The Invisible Influence: Can Endocrine Disruptors Reshape Behaviors Across Generations?

Abstract

Among the numerous compounds released as a result of human activities, endocrine-disrupting chemicals (EDCs) have attracted particular attention due to their widespread detection in human biological samples and their accumulation across various ecosystems. While early research primarily focused on their effects on reproductive health, it is now evident that EDCs may impact neurodevelopment, altering the integrity of neural circuits essential for cognitive abilities, emotional regulation, and social behaviors. These compounds may elicit epigenetic modifications, such as DNA methylation and histone acetylation, that result in altered expression patterns, potentially affecting multiple generations and contribute to long-term behavioral phenotypes. The effects of EDCs may occur though both direct and indirect mechanisms, ultimately converging on neurodevelopmental vulnerability. In particular, the gut–brain axis has emerged as a critical interface targeted by EDCs. This bidirectional communication network integrates the nervous, immune, and endocrine systems. By altering the microbiota composition, modulating immune responses, and triggering epigenetic mechanisms, EDCs can act on multiple and interconnected pathways. In this context, elucidating the impact of EDCs on neurodevelopmental processes is crucial for advancing our understanding of their contribution to neurological and behavioral health risks.

1. Introduction

Nowadays, one of the major challenges in research is investigating the effects of environmental pollutants on human health. Modern ecosystems (including water, air, soil, and biomass) are contaminated by a mixture of substances resulting from anthropogenic activity [1]. These contaminants include industrial chemicals, pesticides, pharmaceuticals, personal care product and food additives, heavy metals, and plastic-associated compounds, and much of our current knowledge about their effects derives from experimental studies using animal models (especially rodents) [2] (Figure 1).

Among the substances that have aroused interest are endocrine-disrupting chemicals (EDCs), which the World Health Organization (WHO) and the International Program on Chemical Safety (IPCS) define as “an exogenous substance or mixture that alters function(s) of the endocrine system and consequently causes adverse health effects in an intact organism, or its progeny or (sub)populations” [8].

EDCs comprise two important groups. The first includes the so-called “dirty dozen”, a list of twelve persistent organic pollutants (POPs) classified by the Stockholm Convention as hazardous to the environment and human health [9]. This group includes dioxins, polychlorinated biphenyls (PCBs), and dichlorodiphenyltrichloroethane (DDT), which are known to be persistent in the environment, bioaccumulate in the food chain, and disrupt endocrine functions, leading to adverse outcomes in development and reproduction [10].

The second group of “emerging contaminants” includes chemicals commonly found in personal care products [11,12] or food and beverage packaging, such as triclosan, parabens, and phthalates. They have importance in the chemical industry as they are added to materials to enhance their quality and confer specific properties (such as flame retardancy, plasticity, antimicrobial activity, or preservation) [13]. By imitating or inhibiting natural hormones, these compounds can also impair thyroid function and reproductive health [2]. Their health risks may extend beyond the disruption of classical nuclear hormone receptors to include membrane steroid hormone receptors, which can induce widespread effects across multiple tissues and organ systems by modulating diverse intracellular pathways through non-genomic signaling mechanisms [14].

Given their widespread use and increasing detection in aquatic environments, they are now recognized as emerging pollutants with substantial endocrine-disrupting potential [15].

Concerns about EDCs stem mostly from their detection in a variety of biological matrices, including those of particularly vulnerable populations [5,16,17,18]. These compounds were found in the serum, urine, amniotic fluid, cord blood, and breast milk of pregnant women [4,19,20], posing questions about their potential impact on the development of fetuses. The nature and extent of the effects depend on several factors, including the type of EDC, the dose, and the timing of exposure throughout critical developmental windows. In this context, exposures during the prenatal and early childhood periods are particularly concerning, as they may interfere with normal growth, resulting in permanent neurological, immunological, reproductive, and developmental abnormalities [21].

In vitro research on human peripheral blood mononuclear cells from both males and females showed EDCs’ ability to directly alter immune functions, pro-inflammatory responses, natural killer cell activity, and lymphocyte differentiation, including the expression of the scaffolding receptor for activated C kinase 1 (RACK1) gene, in a sex-dependent manner [22]. Recent evidence reported that the pesticides vinclozolin, atrazine, and cypermethrin, acting as EDCs, may downregulate RACK1 via anti-androgenic mechanisms (also mediating indirect effects), leading to a decrease in cytokine release and reduced immune cell surface marker expression, exerting a potential immunosuppressive effect [23].

Their documented immunomodulatory action necessitates additional examination, since it may lead to several detrimental health consequences, including an elevated risk of cancer. In this context, RACK1 emerges as a potential point of convergence between endocrine, immune, and oncogenic signaling pathways [24]. Notably, RACK1 is aberrantly expressed in several malignancies, including breast cancer [25]. This dual role highlights a plausible mechanism through which EDCs may elevate cancer susceptibility by targeting signaling hubs such as RACK1, thereby simultaneously impairing immune responses and altering the tumor microenvironment [26].

In rodent models, estrogen-mimicking chemicals such as certain phthalates and bisphenol A (BPA) potentially cause severe reproductive issues in females [27] and affect testosterone levels in males [28,29]. In particular, in utero exposure to BPA (0.5, 20, and 50 µg/kg/day) resulted in the inhibition of germ cell nest breakdown in F1 females, and fertility problems were also observed at low doses [30]. Exposure of adult male rats to 5 and 25 mg BPA/kg/day resulted in decreased testosterone levels, diminished sperm production, and alterations in particular functional parameters [31].

Dietary and lifestyle decisions also contribute to the EDC burden. For instance, daily exposure can occur through the ingestion of contaminated food or the use of personal care products, but unhealthy behaviors such as alcohol consumption and smoking can exacerbate the effects by altering detoxification and metabolic pathways [32]. However, detoxification efficiency and overall resilience to these chemicals could be influenced by genetic factors, which may define individual susceptibility [33].

