EXPERIMENTAL INFECTIONS WITH EUHAPLORCHIS CALIFORNIENSIS AND A SMALL CYATHOCOTYLID INCREASE CONSPICUOUS BEHAVIORS IN CALIFORNIA KILLIFISH (FUNDULUS PARVIPINNIS).

Some parasites manipulate their host's phenotype to enhance predation rates by the next host in the parasite's life cycle. Our understanding of this parasite-increased trophic transmission is often stymied by study-design challenges. A recurring difficulty has been obtaining uninfected hosts with a coevolutionary history with the parasites, and conducting experimental infections that mimic natural processes. In 1996, Lafferty and Morris provided what has become a classic example of parasite-increased trophic transmission; they reported a positive association between the intensity of a brain-infecting trematode (Euhaplorchis californiensis) in naturally infected California killifish (Fundulus parvipinnis) and the frequency of conspicuous behaviors, which was thought to explain the documented 10-30× increase in predation by the final host birds. Here, we address the primary gap in that study by using experimental infections to assess the causality of E. californiensis infection for increased conspicuous behaviors in F. parvipinnis. We hatched and reared uninfected F. parvipinnis from a population co-occurring with E. californiensis, and infected them 1-2 times/week over half their life span with E. californiensis and a small cyathocotylid trematode (SMCY) that targets the host's muscle tissue. At 3 time points throughout the hosts' lives, we quantified several conspicuous behaviors: contorting, darting, scratching, surfacing, and vertical positioning relative to the water's surface. Euhaplorchis californiensis and SMCY infection caused 1.8- and 2.5-fold overall increases in conspicuous behaviors, respectively. Each parasite was also associated with increases in specific conspicuous behaviors, particularly 1.9- and 1.4-fold more darting. These experimental findings help solidify E. californiensis-F. parvipinnis as a classic example of behavioral manipulation. Yet our findings for E. californiensis infection-induced behavioral change were less consistent and strong than those previously documented. We discuss potential explanations for this discrepancy, particularly the idea that behavioral manipulation may be most apparent when fish are actively attacked by predators. Our findings concerning the other studied trematode species, SMCY, highlight that trophically transmitted parasites infecting various host tissues are known to be associated with conspicuous behaviors, reinforcing calls for research examining how communities of trophically transmitted parasites influence host behavior.

Effects of repeated short episodes of environmental acidification on Atlantic salmon (Salmo salar) from a landlocked population.

Chronic or repeated exposure to environmental contaminants may result in allostatic overload, a physiological situation in which the costs of coping affect long-term survival and reproductive output. Continuous measurements in Otra, the largest river in southern Norway, show the occurrence of repeated 24-48 h episodes of acidification. This work investigates the impact of repeated short acidification episodes on a unique land-locked population of normally anadromous Atlantic salmon ("Bleke"). This was done by recording physiological measures of stress and allostatic load in fish exposed for 7 days to continuous or repeated episodes of simulated environmental acidification or untreated Otra water (controls). A standardized acute stress test was performed after these different exposure regimes, with brain and blood samples taken before (baseline) or after the stress test. Treatment effects on stress coping ability were assessed by neuroendocrine indicators, including telencephalic serotonergic activity and plasma cortisol. Continuous exposure to acidification resulted in increased baseline plasma Cl- and Na+ and elevated baseline plasma cortisol compared to episodic exposed fish. However, both episodic and continuous acidification resulted in similar increase in gill Al, indicating similar impact on gill permeability of these two exposures. This suggests a lower impact on the electrolyte homeostasis in episodic compared to continuous exposure and that this effect is not directly related to the effects of Al complexes binding to the gills. Furthermore, there were no treatment induced differences on stress coping ability, suggesting that episodic exposure to the sublethal concentrations of Al in pH 5.5 in the present study do not result in higher allostatic load than in control or continuous exposed Bleke.

Assaying waterborne psychoactive drugs by the response to naturalistic predator cues in the stickleback (Gasterosteus aculeatus).

