury exposure, and dental amalgam. The general population does not face a significant health risk from methyl mercury, although certain groups with high fish consumption may attain blood levels associated with a low risk of neurological damage to adults. Since there is a risk to the fetus in particular, pregnant women should avoid a high intake of certain fish, such as shark, swordfish and tuna; fish (such as pike, walleye and bass) taken from polluted fresh waters should especially be avoided. There has been a debate on the safety of dental amalgams and claims have been made that mercury from amalgam may cause a variety of diseases. However, there are no studies so far that have been able to show any associations between amalgam fillings and ill health. The general population is exposed to lead from air and food in roughly equal proportions. During the last century, lead emissions to ambient air have caused considerable pollution, mainly due to lead emissions from petrol. Children are particularly susceptible to lead exposure due to high gastrointestinal uptake and the permeable blood-brain barrier. Blood levels in children should be reduced below the levels so far considered acceptable, recent data indicating that there may be neurotoxic effects of lead at lower levels of exposure than previously anticipated. Although lead in petrol has dramatically decreased over the last decades, thereby reducing environmental exposure, phasing out any remaining uses of lead additives in motor fuels should be encouraged. The use of lead-based paints should be abandoned, and lead should not be used in food containers. In particular, the public should be aware of glazed food containers, which may leach lead into food. Exposure to arsenic is mainly via intake of food and drinking water, food being the most important source in most populations. Long-term exposure to arsenic in drinking-water is mainly related to increased risks of skin cancer, but also some other cancers, as well as other skin lesions such as hyperkeratosis and pigmentation changes. Occupational exposure to arsenic, primarily by inhalation, is causally associated with lung cancer. Clear exposure-response relationships and high risks have been observed.
Anxiety-related disorders are frequently observed in the population. Because the available pharmacotherapies for anxiety can cause side effects, new anxiolytic compounds have been screened using behavioral tasks. For example, diphenyl diselenide (PhSe)2, a simple organoselenium compound with neuroprotective effects, has demonstrated anxiolytic effects in rodents. However, this compound has not yet been tested in a novelty-based paradigm in non-mammalian animal models. In this study, we assessed the potential anxiolytic effects of (PhSe)2 on the behavior of adult zebrafish under novelty-induced stress. The animals were pretreated with 0.1, 0.25, 0.5, and 1μM (PhSe)2 in the aquarium water for 30min. The fish were then exposed to a novel tank, and their behavior was quantified during a 6-min trial. (PhSe)2 treatment altered fish behavior in a concentration-dependent manner. At 0.01 and 0.25μM, (PhSe)2 did not elicit effects on fish behavior. At 0.5μM, moderate behavioral side effects (e.g., lethargy and short episodic immobility) were noted. At the highest concentration tested (1μM), dramatic side effects were observed, such as burst behavior and longer periods of immobility. The results were confirmed by spatiotemporal analysis of each group. Occupancy plot data showed dispersed homebase formation in the 0.25μM (PhSe)2-treated group compared with the control group (treated with 0.04% DMSO). Furthermore, animals treated with 0.25μM (PhSe)2showed a reduction in latency to enter the top and spent more time in the upper area of the tank. These data suggest that (PhSe)2 may induce an anxiolytic-like effect in situations of anxiety evoked by novelty.
The zebrafish (Danio rerio) is rapidly becoming a popular model organism in pharmacogenetics and neuropharmacology. Both larval and adult zebrafish are currently used to increase our understanding of brain function, dysfunction, and their genetic and pharmacological modulation. Here we review the developing utility of zebrafish in the analysis of complex brain disorders (including, for example, depression, autism, psychoses, drug abuse and cognitive disorders), also covering zebrafish applications towards the goal of modeling major human neuropsychiatric and drug-induced syndromes. We argue that zebrafish models of complex brain disorders and drug-induced conditions have become a rapidly emerging critical field in translational neuropharmacology research.
The effect of preexposure to copper was examined in European eels with respect to uptake and distribution of 64Cu among tissues. Eels were exposed in freshwater to 8 and 64 μg Cu · L−1 for 0, 6 and 28 days before 64Cu incubation. The accumulation of copper in gill, liver and muscle tissue was studied by the uptake of 64Cu during 67 hr of exposure to 8 and 64 μg · L−1. The Cu accumulation rate in muscle tissue was inversely related to bodyweight. Although the accumulation rates in liver and muscle tissues were significantly reduced after 6 and 28 days of preexposure to 64 μg · L−1 (but not to 8 μg · L−1), the accumulation rates in the gills were not affected by preexposure. We suggest that the excretion rate of copper was stimulated by the applied preexposure to 64 μg Cu · L−1.
