Shoal Cohesion Research Articles

Oil exposure alters social group cohesion in fish

Many animal taxa live in groups to increase foraging and reproductive success and aid in predator avoidance. For fish, a large proportion of species spend all or part of their lives in groups, with group coordination playing an important role in the emergent benefits of group-living. Group cohesion can be altered by an array of factors, including exposure to toxic environmental contaminants. Oil spills are one of the most serious forms of pollution in aquatic systems, and while a range of effects of acute oil exposure on animal physiology have been demonstrated, sub-lethal effects on animal behavior are relatively under-studied. Here we used an open-field behavioral assay to explore influence of acute oil exposure on social behavior in a gregarious fish native to the Gulf of Mexico, Atlantic croaker (Micropogonias undulatus). We used two oil concentrations (0.7% and 2% oil dilution, or 6.0 ± 0.9 and 32.9 ± 5.9 μg l−1 ΣPAH50 respectively) and assays were performed when all members of a group were exposed, when only one member was exposed, and when no individuals were exposed. Shoal cohesion, as assessed via mean neighbor distance, showed significant impairment following acute exposure to 2% oil. Fish in oil-exposed groups also showed reduced voluntary movement speed. Importantly, overall group cohesion was disrupted when even one fish within a shoal was exposed to 2% oil, and the behavior of unexposed in mixed groups, in terms of movement speed and proximity to the arena wall, was affected by the presence of these exposed fish. These results demonstrate that oil exposure can have adverse effects on fish behavior that may lead to reduced ecological success.

Shoal familiarity modulates effects of individual metabolism on vulnerability to capture by trawling.

Impacts of fisheries-induced evolution may extend beyond life history traits to more cryptic aspects of biology, such as behaviour and physiology. Understanding roles of physiological traits in determining individual susceptibility to capture in fishing gears and how these mechanisms change across contexts is essential to evaluate the capacity of commercial fisheries to elicit phenotypic change in exploited populations. Previous work has shown that metabolic traits related to anaerobic swimming may determine individual susceptibility to capture in trawls, with fish exhibiting higher anaerobic performance more likely to evade capture. However, high densities of fish aggregated ahead of a trawl net may exacerbate the role of social interactions in determining an individual fish's behaviour and likelihood of capture, yet the role of social environment in modulating relationships between individual physiological traits and vulnerability to capture in trawls remains unknown. By replicating the final moments of capture in a trawl using shoals of wild minnow (Phoxinus phoxinus), we investigated the role of individual metabolic traits in determining susceptibility to capture among shoals of both familiar and unfamiliar conspecifics. We expected that increased shoal cohesion and conformity of behaviour in shoals of familiar fish would lessen the role of individual metabolic traits in determining susceptibility to capture. However, the opposite pattern was observed, with individual fish exhibiting high anaerobic capacity less vulnerable to capture in the trawl net, but only when tested alongside familiar conspecifics. This pattern is likely due to stronger cohesion within familiar shoals, where maintaining a minimal distance from conspecifics, and thus staying ahead of the net, becomes limited by individual anaerobic swim performance. In contrast, lower shoal cohesion and synchronicity of behaviours within unfamiliar shoals may exacerbate the role of stochastic processes in determining susceptibility to capture, disrupting relationships between individual metabolic traits and vulnerability to capture.

Altered non-reproductive behavior and feminization caused by developmental exposure to 17α-ethinylestradiol persist to adulthood in three-spined stickleback (Gasterosteus aculeatus)

