Utility of zebrafish-based models in understanding molecular mechanisms of neurotoxicity mediated by the gut-brain axis.

The gut microbes perform several beneficial functions which impact the periphery and central nervous systems of the host. Gut microbiota dysbiosis is acknowledged as a major contributor to the development of several neuropsychiatric and neurological disorders including bipolar disorder, depression, anxiety, Parkinson's disease, Alzheimer's disease, attention deficit hyperactivity disorder, and autism spectrum disorder. Thus, elucidation of how the gut microbiota-brain axis plays a role in health and disease conditions is a potential novel approach to prevent and treat brain disorders. The zebrafish (Danio rerio) is an invaluable vertebrate model that possesses conserved brain and intestinal features with those of humans, thus making zebrafish a valued model to investigate the interplay between the gut microbiota and host health. This chapter describes current findings on the utility of zebrafish in understanding molecular mechanisms of neurotoxicity mediated via the gut microbiota-brain axis. Specifically, it highlights the utility of zebrafish as a model organism for understanding how anthropogenic chemicals, pharmaceuticals and bacteria exposure affect animals and human health via the gut-brain axis.

Towards zebrafish models to unravel translational insights of obsessive-compulsive disorder: A neurobehavioral perspective.

Obsessive-compulsive disorder (OCD) is a chronic and debilitating illness that has been considered a polygenic and multifactorial disorder, challenging effective therapeutic interventions. Although invaluable advances have been obtained from human and rodent studies, several molecular and mechanistic aspects of OCD etiology are still obscure. Thus, the use of non-traditional animal models may foster innovative approaches in this field, aiming to elucidate the underlying mechanisms of disease from an evolutionary perspective. The zebrafish (Danio rerio) has been increasingly considered a powerful organism in translational neuroscience research, especially due to the intrinsic features of the species. Here, we outline target mechanisms of OCD for translational research, and discuss how zebrafish-based models can contribute to explore neurobehavioral aspects resembling those found in OCD. We also identify possible advantages and limitations of potential zebrafish-based models, as well as highlight future directions in both etiological and therapeutic research. Lastly, we reinforce the use of zebrafish as a promising tool to unravel the biological basis of OCD, as well as novel pharmacological therapies in the field.

The influence of acute dopamine transporter inhibition on manic-, depressive-like phenotypes, and brain oxidative status in adult zebrafish.

Functional changes in dopamine transporter (DAT) are related to various psychiatric conditions, including bipolar disorder (BD) symptoms. In experimental research, the inhibition of DAT induces behavioral alterations that recapitulate symptoms found in BD patients, including mania and depressive mood. Thus, developing novel animal models that mimic BD-related conditions by pharmacologically modulating the dopaminergic signaling is relevant. The zebrafish (Danio rerio) has been considered a suitable vertebrate system for modeling BD-like responses, due to the well-characterized behavioral responses and evolutionarily conservation of the dopaminergic system of this species. Here, we investigate whether GBR 12909, a selective inhibitor of DAT, causes neurobehavioral alterations in zebrafish similar to those observed in BD patients. Behaviors were recorded after a single intraperitoneal (i.p.) administration of GBR 12909 at different doses (3.75, 7.5, 15 and 30 mg/kg). To observe temporal effects on behavior, swim path parameters were measured immediately after the administration period during 30 min. Locomotion, anxiety-like behavior, social preference, aggression, despair-like behavior, and oxidative stress-related biomarkers in the brain were measured 30 min post administration. GBR 12909 induced prominent effects on locomotor activity and vertical exploration during the 30-min period. Hyperactivity was observed in GBR 30 group after 25 min, while all doses markedly reduced vertical drifts. GBR 12909 elicited hyperlocomotion, anxiety-like behavior, decreased social preference, aggression, and induced depressive-like behavior in a behavioral despair task. Depending on the dose, GBR 12909 also decreased SOD activity and TBARS levels, as well as increased GR activity and NPSH content. Collectively, our novel findings show that a single GBR 12909 administration evokes neurobehavioral changes that recapitulate manic- and depressive-like states observed in rodents, fostering the use of zebrafish models to explore BD-like responses in translational neuroscience research.

