Joachim Schjolden

Does Individual Variation in Stress Responses and Agonistic Behavior Reflect Divergent Stress Coping Strategies in Juvenile Rainbow Trout

Individual rainbow trout were transferred to visual isolation in experimental aquaria. As a measure of the speed of acclimation, individual food intake was quantified during the first 6 d following transfer. Following acclimation, aggression was quantified by subjecting the fish to three resident‐intruder tests, with 30 d of recovery between the tests. Moreover, between the resident‐intruder tests (i.e., two times) the fish were exposed to an unfamiliar environment and their cortisol response was measured. The results of this study show that individuals of juvenile rainbow trout differ distinctly in their response to changes in their environment, and that this diversity in behavior is reflected by consistent behavioral traits displayed by individual fish. These traits have proven to be consistent not only over time but also across situations, revealing two distinct behavioral profiles, in the same manner as shown in studies on proactive and reactive mammals. Our results also show that the reactivity of the hypothalamic‐pituitary‐interrenal (HPI) axis, when exposed to a stressor, is a consistent physiological trait in juvenile rainbow trout. We found that difference in HPI axis reactivity is linked to the different behavioral profiles. However, HPI axis reactivity could not be linked directly to the singular behavioral traits measured. In other words, we did not find that the consistent behavioral traits shown by the fish were associated with a difference in HPI axis reactivity in the same manner as the reactivity of the hypothalamic‐pituitary‐adrenocortical axis does in mammals. Taken together, our results show that stress coping strategies akin to what has been described as reactive and proactive stress coping in mammals appear to exist in juvenile rainbow trout.

Divergence in behavioural responses to stress in two strains of rainbow trout (Oncorhynchus mykiss) with contrasting stress responsiveness

The aim of this study was to establish whether two lines of rainbow trout divergent for their plasma cortisol response to a standardized stressor would show consistent differences in their behavioural response to a range of challenging situations. Our results show that the high- and low-responding (HR and LR) lines of rainbow trout did not differ in the aggression shown towards an intruder or in their response to the introduction of a novel object to their home environment. However, there was a difference in behaviour between the two selection lines when they were exposed to two unfamiliar environments. These results suggest that the behaviour of the HR and LR fish differs when they are challenged in unfamiliar environments, while their behaviour does not differ when they are challenged in their home environment. These observations are in agreement with studies on mammals that show that individuals with reactive coping styles perform similarly to proactive animals when they are challenged in a familiar environment, while they show different behaviour when they are challenged in unfamiliar environments. Thus, these results provide further evidence that the HR and LR selection lines of rainbow trout exemplify the two different coping styles described in mammals.

Melanin-based skin spots reflect stress responsiveness in salmonid fish

Within animal populations, genetic, epigenetic and environmental factors interact to shape individual neuroendocrine and behavioural profiles, conferring variable vulnerability to stress and disease. It remains debated how alternative behavioural syndromes and stress coping styles evolve and are maintained by natural selection. Here we show that individual variation in stress responsiveness is reflected in the visual appearance of two species of teleost fish; rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar). Salmon and trout skin vary from nearly immaculate to densely spotted, with black spots formed by eumelanin-producing chromatophores. In rainbow trout, selection for divergent hypothalamus-pituitary-interrenal responsiveness has led to a change in dermal pigmentation patterns, with low cortisol-responsive fish being consistently more spotted. In an aquaculture population of Atlantic salmon individuals with more spots showed a reduced physiological and behavioural response to stress. Taken together, these data demonstrate a heritable behavioural-physiological and morphological trait correlation that may be specific to alternative coping styles. This observation may illuminate the evolution of contrasting coping styles and behavioural syndromes, as occurrence of phenotypes in different environments and their response to selective pressures can be precisely and easily recorded.

