Obesity Impairs Cognitive Function with No Effects on Anxiety-like Behaviour in Zebrafish.

Over the last decade, the zebrafish has emerged as an important model organism for behavioural studies and neurological disorders, as well as for the study of metabolic diseases. This makes zebrafish an alternative model for studying the effects of energy disruption and nutritional quality on a wide range of behavioural aspects. Here, we used the zebrafish model to study how obesity induced by overfeeding regulates emotional and cognitive processes. Two groups of fish (n = 24 per group) were fed at 2% (CTRL) and 8% (overfeeding-induced obesity, OIO) for 8 weeks and tested for anxiety-like behaviour using the novel tank diving test (NTDT). Fish were first tested using a short-term memory test (STM) and then trained for four days for a long-term memory test (LTM). At the end of the experiment, fish were euthanised for biometric sampling, total lipid content, and triglyceride analysis. In addition, brains (eight per treatment) were dissected for HPLC determination of monoamines. Overfeeding induced faster growth and obesity, as indicated by increased total lipid content. OIO had no effect on anxiety-like behaviour. Animals were then tested for cognitive function (learning and memory) using the aversive learning test in Zantiks AD units. Results show that both OIO and CTRL animals were able to associate the aversive stimulus with the conditioned stimulus (conditioned learning), but OIO impaired STM regardless of fish sex, revealing the effects of obesity on cognitive processes in zebrafish. Obese fish did not show a deficiency in monoaminergic transmission, as revealed by quantification of total brain levels of dopamine and serotonin and their metabolites. This provides a reliable protocol for assessing the effect of metabolic disease on cognitive and behavioural function, supporting zebrafish as a model for behavioural and cognitive neuroscience.

Optimizing zebrafish rearing-Effects of fish density and environmental enrichment.

Introduction: Despite its popularity in research, there is very little scientifically validated knowledge about the best practices on zebrafish (Danio rerio) husbandry, which has led to several facilities having their own husbandry protocols. This study was performed to expand knowledge on the effects of enrichment and fish density on the welfare of zebrafish, with hopes of providing a scientific basis for future recommendations and legislations. Methods: Zebrafish were reared at three different stocking densities, (1, 3 or 6 fish/L), in tanks with or without environmental enrichment. Agonistic behavior was observed twice a week for 9 weeks directly in the housing tanks. Aspects of welfare is known to be reflected in neuroendocrine stress responses. Thus, cortisol secretion in response to lowering the water level was analyzed for each group. In addition, we assessed cortisol secretion in response to confinement and risk-taking behavior (boldness) using the novel tank diving test for individual fish. At termination of the experiment fish were subjected to stress by transfer to a novel environment and brain tissue was sampled for analysis of brain monoaminergic activity. Results: Fish kept at the lowest density (1 fish/L) showed a significantly higher level of aggression than fish kept at 3 or 6 fish/L. Moreover, fish kept at this low density showed significantly higher cortisol secretion on a group level than fish kept at the higher stocking densities, when subjected to lowering of the water level. Keeping fish at 1 fish/L also had effects on brain monoamines, these fish showing higher brain dopamine concentrations but lower dopamine turnover than fish kept at higher densities. Neither stocking density or enrichment had any clear effects on the behavior of individual fish in the novel tank diving test. However, fish kept at high densities showed lower and more variable growth rates than fish kept at 1 fish/L. Discussion: Taken together these results suggest that zebrafish should not be kept at a density of 1 fish/L. The optimal stocking density is likely to be in the range of 3-6 fish/L.

Social effects on AVT and CRF systems.

