Serotonergic Control of Gastrointestinal Development, Motility, and Inflammation

Abstract

Although it is most well-known for its roles in central nervous system (CNS) function, the vast majority of serotonin, or 5-hydroxytryptamine (5-HT), is produced in the gastrointestinal (GI) tract. 5-HT is synthesized mostly by enterochromaffin (EC) cells of the GI epithelium and, in small part, by neurons of the enteric nervous system (ENS). The GI tract contains an array of broadly distributed 5-HT receptors, which participate in functions such as motility, sensation, inflammation, and neurogenesis. The roles of 5-HT in these functions are reviewed, as well as its role in the pathophysiology of disorders of gut-brain interaction (DGBIs) and inflammatory bowel diseases (IBD).

Introduction

Although it is most well-known for its roles in central nervous system (CNS) function, serotonin, or 5-hydroxytryptamine (5-HT), was first discovered in the gastrointestinal (GI) tract. Vittorio Erspamer, the pharmacologist who discovered 5-HT, first named it “enteramine” after extracting it from rabbit gastric mucosa in 1937 (67), and later noted that it was present in the GI tract of every vertebrate animal he studied. Ten years later, Maurice Rapport, Arda Green, and Irvine Page identified this molecule as serotonin, named for its origins in serum (“sero”) and its ability to increase tone or vasoconstriction (“tonin”) (189, 190), and published its structure as 5-HT in 1949 (188). This research merged in 1952, when Erspamer confirmed that the enteramine molecule he had discovered in the mucosa of the GI tract was 5-HT (68). 5-HT was then found in the CNS and discovered to have roles in essential functions such as sleep, mood, and appetite (33). Although 5-HT has many important roles in the brain, the vast majority of the body’s 5-HT is produced in the gut, where it participates in functions such as motility, sensation, inflammation, and enteric neurogenesis (28, 74, 92, 158). The roles of 5-HT in these functions are reviewed, as well as its role in the pathophysiology of disorders of gut-brain interaction (DGBIs) and inflammatory bowel diseases (IBD).

5-HT Synthesis and Signaling

The gut contains the vast majority of the body’s 5-HT, synthesized mostly by EC cells of the mucosa and, in small part, by neurons of the enteric nervous system (ENS) (92, 119, 197). EC cells are a subtype of enteroendocrine cell that constitute less than 1% of the GI epithelium yet produce the vast majority of gut 5-HT (92). Serotonergic neurons make up only 2% to 3% of total ENS neurons but project a vast number of fibers throughout the myenteric plexus to elicit broad effects (52) (Figure 1).

Figure 1. Expression and distribution of serotonergic cells and axonal fibers in the colonic mucosa and the enteric nervous system (ENS).

Serotonin (5-hydroxytryptamine, 5-HT)-positive cells (green) are present in both the enterochromaffin (EC) cells of the mucosa (A) and within myenteric plexus of the ENS (B). In the mucosa, 5-HT is expressed in EC cells (starred in inset) which make up less than 1% of gut epithelial cells (stained with nuclei marker, bisbenzimide, magenta). In the myenteric plexus, 5-HT+ neurons (arrow, inset) comprise 2% to 3% of total myenteric neurons (stained with pan-neuronal marker, ANNA-1, magenta) as observed in wholemount laminar preparation of the myenteric plexus (B) and a cross-section of the colon (arrow). In the ENS, 5-HT is also present in continuous presynaptic fibers that run across the enteric ganglia and along interganglionic fiber tracts, thus providing extensive innervation to other neurons within the ganglion. Made in © BioRender - biorender.com.

EC cells that synthesize 5-HT contain tryptophan hydroxylase 1 (TPH1), the rate-limiting enzyme that converts the precursor amino acid l-tryptophan into 5-hydroxytryptophan (5-HTP), which is then converted by a nonspecific aromatic l-amino acid decarboxylase to 5-HT (95, 228). 5-HT is stored in vesicular monoamine transporter 1 (VMAT1) within EC cells (163). Enteric neurons synthesize 5-HT in a similar fashion, but with tryptophan hydroxylase 2 (TPH2) as the rate-limiting enzyme and, within enteric neurons, 5-HT is stored in VMAT2 (193) (Figure 2). TPH2 is also present in serotonergic neurons of the CNS, primarily in the dorsal raphe nucleus (229, 241). A global loss of TPH1, in TPH1 knockout (KO) mice, results in complete depletion of intestinal epithelial 5-HT synthesis without any loss in the brain or the ENS, while knockout of TPH2 results in a loss of both brain and enteric neuronal 5-HT biosynthesis (228). Interestingly, loss of TPH1 does not affect total gastrointestinal transit time but does result in increased stool size and changes in colonic migrator motor complex patterns (107, 153). In contrast, although TPH2 synthesizes a much smaller amount of 5-HT in the gut, its deletion has profound effects on in vivo and ex vivo gut motility and enteric neurogenesis (140), which is explained in greater detail below. These data indicate differing roles of TPH1 and TPH2 in gut function and ENS development as well as differences in how epithelial versus neuronal 5-HT impact motility.

Figure 2. Serotonin (5-HT) biosynthesis and metabolism in the gastrointestinal tract.

Biosynthesis of 5-HT in the gut is accomplished by two different isoforms of the rate-limiting enzyme, tryptophan hydroxylase (TPH). TPH1 is responsible for 5-HT synthesis in the mucosal enterochromaffin cells and TPH2 synthesizes 5-HT in enteric neurons. Both enzymes convert the amino acid l-tryptophan (l-Tryp) to 5-hydroxytryptophan (5-HTP). The enzyme l-amino acid decarboxylase (l-AAD) then converts 5-HTP to 5-HT where it is then released into the extracellular space (solid arrows) to act on 5-HT receptors and initiate downstream serotonergic signaling. Extracellular 5-HT is then taken back up into EC cells or enteric neurons via the 5-HT transporter, SERT (dashed arrows). Once intracellular, 5-HT is inactivated by monoamine oxidase (MAO), in the mitochondria, to 5-hydroxyindoleacetic acid (5-HIAA). Made in © BioRender - biorender.com.

