Caenorhabditis elegans (C. elegans) is a soil-dwelling free-living nematode that has been established as a model system since the 70′s and was popularized by Sydney Brenner, followed by other scientists that worked on mapping its genome and in unraveling its development, behavior and neuronal network (Brenner, 1973, 1974; White et al., 1986). The study published in 1986 by John White and collaborators (a publication also known as “The mind of a worm”, with 340 pages) was a breakthrough for neuroscientists, as they have described how genes determine the structure of the nervous system and how this system converts the signaling into a behavior. They have described the number of neurons in this nematode and their wiring or connectome (White et al., 1986). In the following years, researchers described the neurotransmitters systems and further identified that the neurons would also produce and secrete hormones (Li et al., 1999, Li et al., 1999a; Schinkmann and Li, 1992).

The endocrine system in C. elegans regulates several functions such as molting, development, energetic metabolism, reproduction, neural development and several behaviors such as feeding, defecation and mating (Antebi, 2006). Besides neurons, other cells, such as hypodermic, muscle, and intestinal cells, produce molecules that can act as modulators (Alfhili et al., 2018; Shi et al., 2022; White et al., 1986). In response to an environmental cue, steroid and peptide hormones are produced and released to bind and activate their target receptors (membrane or nuclear receptors) to produce a physiological response. Since the worm does not have a bloodstream, hormones are distributed through intercellular pathways, such as gap junctions, neuronal signaling to neighboring cells or by just acting locally, providing a slower but consistent communication among the cells (Nässel, 2009; Nurrish, 2014). Notably, a dynamic interplay between synaptic and neuromodulatory signaling guarantees flexible but robust neuronal circuits.

There are many differences between the neuroendocrine system between mammals and C. elegans, but also many similarities. In these worms, many receptors share significant sequence homology to those found in humans and are likely to be functionally similar, as conserved peptides and steroids have also been found (Lai et al., 2000; Shaye and Greenwald, 2011). Worms do not synthesize cholesterol, they need to obtain it from the diet (Zečić et al., 2019). When they intake cholesterol, they can produce steroids, such as pregnenolone and dafachronic acid (Aguilaniu et al., 2016; Broué et al., 2007). Peptides are produced from DNA information (transcription and translation process) and the most recent data indicate that worms may produce approximately 300 peptides with hormonal function from 59 predicted neuropeptide precursor (NPP) genes (Ripoll-Sánchez et al., 2023). For example, it has been identified a functional thyrostimulin (TSH)-like signaling system in C. elegans, characterized by the presence of human orthologs of glycoproteins GPA2 and GPB5, which are TSH subunits that bind to the FSHR-1 receptor to trigger the hormone actions, such as controlling the defecation cycle (Kenis et al., 2023) In addition, the worms genome encodes 40 insulin-like genes, and only one insulin-like receptor DAF-2 (Zheng et al., 2018). Of note, the modulation of insulin/IGF signaling (IIS) in C. elegans is the central determinant of the endocrine control of stress response, diapause, and aging (Baumeister et al., 2006).

This particular pathway can be modulated by xenobiotics, which are exogenous chemical molecules to which the worm can be exposed to. Some molecules can modulate DAF-2 receptor and the downstream transcription factor DAF-16 (IIS) pathway and increase worms lifespan, such as sitaglipin, which is a peptidase-4 inhibitor and commonly used in the treatment of type 2 diabetes in humans (Ye et al., 2023). On the other hand, bisphenol S (BPA S), which is used for plastic production in replacement for bisphenol A (BPA A), causes obesogenic effect in worms by hyperactivating DAF-16 and NHR-8, which increases fatty acid synthesis and impairs longevity (Zhou et al., 2021) Similarly, other neuroendocrine pathways can be modulated and have been studied in C. elegans, which will be featured here.

This review aims to shed some light on what is currently known about the C. elegans neuroendocrine system and how it can be positively and negatively modulated. Unraveling the pathways is important to understand the nematodes biochemistry and physiology, to understand evolution of the neuroendocrine system and to validate this model as a screening model to study the effect drugs that can impact the environment (as endocrine disruptors) or that can be promising medicines for metabolic human diseases.

The nematode C. elegans is a well-established model that allows the assessment of toxicological and pharmacological endpoints for the screening of new drugs and xenobiotics, contributing to the transposition of these effects into more complex organisms. Furthermore, it is the first multicellular organism to have its genome completely sequenced with a high level of genetic conservation in mammals, encompassing this similarity to several basic biological functions and conserved pathways, such as apoptosis, stress response and cell signaling (Hunt, 2017; Ruszkiewicz et al., 2018). Notably, the C. elegans nervous system is a well characterized system and considered structurally and functionally similar to mammals. It is the only organism in which all neurons and the main interactions between them are well mapped, presenting approximately 302 neurons for adult hermaphrodites, and 383 in males, divided into 118 morphologically distinct classes and 56 glial cells, forming approximately more than 7600 synapses, 900 gap junctions, and 1500 neuromuscular junctions. Glial cells are fundamental structures of the C. elegans nervous system and although their development is poorly understood, it is known that of the 56 glial cells that are morphologically similar to several other organisms, 24 are called “glial sheath” and sheathe the modified endings of sensory neurons. These cells are associated with sensory neurons and respond to stimuli such as temperature, odors, flavors, high osmolarity solutions, pheromones and touch (Shaham, 2006).

The C. elegans neurons belong to two distinct nervous systems, the somatic and the pharyngeal. Neurons of the somatic system are positioned between the hypodermis and body wall muscle and share a basal lamina of the hypodermis that isolates them from the muscles. The neurons of the pharyngeal system are located directly between the muscles and the pharyngeal systems and, unlike the somatic system, are not separated by the hypodermis. These systems communicate through a single pair of interneurons called the Ring/Pharynx interneuron (RIP). The cell bodies of most neurons are grouped in ganglia present in the head or tail (White et al., 1983). The nervous system also differs between the biological sexes, males have a system larger in number of cells (473), with 79 additional neurons and 36 specific support cells that are involved in the mating process of the males, located in the posterior part of the body. Hermaphrodites presents two classes of specific neurons, the Hermaphrodite Specific motor Neurons (HSN), generated in males that undergo programmed cell death during early development, and the Ventral Cord motor neurons (VC), which are derived from lineages P that give rise to hook sensillum cells in male (Fig. 1A). Males also have three specific classes of motor neurons, sensory neurons and interneurons (Fig. 1B) (Chen et al., 2006; Donald L Riddle, Thomas Blumenthal, Barbara J Meyer, 1997; Emmons, 2005).

