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INTRODUCTION: Embryonic exposure to ethanol leads to a condition of physical, behavioral, and cognitive deficiencies named fetal alcohol spectrum disorders (FASD). The most severe variations are in fetal alcohol syndrome (FAS), which is easier to diagnose and not studied in animal models. On the other side, the pFAS (partial fetal alcohol syndrome) includes cases of alcohol-related congenital disabilities and neurodevelopmental disorder with an inconclusive diagnosis. In recent years, the zebrafish has become a valuable model to study FASD and its variations. METHODS: This study characterizes the zebrafish embryonic and larval development after low and moderate ethanol concentration exposure. Fish eggs were exposed to 0.0%, 0.25%, 0.5%, and 1.0% ethanol at 24 hr postfertilization, and embryonic development was observed every 8 hr up to 120 hpf. It evaluated movements, phenotypic abnormalities, hatching, cardiac function and heartbeat frequency, larvae length at 120 hpf, and the apoptotic cells' fluorescence stained with acridine orange. RESULTS: Embryonic exposure to 0.5% and 1% ethanol presented reduced body size, decreased heartbeat rate, higher numbers of apoptotic cells, and hatching time differences. CONCLUSIONS: Our results suggest any ethanol exposure during embryogenesis can be harmful and reinforces zebrafish as a suitable model for fetal alcohol spectrum disorders (FASD).

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Fetal alcohol spectrum disorder (FASD) is the most common cause of birth defects. The severe variations are in fetal alcohol syndrome (FAS) but the most frequent cases are alcohol-related neurodevelopmental disorder (ARND), which is of a difficult diagnosis. ARND characteristics include impaired social behavior, anxiety and depression prevalence, cognitive deficits, and an increased chance for drug addiction. Here, we aimed to test whether early alcohol exposure leads to later alcohol preference. We hypothesize that early alcohol exposure increases the reinforcing effects on later experiences, raising the chance of addiction in adult life. Lately, the zebrafish has been a valuable model on alcohol research, allowing embryonic exposure and the study of the ontogenetic effects. For this, embryos were exposed to three different alcohol treatments: 0.0%, 0.25% and 0.5%, for 2 hr, at 24-hr post-fertilization. Then we evaluated the effects of embryonic alcohol exposure on conditioned place preference in two developmental stage: fry (10 days post-fertilization (dpf)) and young (90 dpf) zebrafish. Results show that control fish presented alcohol associative learning, which means, changes in place preference due to alcohol exposure, at both ontogenetic phases. However, zebrafish exposed to 0.25 and 0.5% alcohol during embryogenesis did not show conditioning response at any evaluated stage. These results suggest perception and cognitive deficits due to embryonic alcohol exposure that can alter alcohol responsiveness throughout a lifetime. Although low alcohol doses do not provoke malformation, it has been shown to induce several neurological and behavioral changes that are termed as Alcohol-Related Neurodevelopmental Disorders. These results may contribute to future investigations on how embryonic exposure affects the neurocircuitry related to perception and associative learning processing.

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AIMS: Fetal alcohol spectrum disorder (FASD) is an umbrella term to describe the effects of ethanol (Eth) exposure during embryonic development, including several conditions from malformation to cognitive deficits. Zebrafish (Danio rerio) are a translational model popularly applied in brain disorders and drug screening studies due to its genetic and physiology homology to humans added to its transparent eggs and fast development. In this study, we investigated how early ethanol exposure affects zebrafish behavior during the initial growth phase. METHODS: Fish eggs were exposed to 0.0 (control), 0.25 and 0.5% ethanol at 24 h post-fertilization. Later, fry zebrafish (10 days old) were tested in a novel tank task and an inhibitory avoidance protocol to inquire about morphology and behavioral alterations. RESULTS: Analysis of variance showed that ethanol doses of 0.25 and 0.5% do not cause morphological malformations and did not impair associative learning but increased anxiety-like behavior responses and lower exploratory behavior when compared to the control. CONCLUSION: Our results demonstrate that one can detect behavioral abnormalities in the zebrafish induced by embryonic ethanol as early as 10 days post-fertilization and that alcohol increases anxious behavior during young development in zebrafish.