Stress and aggression have negative effects on fish welfare and productivity in aquaculture. Thus, research to understand aggression and stress in farmed fish is required. The neuropeptides arginine-vasotocin (AVT) and corticotropin-releasing factor (CRF) are involved in the control of stress and aggression. Therefore, we investigated the effect of agonistic interactions on the gene expression of AVT, CRF and their receptors in juvenile rainbow trout (Oncorhynchus mykiss). The social interactions lead to a clear dominant-subordinate relationship with dominant fish feeding more and being more aggressive. Subordinate fish had an upregulation of the AVT receptor (AVT-R), an upregulation of CRF mRNA levels, and higher plasma cortisol levels. The attenuating effect of AVT on aggression in rainbow trout is proposed to be mediated by AVT-R, and the attenuating effect of the CRF system is proposed to be mediated by CRF.

Visualization of early oligomeric α-synuclein pathology and its impact on the dopaminergic system in the (Thy-1)-h[A30P]α-syn transgenic mouse model.

Aggregation of alpha-synuclein (α-syn) into Lewy bodies and Lewy neurites is a pathological hallmark in the Parkinson´s disease (PD) brain. The formation of α-syn oligomers is believed to be an early pathogenic event and the A30P mutation in the gene encoding α-syn, causing familial PD, has been shown to cause an accelerated oligomerization. Due to the problem of preserving antigen conformation on tissue surfaces, α-syn oligomers are difficult to detect ex vivo using conventional immunohistochemistry with oligomer-selective antibodies. Herein, we have instead employed the previously reported α-syn oligomer proximity ligation assay (ASO-PLA), along with a wide variety of biochemical assays, to discern the pathological progression of α-syn oligomers and their impact on the dopaminergic system in male and female (Thy-1)-h[A30P]α-syn transgenic (A30P-tg) mice. Our results reveal a previously undetected abundance of α-syn oligomers in midbrain of young mice, whereas phosphorylated (pS129) and proteinase k-resistant α-syn species were observed to a larger extent in aged mice. Although we did not detect loss of dopaminergic neurons in A30P-tg mice, a dysregulation in the monoaminergic system was recorded in older mice. Taken together, ASO-PLA should be a useful method for the detection of early changes in α-syn aggregation on brain tissue, from experimental mouse models in addition to post mortem PD cases.

Chronic Exposure to Oxazepam Pollution Produces Tolerance to Anxiolytic Effects in Zebrafish (Danio rerio).

Environmental concentrations of the anxiolytic drug oxazepam have been found to disrupt antipredator behaviors of wild fish. Most experiments exposed fish for a week, while evidence from mammals suggests that chronic exposure to therapeutic concentrations of benzodiazepines (such as oxazepam) results in the development of tolerance to the anxiolytic effects. If tolerance can also develop in response to the low concentrations found in the aquatic environment, it could mitigate the negative effects of oxazepam pollution. In the current study, we exposed wild-caught zebrafish to oxazepam (∼7 µg L-1) for 7 or 28 days and evaluated behavioral and physiological parameters at both time points. Females showed reduced diving responses to conspecific alarm pheromone after 7 days, but not after 28 days, indicating that they had developed tolerance to the anxiolytic effects of the drug. Zebrafish males were not affected by this oxazepam concentration, in line with earlier results. Serotonin turnover (ratio 5-HIAA/5-HT) was reduced in exposed females and males after 28 days, indicating that brain neurochemistry had not normalized. Post-confinement cortisol concentrations and gene expression of corticotropin-releasing hormone (CRH) were not affected by oxazepam. We did not find evidence that chronically exposed fish had altered relative expression of GABAA receptor subunits, suggesting that some other still unknown mechanism caused the developed tolerance.

Low concentrations of the benzodiazepine drug oxazepam induce anxiolytic effects in wild-caught but not in laboratory zebrafish.

Pollution by psychoactive pharmaceuticals has been found to disrupt anti-predator behaviors of wild fish. The challenge is now to identify which of the many psychoactive drugs pose the greatest threat. One strategy is to screen for behavioral effects of selected pharmaceuticals using a single, widely available fish species such as zebrafish. Here, we show that although such high-throughput behavioral screening might facilitate comparisons between pharmaceuticals, the choice of strain is essential. While wild-caught zebrafish exposed to concentrations of the anxiolytic drug oxazepam as low as 0.57 µg L-1 showed a reduction in the response to conspecific alarm pheromone, laboratory strain AB did not respond to the alarm cue, and consequently, the anxiolytic effect of oxazepam could not be measured. Adaptation to the laboratory environment may have rendered laboratory strains unfit for use in some ecotoxicological and pharmacological studies, since the results might not translate to wild fish populations.