The biochemistry of the C. elegans nervous system is highly conserved, presenting neurotransmitters such as glutamate (Glu), γ-aminobutyric acid (GABA), dopamine (DA), serotonin (5-hydroxytryptamine; 5-HT) and acetylcholine (ACh), ion channels, G-protein coupled receptors, vesicular transporters and other synaptic components analogous to those of mammals. On the other hand, they do not have epinephrine, norepinephrine, histamine signaling and have significant differences in sodium channels (Ruszkiewicz et al., 2018). Little is known about the association of neurotransmitters and the endocrine system in C. elegans. However, it has already been described that serotonin, DA, GABA and Glu are neurotransmitters that undergo changes and are influenced by neuroendocrine modulators that stimulate their synthesis and activate or inhibit neurons that, for the most part, are associated with changes in the worm's behavior (Barrios, 2014; Katsanos and Barkoulas, 2021; Song et al., 2013; Zemkova et al., 2014).

Dopamine is produced in eight neurons in the nematode, each of which is mechanosensory, where any modification in these neurons triggers defects in the animal's ability to respond to changes in its environment. Dopamine modulates locomotion behavior, searching for food, learning and olfactory adaptation. Anterior Lateral Microtubule (ALM) and Posterior Lateral Microtubule (PLM) neurons express the DOP-1 receptor and modulate the response to touch, which may be associated with the animal's habituation to the environment. C. elegans encodes 4 dopamine receptors (DOP-1, DOP-2, DOP-3 and DOP-4) expressed on ventral medulla motor neurons to modulate the rate of locomotion and two of them are homologous receptors in mammals (D1 and D2) that function antagonistically to control the movement (Missale et al., 1998; Sanyal et al., 2004).

Serotonin, like dopamine, is also produced in eight types of neurons and is associated with the behavior modulation. This modulation is slightly similar to that of dopamine, as it is also associated with changes in the environment, interfering with locomotion, feeding, pharyngeal pumping and egg laying. However, serotonin encourages food deprived animals to not abandon the food source once found, unlike dopamine, which encourages well-fed animals to remain close to the food (Chase and Koelle, 2007). Amphid dual neurons (ADF), Neurosecretory-motor Neurons (NSM), HSN and Male specific cell in ventral cord (CPs) express the enzymes that synthesize serotonin, tryptophan hydroxylase (TPH-1) and Aromatic L-Amino Acid Decarboxylase (BAS-1). The nematode has four serotonin receptors, three of which are metabotropic (SER-1, SER-4 and SER-7), a chloride channel controlled by serotonin (MOD-1) and serotonin reuptake transporter (MOD-5), which acts on AIM and RIH neurons, showing that serotonin is a neurotransmitter that acts on neuromuscular synapses. (Fig. 2) (Chase, 2007; Ishita et al., 2020). The ADF and NSM neurons are responsible for modulating pharyngeal pumping depending on the environmental conditions in which the nematode is found, where ADF is related to increased pharyngeal pumping, being associated with feeding and NSM is associated with the regulation of feeding through chemosensory responses.

In ADF neurons, serotonin synthesis is coordinated and plays a fundamental role in modulating the worm's lipid metabolism. The expression of tph-1 is reduced in short-term food deprivation and can be restored when the animal is placed in an environment with food, associating serotonin levels with satiety. Although this mechanism is not well understood, it is already known that the serotonin receptor 5-HT acts directly on the worm's intestinal fat stores, which can lead to increased mitochondrial beta-oxidation (Lee et al., 2011; Palamiuc et al., 2017). 5-HT also regulates adipocyte triglyceride lipase (ATGL-1) which is predominantly expressed in the intestine, once excess 5-HT increases the expression of atgl-1, which decreases body fat, whereas loss of 5-HT decreases atgl-1 expression and increases body fat (Lai et al., 2000; Shaye and Greenwald, 2011). It has also been observed that serotonin signaling combined with octopamine has coordinated effects on body fat loss. It has been determined that 5-HT signaling requires the synthesis of octopamine via tbh-1 in RIC neurons and octopamine requires the synthesis of 5- HT via tph-1 through a possible yet unknown neuroendocrine axis (Srinivasan et al., 2008). In the same sense, it has been observed that serotonin acts to increase longevity. The transcription factor HLH-11 is a repressor of atgl-1, where in animals with a deletion of the hlh-11 gene there is an increase in fat oxidation. The increase in this oxidation triggers a mitochondrial stress response, which in turn will be necessary to maintain lipid oxidation and confer a protective nature on longevity (Littlejohn et al., 2020).

The neurotransmitter octopamine (chemically and functionally similar to epinephrine) is synthesized from tyramine by the tyramine β-hydroxylase TBH-1 in RIC interneurons and gonadal sheath cells. In C. elegans, the ser-3 gene encodes the octopamine receptor in SIA neurons during the absence of food or exogenous octopamine, relating this neurotransmitter to the response to hunger, inhibition of pharyngeal pumping and egg laying (O'Donnell et al., 2020; Suo et al., 2009).

Of the 302 neurons present in C. elegans, 26 express the GABA produced by two classes of neurons present in muscles of the vulva (VC4/5 and HSN). The unc-25 gene encodes the enzyme homologous to glutamic acid decarboxylase – GAD responsible for the synthesis of GABA. GABA performs excitatory and inhibitory functions. Type D ventral cord motor neurons (6DD and 13VD), RME, RIS interneuron and motor neurons (AVL and DVB) that are located respectively in the dorsal and ventral muscles of the body, head muscles and enteric muscles are GABA neurons (da Silva et al., 2022).

