Physiological effects on demography: a long-term experimental study of testosterone's effects on fitness

Understanding physiological and behavioral mechanisms underlying the diversity of observed life-history strategies is challenging because of difficulties in obtaining long-term measures of fitness and in relating fitness to these mechanisms. We evaluated effects of experimentally elevated testosterone on male fitness in a population of dark-eyed juncos studied over nine breeding seasons using a demographic modeling approach. Elevated levels of testosterone decreased survival rates but increased success of producing extra-pair offspring. Higher overall fitness for testosterone-treated males was unexpected and led us to consider indirect effects of testosterone on offspring and females. Nest success was similar for testosterone-treated and control males, but testosterone-treated males produced smaller offspring, and smaller offspring had lower postfledging survival. Older, more experienced females preferred to mate with older males and realized higher reproductive success when they did so. Treatment of young males increased their ability to attract older females yet resulted in poor reproductive performance. The higher fitness of testosterone-treated males in the absence of a comparable natural phenotype suggests that the natural phenotype may be constrained. If this phenotype were to arise, the negative social effects on offspring and mates suggest that these effects might prevent high-testosterone phenotypes from spreading in the population.     

Females competing to reproduce: dominance matters but testosterone may not

The associations among aggression, testosterone (T), and reproductive success have been well studied, particularly in male birds. In many species, males challenged with simulated or real territorial intrusions increase T and levels of aggression, outcomes linked to higher dominance status and greater reproductive success. For females, the patterns are less clear. Females behave aggressively towards one another, and in some species, females respond to a social challenge with increases in T, but in other species they do not. Prior work on female dark-eyed juncos (Junco hyemalis) had shown that experimental elevation of T increases social status and intrasexual aggression. Here, we conducted two experiments designed to answer three questions: Are endogenous concentrations of T associated with dominance status in captive female juncos? Does dominance status influence readiness to breed in female juncos? And do captive females increase T in response to a challenge? In the first experiment, we introduced two females to a breeding aviary, allowed them to form a dominance relationship and then introduced a male. We found that dominant females were more likely to breed than subordinates, but that dominance status was not predicted by circulating T. In the second experiment, we allowed a resident male and female to establish ownership of a breeding aviary (territory) then introduced a second, intruder female. We found that resident females were aggressive towards and dominant over intruders, but T did not increase during aggressive interactions. We suggest that during the breeding season, intrasexual aggression between females may influence reproductive success, but not be dependent upon fluctuations in T. Selection may have favored independence of aggression from T because high concentrations of T could interfere with normal ovulation or produce detrimental maternal effects.     

Effects of captivity and testosterone on the volumes of four brain regions in the dark-eyed junco (Junco hyemalis)

This study investigates the effects of captivity and testosterone treatment on the volumes of brain regions involved in processing visual and spatial information in adult dark-eyed juncos (Junco hyemalis). We treated captive and free-living male juncos with either testosterone-filled or empty implants. Captive juncos had a smaller hippocampal formation (HF) (both in absolute volume and relative to telencephalon) than free-living birds, regardless of hormone treatment. Testosterone-treated males (both captive and free-living) had a smaller telencephalon and nucleus rotundus, but not a smaller HF or ectostriatum, than controls. We found that free-living testosterone-treated males had larger home ranges than free-living controls in agreement with earlier experiments, but we found no corresponding difference in HF volume. We discuss the implications of the effect of captivity on HF volume for past and future laboratory experiments.     

Territorial aggression and song of male European robins (Erithacus rubecula) in autumn and spring: effects of antiandrogen treatment

Male robins aggressively defending a feeding territory in winter have low levels of testosterone, while males defending a breeding territory in spring have elevated levels of testosterone. Song is an integrated part of territorial defense during both phases. We investigated whether testosterone is involved in the expression of these behaviors by treating free-living and captive male robins during both phases with the antiandrogen flutamide. Results suggest that, similar to species in which territoriality is restricted to the reproductive phase, aggressive defense of a breeding territory by male robins is facilitated by androgens. Territorial defense during the nonbreeding season, however, does not require androgenic activity. Singing frequency, on the other hand, was not significantly reduced during either phase by flutamide application. Since the quality of male song changes with season we suggest that the incorporation of sexual signals into male song during breeding depends on testosterone, but that the year-round production of territorial "keep out" signals is independent of testosterone.     