Ecotoxicological effects of psychiatric drugs and drug metabolites released by the human population are of increasing environmental concern. In this study we evaluate behavioral responses to visual predator cues in wild caught three-spined stickleback (Gasterosteus aculeatus) after exposure to water-born citalopram, a widely prescribed selective serotonin reuptake inhibitor with antidepressant and anxiolytic effects. Fish were exposed to ecological relevant concentrations of citalopram (0.15 or 1.5 µg L-1) for 10 or 20 days. After drug exposure, individual fish were moved to a test arena where they were exposed to two naturalistic visual predator cues; a shadow from beneath, which simulated an approaching fish, and an overhead silhouette of a passing gull. Both visual cues resulted in decreased locomotor activity after post cue presentation. Notably, citalopram exposure resulted in a dose dependent suppression in response to the overhead stimulus. These results show that an ecologically relevant stimulus elicits a robust avoidance behavioral in wild caught fish after 25 min of acclimatization in the test arena. This suggests that the gull stimulus can be utilized as a behavioral endpoint in high flow through assays of ecotoxicological effects of psychiatric drugs and drug metabolites. Furthermore, the short acclimation time of wild caught fish in the test arena, opens for behavioral screening by fish living or kept in water bodies which are potentially impacted by psychiatric drugs.

Effects of Pseudoloma neurophilia infection on the brain transcriptome in zebrafish (Danio rerio).

Laboratory zebrafish are commonly infected with the intracellular, brain-infecting microsporidian parasite Pseudoloma neurophilia. Chronic P. neurophilia infections induce inflammation in meninges, brain and spinal cord, and have been suggested to affect neural functions since parasite clusters reside inside neurons. However, underlying neural and immunological mechanisms associated with infection have not been explored. Utilizing RNA-sequencing analysis, we found that P. neurophilia infection upregulated 175 and downregulated 45 genes in the zebrafish brain, compared to uninfected controls. Four biological pathways were enriched by the parasite, all of which were associated with immune function. In addition, 14 gene ontology (GO) terms were enriched, eight of which were associated with immune responses and five with circadian rhythm. Surprisingly, no differentially expressed genes or enriched pathways were specific for nervous system function. Upregulated immune-related genes indicate that the host generally show a pro-inflammatory immune response to infection. On the other hand, we found a general downregulation of immune response genes associated with anti-pathogen functions, suggesting an immune evasion strategy by the parasite. The results reported here provide important information on host-parasite interaction and highlight possible pathways for complex effects of parasite infections on zebrafish phenotypes.

Behavioural effects of the common brain-infecting parasite Pseudoloma neurophilia in laboratory zebrafish (Danio rerio).

Research conducted on model organisms may be biased due to undetected pathogen infections. Recently, screening studies discovered high prevalence of the microsporidium Pseudoloma neurophilia in zebrafish (Danio rerio) facilities. This spore-forming unicellular parasite aggregates in brain regions associated with motor function and anxiety, and despite its high occurrence little is known about how sub-clinical infection affects behaviour. Here, we assessed how P. neurophilia infection alters the zebrafish´s response to four commonly used neurobehavioral tests, namely: mirror biting, open field, light/dark preference and social preference, used to quantify aggression, exploration, anxiety, and sociability. Although sociability and aggression remained unaltered, infected hosts exhibited reduced activity, elevated rates of freezing behaviour, and sex-specific effects on exploration. These results indicate that caution is warranted in the interpretation of zebrafish behaviour, particularly since in most cases infection status is unknown. This highlights the importance of comprehensive monitoring procedures to detect sub-clinical infections in laboratory animals.

Brain-encysting trematodes (Euhaplorchis californiensis) decrease raphe serotonergic activity in California killifish (Fundulus parvipinnis).

Modulation of brain serotonin (5-HT) signalling is associated with parasite-induced changes in host behaviour, potentially increasing parasite transmission to predatory final hosts. Such alterations could have substantial impact on host physiology and behaviour, as 5-HT serves multiple roles in neuroendocrine regulation. These effects, however, remain insufficiently understood, as parasites have been associated with both increased and decreased serotonergic activity. Here, we investigated effects of trematode Euhaplorchis californiensis metacercariae on post-stress serotonergic activity in the intermediate host California killifish (Fundulus parvipinnis). This parasite is associated with conspicuous behaviour and increased predation of killifish by avian end-hosts, as well as inhibition of post-stress raphe 5-HT activity. Until now, laboratory studies have only been able to achieve parasite densities (parasites/unit host body mass) well below those occurring in nature. Using laboratory infections yielding ecologically relevant parasite loads, we show that serotonergic activity indeed decreased with increasing parasite density, an association likely indicating changes in 5-HT neurotransmission while available transmitter stores remain constant. Contrary to most observations in the literature, 5-HT activity increased with body mass in infected fish, indicating that relationships between parasite load and body mass may in many cases be a real underlying factor for physiological correlates of body size. Our results suggest that parasites are capable of influencing brain serotonergic activity, which could have far-reaching effects beyond the neurophysiological parameters investigated here.