After 5-HT is released and exerts effects on its receptors, it must be rapidly inactivated, as excess extracellular 5-HT can cause toxicity and/or receptor desensitization (46, 94). The enzyme, monoamine oxidase (MAO) is required for inactivation of 5-HT (181) but is located intracellularly. Outside of the cell, a high-affinity transporter is thus required to take up 5-HT because it is highly charged at a physiological pH (142). The plasmalemmal serotonin reuptake transporter (SERT) is the primary mechanism by which 5-HT is transported intracellularly. Once inside the cell, 5-HT is deaminated to 5-hydroxyindole acetaldehyde (5-HIAL), then oxidated by MAO to 5-hydroxyindoloacetic acid (5-HIAA) which is excreted by the kidneys (137). SERT is ubiquitously expressed in enterocytes that line the mucosa (98, 226), in some ENS neurons (92, 95), and in immune cells such as macrophages, mast cells, lymphocytes, and dendritic cells (108). Platelets circulating throughout the intestine also express SERT and thus contribute to 5-HT uptake in the gut (15, 138). Through blood circulation, platelets thus act as a vessel for endocrine signaling of 5-HT to targets such as liver (139) and bone (132). Platelets do not synthesize 5-HT because they do not contain TPH. The concentration of 5-HT in the blood is therefore likely an indicator of peripheral gut 5-HT production and SERT efficacy in platelets (77, 165).

5-HT Receptors in the GI Tract

At least 15 different 5-HT receptors have been characterized. Five of these receptor subclasses are present in the intestine: 5-HT₁, 5-HT₂, 5-HT₃, 5-HT₄, and 5-HT₇ (118). These subtypes are all G-protein-coupled receptors except for 5-HT₃, which is ionotropic (118, 157). These receptors are distributed in different cell types throughout the digestive tract, including in intrinsic neurons of the ENS and extrinsic primary afferent neurons (ExPANs) that project to the CNS (89). The roles of these receptors are detailed in the below sections and a summary of 5-HT receptor expression and function can be found in Table 1. Little is currently known about the whether multiple types of 5-HT receptors are present in the same cell; thus, future studies should examine co-expression of 5-HT receptors in ENS neurons and other cell types of the gut.

Table 1.

5-HT Receptor Expression and Function

Receptor	Expression	Function	References
5-HT1A	Enteric neurons	Inhibits neurons involved in peristalsis	(83, 85, 87)
5-HT1P	Enteric neurons	Activates neurons involved in peristalsis	(4, 31, 51, 103, 231)
5-HT2B	Enteric neurons	Activates neurons involved in peristalsis	(30)
		Promotes neuronal development	(76)
	Interstitial cells of Cajal	Promotes ICC proliferation	(216, 236)
	Extrinsic primary afferent neurons	Initiates pain signaling	(112, 178)
5-HT3	Enteric neurons	Activates neurons involved in peristalsis	(110, 111)
	Smooth muscle cells	Evokes muscle contraction	(158, 204)
	Extrinsic primary afferent neurons	Initiates pain signaling	(158, 204)
	Vagal afferent neurons	Evokes nausea and emesis	(168, 222)
	Enterochromaffin cells	Evokes 5-HT release	(158, 204)
	Interstitial cells of Cajal	Unknown	(235)
	Enterocytes	Facilitates secretion	(22)
5-HT4	Enteric neurons	Present on presynaptic nerve terminals, activation evokes release of acetylcholine to initiate peristaltic contractions	(86, 218)
	Smooth muscle cells	Evokes muscle contraction	(215)
	Extrinsic primary afferent neurons	Modulates pain signaling	(198)
	Enterocytes	Facilitates chloride secretion	(114)
	Enterochromaffin cells	Evokes 5-HT release	(114)
	Goblet cells	Evokes mucus discharge	(114)
5-HT7	Enteric neurons	Mediates slow depolarizing responses in intrinsic primary afferent neurons	(59, 162, 219)
	Smooth muscle cells	Inhibits smooth muscle cells, resulting in relaxation of smooth muscle	(44, 121, 184, 219)

5-HT₁ receptors

5-HT₁ receptors (specifically 5-HT_1A and 5-HT_1P) are G-protein-coupled receptors localized to enteric neurons that exert inhibitory or excitatory actions, depending on the subtype. General 5-HT₁ receptor activation with nonspecific agonists results in membrane hyperpolarization of postsynaptic neurons, thus inhibiting neurotransmitter release (83, 85, 87). Specific activation of the 5-HT_1A receptor inhibits neurotransmitter release onto enteric motor neurons and disrupts motility, by diminishing colonic migrating motor complexes (CMMCs) (59). In contrast, the 5-HT_1P receptor exerts excitatory effects; its activation induces neurotransmitter release from intrinsic primary afferent neurons (IPANs), which initiate peristaltic reflexes in the gut (31, 103, 231). 5-HT_1P receptors on IPANs also mediate chloride secretion in the gut (51). A recent study demonstrated the presence of functional 5-HT_1P receptors in the human small and large intestine; application of a 5-HT_1P agonist resulted in long-lasting activation of human submucosal ENS neurons (4).

5-HT₂ receptors

The 5-HT_2B receptor is a G-protein-coupled receptor widely expressed in the GI tract where it is present in enteric neurons, smooth muscle cells, and interstitial cells of Cajal (ICCs) (30, 76). Ex vivo studies have shown that in rodent and human intestine, the excitatory effects of 5-HT are mediated in part by the 5-HT_2B receptor (30). Activation of this receptor also contributes to the development of enteric neurons (76) as well as proliferation of ICCs (216, 236), which are the pacemaker cells of the gut that coordinate smooth muscle function (196). ICCs are required for normal gut motility; decreased ICC volume and/or impaired ICC functioning have been shown to contribute to dysmotility symptoms like constipation and gastroparesis (105, 239). 5-HT_2B receptors are also expressed on ExPANs that innervate the gut and convey pain and other sensory information (112).

5-HT₃ receptors

In contrast to the other 5-HT receptors present in the GI tract, which are G-protein-coupled, 5-HT₃ receptors are ionotropic, enabling ligand-gated entry of cations including Ca²⁺ (180). These excitatory receptors are present in several subtypes of enteric neurons as well as ExPANs and vagal afferent fibers, ICCs, smooth muscle cells, enterocytes, and enterochromaffin cells (158, 215). The stimulation of 5-HT₃ receptors evokes peristaltic contractions through stimulation of enteric neurons (110, 111). 5-HT₃ receptor activation on enterocytes facilitates secretion (22) and the role of 5-HT₃ receptors on ICCs is unknown, although activation results in rapid depolarization and therefore may initiate slow waves in the ICC network (235).

5-HT₃ receptors play a critical role in evoking peristaltic contractions through stimulation of enteric neurons (110, 111); IPANs, which project to the mucosa and are involved in initiating peristaltic contractions, demonstrate responses to 5-HT that are exclusively mediated via 5-HT₃ receptors (21). In isolated colons, addition of the 5-HT₃ receptor antagonist, ondansetron abolishes spontaneous colonic contractions and diminishes peristaltic contractions triggered by mucosal stimulation, indicating that EC cell-released 5-HT activates IPANs via 5-HT₃ receptors (106). These receptors also appear to be present in inhibitory motor neurons, which receive input from serotonergic interneurons and contribute to tonic inhibition of the circular muscle (59).

