Plastic pollution is a growing global emergency and it could serve as a geological indicator of the Anthropocene era. Microplastics are potentially more hazardous than macroplastics, as the former can permeate biological membranes. The toxicity of microplastic exposure on humans and aquatic organisms has been documented, but the toxicity and behavioral changes of nanoplastics (NPs) in mammals are scarce. In spite of their small size, nanoplastics have an enormous surface area, which bears the potential to bind even bigger amounts of toxic compounds in comparison to microplastics. Here, we used polystyrene nanoplastics (PS-NPs) (diameter size at ~70 nm) to investigate the neurobehavioral alterations, tissue distribution, accumulation, and specific health risk of nanoplastics in adult zebrafish. The results demonstrated that PS-NPs accumulated in gonads, intestine, liver, and brain with a tissue distribution pattern that was greatly dependent on the size and shape of the NPs particle. Importantly, an analysis of multiple behavior endpoints and different biochemical biomarkers evidenced that PS-NPs exposure induced disturbance of lipid and energy metabolism as well as oxidative stress and tissue accumulation. Pronounced behavior alterations in their locomotion activity, aggressiveness, shoal formation, and predator avoidance behavior were exhibited by the high concentration of the PS-NPs group, along with the dysregulated circadian rhythm locomotion activity after its chronic exposure. Moreover, several important neurotransmitter biomarkers for neurotoxicity investigation were significantly altered after one week of PS-NPs exposure and these significant changes may indicate the potential toxicity from PS-NPs exposure. In addition, after ~1-month incubation, the fluorescence spectroscopy results revealed the accumulation and distribution of PS-NPs across zebrafish tissues, especially in gonads, which would possibly further affect fish reproductive function. Overall, our results provided new evidence for the adverse consequences of PS-NPs-induced behavioral dysregulation and changes at the molecular level that eventually reduce the survival fitness of zebrafish in the ecosystem.

Leptin, a hormone secreted by peripheral adipose tissues, regulates the appetite in animals. Recently, evidence has shown that leptin also plays roles in behavioral response in addition to controlling appetite. In this study, we examined the potential function of leptin on non-appetite behaviors in zebrafish model. By using genome editing tool of Transcription activator-like effector nuclease (TALEN), we successfully knocked out leptin a (lepa) gene by deleting 4 bp within coding region to create a premature-translation stop. Morphological and appetite analysis showed the lepa KO fish display a phenotype with obese, good appetite and elevation of Agouti-related peptide (AgRP) and Ghrelin hormones, consistent with the canonical function of leptin in controlling food intake. By multiple behavior endpoint analyses, including novel tank, mirror biting, predator avoidance, social interaction, shoaling, circadian rhythm, and color preference assay, we found the lepa KO fish display an anxiogenic phenotype showing hyperactivity with rapid swimming, less freezing time, less fear to predator, loose shoaling area forming, and circadian rhythm and color preference dysregulations. Using biochemical assays, melatonin, norepinephrine, acetylcholine and serotonin levels in the brain were found to be significantly reduced in lepa KO fish, while the levels of dopamine, glycine and cortisol in the brain were significantly elevated. In addition, the brain ROS level was elevated, and the anti-oxidative enzyme catalase level was reduced. Taken together, by performing loss-of-function multiple behavior endpoint testing and biochemical analysis, we provide strong evidence for a critical role of lepa gene in modulating anxiety, aggression, fear, and circadian rhythm behaviors in zebrafish for the first time.