The synthetic estrogen 17α-ethinylestradiol (EE2), ubiquitous in the aquatic environment and commonly detected in sewage effluents, interferes with the endocrine system in multiple ways. Exposure during sensitive windows of development causes persistent effects on fertility, reproductive and non-reproductive behavior in mammals and fish. In the present study, three-spined stickleback (Gasterosteus aculeatus) were exposed to nominal 0 and 20 ng/L EE2 from fertilization to 7 weeks post-hatch. After 8 months of remediation in clean water three non-reproductive behaviors, not previously analyzed in developmentally EE2-exposed progeny of wild-caught fish, were evaluated. Chemical analysis revealed that the nominal 0 and 20 ng/L exposure contained 5 and 30 ng/L EE2, respectively. Therefore, the use of control fish from previous experiments was necessary for comparisons. Fish exposed during development showed significant concentration-dependent reduction in anxiety-like behavior in the scototaxis (light/dark preference) test by means of shorter latency to first entrance to the white compartment, more visits in white, and longer total time in white compared to unexposed fish. In the novel tank test, developmental exposure significantly increased the number of transitions to the upper half of the aquaria. Exposure to EE2 during development did not alter shoal cohesion in the shoaling test compared with unexposed fish but fish exposed to 30 ng/L EE2 had significantly longer latency to leave the shoal and fewer transitions away from the shoal compared to fish exposed to 5 ng/L EE2. Skewed sex ratio with more females, sex reversal in genetic males as well as intersex in males was observed after exposure to 30, but not 5 ng/L EE2. In conclusion, EE2 exposure during development in three-spined stickleback resulted in persistent effects on anxiety-like behaviors. These long-term effects from developmental exposure are likely to be of higher relevance for natural populations than are short-term effects from adult exposure.

Effects of short-term turbidity on sensory preference and behaviour of adult fish

When changing environments make the primary sensory modality less useful or reliable, animals may shift their reliance to a different sensory modality. However, such plasticity in sensory modalities may be unlikely outside of critical developmental periods or may depend on the type of behaviour associated with the sensory response. Animals' previous experience with a particular environment and its influence on the reliability of cues may also constrain the occurrence of sensory shifts. Here, we ask about the impacts of changing environments that occur during adulthood on sensory behaviour. Specifically, we housed adult zebrafish, Danio rerio, for 6 weeks either in constantly clear water, or in water that was periodically turbid. We then compared the two groups in terms of their (1) response to a reflexive optomotor stimulus, (2) choice for visual versus olfactory cues in a foraging context and (3) social behaviour. We found that fish with experiences in the turbid context responded less robustly to a reflexive visual stimulus and responded to olfactory as opposed to visual cues as the first choice, suggesting a shift in their primary sensory modality. However, other behavioural measures (e.g. shoal cohesion, charge rate, activity level) depended more on the context in which they were measured than on recent experience. Taken together, these results suggest that recent experiences in particular physical contexts may impact the use of different sensory modalities and visual physiology, but that these effects depend on the type of behaviour being considered. These compensatory shifts between sensory modalities may help animals to buffer other forms of behaviour from major changes in the environment.

Disrupted-in-Schizophrenia-1 is essential for normal hypothalamic-pituitary-interrenal (HPI) axis function

Psychiatric disorders arise due to an interplay of genetic and environmental factors, including stress. Studies in rodents have shown that mutants for Disrupted-In-Schizophrenia-1 (DISC1), a well-accepted genetic risk factor for mental illness, display abnormal behaviours in response to stress, but the mechanisms through which DISC1 affects stress responses remain poorly understood. Using two lines of zebrafish homozygous mutant for disc1, we investigated behaviour and functioning of the hypothalamic-pituitary-interrenal (HPI) axis, the fish equivalent of the hypothalamic-pituitary-adrenal (HPA) axis. Here, we show that the role of DISC1 in stress responses is evolutionarily conserved and that DISC1 is essential for normal functioning of the HPI axis. Adult zebrafish homozygous mutant for disc1 show aberrant behavioural responses to stress. Our studies reveal that in the embryo, disc1 is expressed in neural progenitor cells of the hypothalamus, a conserved region of the vertebrate brain that centrally controls responses to environmental stressors. In disc1 mutant embryos, proliferating rx3+ hypothalamic progenitors are not maintained normally and neuronal differentiation is compromised: rx3-derived ff1b+ neurons, implicated in anxiety-related behaviours, and corticotrophin releasing hormone (crh) neurons, key regulators of the stress axis, develop abnormally, and rx3-derived pomc+ neurons are disorganised. Abnormal hypothalamic development is associated with dysfunctional behavioural and neuroendocrine stress responses. In contrast to wild type siblings, disc1 mutant larvae show altered crh levels, fail to upregulate cortisol levels when under stress and do not modulate shoal cohesion, indicative of abnormal social behaviour. These data indicate that disc1 is essential for normal development of the hypothalamus and for the correct functioning of the HPA/HPI axis.