RNA sequencing studies have begun to show differences in 5-HT receptor expression throughout the length of the gut; it was recently demonstrated that 5-HT3 and 5-HT4 receptors are more highly expressed in the distal colon compared to proximal colon (61).

5-HT₄ receptors

5-HT₄ receptors are G-protein-coupled receptors widely expressed throughout the GI tract in enteric neurons, ExPANs, EC cells, and smooth muscle cells (215). In the ENS, 5-HT₄ receptor agonists evoke peristaltic reflexes by acting on presynaptic nerve terminals to stimulate the release of excitatory neurotransmitters such as acetylcholine (71, 86, 141, 179, 218). 5-HT₄ receptors are also expressed on colonic epithelial cells and their activation results in 5-HT release from EC cells, mucus discharge from goblet cells, and chloride secretion by enterocytes (114). 5-HT₄ receptors have important neurotrophic roles as well; expression on enteric neurons and enteric neural crest-derived precursor cells (ENCDCs) mediates neuronal proliferation and differentiation (115, 141).

5-HT₇ receptors

5-HT₇ receptors are G-protein-coupled receptors that are present in cell bodies of a subset of enteric neurons and mediate slow depolarizing responses to 5-HT (162, 219). Recent RNAseq data confirm expression of 5-HT₇ receptors in enteric neurons (240). 5-HT₇ receptors have also been identified on smooth muscle cells in guinea pig ileum (44, 219), human ileum (184), and dog stomach (121). Pharmacological studies suggest they serve an inhibitory function, causing relaxation of the smooth muscle; however, functional 5-HT₇ receptors have not been identified in smooth muscle of the mouse intestine (59).

Role of the Gut Microbiome in 5-HT Signaling

The gut microbiome comprises a dynamic population of trillions of microorganisms that reside at the epithelial interface of the gut. The microbiome has recently emerged as an important contributor to metabolism, immunity, and gut-brain signaling (149). Recent studies have demonstrated the presence of a bidirectional communication between 5-HT and gut microbiota. For the most part, EC cells secrete 5-HT basolaterally for purposes of signaling to surrounding neurons; however, some 5-HT is released on the apical side of EC cells directly into the lumen of the intestine (80). A recent study in mice demonstrated that increasing the amount of 5-HT available in the lumen increased the relative abundance of spore-forming microbes of the Clostridiaceae and Turicibacteraceae families (82). Further, a protein was detected on a particular bacterium, Turicibacter sanguinis, that is similar in structure and function to SERT, showing that bacteria can take up 5-HT and thus have a potential role in regulating 5-HT levels in the gut (82). Gut bacteria also secrete metabolites such as short-chain fatty acids (SCFAs) that have been shown to promote colonic motility in a 5-HT₃ receptor-dependent manner (81). A more recent study demonstrated that colonization of bacteria in the mouse colon promoted TPH1 mRNA levels and increased 5-HT concentration and that this occurred via SCFA interactions with EC cells (191). Using calcium imaging techniques in primary murine EC cells, researchers found that SCFAs do not acutely (i.e., within 2 h) activate EC cells to induce 5-HT secretion (155). Microbiota regulation of gut 5-HT is thus more likely the result of ongoing interactions between bacterial metabolites and EC cells. Studies have also demonstrated that other microbial-specific metabolites such as tyramine, p-aminobenzoate, and α-tocopherol also promote TPH1 expression and 5-HT release from EC cells (238). Colonic EC cells express many receptors for these metabolites such as free fatty acid receptor 2 (FFAR2), which senses SCFAs, G-protein-coupled receptor 35 (GPR35), which senses small aromatic acids, and G-protein-coupled receptor 132 (GPR132), which senses lactate and acyl amides (145). Together these data suggest that there are abundant mechanisms by which microbes influence 5-HT production.

5-HT in ENS Development

In addition to its importance in gut function, 5-HT is also recognized for its role as a growth factor in enteric neuron development. Early-born enteric neurons contain 5-HT, which has been shown to promote both neurogenesis as well as the growth of specific subsets of later-born neurons, thus influencing ENS development. For example, in studies of cultured ENCDCs, the addition of 5-HT promotes the development of dopaminergic neurons. Conversely, mice lacking neuronal 5-HT (TPH2 KO mice) display ENS hypoplasia and deficits in later-born neurons, including those expressing gamma-aminobutyric acid (GABA), nitric oxide (NO), calcitonin gene-related peptide (CGRP), and tyrosine hydroxylase (TH, a marker of dopaminergic neurons) (140). ENS hypoplasia was also found in transgenic mice deficient in neuronal 5-HT due to a single nucleotide polymorphism (R439H) in the TPH2 enzyme (TPH2-R439H mice), a defect that was rescued by administration of slow-release 5-hydroxytryptophan (5-HTP) (120). In contrast, TPH1 KO mice do not display these deficits in ENS development, suggesting that mucosal 5-HT is not critical for proper ENS development. SERT, presumably through its modulation of 5-HT inactivation, also modulates ENS development. In transgenic mice with a substitution of glycine for alanine at the SERT locus (SERT Ala56 mice), this mutation results in lower levels of circulating 5-HT, due to excess SERT activity. Consequently, these mice display a hypoplastic ENS (151). Conversely, when SERT is knocked out globally (SERT KO mice) or when mice are exposed to selective serotonin reuptake inhibitors (SSRIs) during fetal development, mice display a hyperplastic ENS (151), supporting the idea that SERT blockade in development leads to enteric neuronal hyperplasia.

The receptors 5-HT_2B and 5-HT₄ have been shown to have important roles in mediating ENS development. The 5-HT_2B receptor is expressed in subsets of enteric neurons and, in cultures of ENCDCs, stimulation of 5-HT_2B enhances neuronal differentiation (76). 5-HT may therefore influence the fate of developing enteric neurons through activation of this receptor. ICCs also express the 5-HT_2B receptor and require it for normal development of the ICC network (236), which acts as an intermediary between ENS neurons and smooth muscle to mediate slow wave propagation involved in gut motility (63). Activation of the 5-HT₄ receptor has also been shown to be neurogenic; 5-HT₄ agonism promotes neuronal survival and differentiation in vitro and neurogenesis in adult mice in vivo (141). The presence of stem cells in the mature gut makes this neurogenesis possible (27, 29, 134). Mice in which 5-HT₄ receptors are knocked out (5-HT₄ KO) show deficiencies in postnatal enteric neurogenesis that are exaggerated with age (141). 5-HT₄ signaling also promotes postnatal enteric neural stem cell (ENSC) proliferation and neuronal differentiation. In colon explants cultured with ENSCs, addition of liposomal nanoparticles containing a 5-HT₄ agonist, RS67506, increased neuronal proliferation and differentiation (115). Transplanting ENSCs along with these nanoparticles to the mouse colon in vivo also led to increased neuronal proliferation (115). The gut microbiota may also play a role in enteric neurogenesis that is dependent on 5-HT, as bacteria residing in the gut can induce 5-HT release from enteric neurons, via the 5-HT₄ receptor, (58), which then influences ENS development.