The nematode also has 10 neurons that express the excitatory neurotransmitter glutamate. Eight are from the non-NMDA class, which would be GLR-1 – GLR-8 neurons, homologous to neurons from the AMPA or kainate subfamilies. Two receptors belong to the NMDA class, the NMR-1 and NMR-2, homologous to mammalian NR1 and NR2A subfamilies. The eat-4 gene in C. elegans encodes a homologue of VGLUT, a glutamate transporter that mediates glutamatergic transmission. EAT-4 controls behaviors such as food seeking, avoidance, osmosensation and mechanosensation. AMPA-type receptors (GLR-1 and GLR-2) contribute to synaptic plasticity, being fundamental for learning and memory, and can even mediate behavioral responses during pathogen infections (da Silveira et al., 2018; Yu and Chang, 2022).

Despite being simple and limited when compared to that of mammals, their nervous system is responsible for performing basic behaviors such as locomotion, feeding and defecation, as well as more complex behaviors, such as discrimination of temperature, food source and chemicals, pheromones and changes in oxygen levels. They also present biological sex-specific behaviors, such as egg laying in hermaphrodites and mating in males. Feeding is a social behavior, which also modulates egg laying, locomotion and olfactory behavior of the nematode. Behaviors in C. elegans are plastic and subject to change through learning and memory (Ruszkiewicz et al., 2018).

In addition to being responsible for behavioral functions, the C. elegans nervous system is also responsible for the worm's physiological functions. Chemosensing functions are physiological functions that control the entry and exit of the worm at the dauer stage, an alternative stage that occurs at the end of the L1 larval stage when the animal is subjected to adverse environmental conditions to resist to stress. At this stage, chemosensory neurons express peptides from the TGF beta family, which is associated with fat storage. Daumone, a pheromone secreted by all animals, assists this process by detecting population density and food in the environment. Amphid sensory neurons ADF, ASI and ASG detect the dauer and the amount of food and are responsible for entry into the dauer stage. ASI produces DAF-7, which acts as a neuroendocrine signal preventing dauer formation and stimulating development, controlling the body size and lifespan of the worm (Bargmann, 2006). Therefore, it is remarkable that there is close relationship between neuronal function and hormonal modulation in this nematode as well, which will be further explored in this review.

The cellular communication in multicellular organisms occurs through signaling mechanisms. In synapses, information is passed through direct contact between the cells of the nervous system, however, when there is a greater distance between the cells that must be communicated, it is the endocrine system that carries out the signaling (Hartenstein, 2006). Endocrine system is composed of specialized cells that transmit signals through the secretion of hormones. The release of specific hormones informs target cells about the state of the organism, influencing growth, metabolism, reproduction and regulating homeostasis mechanisms (Antebi, 2015; Hartenstein, 2006).

In C. elegans, hormones control diverse behaviors, including dauer formation, locomotion, egg laying, and mechanical and chemosensation (Li et al., 1999). Hormones can be synthesized by specialized cells or produced through the proteolytic cleavage of prohormones. In the C. elegans genome, there are 163 neuropeptide genes that encode more than 300 neuropeptides. Forty genes encode insulin-like peptides, such as the ins genes, are expressed in ASI and ASJ neurons. Thirty one genes encode FMRFamide-related peptides, called FMRF-Like Peptide (flp) gene, expressed in the RIG neuron. Still, 42 genes encode neuropeptides unrelated to insulin and unrelated to FMRFamide, known as Neuropeptide-Like Protein (nlp), expressed in the hypodermic cells (Li et al., 1999).

Hormonal synthesis occurs through gene translation and cleavage of the molecule by peptidases located in the endoplasmic reticulum. In the case of FMRF, the addition of the amine group from glycine occurs, forming the active peptide. They are then stored in vesicles and released into the synaptic cleft to carry out endocrine signaling (Fig. 3) (Li et al., 1999).

On the other hand, the synthesis of steroid hormones is dependent on prohormones, such as dietary cholesterol, which acts as a crucial precursor (Höss and Weltje, 2007). The steroids endocrine signaling mechanism occurs when hormonal signals bind to nuclear hormone receptors, and transmit information. These specific receptors are ancient transcription factors that respond to some molecules and regulate gene expression (Antebi, 2015; Scholtes and Giguère, 2022).

In addition to the classic receptors that have their ligands already configured, there are so-called “orphan receptors”, which do not have any identified ligands. After the discovery of ligands for these receptors, they came to be called “adopted orphan receptors” (Scholtes and Giguère, 2022). C. elegans has 284 nuclear hormone receptors, most of them (269) derived from the hepatocyte nuclear factor 4 (HNF4) family and some of its functions are related to fat metabolism, molting, reproduction and cell differentiation (Anderson and Pukkila-Worley, 2020; Antebi, 2015).

Most of the receptors derived from the HNF4 family are orthologous to the alpha or gamma isoforms of the mammalian nuclear receptor hepatocyte nuclear factor 4 (HNF4⍺/γ). The C. elegans ortholog, the nuclear hormone receptor 64 (NHR-64), is located in the nucleus of the cells from the hypodermis, intestine, neurons and pharynx, is directly related to the regulation of fatty acid metabolism, repressing synthesis and promoting beta-oxidation (Antebi, 2015). The mechanism involves the positive regulation of fatty acid desaturases (FAT-5/6/7) and the negative regulation of the acetyl-CoA carboxylase homologue (POD-2) (Yue et al., 2021).

NHR-69 is expressed in the cell nucleus of various structures such as hypodermis, intestine, uterine toroidal epithelial cells and ASI and tail neurons. Its activity is related to binding to steroid hormones, therefore being a putative receptor for androgen hormones (Mendelski et al., 2019). This protein is also involved in glucose metabolism, insulin and transforming growth factor beta (TGF-β) signaling through associated action with other nuclear receptors. Furthermore, it can influence dauer formation and worm longevity, as it is expressed in ASI, an important sensory neuron that controls these two factors (Antebi, 2015; Godoy and Hochbaum, 2023).

DAF-12 is a nuclear receptor expressed in several structures, including gonad, neurons, pharyngeal muscle cell, somatic gonad precursor, and vulva. It is homologous to the human farnesoid-X (FXR), liver X (LXR) and vitamin D (VITD) receptors, which act by binding bile acids, oxysteroids and vitamin D. In C. elegans, DAF-12 is also regulated by steroid-like bile acid called dafachronic acid (DA) or cholestenoic acid. Activation of DAF-12 by DA is linked to the regulation of offspring formation, development time, metabolism, longevity, reproduction and worm reproduction (Antebi, 2015).

