Environmental Contaminants and Endocrine Health

Environmental Contaminants and Endocrine Health focuses specifically on contaminants with hormonal disrupting activities. The book provides insights into the multiple effects of endocrine-disrupting chemicals (EDCs) and their mechanism of action (MoA) on metabolism, reproduction and the multiple physiological roles of the endocannabinoid system which has recently been indicated as new target. The content systematically covers EDC sources and effects, EDCs as sources of disease and health impairment in laboratory models, EDCs as the cause of disease and health impairment in humans and wild species, and the removal of hazardous pollutants from wastewaters to highlight intervention, mitigation and adaptation for reduced threat.

This content will be a foundational resource for academic and research staff in endocrinology and hormone toxicology as well as for professors, researchers and students in these areas.

• Includes important foundational coverage of the endocrine system, definitions of EDC sources and descriptions, model examples and mechanisms of action biological effects
• Provides coverage of EDC effects in humans and animals, from metabolic alterations to epidemiological studies of fertility and metabolism
• Presents insights into the confirmed and suspected human diseases spectrum with origins linked to EDC exposure, including cancers, intellectual disabilities, autism, birth defects of the urethra (hypospadias), decreased sperm count, increased rates of miscarriage, obesity, type 2 diabetes, and more

• Language: English
• Publisher: Academic Press
• Release date: May 30, 2023
• ISBN: 9780323859325

Book preview

Chapter 1

Endocrine-disrupting chemical sources and effects

Chapter 1.1: Endocrine system

Thomas M. Galligana,b,⁎; Alexis M. Temkinb,⁎; Matthew D. Halec,⁎    a Center for Science in the Public Interest, Washington, DC, United States

b Environmental Working Group, Washington, DC, United States

c Department of Biology, University of Virginia, Charlottesville, VA, United States

⁎ All authors contributed equally to this work.

Abstract

Intercellular communication is fundamental to vertebrate life. The body must coordinate a variety of complex physiological and behavioral processes to maintain homeostasis, respond to environmental changes, and reproduce. The endocrine system evolved as a means of intercellular communication operating across spatial scales within the body, integrating the functions of diverse tissues into coordinated systems. It accomplishes this through the production, release, and sensing of hormone signaling molecules. Hormones are secreted by so-called endocrine glands and carried throughout the body via blood to cells expressing hormone receptors. Hormone receptor binding and activation in turn triggers diverse cellular responses in target tissues, which collectively produce coordinated physiological effects at multiple spatial scales. In this chapter, we provide a brief overview of vertebrate endocrine physiology. Section 1 overviews hormone classification, synthesis, and signaling from a molecular perspective, including key biochemical characteristics of hormones and hormone receptors. Section 2 covers basic systemic endocrinology, with focus on the endocrine organs/axes and hormones commonly studied in the context of environmental endocrine disruption.

Keywords

Endocrinology; Hormone; Metabolism; Reproduction; Thyroid; Adrenal; Gonad; Pituitary; Hypothalamus; Pancreas; Adipose

1: Fundamentals of hormones

Hormone-receptor interaction and downstream cellular responses are determined by the molecular characteristics of both the hormone and the receptor. In this section, we overview the biochemical classes of vertebrate hormones and functional classes of hormone receptors and discuss other relevant components of molecular hormone signaling.

1.1: Hormone classes

Hormones can be organized into three major chemical classes: lipid-derived hormones, amine-derived hormones, and peptide or protein hormones.

1.1.1: Lipid-derived hormones

Lipid-derived hormones are small molecular weight, lipophilic molecules derived from cholesterol or arachidonic acid. The major subclasses of lipid-derived hormones are steroid hormones (derived from cholesterol) and eicosanoids (derived from arachidonic acid). Each of these subclasses is composed of numerous functionally diverse, yet structurally similar, hormones.

Steroid hormones

Steroid hormones generally are grouped into classes reflecting their structure and function, namely estrogens (18 carbons), androgens (19 carbons), progestogens (21 carbons), and corticosteroids (21 carbons). The biosynthetic pathways giving rise to each of the four classes of steroid hormones share common elements, including precursors, intermediates, and enzymes, are synthesized from cholesterol, giving them a common backbone structure composed of three cyclohexane rings and one cyclopentane.

Steroid hormones are functionally diverse. Progestogens, estrogens, and androgens are commonly associated with reproductive physiology, though their physiological roles are not limited to reproduction. Progestogens, including progesterone and 17α-hydroxyprogesterone (17OHP4), are typically discussed in the context of pregnancy and the luteal/secretory phase of female reproductive cycles. Further, as the most upstream class of steroid hormones, progestogens serve as precursors for the remaining steroid classes. Although often considered male and female sex hormones, respectively, androgens and estrogens each play critical roles in both sexes, including regulating sexual differentiation and development. In humans, the androgens include precursors, dehydroepiandrosterone (DHEA) and androstenedione (A4), and active androgens, testosterone (T) and 5α-dihydrotestosterone (DHT). The major estrogen in most vertebrates is 17β-estradiol (E2), synthesized from testosterone. Other minor estrogens including estrone and estriol. Lastly, corticosteroids are predominantly discussed in the context of stress, but are important regulators of metabolism and osmoregulation, among other processes, and are subdivided into two groups, glucocorticoids (GCs) and mineralocorticoids (see "Corticosteroids: Glucocorticoids and mineralocorticoids" section). The major GCs in vertebrates are cortisol and corticosterone, and the major mineralocorticoid is aldosterone.

De novo biosynthesis of androgens, estrogens, and corticosteroids primarily occurs in the testes, ovaries, and adrenal glands, respectively, although both androgens and estrogens are also produced by the adrenals. Other tissues, such as the placenta, are also known to perform de novo steroidogenesis. The rate-limiting step in steroidogenesis is the transport of cholesterol to the inner mitochondrial membrane where it is metabolized to pregnenolone, a progestogen and precursor steroid common to all steroid hormone biosynthetic pathways. Thereafter, further metabolic transformations leading to the synthesis of bioactive estrogens, androgens, progestogens, and corticosteroids are carried out by two classes of enzymes, cytochrome P450 (CYP) enzymes that catalyze hydroxylation and carbon bond cleavage reactions, and hydroxysteroid dehydrogenase (HSD) enzymes, which reversibly and bidirectionally convert hydroxysteroids to ketosteroids. As steroid hormone pathways share common precursors and enzymes, the production of specific steroid hormones in a given tissue is determined by tissue-specific expression of specific enzymes, which act to shuttle precursors toward a specific terminal hormone [1].