Over the past few decades, significant knowledge has been acquired by using in vitro and in vivo models in experimental studies. Early investigations on animals revealed serious reproductive impairments linked to PCBs and pesticides, such as DDT, historically associated with the environmental crisis known as the “silent spring”, described by R. Carson in the 1960s [6,34,35,36,37,38]. These early findings provided a foundation for subsequent research exploring broader effects on the immune system, metabolism, and neurodevelopment [18,39,40,41]. This body of work revealed the intricate relationships between biological systems and EDCs, with growing evidence of their involvement in epigenetic changes [42,43,44,45].

The endocrine system is directly involved in shaping the central nervous system, and hormones influence neuronal plasticity and the formation of circuits that are responsible for defining the animal’s basic behaviors [46,47].

In particular, the initial phases of development are easily influenced by external factors, including chemical exposure. EDCs can interfere with the proper formation of neuronal circuits and brain structure, with significant effects on cognitive functions and behaviors. Furthermore, it has been extensively demonstrated that early-life stressors can have durable effects on the individual, increasing the risk for the development of neurological disorders later in life [48]. EDCs can act through numerous mechanisms, including the dysregulation of hormonal pathways in both the fetus and the mother [49], also inducing epigenetic alterations [50].

At this point, a key question is how EDCs modulate the development of the central nervous system and which of these changes may be heritable. All this has a profound importance and would demonstrate how environmental chemical contamination can shape biological outcomes not only at the level of individuals but also across populations.

These investigations emphasize the intricate connections between biological systems and environmental chemicals, contributing to a more thorough understanding of EDC-related impacts across species. This is an integrative approach, in alignment with the One Health framework, which recognizes the interdependence of human, animal, and environmental health.

2. Targets of Endocrine-Disrupting Chemicals in Neurodevelopmental Processes

A balanced hormonal milieu is crucial for optimal brain development, and the synchronized maturation of endocrine, neuronal, and immunological systems facilitates the establishment of a healthy brain architecture and enduring cognitive and behavioral functions. The endocrine system regulates neurogenesis, brain maturation, and immune system development through precise hormone communication [51]. Processes such as microglial activation and immune cell differentiation are indispensable for shaping neural circuits, regulating synaptic pruning, and maintaining central nervous system homeostasis [52].

EDCs can interfere with any component of this interconnected system—hormonal, neural, or immune—potentially triggering problems on multiple fronts (Figure 2).

One of the main ways in which a developing fetus is exposed to EDCs is these substances passing through the placental barrier, putting the baby at risk of developing problems, including later in life. Current knowledge on the placental transfer of EDCs remains limited. Due to their lipophilic properties, certain EDCs can easily pass through the placenta and reach the developing fetus [54,55,56]. While passive diffusion has been documented [56], evidence also suggests that transport mechanisms may depend on the physicochemical properties of individual EDCs [57].

Regarding the direct effects on placental structures, a substantial body of knowledge has been derived from studies employing rodent experimental models.

For instance, placental samples from C57BL6J mouse dams fed 200 μg/kg/day of BPA and bisphenol S (BPS) for 2 weeks before mating (and continuously until E12.5) exhibited altered placental organization and dysregulated expression of genes critical for the spongiotrophoblast and trophoblast giant cell development. Moreover, the lower neurotransmitter levels (including 5-hydroxytryptamine) during critical stages of brain development resulted in behavioral abnormalities [58].

The presence of organochlorine and organophosphorus pesticides in brain samples from stillbirths from Northeast Italy provided remarkable proof of the placental transfer of hazardous substances in humans [56]. Moreover, the early expression of hormone receptors in the brain, by the end of the first trimester in humans and mid-embryonic development in rodents, renders the developing nervous system responsive to hormonal stimulations and EDC exposure [59,60]. Such receptors are involved in different key processes, including neural cell differentiation and migration, synaptogenesis, and myelination [61].

The aryl receptor (Ahr) has been shown to play a crucial role in the very early stages of neurodevelopment [62], and the related pathways were among the first identified targets of EDCs’ activity [63]. It contributes to neural circuit formation, synaptic plasticity, learning and memory, and defense against xenobiotic-induced neurotoxicity [62]. The function of AhR has been primarily elucidated through binding assays involving the persistent environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) [64], and other studies have contributed to shedding light on its activity. For example, early exposure of zebrafish to TCDD at an approximate concentration of 10 ng/mL for 1 h starting at 4 h post-fertilization revealed its ability to induce abnormalities in the development of specific brain regions, impairing morphogenesis, and different signaling pathways regulated by the AhR [65].

Estrogen receptors (ERs) are also required in neural proliferation, differentiation, synaptic plasticity, or protection against neuroinflammation and neurodegeneration [66,67,68,69]. PCBs, pesticides, heavy metals, and 4-nonylphenol have all been shown to bind to them [53].

EDCs also target the hypothalamus–pituitary–thyroid (HPT) axis [70], with obvious consequences for neuronal thyroid-hormone-dependent processes [47]. Exposure to polybrominated diphenyl ether (PBDE) flame retardants, specifically the congeners BDE-47, BDE-99, and BDE-100, has been linked to a heightened risk of thyroid disease in U.S. women, with more pronounced effects noted after menopause, potentially attributable to modified estrogen levels that exacerbate PBDE-induced interference with thyroid signaling [71]. Likewise, per- and polyfluoroalkyl compounds (PFAS) have been associated with thyroid dysfunction. A longitudinal investigation in a PFAS-exposed population revealed that heightened serum PFAS levels were associated with raised thyroid-stimulating hormone (TSH) levels, suggesting possible hypothyroid consequences [72].

