Gender bent but not in the mind of fish

There is strong evidence that environmental exposure to endocrine disrupting chemicals (EDCs) is resulting in significant alterations to the reproductive system of many wildlife populations. Most of these studies measure chemically induced changes to the endocrine system or reproductive morphology and mostly provide simple markers of exposure. The heightened concern over the effects of EDCs is however primarily driven by the hypothesis that this disruption may have serious deleterious consequences on reproductive success. In extensive laboratory studies on breeding populations of zebrafish, I have shown that multigenerational exposure to environmentally relevant levels of endocrine disruptors cause very significant reductions in reproductive success. Lifetime exposure to 5 ng/l of ethynylestradiol, for example, caused complete reproductive failure. These reproductive failures were not caused by exposure proximate to the timing of spawning but by the disruption of development during earlier embryonic and larval sexual differentiation. Histology revealed that male gonads had not differentiated into functional testes. Significantly, these sterile males still initiated spawning in females and resulted in unfertilized eggs. This differential in the sensitivity of behaviour compared to gonadal disruption raises important issues in understanding the implications of endocrine disruption in wild populations. Moreover, the particular mode of reproduction behaviour that is used by a species fundamentally affects the population level impact of endocrine disruptors. This paper will discuss these results and how a greater understanding of the dynamics of group spawning may help in assessing the potential impact of endocrine disruption     

Bridging epidemiology and model organisms to increase understanding of endocrine disrupting chemicals and human health effects

Concerning temporal trends in human reproductive health has prompted concern about the role of environmentally mediated risk factors. The population is exposed to chemicals present in air, water, food and in a variety of consumer and personal care products, subsequently multiple chemicals are found human populations around the globe. Recent reviews find that endocrine disrupting chemicals (EDCs) can adversely affect reproductive and developmental health. However, there are still many knowledge gaps. This paper reviews some of the key scientific concepts relevant to integrating information from human epidemiologic and model organisms to understand the relationship between EDC exposure and adverse human health effects. Additionally, areas of new insights which influence the interpretation of the science are briefly reviewed, including: enhanced understanding of toxicity pathways; importance of timing of exposure; contribution of multiple chemical exposures; and low dose effects. Two cases are presented, thyroid disrupting chemicals and anti-androgens chemicals, which illustrate how our knowledge of the relationship between EDCs and adverse human health effects is strengthened and data gaps reduced when we integrate findings from animal and human studies.     

Endocrine disruption in aquatic systems: up‐scaling research to address ecological consequences

Endocrine‐disrupting chemicals (EDCs) can alter biological function in organisms at environmentally relevant concentrations and are a significant threat to aquatic biodiversity, but there is little understanding of exposure consequences for populations, communities and ecosystems. The pervasive nature of EDCs within aquatic environments and their multiple sub‐lethal effects make assessments of their impact especially important but also highly challenging. Herein, we review the data on EDC effects in aquatic systems focusing on studies assessing populations and ecosystems, and including how biotic and abiotic processes may affect, and be affected by, responses to EDCs. Recent research indicates a significant influence of behavioural responses (e.g. enhancing feeding rates), transgenerational effects and trophic cascades in the ecological consequences of EDC exposure. In addition, interactions between EDCs and other chemical, physical and biological factors generate uncertainty in our understanding of the ecological effects of EDCs within aquatic ecosystems. We illustrate how effect thresholds for EDCs generated from individual‐based experimental bioassays of the types commonly applied using chemical test guidelines [e.g. Organisation for Economic Co‐operation and Development (OECD)] may not necessarily reflect the hazards associated with endocrine disruption. We argue that improved risk assessment for EDCs in aquatic ecosystems urgently requires more ecologically oriented research as well as field‐based assessments at population‐, community‐ and food‐web levels.     