Contrasting neurochemical and behavioral profiles reflects stress coping styles but not stress responsiveness in farmed gilthead seabream (Sparus aurata).

In fish, as well as in other vertebrates, contrasting suites of physiological and behavioral traits, or coping styles, are often shown in response to stressors. However, the magnitude of the response (i.e. stress responsiveness) has been suggested to be independent of stress coping style. One central neurotransmitter that has been associated with both stress responsiveness and differences in stress coping styles is serotonin (5-hydroxytryptamine, 5-HT). In this study, we investigated to what extent stress responsiveness reflects differences in stress coping, and the potential involvement of the 5-HT system in mediating such differences in farmed Gilthead seabream. Initially, fish were classified as proactive or reactive based on their behavioural response to net restraint. Following 1.5 months, fish classified as proactive still showed a higher number of escape attempts and spent longer time escaping than those classified as reactive. These differences were reflected in a generally higher brain stem 5-HT concentration and a lower telencephalic 5-HT activity, i.e. the ratio of 5-hydroxyindoleacetic acid (5-HIAA) to 5-HT, in proactive fish. Independent of stress coping styles, stress responsiveness was reflected in elevated 5-HIAA concentrations and 5-HIAA/5-HT ratios in telencephalon and brain stem together with increased plasma cortisol concentrations at 0.5 and 2 h following the last net restraint. The current results show that 5-HT signaling can reflect different behavioural output to a challenge which are independent of neuroendocrine responses to stress and lend support to the hypothesis that stress coping styles can be independent of stress responsiveness.

Effects of early rearing enrichments on modulation of brain monoamines and hypothalamic-pituitary-interrenal axis (HPI axis) of fish mahseer (Tor putitora).

Enriching rearing environment is the strategy suggested for improving the post release survivorship of captive-reared animals. Here, an attempt has been made to evaluate the impact of early rearing enrichment on the hypothalamic-pituitary-interrenal axis (HPI axis), blood glucose, and brain dopaminergic and serotonergic systems of Tor putitora. Fifteen-day-old hatchlings of T. putitora were reared up to advanced fry stage in barren, semi-natural, and physically enriched environments and compared them with regard to pre-stress and post-stress levels of whole-body cortisol, blood glucose, brain serotonergic activity (5HIAA/5HT ratio), dopaminergic activity (DOPAC/DA and HVA/DA ratios) and norepinephrine (NE) levels. Significantly low basal whole-body cortisol, glucose and brain NE levels were observed in a physically enriched group of fish as compared to the other two groups. However, after acute stress, all rearing groups showed elevated levels of cortisol, blood glucose, brain 5HIAA/5HT, DOPAC/DA and HVA/DA ratios and NE levels but the magnitude of response was different among different rearing groups. The barren reared group showed a higher magnitude of response as compared to semi-natural and physically enriched groups. Similarly, the recovery rate of whole-body cortisol, blood glucose, and whole-brain monoamines were long-lasting in barren-reared mahseer. We illustrate that increased structural complexity (physical enrichment) during the early rearing significantly modulates various physiological and stress-coping mechanisms of mahseer.

Neurobiological and behavioural responses of cleaning mutualisms to ocean warming and acidification.

