Joseph M. Casto

Steroid Hormones and Immune Function: Experimental Studies in Wild and Captive Dark‐Eyed Juncos (Junco hyemalis)

Monogamous and polygynous male songbirds generally differ in their breeding season profiles of circulating testosterone. Testosterone level spikes early in the breeding season of monogamists and then declines, but it remains high in polygynists. Male dark‐eyed juncos (Junco hyemalis) are socially monogamous and exhibit the usual pattern, but experimental maintenance of high testosterone throughout the breeding season alters normal behavior and physiology and affects various components of annual reproductive success but not overall annual success. Because stabilizing selection predicts that alteration of naturally existing phenotypes should reduce lifetime reproductive success, we asked whether prolonged testosterone exposure might impair immune function and perhaps thereby reduce life span. We assessed immune function in captive and wild male juncos that we treated with either testosterone‐filled or empty Silastic implants. Results indicate that prolonged elevation of testosterone suppresses antibody production in captive males and cell‐mediated immunity in wild males. Together these results suggest that testosterone‐treated males may be more susceptible to disease or parasitic infection. As earlier studies have shown, levels of corticosterone as well as testosterone are higher in testosterone‐treated males, so it is unclear whether the immune suppression we observed is due to testosterone’s direct effects on immunity or testosterone’s influence on glucocorticoid production. We discuss results in the context of recent hypotheses regarding life‐history theory and potential endocrine‐immune interactions.

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.

Behavioral and physiological responses to experimentally elevated testosterone in female dark-eyed juncos (Junco hyemalis carolinensis)

Testosterone mediates the expression of many fitness-related traits in male vertebrates and is thought to account for numerous sex differences in trait expression. Testosterone is also secreted by females; however, far less is known regarding its effects on female physiology and behavior. Using a bird species in which the effects of testosterone on males are well characterized, the dark-eyed junco (Junco hyemalis), we tested whether an increase in exogenous testosterone in females would alter the phenotypic expression of a suite of behavioral and physiological traits. We found that increased testosterone levels in female dark-eyed juncos led to decreased cell-mediated immune function and increased intrasexual aggression, hypothalamo-pituitary-adrenal (HPA) axis responsiveness, baseline corticosterone and corticosterone-binding globulin (CBG) levels. Furthermore, immunosuppression following testosterone implantation was negatively correlated with total and free testosterone but did not appear to be related to either total or free corticosterone. These results demonstrate that the phenotypic impact of elevated testosterone is not confined to males in dark-eyed juncos, and that the impact in adults can be similar in males and females. We discuss these results in the context of potential endocrine-immune interactions and the evolution of sexual dimorphism.

Hormones and honest signals: males with larger ornaments elevate testosterone more when challenged.

When male investment in mating varies with quality, reliable sexual signals may evolve. In many songbirds, testosterone mediates mating investment, suggesting that signals should be linked to testosterone production. However, because testosterone may change rapidly during behaviour such as territorial aggression and courtship, efforts to establish such a relationship have proved challenging. In a population of dark‐eyed juncos, we measured individual variation in the production of short‐term testosterone increases by injecting gonadotropin‐releasing hormone (GnRH). We found a positive correlation between the magnitude of these increases and the size of a plumage ornament (‘tail white’) previously shown to be important for female choice and male–male competition. We then measured naturally elevated testosterone levels produced during male–male competition and found that they covaried with those induced by GnRH. We suggest that the association between tail white and testosterone increases may allow conspecifics to assess potential mates and competitors reliably using tail white.

Elevated testosterone reduces choosiness in female dark-eyed juncos (Junco hyemalis): evidence for a hormonal constraint on sexual selection?

Because testosterone (T) often mediates the expression of attractive displays and ornaments, in the absence of constraints sexual selection should lead to an evolutionary increase in male T levels. One candidate constraint would be a genetic correlation between the sexes that leads to a correlated response in females. If increased T in females were to have deleterious effects on mate choice, the effect of sexual selection on male T would be weakened. Using female dark–eyed juncos (Junco hyemalis), we tested whether experimentally enhancing female T would lead to a decrease in discrimination between two classes of males, one treated with T (T–males) and one control (C–males). The two female treatments (T–implanted and C–females) spent equal amounts of time with both classes of males, but T–treated females failed to show a preference for either male treatment, whereas C–females showed a significant preference, albeit in an unexpected direction (for C–males). T–females were less discriminating than C–females, irrespective of the direction of their preference. To our knowledge, this is the first study to show that circulating hormones can alter female choosiness without reducing sexual motivation. Our results suggest that hormonal correlations between the sexes have the potential to constrain sexual selection on males.

