A proposed role of the sulfotransferase/sulfatase pathway in modulating yolk steroid effects

Steroid hormones have long been studied by behavioral ecologistsas a nongenetic means whereby females can influence the developmentof their offspring. In oviparous vertebrates, steroids are presentin the yolk at the time of oviposition and have been shown toaffect numerous traits of the offspring. To date, most studieshave focused on the functional relationship between yolk steroidsand offspring development. In this article we used a mechanisticapproach to investigate the effects of yolk steroids in an attemptto decipher how lipophilic steroids may make it from the lipid-richyolk to the developing embryo. First, we examined the distributionof radioactive and nonradioactive estradiol following the exogenousapplication of each to developing eggs of the red-eared slider.Second, we quantified sulfotransferase activity in various componentsof the egg as a potential mechanism for the metabolism of steroids.Results indicate that exogenous estradiol is converted to awater-soluble form during the first 15 days of development,concurrent with an increase of sulfotransferase activity inthe yolk and extra-embryonic membranes. Based on these data,we propose a mechanistic model based upon the sulfotransferase/sulfatasepathway as a means through which developing eggs can convertsteroids to a water-soluble form that can be transported tothe embryo. These sulfonated steroids may then serve as precursorsfor subsequent steroid production via sulfatase activity. Thismodel utilizes a mechanism known to be important for the modulationof maternal steroid signals in placental mammals, at the sametime addressing several previously unanswered questions regardingthe mechanisms underlying the effects of yolk steroids.     

Ecdysteroids During Embryogenesis in Locusta migratoria

SYNOPSIS. Ovaries of Locusta migratoria synthesize large amountsof ecdysteroids at the end of oöcyte maturation. The predominantecdysteroids in mature ovaries are conjugated 2-deoxyecdysone(100 µM) and conjugated ecdysone (50 µM) which outnumberthe corresponding free compounds by 50–100 fold. Thesevarious ecdysteroids persist during ovulation and are recoveredfrom newly-laid eggs. The conjugated maternal ecdysteroids aregradually metabolized as embryonic development proceeds; theyhave disappeared as such on day 6 after oviposition, that isafter blastokinesis and shortly after dorsal closure. Concomitantlyto this metabolism of the maternal conjugated ecdysteroids,other ecdysteroid conjugates appear in the eggs which have differentchromatographic behaviors and some of which are conjugates ofecdysone metabolites formed by the embryo. The data availableso far are compatible with the hypothesis that the maternalconjugates are hydrolysed to free 2-deoxyecdysone and ecdysoneby the embryo during early stages of development and subsequentlyconjugated to inactivation compounds. During the later stagesof embryonic development however, a de novo synthesis of ecdysoneis probable, the maternal conjugates having been metabolizedduring the earlier stages.     

Elevated corticosterone during egg production elicits increased maternal investment and promotes nestling growth in a wild songbird

Glucocorticoids circulating in breeding birds during egg production accumulate within eggs, and may provide a potent form of maternal effect on offspring phenotype. However, whether these steroids affect offspring development remains unclear. Here, we employed a non-invasive technique that experimentally elevated the maternal transfer of corticosterone to eggs in a wild population of house wrens. Feeding corticosterone-injected mealworms to free-living females prior to and during egg production increased the number of eggs that females produced and increased corticosterone concentrations in egg yolks. This treatment also resulted in an increase in the amount of yolk allocated to eggs. Offspring hatching from these eggs begged for food at a higher rate than control offspring and eventually attained increased prefledging body condition, a trait predictive of their probability of recruitment as breeding adults in the study population. Our results indicate that an increase in maternal glucocorticoids within the physiological range can enhance maternal investment and offspring development.     

Stressed mothers lay eggs with high corticosterone levels which produce low-quality offspring