Cleaning interactions are textbook examples of mutualisms. On coral reefs, most fishes engage in cooperative interactions with cleaners fishes, where they benefit from ectoparasite reduction and ultimately stress relief. Furthermore, such interactions elicit beneficial effects on clients' ecophysiology. However, the potential effects of future ocean warming (OW) and acidification (OA) on these charismatic associations are unknown. Here we show that a 45-day acclimation period to OW (+3 °C) and OA (980 µatm pCO2) decreased interactions between cleaner wrasses (Labroides dimidiatus) and clients (Naso elegans). Cleaners also invested more in the interactions by providing tactile stimulation under OA. Although this form of investment is typically used by cleaners to prolong interactions and reconcile after cheating, interaction time and client jolt rate (a correlate of dishonesty) were not affected by any stressor. In both partners, the dopaminergic (in all brain regions) and serotoninergic (forebrain) systems were significantly altered by these stressors. On the other hand, in cleaners, the interaction with warming ameliorated dopaminergic and serotonergic responses to OA. Dopamine and serotonin correlated positively with motivation to interact and cleaners interaction investment (tactile stimulation). We advocate that such neurobiological changes associated with cleaning behaviour may affect the maintenance of community structures on coral reefs.

The aggressive spiegeldanio, carrying a mutation in the fgfr1a gene, has no advantage in dyadic fights with zebrafish of the AB strain.

Zebrafish which carries a mutation in the fibroblast growth factor receptor 1A (fgfr1a), also known as spiegeldanio (spd), has previously been reported to be bolder and more aggressive than wildtype (AB) zebrafish. However, in previous studies aggression has been quantified in mirror tests. In dyadic fights the behavior of the combatants is modified by the behavior of their opponent, and fighting a mirror has been reported to have different effects on brain gene expression and brain monoaminergic systems. In the present study aggression was quantified in fgfr1a mutants and AB zebrafish using a mirror test after which the fish were allowed to interact in pairs, either consisting of two fgfr1a mutants or one AB and one fgfr1a mutant fish. Following dyadic interaction aggressive behavior was again quantified in individual fish in a second mirror test after which the fish were sacrificed and brain tissue analyzed for monoamines and monoamine metabolites. The results confirm that fgfr1a mutants are more aggressive than AB zebrafish in mirror tests. However, fgfr1a mutant fish did not have any advantage in fights for social dominance, and agonistic behavior of fgfr1a mutants did not differ from that of AB fish during dyadic interactions. Moreover, as the AB fish, fgfr1a mutant fish losing dyadic interactions showed a typical loser effect and social subordination resulted in an activation of the brain serotonergic system in fgfr1a mutants as well as in AB fish. Overall the effects of dyadic interaction were similar in fgfr1a mutant fish and zebrafish of the AB strain.

Bold zebrafish (Danio rerio) express higher levels of delta opioid and dopamine D2 receptors in the brain compared to shy fish.

Individual variation in coping with environmental challenges is a well-known phenomenon across vertebrates, including teleost fish. Dopamine is the major transmitter in the brain reward networks, and important for motivational processes and stress coping. Functions of the endogenous opioid system are not well studied in teleosts. However, in mammals the activity in the brain reward networks is regulated by the endogenous opioid system. This study aimed at investigating if there was a correlation between risk-taking behavior and the expression of dopamine and opioid receptors in the zebrafish (Danio rerio) brain. Risk-taking behavior was assessed in a novel tank diving test, and the most extreme high risk taking, i.e. bold, and low risk taking, i.e. shy, fish were sampled for qPCR analysis of whole brain gene expression. The expression analysis showed a significantly higher expression of the dopamine D2 receptors (drd2a and drd2b) and the delta opioid receptor (DOR; oprd1b) in bold compared to shy fish. Besides reward and reinforcing properties, DORs are also involved in emotional responses. Dopamine D2 receptors are believed to be important for active stress coping in rodents, and taken together the results of the current study suggest similar functions in zebrafish. However, additional experiments are required to clarify how dopamine and opioid receptor activation affect behavior and stress coping in this species.

The variable monoaminergic outcomes of cleaner fish brains when facing different social and mutualistic contexts.

