Enteric nervous system development: what could possibly go wrong?

Abstract

The gastrointestinal tract contains its own set of intrinsic neuroglial circuits - the enteric nervous system (ENS) - which detects and responds to diverse signals from the environment. Here, we address recent advances in the understanding of ENS development, including how neural-crest-derived progenitors migrate into and colonize the bowel, the formation of ganglionated plexuses and the molecular mechanisms of enteric neuronal and glial diversification. Modern lineage tracing and transcription-profiling technologies have produced observations that simultaneously challenge and affirm long-held beliefs about ENS development. We review many genetic and environmental factors that can alter ENS development and exert long-lasting effects on gastrointestinal function, and discuss how developmental defects in the ENS might account for some of the large burden of digestive disease.

Fig. 1 |. Migration of neural-crest-derived progenitors to the primordial gut.

a | A diagram of a developing mouse fetus after the formation of the primordial foregut, midgut and hindgut. The vasculature defines regions of the fetal bowel: the coeliac trunk supplies the foregut, the superior mesenteric artery supplies the midgut and the inferior mesenteric artery, supplies the hindgut. The foregut includes the portion of the bowel from the pharynx to the entry of the pancreatic and bile ducts in the mid-duodenum; the midgut extends to the mid-transverse colon; and the hindgut extends to the ectoderm of the anal canal. The midgut grows rapidly and herniates transiently into the umbilical cord but then folds extensively and rotates upon returning to the abdomen. Neural-crest-derived cells were first shown to migrate to the bowel from the vagal level (corresponding to somites 1–7) and sacral axial level (dark red; caudal to somite 28). A third source of crest-derived precursors of neurons or glia (blue) enter the colon later in ontogeny, among the Schwann cell population found in innervating extrinsic nerve fibres. Most recently, molecular genetic studies have suggested that the vagal level of the crest is more complex than previously suspected. Crest-derived cells from axial levels 1–2 (green) migrate to the oesophagus within the descending fibres of the vagus nerves. Properties of the more caudal vagal crest, at levels 3–7 (red), are more like those of the truncal crest, which gives rise to sympathetic ganglia. This ‘sympatho-enteric’ crest colonizes the entire gut and is the major source of enteric neurons and glia. Sacral-crest-derived cells colonize only the post-umbilical gut. The proportion of enteric neurons derived from the sacral crest (purple) is relatively small and higher distally than proximally. In contrast to vagal-crest-derived cells, which migrate proximo-distally, sacral-crest-derived cells migrate in a distal-to-proximal direction. b | During the folding of the midgut, at embryonic day 11 (E11) to E11.5, the presumptive ileum is transiently located next to a loop of post-caecal bowel (presumptive ascending colon). The dorsal mesentery intervenes between them. A subset of cells within the descending vagal enteric neural-crest-derived cell (ENCDC) population (red) takes a shortcut through the mesentery (pink) to enter the still-to-be colonized gut distal to the caecum. These cells do not have to traverse the caecum. Part a is adapted with permission from REF, Elsevier.

Fig. 2 |. Molecular regulation of cell-type diversification in the enteric nervous system.

Enteric nervous system (ENS) progenitors must proliferate to maintain an adequate progenitor pool while a certain proportion become bipotential progenitors that generate cells that diverge along neurogenic or gliogenic trajectories. Neurogenic commitment requires downregulating transcription factor SOX10 and maintaining receptor tyrosine kinase RET expression, whereas gliogenesis requires maintaining SOX10 and downregulating RET. Paired-like homeobox 2B (PHOX2B) expression is maintained in all enteric neurons and some enteric glia. There are many neuronal subtypes in the ENS, including those marked by vasoactive intestinal peptide (VIP), choline acetyltransferase (ChAT), serotonin (5-HT) and many others. The full extent of enteric glial diversity remains unknown, but morphologically distinct glia are found in at least three distinct locations.