An endocrine disrupter increases growth and risky behavior in threespined stickleback (Gasterosteus aculeatus)

There is considerable concern that endocrine disrupting chemicals (EDCs) can affect wildlife and humans. While several studies have reported that acute exposure to EDCs can cause changes in reproductive traits, we are in the early stages of discerning whether such changes have significant deleterious fitness consequences. In this study, chronic exposure of threespined stickleback (Gasterosteus aculeatus) to an environmentally relevant level of an EDC used in the birth control pill and post-menopausal hormone replacement therapy produced changes in growth and behavior that were related to fitness. Exposure to 100 ng/l ethinyl estradiol accelerated growth rate and increased levels of behavior that makes individuals more susceptible to predation (activity and foraging under predation risk). Moreover, the costs of exposure to ethinyl estradiol took their ultimate toll via mortality later in life, and were particularly high for females and for one population. The ecological approach taken in this work revealed heretofore unexamined effects of EDCs and has direct implications for the way we evaluate the impact of EDCs in the environment.     

Endocrine Toxicants and Reproductive Success in Fish

There is compelling evidence on a global scale for compromised growth and reproduction, altered development, and abnormal behaviour in feral fish that can be correlated or in some cases causally linked with exposure to endocrine disrupting chemicals (EDCs). Attributing cause and effect relationships for EDCs is a specific challenge for studies with feral fish as many factors including food availability, disease, competition and loss of habitat also affect reproduction and development. Even in cases where there are physiological responses of fish exposed to EDCs (e.g., changes in reproductive hormone titres, vitellogenin levels), the utility of these measures in extrapolating to whole animal reproductive or developmental outcomes is often limited. Although fish differ from other vertebrates in certain aspects of their endocrinology, there is little evidence that fish are more sensitive to the effects of EDCs. Therefore, to address why endocrine disruption seems so widespread in fish, it is necessary to consider aspects of fish physiology and their environment that may increase their exposure to EDCs. Dependence on aquatic respiration, strategies for iono-osmotic regulation, and maternal transfer of contaminants to eggs creates additional avenues by which fish are exposed to EDCs. This paper provides an overview of responses observed in feral fish populations that have been attributed to EDCs and illustrates many of the factors that need consideration in evaluating the risks posed by these chemicals.     

Altered reproductive success in rat pairs after environmental-like exposure to xenoestrogen

Endocrine-disrupting compounds (EDCs) have the capacity of altering the normal function of the endocrine system. EDCs have shown dramatic effects on the reproductive biology of aquatic wildlife and may affect human reproduction as well. Studies on EDCs in mammalian species have often investigated the effects of short-term, high doses on male and female reproductive physiology. However, it is difficult to predict from such studies the effects of EDC on populations that are exposed to very low doses throughout their life via contaminated food and water. We studied the effects of EDC on mammalian reproduction with an environmental-like protocol where the endpoint is the reproductive success of exposed pairs. We focused on a subclass of EDC, the xenoestrogens, which mimic the action of natural oestrogen hormones. Male and female rats were exposed to low doses of the pure oestrogen, ethynyloestradiol, during development, by oral administration to their mothers during pregnancy and lactation, and to them until puberty. We evaluated the effects of the exposure on development and reproductive physiology of individuals, and on fertility and fecundity of pairs in which both members had been exposed to the same treatment. We found that low doses caused major reproductive deficits in the experimental animals. Very low, environmentally relevant doses did not have evident effects on exposed animals; however, the fecundity of exposed pairs was substantially altered. Environmentally relevant doses of xenoestrogens which have no evident physiological effects can alter the reproductive success of exposed pairs in natural populations.     

Consequences of in ovo exposure to p,p'-DDE on reproductive development and function in Japanese quail

This study was conduced to assess the effects of a one time embryonic exposure to p,p'-DDE (dichlorodiphenyldichloroethylene; DDE) on the reproductive development and function in Japanese quail (Coturnix japonica). Embryos were exposed at day one of incubation to either 20 or 40 microg DDE or a sesame oil vehicle control (injection volume=20 microl). Onset of puberty, gonadal histopathology, sperm motility, cloacal gland size, and male copulatory behavior were assessed in adults. DDE accelerated onset of puberty in females and reduced male reproductive behaviors. Gonadal morphology and sperm motility appeared to be unaffected. Results from this study provide evidence that the neuroendocrine system may be more sensitive and less resilient to embryonic exposure to contaminants than traditional measures of reproductive success following contaminant exposure in adults. This study further supports the inclusion of behavioral assessments in toxicity tests.     