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Brycon amazonicus is a native Amazonian fish that is important for aquaculture in South America. Larval mortality is high in this species in intensive breeding systems due to aggressiveness among larvae. The present study investigated experimentally the effects of body size heterogeneity on the aggressive behavior and survival of B. amazonicus during the early stages of larval development. Two treatments (larvae groups with homogeneous and heterogeneous body size) were evaluated throughout early larval stages tested at six time points: 12, 24, 36, 48, 60 and 72 hours after hatching (HAH). Two experiments quantified, respectively, aggressive interactions and mortality rates among larvae at each time point. The frequency of aggressive interactions exhibited by the less aggressive larvae in each replicate was higher in the homogeneous size treatment. Aggressiveness was higher at 12 HAH, decreasing thereafter, and increasing again at 72 HAH. The mortality rate significantly increased with the larval stage, and was higher in the homogeneous than in the heterogeneous sized groups. Our results showed that aggressiveness in B. amazonicus larvae is affected by size variability and larval development stage. This knowledge about larval behavior is important to develop measures to improve larval health and survival in intensive production systems for this species.(AU)

Brycon amazonicus é um peixe nativo da Amazônia que apresenta importância para a aquicultura na América do Sul. Em sistemas de criação intensiva, a mortalidade é alta nessa espécie devido à agressividade entre as larvas. O objetivo deste estudo foi testar se o tamanho diferencial entre indivíduos afeta o comportamento agressivo e a sobrevivência de B. amazonicus durante os estágios iniciais do desenvolvimento larval. Dois tratamentos (grupos de larvas com tamanho homogêneo e heterogêneo) foram avaliados ao longo do desenvolvimento larval em seis pontos de observação: 12, 24, 36, 48, 60 e 72 horas pós-eclosão (HPE). Em dois experimentos foram quantificadas, respectivamente, as interações agressivas e a mortalidade em cada ponto de observação. A frequência das interações agressivas exibidas pelas larvas menos agressivas foi maior no tratamento de tamanho homogêneo. A agressividade também foi maior às 12 HPE, diminuiu nos períodos intermediários e aumentou novamente às 72 HPE. A taxa de mortalidade aumentou ao longo dos estágios larvais, sendo observada maior mortalidade total nos grupos de tamanho homogêneo. Os resultados deste estudo indicam que a agressividade em B. amazonicus é modulada pela variação do tamanho dos indivíduos e pelo estágio de desenvolvimento das larvas. O conhecimento sobre o comportamento de larvas é importante para desenvolver medidas que melhorem a saúde e a sobrevivência em sistemas de produção intensiva para esta espécie.(AU)

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Brycon amazonicus is a native Amazonian fish that is important for aquaculture in South America. Larval mortality is high in this species in intensive breeding systems due to aggressiveness among larvae. The present study investigated experimentally the effects of body size heterogeneity on the aggressive behavior and survival of B. amazonicus during the early stages of larval development. Two treatments (larvae groups with homogeneous and heterogeneous body size) were evaluated throughout early larval stages tested at six time points: 12, 24, 36, 48, 60 and 72 hours after hatching (HAH). Two experiments quantified, respectively, aggressive interactions and mortality rates among larvae at each time point. The frequency of aggressive interactions exhibited by the less aggressive larvae in each replicate was higher in the homogeneous size treatment. Aggressiveness was higher at 12 HAH, decreasing thereafter, and increasing again at 72 HAH. The mortality rate significantly increased with the larval stage, and was higher in the homogeneous than in the heterogeneous sized groups. Our results showed that aggressiveness in B. amazonicus larvae is affected by size variability and larval development stage. This knowledge about larval behavior is important to develop measures to improve larval health and survival in intensive production systems for this species.

