The effects of social isolation on steroid hormone levels are modulated by previous social status and context in a cichlid fish

Social isolation is a major stressor which impacts the physiology, behaviour and health of individuals in gregarious species. However, depending on conditional and contextual factors, such as social status and group composition, social isolation may be perceived differently by different individuals or even by the same individuals at different times. Here we tested the effects of social status (territorial vs. non-territorial) and previous group composition (i.e. type of social group: mixed sex group with two territorial males, TT vs. mixed sex group with one territorial and one non-territorial male, TnT) on the hormonal response (androgens and cortisol) to social isolation in a cichlid fish (Oreochromis mossambicus). The different steroid hormones measured responded differentially to social isolation, and their response was modulated by social factors. Social isolation elicited a decrease of 11-keto formation only in territorial males, whereas non-territorial males present a non-significant trend for increasing KT levels. Testosterone did not respond to social isolation. Cortisol only increased in isolated individuals from TnT groups irrespective of social status (i.e. both in territorials and non-territorials). These results suggest that it is the perception of social isolation and not the objective structure of the situation that triggers the hormonal response to isolation.     

From molecule to behavior: Brain aromatase (cyp19a1b) characterization,expression analysis and its relation with social status and male agonistic behavior in a Neotropical cichlid fish

The enzyme aromatase, responsible for the conversion of C19 androgens to C18 estrogens, exists as two paralogue copies in teleost fish: Cyp19a1a mostly expressed in the gonads, referred as gonadal aromatase, and Cyp19a1b, mostly expressed in the brain, accordingly known as brain aromatase. The neural localization of Cyp19a1b is greatly contained within the social behavior network and mesolimbic reward system in fish, suggesting a strong role of estrogen synthesis in the regulation of social behavior. In this work we aimed to analyze the variation in cyp19a1b expression in brain and pituitary of males of a highly social cichlid, Cichlasoma dimerus (locally known as chanchita), and its relation with inter-individual variability in agonistic behavior in a communal social environment. We first characterized chanchita's cyp19a1b mRNA and deduced amino acid sequence, which showed a high degree of conservation when compared to other teleost brain aromatase sequences, and its tissue expression patterns. Within the brain, Cyp19a1b was solely detected at putative radial glial cells of the forebrain, close to the brain ventricles. We then studied the relative expression levels of cyp19a1b by Real Time PCR in the brain and pituitary of males of different social status, territorial vs. non-territorial, and its relationship with an index of agonistic behavior. We found that even though, brain aromatase expression did not differ between types of males, pituitary cyp19a1b expression levels positively correlated with the index of agonistic behavior. This suggests a novel role of the pituitary in the regulation of social behavior by local estrogen synthesis.     

Functional groups in the social behavior of a cichlid fish,the Oscar,Astronotus ocellatus

Dummy conspecifics were presented to isolated adults of the cichlid fish Astronotus ocellatus to investigate the functional organization of cichlid social behavior. Body size and 15 dummy-elicited activities were recorded during 15 min sessions and analyzed by principal components analysis (PCA) to reveal their temporal organization. Five principal components explained almost 80% of the variation in dummy-elicited behavior, and these five factors define functional groups for

• 1.(a) investigation,
• 2.(b) attack,
• 3.(c) nesting,
• 4.(d) boldness,
• 5.(e)distress.

Nest-oriented and attack modal action patterns are not mutually inhibitory during this time frame, and biting does not appear to function exclusively during an attack on a conspecific. Comparison with previous studies of New and Old World cichlids suggests evolutionary conservation of the functional organization of social behavior.     

The relationship between fecundity and fertility in the mouthbrooding cichlid fish Tilapia leucosticta

The development of fry in the mouth of the female mouthbrooder Tilapia leucosticta Trewavas is described. The egg production by fish of various lengths is plotted and a curvilinear regression for standard and total length fitted. The number of eggs produced is approximately equal to the square of the total length in centimetres of the parent fish. The brood numbers of 460 females are plotted against length, and linear regressions for mean brood number and mean fry number fitted. The relationship between the expressions for numbers of fry brooded and egg production gives an estimate of brooding efficiency that decreases with increasing parent length. Individual female fish show signs of reripening and the timing of this process is estimated from gonad state and from the length of fry brooded in the mouth. The relationship between fecundity (egg production) and fertility (fry brooded) is considered and the effect of maturation size, growth rate and spawning frequency assessed.     