Assessing impacts of offshore pile driving noise on the antipredator defense and shoaling behaviors of squid (Doryteuthis pealeii)

Impulsive pile driving occurs during construction of marine platforms, producing intense sounds that may adversely impact animals’ physiology and behavior. Little is known regarding how this noise impacts sound-sensitive invertebrates such as squid, which is surprising given squids’ relative abundance and key ecological role. We quantified how a commercially important squid species (Doryteuthis pealeii) behaviorally responded (in a controlled environment) to pile driving sounds recorded from an offshore windfarm installation within this species' habitat. Both sound pressure and particle motion components of the sound were quantified. Fifteen-minute portions of the recordings were played to individual squid. Body pattern changes, inking, jetting, and startle responses were observed during sound exposure and all squid exhibited at least one response. These responses occurred primarily during the first few noise impulses and diminished quickly over the first minute of playback, indicating short-term habituation. Responses returned after a 24-hr rest, indicating re-sensitization. Separate experiments investigated changes in shoaling behaviors by quantifying shoal cohesion and polarity in groups of squid during ten-minute noise exposures. Rapid habituation of antipredator alarm responses and changes in shoaling dynamics may alter squids’ susceptibility to predation. Noise exposure may also disrupt normal intraspecific communication and ecologically relevant behavioral responses to sounds.Impulsive pile driving occurs during construction of marine platforms, producing intense sounds that may adversely impact animals’ physiology and behavior. Little is known regarding how this noise impacts sound-sensitive invertebrates such as squid, which is surprising given squids’ relative abundance and key ecological role. We quantified how a commercially important squid species (Doryteuthis pealeii) behaviorally responded (in a controlled environment) to pile driving sounds recorded from an offshore windfarm installation within this species' habitat. Both sound pressure and particle motion components of the sound were quantified. Fifteen-minute portions of the recordings were played to individual squid. Body pattern changes, inking, jetting, and startle responses were observed during sound exposure and all squid exhibited at least one response. These responses occurred primarily during the first few noise impulses and diminished quickly over the first minute of playback, indicating short-term habituat...

Is Heightened-Shoaling a Good Candidate for Positive Emotional Behavior in Zebrafish?

Simple SummaryDespite increasing interest in fish welfare, we still know very little about positive emotions in fish. This study had two aims: (1) to characterize a previously undescribed social behavior in zebrafish, “heightened-shoaling”, and (2) to evaluate whether heightened-shoaling may be a good candidate for future research into positive emotional behavior in zebrafish. Observing six groups of zebrafish housed in 110 L tanks furnished with a sloping gravel substrate, rocks, and artificial plants (10 fish/tank), we found that heightened-shoaling is marked by tight group cohesion, high behavioral synchrony (all fish engaging in the same behaviors at the same time), and low agonism. Episodes of heightened-shoaling occurred in all six groups, but rarely at the same time (only once out of 31 episodes), suggesting that the onset of heightened-shoaling is driven by internal group dynamics rather than external influences. Heightened-shoaling also appeared to be self-reinforcing as it had sustained durations—typically lasting for over 7 min and sometimes lasting for nearly half-an-hour. Collectively, these results are similar to the patterns that typify positive emotional behavior in other animals, for example, social grooming and social play. As the first description of heightened-shoaling, this research extends our knowledge of the range of zebrafish social dynamics and suggests a promising area for future research on positive emotions in fish.Zebrafish, a highly-social species of freshwater fish, are widely studied across many fields of laboratory science including developmental biology, neuroscience, and genomics. Nevertheless, as standard housing for zebrafish typically consists of small and simplistic environments, less is known about their social behavioral repertoire in more naturalistic settings. In particular, spontaneously occurring, socio-positive affiliative behaviors (e.g., social coordination and cohesion) that may be indicative of positive emotional experiences have rarely been reported or studied deliberately in zebrafish. Housing adult zebrafish (10 fish/tank) in large semi-natural tanks (110 L; n = 6) with sloping gravel substrate, rocks, and artificial plants, we observed a previously undescribed behavior: Distinct periods of spontaneous, synchronized, compact aggregations, what we call “heightened-shoaling”. This project aimed to quantify the characteristics of this distinctive behavior and compare parameters of heightened-shoaling to baseline periods (normal behavior) and pre-feed periods (feed-anticipatory behavior). First, across 4 days, we selected video-clips (100 s each) from within (i) instances of heightened-shoaling (n = 9), (ii) baseline periods (n = 18), and (iii) pre-feed periods (n = 18). For each of these video clips, we scan sampled every 10 s to determine fish orientations and location within the tank and agonistic behavior. Next, we used an all-occurrence sampling method to record the timing and duration of all episodes of heightened-shoaling behavior when tank-lights were on (8:00 h to 18:00 h) across 10 days. From the scan-sampling data, we found that compared to baseline periods, heightened-shoaling was characterized by increased shoal cohesion (p < 0.0001), increased adherence to the horizontal plane (p < 0.0001), decreased agonism (p < 0.0001), and no diving behavior (lower positions within the water column signal negative effect in zebrafish, p > 0.1). From the all-occurrence sampling data, we found 31 episodes of heightened-shoaling with instances observed in all six tanks and only a single case in which heightened-shoaling occurred in two tanks at the same time. The median episode duration was 7.6 min (Range 1.3–28.6). As the first systematic description of heightened-shoaling behavior, this research contributes to our knowledge of the range of zebrafish social dynamics living in naturalistic environments. Moreover, as a spontaneously occurring, protracted, affiliative behavior, heightened-shoaling appears to be a good candidate for future research into positive emotional behavior in zebrafish.