The monoamines serotonin and dopamine are important neuromodulators present in the central nervous system, known to be active regulators of social behaviour in fish as in other vertebrates. Our aim was to investigate the region-specific brain monoaminergic differences arising when individual cleaners face a client (mutualistic context) compared to when they are introduced to another conspecific (conspecific context), and to understand the relevance of visual assessment compared to the impact of physical contact with any partner. We demonstrated that serotoninergic activity at the diencephalon responds mostly to the absence of physical contact with clients whereas cerebellar dopaminergic activity responds to actual cleaning engagement. We provide first insights on the brain's monoaminergic (region-specific) response variations, involved in the expression of cleaner fishes' mutualistic and conspecific behaviour. These results contribute to a better understanding of the monoaminergic activity in accordance to different socio-behavioural contexts.

Monoaminergic levels at the forebrain and diencephalon signal for the occurrence of mutualistic and conspecific engagement in client reef fish.

Social interactions are commonly found among fish as in mammals and birds. While most animals interact socially with conspecifics some however are also frequently and repeatedly observed to interact with other species (i.e. mutualistic interactions). This is the case of the (so-called) fish clients that seek to be cleaned by other fish (the cleaners). Clients face an interesting challenge: they raise enough motivation to suspend their daily activities as to selectively visit and engage in interactions with cleaners. Here we aimed, for the first time, to investigate the region-specific brain monoaminergic level differences arising from individual client fish when facing a cleaner (interspecific context) compared to those introduced to another conspecific (socio-conspecific context). We show that monoaminergic activity differences occurring at two main brain regions, the diencephalon and the forebrain, are associated with fish clients' social and mutualistic activities. Our results are the first demonstration that monoaminergic mechanisms underlie client fish mutualistic engagement with cleanerfish. These pathways should function as a pre-requisite for cleaning to occur, providing to clients the cognitive and physiological tools to seek to be cleaned.

The brain-gut axis of fish: Rainbow trout with low and high cortisol response show innate differences in intestinal integrity and brain gene expression.

In fish, the stress hormone cortisol is released through the action of the hypothalamic pituitary interrenal axis (HPI-axis). The reactivity of this axis differs between individuals and previous studies have linked this to different behavioural characteristics and stress coping styles. In the current study, low and high responding (LR and HR) rainbow trout in terms of cortisol release during stress were identified, using a repeated confinements stress test. The expression of stress related genes in the forebrain and the integrity of the stress sensitive primary barrier of the intestine was examined. The HR trout displayed higher expression levels of mineralocorticoid and serotonergic receptors and serotonergic re-uptake pumps in the telencephalon during both basal and stressed conditions. This confirms that HPI-axis reactivity is linked also to other neuronal behavioural modulators, as both the serotonergic and the corticoid system in the telencephalon are involved in behavioural reactivity and cognitive processes. Involvement of the HPI-axis in the brain-gut-axis was also found. LR trout displayed a lower integrity in the primary barrier of the intestine during basal conditions compared to the HR trout. However, following stress exposure, LR trout showed an unexpected increase in intestinal integrity whereas the HR trout instead suffered a reduction. This could make the LR individuals more susceptible to pathogens during basal conditions where instead HR individuals would be more vulnerable during stressed conditions. We hypothesize that these barrier differences are caused by regulation/effects on tight junction proteins possibly controlled by secondary effects of cortisol on the intestinal immune barrier or differences in parasympathetic reactivity.

High risk no gain-metabolic performance of hatchery reared Atlantic salmon smolts, effects of nest emergence time, hypoxia avoidance behaviour and size.