Embryonic modulation of maternal steroids in European starlings (Sturnus vulgaris)

In birds, maternally derived yolk steroids are a proposed mechanism by which females can adjust individual offspring phenotype to prevailing conditions. However, when interests of mother and offspring differ, parent–offspring conflict will arise and embryonic interests, not those of the mother, should drive offspring response to maternal steroids in eggs. Because of this potential conflict, we investigated the ability of developing bird embryos to process maternally derived yolk steroids. We examined how progesterone, testosterone and oestradiol levels changed in both the yolk/albumen (YA) and the embryo of European starling eggs during the first 10 days of development. Next, we injected tritiated testosterone into eggs at oviposition to characterize potential metabolic pathways during development. Ether extractions separated organic and aqueous metabolites in both the embryo and YA homogenate, after which major steroid metabolites were identified. Results indicate that the concentrations of all three steroids declined during development in the YA homogenate. Exogenous testosterone was primarily metabolized to an aqueous form of etiocholanolone that remained in the YA. These results clearly demonstrate that embryos can modulate their local steroid environment, setting up the potential for parent–offspring conflict. Embryonic regulation must be considered when addressing the evolutionary consequences of maternal steroids in eggs.

Female ornamentation and male mate choice in dark-eyed juncos

Traits that enhance attractiveness in one sex may or may not influence attractiveness in the other. In the dark-eyed junco, Junco hyemalis, outer tail feathers of males and females are all or partly white and form a sharp contrast with the bird’s mostly grey plumage. The amount of white in these feathers (‘tail white’) is greater in males than in females and, as we report here, is greater in birds that have completed a second prebasic moult than in yearlings. During courtship, male juncos spread their tails, revealing their tail white, and a previous experiment has shown that males with experimentally enhanced tail white are more attractive to females. To determine whether females with experimentally enhanced tail white would be preferred by males, we clipped and replaced tail feathers of females, creating a control group with low to natural levels of tail white and an enhanced group with high levels. We tested preference in a mate choice apparatus like that used previously and found that males courted both control and enhanced females and displayed individual preferences but showed no collective preference for members of either category of females. Because we found neither a preference for trait values that indicate greater age/experience (experimentally enhanced females) nor a preference for less male-like appearance (control females), our results are inconsistent with a role for male mate choice in the maintenance of tail white in females. Female tail white may be subject to selection in another context or persist owing to a genetic correlation between the sexes. Regardless, the sexes apparently prefer different trait values, which suggests that preferences are expressed independently in males and females.

Offspring sex ratio is unrelated to male attractiveness in dark-eyed juncos (Junco hyemalis)

Abstract. Sex allocation theory predicts that parents should bias investment toward the offspring sex that confers higher relative fitness on the parents. When variance in reproductive success is higher in males than females, and some males are more attractive to females than others, thereby achieving higher reproductive success, female parents mated to attractive males are expected to bias reproductive allocation toward sons. Modification of the primary sex ratio is one mechanism by which avian parents may bias allocation. In mate choice trials, captive female dark-eyed juncos (Junco hyemalis) prefer to associate with males whose circulating testosterone level has been elevated experimentally. We asked whether females socially mated to testosterone-treated males in nature might bias the sex ratio of progeny in favor of sons. We determined the primary sex ratio of broods using a sex-linked molecular marker, the CHD gene on the W chromosome. We found no relationship between the hormonal treatment of males and the primary or secondary sex ratios of offspring produced by their social mates. Sex ratio was also unrelated to breeding-season date, study year, male viability, or female age. While unlikely, it is possible that female juncos are not able to manipulate the primary sex ratio of their broods. More likely, possible benefits to female juncos of producing attractive sons may be small because (1) attractive males do not necessarily have higher fitness, depending on the trade-offs involved for males, and (2) when such benefits exist, they may be outweighed for females by the costs of compensating for the reduced paternal care of attractive males.

Hyperthermia exacerbates ischaemic brain injury

Hyperthermia frequently occurs in stroke patients. Hyperthermia negatively correlates with clinical outcome and adversely effects treatment regiments otherwise successful under normothermic conditions. Preclinical studies also demonstrate that hyperthermia converts salvageable penumbra to ischaemic infarct. The present article reviews the knowledge accumulated from both clinical and preclinical studies about hyperthermia and ischaemic brain injury, examines current treatment strategies and discusses future research directions.

The decline in yolk progesterone concentrations during incubation is dependent on embryonic development in the European starling

Oviparous amniotes, particularly birds, have become model systems in which to study how mothers may utilize steroids to adaptively adjust offspring development. Although there is now ample evidence that maternally derived steroids in the egg at oviposition can influence offspring phenotype, very little is known about how these steroids elicit such effects. Of the major avian steroid hormones found in yolk, progesterone is by far the most abundant at oviposition, but has received little research attention to date. In this study, we examine the metabolism of [(3)H]-progesterone injected into freshly laid European starling eggs throughout the first 5 days of development by characterization of radioactivity within the egg homogenate. We also introduce a technique that utilizes a focal, freeze/thaw cycle to prevent embryonic development and allows us to assess the role of the embryo in metabolizing progesterone during early incubation. Two major findings result. First is that [(3)H]-progesterone is metabolized in eggs possessing a developing embryo, but not in eggs with disrupted embryonic development. Second is that the change in the distribution of radioactivity within eggs possessing an embryo is the result of metabolism of [(3)H]-progesterone to a more polar form that is subsequently conjugated. Together, these data suggest live embryos are necessary for metabolism of progesterone during early incubation, underscoring the potentially important contribution of embryos to functional modulation or mediation of maternal yolk steroid effects.