In this study, we evaluated the acute (24, 48, 72, and 96 h) and chronic (21 days) adverse effects induced by low doses (0.1, 0.5, 1, and 1.5 mg/L) of zinc chloride (ZnCl2) exposure in adult zebrafish by using behavioral endpoints like three-dimensional (3D) locomotion, passive avoidance, aggression, circadian rhythm, and predator avoidance tests. Also, brain tissues were dissected and subjected to analysis of multiple parameters related to oxidative stress, antioxidant responses, superoxide dismutase (SOD), neurotoxicity, and neurotransmitters. The results showed that ZnCl2-exposed fishes displayed decreased locomotor behavior and impaired short-term memory, which caused an Alzheimer’s Disease (AD)-like syndrome. In addition, low concentrations of ZnCl2 induced amyloid beta (amyloid β) and phosphorylated Tau (p-Tau) protein levels in brains. In addition, significant induction in oxidative stress indices (reactive oxygen species (ROS) and malondialdehyde (MDA)), reduction in antioxidant defense system (glutathione (GSH), GSH peroxidase (GSH-Px) and SOD) and changes in neurotransmitters were observed at low concentrations of ZnCl2. Neurotoxic effects of ZnCl2 were observed with significant inhibition of acetylcholine (ACh) activity when the exposure dose was higher than 1 ppm. Furthermore, we found that zinc, metallothionein (MT), and cortisol levels in brain were elevated compared to the control group. A significantly negative correlation was observed between memory and acetylcholinesterase (AChE) activity. In summary, these findings revealed that exposure to ZnCl2 affected the behavior profile of zebrafish, and induced neurotoxicity which may be associated with damaged brain areas related to memory. Moreover, our ZnCl2-induced zebrafish model may have potential for AD-associated research in the future.

There is a growing concern for the potential toxicity of engineered nanomaterials that have made their way into virtually all novel applications in the electronics, healthcare, cosmetics, technology, and engineering industries, and in particular, biomedical products. However, the potential toxicity of carbon 60 (C60) at the behavioral level has not been properly evaluated. In this study, we used idTracker, a multitracking algorithm to quantitatively assess behavioral toxicity induced by C60 nanoparticles (C60 NPs) in adult zebrafish. We demonstrated that locomotion, novel tank exploration, aggression, shoaling, and color preference activities of the C60 NPs-treated fish was significantly reduced. In addition, the C60 NPs-treated fish also displayed dysregulation of the circadian rhythm by showing lower locomotion activities in both day and night cycles. The biochemical results showed that C60 NPs exposure at low concentration induced oxidative stress and DNA damage, reduced anti-oxidative capacity and ATP (adenosine triphosphate) levels, and induced stress-associated hormones, hypoxia, as well as inflammation marker upregulation in muscle and gill tissues. Together, this work, for the first time, provide direct evidence showing that the chronic exposure of C60 NPs induced multiple behavioral abnormalities in adult zebrafish. Our findings suggest that the ecotoxicity of C60 NPs towards aquatic vertebrates should be carefully evaluated.

There is an imperative need to develop efficient whole-animal-based testing assays to determine the potential toxicity of engineered nanomaterials. While previous studies have demonstrated toxicity in lung and skin cells after C70 nanoparticles (NPs) exposure, the potential detrimental role of C70 NPs in neurobehavior is largely unaddressed. Here, we evaluated the chronic effects of C70 NPs exposure on behavior and alterations in biochemical responses in adult zebrafish. Two different exposure doses were used for this experiment: low dose (0.5 ppm) and high dose (1.5 ppm). Behavioral tests were performed after two weeks of exposure of C70 NPs. We found decreased locomotion, exploration, mirror biting, social interaction, and shoaling activities, as well as anxiety elevation and circadian rhythm locomotor activity impairment after ~2 weeks in the C70 NP-exposed fish. The results of biochemical assays reveal that following exposure of zebrafish to 1.5 ppm of C70 NPs, the activity of superoxide dismutase (SOD) in the brain and muscle tissues increased significantly. In addition, the concentration of reactive oxygen species (ROS) also increased from 2.95 ± 0.12 U/ug to 8.46 ± 0.25 U/ug and from 0.90 ± 0.03 U/ug to 3.53 ± 0.64 U/ug in the muscle and brain tissues, respectively. Furthermore, an increased level of cortisol was also observed in muscle and brain tissues, ranging from 17.95 ± 0.90 pg/ug to 23.95 ± 0.66 pg/ug and from 3.47 ± 0.13 pg/ug to 4.91 ± 0.51 pg/ug, respectively. Increment of Hif1-α level was also observed in both tissues. The elevation was ranging from 11.65 ± 0.54 pg/ug to 18.45 ± 1.00 pg/ug in the muscle tissue and from 4.26 ± 0.11 pg/ug to 6.86 ± 0.37 pg/ug in the brain tissue. Moreover, the content of DNA damage and inflammatory markers such as ssDNA, TNF-α, and IL-1β were also increased substantially in the brain tissues. Significant changes in several biomarker levels, including catalase and malondialdehyde (MDA), were also observed in the gill tissues. Finally, we used a neurophenomic approach with a particular focus on environmental influences, which can also be easily adapted for other aquatic fish species, to assess the toxicity of metal and carbon-based nanoparticles. In summary, this is the first study to illustrate the adult zebrafish toxicity and the alterations in several neurobehavior parameters after zebrafish exposure to environmentally relevant amounts of C70 NPs.