Expanding the use of homebase-related parameters to investigate how distinct stressful conditions affect zebrafish behaviors.

Stress is a physiological reaction that allows the organisms to cope with challenging situations daily. Thus, elucidating the behavioral outcomes following different stressors is of great importance in translational research. Here, we aimed to characterize the main factors which explain similarities and differences of two stress protocols on zebrafish exploratory activity. To answer this point, we performed behavioral analyses aiming to simplify the data structure associated with homebase-related measurements in an integrated manner. Adult zebrafish were exposed to conspecific alarm substance for 5 min (acute stress protocol - AS) or submitted to 7 days of unpredictable chronic stress (UCS). Immediately after AS or in the subsequent day following UCS (8th day), fish were individually tested in the open field and the behaviors were recorded for 30 min to posterior identification of homebase locations. For both protocols, behavioral clustering revealed two major clusters, grouping homebase- and locomotor-related parameters, respectively. While AS increased both positive and negative correlations between exploratory and locomotor endpoints, a significant increase in negative correlations was found in UCS-challenged fish. Comparison of the principal component analyses data set revealed a reduced exploratory activity using the homebase in AS group, while decreased locomotion in the periphery and anxiety-like behaviors were evidenced in UCS fish. In conclusion, our findings revealed a different structure of behavior in zebrafish following AS and UCS protocols, supporting the existence of distinct behavioral strategies to cope with acute and chronic stress. Furthermore, we expand the use of homebase-related measurements as a valuable tool to investigate complex behavioral modulations in future translational neuropsychiatry research.

Influence of acute and unpredictable chronic stress on spatio-temporal dynamics of exploratory activity in zebrafish with emphasis on homebase-related behaviors.

The open field is a suitable task to analyze the sequential organization of exploratory activity and the homebase formation represents an important feature of environmental recognition. Although the zebrafish can define homebase locations, there are no data reporting how stressful conditions modulate complex behaviors of this aquatic species in the open field so far. Here, we aimed to characterize the spatio-temporal exploratory activity of adult zebrafish in the open field test, as well as to verify the responsiveness of homebase-related parameters to acute stress (AS) and unpredictable chronic stress (UCS) protocols. Animals were exposed to conspecific alarm substance for 5 min or subjected to a 7-days stress protocol using distinct stressors in an unpredictable manner. Immediately after exposure to AS or 24 h after UCS, fish were individually placed in a circular tank and their behaviors were recorded for 30 min to identify the respective homebase for each animal. We observed that UCS, but not AS, increased thigmotaxis compared to the non-stressed fish. Notably, the sequential organization of exploratory activity showed robust differences depending on the stress protocol. After the first 15 min of trial, AS-challenged fish apparently used the homebase to organize briefly explorations to the environment. Conversely, the UCS group was more immobile in the homebase after periodically performing 'swimming bursts' to the periphery with a greater number of stops per trip. Physiological stress responses were confirmed by the increased whole-body cortisol in both AS and UCS groups. In conclusion, our novel findings report a different exploratory profile related to stress responses in adult zebrafish tested in the open field, supporting the sensitivity of homebase-related parameters to manipulations that modulate affective-like states.

Induction of aggression and anxiety-like responses by perfluorooctanoic acid is accompanied by modulation of cholinergic- and purinergic signaling-related parameters in adult zebrafish.