These data highlight the susceptibility of the HPT axis to EDC interference, potentially indicating consequences for neurodevelopmental outcomes. For instance, the biologically active form of the thyroid hormone triiodothyronine (T3) plays a pivotal role in neuronal development, the differentiation and maturation of astrocytes, oligodendrocytes, and microglia [73,74,75,76], and myelin deposition [77].

Maternal and congenital hypothyroidism is well-known to cause significant neurological damage and intellectual incapacity in offspring, including an increased autism risk [78,79]. Other thyroid dysfunctions, such as maternal hypothyroxinemia owing to iodine shortage, are risk factors for neurodevelopmental disabilities, particularly during the early parts of pregnancy [80]. The exposure of pregnant Wistar rats to 0.5 or 5 mg/kg of 4-OH-CB107 (a PCB metabolite) and 25 mg/kg of Aroclor 1254 (a mixture of parent PCB congeners) from gestational day 10 to 16 resulted in a substantial drop in plasma thyroid hormone levels in offspring by postnatal day 4. Neurochemical analysis revealed that Aroclor 1254 disrupted both dopaminergic and serotonergic neurotransmission, while 4-OH-CB107 primarily affected the dopaminergic and noradrenergic systems, with minimal effects on serotonin signaling [81].

In addition to processes such as neuronal proliferation, migration, differentiation, synaptogenesis, and myelination, thyroid hormones play a role in photoreceptor maturation in the retina and are essential for neural circuit formation in the visual cortex [82,83,84]. Certain azole pesticides and PCBs can disrupt thyroid-hormone-mediated pathways, leading to interference with eye growth and vision processing, resulting in visual impairment, and altered ability to traverse the environment, identify social cues, and interact socially. In zebrafish models, larvae exposed to common azoles such as climbazole (0.1 and 10 μg/L) and triadimefon (50 and 500 μg/L) [85], and embryos exposed to PCBs at concentrations ranging from 0.125 to 2.0 mg/L [86,87,88], exhibited impaired retinal development and morphological alterations in photoreceptor cells [89,90], ultimately compromising visual function and behavior [91]. Also, BPA can act as an antagonist for thyroidal hormone receptors by modifying the expression of target genes such as RC3/neurogranin (the rat homolog for human neurogranin) [92], which is involved with the development of synaptic circuits and serves as a biomarker of cognitive activity [93].

The brain’s susceptibility to hormones and EDCs also depends on the expression of hormone-metabolizing enzymes, whose presence is influenced by the developmental stage of the fetus and sex. Enzymes of particular relevance include aromatase, monoamine oxidases (MAO), enzymes involved in thyroid hormone metabolism (such as thyroid peroxidase and deiodinases), acetylcholinesterase, and UDP-glucuronosyltransferase (UGT). They represent potential targets for EDCs, and their disruption may contribute to neurodevelopmental defects and behavioral alterations. Considerable insights have been garnered from teleosts, particularly zebrafish, where the significant expression and activity of aromatase B in the brain [94] modulate neurogenic processes, influencing behavioral patterns and locomotor capabilities [95].

MAOs are crucial for mood regulation, cognitive functions, and emotional responses [96,97], and EDC-induced disruption of monoamine metabolism may lead to altered neurotransmitter levels and behavioral effects [98,99,100].

The disruption of neurotransmitter catabolism may be another mechanism. The inhibition of AChE can lead to acetylcholine accumulation, potentially causing neurodevelopmental and behavioral issues [101]. UGTs, which are involved in glucuronidation and xenobiotic metabolism in the brain, can be affected by dioxin-like compounds [102], altering steroid metabolism and then impacting neurodevelopment [103,104,105].

As previously stated, the normal growth and maintenance of the central nervous system is also regulated by the cross-talk between the endocrine and immune systems, known as the immune–neuroendocrine network (INEN) [106,107]. Recent evidence highlights that EDCs can interfere with the INEN, targeting key components of the network and contributing to neuroinflammation and increased vulnerability to neuropsychiatric and neurodegenerative disorders [108]. It is already well-established that xenoestrogens may contribute to the risk of neurological diseases such as multiple sclerosis, stroke, and Alzheimer’s disease, which are more prevalent in women, also suggesting a sex-dependent mode of action [109].

Buoso et al. (2025) provided valuable insights into the complex interplay between endocrine disruptors and the immune–neuroendocrine network by proposing a hypothesis centered on a common molecular target to explain the detrimental effects of BPA [108]. Specifically, BPA was shown to downregulate the abovementioned scaffolding protein RACK1, involved in key cellular processes, directly or indirectly through the hyperactivation of the hypothalamic–pituitary–adrenal (HPA) axis. This downregulation results in reduced neuronal production of mature brain-derived neurotrophic factor (BDNF) [110], which is associated with anxiety-like behavior, increased neuroinflammation [110], and oxidative stress.

The G protein-coupled estrogen receptor (GPER), a seven-transmembrane domain receptor, has a particular relevance in this context, due to its ability to crosstalk with nuclear receptors, such as AhR and ERs [111,112], and bind environmental EDCs, including 17α-ethynylestradiol (EE), BPA [113] and pesticides [23].

GPER is broadly expressed in the HPA axis as well as in brain regions implicated in behavioral control [114]. Notably, a GPER deficiency in the rat model has been associated with phenotypic alterations, including dysregulated basal and stress-induced glucocorticoid responses, increased anxiety-like behaviors, and deficits in learning and memory [115]. In vitro studies using human THP-1 monocytes further demonstrated that BPA-mediated GPER activation leads to RACK1 up-regulation. These findings also suggested the involvement of the NF-κB signaling pathway in BPA-induced immunomodulatory responses, reinforcing GPER’s role in neuroimmune and endocrine homeostasis [116].