Studies have demonstrated the therapeutic relevance of targeting the 5-HT₄ receptor for neurogenesis (93). In studies where gut transection and anastomosis were carried out in murine models, oral delivery of the 5-HT₄ agonist, mosapride citrate promoted reconstruction of enteric neural circuits, which restored defecation reflexes that had been disrupted (100, 212). These data show the potential mechanisms underlying the partial success of 5-HT₄ receptor agonists as therapies for gut motility disorders. The 5-HT₄ agonist prucalopride has been shown to increase neurogenesis in vivo and prevent deficiencies in ENS development caused by the SERT-Ala56 mutation (151). Prucalopride also exerts protective effects on enteric neurons exposed to oxidative stress challenge (25), indicating that it may have utility in treating degenerative disorders of the ENS, though this implication has not yet been extensively explored.

In addition to neurogenesis, enteric neuronal 5-HT also impacts the proliferation and maintenance of the intestinal mucosa. Mice lacking neuronal 5-HT (TPH2 KO) show diminished mucosal proliferation, as evidenced by reduced villus height, crypt depth, and proliferation index (104). TPH2-R439H mice display similar mucosal defects, which could be reversed by treatment with sustained-release 5-HTP (120). SERT Ala56 mice, which have lower 5-HT availability, also have an underdeveloped mucosa (151). In contrast, mice lacking mucosal 5-HT (TPH1 KO) did not display an under-developed mucosal layer; proliferation index in the gut epithelium of these mice was unchanged, whereas crypt depth and villus height were greater compared to wildtype littermate controls (104). Altogether, the available data are suggestive of the notion that 5-HT derived from ENS neurons, but not from EC cells, contributes to epithelial proliferation. Mice that lack SERT (SERT KO) or adult mice that have been administered SSRIs, show significantly more mucosal growth and proliferation of epithelial precursor cells (104). Administration of the 5-HT_2A receptor antagonist, ketanserin reversed 5-HT-mediated mucosal proliferation; this effect is thus likely mediated by 5-HT_2A receptors expressed on submucosal cholinergic neurons that innervate the mucosal layer (104). Consistent with these findings, mice lacking the 5-HT_2A receptor have a smaller mucosal layer with shorter enterocytes (75). In contrast, mice exposed to SSRIs during embryonic development, through maternal administration, exhibit greater mucosal area (151). Finally, in vivo administration of the 5-HT₄ agonist tegaserod increases proliferation of crypt epithelial cells in mice and promotes epithelial cell migration in vitro, as modeled in Caco-2 cell cultures (207). These data provide support of 5-HT₄ agonists as a therapeutic that may enable growth and healing of the epithelial layer in conditions that damage the gut epithelium such as inflammatory diseases or chemotherapeutic regimens.

Role of 5-HT in Gut Motility

Peristaltic reflexes

Peristalsis is a complex set of interactions within the ENS responsible for propulsive contractions of the GI musculature that propel contents through the gut. Smooth muscle contraction on the oral end of the gut and relaxation on the aboral end is required to efficiently move digestive contents in a sequential manner. In 1899, Bayliss and Starling were the first to describe how a peristaltic reflex is initiated by an increase in intraluminal pressure; they found that they could reliably initiate peristaltic contractions via balloon distension of a dog intestine (13, 14). They observed that these reflexes remained intact after the intestine was severed from all extrinsic nerves and thus attributed the reflexes to a “local nervous mechanism” (13, 14). Trendelenburg coined the term “peristaltic reflex” in 1917, after confirming Bayliss and Starling’s results in isolated guinea pig intestine, also noting the intestine’s ability to produce reflexes independently of the brain (221). The “local nervous mechanism” is now known as the ENS.

Edith Bülbring and colleagues were the first to define the role of 5-HT in peristaltic reflexes and intestinal propulsion. In isolated guinea pig small intestine, they showed that application of 5-HT was sufficient to initiate peristaltic reflexes (35, 36) and that pressure-induced stimulation of the peristaltic reflex resulted in 5-HT release (37). They determined that 5-HT is released from EC cells in an intraluminal pressure-dependent manner (37). Later studies showed that other mechanical stimuli (e.g., brush stroking) applied to the mucosa also result in 5-HT release (19, 128), as do chemical stimuli that selectively activate EC cells (18, 172). 5-HT release from the mucosa has been shown to activate the ascending and descending circuits that comprise the peristaltic reflex and cause contraction and relaxation of smooth muscle, respectively. One way this may occur is via 5-HT released by EC cells, which act on IPANs, initiating neuronal reflexes necessary for contraction and relaxation (Figure 3) (23, 106, 123). The importance of mucosal 5-HT in peristalsis has been debated (201, 205), as neuronal 5-HT is also critical for normal gut motility patterns (140); however, mucosal 5-HT appears to have distinctive roles that are delineated below.

Figure 3. Mucosa and neuronal 5-HT in peristaltic reflexes and intestinal motility.

Mucosal 5-HT is released from enterochromaffin (EC) cells upon chemical or mechanical stimulation of the mucosa. A subpopulation of EC cells expresses the mechanoreceptor, Piezo2, that responds to distention/mechanical force causing 5-HT release (Inset). Mucosal 5-HT release causes downstream neuronal activation where intrinsic primary afferent neurons (IPAN) signal from the mucosa to ascending and descending neural pathways in the ENS to initiate contraction and relaxation respectively. Neuronal 5-HT present in a small population of enteric neurons (2% to 3%) project extensively and innervate different functional classes of neurons that drive intestinal motility. Neuronal 5-HT can mediate slow and fast excitatory synaptic transmission as well as play critical roles in the descending inhibitory pathways which culminates to intestinal motility. (A) A bolus of food in the gut lumen stimulates both ascending (B_i and descending (B_ii) reflex pathways that work cohesively to propel the food through the gut. Ascending interneurons activate excitatory motor neurons upstream of the stimulus to initiate smooth muscle contraction (B_i, red dotted box) while descending interneurons activate inhibitory motor neurons downstream of the stimulus to initiate smooth muscle relaxation (B_ii, blue dotted box). Made in © BioRender - biorender.com.