Serum prolactin and testosterone levels in captive and wild brown kiwi (Apteryx mantelli) during the prebreeding,breeding, and incubation periods

In brown kiwi (Apteryx mantelli), the male is the primary incubator, a trait that is relatively rare among birds. The maintenance of avian incubation behavior is controlled by the protein hormone prolactin (PRL). Although steroid hormone concentrations in both wild and captive kiwi have previously been reported, this study is the first to report levels of PRL in captive and wild male and female kiwi through the prebreeding and breeding seasons, and to directly compare testosterone (T) concentrations between captive and wild males during the breeding and incubation periods. Female PRL concentrations increased at the time of oviposition, whereas male PRL concentrations rose gradually between the prebreeding and incubation periods. Although males are considered the main incubator, an increase in PRL levels could help females maintain behaviors such as nest guarding, or to take over incubation the event of mate loss. A gradual increase in PRL allows the male to be ready for incubation during the long breeding season. Interestingly, T concentrations in captive males did not decrease during incubation and was significantly higher than in wild males. Continual elevated T could have an impact on sperm production through negative feedback, thereby contributing to the low egg fertility seen in captive kiwi. Therefore, determining the underlying reason for the differences in hormone levels could be significant, if not vital, for improving the success of captive kiwi breeding programs.     

Effect of testosterone on T cell-mediated immunity in two species of Mediterranean lacertid lizards

One of the primary assumptions of the immunocompetence handicap hypothesis is that testosterone has an immunosuppressive effect, but conflicting results have been reported in a variety of bird species concerning the effect of testosterone on the humoral and the T cell-mediated components of the immune system. The T cell-mediated component of the immune system is particularly important during the breeding season, because the likelihood of injury during sexual competition is high and T cell-mediated immunity is essential for healing wounds and resisting infection. In this study we examined the effect of experimentally increased levels of testosterone during breeding season on T cell-mediated immunity in male lizards of two Mediterranean lacertid species, Psammodromus algirus and Acanthodactylus erythrurus. The hormonal treatment significantly increased testosterone of the experimental individuals. T cell-mediated responses to phytohemagglutinin stimulation were significantly suppressed in testosterone-treated males of both species. Furthermore, there was a significant negative relationship between individual variability in T cell-mediated responsiveness and plasma testosterone concentration. The present study is the first to demonstrate testosterone-induced suppression of T cell-mediated immunity in lizards.     

Life History Trade-Offs and Behavioral Sensitivity to Testosterone: An Experimental Test When Female Aggression and Maternal Care Co-Occur

Research on male animals suggests that the hormone testosterone plays a central role in mediating the trade-off between mating effort and parental effort. However, the direct links between testosterone, intrasexual aggression and parental care are remarkably mixed across species. Previous attempts to reconcile these patterns suggest that selection favors behavioral insensitivity to testosterone when paternal care is essential to reproductive success and when breeding seasons are especially short. Females also secrete testosterone, though the degree to which similar testosterone-mediated trade-offs occur in females is much less clear. Here, I ask whether testosterone mediates trade-offs between aggression and incubation in females, and whether patterns of female sensitivity to testosterone relate to female life history, as is often the case in males. I experimentally elevated testosterone in free-living, incubating female tree swallows (Tachycineta bicolor), a songbird with a short breeding season during which female incubation and intrasexual aggression are both essential to female reproductive success. Testosterone-treated females showed significantly elevated aggression, reduced incubation temperatures, and reduced hatching success, relative to controls. Thus, prolonged testosterone elevation during incubation was detrimental to reproductive success, but females nonetheless showed behavioral sensitivity to testosterone. These findings suggest that the relative importance of both mating effort and parental effort may be central to understanding patterns of behavioral sensitivity in both sexes.     