Perfluorooctanoic acid (PFOA) is a contaminant of global concern owing to its prevalent occurrence in aquatic and terrestrial environments with potential hazardous impact on living organisms. Here, we investigated the influence of realistic environmental concentrations of PFOA (0, 0.25, 0.5, or 1.0 mg/L) on relevant behaviors of adult zebrafish (Danio rerio) (e.g., exploration to novelty, social preference, and aggression) and the possible role of PFOA in modulating cholinergic and purinergic signaling in the brain after exposure for 7 consecutive days. PFOA significantly increased geotaxis as well as reduced vertical exploration (a behavioral endpoint for anxiety), and increased the frequency and duration of aggressive episodes without affecting their social preference. Exposure to PFOA did not affect ADP hydrolysis, whereas ATP and AMP hydrolysis were significantly increased at the highest concentration tested. However, AChE activity was markedly decreased in all PFOA-exposed groups when compared with control. In conclusion, PFOA induces aggression and anxiety-like behavior in adult zebrafish and modulates both cholinergic and purinergic signaling biomarkers. These novel data can provide valuable insights into possible health threats related to human activities, demonstrating the utility of adult zebrafish to elucidate how PFOA affects neurobehavioral responses in aquatic organisms.

A novel behavioral paradigm to measure anxiety-like behaviors in zebrafish by the concomitant assessment of geotaxis and scototaxis.

Pathological anxiety is a set of diseases characterized by specific clinical manifestations and the use of alternative models may provide novel insights in translational neurobehavioral research. In zebrafish, the separate performance of novel tank and light dark tests in different order to assess anxiety using a same animal may provide conflicting data due to the battery effect and/or time-drug-response and variability across tests. To improve data reliability, we aimed to characterize a novel behavioral paradigm to measure geotaxis and scototaxis as anxiety-like responses in the same trial. The novel apparatus consisted of four colored-compartments, with specific white- and black sections delimited in both bottom and upper areas of the tank. The main baseline responses of zebrafish in the novel apparatus were measured and animals were further exposed to modulators of anxiety. Zebrafish showed robust habituation to novelty stress during the 6-min trial with preference for the black section while exploring the top area. Fluoxetine (100 µg/L, 15 min) reduced geotaxis and scototaxis and ketamine (20 mg/L, 20 min) decreased geotaxis and increased the distance traveled in the black section while exploring the top, possibly due to the increased circling behavior. As anxiogenic modulators, conspecific alarm substance (3.5 mL/L, 5 min) exacerbated risk assessment, geotaxis, and scototaxis, whereas caffeine (10 mg/L, 15 min) increased geotaxis and exploration in the black section of the top area. Since important correlations were also found for relevant anxiety-like behaviors, our findings support the predictive validity of this novel paradigm to simultaneously assess geotaxis and scototaxis in zebrafish. Moreover, it fully adheres to the 3Rs principle of animal experimentation of reducing the number of subjects tested, execution time, also minimizing a potential battery effect.

Zebrafish as a potential non-traditional model organism in translational bipolar disorder research: Genetic and behavioral insights.

Bipolar disorder (BD) is a severe and debilitating illness that affects 1-2% of the population worldwide. BD is characterized by recurrent and extreme mood swings, including mania/hypomania and depression. Animal experimental models have been used to elucidate the mechanisms underlying BD and different strategies have been proposed to assess BD-like symptoms. The zebrafish (Danio rerio) has been considered a suitable vertebrate system for modeling BD-like responses, due to the genetic tractability, molecular/physiological conservation, and well-characterized behavioral responses. In this review, we discuss how zebrafish-based models can be successfully used to understand molecular, biochemical, and behavioral alterations paralleling those found in BD. We also outline some advantages and limitations of this aquatic species to examine BD-like phenotypes in translational neurobehavioral research. Overall, we reinforce the use of zebrafish as a promising tool to investigate the neural basis associated with BD-like behaviors, which may foster the discovery of novel pharmacological therapies.

Influence of acid-sensing ion channel blocker on behavioral responses in a zebrafish model of acute visceral pain.