Brycon amazonicus é um peixe nativo da Amazônia que apresenta importância para a aquicultura na América do Sul. Em sistemas de criação intensiva, a mortalidade é alta nessa espécie devido à agressividade entre as larvas. O objetivo deste estudo foi testar se o tamanho diferencial entre indivíduos afeta o comportamento agressivo e a sobrevivência de B. amazonicus durante os estágios iniciais do desenvolvimento larval. Dois tratamentos (grupos de larvas com tamanho homogêneo e heterogêneo) foram avaliados ao longo do desenvolvimento larval em seis pontos de observação: 12, 24, 36, 48, 60 e 72 horas pós-eclosão (HPE). Em dois experimentos foram quantificadas, respectivamente, as interações agressivas e a mortalidade em cada ponto de observação. A frequência das interações agressivas exibidas pelas larvas menos agressivas foi maior no tratamento de tamanho homogêneo. A agressividade também foi maior às 12 HPE, diminuiu nos períodos intermediários e aumentou novamente às 72 HPE. A taxa de mortalidade aumentou ao longo dos estágios larvais, sendo observada maior mortalidade total nos grupos de tamanho homogêneo. Os resultados deste estudo indicam que a agressividade em B. amazonicus é modulada pela variação do tamanho dos indivíduos e pelo estágio de desenvolvimento das larvas. O conhecimento sobre o comportamento de larvas é importante para desenvolver medidas que melhorem a saúde e a sobrevivência em sistemas de produção intensiva para esta espécie.

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Ayahuasca is a psychoactive brew traditionally used in shamanistic and vegetalistic rituals and has recently received lot of attention due to potential cognitive benefits. Ayahuasca effects are caused by the synergistic interaction of ß-carbolines (harmine, harmaline and tetrahydroarmine) contained in Banisteriopsis caapi stalks combined with the N,N-dimethyltryptamine (DMT) from Psychotria viridis leaves, a potent agonist to serotonin (5-HT) receptors. The present study approaches the effects of chronic and acute exposure to two Ayahuasca concentrations (0.1 and 0.5 ml/L) on the cognitive ability to discriminate objects in a one-trial learning task in zebrafish. Based on the combination of concentrations and exposure regimens, we divided adult zebrafish in five treatment groups: acute 0.1 and 0.5 ml/L, chronic 0.1 and 0.5 ml/L, and control 0.0 (n = 20 for each group). Then we tested them in a memory task of object discrimination. Acute Ayahuasca exposed groups performed similarly to the control group, however chronically treated fish (13 days) presented both impaired discriminative performance and locomotor alterations. Overall, these results indicate that Ayahuasca is a potent psychoactive drug that, in chronic exposure, negatively affects mnemonic parameters in zebrafish. In single exposure it does not affects cognitive performance, but the higher concentration (0.5) affected locomotion. Moreover, we reinforce the importance of the zebrafish for behavioral pharmacological studies of drug screening, in special to psychedelic drug research.

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Personality traits are related to many aspects of one's life, including risk taking, sociability, and behavioral changes caused by psychoactive substances. This study aimed to investigate individual differences and behavioral changes due to alcohol exposure in zebrafish (Danio rerio). To that end, adult animals were separated into two behavioral profiles: bold and shy, according to their risk taking behavior in an emergence test. Bold and shy fish were allowed to explore a 5-chamber tank and were tested for exploration and sociability (shoaling behavior) following alcohol exposure. The acute drug exposure treatments were 0.0%, 0.1% and 0.5% (v/v%) alcohol. The behavioral parameters evaluated were average and maximum swimming speed, total distance traveled, total time spent immobile, and time spent near a shoal or exploring the tank. For the groups that received no alcohol (0.0% alcohol), shy individuals spent more time near the shoal than bold fish. However, 0.5% alcohol increased bold fish responsiveness to the shoal, while both 0.1% and 0.5% alcohol diminished shoaling in shy fish. Our results show that the behavioral profiles of zebrafish are affected differently by alcohol, with shy animals seemingly more sensitive to behavioral change due to drug exposure. Moreover, we confirm zebrafish as a model for alcohol induced functional (exploration and social behavior) changes that could be useful in high throughput drug screens.