Also, BPA exposure was found to stimulate pro-inflammatory cytokines production by microglia and to affect neuronal activity through glucocorticoid receptors (GRα) expressed in the astrocytes [117], further contributing to BPA’s disruptive effects on neurodevelopment and neuroimmune regulation [117]. In particular, the study in E15.5 mouse embryos by Takahashi et al. (2018) revealed that BPA exposure significantly increased microglial numbers in the dorsal telencephalon and hypothalamus and altered the expression of microglial markers (Iba1, CD16, iNOS, CD206), inflammatory cytokines (TNFα, IL-4), signaling molecules (Cx3Cr1, Cx3Cl1), and neurotrophic factor IGF1 [118].

These data imply that BPA inhibits microglial growth and neuroinflammatory pathways during brain development, indicating prospective targets for neurodevelopmental toxicity assessment.

Since neuroinflammatory phenomena can significantly influence the integrity of the blood–brain barrier (BBB) [119] by triggering microglial activation and the release of pro-inflammatory cytokines, EDCs may increase BBB permeability, facilitating the entry of peripheral immune cells and other harmful substances into the brain, thereby exacerbating neurodevelopmental and neurodegenerative risks. A 6-week oral exposure of two-month-old C57BL/6J males to di(2-ethylhexyl) phthalate (DEHP) alone (5 and 50 μg/kg/day) or in an environmental phthalate mixture enhanced the permeability of the BBB due to the affection of the endothelial accessory tight junction protein zona occludens-1 and caveolae protein Cav-1 in the hypothalamic medial preoptic area and the hippocampal CA1 and CA3 areas [120].

3. The Epigenetics of Endocrine Disruptors in Nervous and Neuroendocrine Systems

The concept of endocrine interference includes the ability of certain substances to induce epigenetic changes, i.e., alterations in gene expression that occur without modifying the DNA sequence itself. These processes include methylation, modifications in histones, or gene silencing via noncoding RNAs and can be triggered by various factors, including exposure to xenobiotics. Establishing a direct cause-and-effect correlation between environmental pollutants and health outcomes is challenging in both experimental and epidemiological studies, as the effects are generally cumulative and may manifest years after the initial exposure.

A wide range of substances, including metals (cadmium, arsenic, methylmercury, nickel, and chromium), peroxisome proliferators (trichloroethylene, dichloroacetic acid, and trichloroacetic acid), air pollutants (particulate matter, black carbon, and benzene), and also EDCs (diethylstilbestrol, BPA, persistent organic pollutants, and dioxins), have been shown in in vitro and in vivo experiments to induce epigenetic modifications [121]. Such changes may contribute to the transmission of potentially harmful conditions to future generations (even in the absence of continued exposure to the initial chemical trigger). Notably, epigenetic phenomena within the brain (associated with behavioral and reproductive dysfunction) have advanced our comprehension of EDCs’ role in influencing gene expression [122], complementing information from cancer research on EDC-induced epigenetic changes [7,123]. Although the literature contains little knowledge on how EDCs can act directly on the genome, there is evidence that their exposure is linked to the altered expression of certain epigenetic regulators, such as DNA methyltransferases (DNMTs), methyl-CpG-binding proteins, and histone deacetylases (HDACs), producing outcomes that depend on sex, dose, and the timing of exposure [124].

3.1. Mechanistic Insights into Endocrine Disruptors’ Epigenetic Perspective

The HPA axis is a good illustration of how EDCs can cause epigenetic remodeling due to its involvement in neuroendocrine homeostasis [125]. Evidence has suggested that the offspring phenotype can be profoundly influenced by parental exposure to stress [126,127,128]. In particular, in several human studies, increased maternal stress and cortisol levels have been correlated with neurodevelopmental outcomes in offspring, including increased attention deficit hyperactivity disorder and autistic traits, increased anxiety-related symptomology, and reduced IQ and memory [129,130,131]. In this respect, animal model studies have provided important insights by explaining the underlying mechanisms, mostly connected to epigenetic alterations of genes implicated in HPA axis development and related behaviors. According to research by Grundwald et al. (2015) in rats, the effects of maternal stress during pregnancy can be transferred to F2 generations in a sex-dependent manner [132]. In females, increased HPA responses were linked to alterations in glucocorticoid negative feedback sensitivity, with decreased hippocampal glucocorticoid (GR) and mineralocorticoid (MR) receptor mRNA expression and increased corticotropin-releasing hormone (CRH) expression in the PVN. In F2 males, increased hippocampal Gr expression and attenuated HPA reactivity led to acute stress [133]. Also, adult male mice’s exposure to glucocorticoids before mating resulted in altered offspring behavior, with impairments in fear extinction and memory in females and increased anxiety in males [133,134].

Recently, a study by Di Criscio et al. (2023) on mice revealed that in utero exposure to human-relevant EDCs could alter DNA methylation in the HPA-axis-related genes Nr3c1, Nr3c2, and Crhr1, encoding GR, MR, and CHR receptor 1, respectively. These changes correlated with hyperactivity and decreased social behavior [135].

The modification of DNA methylation by EDCs has been documented in numerous studies. Through these mechanisms, BPA may influence synapse formation and maturation, with potential long-term consequences and the development of nervous system disorders across generations [136].

In the mouse model, maternal exposure to low doses of BPA at 20 µg/kg of body weight administered once daily by means of subcutaneous injection from E0 resulted in hypo- and hyper-methylation events in the fetal forebrain mediated by de novo methyltransferases (Dnmt 3a and 3b) [137].

BPA exposure increased basolateral amygdala Dnmt1 expression and decreased glutamic acid decarboxylase (GAD67) in postnatal day 45 female rats, indicating a potential impact on anxiety-related behaviors [138]. BPA may also disrupt the timing of key neurodevelopmental processes, such as the transition to increased expression of the chloride transporter KCC2 via the epigenetic downregulation of its expression, an effect reversible by HDAC inhibitors [136].