Shoaling behaviour is differentially altered by ethanol and dopamine D1 receptor modulators in tropical marine forage fish

Anchovies are filter-feeding fish that inhabit nearshore environments worldwide. With increasing human pharmaceutical use, drugs that alter neurological functioning are becoming more prevalent in aquatic ecosystems via wastewater effluent, creating the need for tests that can reliably determine sublethal effects of these drugs on coastal fish populations. In this study, we used Caribbean anchovies (Anchoa spp.) as a tropical marine fish model to test drug-induced alterations of locomotion and shoaling behaviour with a video-based analysis system. Consistent with its anxiolytic effects in zebrafish (Danio rerio), ethanol decreased shoal cohesion in anchovies. We also characterized the effects of drugs known to modulate the dopaminergic system in zebrafish and rodents. A D1 receptor agonist (SKF 38393) and a D1 receptor antagonist (SCH 23390) increased the time anchovy spent in the center of the arena, but neither drug had an impact on shoal cohesion. Finally, the D1 receptor agonist caused significantly lower meandering compared with fish treated with the D1 receptor antagonist and ethanol. This study suggests that anchovy is a suitable Caribbean marine model for toxicology studies.

Differential behavioural responses to venlafaxine exposure route, warming and acidification in juvenile fish (Argyrosomus regius)

Antidepressants, such as venlafaxine (VFX), which are considered emerging environmental pollutants, are increasingly more present in the marine environment, and recent evidence suggest that they might have adverse effects on fish behaviour. Furthermore, altered environmental conditions associated to climate change (e.g. warming and acidification) can also have a determinant role on fish behaviour, fitness and survival. Yet, the underlying interactions between these environmental stressors (pharmaceuticals exposure and climate change) are still far from being fully understood. The aim of this study was to assess behavioural responses (in juvenile meagre (Argyrosomus regius) exposed to VFX via water ([VFX] ~20μgL−1) and via dietary sources ([VFX] ~160μgkg−1 dry weight), as well as to increased temperature (ΔT°C=+5°C) and high CO2 levels (ΔpCO2 ~1000μatm; equivalent to ΔpH=−0.4units). Overall, VFX bioaccumulation in fish plasma was enhanced under the combination of warming and acidification. VFX triggered fish exploration, whereas fish activity and shoal cohesion were reduced. Acidification alone decreased fish exploration and shoal cohesion, and reversed fish preference to turn leftwards compared to control conditions. Such alterations were further enhanced by VFX exposure. The combination of warming and acidification also reduced shoal cohesion and loss of lateralization, regardless of VFX exposure. The distinct behaviour observed when VFX contamination, acidification and warming acted alone or in combination highlighted the need to consider the likely interactive effects of seawater warming and acidification in future research regarding the toxicological aspects of chemical contaminants.