Acid-sensing ion channels (ASICs) play significant roles in numerous neurological and pathological conditions, including pain. Although acid-induced nociception has been characterized previously in zebrafish, the contribution of ASICs in modulating pain-like behaviors is still unknown. Here, we investigated the role of amiloride, a nonselective ASICs blocker, in the negative modulation of specific behavioral responses in a zebrafish-based model of acute visceral pain. We verified that intraperitoneal injection (i.p.) of 0.25, 0.5, 1.0, and 2.0 mg/mL amiloride alone or vehicle did not change zebrafish behavior compared to saline-treated fish. Administration of 2.5% acetic acid (i.p.) elicited writhing-like response evidenced by the abnormal body curvature and impaired locomotion and motor activity. Attenuation of acetic acid-induced pain was verified at lower amiloride doses (0.25 and 0.5 mg/mL) whereas 1.0 and 2.0 mg/mL abolished pain-like responses. The protective effect of the highest amiloride dose tested was evident in preventing writhing-like responses and impaired locomotion and vertical activity. Collectively, amiloride antagonized abdominal writhing-like phenotype and aberrant behaviors, supporting the involvement of ASICs in a zebrafish-based model of acute visceral pain.

Taurine modulates behavioral effects of intermittent ethanol exposure without changing brain monoamine oxidase activity in zebrafish: Attenuation of shoal- and anxiety-like responses, and abolishment of memory acquisition deficit.

Prolonged alcohol consumption has been considered as an important risk factor for various diseases. Chronic ethanol (EtOH) intake is associated with deleterious effects on brain functions culminating in robust behavioral changes. Notably, drugs available to treat the effects of EtOH have low therapeutic efficacy so far. Taurine (TAU) appears as a promising neuroprotective molecule due to its pleiotropic action in the brain. Here, we investigated whether TAU plays a beneficial role in different behavioral domains of zebrafish submitted to an intermittent EtOH exposure model, specially focusing on social behavior, anxiety-like responses, and memory. Moreover, since monoamines play a role in EtOH-mediated responses, we also evaluated the influence of both TAU and EtOH exposures on brain monoamine oxidase (Z-MAO) activity. Fish were exposed to non-chlorinated water or 1% EtOH for 8 consecutive days (20 min per day). From the 5th day until the end of the experimental period (8th day), animals were kept in the absence or presence of TAU (42, 150, or 400 mg/L) 1 h per day immediately after EtOH exposure. Behavioral measurements started 24 h after the last EtOH exposure. We observed that TAU showed modest attenuating effects on shoaling behavior and anxiety-like responses, while 42 and 150 mg/L TAU abolished the memory acquisition deficit in the inhibitory avoidance task. Biochemical analysis revealed that TAU did not modulate EtOH-induced increase on brain Z-MAO activity. Collectively, our novel data show a potential beneficial effect of TAU in an intermittent EtOH exposure model in zebrafish. Moreover, these findings foster the growing utility of this aquatic species to investigate the neurobehavioral basis of EtOH- and TAU-mediated responses in vertebrates.

Taurine prevents MK-801-induced shoal dispersion and altered cortisol responses in zebrafish.

Schizophrenia is a chronic neuropsychiatric disorder characterized by a shortened lifespan and significant impaired social and vocational functioning. Schizophrenic patients can present hypothalamic-pituitary-adrenal (HPA) axis dysfunctions and cortisol dysregulation, which play an important role on the etiology onset, exacerbation, and relapsing of symptoms. Based on its intrinsic neuroprotective properties, taurine is considered a promising substance with beneficial role on various brain disorders, including schizophrenia. Here, we evaluated the effects of taurine on shoaling behavior and whole-body cortisol levels in zebrafish treated with dizocilpine (MK-801), which elicits schizophrenia-like phenotypes in animal models. Briefly, zebrafish shoals (4 fish per shoal) were exposed to dechlorinated water or taurine (42, 150, or 400 mg/L) for 60 min. Then, saline (PBS, pH 7.4 or 2.0 mg/kg MK-801) were intraperitoneally injected and zebrafish behavior was recorded 15 min later. In general, MK-801 disrupted shoaling behavior and reduced whole-body cortisol levels in zebrafish. All taurine pretreatments prevented MK-801-induced increase in shoal area, while 400 mg/L taurine prevented the MK-801-induced alterations in neuroendocrine responses. Moreover, all taurine-pretreated groups showed increased geotaxis, supporting a modulatory role in the overall dispersion pattern of the shoal. Collectively, our novel findings show a potential protective effect of taurine on MK-801-induced shoal dispersion and altered neuroendocrine responses, fostering the use of zebrafish models to assess schizophrenia-like phenotypes.