When animals are reared for conservational releases it is paramount to avoid reducing genetic and phenotypic variation over time. This requires an understanding of how diverging behavioural and physiological traits affect performance both in captivity and after release. In Atlantic salmon, emergence time from the spawning gravel has been linked to certain behavioural and physiological characteristics and to the concept of stress coping styles. Early emerging fry has for example been shown to be bolder and more aggressive and to have higher standard metabolic rates compared to late emerging fry. The first aim was therefore to examine if emergence latency affect the behavioural stress coping response also beyond the fry and parr stage. This was done using a hypoxia avoidance test, where an active behavioural avoidance response can be related to higher risk taking. No behavioural differences were found between the two emergence fractions either at the parr or pre-smolt stage, instead smaller individuals were more prone to express an "active" hypoxia avoidance response. Further, an individual expressing a "passive" response as parr were also more prone to express this behaviour at the pre-smolt stage. While there are some previous studies showing that early emerging individuals with a bolder personality may be favored within a hatchery setting it is not known to what extent these early differences persist to affect performance after release. The second aim was therefore to compare the physiological performance at the time of release as smolts using the two subgroups; 1) early emerging fish showing active hypoxia avoidance (Early+Bold) and 2) late emerging fish showing a passive hypoxia response (Late+Shy). The Early+Bold group showed a higher red blood cell swelling, suggesting a higher adrenergic output during stress, whereas there was no difference in post-stress plasma cortisol or physiological smolt status. While there was no difference in standard metabolic rate between the groups, the Early+Bold group exhibited a lower maximum metabolic rate and aerobic scope following strenuous swimming. In captivity this may have no clear negative effects, but in the wild, a more risk prone behavioural profile linked to a lower aerobic capacity to escape from e.g. a predator attack, could clearly be disadvantageous.

How do individuals cope with stress? Behavioural, physiological and neuronal differences between proactive and reactive coping styles in fish.

Despite the use of fish models to study human mental disorders and dysfunctions, knowledge of regional telencephalic responses in non-mammalian vertebrates expressing alternative stress coping styles is poor. As perception of salient stimuli associated with stress coping in mammals is mainly under forebrain limbic control, we tested region-specific forebrain neural (i.e. mRNA abundance and monoamine neurochemistry) and endocrine responses under basal and acute stress conditions for previously characterised proactive and reactive Atlantic salmon. Reactive fish showed a higher degree of the neurogenesis marker proliferating cell nuclear antigen (pcna) and dopamine activity under basal conditions in the proposed hippocampus homologue (Dl) and higher post-stress plasma cortisol levels. Proactive fish displayed higher post-stress serotonergic signalling (i.e. higher serotonergic activity and expression of the 5-HT1A receptor) in the proposed amygdala homologue (Dm), increased expression of the neuroplasticity marker brain-derived neurotropic factor (bdnf) in both Dl and the lateral septum homologue (Vv), as well as increased expression of the corticotropin releasing factor 1 (crf1 ) receptor in the Dl, in line with active coping neuro-profiles reported in the mammalian literature. We present novel evidence of proposed functional equivalences in the fish forebrain with mammalian limbic structures.

Characterization of the γ-aminobutyric acid signaling system in the zebrafish (Danio rerio Hamilton) central nervous system by reverse transcription-quantitative polymerase chain reaction.

In the vertebrate brain, inhibition is largely mediated by γ-aminobutyric acid (GABA). This neurotransmitter comprises a signaling machinery of GABAA, GABAB receptors, transporters, glutamate decarboxylases (gads) and 4-aminobutyrate aminotransferase (abat), and associated proteins. Chloride is intimately related to GABAA receptor conductance, GABA uptake, and GADs activity. The response of target neurons to GABA stimuli is shaped by chloride-cation co-transporters (CCCs), which strictly control Cl- gradient across plasma membranes. This research profiled the expression of forty genes involved in GABA signaling in the zebrafish (Danio rerio) brain, grouped brain regions and retinas. Primer pairs were developed for reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The mRNA levels of the zebrafish GABA system share similarities with that of mammals, and confirm previous studies in non-mammalian species. Proposed GABAA receptors are α1ß2γ2, α1ß2δ, α2bß3γ2, α2bß3δ, α4ß2γ2, α4ß2δ, α6bß2γ2 and α6bß2δ. Regional brain differences were documented. Retinal hetero- or homomeric ρ-composed GABAA receptors could exist, accompanying α1ßyγ2, α1ßyδ, α6aßyγ2, α6aßyδ. Expression patterns of α6a and α6b were opposite, with the former being more abundant in retinas, the latter in brains. Given the stoichiometry α6wßyγz, α6a- or α6b-containing receptors likely have different regulatory mechanisms. Different gene isoforms could originate after the rounds of genome duplication during teleost evolution. This research depicts that one isoform is generally more abundantly expressed than the other. Such observations also apply to GABAB receptors, GABA transporters, GABA-related enzymes, CCCs and GABAA receptor-associated proteins, whose presence further strengthens the proof of a GABA system in zebrafish.