To produce corticosteroids, pregnenolone synthesized in the adrenal is converted by HSD3B2 to progesterone, which can be converted to mineralocorticoids or GCs. To produce aldosterone, CYP21A2 metabolizes progesterone to 11-deoxycorticosterone (DOC), and then CYP11B1/2 catalyzes three additional conversions: DOC to corticosterone, corticosterone to 18-hydroxycorticosterone, and 18-hydroxycorticosterone to aldosterone. GCs are generated through this same process—as noted above, corticosterone is produced as an intermediate in the aldosterone biosynthesis pathway—or, in the case of cortisol, a very similar process involving these same enzymes. One additional preliminary step is required for cortisol production; progesterone is first metabolized to 17OHP4 by CYP17A1. Thereafter, the pathway proceeds as above, with 17OHP4 being converted to 11-deoxycortisol by CYP21A2 and then 11-deoxycortisol to cortisol by CYP11B1 [1].

Androgens (and thus, estrogens) can be synthesized via one of two parallel pathways, the Δ4 pathway or the Δ5 pathway. The Δ4 pathway proceeds from 17OHP4, which is metabolized to A4 by CYP17A1 and then to T by HSD17B. The Δ5 pathway proceeds from pregnenolone. CYP17A1 metabolizes pregnenolone to 17α-hydroxypregnenolone and then to DHEA. T is produced from DHEA through one of two intermediates, A4, produced via HSD3B2, or androstenediol, produced through HSD17B. From here, testosterone may be converted to DHT by the testes as well as other peripheral tissues. Androgen synthesis occurs in the adrenal glands, testicular Leydig cells, and ovarian theca cells. Testosterone and DHT are secreted by the testes, while DHEA and DHEA-sulfate are the major androgenic products of the adrenal glands and can later be metabolized to active androgens in peripheral tissues. Theca cells of the ovary produce A4and T which is then converted to E2 by aromatase (CYP19A1) in ovarian granulosa cells, secreted during the follicular phase of the estrous cycle (proliferative phase of the menstrual cycle) [1].

Across vertebrate species, steroid hormone structure is chemically identical therefore highly conserved in structure, yet certain hormone functions differ considerably across different taxa. In teleost fishes, the major androgen is 11-ketotestosterone (11-KT), which is produced from 11-β-hydroxy-androgens, metabolites unique to fishes [2] (though 11-keto-androgens have also been observed in amphibians) [3]. In elephants, the dominant progestogens are 5α-reduced metabolites instead of progesterone, including 5α-pregnane-3,20-dione and 5α-pregnane-3α-ol-20-one [4]. In horses, the major circulating estrogen during pregnancy is not estradiol but estrone and other unique estrogens, equilin and equilenin, are observed as well [5]. Lastly, while cortisol is the dominant GC in humans, many other mammals, and fish, corticosterone fulfills the same role in rodents, birds, reptiles, and most amphibians.

Eicosanoids

Eicosanoids, synthesized primarily from arachidonic acid, include prostaglandins and thromboxane (produced through the cyclooxygenase enzymatic pathway [COX]), and leukotrienes and lipoxins (produced through the lipoxygenase enzymatic pathway [LOX]). Additionally, cytochrome P450-catalyzed reactions yield epoxides, an additional subset of eicosanoids. Other fatty acids can give rise to eicosanoids, including eicosapentaenoic- and dihomo-ɣ-linoleic acid. Eicosanoids are involved in regulating angiogenesis, vascularization, inflammation, and immune function, and, furthermore, prostaglandins, are important in late-stage pregnancy to initiate parturition [6,7]. Unlike steroid hormones that typically travel to target organs via the circulatory system, eicosanoids are involved in local signaling, acting on or near the cell in which they were produced, in an autocrine and paracrine manner.

To produce eicosanoids, cells first produce arachidonic acid, a -20-carbon polyunsaturated fatty acid, from membrane phospholipids by phospholipases. Eicosanoids can be metabolized entirely within a single cell or in a specialized multicellular process, known as transcellular metabolism, where one cell produces an intermediary and another cell produces the terminal molecule [8]. Three major enzymatic pathways convert arachidonic acid to eicosanoids. The cyclooxygenase enzymes (COX1 and COX2) catalyze reactions to produce prostaglandins (e.g., PGE2, PGD2, and thromboxanes) via intermediary PGG2 and PGH2 compounds. Prostaglandin structure includes a hydroxylated five carbon ring connected via C8 and C12 of arachidonic acid with varying: hydrocarbon sides chains, carbon–carbon double bonds, and functional groups on the ring structure [9]. Thromboxane species differ by the addition of an oxygen atom in the ring. COX2 is typically upregulated in response to cellular stimuli leading to the production of inflammatory prostaglandins while COX1 is constitutively expressed. Given the role of these molecules in the inflammation response, COX2 is the target of nonsteroidal antiinflammatory drugs (NSAIDs).

Lipoxygenase enzymes (LOX) catalyze arachidonic acid into lipoxins and leukotrienes via hydroperoxyeicosatetraenoic acids (HPETEs) and hydroxyeicosatetraenoic acid intermediates. There are over a dozen different lipoxygenase enzymes, which are expressed in different types of immune cells, yielding structurally similar yet distinct lipoxins [10]. Physiologically, lipoxins play an important role in antiinflammatory processes, while leukotrienes possess proinflammatory activity [11].

While some eicosanoids are well studied, and the majority of their biological functions well described, there are several eicosanoids with limited data and novel eicosanoids are still being identified [12]. Furthermore, the role of eicosanoids in nonmodel organisms is poorly characterized. Evidence suggests prostaglandins are important for development in reptiles and birds and have been identified in the chorioallantoic membrane of the American alligator and chicken [13]. Other work has identified prostaglandins and thromboxanes in marine organisms including teleosts, elasmobranchs, corals, mollusks, and crustaceans, with suspected biological activities impacting reproduction and defense [14].

1.1.2: Amino acid-derived hormones

Amine-derived hormones include thyroid hormones (THs; Section 2.3), melatonin, and catecholamines (Section 2.2). Catecholamines and THs are produced from tyrosine, and melatonin is derived from tryptophan. Like lipid-derived hormones, amine-derived hormones are chemically identical across many species.

Catecholamines are synthesized in the adrenal glands as well as in dopaminergic and noradrenergic neurons. In the adrenal gland, tyrosine hydroxylase catalyzes the addition of a hydroxyl group to tyrosine's six carbon ring to form DOPA. Decarboxylation of DOPA by aromatic l-amino acid decarboxylase yields dopamine. Dopamine β-hydroxylase adds another hydroxyl group to the carbon side chain to form norepinephrine (NE). Finally, phenylethanolamine N-methyltransferase adds a terminal methyl group to yield epinephrine. Different enzymes catalyze similar reactions to form catecholamines in neuroendocrine cells [15].