Effects of 4-n-nonylphenol and 17beta-oestradiol on early development of the barnacle Elminius modestus

Pollutants that are present in the aquatic environment and cause abnormal endocrine function in wildlife populations have been termed endocrine disrupting chemicals (EDCs). The impacts of these chemicals on the reproduction and development of vertebrates has been shown to be significant in both field studies and laboratory experiments. Over the past decade the number of investigations into the impacts of EDCs that affect reproductive and sexual characteristics (reproductive EDCs) has increased and evidence of their potency is evident in numerous wildlife species and through data from in vitro tests. However, little information is available on whether chemicals which act as EDCs in vertebrate species affect aquatic invertebrates. The case of imposex in archeogastropods following exposure to tributyltin (TBT) is a notable exception. Moreover, a number of studies have shown that development, fecundity and reproductive output of some aquatic invertebrates are affected significantly by exposure to pollutants. In order to determine whether external signs of exposure to vertebrate EDCs can be observed and monitored in invertebrate species, we exposed larvae of the barnacle Elminius modestus to environmentally realistic concentrations of the xeno-oestrogen, 4-n-nonylphenol (NP), and the natural oestrogen, 17beta-oestradiol (E(2)). Early life stages (nauplii and cyprids) were also exposed in the laboratory to determine whether there were effects on the timing of larval development and settlement. Ovary development and size of juveniles was measured following chronic exposure. Exposure to NP in the concentration range 0.01-10 μg l(-1) resulted in disruption of the timing of larval development. Similar results were obtained with E(2). Pulse exposures showed that the timing of exposure is critical and exposures for a period of 12 months caused long-term effects. A linear, concentration-dependent response was not evident.     

A Computational Model to Predict Rat Ovarian Steroid Secretion from In Vitro Experiments with Endocrine Disruptors

A finely tuned balance between estrogens and androgens controls reproductive functions, and the last step of steroidogenesis plays a key role in maintaining that balance. Environmental toxicants are a serious health concern, and numerous studies have been devoted to studying the effects of endocrine disrupting chemicals (EDCs). The effects of EDCs on steroidogenic enzymes may influence steroid secretion and thus lead to reproductive toxicity. To predict hormonal balance disruption on the basis of data on aromatase activity and mRNA level modulation obtained in vitro on granulosa cells, we developed a mathematical model for the last gonadal steps of the sex steroid synthesis pathway. The model can simulate the ovarian synthesis and secretion of estrone, estradiol, androstenedione, and testosterone, and their response to endocrine disruption. The model is able to predict ovarian sex steroid concentrations under normal estrous cycle in female rat, and ovarian estradiol concentrations in adult female rats exposed to atrazine, bisphenol A, metabolites of methoxychlor or vinclozolin, and letrozole.     

The effects of endocrine disruptors on the male germline: an intergenerational health risk

Environmental pollution is becoming one of the major concerns of society. Among the emerging contaminants, endocrine-disrupting chemicals (EDCs), a large group of toxicants, have been the subject of many scientific studies. Besides the capacity of these compounds to interfere with the endocrine system, they have also been reported to exert both genotoxic and epigenotoxic effects. Given that spermatogenesis is a coordinated process that requires the involvement of several steroid hormones and that entails deep changes in the chromatin, such as DNA compaction and epigenetic remodelling, it could be affected by male exposure to EDCs. A great deal of evidence highlights that these compounds have detrimental effects on male reproductive health, including alterations to sperm motility, sexual function, and gonad development. This review focuses on the consequences of paternal exposure to such chemicals for future generations, which still remain poorly known. Historically, spermatozoa have long been considered as mere vectors delivering the paternal haploid genome to the oocyte. Only recently have they been understood to harbour genetic and epigenetic information that plays a remarkable role during offspring early development and long-term health. This review examines the different modes of action by which the spermatozoa represent a key target for EDCs, and analyses the consequences of environmentally induced changes in sperm genetic and epigenetic information for subsequent generations.     