Regional Distribution of a Brain-Encysting Parasite Provides Insight on Parasite-Induced Host Behavioral Manipulation.

Some parasite species alter the behavior of intermediate hosts to promote transmission to the next host in the parasite's life cycle. This is the case for Euhaplorchis californiensis, a brain-encysting trematode parasite that causes behavioral changes in the California killifish (Fundulus parvipinnis). These manipulations increase predation by the parasite's final host, piscivorous marsh birds. The mechanisms by which E. californiensis achieves this manipulation remain poorly understood. As E. californiensis cysts reside on the surface of the killifish's brain, discerning regional differences in parasite distribution could indicate mechanisms for host control. In this study, we developed a method for repeated experimental infections. In addition, we measured brain-region specific density using a novel methodology to locate and quantify parasite infection. We show that E. californiensis cysts are non-randomly distributed on the fish brain, aggregating on the diencephalon/mesencephalon region (a brain area involved in controlling reproduction and stress coping) and the rhombencephalon (an area involved in controlling locomotion and basal physiology). Determining causal mechanisms behind this pattern of localization will guide future research examining the neurological mechanisms of parasite-induced host manipulation. These findings suggest that parasites are likely targeting the reproductive, monoaminergic, and locomotor systems to achieve host behavioral manipulation.

Physiological responses during acute stress recovery depend on stress coping style in European sea bass, Dicentrarchus labrax.

Individual stress coping style (reactive, intermediate and proactive) was determined in 3 groups of 120 pit tagged European seabass using the hypoxia avoidance test. The same three groups (no change in social composition) were then reared according to the standards recommended for this species. Then, 127 days later, individuals initially characterized as reactive, intermediate or proactive were submitted to an acute confinement stress for 30 min. Blood samples were taken to measure plasma cortisol levels 30 min (Stress30) or 150 min (Stress150) after the end of the confinement stress. Individuals were then sacrificed to sample the telencephalon in order to measure the main monoamines and their catabolites (at Stress30 only). Individuals from Stress150 were sampled for whole brain for a transcriptomic analysis. The main results showed that reactive individuals had a lower body mass than intermediate individuals which did not differ from proactive individuals. The physiological cortisol response did not differ between coping style at Stress30 but at Stress150 when intermediate and proactive individuals had recovered pre stress levels, reactive individuals showed a significant higher level illustrating a modulation of stress recovery by coping style. Serotonin turnover ratio was higher in proactive and reactive individuals compared to intermediate individuals and a significant positive correlation was observed with cortisol levels whatever the coping style. Further, the confinement stress led to a general increase in the serotonin turnover comparable between coping styles. Stress150 had a significant effect on target mRNA copy number (Gapdh mRNA copy number decreased while ifrd1 mRNA copy number increased) and such changes tended to depend upon coping style.

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.

Neuroendocrine indicators of allostatic load reveal the impact of environmental acidification in fish.

When mobilized from surrounding soils and binding to gills at moderately low pH, aluminum (Al) cations can adversely affect fish populations. Furthermore, acidification may lead to allostatic overload, a situation in which the costs of coping with chronic stress affects long-term survival and reproductive output and, ultimately, ecosystem health. The brain's serotonergic system plays a key role in neuroendocrine stress responses and allostatic processes. Here, we explored whether sublethal effects of Al in acidified water affects serotonergic neurochemistry and stress coping ability in a unique land-locked salmon population from Lake Bygelandsfjorden, in southern Norway. Fish were exposed to untreated water with pH 6.5 and 74 µg Al l-1 or acidified (pH 5.5) water with different aluminum concentrations ([Al]; 74-148 µg l-1) for 5-6 days. Afterward, effects on stress coping ability were investigated by analyzing plasma cortisol levels and telencephalic serotonergic neurochemistry before and after a standardized acute stress test. Before the stress test, positive dose-response relationships existed between [Al], serotonergic turnover rate and plasma cortisol. However, in acutely stressed fish, exposure to the highest [Al] resulted in reduced cortisol values compared with those exposed to lower concentrations, while the positive dose-response relationship between Al concentrations and serotonergic turnover rate persisted in baseline conditions. This suggests that fish exposed to the highest Al concentration were unable to mount a proper cortisol response to further acute stress, demonstrating that neuroendocrine indicators of allostatic load can be used to reveal sublethal effects of water acidification-and potentially, the environmental impacts of other factors.