THs, thyroxine (T4), and triiodothyronine (T3) are synthesized from tyrosine in the thyroid gland. A unique characteristic of THs synthesis and structure is the incorporation of iodine. See Section 2.3.3 for a more detailed overview of TH biosynthesis.

Melatonin is synthesized in the pineal gland in the brain, with synthesis occurring at night, as it plays a role in circadian rhythms. From tryptophan, serotonin is produced through hydroxylation by tryptophan hydroxylase and subsequent decarboxylation by aromatic amino acid decarboxylase. Serotonin is then acetylated by serotonin N-acetyltransferase and then methylated by N-acetylserotonin O-methyltransferase to form melatonin. Melatonin is found across vertebrates and the animal kingdom, as well as in plants and bacteria [16].

1.1.3: Peptide hormones

Peptide hormones are typically high molecular weight, hydrophobic structures that may or may not contain post translational modifications such as glycosylation. Unlike lipid- and amine-derived hormones, peptide hormones are encoded by genes and must therefore be transcribed and translated. As such there is greater interspecies variability in structure for these hormones. Despite this relative increase in variation across species, gene homologs for peptide hormones exist across vertebrates, especially mammals, and many peptide hormones fall into conserved families.

Peptide hormone biosynthesis proceeds according to standard transcriptional and translational processes Alternative splicing of transcripts permits a single gene to produce more than one peptide hormone from a single gene, as has been observed for the peptide growth factor, IGF1 [17]. Further, many peptide hormones are heterodimers, such as TSH, LH, and FSH. In these instances, individual subunits are derived from two separate genes, the products of which must physically associate. Posttranslational modification of certain peptide hormones can influence bioactivity and stability (see Section 2.3.2 for example). Peptide hormones are also commonly secreted as inactive precursor peptides, which can be cleaved into the active hormone by peptidases at specific signal recognition sites. These precursor peptides are referred to as prehormones when one cleavage event is required, or preprohormones, when two cleavage events occur before the terminal hormone is produced, as is the case with insulin., We discuss a wide variety of peptide hormones, including their structures and physiological roles, in Section 2.

1.2: Hormone receptors: Nuclear and membrane receptors

Hormones enact their cellular functions by binding receptors expressed by cells. Hormone receptors are ligand-activated proteins expressed by hormone-responsive, or target, cells. Each hormone or class of hormone generally binds a specific, cognate receptor (e.g., T and DHT bind the androgen receptor), though a hormone may associate with multiple receptors with varying affinity and single receptors may also bind multiple hormones. There are instances where multiple receptor isoforms occupy distinct functional niches. Activation of a hormone receptor alters cellular physiology through a wide variety of intracellular signaling pathways and mechanisms. The nuclear receptors, commonly known as steroid and nonsteroid nuclear hormone receptors, are transcription factors that directly regulate gene expression via binding to gene promoter and/or enhancer regions. Transmembrane receptors activate signaling cascades that influence cellular function through both genomic and nongenomic mechanisms. Thus, transmembrane receptors may ultimately lead to changes in gene and protein expression, but do not associate with DNA themselves.

1.2.1: The nuclear receptor superfamily

The nuclear receptor superfamily, which includes receptors for steroid and THs, shares a general structure including ligand-binding domains and DNA-binding domains. Unbound receptors reside in cytosolic compartments of the cell and, upon ligand binding, translocate to the nucleus to bind DNA at receptor-specific sequence motifs (referred to as response elements), recruit additional coregulating proteins, and ultimately regulate gene transcription. These receptors fall into three classes. Class I receptors form homodimers to control transcription. Class II receptors are those that heterodimerize with the retinoid-X-receptor (RXR) or another receptor. A third class of receptors are commonly referred to as the orphan receptors, originally named due to the lack of identification for endogenous ligands [18]. Table 1 lists the major nuclear hormone receptors within each of these classes, their endogenous ligands, experimental ligands commonly used as positive controls or developed as therapeutics, as well as tissues with receptor expression.

Table 1

a Tissue expression information summarized from The Human Protein Atlas ([19]; https://www.proteinatlas.org) and references cited throughout the text.

b Synthetic ligands developed as drug treatments or for experimental purposes.

Class I and class II nuclear receptors are structurally similar in that they are made up of two major subunits. The C-terminal domain of the receptor contains a ligand-binding domain (LBD), the N-terminal contains the DNA-binding domain (DBD), and the two are separated by a variable hinge sequence. Within the LBD domain, there are binding sites for receptor-associated ligands as well as coactivator proteins, which assist in gene transcription through interaction with chromatin remodeling proteins and other transcriptional machinery. The DBD is highly conserved within the receptor family and functions to bind hexanucleotide response elements located in the enhancer and promoter regions of genes [20]. Most steroid receptors (AR, MR, PR, GR) recognize variations of a shared palindromic motif composed of inverted repeats, separated by a short linker region (GGTACAnnnTGTTCT). Estrogen receptors are an exception, recognizing the sequence AGGTCAnnnTGACCT [21]. Steroid receptors can also regulate transcription via binding to response element half sites and direct, as opposed to inverted, repeat motifs. Class II receptors including peroxisome proliferator-activated receptors (PPARs) recognize a specific response element as well, commonly referred to as PPRE, which resembles estrogen response element half site (AGGTCA) tandem repeats [22]. Not only is there strong conservation between sequences of nuclear receptors within the same species, but the same receptors are highly conserved across different species within the vertebrate class. In a recent analysis of nuclear receptors from 12 vertebrate genomes, the most conserved receptors, based on greater than 90% similarity to humans, were THRs, RARs, and RXRs, while the similarity of ERs, ARs, and PPARs had sequence similarities of 60%–100% [23].

1.2.2: Transmembrane receptors

Transmembrane receptors span the lipid bilayer of cell membranes. These are commonly G-protein-coupled receptors (GPCR) but also receptor tyrosine kinases (RTKs) and type I cytokine receptors, each of which has unique signal transduction pathways that occur after ligand binding. Generally, despite falling into different receptor subtypes, these receptors all consist of an extracellular domain in which ligand binding occurs, a membrane bound domain, and an intracellular domain involved in activating downstream signaling pathways through recruitment of other signaling molecules.