Sediment flushing may be effective in mitigating loss of reservoir storage due to siltation, but flushing must be controlled to limit the impact on the downstream environment. A reliable prediction of the environmental effects of sediment flushing is hindered by the limited scientific information currently available. Consequently, there may be some controversy as regards to management decisions, planning the work, and monitoring strategies. This paper summarizes the main results of a monitoring campaign on the stream below a small alpine hydropower reservoir subjected to annual flushing between 2006 and 2009. The removed sediment was essentially silt, and the suspended solid concentration (SSC) of the discharged water was controlled to alleviate downstream impact. Control was achieved through hydraulic regulation and mechanical digging, alternating daytime sediment evacuation, and nocturnal clear water release. The four operations lasted about two weeks each and had an average SSC of about 4 g L(-1). Maximum values of SSC were generally kept below 10 g L(-1). Downstream impact was quantified through sampling of fish fauna (brown trout) and macroinvertebrate in the final reach of the effluent stream. The benthic community was severely impaired by the flushing operations, but recovered to pre-flushing values in a few months. As expected, the impact on brown trout was heavier on juveniles. While data biasing due to fish removal and re-stocking cannot be ruled out, the fish community seems to have reached a state of equilibrium characterized by a lower density than was measured before the flushing operations.
Taurine (TAU) is an amino sulfonic acid that plays protective roles against neurochemical impairments induced by ethanol (EtOH). Mounting evidence shows the applicability of zebrafish for evaluating locomotor parameters and anxiety-like behavioral phenotypes after EtOH exposure in a large scale manner. In this study, we assess the effects of TAU pretreatment on the behavior of zebrafish in the open tank after acute 1% EtOH (v/v) exposure (20 and 60 min of duration) and on brain alcohol contents. The exposure for 20 min exerted significant anxiolytic effects, which were prevented by 42, 150, and 400 mg/L TAU. Conversely, the 60-min condition induced depressant/sedative effects, in which the changes on vertical activity were associated to modifications on the exploratory profile. Although all TAU concentrations kept locomotor parameters at basal levels, 150 mg/L TAU, did not prevent the impairment on vertical activity of EtOH[60]. Despite the higher brain EtOH content detected in the 60-min exposure, 42, 150, and 400 mg/L TAU attenuated the increase of alcohol content in EtOH[60] group. In conclusion, our data suggest that both protocols of acute EtOH exposure induce significant changes in the spatio-temporal behavior of zebrafish and that TAU may exert a preventive role by antagonizing the effects induced by EtOH possibly due to its neuromodulatory role and also by decreasing brain EtOH levels. The hormetic dose-response of TAU on vertical exploration suggests a complex interaction between TAU and EtOH in the central nervous system.
Pesticides such as chlorpyrifos (CPF) and metals such as copper can impair swimming behavior in fish. However, the impact to swimming behavior from exposure to mixtures of neurotoxicants has received little attention. In the current study, we analyzed spontaneous swimming rates of adult zebrafish (Danio rerio) to investigate in vivo mixture interactions involving two chemical classes. Zebrafish were exposed to the neurotoxicants copper chloride (CuCl, 0.1 muM, 0.25 muM, 0.6 muM, or 6.3, 16, 40 ppb), chlorpyrifos (CPF, 0.1 muM, 0.25 muM, 0.6 muM, or 35, 88, 220 ppb) and binary mixtures for 24h to better understand the effects of Cu on CPF neurotoxicity. Exposure to CPF increased the number of animals undergoing freeze responses (an anti-predator behavior) and, at the highest CPF dose (0.6 muM), elicited a decrease in zebrafish swimming rates. Interestingly, the addition of Cu caused a reduction in the number of zebrafish in the CPF exposure groups undergoing freeze responses. There was no evidence of additive or synergistic toxicity between Cu and CPF. Although muscle AChE activity was significantly reduced by CPF, there was a relatively poor relationship among muscle AChE concentrations and swimming behavior, suggesting non-muscle AChE mechanisms in the loss of swimming behavior. In summary, we have observed a modulating effect of Cu on CPF swimming impairment that appears to involve both AChE and non-AChE mechanisms. Our study supports the utility of zebrafish in understanding chemical mixture interactions and neurobehavioral injury.