Endocrine disruptors and female fertility: Focus on (bovine) ovarian follicular physiology

Throughout the previous century, the production, use and, as a result, presence of chemicals in the environment increased enormously. Consequently, humans and animals are exposed to a wide variety of chemical substances of which some possess the ability to disrupt the endocrine system in the body, thereby denominated as “endocrine disrupting chemicals” (EDCs) or “endocrine disruptors”. Because the reproductive system is a target organ for endocrine disruption, EDCs are postulated as one of the possible causes of human subfertility. Within the reproductive system, the ovarian follicle can be considered as an extremely fragile microenvironment where interactions between the oocyte and its surrounding somatic cells are essential to generate a fully competent oocyte. In this review, we explore how EDCs can interfere with the well-balanced conditions in the ovarian follicle. In addition, we highlight the bovine ovarian follicle as an alternative in vitro model for EDC and broader toxicology research.     

Sex in troubled waters: Widespread agricultural contaminant disrupts reproductive behaviour in fish

Chemical pollution is a pervasive and insidious agent of environmental change. One class of chemical pollutant threatening ecosystems globally is the endocrine disrupting chemicals (EDCs). The capacity of EDCs to disrupt development and reproduction is well established, but their effects on behaviour have received far less attention. Here, we investigate the impact of a widespread androgenic EDC on reproductive behaviour in the guppy, Poecilia reticulata. We found that short-term exposure of male guppies to an environmentally relevant concentration of 17β-trenbolone—a common environmental pollutant associated with livestock production—influenced the amount of male courtship and forced copulatory behaviour (sneaking) performed toward females, as well as the receptivity of females toward exposed males. Exposure to 17β-trenbolone was also associated with greater male mass. However, no effect of female exposure to 17β-trenbolone was detected on female reproductive behaviour, indicating sex-specific vulnerability at this dosage. Our study is the first to show altered male reproductive behaviour following exposure to an environmentally realistic concentration of 17β-trenbolone, demonstrating the possibility of widespread disruption of mating systems of aquatic organisms by common agricultural contaminants.     

Determining Indicators of Exposure and Effects for Endocrine Disrupting Chemicals (EDCs): An Introduction

Endocrine disruptors are characterized by their influence on animal endocrine systems resulting in reproductive, developmental, neurological, and immune dysfunction. The purpose of this overview is to provide the reader with a sense of the activities within the U.S. Environmental Protection Agency (USEPA), in particular NHEERL, that address the many facets of research on endocrine disrupting chemicals (EDCs) and to highlight the approach being taken at the different organizational levels within the USEPA, including screening, testing and evaluating endocrine disrupting chemicals. As a part of this endeavor, the USEPA continues to evaluate the current research activities in order to better understand and refine the process of risk characterization of EDCs. Thus, the participants in this session were asked to review their research within the framework of a better identification of EDC effects, better characterization of those compounds that have endocrine disrupting activity and how to incorporate this information into the risk assessment paradigm. Specifically, the goals of the ensuing papers were to compare individual vs. population indicators of endocrine disrupting effects, examine comparable and multiple mechanisms of toxicity, and describe the use of effects as indicators to identify toxicants and their sources. Mammalian and fish reproductive endpoints served as models to emphasize commonalities between human and wildlife risks.     