Organisms frequently encounter stressful ecological conditions. In vertebrates, a major mechanism of physiological response to stress is mediated by the hypothalamic-pituitary-adrenal axis and results in increased secretion of glucocorticosteroids, which can have adverse consequences on diverse phenotypic traits affecting fitness. Maternal stress may thus have carry-over effects on progeny if it influences pre-natal offspring environment in terms of glucocorticosteroid concentration, although this hypothesis has never been tested in any species under field conditions. We manipulated stress experienced by female barn swallows Hirundo rustica, by exposing them to a predator during laying and measured egg corticosterone concentration. Stressed females laid eggs with greater corticosterone concentration than controls exposed to a herbivore. In another experiment, we injected physiological doses of corticosterone in the egg albumen and compared the phenotype of offspring originating from these eggs with their control siblings originating from either sham-inoculated or unmanipulated eggs and reared in the same nest. Eggs injected with corticosterone had lower hatchability and produced fledglings with smaller body size and slower plumage development than did control eggs. Nestling body size in our study population predicts long-term survival. Thus, maternal stress impaired offspring phenotype and viability by increasing transmission of glucocorticosteroids to the eggs. This study identifies a novel mechanism mediating early maternal effects whereby maternal stress affects offspring quality. These results are relevant to biological conservation because they disclose a mechanism that can link environmental conditions to population productivity and viability.     

Maternal deposition of yolk corticosterone in clutches of southern rockhopper penguins (Eudyptes chrysocome chrysocome)

High corticosterone levels can have deleterious effects in developing avian embryos and chicks. Therefore, it may be adaptive for avian mothers to reduce corticosterone transfer to their eggs. However, until now, data about the active or/and passive role of mothers in corticosterone transfer to eggs are inconclusive. Here, we study maternal investment into A- and B-eggs of southern rockhopper penguins (Eudyptes chrysocome chrysocome). This species exhibits reversed hatching asynchrony and provides a unique model to test whether there is a strategic investment in corticosterone favoring the B-chick, which is most likely to survive. We found that rockhopper penguins had the highest yolk concentrations of any wild bird species studied so far. Contrary to our expectations, B-eggs had more yolk corticosterone both in concentration and in quantity than A-eggs, independently of the laying period and the level of human disturbance. Additionally, females deposited more yolk corticosterone in their eggs when they were disturbed. However, this disturbance effect was particularly strong for A-eggs and for late-laid eggs. The present data support neither the predictions for an active regulation nor for a passive deposition, and hormone deposition mechanisms still need to be explored. The adaptive value, if any, of high yolk corticosterone is presently unknown.     

Direct Exposure to Corticosterone During Embryonic Development Influences Behaviour in an Ovoviviparous Lizard

It is becoming increasingly clear that conditions experienced during embryonic development can be of major importance for traits subsequent to parturition or hatching. For example, in mammals, offspring from stressed mothers show a variety of changes in behavioural, morphological, and life‐history traits. The effects of maternal stress on trait development are believed to be mediated via transfer of glucocorticoids, the main hormones released during the stress response, from mother to offspring. However, also other physiological maternal responses during stress could be responsible for changes in offspring phenotype. We investigated the direct effects of corticosterone on offspring development, without other confounding factors related to increased maternal stress, by injection of corticosterone in eggs of the ovoviviparous lizard Lacerta vivipara. Corticosterone‐manipulated offspring did not show impaired development, reduced body size or body condition at parturition. However, corticosterone‐treated offspring showed altered anti‐predator behaviour, as measured by the time required to emerge from shelter after a simulated predator attack. Differential steroid exposure during development, possibly mediated by maternal stress response, may explain some of the variation in behaviour among individuals in natural populations.     

Eggshell Appearance Does Not Signal Maternal Corticosterone Exposure in Japanese Quail: An Experimental Study with Brown-Spotted Eggs

Reproduction is a critical period for birds as they have to cope with many stressful events. One consequence of an acute exposure to stress is the release of corticosterone, the avian stress hormone. Prolonged stress can have negative impacts on the immune system, resulting in, for example, increased oxidative stress. Through maternal effects, females are known to modulate their investment in eggs content according to their own physiological condition. Less is known about maternal investment in eggshells, especially in pigments. The two main eggshell pigments may possess opposite antioxidant properties: protoporphyrin (brown) is a pro-oxidant, whereas biliverdin (blue-green) is an antioxidant. In Japanese quail, we know that the deposition of both pigments is related to female body condition. Thus, a chronic stress response may be reflected in eggshell coloration. Using female Japanese quails that lay brown-spotted eggs, we explored whether physiological exposure to corticosterone induces a change in female basal stress and antioxidant factors, and eggshell pigment concentration, spectrophotometric reflectance, and maculation coverage. We supplemented adult females over a 2 week period with either peanut oil (control) or corticosterone (treatment). We collected pre- and post-supplementation eggs and analysed the effect of corticosterone treatment on female physiology and eggshell appearance parameters. Except for corticosterone-fed birds which laid eggs with brighter spots, supplementation had no significant effect on female physiology or eggshell pigment concentration, reflectance and maculation. The change in eggshell spot brightness was not detected by a photoreceptor noise-limited color opponent model of avian visual perception. Our data confirms that eggshell reflectance in spotted eggs varies over the laying sequence, and spot reflectance may be a key factor that is affected by females CORT exposure, even if the changes are not detected by an avian visual model.     