Fig. 3 |. The ‘outside-in’ development and columnar organization of the enteric plexuses.

a | Schematic of the murine bowel during embryonic development illustrates the radial migration of enteric neural-crest-derived cells (ENCDCs) from the muscularis externa into the submucosa. The rudimentary mucosal epithelium expresses netrins, which serve as chemoattractants, and sonic hedgehog (SHH), which serves as a chemorepellent. The subset of ENCDCs that expresses the netrin receptor DCC migrates into the submucosa to populate the submucosal plexus (SMP). SHH signalling through the growth arrest-specific protein 1 (GAS1) receptor prevents them from migrating into the mucosa and extending premature projections to the epithelium. Laminin 111, which can convert netrin-DCC signalling from attractive to repulsive, is expressed in the basal lamina immediately underneath the epithelium and might serve as an additional cue that keeps ENCDCs from migrating beyond the submucosa into the epithelium. The timelines for myenteric plexus (MP) and SMP development in mice and humans are illustrated below from embryonic day 8 (E8) through postnatal day 14 (P14) and from week 4 of gestation, respectively. The timelines are based on birth-dating of enteric neuronal subtypes in mice; human data are much more limited and suggest that some ENCDCs have colonized the hindgut as early as week 8 of gestation and that SMP development lags behind MP development by ~3 weeks. Timelines are illustrated as continuums without definitive end points because recent reports suggest that there is robust enteric neurogenesis in the mature murine gut, and it remains unknown to what extent this occurs in humans. b | Schematic of the mature small intestine illustrating the laminar organization of the bowel and its integrated MP and SMP. The MP is located in between the circular (CM) and longitudinal (LM) layers of smooth muscle, and the SMP is located in the submucosa. Clonally related neurons and glia (shown in the same colours) are distributed in columns along the radial axis, as highlighted in the boxed area. c | The boxed area in part b is expanded to illustrate the observations made by lineage tracing of the progeny of individual SoxlO-expressing ENCDCs using the multicolour Confetti reporter and examining clonally related cells (which all express the same reporter colour) in mature bowel. Three types of clones were observed: neuronal (N; depicted in blue), glial (G; green) and those that contained both neurons and glia (NG; red). The majority of clonally related cells were distributed in a columnar fashion, similar to the CNS. All three types of clones can be observed in the MP, but only NG and G contribute to the SMP (neurons and/or glia) and mucosa (glia). Sister cells are distributed in register along the radial (serosa-to-mucosa) axis of the bowel.

Fig. 4. Neuroglial diversification and cellular interactions in the enteric nervous system.

| a | Schematic illustrating the major neuronal subtypes found in the enteric plexuses and highlighting the subset that forms the peristaltic reflex microcircuit. Luminal distention or mucosal deformation triggers direct activation of mechanoreceptive endings of intrinsic primary afferent neurons (IPANs) as well as indirect activation of IPANs upon serotonin (5-HT) release by enterochromaffin cells (ECs) in the epithelium. IPANs release acetylcholine (ACh) to activate ascending and descending interneurons, which stimulate excitatory and inhibitory motor neurons, respectively. Motor neuron activity leads to oral contraction and anal relaxation of intestinal smooth muscle, which propels luminal contents in the proximal-distal direction. Ascending interneurons express enkephalin (Enk) and descending interneurons release ACh and 5-HT. Excitatory motor neurons secrete ACh and substance P (SP), whereas inhibitory motor neurons secrete nitric oxide (NO), vasoactive intestinal peptide (VIP) and the purine β-nicotinamide adenine dinucleotide (βNAD). Secretomotor and vasomotor neurons of the submucosal plexus (SMP) secrete ACh or VIP. GABAergic and dopaminergic (DA) neurons are also found in enteric plexuses, but their synaptic targets are unknown. b | Schematic of enteric glial subpopulations and their interactions with other cell types. Mucosal glia are found in the lamina propria immediately underneath the epithelium and have been reported to interact with enteroendocrine cells (EECs; specialized epithelial cells that release peptides and other signals in response to luminal stimuli, such as ingested nutrients or mechanical distention), a subset of immune cells known as group 3 innate lymphoid cells (ILC3s; important for antimicrobial defence and maintaining tolerance to commensal microbiota), nerve fibres and blood vessels. Intraganglionic glia are found within the myenteric plexus (MP) and SMP, closely apposing and partially ensheathing neurons (depicted as blue circles). A subset of intraganglionic glia is connected by gap junctions. Long, bipolar, intramuscular glia are found in the circular and longitudinal muscle layers (CM and LM, respectively), in close association with nerve fibres (not shown) that innervate the smooth muscle. CGRP, calcitonin gene-related peptide. Part a is adapted from REF, Springer Nature Limited.