The relationship between fecundity and fertility in the mouthbrooding cichlid fish Tilapia leucosticta

The development of fry in the mouth of the female mouthbrooder Tilapia leucosticta Trewavas is described. The egg production by fish of various lengths is plotted and a curvilinear regression for standard and total length fitted. The number of eggs produced is approximately equal tothe square of the total length in centimetres of the parent fish. The brood numbers of 460 femalesare plotted against length, and linear regressions for mean brood number and mean fry number fitted. The relationship between the expressions for numbers of fry brooded and egg production gives anestimate of brooding efficiency that decreases with increasing parent length. Individual female fish show signs of reripening and the timing of this process is estimated from gonad state and from the length of fry brooded in the mouth. The relationship between fecundity (egg production) and fertility (fry brooded) is considered and the effect of maturation size, growth rate and spawning frequency assessed.     

Steroid hormones and agonistic behavior in a cichlid teleost, Aequidens pulcher

The effects of three steroid hormones on the agonistic behavior of female Aequidens pulcher have been evaluated. Testosterone, estradiol, and cortisol were tested using an immersion technique to minimize trauma, and we also examined metyrapone, a blocker of cortisol biosynthesis. Two different experimental protocols were employed, the first investigating agonistic interactions within groups of fish, and the second examining the responses of isolated fish to models and mirrors. Differences between replicates were small, and both protocols supported similar conclusions. Each of the three hormones produced a characteristically different spectrum of behaviors when compared to the controls. Testosterone increased agonistic behavior in all experimental situations, while estradiol had a generally opposite effect; this may reflect the natural modulation of behavior by hormones during the reproductive cycle of A. pulcher. Cortisol also had distinct behavioral effects; available evidence suggests that this steroid increases submissive components of agonistic behavior, and that observed increases in some aggressive components are an indirect consequence, dependent upon the feedback of social information received by each fish. Metyrapone treatment greatly reduced all agonistic behaviors, groups of fish forming shoals typical of juveniles. This was not reversed by replacement therapy with cortisol, which suggests that metyrapone affects behavior by an alternative, possibly toxic, mechanism.     

Sex differences and similarities in the neuroendocrine regulation of social behavior in an African cichlid fish

An individual's position in a social hierarchy profoundly affects behavior and physiology through interactions with community members, yet little is known about how the brain contributes to status differences between and within the social states or sexes. We aimed to determine sex-specific attributes of social status by comparing circulating sex steroid hormones and neural gene expression of sex steroid receptors in dominant and subordinate male and female Astatotilapia burtoni, a highly social African cichlid fish. We found that testosterone and 17β-estradiol levels are higher in males regardless of status and dominant individuals regardless of sex. Progesterone was found to be higher in dominant individuals regardless of sex. Based on pharmacological manipulations in males and females, progesterone appears to be a common mechanism for promoting courtship in dominant individuals. We also examined expression of androgen receptors, estrogen receptor α, and the progesterone receptor in five brain regions that are important for social behavior. Most of the differences in brain sex steroid receptor expression were due to sex rather than status. Our results suggest that the parvocellular preoptic area is a core region for mediating sex differences through androgen and estrogen receptor expression, whereas the progesterone receptor may mediate sex and status behaviors in the putative homologs of the nucleus accumbens and ventromedial hypothalamus. Overall our results suggest sex differences and similarities in the regulation of social dominance by gonadal hormones and their receptors in the brain.     

The relationship between social status, behaviour, growth and steroids in male helpers and breeders of a cooperatively breeding cichlid

We tested whether subordinate helper males of the Lake Tanganyika cooperatively breeding cichlid Neolamprologus pulcher show elevated excretion levels of the stress hormone cortisol and reduced levels of 11-ketotestosterone and testosterone when living in groups with a small breeder male, compared to similar helper males living in groups with a large breeder male, in a full-factorial repeated measures experimental design. We also measured the same hormones in breeder males with and without helper males. Previous research showed that the size difference between large male helpers and male breeders in groups of this species influences behaviour and growth decisions. Contrary to our expectation, no effect of the size-difference between helper males and breeder males on helper hormone levels was detected. Furthermore, helper males had similar hormone excretion levels to those of size-matched breeder males without helpers, and to small breeder males. There was no influence of egg laying on breeder male and helper hormone levels during the experiment. Interestingly, all three hormone levels were significantly lower in helpers showing elevated levels of submissive behaviour towards the breeders, independently of the size of the breeder males. The low cortisol levels suggest that helper males can successfully reduce stress by appeasing breeder males through submission. Furthermore, helper males showing a high level of submissive behaviour had lower levels of androgens than less submissive helpers, suggesting a lower reproductive potential in submissive helpers. We propose that helper submission may be used as an honest signal of reduced interest in reproduction towards the breeder male in this species.     