Genetically Determined Variation in Stress Responsiveness in Rainbow Trout: Behavior and Neurobiology

It is becoming increasingly recognized that the diversity in stressors, their intensity, predictability and the context in which they are experienced, will result in behavioral and physiological responses just as diverse. In addition, stress responses are characterized by individual variations where the physiological and behavioral reactions are associated in such a manner that distinct stress coping styles encompassing suites of correlated traits can be identified. These are often referred to as proactive and reactive stress coping styles. Proactive coping is characterized by more aggression, higher general activity and higher sympathetic activation, whereas reactive coping is characterized by immobility, lack of initiative and a higher parasympathetic/hypothalamic activation. Stable coping styles appear to coexist within populations, and these strategies appear to be largely innate. Moreover, the physiological and behavioral traits of coping styles appear to be heritable. These stress coping styles have proven to play a major role in competitive ability and subsequent social position in different species of vertebrates. However, there are also studies showing that social position can affect parameters encompassing the stress coping style of individuals. In this regard it is important, but not always easy, to distinguish between causes and effects of behavioral and physiological responses to stressors. The question raised is to what extent and rigidness stress coping styles are guided by genetic factors.

Serotonergic characteristics of rainbow trout divergent in stress responsiveness

Juvenile rainbow trout divergent in their cortisol response to confinement stress (HR: high responsive or LR: low responsive fish) were exposed to either 1 or 3 h of confinement stress. Untreated fish served as control. After the exposure blood and brain samples were collected. From the blood samples, the levels of cortisol and catecholamines were determined, while the brain serotonergic and monoamineoxidase (MAO) activity was determined in four different brain areas (brain stem, hypothalamus, telencephalon and optic tectum). Our results show that the LR fish responds to handling stress with a higher increase in plasma epinephrine compared to HR fish. Our results also show that confinement stress leads to a larger increase in the serotonergic activity in the brain stem and telencephalon in LR fish compared to HR fish. These results support the hypothesis that stress coping strategies similar to those described in mammals also exists in fish. Further, our results have shown that the MAO activity increases in optic tectum and hypothalamus of rainbow trout during confinement stress, while it remains unchanged or decreases in brain stem and telencephalon. Moreover, the MAO activity does not differ between the two selection lines. This indicates that MAO participates actively in the stress response without contributing to the differences in stress coping strategies.

Stress effects on AVT and CRF systems in two strains of rainbow trout (Oncorhynchus mykiss) divergent in stress responsiveness

The aim for this study was to examine whether the F4 generation of two strains of rainbow trout divergent in their plasma cortisol response to confinement stress (HR: high responder or LR: low responder) would also differ in stress-induced effects on forebrain concentrations of mRNA for corticotropin-releasing factor (CRF), arginine vasotocin (AVT), CRF receptor type 1 (CRF-R1), CRF receptor type 2 (CRF-R2) and AVT receptor (AVT-R). In addition, plasma cortisol concentrations, brainstem levels of monoamines and monoamine metabolites, and behaviour during confinement were monitored. The results confirm that HR and LR trout differ in their cortisol response to confinement and show that fish of these strains also differ in their behavioural response to confinement. The HR trout displayed significantly higher locomotor activity while in confinement than LR trout. Moreover, following 180 min of confinement HR fish showed significantly higher forebrain concentrations of CRF mRNA than LR fish. Also, when subjected to 30 min of confinement HR fish showed significantly lower CRF-R2 mRNA concentrations than LR fish, whereas there were no differences in CRF-R1, AVT or AVT-R mRNA expression between LR and HR fish either at 30 or 180 min of confinement. Differences in the expression of CRF and CRF-R2 mRNA may be related to the divergence in stress coping displayed by these rainbow trout strains.

Aggression in rainbow trout is inhibited by both MR and GR antagonists.

The present study has investigated the effect of exogenous cortisol on aggression in juvenile rainbow trout, along with the involvement of mineralocorticoid (MR) and glucocorticoid receptors (GR) mediating the effects of cortisol. Fish were fed pellets supplemented with cortisol, the GR antagonist mifepristone (RU486) in combination with cortisol, the MR antagonist spironolactone (SA) in combination with cortisol or both antagonists in combination with cortisol. Aggressive behaviour was then assessed 1h subsequent to feeding. Our results showed that the attack latency was increased by exogenous cortisol, an effect that was not abolished by the antagonists. The intensity of aggression was not changed by exogenous cortisol. However, the intensity of aggression was significantly reduced by both antagonists. These results are discussed with regard to cortisol affecting aggressive behaviour through genomic and non-genomic pathways. Our results have demonstrated the involvement of both MR and GR in regulating behavioural responses during social interactions in teleost fish. The intensity of aggression seen in control and cortisol treated fish is probably mediated by the basal levels of cortisol through the intracellular MRs and GRs. We conclude that the initiative to engage in social confrontations is mediated through a non-genomic pathway, which could involve extracellular corticoid receptors. Further, the majority of arguments lean towards the MR and GR antagonists blocking the effect of cortisol on aggressive intensity through intracellular receptors. If this is the case, then it is probable that these two aspects of aggressive behaviour are based on different neuronal mechanisms.