Future Directions for Personality Research: Contributing New Insights to the Understanding of Animal Behavior.

As part of the European Conference on Behavioral Biology 2018, we organized a symposium entitled, "Animal personality: providing new insights into behavior?" The aims of this symposium were to address current research in the personality field, spanning both behavioral ecology and psychology, to highlight the future directions for this research, and to consider whether differential approaches to studying behavior contribute something new to the understanding of animal behavior. In this paper, we discuss the study of endocrinology and ontogeny in understanding how behavioral variation is generated and maintained, despite selection pressures assumed to reduce this variation. We consider the potential mechanisms that could link certain traits to fitness outcomes through longevity and cognition. We also address the role of individual differences in stress coping, mortality, and health risk, and how the study of these relationships could be applied to improve animal welfare. From the insights provided by these topics, we assert that studying individual differences through the lens of personality has provided new directions in behavioral research, and we encourage further research in these directions, across this interdisciplinary field.

Tryptophan Metabolic Pathways and Brain Serotonergic Activity: A Comparative Review.

The essential amino acid L-tryptophan (Trp) is the precursor of the monoaminergic neurotransmitter serotonin (5-hydroxytryptamine, 5-HT). Numerous studies have shown that elevated dietary Trp has a suppressive effect on aggressive behavior and post-stress plasma cortisol concentrations in vertebrates, including teleosts. These effects are believed to be mediated by the brain serotonergic system, even though all mechanisms involved are not well understood. The rate of 5-HT biosynthesis is limited by Trp availability, but only in neurons of the hindbrain raphe area predominantly expressing the isoform TPH2 of the enzyme tryptophan hydroxylase (TPH). In the periphery as well as in brain areas expressing TPH1, 5-HT synthesis is probably not restricted by Trp availability. Moreover, there are factors affecting Trp influx to the brain. Among those are acute stress, which, in contrast to long-term stress, may result in an increase in brain Trp availability. The mechanisms behind this stress induced increase in brain Trp concentration are not fully understood but sympathetic activation is likely to play an important role. Studies in mammals show that only a minor fraction of Trp is utilized for 5-HT synthesis whereas a larger fraction of the Trp pool enters the kynurenic pathway. The first stage of this pathway is catalyzed by the hepatic enzyme tryptophan 2,3-dioxygenase (TDO) and the extrahepatic enzyme indoleamine 2,3-dioxygenase (IDO), enzymes that are induced by glucocorticoids and pro-inflammatory cytokines, respectively. Thus, chronic stress and infections can shunt available Trp toward the kynurenic pathway and thereby lower 5-HT synthesis. In accordance with this, dietary fatty acids affecting the pro-inflammatory cytokines has been suggested to affect metabolic fate of Trp. While TDO seems to be conserved by evolution in the vertebrate linage, earlier studies suggested that IDO was only present mammals. However, recent phylogenic studies show that IDO paralogues are present within the whole vertebrate linage, however, their involvement in the immune and stress reaction in teleost fishes remains to be investigated. In this review we summarize the results from previous studies on the effects of dietary Trp supplementation on behavior and neuroendocrinology, focusing on possible mechanisms involved in mediating these effects.

Proactive avoidance behaviour and pace-of-life syndrome in Atlantic salmon.

Individuals in a fish population differ in key life-history traits such as growth rate and body size. This raises the question of whether such traits cluster along a fast-slow growth continuum according to a pace-of-life syndrome (POLS). Fish species like salmonids may develop a bimodal size distribution, providing an opportunity to study the relationships between individual growth and behavioural responsiveness. Here we test whether proactive characteristics (bold behaviour coupled with low post-stress cortisol production) are related to fast growth and developmental rate in Atlantic salmon, Salmo salar. Boldness was tested in a highly controlled two-tank hypoxia test were oxygen levels were gradually decreased in one of the tanks. All fish became inactive close to the bottom at 70% oxygen saturation. At 40% oxygen saturation level a fraction of the fish actively sought to avoid hypoxia. A proactive stress coping style was verified by lower cortisol response to a standardized stressor. Two distinct clusters of bimodal growth trajectories were identified, with fast growth and early smoltification in 80% of the total population. There was a higher frequency of proactive than reactive individuals in this fast-developing fraction of fish. The smolts were associated with higher post-stress plasma cortisol than parr, and the proactive smolts leaving hypoxia had significant lower post-stress cortisol than the stayers. The study demonstrated a link between a proactive coping and fast growth and developmental ratio and suggests that selection for domestic production traits promotes this trait cluster.