Social context influences aggressive and courtship behavior in a cichlid fish

Social interactions require knowledge of the environment and status of others, which can be acquired indirectly by observing the behavior of others. When being observed, animals can also alter their signals based on who is watching. Here we observed how male cichlid fish (Astatotilapia burtoni) behave when being watched in two different contexts. In the first, we show that aggressive and courtship behaviors displayed by subordinate males depends critically on whether dominant males can see them, and in the second, we manipulated who was watching aggressive interactions and showed that dominant males will change their behavior depending on audience composition. In both cases, when a more dominant individual is out of view and the audience consists of more subordinate individuals, those males signal key social information to females by displaying courtship and dominant behaviors. In contrast, when a dominant male is present, males cease both aggression and courtship. These data suggest that males are keenly aware of their social environment and modulate their aggressive and courtship behaviors strategically for reproductive and social advantage.     

Wasteful carotenoid coloration and its effects on territorial behavior in a cichlid fish

The famously diverse body coloration of cichlid fish serves communicative functions in mating and social interactions including competition for resources. Here, we examined the effects of a color pattern trait—the width of a yellow bar on a black body—on territorial competition in males and females of a color variant (“Ikola”) of the Lake Tanganyika cichlid Tropheus. First, measuring integumentary carotenoid concentrations in the yellow and black body regions, we established that wider yellow bars require more carotenoids allocated to body coloration. However, we also detected high carotenoid concentrations in the black body regions (>?100 µg/g fresh skin), raising questions about the function of non-displayed integumentary carotenoids. Behavioral experiments showed that fish with wider bars were quicker to explore an unfamiliar area of the tank. In experiments including presentations of fish dummies, the bar width of ‘territorial’ dummies had no effect on the latency time which test fish took to intrude into the dummies’ territories. However, male test fish performed fewer aggressive acts against wide-barred than against narrow-barred dummy competitors. Our results suggest that intimidation by wide bars as well as correlations between bar width and explorative behavior may contribute to mediating success in territorial Tropheus “Ikola”.

     

Visual information alone changes behavior and physiology during social interactions in a cichlid fish (Astatotilapia burtoni)

Social behavior can influence physiological systems dramatically yet the sensory cues responsible are not well understood. Behavior of male African cichlid fish, Astatotilapia burtoni, in their natural habitat suggests that visual cues from conspecifics contribute significantly to regulation of social behavior. Using a novel paradigm, we asked whether visual cues alone from a larger conspecific male could influence behavior, reproductive physiology and the physiological stress response of a smaller male. Here we show that just seeing a larger, threatening male through a clear barrier can suppress dominant behavior of a smaller male for up to 7 days. Smaller dominant males being "attacked" visually by larger dominant males through a clear barrier also showed physiological changes for up to 3 days, including up-regulation of reproductive- and stress-related gene expression levels and lowered plasma 11-ketotestesterone concentrations as compared to control animals. The smaller males modified their appearance to match that of non-dominant males when exposed to a larger male but they maintained a physiological phenotype similar to that of a dominant male. After 7 days, reproductive- and stress- related gene expression, circulating hormone levels, and gonad size in the smaller males showed no difference from the control group suggesting that the smaller male habituated to the visual intruder. However, the smaller male continued to display subordinate behaviors and assumed the appearance of a subordinate male for a full week despite his dominant male physiology. These data suggest that seeing a larger male alone can regulate the behavior of a smaller male but that ongoing reproductive inhibition depends on additional sensory cues. Perhaps, while experiencing visual social stressors, the smaller male uses an opportunistic strategy, acting like a subordinate male while maintaining the physiology of a dominant male.     

Behavioral coping strategies in a cichlid fish: the role of social status and acute stress response in direct and displaced aggression

The African cichlid fish, Astatotilapia burtoni, has a complex social system with a sophisticated social hierarchy that offers unique opportunities to understand how social rank and its physiological substrates relate to behavioral strategies. In A. burtoni, a small fraction of the males are dominant (T, territorial), as distinguished by being large, brightly colored, reproductively active, and aggressively defending territories. In contrast, the majority of males are non-dominant (NT, non-territorial), being smaller, drably colored, sexually immature, and typically schooling with females. The social system is regulated by aggressive interactions between males and behavioral responses to aggression can be direct or displaced with respect to the animal that acts. To determine whether direct and displaced behaviors are differentially exhibited by T and NT males, individuals were shown a video presentation of a dominant male displaying aggressively. Analysis of aggressive acts toward the video display and displaced activity toward a tank mate revealed that T males exhibited more direct behavior (toward the video display), while NT males engaged in more displaced behavior (toward tank mates). Because similar experiments with primates suggest that shifts in behavioral strategies are linked to changes in the stress response (as measured by circulating cortisol levels), we measured cortisol levels of T and NT males following exposure to the aggressive stimulus. Although in some animals subordinate males are reported to have higher cortisol levels, here we show that in A. burtoni the endocrine response to specific situations can vary considerably even among animals of the same status. Interestingly, NT males with intermediate cortisol levels showed more directed behavior while NT males with both high and low cortisol levels showed more displaced. This suggests an optimal physiological stress response in NT males that predisposes them to challenge aggressors perhaps making it more likely for them to ascend in status.     