GPCRs are made up of seven membrane-bound α-helical subunits, connected via intra and extracellular loop regions, and make up the largest group of receptors for peptide hormones, many of which are the family b subtype of GPCRs, but other hormone types also activate GPCRs [24]. One notable GPCR that binds a steroid hormone is the G-protein estrogen receptor (GPER or GPR30), which mediates nongenomic estrogen signaling. Other GPCRs include the ghrelin receptor, angiotensin receptors I and II, and receptors that bind prostaglandins and thromboxanes. A distinguishing characteristic of GPCR signaling is their coordination with G proteins, which are membrane-bound proteins made up of three subunits, α, β, and γ. When inactive, the trimeric G protein complex binds GDP and associates with a GPCR. Ligand binding induces a conformational shift that activates the G protein and induces the replacement of GDP with GTP. This subsequently causes the α subunit of the G protein to dissociate from the β and γ subunits, and both α and βγ subunits can continue cellular signaling through the involvement of secondary messenger molecules [25]. These secondary messengers are ligand/receptor specific but often include cyclic AMP (cAMP), formed through activation of adenylyl cyclase by the GTP bound α subunit, diacylglycerol (DAG) and inositol 1,4,5-triphosphate (IP3) both formed through activation of phospholipase C [26]. G proteins can also impact ion channels in the plasma membrane, changing the intracellular concentrations of ions like calcium which catalyze several downstream signals [27].

Adiponectin receptors are similar to G-protein-coupled receptors in that they have seven transmembrane sections, yet differ in that the N terminus resides in the intracellular domain, while the C terminus can be found in the extracellular region (opposite to the orientation of GPCRs) [28]. Additionally, they do not interact with G-proteins and have unique signal transduction pathways.

The RTK family includes the insulin receptor and growth factor receptors, which while not activated by hormones are activated by growth factor signaling molecules and typically control cellular processes such as proliferation, differentiation, cell migration, and cell cycle control. RTKs are composed of an extracellular ligand-binding domain, a single transmembrane domain, and an intracellular domain that contains tyrosine kinase. The insulin receptor is unique within this receptor superfamily in that it is expressed in the cell membrane as a dimer, while other RTKs typically dimerize or oligomerize after ligand activation [29]. After ligand binding, the receptor autophosphorylates tyrosine residues in the intracellular region which then serve as binding sites for subsequent docking and signaling molecules, for example, IRS1-4 in the case of insulin receptor serves to bind PI3K to generate phosphorylated inositol and activate the serine/threonine kinase AKT [30].

Leptin receptor has several different isoforms; however, the dominant form responsible for well-described neuroendocrine function is a type I cytokine receptor that can undergo phosphorylation at three tyrosine residues. Leptin binding initiates phosphorylation events mediated by janus kinase 2 (JAK2) followed by distinct signaling pathways and physiological responses mediated by mitogen-activated protein kinase (MAPK) and signal-transducer-and-activator-of-transcription 3 and 5 (STAT3, STAT5) [31].

1.3: Hormone transport, peripheral metabolism, and clearance

Once secreted, hormones must be transported to target cells—where they may be activated through further metabolism before binding their cognate receptor—and are eventually cleared from the body. In this section, we outline general themes of hormonal transport and metabolism/clearance physiology. We also highlight the involvement of the circulatory and hepatobiliary systems in these processes and discuss practical implications for the study of endocrine physiology in vertebrates.

1.3.1: Hormone transport, binding proteins, and the role of the circulatory system

Hormones operate across numerous spatial scales within the body. At the shortest scale, hormones can engage in autocrine signaling, a signaling pattern in which hormones act on the cell from which they were secreted. Paracrine signaling occurs at a slightly longer scale by acting between cells that are physically proximal to one another. Both autocrine and paracrine signaling involve hormonal transport across relatively short distances through the extracellular space. Conversely, endocrine signaling specifically refers to hormonal signaling between distal cells, mediated by transport through the circulatory system, and thus is the broadest spatial scale of hormone action in the body. Transport via the circulatory system also delivers hormones to tissues involved in hormone metabolism and clearance (e.g., the liver). Circulatory transport necessitates specialized transport for certain hormones.

Steroid and THs predominantly exist in circulation bound to proteins because they are poorly soluble in plasma due to their lipophilicity. Among these proteins are several that specifically bind hormones, including sex hormone-binding globulin (SHBG), which binds estrogens and androgens; cortisol-binding globulin (CBG), which binds corticosteroids, progestogens, and androgens in some species; and transthyretin (TTR) and thyroxine-binding globulin (TBG), both of which bind THs. Additionally, albumin, the most abundant protein in circulation in humans, can nonspecifically bind steroid hormones and THs with relatively low affinity. Collectively, albumin, SHBG, CBG, TTR, and TBG allow the aqueous blood matrix to transport higher total concentrations (free + bound) of these lipophilic hormones than would be possible through dissolution alone (free only). The majority of steroid hormones and THs in circulation are bound, but only the free fractions are bioavailable to target tissues (i.e., able to diffuse from blood to target tissues). Circulating hormone-binding proteins function to regulate the bioavailable fractions of steroid hormones and THs and thereby regulate hormone signaling in target tissues. All of these proteins are secreted by the liver, providing one mechanism through which the liver indirectly regulates endocrine hormone homeostasis physiology [32,33]. Binding proteins also integral to circulatory transport of and signaling of insulin-like growth factors (IGFs), which are effectors of growth hormone action in the liver.

1.3.2: Peripheral hormone metabolism and clearance

Hormones in circulation are metabolized by peripheral tissues for three purposes: activation, inactivation, and clearance. Some hormones are activated through peripheral metabolic conversion into more potent metabolites, such as T, which is metabolized to DHT, a more potent androgen in some tissues. Similarly, peripheral tissues can metabolically inactivate hormones and thereby downregulate hormone signaling, such as the conversion of cortisol to cortisone, an inactive GC. Finally, hormone metabolism is necessary for hormone clearance/excretion. While many peripheral tissues can remove hormones from circulation, the liver is the primary site of hormone clearance. Clearance/excretion mechanisms differ by hormone class/structure. Peptide hormones are degraded via proteolysis, sometimes by hormone-specific enzymes, such as insulin degrading enzyme, primarily in the liver and kidneys [34–39]. Steroid hormones and THs undergo phase I/II biotransformation, performed by a suite of enzymes expressed in the liver and other peripheral tissues. Phase I biotransformation inactivates the active hormone while phase II biotransformation conjugates the inactivated hormone to polar moieties, thereby increasing water solubility and facilitating excretion via urine and bile [1,40].