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.     

Hormone-mediated maternal effects in birds: mechanisms matter but what do we know of them?

Over the past decade, birds have proven to be excellent models to study hormone-mediated maternal effects in an evolutionary framework. Almost all these studies focus on the function of maternal steroid hormones for offspring development, but lack of knowledge about the underlying mechanisms hampers further progress. We discuss several hypotheses concerning these mechanisms, point out their relevance for ecological and evolutionary interpretations, and review the relevant data. We first examine whether maternal hormones can accumulate in the egg independently of changes in hormone concentrations in the maternal circulation. This is important for Darwinian selection and female physiological trade-offs, and possible mechanisms for hormone accumulation in the egg, which may differ among hormones, are reviewed. Although independent regulation of plasma and yolk concentrations of hormones is conceivable, the data are as yet inconclusive for ovarian hormones. Next, we discuss embryonic utilization of maternal steroids, since enzyme and receptor systems in the embryo may have coevolved with maternal effect mechanisms in the mother. We consider dose-response relationships and action pathways of androgens and argue that these considerations may help to explain the apparent lack of interference of maternal steroids with sexual differentiation. Finally, we discuss mechanisms underlying the pleiotropic actions of maternal steroids, since linked effects may influence the coevolution of parent and offspring traits, owing to their role in the mediation of physiological trade-offs. Possible mechanisms here are interactions with other hormonal systems in the embryo. We urge endocrinologists to embark on suggested mechanistic studies and behavioural ecologists to adjust their interpretations to accommodate the current knowledge of mechanisms.     

Maternal source of variability in the embryo development of an annual killifish

Organisms inhabiting unpredictable environments often evolve diversified reproductive bet‐hedging strategies, expressed as production of multiple offspring phenotypes, thereby avoiding complete reproductive failure. To cope with unpredictable rainfall, African annual killifish from temporary savannah pools lay drought‐resistant eggs that vary widely in the duration of embryo development. We examined the sources of variability in the duration of individual embryo development, egg production and fertilization rate in Nothobranchius furzeri. Using a quantitative genetics approach (North Carolina type II design), we found support for maternal effects rather than polyandrous mating as the primary source of the variability in the duration of embryo development. The number of previously laid eggs appeared to serve as an internal physiological cue initiating a shift from rapid‐to‐slow embryo developmental mode. In annual killifish, extensive phenotypic variability in progeny traits is adaptive, as the conditions experienced by parents have limited relevance to the offspring generation. In contrast to genetic control, with high phenotypic expression and heritability, maternal control of traits under natural selection prevents standing genetic diversity from potentially detrimental effects of selection in fluctuating environments.     

Steroid hormone metabolism by the chorioallantoic placenta of the mountain spiny lizard Sceloporus jarrovi as a possible mechanism for buffering maternal-fetal hormone exchange

The placenta provides a maternal-fetal exchange interface that maximizes the diffusion of gases, nutrients, and wastes. However, the placenta also may permit diffusion of lipid-soluble steroid hormones that influence processes such as sex-specific fetal development and maternal pregnancy maintenance. In mammals, placental steroid metabolism contributes to regulation of maternal and fetal hormone levels. Such mechanisms may be less highly developed in species that have recently evolved placentation, such as many reptiles. We therefore chose to investigate placental metabolism of steroids in the viviparous lizard Sceloporus jarrovi. In vitro tissue incubations tested the abilities of the chorioallantoic placenta to clear progesterone and corticosterone by converting them to other metabolites and to synthesize progesterone. Placental tissue rapidly cleared progesterone and corticosterone added to the incubation media, indicating that the tissue had converted the steroids to other products. Placental tissue also synthesized substantial concentrations of progesterone from the prohormone pregnenolone. Thus, even in a species with a simple, recently evolved placenta, steroid metabolism appears to be highly developed and could be critical for regulation of maternal and fetal hormone levels. This finding suggests that placental hormone metabolism may be critical to the successful evolution of placentation.     