Heterospecific shoaling in an invasive poeciliid: shared history does not affect shoal cohesion

Social behaviour potentially plays an important role in invasion success. New colonists, for example, may glean useful information about predators and food by interacting with native heterospecifics. The extent to which invaders benefit from such social interactions could hinge on their prior exposure to other species. Here we asked how the shoaling decisions of the Trinidadian guppy, Poecilia reticulata, a successful invasive species, are mediated by their shared history with a heterospecific, the phenotypically similar Micropoecilia picta. To do this, we monitored shoal cohesion in single-species treatments and in treatments where M. picta was present. We predicted that shoal cohesion would be greater in single- than in mixed-species shoals. We also hypothesized that mixed-species shoals consisting of fish with a shared history would be more cohesive than those where the two species had hitherto occurred allopatrically. We found that shoal cohesion did not differ between single- and two-species treatments, or in relation to shared history with M. picta. However, while guppies were more often found in mixed-species than single-species shoals, they were more likely to have a conspecific individual as their nearest neighbour within mixed-species shoals. These results show that guppies willingly shoal with heterospecifics, even in the absence of a shared history, but also that the resulting shoals are not randomly assembled. This flexibility in shoaling may confer a crucial advantage in the initial stages of invasion.

Free-choice exploration increases affiliative behaviour in zebrafish

Cognitive stimulation has been shown to be rewarding and capable of eliciting positive emotions in several species. In contrast to the abundant learning and exploration opportunities available in nature, captive environments can be under-stimulating—with the potential to induce anhedonia and reduce welfare. Zebrafish are now a popular scientific model, in part because of their high cognitive function and sensitivity to environmental manipulations, yet little is known regarding their response to free-choice opportunities to explore. To begin to understand the effect that cognitive enrichment may have on zebrafish behaviour, we housed zebrafish within six tanks (10 fish/110 L tank) for nine months and then removed a divider to expose 10 cm of additional novel tank space. We measured free-choice exploratory behaviour (latency and number of inspections), as well as anxiety/fear (bottom-dwelling) and social behaviour (agonistic behaviour, group cohesion and coordination). We collected video data on each of four days: the day before (baseline), the day of, the day after, and two weeks after divider removal. Using multilevel models, we found that after the divider was removed, zebrafish moved into the novel space on average within 9.7 ± 7.6 s (mean ± SD), and the number of inspections increased on each day observed (p < 0.003). We found no evidence of bottom-dwelling (p > 0.73), indicating that the manipulation was not likely to have induced a negative emotional response. Furthermore, the opportunity to explore altered social behaviour: reducing agonistic behaviour (p = 0.02), and increasing shoal cohesion (p = 0.04) and coordination (p = 0.04) for up to a day relative to baseline. In light of the fact that their natural habitats would normally include such information-gain opportunities, these results indicate that free-choice exploration may be beneficial for zebrafish welfare. This study thus adds to the growing body of literature on the role cognitive stimulation plays in welfare and indicates that zebrafish are good candidates for further cognitive enrichment research.

Developmental social isolation affects adult behavior, social interaction, and dopamine metabolite levels in zebrafish.

The zebrafish is a social vertebrate and an excellent translational model for a variety of human disorders. Abnormal social behavior is a hallmark of several human brain disorders. Social behavioral problems can arise as a result of adverse early social environment. Little is known about the effects of early social isolation in adult zebrafish. We compared zebrafish that were isolated for either short (7 days) or long duration (180 days) to socially housed zebrafish, testing their behavior across ontogenesis (ages 10, 30, 60, 90, 120, 180 days), and shoal cohesion and whole-brain monoamines and their metabolites in adulthood. Long social isolation increased locomotion and decreased shoal cohesion and anxiety in the open-field in adult. Additionally, both short and long social isolation reduced dopamine metabolite levels in response to social stimuli. Thus, early social isolation has lasting effects in zebrafish, and may be employed to generate zebrafish models of human neuropsychiatric conditions.

Combinatory effects of low concentrations of 17α-etinylestradiol and citalopram on non-reproductive behavior in adult zebrafish (Danio rerio)