2: Vertebrate endocrine physiology: Organs, axes, and hormones

Considering the large number of hormones involved in vertebrate physiology and the diverse functional roles (Table 2) and biochemical characteristics of these hormones, it is unsurprising that the systems and mechanisms regulating hormone synthesis and release are complex. Despite this complexity, however, there are generalized themes in how endocrine organs/tissues and hormones are regulated. Chief among these themes is the close integration of the nervous and endocrine systems. The brain plays a central role in regulating endocrine physiology, integrating a wide variety of internal and external stimuli and transducing this information into signals that influence the function of diverse endocrine tissues. The hypothalamus, through its function as an endocrine gland, plays a major role in regulating other endocrine tissues, which raises another unifying theme of vertebrate endocrine physiology: the endocrine axis framework. An endocrine axis represents a hierarchical subdivision of the endocrine system. These axes are composed of a central regulator (the hypothalamus), an intermediate endocrine gland (the anterior pituitary gland), and peripheral endocrine glands (gonads, adrenal glands, thyroid gland, or liver). Within any given axis, the general flow pattern of information is conserved: hypothalamic hormones (releasing hormones) modulate the secretion of pituitary hormones (trophic hormones), which in turn regulate peripheral glands. Finally, hormones secreted by peripheral glands exert both positive and negative feedback on the hypothalamus and/or anterior pituitary to regulate overall axis function. The hormones of the anterior pituitary (more specifically, those involved in an endocrine axis) are generally named based on the peripheral gland on which they exert trophic action, such as gonadotropins and thyrotropin (thyroid-stimulating hormone; TSH). Upstream hypothalamic hormones are then named for the trophic hormone they regulate, such as gonadotropin-releasing hormone (GnRH) and thyrotropin-releasing hormone (TRH).

Table 2

In Sections 2.1–2.4, we briefly review the major vertebrate endocrine axes: the hypothalamic-pituitary-gonadal (HPG) axis (Section 2.1), the HP-adrenal (HPA) axis (The hypothalamic-pituitary-adrenal (HPA) axis section), the HP-thyroid (HPT) axis (Section 2.3), and the HP-somatotropic (HPS) axis (Section 2.4). There are other multifaceted physiological systems that regulate some hormones, such as the renin-angiotensin-aldosterone system (RAAS) (The renin-angiotensin-aldosterone system (RAAS) section), and there is substantial cross-talk among the endocrine axes and throughout the endocrine system in general, which we mention only briefly throughout Section 2. In Section 2.5, we discuss metabolic and nutritional endocrinology. In Section 2.6, we briefly mention other hormones that are involved in vertebrate physiology but otherwise not mentioned elsewhere in this chapter.

2.1: Hypothalamic-pituitary-gonadal (HPG) axis

Postnatal reproductive maturation and gamete production in adulthood are the product of highly coordinated regulatory pathways that initiate in the brain and ultimately end in the gonads. Chief regulatory centers and signaling components of this axis include the hypothalamus, where neuroendocrine regulatory control is initiated via the production of GnRH; the anterior pituitary, where hypothalamic GnRH dictates the production and secretion of gonadotropin hormone intermediaries, follicle-stimulating hormone (FSH) and luteinizing hormone (LH); and the gonads, where gonadotropins induce steroidogenesis and gametogenesis. Finally, gonadal hormones, especially estrogens, androgens, and progestogens, secreted into circulation, act upstream on both the hypothalamus and pituitary to establish both positive and negative feedback mechanisms and to regulate overall axis function while also exerting control over physiological and behavioral endpoints through effects on various peripheral tissues. A variety of other internal and external factors influence HPG axis regulation, mediated through neuronal and hormonal signaling. In seasonally breeding organisms, photoperiod, temperature, or other environmental cues are involved in stimulating GnRH secretion. Similarly, in induced ovulators (e.g., cats), stimuli associated with copulation influence HPG function. Energy balance also influences GnRH secretion and HPG/reproductive physiology. For example, energy balance and body condition are determinants of pubertal timing in girls, and this effect is seemingly mediated through leptin. The link between energy balance and reproduction/HPG axis physiology is also mediated through insulin and insulin-like growth factor 1 (IGF1). Melatonin regulates GnRH mRNA levels, providing a molecular mechanism linking photoperiod and reproduction in seasonal breeders. Other hormones, such as THs and GCs, also influence the HPG axis and reproduction.

2.1.1: The hypothalamus and GnRH

The initiating event of the HPG axis is the synthesis and release of GnRH, a decapeptide protein hormone, from GnRH neurons within the hypothalamus. Although the exact location and number of GnRH neurons vary by species, they are a relatively small population of neurons, composed of 1000 to 1500 nuclei in humans, that are spread throughout preoptic and mediobasal regions of the hypothalamus. Production of GnRH from these cells is complex and exhibits species specificities; however, its secretion is likely the result of joint activity of multiple regulatory factors, including catecholamines (NE, epinephrine, serotonin, and dopamine) and chiefly the neuropeptide, kisspeptin. The GnRH gene itself encodes a large 92 amino acid polypeptide that must be modified posttranscriptionally to produce the mature signaling peptide GnRH, which is secreted into the hypophyseal portal system to reach the anterior pituitary. The structure of the mature peptide is conserved across vertebrates [41]. Numerous additional forms of GnRH also occur that differ in location of expression and function. This includes a second GnRH gene, GnRH-II (encoding GnRH-II), but its expression occurs outside the hypothalamus and its primary function is not the stimulation of pituitary gonadotropin secretion [42]. Rather, mature GnRH-I (hereafter GnRH) is the major vertebrate form of GnRH regulating HPG axis function.

A critical facet of the regulation of GnRH expression/secretion is that the production occurs in a pulsatile manner in spontaneously ovulating species. Pulsatile GnRH secretion is responsible for establishing distinct patterns of gonadotropin release from the pituitary and ultimately generates patterns of reproductive cyclicity. Although the entire picture of regulatory mechanisms controlling pulsatile GnRH release, or the GnRH pulse generator remains to be elucidated, kisspeptin and steroid hormones (estradiol and progesterone) are involved, as are additional peptide hormones including neurokinin B and gonadotropin inhibitory hormone (GnIH) [43–45].

Within the hypothalamus, decapeptide GnRH travels from cell bodies in the mediobasal and preoptic areas to the median eminence where it is secreted into the hypophyseal portal system. This system is composed of a network of capillary beds, which branch from the superior hypophyseal artery, in the median eminence region. They drain into sinusoids that constitute the major blood supply to the anterior pituitary. Once in the pituitary, GnRH binds to its cognate receptor, GnRH-receptor or GnRHR. This stimulates intracellular signaling cascades in a population of cells termed gonadotropes, which are responsible for the synthesis and secretion of gonadotropins and express GnRHR on their cell membranes. GnRHR is a G-protein coupled receptor (GPCR) and while three types of GnRHR have been identified across vertebrates, GnRHR-I is the predominant receptor on gonadotropes [46].