Additionally, BDNF, a critical neurotrophin for synaptic plasticity and neurodevelopment, has been identified as a target of EDC-induced epigenetic modifications. Administration of PCBs at 1 mg/kg to pregnant rats via intraperitoneal injection on E16 and E18 led to significant changes in offspring gene expression, affecting regulators of hypothalamic development, including BDNF and genes encoding GABA_B receptor subunits [139].

However, epigenetic alterations can be mediated by microRNA (miRNA), short noncoding endogenous RNA transcripts. Some modifications in microRNA levels may also exhibit variability across both sex and brain regions, as observed in the offspring of exposed pregnant rat dams [140].

Phthalate exposure was associated with modified miRNA expression, leading to different health conditions, such as neurodevelopmental outcomes. For instance, maternal urine concentrations of phthalate metabolites were correlated with detrimental cognitive and behavioral development in children, exhibiting sex-specific effects [141].

Research conducted on rodent and zebrafish models revealed modifications in the expression of miRNA related to BDNF, cAMP response element binding protein (CREB) [142], synapsin-1, and PSD95 [143]. In particular, to study hippocampus pubertal effects, 5-week-old male C57BL/6 mice were exposed to DEHP, 0.5, 5, and 50 mg/kg/day for 28 days. DEHP decreased miR-93 and increased TNFAIP1, resulting in CK2β degradation and inhibition of the Akt/CREB pathway, leading to neuronal death (via downregulation of Bcl-2). Learning and memory impairments were also discovered [143].

As discussed in the previous section, inflammatory processes play a central role in the mechanisms underlying neurodevelopmental disruptions. Emerging evidence indicates that EDC exposure can induce epigenetic modifications in genes regulating inflammation. For example, studies in human myeloid dendritic cells have demonstrated that exposure to nonylphenol and 4-octylphenol could alter T-cell cytokine responses through pathways involving estrogen receptors, the MKK3/6-p38 MAPK signaling cascade, and histone modifications [144]. Additional findings in human monocytes have shown that these same compounds could modulate chemokine expression by altering histone H4 acetylation [145].

Early BPA exposure induces long-term epigenetic and inflammatory alterations in immune cells. Prenatal exposure to BPA was found to enhance the production of pro-inflammatory mediators (CysLTs, PGD2, TNFα, IL-13) and reduce global DNA methylation levels in bone-marrow-derived mast cells (BMMCs) of adult mice [146].

These observations suggest a plausible hypothesis: if EDCs can drive the epigenetic reprogramming of inflammatory gene networks, they may contribute to neurodevelopmental processes where inflammation plays a critical role, potentially participating in the pathogenesis of neurodevelopmental and neurodegenerative disorders.

3.2. Endocrine Disruptors, Microbiota, and Epigenetic Changes: A Warning Triangle for Neuromodulation

Emerging studies suggest that EDCs may contribute to inflammatory conditions in the digestive system by interfering directly or indirectly with different pathways regulating the immune responses and gastrointestinal functions [147,148]. The intestinal immune system, with innate and adaptive components, plays a crucial role in protecting the host from external stressors, including chemical contaminants [149]. By regulating tolerance to dietary antigens and commensal microbiota while mounting defenses against pathogens and xenobiotics, it serves as a first line of defense against chemical stressors, which can otherwise compromise gut homeostasis and overall health.

To understand the ability of EDCs to interfere with proper immune system function, both in vitro and in vivo models have been employed, providing valuable insights into the underlying mechanisms of their immunomodulatory effects. Several studies have shown the ability of certain compounds, such as BPA, to interfere with the differentiation and proliferation of immune cells [108,150,151,152] and modulate histamine release [153].

Perinatal exposure to BPA in rats can reduce the population of helper T cells (Th), regulatory T cells (Treg), and dendritic cells (DC) in the mesenteric lymph nodes and spleen, impacting the appropriate production of cytokines [154].

Certain ECDs have been shown to alter the intestinal microbiota, which can exacerbate intestinal inflammation, increasing gut permeability and enhancing susceptibility to infections [155].

Additionally, intestinal bacteria can metabolize EDCs, leading to the production of equally or more toxic by-products, further altering microbiota composition and function [147]. All of this has significant neurological implications, as dysbiosis has been associated with the onset of neurobehavioral diseases [156].

According to the established theory of the “gut–brain axis”, brain function and behavior are intimately connected to gut microbiota health through bidirectional communication [157] mediated by immune signaling, hormones, and the vagus nerve [158]. Mediators synthesized by the gut microbiota [159], once released into the bloodstream, can impact neuronal activity and potentially trigger cognitive impairments [160]. Importantly, there is growing but still emerging evidence that microbiota-derived metabolites can act as epigenetic regulators, such as short-chain fatty acids (SCFAs), which can influence histone acetylation and DNA methylation, altering chromatin accessibility [161,162,163]. In particular, bacterial butyrate has been linked to altered DNA methylation patterns associated with neurodegenerative diseases and depressive-like symptoms [164].

Dietary nutrients processed by the gut microbiota serve as substrates to generate metabolites involved in epigenetic regulation [165]. For example, choline acts as a donor of methyl groups in methylation reactions, and its bioavailability is influenced by both diet and gut flora composition [166]. In rodents, deficits in maternal choline have been associated with changes in the cholinergic system and changes in the DNA methylation within the offspring’s hippocampus [167,168], a critical brain region involved in memory, creativity, decision-making, social behavior, and other cognitive functions, whose alterations can result in rigid and maladaptive behaviors [169].