Season and testosterone affect contractile properties of fast calling muscles in the gray tree frog Hyla chrysoscelis

In anurans, circulating levels of androgens influence certain secondary sexual characteristics that are expressed only during the breeding season. We studied the contractile properties of external oblique muscles (used to power sound production) in a species of North American gray tree frog, Hyla chrysoscelis, during the breeding season and also in testosterone-treated captive males and females after the breeding season. Compared with the muscles of breeding-season males, the trunk muscles of postbreeding-season males have 50% less mass, 60% longer twitches, and 40% slower shortening velocities. Testosterone levels similar to those found in breeding-season male hylid frogs restore the contractile speed and mass of male trunk muscles and also convert the small slow trunk muscles of females into larger fast-contracting muscles. We conclude that androgens likely play a key role in altering the contractile properties of these muscles in males during the annual cycle, allowing them to operate in the breeding season at the frequencies required to produce the characteristic rapidly pulsed calls of this species. Females as well as nonbreeding-season males do not produce advertising calls, and therefore the slower muscles found in these animals may allow more economic operation of these muscles. The effects of testosterone on female trunk muscles indicate the potential of this hormone in contributing to the sexual dimorphism in size and contractile properties of these muscles, but this dimorphism is likely due to the interaction of more than one hormone.     

Testosterone increases activity but not daily energy expenditure in captive male dark-eyed juncos, Junco hyemalis

Plasma testosterone (T) levels in male dark-eyed juncos peak early in the breeding season, then decline. If T enhances opportunities for reproductive success, as suggested by previous experiments, why does elevated T not occur naturally? To address this question, we prolonged the early peak level throughout the breeding season and explored potential energetic costs of maintaining elevated T. We measured daily energy expenditure (DEE) of treated males (T-males) and controls (C-males) using doubly labelled water (DLW). We also conducted behaviour scans of T- and C-males housed in outdoor aviaries. DEE was not higher in T-males than in C-males. However, T-males did increase locomotion and foraging and decrease rest and self-maintenance. These results suggest that elevated T may increase the contribution of some components of DEE and lower the contribution of others. Furthermore, the T-induced decrease in allocation of time to rest and maintenance may represent a long-term cost that has led to selection against the maintenance of elevated T beyond the natural early spring peak. Copyright 2000 The Association for the Study of Animal Behaviour.     

Testosterone affects reproductive success by influencing extra-pair fertilizations in male dark-eyed juncos (Aves: Junco hyemalis)

Monogamous male birds typically allocate less effort to courtship and more to parental behaviour than males of polygynous species. The seasonal pattern of testosterone (T) secretion varies accordingly. Monogamous males exhibit a spring peak in plasma T followed by lower levels during the parental phase, while males of polygynous species continue to court females and maintain T at higher levels. To determine whether testosterone underlies the trade-off between mating and parental effort, we treated male dark-eyed juncos (Junco hyemalis) with exogenous T and compared the reproductive success (RS) of T-treated males (T-males) to that of controls. T-males had lower apparent annual RS than controls, probably because elevated T reduced parental care. Nevertheless, annual genetic RS of the treatment groups was similar because (i) T-males suffered fewer losses in genetic RS due to extra-pair fertilizations (EPFs), and (ii) T-males gained more genetic RS through their own EPFs. This is the first hormonal manipulation of an avian phenotype shown to have influenced male RS through EPFs. Together with other studies, it suggests that testosterone may have mediated the evolution of inter- and intraspecific differences in allocation of reproductive effort to mate attraction and parental care.     