2.1.2: The anterior pituitary and gonadotropins

The major functional outcome of GnRH association with GnRHR-I on pituitary gonadotropes is the synthesis and secretion of FSH and LH. These hormones share a common α subunit, encoded by the CGA gene, and have unique ꞵ subunits, encoded by LHB and FSHB genes, respectively. Transcriptional control of these genes is dynamic and varies with GnRH levels. Notably, the α subunit is also shared with TSH and mammalian placental glycoprotein hormones (e.g., human chorionic gonadotropin, [hCG]).

The full regulatory dynamics of gonadotropins that follow from GnRH signaling will not be examined herein; however, it is important to note that patterns of GnRH synthesis/secretion in the hypothalamus drive distinct expression and secretion patterns of LH and FSH. GnRH secretion is pulsatile, and both the frequency and magnitude of pulses vary across reproductive cycles and between sexes. In order to drive reproductive function in the gonads, FSH and LH must be exocytosed from pituitary gonadotrope cells; this process occurs under distinct regulatory paradigms in the context of GnRH stimulation. FSH release is generally constitutive, occurring in tight association with its synthesis, and both synthesis and secretion occur under low pulse frequencies of GnRH. However, GnRH regulates FSH synthesis, whereas other factors control secretion including the protein hormones activin, inhibin, and follistatin. Activin promotes secretion, whereas inhibin and follistatin inhibit the stimulatory effects of activin, blocking interaction of activin with its receptor [47]. In contrast, LH secretion is more pulsatile than FSH and occurs predominantly at higher GnRH pulse frequencies. Finally, GnRH signaling also plays a more direct role in stimulating LH secretion from gonadotropes than FSH.

2.1.3: The gonads, estrogens, androgens, and progestogens

Both FSH and LH are secreted directly into circulation and travel through plasma to the gonads. FSH and LH work in concert to promote steroidogenesis and gamete maturation but occupy distinct functional niches in these processes. In the ovary, FSH primarily regulates follicle maturation and growth (folliculogenesis), follicle selection for ovulation, and estrogen biosynthesis. The bulk of these actions result from interaction of FSH with its cognate receptor, FSHR, a GPCR expressed on the surface of granulosa cells surrounding individual oocytes. Activation of FSHR on granulosa cells triggers PKA/cAMP signaling to drive expression of the steroidogenic enzyme aromatase (CYP19A1). Aromatase subsequently is responsible for synthesis of E2 via conversion from theca-derived androgens (discussed below). FSHR activation also promotes expression of cell-cycle progression genes, including cyclins, through ERK signaling to drive granulosa proliferation [48]. Almost paradoxically, FSH signaling also activates apoptotic pathways, depending on the level of FSHR expression and activation. The balancing act of FSH-mediated proliferative and apoptotic signals is critical to the selective growth of dominant follicles for ovulation and culling, or atresia, of subordinate follicles.

Similar to FSH, LH signaling also regulates steroidogenesis and cell proliferation in the ovary, albeit in distinct fashion from FSH. Like FSH, LH action first requires binding to its cognate receptor, LHCGR. LHCGR is a cell-surface GPCR, and its activation stimulates cAMP/PKA and ERK1/2 signaling to regulate transcription of LH target genes. This includes inducing expression of CYP17A1, responsible for the biosynthesis of androgens. Androgens derived from thecal cells diffuse into adjacent granulosa cells, forming the substrate for FSH-induced E2 synthesis. LH also stimulates the expression of CYP11A1, leading to production of progesterone in both theca and granulosa cells after ovulation. In addition to its stimulatory effects on steroidogenesis, LH signaling also regulates cellular growth and differentiation in the ovary. These effects include inducing terminal differentiation of granulosa cells shortly before ovulation, a process termed luteinization, and stimulates resumption of meiosis in the oocyte [49].

In the testis, FSH acts predominantly on Sertoli cells; however, its functions in this cell population do not include directly stimulating steroidogenesis, as occurs in the ovary. Instead, FSH works independently and in concert with androgens to stimulate Sertoli proliferation, critical to both testicular development during puberty and to spermatogenesis in adulthood. Proliferative effects in the testes similarly involve ERK signaling and downstream cyclin activation; however, these responses are dynamic throughout reproductive development and do not occur in mature Sertoli cells. FSH also stimulates Sertoli cell maturation and function related to germ cell support and development, ultimately stimulating progression of spermatogenesis [50]. In contrast to FSH, LH plays a similar role in the testis as the ovary and is primarily responsible for stimulating androgen production in Leydig cells. LH binding to LHCGR in Leydig cells triggers cAMP-mediated PKA activation, ultimately regulating expression of genes controlling translocation of cholesterol into mitochondria and subsequent conversion to T and intermediates. T in turn positively regulates spermatogenesis within the testes through its actions on Sertoli cells and further regulates numerous physiological processes throughout the body [51].

Gonadal steroid hormones produced in response to gonadotropins play an immediate role in modulating GnRH release from the hypothalamus and gonadotropin production in the pituitary. Estrogen action is bimodal in the hypothalamus, and results from its actions on kisspeptin neurons. At high levels, estrogen enhances GnRH production whereas it suppresses production at low levels. This dual regulatory control over GnRH contributes to the modulation of gonadotropin secretion, particularly suppression of LH, throughout the estrous cycle and ultimately drives a surge of LH that is necessary for ovulation. Estrogen also acts directly on pituitary gonadotrophs to modulate gonadotropin secretion, contributing to both positive and negative feedback [52]. Finally, estrogens are also necessary within the ovary to promote follicle growth and maturation [53]. Similar to estrogens, androgens function across the HPG axis to regulate its function, acting to negatively regulate GnRH production in the hypothalamus and gonadotropin action and spermatogenesis in the testes. Direct androgen action within the pituitary is less clear; however, lines of evidence suggest androgens can both augment and inhibit gonadotropin production—likely dependent on the species in question [54]. Finally, progestogens are an important factor produced following gonadotropin action, particularly in the ovary. Although not discussed in great detail herein, gonadal progestogens play a critical role in regulating the HPG axis and occupy a dual-natured role similar to estrogen. By negatively regulating GnRH secretion, they dictate gonadotropin production and inhibit follicle maturation. However, progesterone is also needed for positive GnRH feedback, as loss of progesterone receptors in the hypothalamus disrupts the LH surge.

Lastly, the steroid hormones produced in the testes and ovaries have been described as a male/female binary system, similar to traditional representations of androgens as male hormones and estrogens as female. It is important to note, however, that androgens are integral to ovarian function and female HPG axis function, as are estrogens to testis function and the male HPG axis. Further, these hormones, including progestogens, act as pleiotropic regulators, regulating the expression of diverse phenotypes across multiple tissues. As such, the net actions of the HPG axis extend beyond the bounds of gamete production and reproduction, and instead include expression of primary and secondary sex characteristics, somatic growth, behavior, maintenance of pregnancy, and modulation of immune function and energy balance. These are only a few of the numerous physiological roles of gonadal steroids.