Gut microbiota can influence brain neurotransmitter levels, affecting brain function and cognition. Animal studies have demonstrated that the gut microbiota can modulate neurotransmitter systems, with germ-free and antibiotic-treated mice exhibiting altered neurotransmitter levels and receptor expression in the brain [170].

In this context, the relationship between dietary exposure to EDCs and detrimental effects on the nervous system may represent another key point due to their ability to impact the microbiota. It was reported that long-term BPA exposure in women results in a pro-inflammatory gut microbiota and a decrease in Bacteroidales and Lactobacillus [171], with the latter genus appearing to exert a protective role against EDC-induced oxidative stress [172]. A study by Kong and colleagues further supported the role of the gut–brain axis by showing that daily consumption of Lactobacillus reduced autistic-like behaviors in rats treated with valproic acid during sexual maturation and weaning by resolving disruptions in 5-hydroxytryptamine metabolism [173].

However, in the rodent model, parental exposure to BPA and ethinyl estradiol (EE) was shown to affect the gut microbiota, leading to later behavioral and metabolic disruptions in F1 offspring, highlighting the transgenerational issues of dysbiosis [174].

It is essential to consider the gut–brain axis as a comprehensive network without neglecting the HPA axis, whose dysregulation can also be associated with alterations in the nervous system [175].

For example, in response to stress, activation of the HPA axis leads to the release of CRH, adrenocorticotropic hormone (ACTH), and GC, which can directly influence gut physiology by altering intestinal permeability, motility, and immune responses [176]. These hormonal signals also modulate gut microbiota composition and activity, creating a feedback loop in which gut-derived signals, including microbial metabolites and inflammatory cytokines, can in turn influence HPA axis function. For example, the gut microbiome’s enzymatic repertoire comprises 11β-hydroxysteroid dehydrogenase type 1, which modulates glucocorticoid levels and is influenced by several EDCs [177].

Therefore, considering that EDCs can interfere with the health of our gut microbiota, it is reasonable to propose a mechanistic link between EDCs, microbiota dysregulation, and epigenetic modifications (Figure 3). Understanding this complex interaction is critical for elucidating how environmental chemicals influence neurodevelopment and behavioral outcomes across generations.

4. From Individual Behavior to Population Dynamics: New Challenges for Behavioral Endocrinology

One important consequence of aberrant neurodevelopment is the emergence of behavioral alterations, highlighting the need to identify the temporal windows of susceptibility to toxic exposures, particularly during gestation. Early-life exposure can result in transgenerational effects, as fetal exposure also impacts the developing germ cells. Epigenetic modifications triggered during this period may lead to heritable increases in neurological disease susceptibility, behavioral changes, and altered adaptability to environmental challenges across generations [28,178,179,180,181,182,183].

For instance, Tran and colleagues (2023) demonstrated that placental DNA hypomethylation resulting from exposure to the DEHP plasticizer in rodents primarily affected genes implicated in neurological disorders, including dementia and autism [182]. The inheritance of such behavioral phenotypes is often sex-dependent. For example, F3-generation rats, descendants of vinclozolin-exposed progenitors, exhibited sex-specific behavioral differences. Male offspring displayed reduced anxiety behavior, whereas females exhibited an increase [181,184,185]. These effects correlated with distinct genetic patterns, especially in the mRNA levels of genes encoding for hormone receptors (estrogen receptor α, Esr1, and androgen receptor, Ar) across different brain regions [186,187].

Similarly, Hatcher et al. (2019) revealed that multigenerational exposure to phthalates (DEHP) reduced anxiety in female F3 mice from high-dose backgrounds, with no effect in males. Moreover, sex- and dose-dependent changes in estrogen receptor gene expression were found in the amygdala, notably with reduced Esr1 mRNA in F3 females with ancestral DEHP exposure [188].

In the field of behavioral endocrinology, there is a growing interest in the intricate relationship between environmental factors and behavioral development. This perspective is driven by the organization/activation hypothesis, which posits that gonadal hormones can shape nervous system development, acting both during early life and adulthood. Indeed, EDC exposure can affect reproductive health and neurodevelopment, often in a sex-specific manner [189].

An interesting example comes from Carbone et al. (2019), who reported a dual effect following chronic DEHP exposure in the rat model. Adult males showed an increase in plasma levels of luteinizing (LH) and follicle-stimulating (FSH) hormones, reduced testosterone levels, and diminished hypothalamic GABA content. These changes were accompanied by anxiety-like behaviors, suggesting simultaneous modulation of endocrine and behavioral systems [190]. Similarly, in utero exposure to a human-relevant mixture of EDCs altered the methylation pattern of HPA axis genes in mice, with subsequent effects on behavior, such as increased hyperactivity and reduced sociability, in adult males [135].

Experimental models clearly show that EDCs can interfere with normal neurodevelopment, shaping primary behavioral responses. Studying these effects in humans requires epidemiological research, which presents unavoidable challenges due to the ubiquity of chemical exposure and mixture effects.

Nonetheless, past incidents raised public awareness, for example, the “Yu-Cheng Rice Oil Poisoning” episode in the late 1970s, in which approximately 2000 individuals were exposed to PCBs, polychlorinated dibenzofurans (PCDFs), and ter- and quaterphenyls via contaminated rice oil. In addition to skin and ocular lesions, developmental abnormalities, such as ectodermal defects, cognitive delays, and behavioral impairments, were observed in the offspring of exposed mothers; these effects persisted in children born up to six years after exposure [191].

Recent studies continue to link EDC exposure to neurodevelopmental abnormalities, such as altered internalizing and externalizing behaviors (including anger, anxiety, and depression) [192,193,194,195]. Thus, it would be more accurate to refer to EDCs as exogenous substances capable of interfering with normal neuroendocrine regulation at both the individual and population levels [196].