Sewage effluents contain pharmaceuticals, personal care products and industrial chemicals, exposing aquatic organisms to complex mixtures. The consequences of exposure to combinations of different classes of drugs in fish are largely unknown. In this study, we exposed adult zebrafish (Danio rerio) males and females for two weeks to low, environmentally relevant concentrations of the endocrine disrupting chemical 17α-etinylestradiol (EE2) and the selective serotonin re-uptake inhibitor (SSRI) citalopram, alone and in combination, and analyzed behaviors of importance for population fitness, scototaxis (light/dark preference), the novel tank test and shoal cohesion. Control water contained 0.4ng/L EE2 and the measured exposure concentrations were 0.9ng/L EE2 (nominal 0.1) and 1ng/L EE2 (nominal 0.5). The measured concentrations of citalopram were 0.1 (nominal 0.1) and 0.4μg/L (nominal 0.5). Both EE2 exposures increased anxiety in males in the scototaxis test, with significantly longer latency periods before entering and fewer visits to the white zone of the tank. The combined exposures (0.9ng/L EE2+0.1μg/L citalopram and 1ng/L EE2+0.4μg/L citalopram) resulted in abolishment of effects of EE2, with shorter latency period and more transitions to white than for fish exposed to EE2 alone. In the novel tank test, the results surprisingly indicated lower anxiety after both EE2 and citalopram exposure. Significantly more transitions to the upper half of the tank observed in males exposed to 0.1μg/L citalopram alone compared to control males. Males exposed to EE2 (0.9ng/L) had shorter latency period to the upper half. Combination exposure resulted in a longer latency and fewer transitions to the upper half compared to both control, EE2- and citalopram-exposed males. Males exposed to the combination spent significantly less time in the upper half than males EE2 or citalopram-exposed males. Females exposed to 1ng/L EE2 had fewer transitions to the upper half than the control group and females exposed to 0.4μg/L citalopram. In the shoaling test, males exposed to 0.1μg/L citalopram+0.9ng/L EE2 showed more transitions away from peers than males exposed to 0.1μg/L citalopram alone. In conclusion, low concentrations of EE2, closely above the predicted no effect concentration (NOEC) of 0.1ng/L, created anxiety-like behavior in zebrafish males. Citalopram showed marginal effects at these low concentrations but in the combination exposure the behavioral effects of EE2 were abolished. This is an initial effort to understand the effects of cocktails of anthropogenic substances contaminating aquatic environments.

Repeated ethanol exposure alters social behavior and oxidative stress parameters of zebrafish

Repeated ethanol (EtOH) consumption induces neurological disorders in humans and is considered an important public health problem. The physiological effects of EtOH are dose- and time-dependent, causing relevant changes in the social behavior. In addition, alcohol-induced oxidative stress has been proposed as a key mechanism involved in EtOH neurotoxicity. Here we investigate for the first time whether repeated EtOH exposure (REE) alters the social behavior of zebrafish and influences brain oxidation processes. Animals were exposed to water (control group) or 1% (v/v) EtOH (EtOH group) for 8 consecutive days (20min per day). EtOH was added directly to the tank water. At day 9, the social behavior and biochemical parameters were assessed. REE increased shoal cohesion by reducing inter-fish and farthest neighbor distances. SOD and CAT activities, as well as NPSH levels decreased in brain tissue. Moreover, REE increased lipid peroxidation suggesting oxidative damage. In summary, changes in oxidation processes may play a role in the CNS effects of EtOH, influencing the social behavior of zebrafish. Furthermore, in a translational neuroscience perspective, our data reinforces the utility of zebrafish to clarify the biochemical and behavioral effects of intermittent EtOH administration.

Persistent Effects of Developmental Exposure to 17α-Ethinylestradiol on the Zebrafish (Danio rerio) Brain Transcriptome and Behavior.

The synthetic estrogen 17α-ethinylestradiol (EE2) is an endocrine disrupting compound of concern due to its persistence and widespread presence in the aquatic environment. Effects of developmental exposure to low concentrations of EE2 in fish on reproduction and behavior not only persisted to adulthood, but have also been observed to be transmitted to several generations of unexposed progeny. To investigate the possible biological mechanisms of the persistent anxiogenic phenotype, we exposed zebrafish embryos for 80 days post fertilization to 0, 3, and 10 ng/L EE2 (measured concentrations 2.14 and 7.34 ng/L). After discontinued exposure, the animals were allowed to recover for 120 days in clean water. Adult males and females were later tested for changes in stress response and shoal cohesion, and whole-brain gene expression was analyzed with RNA sequencing. The results show increased anxiety in the novel tank and scototaxis tests, and increased shoal cohesion in fish exposed during development to EE2. RNA sequencing revealed 34 coding genes differentially expressed in male brains and 62 in female brains as a result of EE2 exposure. Several differences were observed between males and females in differential gene expression, with only one gene, sv2b, coding for a synaptic vesicle protein, that was affected by EE2 in both sexes. Functional analyses showed that in female brains, EE2 had significant effects on pathways connected to the circadian rhythm, cytoskeleton and motor proteins and synaptic proteins. A large number of non-coding sequences including 19 novel miRNAs were also differentially expressed in the female brain. The largest treatment effect in male brains was observed in pathways related to cholesterol biosynthesis and synaptic proteins. Circadian rhythm and cholesterol biosynthesis, previously implicated in anxiety behavior, might represent possible candidate pathways connecting the transcriptome changes to the alterations to behavior. Further the observed alteration in expression of genes involved in synaptogenesis and synaptic function may be important for the developmental modulations resulting in an anxiety phenotype. This study represents an initial survey of the fish brain transcriptome by RNA sequencing after long-term recovery from developmental exposure to an estrogenic compound.