Effects of shell status and social context on the agonistic behavior of the tropical hermit crab,Clibanarius signatus

In the hermit crabClibanarius signatus, the influence of both shell status and social context on agonistic behavior was examined. The experimental hermits were of the same size and sex, but differed in the status of the inhabited shell, which was either optimal, sub-optimal or damaged. The evidence from this experiment suggests that the agonistic behavior ofC. signatus is influenced to a great extent by asymmetries in the resource value; attacks are mainly performed by crabs which inhabit shells in a relatively poor condition and are mostly directed towards opponents with better-fitting shells. However, this effect differs according to the social context, being more pronounced when the potential gain is greater. Shell fights are decidedly more complex than most animal combats; more work is certainly needed to understand the mechanisms underlying resource assessment and decisions made during hermit crabs' interactions.     

Females of an African cichlid fish display male-typical social dominance behavior and elevated androgens in the absence of males

Social environment can affect the expression of sex-typical behavior in both males and females. Males of the African cichlid species Astatotilapia burtoni have long served as a model system to study the neural, endocrine, and molecular basis of socially plastic dominance behavior. Here we show that in all-female communities of A. burtoni, some individuals acquire a male-typical dominance phenotype, including aggressive territorial defense, distinctive color patterns, and courtship behavior. Furthermore, dominant females have higher levels of circulating androgens than either subordinate females or females in mixed-sex communities. These male-typical traits do not involve sex change, nor do the social phenotypes in all-female communities differ in relative ovarian size, suggesting that factors other than gonadal physiology underlie much of the observed variation. In contrast to the well-studied situation in males, dominant and subordinate females do not differ in the rate of somatic growth. Dominant females are not any more likely than subordinates to spawn with an introduced male, although they do so sooner. These results extend the well known extraordinary behavioral plasticity of A. burtoni to the females of this species and provide a foundation for uncovering the neural and molecular basis of social dominance behavior while controlling for factors such as sex, gonadal state and growth.     

Immune activation is inversely related to, but does not cause variation in androgen levels in a cichlid fish species

Studies on birds and mammals indicate that sexual traits may signal superior health because active immunity, like inflammatory responses to infections, is suppressive to the production of androgens that facilitate the expression of these traits. Here we test this possible pathway for honest signaling in a teleost species, Sarotherodon galilaeus, by activating the immune system with sheep red blood cells (SRBC), which is a non-pathogenic T- and B-cell stimulating antigen. Two weeks after the start of treatment adult males injected with SRBC showed a significant increase in antibody production in comparison with control males. The variation in specific antibody production was negatively related with variation in both testosterone and 11-ketotestosterone levels. This suggests that investment in immune protection is incompatible with increased activity of the hypothalamic-pituitary-gonadal axis. However, opposite to our expectation no difference in androgen levels was found between placebo and SRBC treatment suggesting that immune activation did not cause androgen suppression in our studied species.     

The influence of status and the social environment on energy stores in a social fish

This study explores how muscle and liver energy stores are linked with social status and the social environment in Neolamprologus pulcher, a cooperatively breeding fish that lives in colonies comprised of up to 200 distinct social groups. Subordinate muscle energy stores were positively correlated with the number of neighbouring social groups in the colony, but this pattern was not observed in dominant N. pulcher. Furthermore, liver energy stores were smaller in dominants living at the edge of the colony compared with those living in the colony centre, with no differences among subordinates in liver energy stores. Subordinate N. pulcher may build up large energy stores in the muscles to fuel rapid growth after dispersal, which could occur more frequently in high‐density environments. Dominant N. pulcher may use the more easily mobilized energy stores in the liver to fuel daily activities, which could be more energetically demanding on the edge of the colony as a result of the increased predation defence needed on the edge. Overall, this study demonstrates that both subordinate and dominant physiology in N. pulcher varies with characteristics of the social environment. Furthermore, dominant and subordinate energy storage strategies appear to differ due to status‐dependent variation in daily activities and variation in the need to prepare for future reproductive or dispersal opportunities.