Beyond individual outcomes, EDC-driven behavioral changes can impact fertility and reproductive dynamics. For instance, women exposed to high levels of PCBs through the consumption of contaminated fish have exhibited shortened menstrual cycles [197], while other PCB congeners have been associated with a decreased gestation duration [198] and reduced couple fertility [199].

In wildlife, EDC-induced behavioral alterations may destabilize social structures essential for survival and procreation, perhaps impacting group cohesion and survival strategies. For example, the exposure of honeybee larvae to the hormone analog pyriproxyfen impaired social integration, impacting colony balance and population growth [200]. Similarly, EDC-driven changes in predator–prey dynamics can adversely affect the ecosystem equilibrium. In larval fish, the exposure to environmental estrogen estrone (E1) impaired anti-predator behavior, increasing vulnerability to predators, though paradoxically reducing predation success in piscivorous sunfish [201,202]. Changes in migratory activity [203], foraging, pollination, and seed dispersal may also result from EDC exposure, impairing ecosystem services and contributing to biodiversity loss [204,205,206].

In humans, EDC-driven behavioral alterations in humans may affect social functioning and mental health, contributing to the increasing prevalence of neurodevelopmental disorders [207], psychiatric conditions, and social relationship impairments. In Poland, early school-age children exposed to certain phthalates exhibited lower cognitive performance (as measured by the Intelligence and Development Scale) and lower behavior scores (Strengths and Difficulties Questionnaire) in association with a metabolite of dimethyl phthalate (MMP) and di-n-butyl phthalate exposure [208]. A French cohort study of 246 families found impaired verbal comprehension in six-year-old children exposed to greater concentrations of the PBDEs BDE 99 and BDE 209 after delivery [209]. Despite limitations in these studies, recent systematic reviews of epidemiological studies show links between early exposure to EDCs and the onset of autistic spectrum symptoms [210]. This reinforces the need to reframe research paradigms to better capture the role of environmental exposure in neurodevelopmental outcomes.

Given the multidimensional nature of EDC effects,

Table 1. Effects of endocrine-disrupting chemicals across epigenetic, neurodevelopmental, behavioral, and ecosystem levels.

Effect Level	Main Mechanism Involved	Observed Outcomes	Type of Study/Model System Investigated
Epigenetic level	DNA methylation, histone modifications, and altered noncoding RNAs	Altered gene expression patterns, transgenerational inheritance of phenotypes [135,181,182,183,184,185,186,187]	Mechanistic studies on rodent models
Neurodevelopmental level	Disruption of neural circuit formation, altered neurotransmitter systems, and hormone receptor modulation	Cognitive deficits, anxiety, and social behavior alterations [135,181,188,190,192,193,194,195,208,209,210]	Mechanistic studies on rodent models; Observational studies on human cohorts
Behavioral level (individual)	Neuroendocrine axis disruption (hypothalamic–pituitary–gonadal axis, hypothalamic–pituitary–adrenal axis), synaptic plasticity impairment	Anxiety, hyperactivity, altered sociability, attention deficits [135,181,188,190,192,193,194,195,208,209,210]	Observational studies on human cohorts; mechanistic studies on rodent models
Behavioral level (population/ ecosystem)	Disrupted mating, social structures, predator-prey dynamics	Reduced fertility, altered colony stability, increased predation risk [197,198,199,200,201,202,204,205,206]	Human observational (epidemiological); observational/speculative on various wildlife species
Ecosystem functions	Cumulative behavioral alterations in key species	Loss of ecosystem services (pollination, seed dispersal), and biodiversity decline [200,201,202,203,204,205,206]	Observational/speculative/mechanistic studies on wildlife models

provides an integrated overview of the elements reported in the text, spanning from epigenetic mechanisms to behavioral alterations and ecological impacts.

5. Methodological Challenges and Translational Barriers in Endocrine Disruptor Research

A comprehensive analysis of the EDC literature highlights considerable methodological heterogeneity arising from differences in species, strains, developmental windows, exposure routes, and dosing regimens. This variability undermines reproducibility and poses significant challenges for drawing consistent conclusions across studies, even when examining the same compound. Biases inherent to animal models present major limitations for extrapolating EDC findings to humans, as species-specific differences in hormone receptor profiles, metabolism, placental structure, and neurodevelopmental timing can influence susceptibility. Rodents (e.g., rats and mice) and fish species (e.g., zebrafish and medaka) are primarily employed to investigate EDCs’ effects on neurodevelopment and behavior; however, the reliance on controlled, high-dose exposures in these models often fails to replicate the chronic, low-level exposures typical in human populations [211].

Toxicological studies must also recognize that experimental settings cannot replicate the multiple concurrent exposures experienced in real-world environments, where organisms are simultaneously exposed to diverse contaminants, thus limiting the ability to assess potential synergistic effects [212].

Another major point in EDC research is represented by non-monotonic dose–response relationships, wherein low doses elicit effects that are not predicted by high-dose exposures or even provoke opposite outcomes. These non-linear dynamics challenge the classical toxicological paradigm that “the dose makes the poison” and complicate risk assessment processes. Despite increasing recognition of this phenomenon, many studies continue to omit sufficiently low doses or fail to include adequate dose ranges to detect non-monotonic responses [213].

Also, epigenetic consequences, suggested as important mediators of transgenerational impacts, represent another area of ambiguity [214]. Most research on epigenetic inheritance has centered on DNA methylation, histone modifications, and small noncoding RNAs. Additional potential carriers (prions [215], maternally inherited proteins, the microbiome, lipids [216], and other regulatory RNAs) may also mediate transgenerational information transfer. The field is rapidly evolving, and it remains to be determined whether these diverse mechanisms converge into a unified epigenetic signaling system or function as interconnected yet distinct pathways of heritable non-genetic information.