Relative abundance of males to females affects behaviour, condition and immune function in a captive population of dark-eyed juncos Junco hyemalis

Relative numbers of males and females in breeding groups may vary from expected values owing to a variety of factors. To determine how sex ratio might influence individual phenotypes in a captive population of dark-eyed juncos Junco hyemalis during the breeding season, we established three treatment groups: a male-biased (2:1), equal (1:1), and female-biased group (1:2). Within-group density (birds/m²) was constant across groups. We assessed the frequency of flight chases (a proxy for social instability), measured changes in body mass and pectoral muscle condition, assayed plasma levels of testosterone (T) and compared cell-mediated immunity of individuals. We found significantly more chases in the male-biased group than in the female-biased group. Birds in the male-biased group lost more mass and displayed poorer pectoral-muscle condition than birds in the equal group. Cell-mediated immune responses were reduced in individuals in the male-biased group in comparison to the female-biased group. Plasma T levels in both sexes did not vary with sex ratio. Collectively, these results suggest that during the breeding season, social instability is greater in male-biased populations, and instability may lead to decreased general health and vigour.     

Factors influencing reproductive success and litter size in captive island foxes

A severe decline of island foxes (Urocyon littoralis) on the northern Channel Islands in the 1990s prompted the National Park Service to begin a captive breeding program to increase their numbers. Using detailed records of all the fox pairs (N = 267) that were part of the program on San Miguel, Santa Rosa, and Santa Cruz Islands from its inception in 2000 through 2007, we identified factors influencing the breeding success of pairs in captivity in the interest of formulating strategies that could increase captive productivity. We compiled a database of variables including litter size, reproductive success, distance to nearest occupied pen during the breeding season, subspecies, exposure, female age, male age, age difference, female and male origin (wild vs. captive born), same versus different origin, years paired, previous reproductive success by the pair, previous reproductive success by the female, mate aggression-related injuries, male previous involvement in a pair with mate aggression, and female previous involvement in a pair with mate aggression. We used multiple linear regression to identify factors predictive of litter size, and logistic regression to predict the probability of reproductive success. A larger inter-pen distance, older male age, less exposure, and a smaller intra-pair age difference positively affected litter size. The probabilities of reproductive success increased with fewer years paired and less exposure. Comparatively, pairs with wild born females (vs. captive born females), and previously successful pairs (vs. previously unsuccessful and new pairs) were most likely to be successful. These results indicate that the optimal situation was to pair wild-caught females with older males in sheltered pens that were as far from other pens as possible, to maintain successful pairs and repair unsuccessful ones. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.     

Consequences of elevating plasma testosterone in females of a socially monogamous songbird: evidence of constraints on male evolution?

To explore whether selection for testosterone-mediated traits in males might be constrained by costs of higher testosterone to females, we examined the effects of experimental elevation of plasma testosterone on physiological, reproductive, and behavioral parameters in a female songbird, the dark-eyed junco (Junco hyemalis). We used subcutaneous implants to elevate testosterone (T) in captive and free-living female juncos. In captive birds, we measured the effects of high T on body mass, feather molt, and brood patch formation. In the field, we monitored its effects on the timing of egg laying, clutch size, egg size, egg steroid levels, incubation, and nest-defense behavior. Females implanted with testosterone (T-females) had significantly higher circulating levels of testosterone than did control females (C-females). Captive T-females had lower body mass, were less likely to develop brood patches, and delayed feather molt relative to C-females. Among free-living females, the interval between nest completion and appearance of the first egg was longer for T-females than for C-females and egg yolk concentrations of testosterone were higher, but there were no significant differences in estradiol levels, clutch size, or egg size. Incubation and nest defense behavior were also similar between T- and C-females. Our results suggest that selection on males for higher testosterone might initially lead to a correlated response in females producing changes in body mass and feather molt, both of which could be detrimental. Other possible female responses would be delayed onset of reproduction, which might reduce reproductive success, and higher yolk testosterone, which might have either positive or negative effects on offspring development. We found no reason to expect reduced parental behavior by females as a negative fitness consequence of selection for higher testosterone in males.     