The fitness consequences of environmental sex reversal in fish: a quantitative review

Environmental sex reversal (ESR) occurs when environmental factors overpower genetic sex-determining factors. The phenomenon of ESR is observed widely in teleost species, where it can be induced by exposing developing fish to endocrine disrupting chemicals (EDCs). EDC-induced ESR has been exploited by the aquaculture industry, while ecological and evolutionary models are also beginning to elucidate the potential roles that sex-reversed individuals play in influencing population dynamics. However, how EDC exposure affects individual fitness remains relatively unknown. To date, many experimental studies have induced sex reversal in fish and measured fitness-as indicated by related traits such as size, survival and gonadal somatic index (GSI), but the reported results vary. Here, we meta-analytically combine the results of 78 studies of induced ESR to gain insight into the fitness of sex-reversed individuals. Overall, our results suggest that the fitness of fish exposed to EDCs is reduced at the time of exposure, with exposed individuals having a smaller size and likely a smaller GSI. Given a period of non-exposure, fish treated with EDCs can regain a size equal to those not exposed, although GSI remains compromised. Interestingly, survival does not appear to be affected by EDC treatment. The published reports that comprise our dataset are, however, based on captive fish and the general small size resulting from exposure is likely to lead to reduced survival in the wild. Additionally, reduced fitness-related parameters are likely to be due to exposure to EDCs rather than ESR itself. We suggest that theoretical models of ESR should account for the fitness-related effects that we report. Whilst we are able to shed light on the physical fitness of EDC-exposed fish, the behaviour of such individuals remains largely untested and should be the focus of future experimental manipulation.     

Molecular mechanism(s) of endocrine‐disrupting chemicals and their potent oestrogenicity in diverse cells and tissues that express oestrogen receptors

Endocrine‐disrupting chemicals (EDCs) are natural or synthetic compounds present in the environment which can interfere with hormone synthesis and normal physiological functions of male and female reproductive organs. Most EDCs tend to bind to steroid hormone receptors including the oestrogen receptor (ER), progesterone receptor (PR) and androgen receptor (AR). As EDCs disrupt the actions of endogenous hormones, they may induce abnormal reproduction, stimulation of cancer growth, dysfunction of neuronal and immune system. Although EDCs represent a significant public health concern, there are no standard methods to determine effect of EDCs on human beings. The mechanisms underlying adverse actions of EDC exposure are not clearly understood. In this review, we highlighted the toxicology of EDCs and its effect on human health, including reproductive development in males and females as shown in in vitro and in vivo models. In addition, this review brings attention to the toxicity of EDCs via interaction of genomic and non‐genomic signalling pathways through hormone receptors.     

Neurosteroid-induced hyperalgesia through a histamine release is inhibited by progesterone and p,p'-DDE,an endocrine disrupting chemical

The intraplantar injection of dehydroepiandrosterone sulfate (DHEAS), a representative neurosteroid, showed hyperalgesia in the Hargreaves' thermal or automatic paw-pressure mechanical nociception test. The DHEAS-induced hyperalgesia was abolished by diphenhydramine (DPH), a H(1) histamine (His) receptor antagonist, as well as the hyperalgesia induced by His or compound 48/80, a mast cell degranulating agent. The DHEAS-induced hyperalgesia was also blocked by progesterone (PROG), another type of neurosteroid and a putative neurosteroid receptor antagonist. Neither DPH nor PROG showed any changes in the thermal threshold. On the other hand, endocrine disrupting chemicals (EDCs) are known to disrupt reproductive system in wild-lives and humans through the disturbance of the endocrine homeostasis. In this study, the flexor responses induced by intraplantar injection of DHEAS were blocked by p,p'-DDE, an EDC as well as by PROG in the algogenics-induced nociceptive flexor responses test (ANF test) in mice. Similarly, p,p'-DDE blocked the DHEAS-induced hyperalgesia in Hargreaves' thermal nociception test. Besides the hyperalgesic actions, DHEAS increased vascular permeability as measured with Evans blue plasma extravasation. Consistent with behavioral studies, it was blocked by DPH, PROG, and p,p'-DDE. These results suggest that DHEAS has significant hyperalgesic and vasodilatory actions through histamine release, and these actions were reversible by PROG and an EDC.     