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The zebrafish (Danio rerio) has become a valuable model organism for behavioral studies examining learning and memory. Its diurnal circadian rhythm and characterized sleep-like state make it comparable to mammals, features that have contributed to establishing this small vertebrate as a translational model for sleep research. Despite sleep being an evolutionarily conserved behavior, its mechanisms and functions are still debated. Sleep deprivation is commonly associated with decreased attention, reduced responsiveness to external stimuli, altered locomotor activity and impaired performance on cognitive tasks. In the current study, we examined the effects of partial and total sleep deprivation on zebrafish learning performance in an active avoidance conditioning paradigm. In addition, we examined the effects of two drugs known to alter sleep (alcohol and melatonin) on learning performance in sleep deprived animals. Our results suggest that although partial sleep deprivation did not alter learning performance, total sleep deprivation was found to significantly impair behavioral responses to the electric shock as well as avoidance learning. However, when sleep deprived fish were treated with alcohol the night before the learning task, learning performance was similar to the control group. In contrast, melatonin treatment did not alter learning performance in sleep deprived animals. We conclude that the zebrafish is a sensitive tool for investigating the effects of sleep deprivation on cognitive performance and may be a useful model for dissecting the mechanisms underlying learning and memory.

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The combined infusion of Banisteriopsis caapi stem and Psychotria viridis leaves, known as ayahuasca, has been used for centuries by indigenous tribes. The infusion is rich in N, N-dimethyltryptamine (DMT) and monoamine oxidase inhibitors, with properties similar to those of serotonin. Despite substantial progress in the development of new drugs to treat anxiety and depression, current treatments have several limitations. Alternative drugs, such as ayahuasca, may shed light on these disorders. Here, we present time-course behavioral changes induced by ayahuasca in zebrafish, as first step toward establishing an ideal concentration for pre-clinical evaluations. We exposed adult zebrafish to five concentrations of the ayahuasca infusion: 0 (control), 0.1, 0.5, 1, and 3 ml/L (n = 14 each group), and behavior was recorded for 60 min. We evaluated swimming speed, distance traveled, freezing and bottom dwelling every min for 60 min. Swimming speed and distance traveled decreased with an increase in ayahuasca concentration while freezing increased with 1 and 3 ml/L. Bottom dwelling increased with 1 and 3 ml/L, but declined with 0.1 ml/L. Our data suggest that small amounts of ayahuasca do not affect locomotion and reduce anxiety-like behavior in zebrafish, while increased doses of the drug lead to crescent anxiogenic effects. We conclude that the temporal analysis of zebrafish behavior is a sensitive method for the study of ayahuasca-induced functional changes in the vertebrate brain.

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Learning and memory are vital to an animal's survival, and numerous factors can disrupt cognitive performance. Sleep is an evolutionarily conserved physiological process known to be important for the consolidation of learning and memory. The zebrafish has emerged as a powerful model organism sharing organizational and functional characteristics with other vertebrates, providing great translational relevance. In our study, we used a simple spatial associative learning task to quantify the effects of sleep deprivation (partial vs. total) on learning performance in zebrafish, using an animated conspecific shoal image as a reward. Control animals maintained on a regular light:dark cycle were able to acquire the association between the unconditioned and conditioned stimulus, reinforcing zebrafish as a valid and reliable model for appetitive conditioning tasks. Notably, sleep deprivation did not alter the perception of and response to the conspecific image. In contrast, although partial sleep deprivation did not impair cognitive performance, total sleep deprivation significantly impaired performance on the associative learning task. Our results suggest that sleep is important for learning and memory, and that the effects of sleep deprivation on these processes can be investigated in zebrafish.

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The zebrafish is an ideal vertebrate model for neurobehavioral studies with translational relevance to humans. Many aspects of sleep have been studied, but we still do not understand how and why sleep deprivation alters behavioral and physiological processes. A number of hypotheses suggest its role in memory consolidation. In this respect, the aim of this study was to analyze the effects of sleep deprivation on memory in zebrafish (Danio rerio), using an object discrimination paradigm. Four treatments were tested: control, partial sleep deprivation, total sleep deprivation by light pulses, and total sleep deprivation by extended light. The control group explored the new object more than the known object, indicating clear discrimination. The partially sleep-deprived group explored the new object more than the other object in the discrimination phase, suggesting a certain degree of discriminative performance. By contrast, both total sleep deprivation groups equally explored all objects, regardless of their novelty. It seems that only one night of sleep deprivation is enough to affect discriminative response in zebrafish, indicating its negative impact on cognitive processes. We suggest that this study could be a useful screening tool for cognitive dysfunction and a better understanding of the effect of sleep-wake cycles on cognition.

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