2.2: The adrenal glands and associated hormones and systems

The adrenal glands (or interrenal glands in amphibians and teleosts) secrete four classes of hormones—GCs, mineralocorticoids, catecholamines, and androgens. In so doing, they regulate a variety of physiological and behavioral processes, especially those pertaining to metabolism, stress, immune function, reproduction, development, and osmoregulation (Table 2). Due to the diversity of hormones they secrete, the functional morphology of the adrenal glands and the upstream processes regulating their secretory function are complex. However, these regulatory processes fall broadly into three major, interrelated systems: the hypothalamic-pituitary-adrenal (HPA) axis, the renin-angiotensin-aldosterone system (RAAS), and the sympathetic nervous system.

2.2.1: Overview of adrenal hormones

Corticosteroids: Glucocorticoids and mineralocorticoids

GCs and mineralocorticoids are collectively referred to as corticosteroids. GCs are named for their role regulating glucose physiology, but their functions are diverse. GCs primarily act by binding the GC receptor (GR) and the mineralocorticoid receptor (MR), but they also mediate nongenomic actions via membrane-bound receptors [55,56]. GCs are often called stress hormones due to their role in regulating aspects of physiological/behavioral stress responses. Acute exposure to stress stimuli induces GC secretion, which act canonically to promote survival (e.g., mobilization of energy stores for increased energetic demands). As such, GCs are frequently used as biomarkers of stress in vertebrates, but this designation as stress hormones oversimplifies both the complexity of stress endocrinology, the diverse roles of GCs beyond stress, and other non-GC hormones involved in stress responses (Table 2). GCs are also secreted in the absence of acute stressors and have physiological roles regulating metabolic, immunological, reproductive, and developmental processes, among others [57,58]. Because they bind the MR, GCs also regulate processes associated with mineralocorticoid signaling, such as osmo- and ionoregulation [55]. GCs are used therapeutically as immunosuppressive/antiinflammatory drugs in human and animal medicine [56].

Aldosterone is the major mineralocorticoid in most vertebrates. However, the MR binds aldosterone and GCs with similar affinity, and because GCs circulate at much basal higher levels than aldosterone, most MR are occupied by GCs rather than aldosterone [59]. The apparent specificity of the MR for aldosterone in some tissues is likely mediated by the enzymatic conversion of active GCs to inactive GC metabolites in those tissues, thereby reducing GC-binding to the MR in a tissue-specific manner [55,59]. The MR is primarily involved in osmoregulation and ionoregulation, acting in the kidney, colon, and salivary gland to regulate fluid and ion transport, as well as tissue repair and inflammation [55,60]. It is widely expressed in other tissues, such as the brain (particularly the hippocampus), heart, and vasculature, and MR activation and/or aldosterone concentrations have been implicated in the pathophysiology of cardiovascular disease, kidney disease, metabolic syndrome, and depression [55,59].

Catecholamines

Epinephrine, NE, and dopamine are the catecholamines secreted by the adrenal gland. NE and dopamine are also secreted systemically by postganglionic sympathetic neurons and, for dopamine exclusively, nonneuronal sources. Epinephrine and NE are rapidly secreted following exposure to acute stressors and induce physiological and behavioral changes associated with the fight-or-flight response (e.g., increased heart rate, blood pressure, bronchodilation, gluconeogenesis, lipolysis, etc.) to promote survival (e.g., increasing delivery of oxygen and glucose to skeletal muscles, heart, and brain). Therefore, catecholamines can also be classified as stress hormones along with GCs. Epinephrine and NE mediate their effects on target tissues via adrenergic receptors, while dopamine functions through dopamine receptors, both of which are families of GPCRs expressed in a wide variety of tissues. In addition to functioning as hormones, catecholamines are important neurotransmitters. Drugs targeting catecholamine receptors (agonists and antagonists) are used clinically to treat some cardiovascular conditions (e.g., hypertension, failure, arrhythmia, and angina), anaphylaxis, glaucoma, and others [61].

Adrenal androgens

Adrenal androgens include DHEA, DHEA-sulfate (DHEAS), and A4, among others. In general, adrenal androgens are weakly androgenic, activating the AR with much lower potency than T or DHT. Instead, adrenal androgens primarily serve as precursors for the production of more potent androgens and estrogens [62]. Adrenal androgens do, however, affect cellular physiology through mechanisms independent of bioconversion to other androgens or estrogen, and can directly regulate physiological processes (e.g., DHEA has been implicated in modulating endothelial and immune function) [63]. DHEAS in particular is the major product of the adrenal gland and exists in circulation at concentrations orders of magnitude higher than any other steroid [64]. It also induces the onset of pubic hair growth, or pubarche, during puberty. This occurs due to increased DHEAS secretion during the peripubertal period, a process called adrenarche. Adrenal androgens may be involved in the hyperandrogenism commonly observed in women with polycystic ovarian disease (PCOS) [62]. Finally, age-related declines in DHEA may contribute to the pathogenesis of cardiovascular disease [63].

2.2.2: Adrenal gland morphology

The sites of hormone synthesis within the adrenal gland vary by hormone class, as do the upstream systems regulating these processes (Section 2.2.3). The adrenal gland is anatomically divided into the medulla and the cortex. Catecholamines are secreted by the medulla, while adrenal steroids are secreted by the cortex, which is further subdivided into three zones. These zones, defined by distinct cellular morphology, are called the zona reticularis, zona fasciculata, and zona glomerulosa. This zonation is often discussed in terms of function: the zona reticularis is generally considered the site of androgen production; the zona fasciculata is considered the site of GC production; and the zona glomerulosa is considered the site of mineralocorticoid production. However, biosynthetic functions only partially correspond to these zonal delineations, and steroid biosynthesis may require interactions among the zones. In rats, for example, aldosterone synthase is expressed in the zona glomerulosa, but the full suite of enzymes required for de novo steroidogenesis is not, suggesting that aldosterone precursors are transported to the zona glomerulosa from the zona fasciculata, where de novo steroidogenesis occurs. Zonation of the adrenal cortex is primarily induced by the canonical upstream regulators of GC and mineralocorticoid secretion, specifically adrenocorticotropic hormone (ACTH) and angiotensin II [65].