Short-term high-fat diet induces cognitive decline, aggression, and anxiety-like behavior in adult zebrafish.

Obesity is a global health problem with high prevalence and defined by a high body mass index (BMI). Several comorbidities affecting the central nervous system (CNS) are associated with obesity (e.g., neurodegenerative diseases, cognitive deficit, and psychobehavioral disturbs). The zebrafish (Danio rerio) has been considered a suitable model organism to investigate the neurobehavioral features of various human diseases. Here, we verify the impact of a high-fat diet (HFD) on the CNS by specifically assessing the effects of short-term HFD on anxiety-like responses, aggression, social preference, and memory, which are essential behaviors for survival and reproduction. Animals were separated in three experimental groups. The standard diet group (SD) received 7.5 mg/fish of dry food, while HFD groups received 5 mg/fish dry food plus 7.5 (HFD-7.5) or 15 mg/fish (HFD-15) of chicken egg yolk daily. Dietary fat content (w/w) was approximately 6.5%, 16.9%, and 21.1%, respectively. We performed behavioral tests and morphometric analyses after two weeks of HFD. In comparison to SD animals, HFD groups showed typical obesogenic responses with increases in BMI, abdominal length, and body weight. HFD individuals also showed increased aggression and anxiety-like behaviors in the mirror-induced aggression and novel tank diving tests, respectively. Interestingly, HFD did not change the social preference behavior, mean swimming speed or spontaneous activity levels, while the HFD-15 group showed cognitive deficits in the inhibitory avoidance test. Collectively, this "proof-of-concept" study is the first report to characterize the effects of short-term HFD on different behavioral domains of zebrafish with high degree of face validity. Moreover, our data reinforce the growing utility of zebrafish to explore the neurobehavioral basis of obesity, providing clinically translatable data, complementing the existing rodent models and supporting future mechanistic studies.

Stress increases susceptibility to pentylenetetrazole-induced seizures in adult zebrafish.

Stress is the body's reaction to any change that requires adaptive responses. In various organisms, stress is a seizure-related comorbidity. Despite the exposure to stressors eliciting aversive behaviors in zebrafish, there are no data showing whether stress potentiates epileptic seizures in this species. Here, we investigated whether a previous exposure to an intense acute stressor positively modulates the susceptibility to seizures in pentylenetetrazole (PTZ)-challenged zebrafish. The conspecific alarm substance (CAS) was used to elicit aversive responses (3.5 mL/L for 5 min), observed by increased bottom dwelling and erratic movements. Then, fish were immediately exposed to 7.5 mM PTZ for 10 min to induce seizure-like behaviors. Stress increased the seizure intensity, the number of clonic-like seizure behaviors (score 4), as well as facilitated the occurrence of score 4 episodes by decreasing the latency in which fish reached the score 4. Moreover, fish with heightened anxiety showed increased susceptibility to PTZ, since positive correlations between anxiety- and seizure-like behaviors were found. Overall, since CAS also increased whole-body cortisol levels in zebrafish, our novel findings show a prominent response to PTZ-induced seizures in previously stressed zebrafish. Moreover, we reinforce the growing utility of zebrafish models to assess seizure-related comorbidities aiming to elucidate how stress can affect epileptic seizures in vertebrates.

Three- and bi-dimensional analyses of the shoaling behavior in zebrafish: Influence of modulators of anxiety-like responses.