Effects of acute and chronic stress on telencephalic neurochemistry and gene expression in rainbow trout (Oncorhynchus mykiss).

By filtering relevant sensory inputs and initiating stress responses, the brain is an essential organ in stress coping and adaptation. However, exposure to chronic or repeated stress can lead to allostatic overload, where neuroendocrinal and behavioral reactions to stress become maladaptive. This work examines forebrain mechanisms involved in allostatic processes in teleost fishes. Plasma cortisol, forebrain serotonergic (5-HTergic) neurochemistry, and mRNA levels of corticotropin-releasing factor (CRF), CRF-binding protein (CRF-BP), CRF receptors (CRFR1 and CRFR2), mineralocorticoid receptor (MR), glucocorticoid receptors (GR1 and GR2) and serotonin type 1A (5-HT1A) receptors (5-HT1Aα and 5-HT1Aß) were investigated at 1 h before and 0, 1 and 4 h after acute stress, in two groups of rainbow trout held in densities of 25 and 140 kg m-3 for 28 days. Generally, being held at 140 kg m-3 resulted in a less pronounced cortisol response. This effect was also reflected in lower forebrain 5-HTergic turnover, but not in mRNA levels in any of the investigated genes. This lends further support to reports that allostatic load causes fish to be incapable of mounting a proper cortisol response to an acute stressor, and suggests that changes in forebrain 5-HT metabolism are involved in allostatic processes in fish. Independent of rearing densities, mRNA levels of 5-HT1Aα and MR were downregulated 4 h post-stress compared with values 1 h post-stress, suggesting that these receptors are under feedback control and take part in the downregulation of the hypothalamic-pituitary-interrenal (HPI) axis after exposure to an acute stressor.

Role of brain serotonin in modulating fish behavior.

The organization of the brain serotonergic system appears to have been highly conserved across the vertebrate subphylum. In fish as well as in other vertebrates, brain serotonin (5-HT), mainly acts as a neuromodulator with complex effects on multiple functions. It is becoming increasingly clear that acute and chronic increase in brain 5-HT neurotransmission have very different effects. An acute 5-HT activation, which is seen in both winners and losers of agonistic interactions, could be related to a general arousal effect, whereas the chronic activation observed in subordinate fish is clearly linked to the behavioral inhibition displayed by these individuals. Fish displaying divergent stress coping styles (proactive vs. reactive) differ in 5-HT functions. In teleost fish, brain monoaminergic function is also related to life history traits.

Zebrafish (Danio rerio) behaviour is largely unaffected by elevated pCO2.

Ocean acidification, the decrease in ocean pH caused by anthropogenic emission of carbon dioxide, can cause behavioural disturbances in marine teleost species. We investigated whether AB-strain zebrafish (Danio rerio) show similar behavioural disturbances in the presence of elevated CO2, because this model species could open up a toolbox to investigate the physiological and neurological mechanisms of CO2 exposure. We found no effect of elevated CO2 (~1600 µatm) on the behaviour of zebrafish in the open field test, indicating that zebrafish are largely insensitive to this elevated CO2 level. In the detour test of lateralization, however, zebrafish exposed to elevated CO2 swam more often to the right, whereas individuals exposed to control CO2 (~400 µatm) had no preference for left or right. This may indicate that some behaviours of some freshwater fishes can be altered by elevated CO2 levels. Given that elevated CO2 levels often occur in recirculating aquaculture and aquarium systems, we recommend that dissolved CO2 levels are measured and, if necessary, the aquarium water should be aerated, in order to exclude CO2 level as a confounding factor in experiments.