Additionally, NHR-8, expressed in the nucleus of intestinal cells, shares homology with VITD, FXR, and LXR receptors as DAF-12. Broadly speaking, its role encompasses the regulation of cholesterol homeostasis, specifically overseeing the production of bile acids - like DA, and the disposition of cholesterol (Anderson and Pukkila-Worley, 2020). Moreover, NHR-48 is another receptor homologous to VITD that acts together with DAF-12 and NHR-8. Its expression is notable in various structures, such as the anterior gonad arm, pharyngeal gland cells, and the spermatheca, where it plays a significant role in cell differentiation (Lu et al., 2016).

Steroidogenic factor 1 (SF-1) plays a significant role in the mammalian reproductive system, such as in sexual differentiation, through the NR5A1 receptor. In nematodes, the corresponding homologue of this receptor is NHR-25, located in several structures, including the excretory system, gonad, hypodermic cell, tail, and vulvar precursor cell. This receptor is involved in the positive regulation of tail tip morphogenesis and the molting cycle. The relationship with molting occurs through collaborative action with specific genes, such as collagenase, acting in the differentiation of epidermal cells (Antebi, 2015; Katsanos and Barkoulas, 2021).

Furthermore, the receptor homologous to the human retinoic acid receptor-related orphan receptor (ROR), NHR-23, plays a crucial role in spermatogenesis, the timing of maturation spermatocyte in response to nutrients and the molting process. This receptor is expressed in hypodermis, oocyte, spermatocyte, tail, and vulva precursor cell. Some studies have shown that depletion of NHR-23 affects the development and regulation of spermatogenesis in C. elegans (Johnson et al., 2023; Ragle et al., 2020).

The molting process is regulated by negative feedback between NHR-23 and the let-7 family. NHR-23 binds response elements in the let-7 promoter and activates transcription. In turn, let-7 decreases nhr-23 expression throughout development through a complementary let-7 binding site (LCS). NHR-23 and let-7 coordinate the expression of factors involved in molting such as lin-42, which are enriched in cuticle components (Patel et al., 2022).

Still regarding the reproductive system, the receptor involved in ovulation and morphogenesis of the spermatheca is called NHR-6, a homologue of human neuron-derived orphan receptor-1 (NOR1). It is expressed in BAGL, BAGR, chemosensory neurons, head neurons, and spermatheca. In addition, NHR-67, located in excretory cells, head ganglion, neurons, rectal valve cell, and vulvar cell, participates in processes such as gonad morphogenesis and in the negative regulation of the G1/S transition of the mitotic cell cycle (Antebi, 2015). Vulvar development leads to the generation of seven different types of vulvar cells (vulA, vulB1, vulB2, vulC, vulD, vulE, and vulF). This process occurs through feedback between NHR-67 and cog-1 or lin-1, acting on the increase or reduction in gene expression (Fernandes and Sternberg, 2007).

Some of the hormone receptors are implicated in cellular differentiation processes. UNC-55, which shares homology with COUP receptors in humans, exerts regulatory control over the synaptic remodeling of GABAergic motor neurons during the L1 developmental stage. Consequently, it is localized within motor neurons, the somatic nervous system, and spicular muscle tissues. NHR-85 is involved in cellular differentiation, dauer molt, and egg laying processes. It exhibits expression in excretory duct cells, the hypodermis, rectal epithelium, and vulva, and serves as an ortholog of Rev-Erb. Another example is NHR-41 which, in addition to its role in cellular differentiation, also contributes to the regulation of the molt cycle. Its expression is detected in excretory cells, the hypodermis, neurons, the straight, and the vulva, and it functions as an ortholog of human testicular receptor 2/4 (TR2/4) (Antebi, 2015).

Because endocrine receptors do not have affinity only for specific hormones, some molecules with structure and characteristics similar to these hormones may be capable of inducing an endocrine response in the body, whether positive or negative response. Some can be used as medicines (when positive modulations are observed), whereas others are called endocrine disruptors, when causing damage to the organisms. These endocrine modulators will be further discussed in this review.

Tissue-tissue communications are essential for the organism's homeostasis, allowing physiological processes to occur properly. The nervous system plays a fundamental role in this communication process, assimilating environmental signals and communicating to tissues. Intercellular communications occur throughout the nervous system through synaptic connections and neuroendocrine signaling (Anderson and Pukkila-Worley, 2020).

Neuromodulators such as biogenic amines and neuropeptides, participate in peptidergic signaling and act differently from neurotransmitters, being released by dense central vesicles (DCVs) either perisynaptically or volumetrically, diffusing over long distances and even presenting long half-lives. Consequently, they can evoke prolonged and distant responses through metabotropic G protein-coupled receptors. (Lim et al., 2016). In vertebrates, the hypothalamus is responsible for producing and releasing many hormones to control various functions, such as appetite, reproduction, circadian rhythms and social behaviors (Graebner et al., 2015; Selcho et al., 2012). In invertebrates, several neuromodulators that act in circuits appear to be associated with diverse behaviors, influencing movement, sleep, arousal and learning behaviors (Frooninckx et al., 2012; Güiza et al., 2018; Taghert and Nitabach, 2012).

The nematode C. elegans has characteristics that make it an excellent model for studies on the invertebrates neuroendocrine system, mainly because its nervous system is invariable. This is because its anatomical and morphological structure does not vary between individuals (isogeny by hermaphrodite reproduction), which makes it easy to detect small behavioral and developmental defects, in addition to having favorable genetics and being a great tool for molecular biology. However, a better understanding of the mechanisms associated with the responses of the neuroendocrine system to the environment in which the nematode is exposed is still necessary, as well as the morphology of the peptidergic system (Lim et al., 2016). As previously stated, the three classes of peptides synthesized by the neuroendocrine system of nematodes are associated with different functions. Many of these peptides have already been assigned functions, affecting basic behaviors such as locomotion, feeding and egg laying, or more complex behaviors such as mating, lethargy and learning (Lim et al., 2016).