Determinants of within- and among-clutch variation in yolk corticosterone in the European starling

Maternal glucocorticoids are known to affect offspring phenotype in numerous vertebrate taxa. In birds, the maternal transfer of corticosterone to eggs was recently proposed as a hormonal mechanism by which offspring phenotype is matched to the relative quality of the maternal environment. However, current hypotheses lack supporting information on both intra- and inter-clutch variation in yolk corticosterone for wild birds. As such, we examined variation in yolk corticosterone levels in a wild population of European starlings (Sturnus vulgaris). Maternal condition, clutch size and nesting density were all negatively related to yolk corticosterone deposition; females with high condition indices, those laying larger clutches and those nesting in high-density associations deposited lower amounts of the hormone into eggs than those with low condition indices, laying small clutches and nesting in isolation. Alternatively, we found no effects of maternal age or human disturbance on yolk corticosterone deposition. Intra-clutch variation of yolk corticosterone was significant, with levels increasing across the laying sequence in all clutch sizes examined, with the difference between first and last-laid eggs being greater in large versus small clutches. Given the reported effects of yolk corticosterone on offspring size and growth, intra-clutch variation in yolk corticosterone has the potential to alter the competitive environment within a brood. Furthermore, our results indicate that variation in yolk corticosterone can originate from variation in both the mother's quality as well as the quality of her breeding environment. The presence of inter-female variation in particular is an important pre-requisite in testing whether the exposure of offspring to maternally-derived corticosterone is a mechanistic link between offspring phenotypic plasticity and maternal quality.     

Maternal allocation of androgens and antagonistic effects of yolk androgens on sons and daughters

Mothers can influence the phenotype of their offspring by adjustingthe quality of their eggs in relation to sex and reproductivevalue of the progeny. Maternal androgens in the eggs of vertebratesmay mediate such adaptive early maternal effects. However, theevolution of early maternal effects mediated by egg androgensmay be constrained by the inability of mothers to differentiallyallocate androgens to eggs with a male or a female if androgenshave different effects on sons and daughters. In this study,we increased the concentration of androgens in the eggs of barnswallows (Hirundo rustica) within the physiological range ofvariation and analyzed the effect on nestling growth and beggingbehavior. Egg androgens increased body size and mass of sonsbut reduced these characters in daughters when compared to twocontrol groups in a repeated-measures analysis of variance ofdata collected at different ages. However, the differentialeffect of androgen on the two sexes was no longer significantwhen the analysis was restricted to the age of 12 days, whenfinal body size is attained. In a second experiment, we testedwhether mothers differentially allocated androgens to eggs withsons rather than daughters while manipulating a paternal secondarysexual character. Androgen concentration did not vary in relationto paternal ornamentation or embryo sex. Hence, antagonisticeffects of egg androgens on sons and daughters may exist inthe very early posthatching life and may constrain the evolutionof adaptive maternal effects because mothers do not differentiallyallocate androgens in relation to embryo sex.     

Dynamics of maternal and paternal effects on embryo and seed development in wild radish (Raphanus sativus)

Background and Aims

Variability in embryo development can influence the rate of seed maturation and seed size, which may have an impact on offspring fitness. While it is expected that embryo development will be under maternal control, more controversial hypotheses suggest that the pollen donor and the embryo itself may influence development. These latter possibilities are, however, poorly studied. Characteristics of 10-d-old embryos and seeds of wild radish (Raphanus sativus) were examined to address: (a) the effects of maternal plant and pollen donor on development; (b) the effects of earlier reproductive events (pollen tube growth and fertilization) on embryos and seeds, and the influence of embryo size on mature seed mass; (c) the effect of water stress on embryos and seeds; (d) the effect of stress on correlations of embryo and seed characteristics with earlier and later reproductive events and stages; and (e) changes in maternal and paternal effects on embryo and seed characteristics during development.

Methods

Eight maternal plants (two each from four families) and four pollen donors were crossed and developing gynoecia were collected at 10 d post-pollination. Half of the maternal plants experienced water stress. Characteristics of embryos and seeds were summarized and also compared with earlier and later developmental stages.

Key Results

In addition to the expected effects of the maternal plants, all embryo characters differed among pollen donors. Paternal effects varied over time, suggesting that there are windows of opportunity for pollen donors to influence embryo development. Water-stress treatment altered embryo characteristics; embryos were smaller and less developed. In addition, correlations of embryo characteristics with earlier and later stages changed dramatically with water stress.