Depression-like state behavioural outputs may confer beneficial outcomes in risky environments.

Recent theories in evolutionary medicine have suggested that behavioural outputs associated with depression-like states (DLS) could be an adaptation to unpredictable and precarious situations. In animal models, DLS are often linked to diverse and unpredictable stressors or adverse experiences. Theoretically, there are a range of potential fitness benefits associated with behavioural inhibition (typical to DLS), as opposed to more active/aggressive responses to adverse or uncontrollable events. This stance of evolutionary medicine has to our knowledge not been tested empirically. Here we address a possible key benefit of behavioural inhibition in a comparative model for social stress (territorial rainbow trout). By treating fish with the fast-acting antidepressant ketamine, we reversed the behavioural inhibition (i.e. stimulated an increase in activity level) in subordinate fish. During confrontation with a previously unfamiliar larger, aggressive and dominant individual, this increase in activity led to higher amounts of received aggression compared to sham-treated subordinates. This suggests that the behavioural inhibition characterizing animal models of DLS is indeed an effective coping strategy that reduces the risk of injuries in vulnerable social situations.

Contrasting Coping Styles Meet the Wall: A Dopamine Driven Dichotomy in Behavior and Cognition.

Individual variation in the ability to modify previously learned behavior is an important dimension of trait correlations referred to as coping styles, behavioral syndromes or personality. These trait clusters have been shaped by natural selection, and underlying control mechanisms are often conserved throughout vertebrate evolution. In teleost fishes, behavioral flexibility and coping style have been studied in the high (HR) and low-responsive (LR) rainbow trout lines. Generally, proactive LR trout show a behavior guided by previously learned routines, while HR trout show a more flexible behavior relying on environmental cues. In mammals, routine dependent vs. flexible behavior has been connected to variation in limbic dopamine (DA) signaling. Here, we studied the link between limbic DA signaling and individual variation in flexibility in teleost fishes by a reversal learning approach. HR/LR trout were challenged by blocking a learned escape route, previously available during interaction with a large and aggressive conspecific. LR trout performed a higher number of failed escape attempts against the transparent blockage, while HR trout were more able to inhibit the now futile escape impulse. Regionally discrete changes in DA neurochemistry were observed in micro dissected limbic areas of the telencephalon. Most notably, DA utilization in the dorsomedial telencephalon (DM, a suggested amygdala equivalent) remained stable in HR trout in response to reversal learning under acute stress, while increasing from an initially lower level in LR trout. In summary, these results support the view that limbic homologs control individual differences in behavioral flexibility even in non-mammalian vertebrates.

Uncontrollable chronic stress reduces growth disparities in farmed Atlantic salmon.

Individual variation in behavior and physiological traits in a wide variety of animals has been the focus of numerous studies in recent years. In this context, early life experiences shape responses that individuals have to subsequent environments, i.e. developmental plasticity. In this experiment, we subjected 10-month old fish to an unpredictable chronic stress (UCS) regime or no stress (control) for 3weeks. These individuals then underwent the parr-smolt transformation, when salmonids become adapted for the seawater environment, and were subsequently transferred into seawater before the final sampling. Biometric data was collected at the end of each period. Sampling on the final day was conducted in order to analyze basal monoaminergic activity in the brain stem and hypothalamus, as well as gene expression of target genes in the telencephalon. We found that post-hoc sorting of individuals by their serotonergic activity (high and low) resulted in the elucidation of growth and gene expression differences. UCS groups were found to have less growth disparities throughout the experiment, compared to control fish. Furthermore, we found brain serotonergic signaling and corticotropic releasing factor binding protein expression were positively associated with brain stem serotonergic activity, which is consistent with fish showing a stress reactivity neurophysiological profile. In conclusion, we here submit evidence that sorting individuals by their basal serotonergic activity levels may be a useful tool in the study of developmental plasticity. These results may thus apply directly to improving husbandry practices in aquaculture and elucidating neural mechanisms for coping behavior.