Melanocortin peptides affect the motivation to feed in rainbow trout (Oncorhynchus mykiss).

In this study, we investigated the effects of one melanocortin receptor (MCR) agonist and two antagonists on food intake in juvenile rainbow trout. Baseline food intake was established prior to 1 microl intracerebroventricular injection (ICV) of the non-specific agonist MTII, the MC4R antagonist HS024 and the MC3/4R antagonist SHU9119 at concentrations of 0.3, 1 or 3 nM. Saline-injected fish and untreated fish served as controls. Changes in food intake were observed 1h after the ICV injections. Our results showed that treatment with MTII significantly decreased food intake at 3 nM compared to control, HS024 significantly increased food intake at 3 nM compared to control and saline-treated fish, and SHU9119 significantly increased food intake at 3 nM compared to saline-treated fish. In conclusion, our study provides further evidence, and hence strengthens the hypothesis, that MC4R participates in the control of energy balance in fish in the same manner as in mammals. Our findings that HS024 is more potent than SHU9119 in increasing food intake suggest that the effects of melanocortin on energy balance in rainbow trout are mainly regulated by activation of MC4R. Hence, HS024 seems an excellent tool as a MC4R antagonist in rainbow trout.

Context-dependent responses to novelty in Rainbow trout (Oncorhynchus mykiss), selected for high and low post-stress cortisol responsiveness

Previous studies in a rainbow trout model, selectively bred for high (HR) and low (LR) post stress plasma cortisol levels, have yielded data that are indicative of contrasting stress coping styles. Fish from the HR line have been suggested to display a more diverse behavioral repertoire in challenging situations than the LR counterpart. The present study addressed whether such variation in behavioral flexibility traits was evident in different experimental settings using these selection lines. The fish were subjected to three sets of challenges (novel object test, resident-intruder test and confinement stressor test), all which were repeated a week later. Introducing a novel object evoked a divergent behavioral response in association with feeding: fish from the LR line displayed consistently suppressed feed intake while the HR fish remained unaffected. This observation was found to be repeatable along with attack latency and movement activity from the resident-intruder and confinement stressor tests. These results indicate that the behavioral responses in this animal model are context-dependent and shed new light on the expression of behavioral flexibility.

Changes in regional brain monoaminergic activity and temporary down-regulation in stress response from dietary supplementation with L-tryptophan in Atlantic cod (Gadus morhua)

The brain monoamines serotonin (5-hydroxytryptamine; 5-HT) and dopamine (DA) both play an integrative role in behavioural and neuroendocrine responses to challenges, and comparative models suggest common mechanisms for dietary modulation of transmission by these signal substances in vertebrates. Previous studies in teleosts demonstrate that 7 d of dietary administration with L-tryptophan (Trp), the direct precursor of 5-HT, suppresses the endocrine stress response. The present study investigated how long the suppressive effects of a Trp-enriched feed regimen, at doses corresponding to two, three or four times the Trp levels in commercial feed, last in juvenile Atlantic cod (Gadus morhua) when the fish are reintroduced to a diet with standard amino acid composition. We also wanted to determine whether Trp supplementation induced changes in brain monoaminergic neurochemistry in those forebrain structures innervated by DA and 5-HTergic neurons, by measuring regional activity of DA and 5-HT in the lateral pallial regions (Dl) of the telencephalon and nucleus lateralis tuberis (NLT) of the hypothalamus. Dietary Trp resulted in a dose-dependent suppression in plasma cortisol among fish exposed to confinement stress on the first day following experimental diet; however, such an effect was not observed at 2 or 6 d after Trp treatment. Feeding the fish with moderate Trp doses also evoked a general increase in DA and 5-HT-ergic activity, suggesting that these neural circuits within the NLT and Dl may be indirectly involved in regulating the acute stress response.