Roles of neuronal versus mucosal 5-HT in motility

Recent studies using a combination of genetic and pharmacological techniques have sought to define how both neuronal and mucosal 5-HT contribute to gut motility. Experiments have revealed that neuronal 5-HT is key for normal motility; in TPH2 KO mice, in vivo GI transit is significantly slower in terms of total transit time, colonic transit, and small intestinal transit (120, 140). This slower transit is also observed in mice lacking both TPH1 and TPH2 (TPH1/2 double KO), and in TPH2-R439H mice, which are deficient in neuronal 5-HT (85, 94). As previously mentioned, mice lacking TPH2 have multiple defects in ENS and mucosal development; therefore, it is difficult to decipher whether the slowed motility demonstrated in these animals is due to a lack of neuronal 5-HT or to an underdeveloped ENS. TPH2 is also present in neurons of the CNS; therefore, it is possible that lack of 5-HT in the CNS alters extrinsic pathways that could contribute to these defects in motility. Studies have shown that 5-HT contributes to slow and fast excitatory synaptic transmission and the resulting propagating contractions, suggesting that 5-HT in the ENS is a critical mediator of motility (66, 169, 213, 227). Although only a small proportion of neurons release 5-HT (2%–3%), these neurons have vast projections throughout the ENS, making connections with other 5-HT neurons, as well as other key mediators of motility including inhibitory nitric oxide synthase (NOS)-containing neurons, and ICCs (176). It is thus possible that the slowed motility observed in TPH2 deficient mice results from both developmental defects and/or the ongoing lack of neuronal 5-HT signaling. 5-HT neurons, which reside only in the myenteric plexus of the ENS, are thought of as interneurons that receive sensory input and signal to motor neurons to facilitate motility, but the close associations of 5-HT neuronal fibers with ICCs and blood vessels suggest that they have a motor function in signaling to these cell types (176). 5-HT neurons in the ENS also project to the submucosal layer, where they have a role in coupling motility with secretion (202). Recent RNA-seq data have classified 5-HT neurons with multiple morphological types, suggesting additional roles that require more study (164). A recently published study demonstrated that a pharmacological increase of enteric neuronal 5-HT availability does not affect CMMCs, suggesting that more research is also needed in this area to fully elucidate the roles of neuronal 5-HT in normal motility (154).

In contrast to TPH2-deficient mice, mice lacking mucosal 5-HT do not display defects in in vivo GI transit time, as measured in both TPH1 KO mice (140) and mice that were administered nonabsorbable TPH inhibitor drugs that restrict peripheral 5-HT production (24, 153). Further, with the mucosa removed from ex vivo colon preparations, peristaltic contractions could still be initiated with adequate stretching of the tissue, demonstrating that depletion of mucosal 5-HT does not eradicate CMMCs (126, 204). When peristaltic contractions were measured ex vivo, however, it was demonstrated that TPH1 KO mice displayed irregular CMMCs that often propagated in the retrograde direction (9, 107, 203), suggesting that mucosal 5-HT has a more subtle, yet important, role in regulating peristalsis. Taken together with previous data showing that EC cell-derived 5-HT can initiate contractions (38), this research suggests that mucosal 5-HT is not necessary for initiating propulsive contractions but is important for ensuring that these contractions are fully effective. A recent study in which EC cells were selectively ablated in adult mice showed that loss of EC cells resulted in delayed gastric emptying, total GI transit, and colonic transit, as well as reduced CMMC frequency (232), suggesting that EC cells are important for normal motility postdevelopment. Finally, studies have also shown that applying 5-HT receptor antagonists can block peristaltic contractions even in colons devoid of a mucosal layer, raising questions about the importance of endogenous signaling between EC cells and surrounding ENS neurons in motility (200).

Studies are now beginning to focus on how EC cells sense contents passing through the gut and, in turn, release 5-HT. It was recently discovered that subpopulations of EC cells express Piezo2 (2, 230), a mechanoreceptor that is also expressed in peripheral sensory neurons as well as specialized cells of the skin epidermis (53, 187, 234). Piezo2 senses mechanical stimulation, endowing gut epithelial cells with the ability to respond to distension and small mechanical forces imposed by luminal contents. EC cells release 5-HT downstream of Piezo2 activation (2), likely communicating with surrounding ENS neurons responsible for initiating peristaltic contractions. A more recent study used single-cell RNAseq to show that the vast majority of Piezo2-expressing cells in the gut epithelium also express TPH1, highlighting the significance of 5-HT in gut mechanosensation (145). Mice lacking mucosal Piezo2 displayed irregular small intestinal and colonic transit depending on the size of the intraluminal contents (220), suggesting that Piezo2 is required for modulating motility patterns in response to smaller intraluminal stimuli. Piezo2-expressing EC cells therefore contribute to the “fine-tuning” necessary for effective motility.

EC cells express synaptic proteins (18, 220), indicating that they may communicate rapidly with intrinsic and extrinsic neurons rather than via slower paracrine mechanisms; however, a recent anatomical study in which spinal ExPAN fibers were anterogradely labeled suggested that these fibers terminate at a distance from EC cells that would not enable synaptic communication (60). Optogenetic studies have shown that specific activation of all colonic epithelial cells, including EC cells, initiates ExPAN firing and local colonic contractions, which could be blocked by a combination of 5-HT₃ and 5-HT₄ receptor antagonists (146, 167). The latency between epithelial stimulation and neuronal activation varied greatly in these studies, indicating that communication between colonic epithelial cells and surrounding neurons occurred via fast and slow paracrine mechanisms. Future studies should investigate whether the pattern and numbers of EC cells innervated by extrinsic and intrinsic neurons differ depending on region of the gut.

Role of SERT in motility

Several transgenic mouse models and pharmacological interventions have shown the importance of SERT in gut motility. Mice in which SERT is genetically ablated (SERT KO mice) display abnormalities in GI transit, varying from slowed transit to faster transit, indicative of alternating constipation and diarrhea-like phenotypes, respectively (45). This variation may be due to desensitization of 5-HT receptors involved in producing motility. When averaged, SERT KO mice do not display changes in total GI transit time compared to wild-type littermates but do show slowed in vivo colonic motility as well as ex vivo peristaltic abnormalities (reduced CMMC length) (151). Mice exposed to the SSRI fluoxetine during development had slower intestinal transit compared to control-treated mice, including longer total GI transit and colonic transit times (151). The SERT KO and SSRI treatments have similar effects in inhibiting 5-HT inactivation and thus increasing 5-HT availability; however, SERT KO mice have a congenital deletion of SERT lasting into adulthood and the SSRI-treated mice were only exposed to fluoxetine during development (in utero and up to postnatal day 21) and not in adulthood when motility was assessed, which could explain the somewhat diverse results observed in these studies. The motility abnormalities in both SERT KO mice and fluoxetine-exposed mice were reversed by chemical sympathectomy with systemic 6-hydroxydopamine administration (151), suggesting that deletion or inhibition of SERT can increase central sympathetic input to the intestine, likely contributing to the slowing of motility (1). In a recent study of adult mouse colonic motility, ex vivo CMMC frequency was decreased after bath application of the SSRI fluoxetine, an effect that was abolished when the mucosa was removed, suggesting the importance of mucosal 5-HT in regulating peristaltic contractions (154).