Given the critical need to address the limitations inherent in extrapolating findings from animal studies to humans, enhancing the translatability of research on EDCs requires the integration of epidemiological data with animal and cellular models, alongside explicit consideration of interspecies differences and systematic evaluation of study quality. The incorporation of advanced in vitro systems utilizing human-derived cells, such as brain organoids, in conjunction with computational models of human exposure, can offer valuable complementary insights to those obtained from traditional animal studies [217].

To comprehensively assess the translational relevance of experimental models and their alignment with epidemiological data, we compared findings from animal studies and major human cohorts investigating the neurodevelopmental impacts of EDCs. In this context, the SELMA (Swedish Environmental Longitudinal, Mother and child, Asthma and allergy study), CHAMACOS (Center for the Health Assessment of Mothers and Children of Salinas), and ELEMENT (Early Life Exposure in Mexico to Environmental Toxicants) studies are examples of key longitudinal birth cohorts that examine early-life exposure to EDCs and their effects on child neurodevelopment.

Regarding the SELMA study (https://www.kau.se/en/selma-study (accessed on 4 July 2025)), it enrolled over 2000 pregnant women in Värmland, Sweden, collecting maternal and child blood and urine samples for the analysis of approximately 20 EDCs. Recently, SELMA shed light on significant associations between bisphenol F (BPF) exposure and reduced cognitive function in children at seven years of age [218]. Additionally, prenatal phthalate exposure was linked to sex-specific differences in play behavior at the same age: higher di-isononyl phthalate concentrations were associated with decreased masculine play behavior in boys, although conclusive results were not observed in girls, highlighting potential gender-specific effects [219].

Some investigations within the CHAMACOS cohort have extended assessments into adolescence, reporting no significant associations between prenatal phthalate exposure and neurodevelopmental scores—a notable contribution, as no prior studies had examined these relationships in this critical developmental period [220]. However, despite the importance of CHAMACOS, which has followed over 800 children and their mothers since 1999, discrepancies with findings from other studies highlight how such research can yield unexpected results, possibly reflecting inconsistencies between parent- and self-reported behaviors, with self-report reliability increasing as participants age.

The ELEMENT investigations have followed three birth cohorts recruited from maternity hospitals in Mexico City for over two decades to elucidate how maternal and early-life exposure to metals and environmental chemicals affects pregnant women and child health outcomes. Of particular interest, prenatal exposure to the organophosphate pesticide chlorpyrifos—known to exhibit endocrine-disrupting activity—was associated with an increased risk of attention deficit hyperactivity disorder symptoms in children [3]. However, it is important to acknowledge that these findings are derived from studies with relatively modest sample sizes, underscoring potential limitations and the need for cautious interpretation in this type of research.

Unlike animal models—which, despite being invaluable, have limitations in reflecting human physiology—these cohorts provide direct evidence of EDC impacts in human populations. However, the limitations of cohort studies such as ELEMENT, SELMA, and CHAMACOS may include potential confounding by socioeconomic and lifestyle factors, reliance on spot urine samples or single-time-point measurements that may not accurately capture long-term or fluctuating exposures, and modest sample sizes that limit statistical power for detecting subtle neurodevelopmental effects.

Additionally, real-world exposure usually involves complex mixtures of multiple EDCs that may interact synergistically or antagonistically, and disentangling these combined effects remains challenging both in human epidemiological research and in experimental animal models. Such complexities highlight the difficulty in extrapolating isolated chemical findings to real-life exposure scenarios, underscoring the need for advanced analytical approaches and mixture-focused studies.

Therefore, integrating findings from such studies is essential for accurately assessing the neurodevelopmental risks of EDCs and should complement animal data in the broader evaluation of these chemicals.

Future Directions and Research Priorities

To address these complex challenges, future research should embrace integrative, cross-disciplinary approaches that extend beyond traditional toxicological paradigms. A key priority involves the advancement of human-relevant experimental models, such as brain organoids or induced pluripotent stem cell (iPSC)-derived systems, which more accurately recapitulate human neurodevelopmental processes and reduce reliance on animal models with limited translational validity.

The extension and standardization of longitudinal cohort studies are equally important. These efforts should include frequent exposure assessments and standardized neurodevelopmental outcome measurements to improve reliability and allow for meaningful comparisons between studies. Strengthening the epidemiological framework will also allow for deeper interpretations of exposure–outcome correlations, particularly in terms of developmental windows of susceptibility.

Future research should concentrate on chemical combinations to identify additive, synergistic, or antagonistic interactions, recognizing that environmental exposures rarely occur in isolation. This calls for the incorporation of innovative computational techniques and mixture-oriented risk assessment frameworks that can handle the intricacy of exposures encountered in the actual world.

Simultaneously, as non-monotonic dose–response relationships contradict the traditional presumptions that support current safety standards, they ought to be investigated and included in regulatory risk assessments.

Thus, the successful mitigation of the dangers associated with EDCs necessitates concerted efforts from the public health, scientific, and regulatory domains. Developing well-informed policies and encouraging interdisciplinary collaboration will be essential to converting research findings into effective public health initiatives.

6. Conclusions

Environmental contaminants, particularly EDCs, exert effects that must be assessed not only in terms of acute or short-term damage but especially in terms of chronic exposure and transgenerational impacts.

Initial research focused on the reproductive effects and dysregulation of hormonal homeostasis, but it is now clear that the mechanisms of action can also involve epigenetic modifications and interference with proper neurodevelopment. Disruption of neurodevelopment affects the formation of brain circuits that govern cognitive activities and behavior.

To advance understanding of these complex effects, future research must go beyond the limitations of traditional toxicology by prioritizing robust experimental designs, comprehensive population-level studies, and systematic evaluations of study quality to better inform risk assessments and public health policies.