The promiscuous expression in the nervous system of the genes responsible for coding neuropeptides in C. elegans has led to the belief that all nematode neurons are capable of secreting neuromodulators, suggesting distributed peptidergic signaling (Holden-Dye and Walker, 2013). Subsequently, it was possible to elucidate that neurons have mixed characteristics (synapsis and secretion) or predominantly secretory characteristics, the difference is in the amount of DCVs available, which are located on the periphery of the presynaptic terminals (Lim et al., 2016). The release of neuropeptides from DCVs is dependent on an increase in calcium in the nerve endings driven by high levels of stimulation, provoking different responses in different tissues. The functions of different neuropeptides remain largely unknown in C. elegans (Li, 2008). However, in the last decade studies on the behavioral effects caused by neuropeptides in response to different environmental responses have aroused interest among scientists.

It was in this sense that (Lim et al., 2016), characterized a peptidergic neuron in the motor system of C. elegans, called RID, which had its anatomy, development and function well characterized by the authors. Moreover, it was observed that neuron activity increased during forward movement RID harbors DCVs that cluster periodically along the axon and expresses several neuropeptides, including the FMRF amide-related FLP-14. This neuronal circuit is well preserved in other invertebrates, including the nematode Ascaris suum.

RID neurons have also been attributed to participate in a neuropeptide signaling cascade associated with behavioral arousal in C. elegans. The authors determined that locomotor and sensory arousal behaviors require FLP-20 neuropeptides released directly from primary mechanosensory neurons (ASH) and their receptor FRPR-3. FRPR-3 modulates the activity of the RID endocrine neuron, which in turn is activated in response to mechanical stimulation in a FLP-20 and FRPR-3-dependent manner. Thus, it was established that the peptides secreted by the RID neuron are necessary for both locomotion and sensory excitation in C. elegans (Chew et al., 2018; Lim et al., 2016; Rabinowitch et al., 2016).

Other neuroendocrine modulations commonly observed in several species, including mammals, have been described in C. elegans. The “brain-gut” relationship previously described as the neuroendocrine basis for different behavioral responses in mammals has already been described in studies with C. elegans for its ability to regulate longevity in the worm (Wang and Wang, 2016). Longevity is regulated bidirectionally through neuroendocrine signaling in C. elegans. Two brain neuroendocrine signaling circuits have been identified where worms detect and transform environmental temperature inputs into opposing longevity outputs. The “cold” circuit depends on small neurotransmitters, while the “hot” circuit depends on neuropeptides, where it was confirmed that ASJ neurons signal the gut to shorten lifespan at higher temperatures, in a daf-16-dependent manner (Zhang et al., 2018).

The genetic role of the intestine in neurodevelopment was determined by discovering that Wnt signaling, a well-conserved pathway between mammals and C. elegans, actively participates in synaptogenesis, being able to promote synaptic assembly through positive regulation of the expression of PNL-40 peptides secreted in the intestine, which in turn act through the GPCR/AEX-2 receptor expressed in neurons. Thus, the Wnt pathway in the intestine regulates presynaptic formation in the nerve ring allowing synaptic formation (Shi et al., 2022).

The role of 5-HT in fat oxidation in C. elegans has been attributed to the neuroendocrine receptor FLP-7. This neuropeptide is secreted by ASI sensory neurons in response to 5-HT neural signaling and acts in the intestine through the NPR-22 receptor, mobilizing fat reserves. Through the results found, it is suggested that ASI sensory neurons integrate nutritional information to regulate multiple physiological outputs. The neuroendocrine axis for lipid metabolism mediated by 5-HT was confirmed by the need for the presence of the NPR-22 receptor and the FLP-7 ligand for the oxidative effect on fat storage to be observed. Furthermore, this mechanism does not influence other 5-HT-dependent behaviors such as locomotion, reproduction and feeding (Littlejohn et al., 2020; Palamiuc et al., 2017).

Temperature and diet are factors that determine the processing of environmental signals in the nematodes. In a study evaluating the effects of temperature on the neuroendocrine system, it was proven that the life expectancy of worms is reduced when animals are exposed to high temperatures through the signaling of a sensory neuron, ASJ, capable of secreting two insulin-like peptides, INS -6 and DAF-28. In turn, these can associate with the DAF-2 in the intestine, activating pathways responsible for reducing the lifespan of the worms (Zhang et al., 2018).

Another study demonstrated that the worm's ability to extend lifespan in response to food restriction occurs through signaling from the daf-7 gene that encodes a TGFβ ligand, which is secreted by ASI sensory neurons to control various behaviors in C. elegans. The authors observed that DAF-7 secreted by chemosensory neurons appears to be a key neuroendocrine signal responsible for allowing worms to adequately sense reductions in food availability resulting in activation of the DAF-16/FoxO pathway in the gut under food deprivation. Consequently, this allows the regulation of several genes associated with longevity. An age-dependent decline in neuronal daf-7 expression was also observed, which would be associated with decreased sensitivity of aged animals to the effects of dietary restriction throughout life, linking the reduction in neuroendocrine function to the loss of efficacy of dietary restriction with advancing age (Fletcher and Kim, 2017).

The DAF-7/TGF-β pathway has been also attributed to the neuroendocrine regulation of several aspects of worm development and physiology, including dauer development decision, foraging and aggregation behaviors, quiescence, metabolism and longevity (Gallagher et al., 2013; Milward et al., 2011; Shaw et al., 2007; Meisel et al., 2014) demonstrated that chemosensory recognition of the pathogenic bacterium P. aeruginosa intensely alters the neuronal expression pattern of DAF-7, promoting avoidance behavior. Furthermore, sexual differences were also observed for the expression of DAF-7/TGF-β. It has been reported that C. elegans males display an altered, male-specific expression pattern of daf-7 in the ASJ sensory neuron pair with the onset of reproductive maturity, which functions to promote mate-seeking behavior, thus participating in the worm's decision-making behavior (Hilbert and Kim, 2017).

Sensory mechanisms in C. elegans are also associated with expulsive behavior, defecation and egg laying, which act in a coordinated way in worms. In this study, it was established that HSNs neurons signal changes in the internal hydrostatic pressure of nematodes. This pressure increases as food intake and fertilization occurs, activating mechanoreceptors that facilitate the initiation of defecation and egg laying (Wilcox et al., 1981).