Mice harboring the SERT Ala56 mutation, in which SERT is hyperfunctional and less 5-HT is available, display slower in vivo GI transit as well as slower ex vivo colonic motility, compared to wild-type littermates (151). As mentioned above, these mice develop a hypo-ganglionated ENS, which could partially explain these deficiencies in motility (151). The impact of SERT on gastrointestinal motility has relevance in several clinical areas. For example, multiple SERT mutations have been identified in autism spectrum disorder (ASD) (182, 183, 208), with SERT Ala56 being the most common (224). GI motility problems, especially constipation, are approximately four-fold more common in children with ASD (160). In addition to slowed motility, SERT Ala56 mice also display communication deficits and repetitive behaviors reminiscent of ASD (130, 208, 223, 224), suggesting that the SERT Ala56 mutation may result in developmental perturbations of 5-HT signaling relevant to multiple features of ASD. SERT Ala56 mice also have hyperserotonemia (224), which results from high levels of 5-HT in the blood and is common in individuals with ASD (50, 88, 166, 192). Another area of clinical relevance is exposure to SSRIs in utero. Several studies have supported the hypothesis that gestational exposure to anti-depressants, including SSRIs, impacts ENS development and GI function in humans; retrospective and prospective studies have shown that this exposure increases the likelihood of GI disturbances in childhood (170, 171, 194).

Therapeutic Targets for 5-HT in GI Function

TPH1 and SERT

DGBIs such as irritable bowel syndrome (IBS) are characterized by recurring abdominal pain or discomfort and altered stool patterns (144). No consistent biomarkers have been identified for IBS and thus it is diagnosed based on symptoms and according to stool patterns, as designated by the Rome IV criteria, as constipation-predominant (IBS-C), diarrhea-predominant (IBS-D), or mixed phenotype (IBS-M) (55).

Altered serotonergic signaling may contribute to the disordered motility and/or pain associated with IBS. Several studies that have examined levels of 5-HT or its modulatory factors (SERT, TPH1, EC cells) in IBS patients have yielded inconsistent results (Table 2). One study demonstrated that SERT expression is decreased in colon biopsies from both IBS-D and IBS-C patients (48); however, EC cell density and tissue agitation-evoked 5-HT release did not differ from healthy controls. Subsequent studies have confirmed the finding that SERT mRNA levels in both the colon and small intestine are lower in IBS patients (72, 77, 129), although one study found that there were no differences (41). In patients with IBS-D, mucosal 5-HT content and TPH1 mRNA levels were also found to be decreased (48). Another study confirmed that colon samples from IBS-D and IBS-C patients did not have differences in EC cell density compared to healthy controls; however, postinfectious IBS patient samples showed significantly higher EC cell density, indicating that gut 5-HT may play a more prominent role in this subtype of IBS (136). In another analysis of IBS patient samples of all phenotypes, there were no differences in EC cell count, SERT mRNA levels, or TPH1 mRNA levels when comparing patients with and without hypersensitivity, demonstrating that mucosal 5-HT content may not directly correlate with the painful symptoms of IBS (129).

Table 2.

Alterations of 5-HT in Gastrointestinal Disorders

GI disorder	EC cell density	TPH1 expression	SERT expression	Postprandial 5-HT levels	References
IBS-D	No difference	Decrease	Decrease		(48)
			Decrease		(72, 77, 129)
			No difference		(41)
				Increase	(7, 64, 116)
IBS-C	No difference		Decrease		(48)
			Decrease		(72, 77, 129)
			No difference		(72)
				Decrease	(64)
				No difference	(7)
Postinfectious IBS	Increase				(136)
				Increase	(64)
Ulcerative colitis	Increase	Increase	Decrease		(16, 48, 65)
Crohn’s disease	Increase	Increase	Decrease		(16, 48, 65)

Mucosal SERT impacts circulating 5-HT levels; therefore, blood 5-HT concentration can also indicate gut mucosal 5-HT availability. Postprandial 5-HT levels in platelet-poor plasma samples are reported to be increased in patients with IBS-D (7, 64, 116) or postinfectious IBS (64), but decreased (64) or unchanged (7) in IBS-C patients. These studies may be challenging to interpret, however, because the vast majority of 5-HT in the blood is localized to the platelets (161). Studies have also shown that fasting blood 5-HT levels are reduced in IBS-D patients (79) but drastically increased in IBS-C patients compared to healthy controls (7). A more recent study of patients of all IBS phenotypes demonstrated that fasting plasma 5-HT concentrations did not differ between healthy controls and IBS patients; however, levels of the metabolite 5-hydroxy indoleacetic acid (5-HIAA) were significantly decreased in IBS patients (217). Further studies are necessary to understand how 5-HT release in the GI tract is altered in IBS and whether disordered motility is a cause or the consequence of altered 5-HT signaling.

As described above, 5-HT released from EC cells initiates sensory and motor signaling in the gut; therefore, inhibiting 5-HT release may attenuate symptoms of IBS-D. Drugs have been developed to inhibit TPH synthesis in a peripherally restricted manner, so that 5-HT release is prevented at the level of the intestinal epithelium (i.e., from EC cells). Compounds such as LX-1031 have been shown to lower 5-HT levels in the mouse GI tract without affecting brain and enteric neuronal 5-HT levels (143, 153). In human trials, LX-1031 improved stool consistency as well as pain and discomfort in IBS-D and IBS-M patients who displayed a dose-dependent decrease in urine 5-HIAA levels (32, 40).

SSRI drugs, such as fluoxetine, have commonly been used in the treatment of IBS, but the mechanism by which they improve symptoms is unclear, as they inhibit SERT in both the brain and gut (214). SSRIs act centrally and alleviate anxiety and depression, which are often coincident with IBS and may exacerbate IBS symptoms (62, 109). SSRIs may also increase 5-HT availability in the gut and therefore affect sensorimotor function, potentially slowing gut transit time in IBS-D patients (99). Human trials show conflicting results about their efficacy in alleviating abdominal pain (39, 156), with one study showing that a limited course of SSRIs did not affect rectal sensitivity (135). Further studies are required to establish the efficacy of SSRIs and determine the differential effects of inhibiting SERT in the gut versus the CNS.