Conclusions

The expected maternal effects on embryo development were observed, but there was also evidence for an early paternal role. The relative effects of these controls may change over time. Thus, there may be times in development when selection on the maternal, paternal or embryo contributions to development are more and less likely.     

Maternally derived hormones,neurosteroids and the development of behaviour

In a wide range of taxa, there is evidence that mothers adaptively shape the development of offspring behaviour by exposing them to steroids. These maternal effects have major implications for fitness because, by shaping early development, they can permanently alter how offspring interact with their environment. However, theory on parent–offspring conflict and recent physiological studies showing that embryos rapidly metabolize maternal steroids have placed doubt on the adaptive significance of these hormone-mediated maternal effects. Reconciling these disparate perspectives requires a mechanistic understanding of the pathways by which maternal steroids can influence neural development. Here, we highlight recent advances in developmental neurobiology and psychiatric pharmacology to show that maternal steroid metabolites can have direct neuro-modulatory effects potentially shaping the development of neural circuitry underlying ecologically relevant behavioural traits. The recognition that maternal steroids can act through a neurosteroid pathway has critical implications for our understanding of the ecology and evolution of steroid-based maternal effects. Overall, compared to the classic view, a neurosteroid mechanism may reduce the evolutionary lability of hormone-mediated maternal effects owing to increased pleiotropic constraints and frequently influence long-term behavioural phenotypes in offspring.     

Fate of maternal conjugated ecdysteroids during embryonic development in Locusta migratoria

Newly laid eggs of Locusta migratoria contain impressively high concentrations of conjugated 2-deoxyecdysone and conjugated ecdysone of maternal origin. These molecules are metabolized during embryonic development, the changes concerning not only the ecdysteroid genins but also the conjugating moieties. In the present paper the fates of the maternal conjugates were followed during embryogenesis in the eggs. The conjugates were separated both by silica gel TLC and reverse-phase HPLC and measured, before and after hydrolysis, by RIA. Fluctuations of radioactive ecdysteroid conjugates were also investigated in eggs laid by females subjected to massive injections of tritiated cholesterol. The results are discussed in relation to recent data on identification of ecdysteroid conjugates in Locusta and a model for the sequences of metabolic events leading from maternal ecdysteroid conjugates to the embryonic ecdysteroids is proposed.     

Maternal effects mediated by egg quality in the Yellow-legged Gull Larus michahellis in relation to laying order and embryo sex

Background

Maternal effects mediated by egg size and quality may profoundly affect offspring development and performance, and mothers may adjust egg traits according to environmental or social influences. In avian species, context-dependency of maternal effects may result in variation in egg composition, as well as in differential patterns of covariation among selected egg components, according to, for example, position in the laying sequence or offspring sex. We investigated variation in major classes of egg yolk components (carotenoids, vitamins and steroid hormones) in relation to egg size, position in the laying sequence and embryo sex in clutches of the Yellow-legged Gull (Larus michahellis). We also investigated their covariation, to highlight mutual adjustments, maternal constraints or trade-offs in egg allocation.

Results

Laying sequence-specific patterns of allocation emerged: concentration of carotenoids and vitamin E decreased, while concentrations of androgens increased. Vitamin A, estradiol and corticosterone did not show any change. There was no evidence of sex-specific allocation or covariation of yolk components. Concentrations of carotenoids and vitamins were positively correlated. Egg mass decreased along the laying sequence, and this decrease was negatively correlated with the mean concentrations of carotenoids in clutches, suggesting that nutritionally constrained females lay low quality clutches in terms of carotenoid content. Finally, clutches with smaller decline in antioxidants between first- and last-laid eggs had a larger increase in yolk corticosterone, suggesting that a smaller antioxidant depletion along the laying sequence may entail a cost for laying females in terms of increased stress levels.

Conclusions

Since some of the analyzed yolk components (e.g. testosterone and lutein) are known to exert sex-specific phenotypic effects on the progeny in this species, the lack of sex-specific egg allocation by mothers may either result from trade-offs between contrasting effects of different egg components on male and female offspring, or indicate that sex-specific traits are controlled primarily by mechanisms of sexual differentiation, including endogenous hormone production or metabolism of exogenous antioxidants, during embryonic development.