Furthermore, aversive sensory signals inhibit egg-laying circuit behavior through activation of the major G protein Gα -o (GOA-1) in presynaptic HSNs. Additionally, in the same way as endocrine cells, cholinergic neurons in the vulvar muscle are mechanically activated when the muscle contraction begins to open the vulva, releasing acetylcholine that will participate in the total opening of the vulva for the efficient release of the egg. This muscular activity of the vulva is also related to mating behavior during the insertion of the male spike, allowing the withdrawal of the spike and the ejection of male sperm (Ravi et al., 2021).

The neuroendocrine relationship of the olfactory circuit to peripheral transcriptional control and lipid metabolism has recently been determined. A study conducted by Mutlu et al., (2020) demonstrated that AWCON neurons and their corresponding environmental odor specifically can directly control the rate of catabolism of lipids stored in peripheral tissues in C. elegans. Through pulsed SRS microscopy and lipid chase labeled with deuterium, it was possible to determine the response to the chemical agent 2-butanone in the activation of a specific neuronal circuit in AWCON neurons. As a result, it was possible to observe the activation of the neuronal circuit that allows the control of the rate of lipid catabolism in peripheral tissues. 2-Butanone can be produced by the decarboxylation of β-keto acids that form during the oxidation process of fatty acids in bacteria and fungi, being associated with the scarcity of lipid sources in the environment. Therefore, the olfactory signaling caused by the chemical agent signaled by the low availability of fat in the environment triggers the animals' metabolic adjustment towards fat accumulation.

The autophagy gene atg-18 has also been described as a neuroendocrine modulator of fat metabolism in C. elegans at the dauer stage. In this study it was demonstrated that through chemosensory neurons, with the exception of ASH neurons, present in the intestine, the atg-18 gene acts as a neuroendocrine modulator signaling daf-2 and consequently increasing lipid accumulation in daf-2 dauer stage worms. However, the role of the atg-18 gene in the lipid metabolism of worms at other larval stages has not yet been elucidated (Jia et al., 2019).

Although many gaps still need to be filled regarding the relationship between the nervous and endocrine systems in C. elegans, the available studies provide evidence that demonstrates the nematode's high capacity to be a good model for investigating environmental influences on the neuroendocrine system. This fact is attributed to the simplicity of the model in a complete neuronal network and transparency of the body, complementing the genetic tool available for study. Furthermore, C. elegans shares neuroendocrine modulation pathways with other species, including mammals, allowing to evidence and understand circuits that have not been clarified in these models yet.

The nematode C. elegans is a well-standardized model for pharmacological research, both due to its easy of obtaining, maintenance and manipulation. As the nematode has a fully known genetic sequence, made up of approximately 19,000 genes, it is worth highlighting that this is an advantage of the model in the investigation of more complex biological processes, due to the availability of strains with loss (mutant) or gain of gene function, and target proteins tagged with fluorescent proteins (transgenic). This makes possible elucidating biochemical pathways and mechanisms of drug action through the fast and low-cost screening of new molecules with therapeutic potential, or through drug repositioning. Furthermore, the model brings a variety of behavioral tests that are directly related to the functioning of the neuroendocrine system (Bettinger et al., 2004; Tejeda-Benitez and Olivero-Verbel, 2016).

Firstly, it is important to highlight that hormonal signaling is mainly involved with the lifespan of C. elegans, therefore steroid hormones are involved in this regulation. As an example, we can bring a study carried out with pregnenolone that regulates aging. This is because in concentrations between 7.5 and 60 μg mL-1 of pregnenolone, its useful life increases by up to 20%. It is suggested as a possible mechanism of action that pregnenolone is in the synthesis pathway of another lipophilic hormone that acts directly on DAF-12, a steroid receptor that, when activated, increases the life expectancy of the nematode (Broué et al., 2007).

Mostly known as a neurotransmitter, serotonin is also a peripheral hormone, being produced in the enterochromaffin cells of the intestinal mucosa. Notably, the gut produces about 95% of the serotonin in the human body (Kanova and Kohout, 2021). Hence, serotonin controls energy balance, from food intake behavior, satiety and fat regulation in vertebrates and invertebrates (Morton et al., 2006). In humans, drugs that increase central serotonergic signaling are approved for weight loss (Weintraub et al., 1984). Serotonin reuptake inhibitors have also been studied as nervous system modulators in C. elegans. For instance, fluoxetine administration reduced body fat accumulation and increased peristaltic speed in treated worms. These effects were independent of the insulin-like receptor DAF-2, but dependent on the serotonin receptor ser-1 and also serotonin synthesis enzyme tph-1 (Almotayri et al., 2021).

Drugs like fluoxetine act by blocking the reuptake of the neurotransmitter serotonin, making it more available in the synaptic cleft. Fluoxetine, like other drugs in the same class, also has a longer half-life, which contributes to an inhibitory action on cytochrome P450 isoenzymes (Lochmann and Richardson, 2019). They are prescribed for the treatment of depression, panic disorders, anxiety, and even appetite control. It is already known that the worm has a serotonergic system that controls some behaviors, such as egg laying stimulation, activates two feeding movements (pharyngeal pumping and isthmus peristalsis), locomotion inhibition and promotes lipid breakdown (Almotayri et al., 2021). For instance, the tph-1 mutant, presenting a defect in serotonin synthesis, exhibits hunger-like behaviors, entering the dauer stage even in the presence of food, and increasing its fat storage capacity.

Interestingly the effect of fluoxetine on reducing the formation of beta-amyloid aggregates (Ab) and the paralysis in transgenic strains, was observed. After exposure to fluoxetine and paroxetine, it was possible to observe that they delayed the paralysis of these worms compared to the control group, in addition to reducing the species of Ab oligomers in the strain. In contrast, citalopram did not show the same effect. It was also observed that the expression of the daf-16 and tph-1 genes is necessary for fluoxetine effect in these animals (Keowkase et al., 2010).