Ethanol (EtOH) is a drug widely consumed throughout the world that promotes several neurochemical disorders. Its deleterious effects are generally associated with modifications in oxidative stress parameters, signaling transduction pathways, and neurotransmitter systems, leading to distinct behavioral changes. Taurine (2-aminoethanesulfonic acid) is a β-amino acid not incorporated into proteins found in mM range in the central nervous system (CNS). The actions of taurine as an inhibitory neurotransmitter, neuromodulator, and antioxidant make it attractive for studying a potential protective role against EtOH-mediated neurotoxicity. In this study, we investigated whether acute taurine cotreatment or pretreatment (1 h) prevent EtOH-induced changes in acetylcholinesterase (AChE) activity and in oxidative stress parameters in zebrafish brain. The results showed that EtOH exposure (1% in volume) during 1 h increased AChE activity, whereas the cotreatment with 400 mg·L−1 taurine prevented this enhancement. A similar protective effect of 150 and 400 mg·L−1 taurine was also observed when the animals were pretreated with this amino acid. Taurine treatments also prevented the alterations promoted in superoxide dismutase and catalase activities by EtOH, suggesting a modulatory role in enzymatic antioxidant defenses. The pretreatment with 150 and 400 mg·L−1 taurine significantly increased the sulfydryl levels as compared to control and EtOH groups. Moreover, 150 and 400 mg·L−1 taurine significantly decreased thiobarbituric acid reactive species (TBARS) levels, but the cotreatment with EtOH plus 400 mg·L−1 taurine did not prevent the EtOH-induced lipoperoxidation. In contrast, the pretreatment with 150 and 400 mg·L−1 taurine prevented the TBARS increase besides decreased the basal levels of lipid peroxides. Altogether, our data showed for the first time that EtOH induced oxidative stress in adult zebrafish brain and reinforce the idea that this vertebrate is an attractive alternative model to evaluate the beneficial effect of taurine against acute EtOH exposure.
64Cu and total Cu accumulation were measured in gills, plasma, liver, kidney, bile and urine during 72 h of exposure to 64Cu at 20 μg Cu l−1, in non-acclimated and Cu-acclimated (28 days of pre-exposure) rainbow trout (Oncorhynchus mykiss) fitted with urinary bladder catheters. Renal Cu excretion gradually declined from 0.03 μg Cu kg−1 h−1 in non-exposed fish to 0.01 μg Cu kg−1 h−1 after 28 days of Cu exposure. A comparison of the 64Cu-labelled Cu and the total Cu excretion rates and the corresponding renal clearance revealed apparent differences in Cu binding to plasma protein depending on whether the Cu is derived from recent branchial uptake or is already present in the plasma prior to 64Cu exposure. The plasma Cu pool derived from recent branchial uptake and the Cu pool present in the plasma prior to 64Cu exposure is accessible to renal excretion to different extents, whereas the pools seem equally accessible to hepatic accumulation and elimination. The renal Cu excretion is of minor importance compared with the hepatic Cu excretion, which was estimated to be 0.5–0.75 μg Cu kg−1 h−1 and 1.1–1.6 μg Cu kg−1 h−1 for non-acclimated and Cu-acclimated fish, respectively. Based on the biliary Cu concentration, hepatic Cu elimination appeared to be stimulated in the Cu-acclimated relative to the non-acclimated fish. Only 17% and 12% of the hepatic Cu could be accounted for by metallothionein in the control and Cu-acclimated fish, respectively. Renal Na+ efflux decreased by 40%, which was largely due to increased tubular Na+ reabsorption. Renal compensation for the impaired branchial Na+ uptake, seen during Cu exposure, thus seems to be involved in Cu acclimation in rainbow trout.
64Cu accumulation and total Cu concentration were measured in plasma, red blood cells, gills, liver, kidney and bile during 65 h of exposure to 64Cu at 20 μg of Cu per liter, in non acclimated and Cu acclimated (28 days of preexposure) rainbow trout (Oncorhynchus mykiss) fitted with a dorsal aortic catheter. By measuring both 64Cu accumulation and total Cu concentrations, we were able to analyse the ongoing uptake and turnover of ambient Cu, independent of any Cu already present in the fish.Plasma accounted for at least 90% of the 64Cu labelled Cu present in the blood and Cu-acclimation clearly involves changes in copper accumulation kinetics in the plasma. The acclimated fish showed a 65% reduced 64Cu accumulation after 65 h and an increased turnover of Cu in the plasma compared to the non-acclimated fish. Total Cu in the plasma increased by 59% after 3 h of exposure in the non-acclimated fish but was recovered during the following 24 h and remained at control levels throughout 65 h; even after 28 days the acclimated fish showed no increase in total plasma [Cu]. Apparently Cu acclimation involves an increased clearing of plasma Cu, primarily to the liver, stimulated during the first 12 h of exposure.Acclimation did not have an unambiguous effect on branchial Cu uptake and differences in branchial uptake could not explain the reduced accumulation in the plasma. The rapidly exchangeable Cu pools were 54% in the gills and 33% in the liver, suggesting a considerable hepatic Cu elimination. No increase in the total [Cu] in the kidney was observed, but the kidney did show substantial 64Cu accumulation and thus also a potential renal Cu excretion.