A 15-year study of the association between dominance rank and reproductive success of male rhesus macaques

The reproductive success (RS) of 32 males in a captive group of rhesus macaques (Macaca mulatta) between 1978 and 1992 was determined using paternity exclusion analysis. Dominance rank of each male over age 4 was assessed at the end of each breeding season based on agonistic dyadic interactions. The dominance rank and RS of these males were strongly correlated whether or not subadult males were included. The high reproductive success of males that eventually reached alpha rank is primarily responsible for this outcome. These results support the theory that social dominance has evolved in genusMacaca by sexual selection but some changes in male dominance rank and RS during the 15-year period suggest that priority of access is not the sole focus for such selection.     

Effects of captivity and testosterone on the volumes of four brain regions in the dark‐eyed junco (Junco hyemalis)

This study investigates the effects of captivity and testosterone treatment on the volumes of brain regions involved in processing visual and spatial information in adult dark‐eyed juncos (Junco hyemalis). We treated captive and free‐living male juncos with either testosterone‐filled or empty implants. Captive juncos had a smaller hippocampal formation (HF) (both in absolute volume and relative to telencephalon) than free‐living birds, regardless of hormone treatment. Testosterone‐treated males (both captive and free‐living) had a smaller telencephalon and nucleus rotundus, but not a smaller HF or ectostriatum, than controls. We found that free‐living testosterone‐treated males had larger home ranges than free‐living controls in agreement with earlier experiments, but we found no corresponding difference in HF volume. We discuss the implications of the effect of captivity on HF volume for past and future laboratory experiments. © 2000 John Wiley & Sons, Inc. J Neurobiol 43: 244–253, 2000     

Territorial aggression does not feed back on testosterone in a multiple-brooded songbird species with breeding and non-breeding season territoriality,the European stonechat

Testosterone mediates reproductive behaviours in male vertebrates. For example, breeding season territoriality depends on testosterone in many species of birds and in some, territorial interactions feed back on testosterone concentrations. However, the degree to which territorial behaviour and testosterone are associated differs even between species with seemingly similar life histories, especially between species that also defend territories outside the breeding season. Here, we investigate the link between territorial behaviour and testosterone in European stonechats. Previous studies found that territorial aggression in stonechats depends on testosterone in a breeding, but not in a non-breeding context. We investigated whether stonechats show a rise in testosterone during simulated territorial intrusions (STI) during the breeding season. Post-capture testosterone concentrations of males caught after an STI were not higher than those of males caught in a control situation regardless of breeding stage. However, most of the males would have been able to mount a testosterone response because the same individuals that did not increase testosterone during the STI showed a substantial increase in testosterone after injections of gonadotropin-releasing hormone (GnRH). GnRH-induced and post-capture testosterone concentrations were positively correlated and both decreased with successive breeding stages. Further, territory owners with a short latency to attack the decoy expressed higher post-capture testosterone concentrations than males with a longer latency to attack the decoy. Thus, there is no evidence for behavioural feedback on testosterone concentrations during male-male interactions in stonechats. In combination with previous studies our data suggest that testosterone functions as an on/off switch of high intensity territorial aggression during the breeding season in stonechats. The among-species variation in the androgen control of territorial behaviour may be only partly a result of environmental differences. Instead, potential differences in how territoriality evolved in different species may have influenced whether and how a reproductive hormone such as testosterone was co-opted into the mechanistic control of territorial behaviour.     

Time course of androgenic modulation of odor preferences and odor cues in male meadow voles, Microtus pennsylvanicus.

During the breeding season, male meadow voles prefer female over male odors and females prefer male over female odors. Testosterone control of males' odor preferences and production of odors attractive to females differ. A male meadow vole's preference for female versus male odor was still evident 1 week after castration, but not 1 week later. This preference was reinstated in testosterone-treated male voles 2 weeks after the onset of hormone replacement. The attractiveness of male odors to females did not disappear until 3 weeks after castration. The attractiveness of male odors was reinstated 1 week after castrated males were treated with testosterone. The time course for the androgenic modulation of production of odors attractive to females may facilitate breeding. For example, at the end of the breeding season males may emit an odor that is still attractive to females. Similarly, at the beginning of the breeding season males may emit an odor that is attractive to females.     