Neuroendocrine Systems and Avian Sexual Differentiation

Sexual differentiation of the endocrine and behavioral componentsof the avian reproductive system occurs throughout embryonicdevelopment, initiating some of the early cellular events thatform the central nervous system (CNS) and culminating in theorganization of male or female neuroendocrine responses. Earlycellular events have been studied intensively in recent yearsand these developmental processes appear to involve specificgrowth factors in the development of certain tissues. Subsequentto cellular differentiation, primordial germ cells migrate tothe appropriate anatomical location and contribute to the developmentof the single ovary or testes; steroidogenesis begins soon thereafter.Other portions of the hypothalamo-pituitary-gonad (HPG) axisalso appear during early embryonic development with migrationof the gonadotropin-releasing hormone (GnRH) neurons from theolfactory region of the CNS to the midbrain and separate formationof the pituitary gland. The gonadal steroid hormones affectdevelopment of accessory sex structures as well as the laterorganization of neuroendocrine regulatory systems and secondarysex characteristics. Manipulation of steroids during embryonicand early posthatch periods results in altered endocrine andbehavioral responses in adult birds. There are marked speciesdifferences in the timing of these events, especially when precocialand altricial species are compared. Altricial species hatchin a less developed state and as such are more dependent onparental care. Of necessity, the precocial species must be capableof feeding and other motor capabilities at hatch and coincidentallytheir other physiological systems also appear to be more matureat this time. Finally, there is the separate issue of song birdsversus those avian species that do not have elaborate learnedsongs. It appears that most of the species with elaborate neuralcircuitry responsible for song are altricial. This may benefitthem from the standpoint of gaining more time and contact withthe parents to enable them to learn the appropriate song. Thereare also hormonal and neuroendocrine components critical inthe process of song development. Finally, the effects of environmentalfactors, such as endocrine disrupting chemicals take on addedsignificance when viewed in the context of exerting permanentorganizational effects which are likely to alter endocrine andbehavioral components of reproduction in the adult     

Endocrine disruptors: present issues, future directions

A variety of natural products and synthetic chemicals, known collectively as endocrine-disrupting compounds (EDCs), mimic or interfere with the mechanisms that govern vertebrate reproductive development and function. At present, research has focused on (i) the morphological and functional consequences of EDCs; (ii) identifying and determining the relative potencies of synthetic and steroidal compounds that have endocrine-disrupting effects; (iii) the mechanism of action of EDCs at the molecular level; and (iv) the recognition that in "real life," contamination usually reflects mixtures of EDCs. Future research must examine (i) the interactive nature of EDCs, particularly whether the threshold concept as developed in traditional toxicological research applies to these chemicals; (ii) when and how EDCs act at the physiological level, particularly how they may organize the neural substrates of reproductive physiology and behavior; (iii) the various effects these compounds have on different species, individuals, and even tissues; and (iv) how adaptations may evolve in natural populations with continued exposure to EDCs. Several predictions are offered that reflect these new perspectives. Specifically, (i) the threshold assumption will be found not to apply to EDCs because they mimic the actions of endogenous molecules (e.g., estrogen) critical to development; hence, the threshold is automatically exceeded with exposure. (ii) Behavior can compound and magnify the effects of EDCs over successive generations; that is, bioaccumulated EDCs inherited from the mother not only influence the morphological and physiological development of the offspring but also the offsprings' reproductive behavior as adults. This adult behavior, in turn, can have further consequences on the sexual development of their own young. (iii) The sensitivity of a species or an individual to a compound is related to species (individual)-typical concentrations of circulating gonadal steroid hormones. Related to this is the recent finding that alternate forms of the putative receptors are differentially distributed, thereby contributing to the different effects that have been observed. (iv) Except in extraordinary situations, populations often continue to exist in contaminated sites. One possible explanation for this observation that needs to be considered is that animals can rapidly adapt to the nature and level of contamination in their environment. It is unlikely that successive generations coincidentally become insensitive to gonadal steroid hormones fundamentally important as biological regulators of development and reproduction. Rather, adaptive alterations in the genes that encode steroid receptors may occur with chronic exposure to EDCs, allowing the sex hormone receptor to discriminate natural steroids from EDCs.