Social behaviors are key components that play adaptive roles in various species, including humans. The zebrafish (Danio rerio) is a social species and the shoaling behavior can be pharmacologically manipulated either by anxiogenic or anxiolytic substances, providing translatable data in neuropsychiatric research. Here, we aimed to characterize the shoaling behavior in zebrafish under different pharmacological manipulations in a three-dimensional (3D) perspective using the spatial coordinates of the fish positions. Temporal and spatial reconstructions of shoal occupancy were performed after exposure to conspecific alarm substance (CAS) and caffeine (CAF) (anxiogenic substances) or diazepam (DZP) (a classical anxiolytic drug). Behavioral 3D analyses and spatiotemporal reconstructions of the shoaling behavior revealed that both CAS and CAF decreased the shoal volume, the average fish distance to the centoid point, and increased shoal geotaxis, but only CAS reduced the inter-fish distance when compared to control (CTRL). Conversely, DZP group showed increased shoal volume and inter-fish distance. Because substantial differences were verified when the shoaling response was analyzed in 3D and 2D perspectives, we reinforce the use of 3D reconstructions of fish positions to assess how different manipulations affect the social behavior of zebrafish. The novel procedure described here represents an easy-to-use, inexpensive, and alternative tool to perform a spatiotemporal reconstruction of the shoal occupancy under different pharmacological manipulations, complementing the existing quantification of locomotion activity of multiple fish.

Understanding the neurobiological effects of drug abuse: Lessons from zebrafish models.

Drug abuse and brain disorders related to drug comsumption are public health problems with harmful individual and social consequences. The identification of therapeutic targets and precise pharmacological treatments to these neuropsychiatric conditions associated with drug abuse are urgently needed. Understanding the link between neurobiological mechanisms and behavior is a key aspect of elucidating drug abuse-related targets. Due to various molecular, biochemical, pharmacological, and physiological features, the zebrafish (Danio rerio) has been considered a suitable vertebrate for modeling complex processes involved in drug abuse responses. In this review, we discuss how the zebrafish has been successfully used for modeling neurobehavioral phenotypes related to drug abuse and review the effects of opioids, cannabinoids, alcohol, nicotine, and psychedelic drugs on the central nervous system (CNS). Moreover, we summarize recent advances in zebrafish-based studies and outline potential advantages and limitations of the existing zebrafish models to explore the neurochemical bases of drug abuse and addiction. Finally, we discuss how the use of zebrafish models may present fruitful approaches to provide valuable clinically translatable data.

Role of the serotonergic system in ethanol-induced aggression and anxiety: A pharmacological approach using the zebrafish model.

Acute ethanol (EtOH) consumption exerts a biphasic effect on behavior and increases serotonin levels in the brain. However, the molecular mechanisms underlying alcohol-mediated behavioral responses still remain to be fully elucidated. Here, we investigate pharmacologically the involvement of the serotonergic pathway on acute EtOH-induced behavioral changes in zebrafish. We exposed zebrafish to 0.25, 0.5, 1.0% (v/v) EtOH for 1 h and analyzed the effects on aggression, anxiety-like behaviors, and locomotion. EtOH concentrations that changed behavioral responses were selected to the subsequent experiments. As a pharmacological approach, we used pCPA (inhibitor of tryptophan hydroxylase), WAY100135 (5-HT1A antagonist), buspirone (5-HT1A agonist), CGS12066A and CGS12066B (5-HT1B antagonist and agonist, respectively), ketanserin (5-HT2A antagonist) and (±)-DOI hydrochloride (5-HT2A agonist). All serotonergic receptors tested modulated aggression, with a key role of 5-HT2A in aggressive behavior following 0.25% EtOH exposure. Because CGS12066B mimicked 0.5% EtOH anxiolysis, which was antagonized by CGS12066A, we hypothesized that anxiolytic-like responses are possibly mediated by 5-HT1B receptors. Conversely, the depressant effects of EtOH are probably not related with direct changes on serotonergic pathway. Overall, our novel findings demonstrate a role of the serotonergic system in modulating the behavioral effects of EtOH in zebrafish. These data also reinforce the growing utility of zebrafish models in alcohol research and help elucidate the neurobiological mechanisms underlying alcohol abuse and associated complex behavioral phenotypes.

Concomitant taurine exposure counteracts ethanol-induced changes in locomotor and anxiety-like responses in zebrafish.