Dietary l-tryptophan leaves a lasting impression on the brain and the stress response.

Comparative models suggest that effects of dietary tryptophan (Trp) on brain serotonin (5-hydroxytryptamine; 5-HT) neurochemistry and stress responsiveness are present throughout the vertebrate lineage. Moreover, hypothalamic 5-HT seems to play a central role in control of the neuroendocrine stress axis in all vertebrates. Still, recent fish studies suggest long-term effects of dietary Trp on stress responsiveness, which are independent of hypothalamic 5-HT. Here, we investigated if dietary Trp treatment may result in long-lasting effects on stress responsiveness, including changes in plasma cortisol levels and 5-HT neurochemistry in the telencephalon and hypothalamus of Atlantic salmon. Fish were fed diets containing one, two or three times the Trp content in normal feed for 1 week. Subsequently, fish were reintroduced to control feed and were exposed to acute crowding stress for 1 h, 8 and 21 d post Trp treatment. Generally, acute crowding resulted in lower plasma cortisol levels in fish treated with 3×Trp compared with 1×Trp- and 2×Trp-treated fish. The same general pattern was reflected in telencephalic 5-HTergic turnover, for which 3×Trp-treated fish showed decreased values compared with 2×Trp-treated fish. These long-term effects on post-stress plasma cortisol levels and concomitant 5-HT turnover in the telencephalon lends further support to the fact that the extrahypothalamic control of the neuroendocrine stress response is conserved within the vertebrate lineage. Moreover, they indicate that trophic/structural effects in the brain underlie the effects of dietary Trp treatment on stress reactivity.

Bigger is not better: cortisol-induced cardiac growth and dysfunction in salmonids.

Stress and elevated cortisol levels are associated with pathological heart growth and cardiovascular disease in humans and other mammals. We recently established a link between heritable variation in post-stress cortisol production and cardiac growth in salmonid fish too. A conserved stimulatory effect of the otherwise catabolic steroid hormone cortisol is probably implied, but has to date not been established experimentally. Furthermore, whereas cardiac growth is associated with failure of the mammalian heart, pathological cardiac hypertrophy has not previously been described in fish. Here, we show that rainbow trout (Oncorhynchus mykiss) treated with cortisol in the diet for 45 days have enlarged hearts with lower maximum stroke volume and cardiac output. In accordance with impaired cardiac performance, overall circulatory oxygen-transporting capacity was diminished as indicated by reduced aerobic swimming performance. In contrast to the well-known adaptive/physiological heart growth observed in fish, cortisol-induced growth is maladaptive. Furthermore, the observed heart growth was associated with up-regulated signature genes of mammalian cardiac pathology, suggesting that signalling pathways mediating cortisol-induced cardiac remodelling in fish are conserved from fish to mammals. Altogether, we show that excessive cortisol can induce pathological cardiac remodelling. This is the first study to report and integrate the etiology, physiology and molecular biology of cortisol-induced pathological remodelling in fish.

Brain cortisol receptor expression differs in Arctic charr displaying opposite coping styles.

Individually consistent behavioral and physiological responses to stressful situations (often referred to as coping styles) has been reported in many animal species. Differences in hypothalamic-pituitary axis reactivity characterize individuals, and it has been proposed that the glucocorticoid (gr) and mineralocorticoid (mr) receptors are fundamental in regulating coping styles. We sorted individuals into reactive and proactive coping styles by collapsing behavioral outputs from net restraint and confinement stress tests in a principal component analysis. We then analyzed plasma cortisol levels, serotonin neurochemistry and the relative mRNA expression of gr1 and mr in stressed individuals per coping style. Proactive fish were characterized as having a lower serotonergic activity and being more active under stress. In addition, proactive fish had higher hypothalamic gr1 and mr abundance and a higher mr/gr1 ratio, compared to reactive fish. We found no significant differences in cortisol or telencephalic mRNA, gr1 and mr expression, or their ratio. Brain MR and GR have been proven to have an important role in the appraisal, coping and adaptation to stressful stimuli, so that a higher expression of these receptors in proactive fish suggests increased tolerance and performance under stress, compared to reactive individuals. We present evidence of a conserved neuroendocrine mechanism associated with coping styles in a fish species which is ecologically very diverse and considered to be the most cold-adapted fish in freshwater. We propose that this may be a first step into exploiting this model in order to better understand climate-change related effects in sub populations and ecophenotypes.

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.