Magnetic Nanoparticles (MNPs) are widely being investigated as novel promising multifunctional agents, specifically in the fields of development for theranostics, electronics, waste water treatment, cosmetics, and energy storage devices. Unique, superior, and indispensable properties of magnetization, heat transfer, and melting temperature make MNPs emerge in the field of therapeutics in future healthcare industries. However, MNPs ecotoxicity as well as behavioral toxicity is still unexplored. Ecotoxicity analysis may assist investigate MNPs uptake mechanism and its influence on bioavailability under a given set of environmental factors, which can be followed to investigate the biomagnification of MNPs in the environment and health risk possessed by them in an ecological food chain. In this study, we attempted to determine the behavioral changes in zebrafishes at low (1 ppm) or high (10 ppm) concentration levels of Fe3O4 MNPs. The synthesized Fe3O4 MNPs sized at 15 nm were characterized by the transmission electron microscope (TEM), the superconducting quantum interference device (SQUID) magnetometer, and the multiple behavior tests for novel tank, mirror biting, conspecific social interaction, shoaling, circadian rhythm, and short-term memory of zebrafish under MNPs chronic exposure were demonstrated. Low concentration MNP exposure did not trigger alteration for majority behavioral and biochemical tests in adult zebrafish. However, tight shoal groups were observed at a high concentration of MNPs exposure along with a modest reduction in fish exploratory behavior and a significant reduction in conspecific social interaction behavior. By using enzyme-linked immunosorbent assays (ELISA), we found a high dose of MNPs exposure significantly elevated cortisol, acetylcholine, and catalase levels while reducing serotonin, acetylcholine esterase, and dopamine levels in the brain. Our data demonstrates chronic MNPs exposure at an environmentally-relevant dose is relatively safe by supporting evidence from an array of behavioral and biochemical tests. This combinational approach using behavioral and biochemical tests would be helpful for understanding the MNPs association with anticipated colloids and particles effecting bioavailability and uptake into cells and organisms.

Animal models play important roles in investigating the pathobiology of cancer, identifying relevant pathways, and developing novel therapeutic tools. Despite rapid progress in the understanding of disease mechanisms and technological advancement in drug discovery, negative trial outcomes are the most frequent incidences during a Phase III trial. Skin cancer is a potential life-threatening disease in humans and might be medically futile when tumors metastasize. This explains the low success rate of melanoma therapy amongst other malignancies. In the past decades, a number of skin cancer models in fish that showed a parallel development to the disease in humans have provided important insights into the fundamental biology of skin cancer and future treatment methods. With the diversity and breadth of advanced molecular genetic tools available in fish biology, fish skin cancer models will continue to be refined and expanded to keep pace with the rapid development of skin cancer research. This review begins with a brief introduction of molecular characteristics of skin cancers, followed by an overview of teleost models that have been used in the last decades in melanoma research. Next, we will detail the importance of the zebrafish (Danio rerio) animal model and other emerging fish models including platyfish (Xiphophorus sp.), and medaka (Oryzias latipes) in future cutaneous malignancy studies. The last part of this review provides the recent development and genome editing applications of skin cancer models in zebrafish and the progress in small molecule screening.

The zebrafish (Danio rerio) is a versatile model organism that has been used in biomedical research for several decades to study a wide range of biological phenomena. There are many technical advantages of using zebrafish over other vertebrate models. They are readily available, hardy, easy, and inexpensive to maintain in the laboratory, have a short life cycle, and have excellent fecundity. Due to its optical clarity and reproducible capabilities, it has become one of the predominant models of human genetic diseases. Zebrafish research has made rapid strides in the United States and Europe, but in India the field is at an early stage and many researchers still remain unaware of the full research potential of this tiny fish. The zebrafish model system was introduced into India in the early 2000s. Up to now, more than 200 scientific referred articles have been published by Indian researchers. This review gives an overview of the current state of knowledge for zebrafish research in India, with the aim of promoting wider utilization of zebrafish for high level biological studies.