5-HT₁ receptors

The 5-HT₁ receptor plays a role in gastric motility; in humans, the 5-HT₁ agonist, sumatriptan delays gastric emptying and causes relaxation of the gastric fundus, enabling accommodation of larger volumes of food (54, 117, 210). In rodent studies, peripheral administration of the 5-HT_1A agonist buspirone had an inhibitory effect on fundic tone (237). Although 5-HT₁ receptor agonist drugs showed promise for treating the impaired fundic tone and the early satiety symptoms associated with functional dyspepsia, their use in dyspeptic patients has been accompanied by undesired off-target effects such as enhanced visceral sensitivity and constriction of coronary arteries (57). In contrast, the 5-HT_1P receptor exerts excitatory effects; its activation induces neurotransmitter release from IPANs, which initiate peristaltic reflexes in the gut (31, 103, 231). 5-HT_1P expression on IPANs also mediates chloride secretion in the gut (51). A recent study demonstrated functional 5-HT_1P receptors in human small and large intestines; application of a 5-HT_1P agonist resulted in long-lasting activation of human submucosal ENS neurons (4).

5-HT₂ receptors

Recent studies have shown that 5-HT_2B receptors on ICCs play an important role in colonic motility (122). Diabetic mice with a constipation-like phenotype displayed significant decreases in ICC 5-HT_2B receptor expression compared to nondiabetic mice, providing a possible explanation for slowed colonic motility in these mice (122). Conversely, ICC activity was stimulated by the 5-HT_2B receptor agonist BW723C86, and treatment with this drug increased colonic transit speed in vivo, correcting the slowed motility in diabetic mice (122). 5-HT_2B receptor antagonists can also modulate colonic motility; in studies performed in mice, high doses of the 5-HT_2B receptor antagonist RS-127445 reduced the frequency and amplitude of colonic contractions and reduced fecal output (12). 5-HT_2B receptors are also expressed on ExPANs and are involved in mediating pain (112). Visceral pain is a common symptom of IBS and studies performed in rats have shown that 5-HT_2B receptor antagonists can diminish stress-induced visceral hypersensitivity, as measured by visceromotor responses to colorectal distension (178).

5-HT₃ receptors

5-HT₃ receptor antagonists, such as alosetron, have shown efficacy in alleviating both diarrhea and abdominal pain associated with IBS-D (3, 186). Such drugs slow motility by blocking 5-HT₃ receptors on IPANs that are stimulated by EC cell-released 5-HT and also by inhibiting the 5-HT₃ receptors on interneurons and motor neurons that contribute to fast EPSPs (84). These actions result in decreased propulsive motility and local secretion, which alleviates diarrhea. The 5-HT₃ receptor antagonists likely inhibit pain by blocking 5-HT₃ receptors on extrinsic primary afferent nerves that convey pain and discomfort (10, 124, 147). Rodent studies have shown that alosetron inhibits nociceptive responses to colorectal distension and decreases activation of dorsal horn neurons in the spinal cord that are involved in pain signaling (133). Alosetron has been found to be more effective in female IBS-D patients than in male patients; clinical studies show that alosetron alleviated pain and discomfort and improved stool consistency in females yet did not improve symptoms in male patients (11, 43). Alosetron was removed from the market in 2000 due to GI side effects such as ischemic colitis but later reintroduced for treating only severe cases of IBS-D in female patients (73). The 5-HT₃ receptor also has a role in mediating nausea and vomiting. Stimulation of 5-HT₃ receptors present on the vagus nerve can lead to nausea and thus specific 5-HT₃ receptor antagonists, such as granisetron, have been used to treat nausea and emesis induced by chemotherapy and radiation therapy (168, 222).

To increase propulsive motility and secretion in the gut to alleviate constipation, 5-HT₃ receptor agonists, such as pumosetrag, have been developed (69). Efforts have been focused on partial agonists because 5-HT₃ receptor activation on extrinsic primary afferent fibers and vagal fibers can lead to pain and nausea, respectively. Results from clinical trials showed that pumosetrag was effective in relieving constipation and also reducing reflux events in individuals with gastroesophageal reflux disease (GERD) (47, 69).

5-HT₄ receptors

5-HT₄ receptor agonist drugs promote motility in the ENS by evoking peristaltic reflexes by acting on presynaptic nerve terminals to stimulate the release of neurotransmitters that mediate peristaltic contractions (71, 86, 141, 179, 218). This indirect action is thought to enhance naturally occurring reflexes rather than directly generate neurotransmission, which can ultimately inhibit propulsive motility (113). Transgenic mice that lack 5-HT₄ receptors display slowed colonic motility (141), suggesting that these receptors are critical in propulsive motility; however, these defects may also result from altered ENS development, as 5-HT₄ receptors have important neurotrophic roles as well.

The 5-HT₄ receptor has been exploited for treating IBS; 5-HT₄ receptor agonists have proven to be effective in relieving pain and constipation associated with IBS-C and can also accelerate the rate of gastric emptying (70, 159). These agonists activate 5-HT₄ receptors on presynaptic nerve terminals, evoking acetylcholine release and enhancing propulsive contractions (71, 86, 141, 179, 218). They likely also promote gut motility and alleviate constipation by acting on 5-HT₄ receptors on the intestinal epithelium to cause increased mucus discharge and chloride secretion (114). 5-HT₄ receptors are also present on ExPANs, but it is not clear how 5-HT₄ agonists alleviate visceral pain. Studies show that these agonists have an inhibitory effect on mechanosensory signaling, as they reduce firing of ExPANs in response to distension (198) and reduce visceral hypersensitivity (114).

The earlier developed 5-HT₄ receptor agonists, such as cisapride and tegaserod, were shown to be effective in treating constipation but were accompanied by cardiovascular side effects due to effects on other targets such as hERG potassium channels (56, 209). More targeted agonists, such as prucalopride, have since been developed for treatment of IBS-C (42, 185, 211), which have a favorable safety profile and show efficacy in both male and female patients with constipation (177). Studies have shown that 5-HT₄ receptor agonists have maximal effects when applied to the mucosal, rather than serosal, surface of the colon in ex vivo motility assays (78, 102, 114, 123); thus, studies are now aimed at developing nonabsorbable agonists that would only act on cells of the mucosal layer, which may be a better-targeted treatment.

5-HT and inflammation

The intestine provides an important interface between the body and the external environment and, as such, contains major components of the body’s immune system. The gut is equipped to simultaneously defend against numerous potentially pathogenic organisms and to also maintain a commensal microbiome (101, 127, 195). Although the gut microbiome is an important contributor to functions such as GI motility (191) and nervous system development (175), the immune system must constantly defend against microbial invasion (150). Interactions between neurons and immune cells in the gut are important in regulating these immune responses. 5-HT is an important paracrine messenger that participates in crosstalk between enteric neurons and immune cells and has a significant role in intestinal inflammation (34, 225). Immune cells express the 5-HT receptors 2, 3, 4, and 7 (199) and, via these receptors, 5-HT influences immune cell trafficking, chemotaxis, activation, and proliferation (5, 6, 8, 96). Some immune cells secrete 5-HT themselves, namely macrophages and T cells (8) and, in mice and rats, mast cells synthesize and release 5-HT and take up 5-HT via SERT (148).