Another class of drugs that has been studied are antipsychotics. A study by Dwyer et al., (2015) aimed to evaluate the social feeding behavior of C. elegans, with npr-1 knockout strains, after exposure to drugs of this class. This strain presents changes in the feeding behavior of the nematode with aggregation behavior, different from the wild type strain. Chlorpromazine and trifluoperazine have been shown to significantly inhibit social eating, as well as first-generation (fluphenazine and haloperidol) and second-generation (olanzapine) antipsychotics. To prove the effect presented, treatment with antagonists (sulpiride and raclopride) was also tested and a similar response was observed, although slightly less effective than medications.

The drugs of abuse can also be investigated in the nematode. A study exposed worms to methamphetamine (METH) and ketamine (KET) at concentrations already found in the environment (0.05–0.5 μg L-1) and demonstrated that, at the lowest concentrations, toxicity was seen in the rate of feeding, locomotion, chemosensation and changes in the vulvar morphology (Wang et al., 2019). In addition, they caused changes in the levels of neurotransmitters responsible for these behaviors, such as octopamine, dopamine and serotonin.

The food avoidance behavior assay is an important endpoint that can be assessed. Caffeine exposure to the nematodes causes a modulation of neuroendocrine signals, and at high concentrations it has toxic effects on development, in addition to inducing food avoidance, directly affecting the nematode's survival. This behavior is a form of defense in aggressive environments and is associated with cellular stress responses, involving the serotonergic, dopaminergic and JNK/MAPK kinase signaling pathways. The lack of these neuroendocrine signals suppresses the behavior, which were recovered after treatment with caffeine. Dopamine biosynthesis and transport are also determinants for its homeostasis, both in humans and C. elegans, but its transcription factors still remain obscure. Furthermore, what is known is that it promotes induction of HSP-16.2 and expression of the jnk-1, hsp-16.2, cyp-35A2 and cyp-35A4 genes (Min et al., 2017).

With a greater focus on the endocrine system, pharmacological interventions that perform actions such as peptides and their possible signaling pathways are now being speculated. An important pathway related to the endocrine system is the insulin signaling, where the genes age-1 (which encodes a PI3 kinase), daf-2 and daf-16 are involved. These are directly involved with the life span and larval development of the nematode, which can be modulated by environmental factors such as temperature, food availability, population density and also drugs. For instance, superoxide dismutase and catalase mimetics, such as euk-134, are being studied. Anticonvulsant drugs are also being investigated, molecules that target the acetylation status of histones. Furthermore, this study again mentions the relationship between the signaling of the nuclear hormone receptor DAF-12, which is responsible for the secondary hormonal response and is the only insulin and growth factor receptor similar to that of mammals. But it is not only this pathway that is involved, but also that of TGF-β, and, therefore, its reduced signaling resulted in the formation of dauer larvae (Gill, 2006).

Therefore, the perspectives for the search for new therapies such as the use of hormones and the use of nematodes as a model should be further explored, such as in the context of treating metabolic diseases and aging. Furthermore, discover the mechanisms by which these steroid hormones act and find the receptors responsible for their activity, such as NHR, in the case of DAF-12, which regulate gene expression, and the selective modulators of these receptors, these answers being more specific to find a promising molecule. This model is important for these findings mainly because it has a variety of receptors, such as the NHR with 284 receptors in nematodes, whereas only 48 in humans, and seeks the similarities of each one in search of new pharmacological interventions (Gill, 2006).

The endocrine system coordinates communication between the body cells for the transport of information through chemical stimuli. This exchange of information occurs through the secretion of hormones that act as chemical messengers, interacting with specific receptors. The cellular response occurs according to the specific receptor, which are macromolecular structures of a non-protein nature. They are found in the membranes, cytoplasm and cell nucleus, and through them the genetic and biochemical mechanism can function (Tejada et al., 2014). The endocrine system is basically responsible for controlling and coordinating the functioning of the organism, participating in the processes of growth, development, reproduction and energy levels (Chrousos, 2007).

As previously mentioned in this review, endocrine disruptors are molecules that can be chemically similar or not to hormones, which can generate an endocrine response in the organism. Therefore, they are substances that can interfere with the natural hormonal processes such as hormone production, release, transport, metabolism, action and elimination. These disruptors can be classified according to their origin: natural (estrogens and androgens), semi-synthetic (contraceptives) or synthetic (xenoestrogens) (Diamanti-Kandarakis et al., 2009). Endocrine disruptors can act as agonists, presenting additive or synergistic effects on the hormone's action, neutralizing the actions and decreasing their activity. In addition they can act by altering hormonal metabolism, modifying the metabolic routes of hormone synthesis and degradation and modifying levels of hormone receptors and their activity (Domańska et al., 2021).

Some of these disruptors are synthesized and produced with commercial purposes such as controlling pests in agriculture, stabilizing aesthetic products, and forming the structure of certain plastics. However, the harmful health and environmental effects of these molecules following chronic exposure has been reported. We can highlight pesticides, such as dichlorodiphenyltrichloroethane (DDT) and dichlorodiphenyldichloroethane (DDE), industrial detergents that contain nonylphenol and octylphenols, compounds used in the production of plastics such as BPA A, and also in industrial waste such as chromium, trichloroethylene and tetrachloroethylene (Vieira et al., 2005). Some of them have been called emerging pollutants, as they have been recently identified as hazardous and little or no regulatory control is established (Morris, 1995). These micropollutants that act as endocrine disruptors can also cause adverse effects on aquatic life, even at low concentrations. The main sources from where these micropollutants can reach surface water and harm aquatic life are through sanitary sewage, water treatment station, industrial waste and agricultural runoff. Notably, the lack of regulation in the reference levels of these compounds in aquatic environments increases the pollution in these environments (de Rezende and Mounteer, 2023).

Several studies have demonstrated that C. elegans is an important model for the study of endocrine disruptors, mainly through toxicological analysis of survival, size, behavior and reproduction. Through these analysis, Nagar et al., (2020) developed a study on the action of endocrine disruptors in C. elegans, through exposure to parabens, where they demonstrated an endocrine disruptive activity affecting the expression of the vitellogenin gene and causing oxidative stress, in addition to affecting growth, behavior and reproduction of the worms. In another study, a mixture of six parabens (methyl p-hydroxybenzoate [MP], ethyl p-hydroxybenzoate [EP], propyl p-hydroxybenzoate [PP], isopropyl p-hydroxybenzoate [IPP], butyl p-hydroxybenzoate [BP], and isobutyl p-hydroxybenzoate [IBP]) caused reduced egg maturation and fertility rate. Parabens are common phenolic preservatives added in pharmaceuticals, cosmetics and children's products, but they are also associated with estrogenic effects and several concerns for human health in general, including impacts on hormonal and reproductive systems.