Taurine (TAU) is a ß-amino sulfonic acid with pleiotropic roles in the brain, including the neuromodulatory activity via GABAergic and glycinergic agonism. This molecule is found at high concentrations in energy drinks and is often mixed with alcohol in beverages. Although TAU has a neuroprotective role in the brain, the putative risks of mixing TAU and EtOH are not fully understood. Here, we investigated whether TAU modulates locomotor and anxiety-like behavior in adult zebrafish by using the novel tank and light-dark tests following acute EtOH exposure at anxiogenic and anxiolytic concentrations. Zebrafish were individually exposed to water (control), TAU (42, 150, and 400 mg/L), and EtOH (0.25% (v/v) and 1% (v/v)) both independently and cotreated for 1 h. EtOH 0.25% and TAU produced U-shape anxiolytic-like behavior in the light-dark test, TAU 42 and 400 positively modulated EtOH effects, and TAU 150 exerted a protective effect. All TAU concentrations counteracted EtOH 1%-induced locomotion impairment, as well as the anxiogenic-like behavior. Finally, all TAU concentrations when given independently or cotreated with EtOH 0.25% and 1% decreased the risk assessment of the lit compartment. Principal component analyses revealed that exploration and anxiety-like responses were the main behaviors that contribute to the effects of TAU and EtOH. Overall, we demonstrate that TAU differently modulates EtOH-induced anxiolytic- and anxiogenic-like behaviors depending on the concentration, suggesting a complex mechanism underlying TAU and EtOH interactions.

Modeling psychiatric comorbid symptoms of epileptic seizures in zebrafish.

Epilepsy is a debilitating neurological disorder characterized by recurrent unprovoked seizures. Anxiety, cognitive deficits, depressive-like symptoms, and social dysfunction are psychiatric comorbidities with high prevalence in epileptic patients. Due to the genetic and behavioral tractability, the zebrafish is a promising model organism to understand the neural bases involved in epilepsy-related comorbidities. Here, we aimed to characterize some behavioral phenotypes paralleling those observed in epilepsy-related comorbidities after a single pentylenetetrazole (PTZ) exposure in zebrafish. We also analyzed the influence of whole-body cortisol levels in the behavioral responses measured. Fish were exposed to 10 mM PTZ for 20 min to induce epileptic seizures. After 24 h recovery period, locomotion and anxiety-like responses (novel tank and light-dark tests), social interaction (shoaling behavior task), and memory retention (inhibitory avoidance protocol) were assessed. Basically, PTZ impaired habituation to novelty stress, evoked anxiogenic-like behaviors, disrupted shoaling, and caused memory consolidation deficits in zebrafish without changing whole-body cortisol levels. In conclusion, our novel findings further validate the use of zebrafish as a suitable tool for modeling epilepsy-related comorbidities in translational neuropsychiatric research.

Naloxone prolongs abdominal constriction writhing-like behavior in a zebrafish-based pain model.

The ability to detect noxious stimuli is essential to survival. However, pathological pain is maladaptive and severely debilitating. Endogenous and exogenous opioids modulate pain responses via opioid receptors, reducing pain sensibility. Due to the high genetic and physiological similarities to rodents and humans, the zebrafish is a valuable tool to assess pain responses and the underlying mechanisms involved in nociception. Although morphine attenuates pain-like responses of zebrafish, there are no data showing if the antagonism of opioid receptors prolongs pain duration in the absence of an exogenous opioid. Here, we investigated whether a common opioid antagonist naloxone affects the abdominal constriction writhing-like response, recently characterized as a zebrafish-based pain behavior. Animals were injected intraperitoneally with acetic acid (5.0%), naloxone (1.25 mg/kg; 2.5 mg/kg; 5.0 mg/kg) or acetic acid with naloxone to investigate the changes in their body curvature for 1 h. Acetic acid elicited a robust pain-like response in zebrafish, as assessed by aberrant abdominal body curvature, while no effects were observed following PBS injection. Although naloxone alone did not alter the frequency and duration of this behavior, it dose-dependently prolonged acetic acid-induced abdominal curvature response. Besides reinforcing the use of the abdominal writhing-like phenotype as a behavioral endpoint to measure acute pain responses in zebrafish models, our novel data suggest a putative role of endogenous opioids in modulating the recovery from pain stimulation in zebrafish.