Disrupted-in-Schizophrenia-1 is essential for normal hypothalamic-pituitary-interrenal (HPI) axis function.

Psychiatric disorders arise due to an interplay of genetic and environmental factors, including stress. Studies in rodents have shown that mutants for Disrupted-In-Schizophrenia-1 (DISC1), a well-accepted genetic risk factor for mental illness, display abnormal behaviours in response to stress, but the mechanisms through which DISC1 affects stress responses remain poorly understood. Using two lines of zebrafish homozygous mutant for disc1, we investigated behaviour and functioning of the hypothalamic-pituitary-interrenal (HPI) axis, the fish equivalent of the hypothalamic-pituitary-adrenal (HPA) axis. Here, we show that the role of DISC1 in stress responses is evolutionarily conserved and that DISC1 is essential for normal functioning of the HPI axis. Adult zebrafish homozygous mutant for disc1 show aberrant behavioural responses to stress. Our studies reveal that in the embryo, disc1 is expressed in neural progenitor cells of the hypothalamus, a conserved region of the vertebrate brain that centrally controls responses to environmental stressors. In disc1 mutant embryos, proliferating rx3+ hypothalamic progenitors are not maintained normally and neuronal differentiation is compromised: rx3-derived ff1b+ neurons, implicated in anxiety-related behaviours, and corticotrophin releasing hormone (crh) neurons, key regulators of the stress axis, develop abnormally, and rx3-derived pomc+ neurons are disorganised. Abnormal hypothalamic development is associated with dysfunctional behavioural and neuroendocrine stress responses. In contrast to wild type siblings, disc1 mutant larvae show altered crh levels, fail to upregulate cortisol levels when under stress and do not modulate shoal cohesion, indicative of abnormal social behaviour. These data indicate that disc1 is essential for normal development of the hypothalamus and for the correct functioning of the HPA/HPI axis.

Conspecific alarm substance differently alters group behavior of zebrafish populations: Putative involvement of cholinergic and purinergic signaling in anxiety- and fear-like responses

The zebrafish (Danio rerio) is an emergent model organism for assessing fear and anxiety-like phenotypes. The short fin wild type (WT), and leopard (leo) are two zebrafish populations that present several behavioral differences, in which leo displays pronounced defensive responses. Mounting evidence suggests a modulatory role for cholinergic and purinergic signaling in fear and anxiety, but the involvement of these neurotransmitter systems in the behavioral profile of zebrafish is obscure. Here we tested whether the acute exposure to conspecific alarm substance (AS), an experimental protocol that induces fear, alters shoaling behavior, diving response, acetylcholinesterase (AChE) activity, and nucleotide hydrolysis in brain tissue of WT and leo. When four fish were concomitantly exposed to AS extracted from a donor fish of similar phenotype, both populations presented a significant increase of erratic movements without changes in freezing bouts. An increased shoal cohesion and a decreased vertical distribution were observed only in WT exposed to AS. The respective population also revealed a significant increase in AChE and ecto-5′-nucleotidase activities after the exposure period. The comparison of basal endpoints between populations showed that leo displays a higher social cohesion, few vertical transitions and enhanced AChE and ecto-5′-nucleotidase activities. In conclusion, we suggest that the effects of AS on defensive behaviors depend on the population, indicating the existence of distinct neurochemical mechanisms involved. Furthermore, this report shows the first evidence of a potential role of cholinergic and purinergic systems in fear- and anxiety-like responses of zebrafish populations.