A photometric method for determining acetylcholinesterase activity of tissue extracts, homogenates, cell suspensions, etc., has been described. The enzyme activity is measured by following the increase of yellow color produced from thiocholine when it reacts with dithiobisnitrobenzoate ion. It is based on coupling of these reactions: The latter reaction is rapid and the assay is sensitive (i.e. a 10 μ1 sample of blood is adequate). The use of a recorder has been most helpful, but is not essential. The method has been used to study the enzyme in human erythrocytes and homogenates of rat brain, kidney, lungs, liver and muscle tissue. Kinetic constants determined by this system for erythrocyte eholinesterase are presented. The data obtained with acetylthiocholine as substrate are similar to those with acetylcholine.
The many documents on metallothioneins (MTs) in aquatic organisms focus especially on their use as biomarkers in environmental monitoring programs, but there are a few papers that summarize the physiological role of MTs in aquatic organisms especially in their development. The multifaceted role of MTs include involvement in homeostasis, protection against heavy metals and oxidant damage, metabolic regulation, sequestration and/or redox control. MTs could be induced by heavy metals which are able to hinder gametogenesis, suppress embryogenesis, and hamper development. Here we pay more attention on the non-essential metal cadmium, which is the most studied heavy metal regarding MTs, and its effects on the development of aquatic invertebrates. In this paper, we have collected published information on MTs in aquatic organisms - mollusks, crustaceans, etc., and summarize its functions in aquatic invertebrates, especially those related to their development.
Recent advances in neurobiology have emphasized the study of brain structure and function and its association with numerous pathological and toxicological events. Neurotransmitters are substances that relay, amplify, and modulate electrical signals between neurons and other cells. Neurotransmitter signaling mediates rapid intercellular communication by interacting with cell surface receptors, activating second messenger systems and regulating the activity of ion channels. Changes in the functional balance of neurotransmitters have been implicated in the failure of central nervous system function. In addition, abnormalities in neurotransmitter production or functioning can be induced by several toxicological compounds, many of which are found in the environment. The zebrafish has been increasingly used as an animal model for biomedical research, primarily due to its genetic tractability and ease of maintenance. These features make this species a versatile tool for pre-clinical drug discovery and toxicological investigations. Here, we present a review regarding the role of different excitatory and inhibitory neurotransmitter systems in zebrafish, such as dopaminergic, serotoninergic, cholinergic, purinergic, histaminergic, nitrergic, glutamatergic, glycinergic, and GABAergic systems, and emphasizing their features as pharmacological and toxicological targets. The increase in the global knowledge of neurotransmitter systems in zebrafish and the elucidation of their pharmacological and toxicological aspects may lead to new strategies and appropriate research priorities to offer insights for biomedical and environmental research.
Ethanol has been suggested to have an anxiolytic effect on zebrafish, primarily based on its disruption of the novel tank diving response and of some social behaviors. The light/dark preference test offers a complementary measure of anxiety-like behavior in fish, and the purpose of the current study was to determine the effects of acute ethanol exposure on behavior in the light/dark task. In Experiment 1, the stimuli used to induce light/dark preference in zebrafish were varied in order to determine how best to measure the behavior. Subjects exhibited phototaxis (preference for light) when illumination was manipulated, but scototaxis (preference for dark) when wall and substrate color were manipulated. There was a clear interaction between locomotor activity and color preference, with animals preferentially freezing in darker locations. Because of ambiguity in interpreting behavior in the open/covered version of the test, the black/white version was used in Experiment 2. In Experiment 2, zebrafish were exposed to ethanol (0.25%, 0.5%, or 1.0%) or water for 30 minutes, and then placed in a black/white preference tank containing either ethanol (same doses) or water for a 30-minute test. Ethanol exposure increased locomotor activity and reduced freezing. Additionally, there was a significant interaction between ethanol treatment and locomotor activity on side preference. Low doses of ethanol increased white avoidance in normally swimming fish, while high doses did not.