2.2.3: Regulation of adrenal hormone secretion

The secretory activity of the adrenal medulla and cortex is differentially regulated. In the adrenal medulla, chromaffin cells, which secrete catecholamines, are directly innervated and activated by sympathetic stimuli [66]. Alternatively, the adrenal cortex is regulated as part of the HPA axis and the RAAS. The HPA axis (The hypothalamic-pituitary-adrenal (HPA) axis section) exerts control over all three classes of adrenal steroids, though the extent of its influence on androgens and mineralocorticoids is limited relative to its control over GCs [65]. Mineralocorticoid secretion is also regulated by the RAAS (The renin-angiotensin-aldosterone system (RAAS) section) and stimulated by extracellular potassium [60]. The additional factors regulating adrenal androgens are not yet fully resolved [62,65]. Some evidence suggests that there is a specific adrenal androgen-stimulating hormone, but studies to date have yet to conclusively identify any such hormone. Others suggest that intra-adrenal steroidal paracrine signaling may be involved [62].

The hypothalamic-pituitary-adrenal (HPA) axis

The HPA axis primarily functions to regulate GC secretion and, to a lesser extent, mineralocorticoid and adrenal androgen secretion. The hypothalamus secretes corticotropin-releasing hormone (CRH) in response to internal and external stimuli, such as stress. CRH is synthesized in the neurons of the paraventricular nucleus (PVN) of the hypothalamus, though other regions of the brain also produce CRH at a lower level. CRH is transported to the anterior pituitary via the hypophyseal portal system where it activates its receptors (CRF-R1 and CRF-R2) expressed by corticotrophic cells. CRF-R activation in the anterior pituitary upregulates the expression of pro-opiomelanocortin (POMC), the precursor to ACTH (and several other hormones), and ultimately stimulates the secretion of ACTH into peripheral circulation [66]. Within the adrenal cortex, ACTH binds and activates the melanocortin type 2 receptor (MC2R), inducing de novo steroidogenesis and the secretion of GCs, mineralocorticoids, and adrenal androgens [67,68]. Both POMC and pro-ACTH, an intermediate cleavage product, are also observed in circulation and may act to inhibit ACTH function [69]. Additionally, within the anterior pituitary, CRH activity is modulated by CRH-binding protein (CRH-BP), which downregulates CRH action. GCs in circulation exert negative feedback on both the hypothalamus and the anterior pituitary, suppressing the secretion of CRH and ACTH; since GCs can activate both the GR and the MR, both receptor types are involved in transducing GC-mediated negative feedback signals. Adrenal androgens do not exert negative feedback on the HPA axis. In addition to CRH and the negative feedback mediated by GCs, vasopressin and oxytocin regulate ACTH secretion from the anterior pituitary [66].

The renin-angiotensin-aldosterone system (RAAS)

The RAAS is a physiological system that regulates aldosterone and angiotensin II (AngII), two major regulators of electrolyte and fluid homeostasis. AngII formation results from coordination between the liver, kidneys, and vascular endothelial cells involving several intermediate enzymatic transformations. Renin, an enzyme secreted into circulation by the kidneys, acts on liver-derived angiotensinogen, to produce angiotensin I (AngI). While angiotensinogen secretion is constitutive, renin secretion is stimulated by low blood pressure and volume, low sodium concentrations, and sympathetic nervous system stimulation. AngI is further metabolized into AngII by angiotensin-converting enzyme (ACE) expressed by endothelial cells. The effect of AngII signaling depends on which receptor type it binds. The angiotensin type-1 receptor (AT1R) is the primary mediator of the physiological effects and health outcomes associated with AngII activity. AngII activation AT1R stimulates elevated sodium and fluid retention and intake (i.e., salt appetite and thirst) as well as vasoconstriction and sympathetic nervous system activity. AT1R activation also induces aldosterone secretion from the adrenal, which reinforces the physiological effects associated with AT1R activation, as aldosterone similarly stimulates sodium and fluid retention via the MR. Therefore, excessive AngII and aldosterone can produce a suite of harmful effects, particularly in the cardiovascular and renal systems, such as hypertension, inflammation, reactive oxygen species (ROS) generation, and fibrosis. The RAAS has been linked to cardiovascular, renal, and metabolic diseases, among others, and is a common therapeutic target in clinical medicine (e.g., ACE inhibitors, AT1R inhibitors, and MR antagonists) [60,70].

The production of AngI and AngII likely occurs close to or within the final target tissue, and both AngI and AngII have short half-lives. For this reason, some have argued that AngII should not be considered a circulating hormone, while renin should be as it circulates systemically [71]. While renin and its precursor, prorenin, can act as hormones, binding the prorenin receptor ((P)RR), renin stimulates downstream angiotensin and aldosterone signaling via its enzymatic activity. Therefore, in its canonical role within the RAAS, renin cannot be considered a hormone because it does not act via a receptor. (P)RR activation is also directly involved in regulating fluid/sodium physiology as well as other physiological processes [60,72].

2.3: The hypothalamic-pituitary-thyroid (HPT) axis

THs are critical to numerous developmental and metabolic processes in vertebrates (Table 2). Some of the most notable regulatory roles of THs include thermogenesis in endothermic animals; protein, lipid, and carbohydrate metabolism (both catabolism and anabolism); somatic growth and development (e.g., tissue differentiation); hatching in birds and reptiles; and metamorphosis in amphibians and teleosts. In fact, disruption of metamorphosis by exogenous chemicals is regarded as a hallmark of TH-disrupting chemicals, hence the use of the amphibian metamorphosis assay as a screening tool for thyroid-disrupting activity. Thyroid gland secretory function is regulated within the hypothalamic-pituitary-thyroid (HPT) axis, functioning in an analogous manner to the HPG and HPA axes, with the hypothalamus acting as a central regulator of via secretion of thyrotropin-releasing hormone (TRH). TRH then stimulates secretion of TSH into circulation, which in turn acts on the thyroid gland to stimulate release of THs [33].

2.3.1: The hypothalamus and thyrotropin-releasing hormone (TRH)

The most central hormonal regulator of HPT axis physiology is TRH, a tripeptide hormone synthesized and secreted by the PVN. The release of TRH from the hypothalamus initiates the signaling cascade that ultimately results in the secretion of THs from the thyroid gland. TRH expression is primarily regulated by THs via negative feedback, with TRH neurons expressing THRs and the TRH gene possessing TH response elements in its promoter region. Additional factors conveying metabolic and nutritional information also regulate TRH expression and secretion. Leptin and neuropeptide y (NPY) are particularly important, regulating TRH neurons directly (i.e., TRH neurons express leptin and NPY receptors). Neuronal projections to TRH neurons in the PVN originate from other regions of the hypothalamus (the arcuate nucleus [ARC] and the dorsomedial nucleus [DMN]) and the medulla oblongata). Those neurons originating in the ARC and DMN are also responsive to leptin, meaning leptin regulates TRH production through direct and indirect mechanisms. Upon secretion, TRH is transported to the anterior pituitary gland via the hypophyseal portal system