Turbidity influences individual and group level responses to predation in guppies, Poecilia reticulata

Increasing turbidity (either sedimentary or organic) from anthropogenic sources has significant negative impacts on aquatic fauna, both directly and indirectly by disrupting behaviour. In particular, antipredator responses of individuals are reduced, which has been attributed to a reduced perception of risk. Here, we explored the effect of turbidity on shoaling behaviour, which is known to carry important antipredator benefits, predicting that fish in turbid water should show reduced shoal cohesion (increased interindividual distances) and reduced responses to a simulated predatory threat. We explored both the individual and shoal level responses to a predation threat at four different levels of turbidity. At the shoal level, we found that shoals were less cohesive in more turbid water, but that there was no effect of turbidity on shoal level response to the predation threat. At an individual level, guppies in turbid water were more likely to freeze (rather than dart then freeze), and those that darted moved more slowly and over a shorter distance than those in clear water. Fish in turbid water also took longer to recover from a predation threat than fish in clear water. We suggest that because fish in turbid water behaved in a manner more similar to that expected from lone fish than to those in a shoal, the loss of visual contact between individuals in turbid water explains the change in behaviour, rather than a reduced perception of individual risk as is widely supposed. We suggest that turbidity could lead to a reduced collective response to predators and a loss of the protective benefits of shoaling.

Protective effect of rutin on impairment of cognitive functions of due to antiepileptic drugs on zebrafish model.

Aim:The severity of adverse reactions due to antiepileptics is observed during initiation and early treatment in which impairment of cognitive effects are common. Since long time, herbal medicine is a natural remedy to treat neural symptoms. Phytochemicals have been proven to be potent neuro-protective agents. Rutin, a bioflavonoid is established to be nootropic in many studies. In this study, we aimed to determine the protective effect of rutin in zebrafish against the side effects produced by AEDs.Materials and Methods:Seizures were induced in zebrafish by phenylenetetrazole. Rutin pretreatment (50 mg/kg, single injection, i.p.) was done before commencement of the study. Behavioral studies were performed as: latency to move high in the tank, locomotion effects, color effect, shoal cohesion, light/dark test on Zebrafish.Results:Treatment with rutin reverted the locomotor behavior to normal. Treatment with AEDs caused fishes to move in all regions while, in case of treatment with rutin, the response reverted to normal. Treatment with AEDs altered swimming behavior of zebrafish, however, rutin showed a positive effect over this behavior. Treatment with AEDs resulted in restricted movement of zebrafish to the dark zone. Treatment with rutin caused increased latency of zebrafish to move in the light compartment. Similarly, time spent in the light compartment by zebrafish treated with rutin was significantly (P < 0.01) higher as compared to zebrafish treated with AEDs.Conclusion:The results suggest a protective role of rutin on cognition impaired by AEDs.

Combined effects of flow condition and parasitism on shoaling behaviour of female guppies Poecilia reticulata.

Group living in fish can provide benefits of protection from predators and some parasites, more efficient foraging for food, increased mating opportunities and enhanced energetic benefit when swimming. For riverine species, shoaling behaviour can be influenced by various environmental stressors, yet little is known how flow rate might influence the shoaling of diseased fish shoals. In view of the increasingly unpredictable flow rates in streams and rivers, this study aimed to assess the combined effect of flow condition and parasitism on the shoaling behaviour of a model fish species. Shoal size, shoal cohesion and time spent shoaling of female guppies Poecilia reticulata were compared when infected with the directly transmitted ectoparasite Gyrodactylus turnbulli under flow and static conditions. Flow condition was an important factor in influencing shoaling behaviour of guppies with the fish forming larger shoals in the absence of flow. When a shoal member was infected with G. turnbulli, shoal cohesion was reduced, but the magnitude of this effect was dependent on flow condition. In both flow and static conditions, bigger fish formed larger shoals than smaller counterparts. Future changes to stream hydrology with more frequent flooding and drought events will affect the shoaling tendency of fish. During high-flow events, diseased fish may not be able to keep up with shoal mates and therefore have a higher risk of predation. Additionally, these findings may be important for aquaria and farmed species where an increase in flow rate may reduce aggregation in fish.