BPA is a synthetic chemical substance belonging to the category of diphenyl calcane polycarbonates and is widely present in plastics and epoxy resins. Its presence raises concerns due to possible negative effects on human health and the environment. Although research into the effects of BPA is still new, several studies indicate that it may have an unfavorable impact on fertility and reproduction in women and certain animals, including fish and mammals (Mukherjee et al., 2024). As in other animal models, BPA caused embryonic lethality, germinal apoptosis and reproductive toxicity in C. elegans, but notably, this was reversed by exogenous cholesterol supplementation, revealing another mechanism of BPA germinal toxicity through the alteration of cholesterol transport. Notably, cholesterol is a precursor of steroid hormones and necessary for molting and development in C. elegans (Chen et al., 2019).

As the recent studies with BPA have led to its ban in several countries, industries have used other BPA analogues, such as BPS and BPF, which are currently being assessed. These new compounds have been found in various environments and have demonstrated a certain harm to the reproductive system and male fertility. In C. elegans, the toxicity effects vary from increasing stress response protein expression (SOD-3 and GST-4), inducing genotoxicity and apoptosis and an indirect reaction occurs through endocrine dysregulation Ficociello et al. (2021); Zhou et al., (2021) demonstrated in his study that BPS can increase the obesogenic effects of a high glucose diet through regulating the lipid metabolism. This effect was noted after an analysis of fat accumulation in wild-type worms, while mutant worms deficient in the daf-16 gene showed no change, emphasizing that this effect may be mediated by the DAF-16 pathway in C. elegans. In addition, BPS positively regulates the fat-7 gene, which acts in lipid synthesis and is regulated by daf-16. The study also revealed that BPS can affect nhr-49 by negatively regulating this gene, which is an important regulator of lipid metabolism, also reducing the expression of the acs-2 gene, which is involved in the fatty acids β-oxidation. This effect reduces energy consumption, generating an accumulation of fat in C. elegans, which is transferred to the next generations as well, characterizing a multigenerational effect.

As previously mentioned in this literature, neurotransmitters such as serotonin, dopamine, GABA and glutamate undergo changes and are influenced by endocrine modulators, which can stimulate their synthesis, activate or inhibit neurons that are associated with the worm's behavior. Carboxyl-modified polystyrene microplastics (PS–COOH) have been shown to have a significant impact on locomotion behavior, neuronal development, neurotransmitter levels and gene expression in C. elegans (Yu et al., 2023). Chen et al. (2021) investigated the impact of chronic exposure to aged microplastics under UV radiation, which damaged dopaminergic, glutamatergic and serotonergic neurons, affecting the worm's locomotion. Furthermore, glutamate and dopamine levels decreased, while serotonin increased, suggesting a disturbance in the worm's neurotransmission system mainly due to alterations in genes related to this system (eat-4, glt-3, glt-7, dat-1, dop-1, mod-1, tph-1) (Yu et al., 2023).

Steroids have an important function in nematodes. Ghosh et al. (2021) highlights that the endocrine system in C. elegans is regulated by hormonal pathways, and mentions the potential of endocrine-disrupting chemicals (EDCs) to act on this system at molecular levels. The study highlights that these chemicals can only be called endocrine disruptors if their dysregulation mechanism is elucidated, requiring knowledge about the hormonal regulation processes in C. elegans. Substances such as azasteroids and long-chain alkylamines can disrupt the metabolism of steroids in worms, inhibiting growth, reproduction and egg laying, in addition to inducing changes in larval development (Höss and Weltje, 2007).

Triclosan (TCS) is an antimicrobial chemical with endocrine-disrupting properties, and also demonstrated a disruptive potential using the C. elegans model. This disruptor was able to inhibit the nuclear localization of SKN-1 and the expression of its target genes, which have been associated with oxidative stress. SKN-1 normally translocates from the cytoplasm to the nucleus, and there activates target genes such as gcs-1 and pmp-3, responsible for detoxification mechanisms and response to oxidative stress. However, TCS interrupts this process by inhibiting the translocation of SKN-1 to the nucleus, leading to a suppression of the expression of these genes Yoon et al. (2017); Alfhili et al., (2018) also evaluated the effect of TCS, showing that short-term treatment can cause mortality in a dose-dependent manner.

Pesticides are chemical substances used for pest control in agriculture and play an important role in food production and preservation, but they can also have side effects on human health and to the environment. Concerns about their adverse effects on human health have been mainly attributed to the neurotoxic effects, immune system disorders and interference with hormonal signaling. These pesticides can act as EDCs causing reproductive problems. The assessment of EDCs is complex due to their mechanisms of action and variability in concentration and life stages of organisms (Chen et al., 2019). The nematode C. elegans has been an alternative for this type of analysis due to its short life cycle and the use of fluorescent markers to study these biological processes. Moya et al., (2022) demonstrated the reproductive toxicity of these EDCs when compared to an estrogenic control 17β-estradiol. The study tested the reproductive toxicity of several pesticides including Atrazine, 2,4-dichlorophenoxyacetic acid, Chlorpyrifos, Cypermethrin and Mancozeb, and demonstrated that these can cause a decreased brood size, vitellogenin disruption and gene expression like vit-2 and hus-1, showing that C. elegans it can be a valuable model for assessing ecological risks of EDCs, aiding environmental and ecosystem protection.

According to the studies, it is possible to observe the importance of the C. elegans model for the analysis of endocrine disruptors through toxicological analyses, in addition to the possibility of being used as a screening model in more complex processes. Although nematodes are often neglected in ecotoxicological studies, C. elegans has gained space in studies mainly on endocrine disruption, and due to its genetic sequencing, researcher's interest in working with this nematode has increased. Nematodes can act as bioindicators to evaluate endocrine disruption in various types of ecosystems, due to their ease in carrying out studies in sediments, soils and aqueous media, in addition to evaluating genomic responses using mutant strains with specific dysfunctions (Höss and Weltje, 2007).