• Energy Research

AbstractFetal 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.

Many studies regarding the effects of drugs investigate the acute and chronic use of alcohol, but only a few address the effects of caffeine and alcohol combined to the performance of the zebrafish in cognitive tasks. The zebrafish is an important model for studying the effects of drugs on learning, because it has large genetic similarities to humans and the non-invasive administration of the substances favors translational bias of research. In this study, we observed the effects of alcohol and caffeine on zebrafish cognition through an object discrimination test. We noticed that animals subjected to acute alcohol dose and those under alcohol or caffeine withdrawal did not show discrimination. When fish were treated with associated alcohol and caffeine, those chronically treated with alcohol and subjected to moderate acute dose of caffeine showed learning of the task. Our results reinforce the harmful effects of the alcohol use on cognitive tasks, and suggest that continued use of high doses of caffeine cause cognitive impairment during withdrawal of the substance. However, the acute use of caffeine appears to reverse the harmful effects of alcohol withdrawal, allowing discriminative performance equivalent to control fish. Finally, we reiterate the use of zebrafish as a model for drug effects screening and search for active compounds that modulate the alcohol and caffeine effects.

Alcohol abuse is a major medical problem. Zebrafish have been proposed to model alcohol related human disorders. Alcohol impairs learning and memory. Here, we analyze the effects of alcohol on performance of zebrafish in a recently developed latent learning paradigm. We employ a 2×3×2 experimental design (chronic×acute alcohol treatment×path blocked). The latent learning task had two phases: one, 30min long exploration trials (16 days, 1 trial/day) with left or right path of a complex maze blocked, and two, a subsequent probe trial with all paths open leading to a goal box that now contained stimulus fish. During the 16 days each fish received one of two chronic treatments: freshwater or 0.50% (v/v%) alcohol. Subsequently, fish were immersed for 1h in one of the following solutions: 0.00 (freshwater), 0.50% or 1.00% alcohol, the acute challenge. Behavior of fish was recorded during the probe trial that commenced immediately after the acute treatment. Path choices, latency to leave the start box and to enter the goal box, time spent in the goal box, distance traveled, and duration of freezing were quantified. We found that acute exposure to 1.00% alcohol after chronic freshwater disrupted learning performance, so did exposure to freshwater after chronic alcohol treatment (withdrawal). We also found exposure to chronic alcohol to diminish the effect of subsequent acute alcohol suggesting development of tolerance. Our results demonstrate that analysis of learning performance of zebrafish allows detection of alcohol-induced functional changes. The simplicity and scalability of the employed task also imply the utility of the zebrafish in high throughput drug screens.

This study aimed to test seeking behavior caused by alcohol and the drug effects on learning in the zebrafish, Danio rerio. Three treatments were conducted: acute, chronic and withdrawal, using 0.10%, 0.25%, and 1.00% alcohol and control (0.00%) (vol/vol.%). For the drug seeking behavior, we used a place preference paradigm (shuttle box tank) before and after alcohol exposure in acute (single exposure) and chronic (7 days) treatments. We observed a change in the basal preference due to the association with alcohol only for 0.25% and 1.00% doses in both acute and chronic offering, indicating an alcohol-seeking behavior after the drug exposure. For the learning task, two treatments were tested: chronic alcohol exposure (26 days including the learning period) and alcohol withdrawal (15 days of alcohol exposure before the learning period). During the learning period, fish received light stimulus followed by food in a pre-defined area of the tank for 8 consecutive days. The low dose group (0.10%) learned the task by the 3rd day both in chronic and withdrawal treatments. The higher doses (0.25% and 1.00%) caused a learning impairment in the chronic treatment group, while fish from the alcohol withdrawal treatment displayed learning on the final testing day. Therefore, we suggest that high alcohol doses impair learning and cause drug seeking behavior, even after drug exposure cessation, while low doses positively affect learning and do not cause seeking behavior. Given our results we propose that the zebrafish is a promising model for identifying active compounds, antibodies or genes which modulate the alcohol dual effects: learning improvement and reinforcing behavior.

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.

There are significant individual differences in response to alcohol: some people seem to exhibit higher alcohol sensitivity, while others are more resistant. These differences are related to alcohol metabolism, inherited traits, environmental/social pressure, personal habits and other indeterminate causes. In order to test how individual differences in hatching time are related to behavioral response to different alcohol concentrations, we separated zebrafish larvae into two categories according to egg emergence time: eggs hatched between 48 and 72 hours post-fertilization (hpf) were considered early emerging (EE), while those hatched from 72 to 96 hpf were considered late emerging (LE). On the 30th day post fertilization, EE and LE fish were exposed to four alcohol concentrations: 0.00% (control), 0.10%, 0.25% and 0.50%, and behavior was recorded for 60 min. We observed average and maximum swimming speed, distance traveled, and freezing time (immobility that indicates state of anxiety). For EE fish, 0.10% alcohol did not change behavior, while 0.25% and 0.50% increased freezing and decreased locomotion. By contrast, LE fish increased locomotion when exposed to both 0.10 and 0.25% alcohol, and increased freezing time at 0.50% alcohol. These results show that zebrafish behavioral profiles exhibit different sensitivities to alcohol, likely due to traits that can be tracked from early life stages and may indicate individuals' predisposition to alcohol tolerance and dependence.

AbstractIntroductionEmbryonic 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.MethodsThis 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.ResultsEmbryonic exposure to 0.5% and 1% ethanol presented reduced body size, decreased heartbeat rate, higher numbers of apoptotic cells, and hatching time differences.ConclusionsOur results suggest any ethanol exposure during embryogenesis can be harmful and reinforces zebrafish as a suitable model for fetal alcohol spectrum disorders (FASD).