Effects of capture stress on free-ranging, reproductively active male Weddell seals

Physiological stress responses to capture may be an indicator of welfare challenges induced by animal handling. Simultaneously, blood chemistry changes induced by stress responses may confound experimental design by interacting with the biological parameters being measured. Cortisol elevation is a common indicator of stress responses in mammals and reproductive condition can profoundly influence endocrine response. We measured changes in blood cortisol and testosterone induced by handling reproductively active male Weddell seals (Leptonychotes weddellii) early and late in the breeding season. Weddell seals have the highest resting cortisol levels of all mammals yet showed a clear, prolonged elevation in cortisol in response to capture. Responses were similar when first caught and when caught a second time, later in the breeding season. Baseline testosterone levels declined over the breeding season but were not altered by capture. Administering a light dose of diazepam significantly ameliorated the cortisol response of handled animals without affecting testosterone levels. This may be an effective way of reducing acute capture stress responses. Male breeding success in years males were handled was no different to the years they were not, despite the acute capture response, suggesting no long-term impact of handling on male reproductive output.     

Research on reproductive biology of gorillas

Laboratory research has implicated several variables which contribute to the regulation of reproductive behaviour of captive gorillas. Females were found to be increasingly attractive to males during the follicular phase of the menstrual cycle when estrogen and testosterone concentrations in the female were increasing. Females solicited mating from the males primarily at midcycle, about the time when the concentration of testosterone was maximal. No mating occurred during the mid- to late luteal phase after progesterone concentrations were elevated. Both males and females initiated mating during the midcycle, periovulatory period, but male initiative accounted for most mating that was temporally dissociated from that period. Individual differences between males and among females contributed to the variability in results. Confinement of a male and female in relatively small quarters appears to interact with certain aspects of species-typical behaviour to distort patterns of mating in laboratory tests. Data on behaviour of gorillas in the wild contributed to interpretation of the laboratory results and suggest an enlightened approach to the captive maintenance and breeding of gorillas. An important consideration in promoting captive breeding of gorillas seems to be the provision of options to the female for regulating the frequency and distribution of mating in the cycle.     

Developmental programming: deficits in reproductive hormone dynamics and ovulatory outcomes in prenatal, testosterone-treated sheep

Prenatal testosterone excess leads to neuroendocrine, ovarian, and metabolic disruptions, culminating in reproductive phenotypes mimicking that of women with polycystic ovary syndrome (PCOS). The objective of this study was to determine the consequences of prenatal testosterone treatment on periovulatory hormonal dynamics and ovulatory outcomes. To generate prenatal testosterone-treated females, pregnant sheep were injected intramuscularly (days 30-90 of gestation, term=147 days) with 100 mg of testosterone-propionate in cottonseed oil semi-weekly. Female offspring born to untreated control females and prenatal testosterone-treated females were then studied during their first two breeding seasons. Sheep were given two injections of prostaglandin F2alpha 11 days apart, and blood samples were collected at 2-h intervals for 120 h, 10-min intervals for 8 h during the luteal phase (first breeding season only), and daily for an additional 15 days to characterize changes in reproductive hormonal dynamics. During the first breeding season, prenatal testosterone-treated females manifested disruptions in the timing and magnitude of primary gonadotropin surges, luteal defects, and reduced responsiveness to progesterone negative feedback. Disruptions in the periovulatory sequence of events during the second breeding season included: 1) delayed but increased preovulatory estradiol rise, 2) delayed and severely reduced primary gonadotropin surge in prenatal testosterone-treated females having an LH surge, 3) tendency for an amplified secondary FSH surge and a shift in the relative balance of FSH regulatory proteins, and 4) luteal responses that ranged from normal to anovulatory. These outcomes are likely to be of relevance to developmental origin of infertility disorders and suggest that differences in fetal exposure or fetal susceptibility to testosterone may account for the variability in reproductive phenotypes.