Animal models of IBD have revealed how mucosal and enteric neuronal 5-HT may differentially modulate inflammatory processes in the gut. The most studied rodent IBD models include colitis induced by trinitrobenzene sulfonic acid (TNBS), which mimics pathological features of Crohn’s disease, and dextran sulfate sodium (DSS), which more closely resembles ulcerative colitis. Infections caused by Citrobacter rodentium, Trichinella spiralis, and Escherichia coli have also been studied (20, 34, 173, 174, 233). All these inflammation-based models result in increased EC cell density and decreased levels of epithelial SERT in the intestine. Studies have demonstrated that mucosal 5-HT is proinflammatory; in TPH1 KO mice and mice administered a peripherally restricted TPH inhibitor, colitis induced by TNBS or DSS is significantly diminished (97, 153, 199). Downregulation of SERT could be either a cause or a consequence of gut inflammation. In cultures of Caco-2 human epithelial cells, addition of the proinflammatory cytokines interferon-gamma and tumor necrosis factor reduced levels of SERT (14). In SERT KO mice, TNBS-induced colitis is exacerbated, suggesting that increased 5-HT availability in the gut contributes to inflammation (26).

In contrast to mucosal 5-HT, enteric neuronal 5-HT appears to have anti-inflammatory actions in the gut. This is best demonstrated in mice lacking neuronal 5-HT (TPH2 KO), which, after DSS-induced colitis, had increased mortality, greater inflammation severity, and higher pro-inflammatory cytokine levels in the colon, compared to wild-type littermates (91). The function of neuronal 5-HT may therefore be to counteract inflammation-induced damage to ENS neurons; however, this area of study should be investigated using more targeted techniques.

Studies show that 5-HT may modulate gut inflammation via the 5-HT₄ receptor. In animal models of colitis, activation of 5-HT₄ receptors on the colonic epithelium reduces the development of colitis and accelerates recovery (207) Colitis also promotes 5-HT₄ receptor-mediated neurogenesis that drives glial cells to transdifferentiate into neurons (17). Consistent with these findings, ENS hyperplasia is found in inflamed bowel segments of IBD patients and, in animal models, ENS hyperplasia increases predisposition to both TNBS and DSS-induced colitis (152). Although hyperplasia appears to increase susceptibility to inflammation, neurogenesis may also be protective, as 5-HT₄ receptors promote neuroprotection against oxidative stress associated with inflammation (17, 100, 141). To develop future therapies for IBD, it is critical to further investigate how 5-HT₄ receptor-mediated neurogenesis impacts inflammatory states.

Several aspects of 5-HT signaling have been noted to be altered in patients with IBD (Table 2). The density of EC cells is increased in intestinal biopsies taken from individuals with either Crohn’s disease (CD) or ulcerative colitis (UC) (16, 48, 65). EC cells isolated from such biopsies secrete significantly more of the proinflammatory cytokine interleukin 1p, and release more 5-HT in response to treatment with lipopolysaccharide, compared to healthy controls (131). Ileum and colon biopsies from patients with either UC or CD also displayed higher TPH1 mRNA levels and significantly lower SERT mRNA levels compared to healthy controls (125). Patients with celiac disease and postinfectious IBS also display increases in EC cell number, as well as increases in postprandial 5-HT levels (49, 90, 206). All these conditions are also associated with a decrease in epithelial SERT expression levels (48), further supporting the idea that increased mucosal 5-HT availability may contribute to intestinal inflammation.

Conclusion

Research over the last 60 years has revealed the important roles of 5-HT in GI function. Early studies demonstrated that addition of 5-HT to the intestine evokes motility patterns and the mechanisms underlying this response have proven to be increasingly complex. EC cells of the GI epithelium and some ENS neurons release 5-HT, and multiple cell types that reside in the gut modulate 5-HT signaling via SERT and the expression of 5-HT receptors. This diversity of cell types and receptor functions has made it difficult to determine precisely how 5-HT mediates GI motility while simultaneously contributing to an array of other functions such as sensory signaling, epithelial homeostasis, inflammation, and neurogenesis. Studies should further examine the distribution of 5-HT receptors throughout the gut and their physiological relevance to these functions. This knowledge could reveal cell type-specific treatments for DGBIs; for example, 5-HT₃ receptor activation stimulates peristalsis and also mediates pain signaling, and it is not yet known whether treatments can target one function but not the other. Further, although studies have revealed potential functions of each 5-HT receptor, much of this knowledge has not been translated into effective therapies. For example, activation of the 5-HT₄ receptor ameliorates inflammation in mouse models of colitis, but there is not yet an effective 5-HT₄ receptor agonist drug for IBD. Similarly, 5-HT is known to be a potent growth factor in the ENS, but it is unclear how its signaling may be targeted to ENS regeneration in aging and neurodegenerative disorders. The challenge of future research will be to better understand the different components of the serotonergic system of the GI tract, how they work together, and how they contribute to pathophysiology underlying GI diseases.

Didactic Synopsis.

Major teaching points

• The gut contains the vast majority of the body’s 5-HT, synthesized mostly by enterochromaffin (EC) cells of the mucosa and, in small part, by neurons of the enteric nervous system (ENS).

• 5-HT availability in the gut is modulated by the serotonin reuptake transporter (SERT), which is expressed throughout the gut epithelium, in ENS neurons, and in circulating platelets.

• Enteric neuronal 5-HT acts as a growth factor in development of the ENS and proliferation of the intestinal mucosa.

• Both mucosal and neuronal 5-HT are involved in gastrointestinal (GI) motility; mucosal 5-HT is not necessary for initiating peristaltic contractions but is important for ensuring these contractions are fully effective.

• The 5-HT receptors present throughout the GI tract that have roles in motility are 5-HT₁, 5-HT₂, 5-HT₃, 5-HT₄, and 5-HT₇, which are G-protein-coupled receptors except for the 5-HT₃ receptor, which is ionotropic.

• 5-HT signaling appears to be altered in disorders of gut-brain interaction (DGBIs), such as irritable bowel syndrome (IBS), and has been targeted for treatment of abdominal pain and disordered gut motility, the two main symptoms of DGBIs.

• Mucosal 5-HT promotes inflammation in the gut and is upregulated in inflammatory bowel diseases.