Mechanisms of enteric neuropathy in diverse contexts of gastrointestinal dysfunction

Abstract

The enteric nervous system (ENS) commands moment‐to‐moment gut functions through integrative neurocircuitry housed in the gut wall. The functional continuity of ENS networks is disrupted in enteric neuropathies and contributes to major disturbances in normal gut activities including abnormal gut motility, secretions, pain, immune dysregulation, and disrupted signaling along the gut–brain axis. The conditions under which enteric neuropathy occurs are diverse and the mechanistic underpinnings are incompletely understood. The purpose of this brief review is to summarize the current understanding of the cell types involved, the conditions in which neuropathy occurs, and the mechanisms implicated in enteric neuropathy such as oxidative stress, toll like receptor signaling, purines, and pre‐programmed cell death.

Key points.

• Enteric neurons command ongoing gut functions such as motility and secretions.

• Enteric neurons die due to multiple types of insults such as inflammation, exposure to drugs, infection, metabolic stress, autoimmune targeting, and old age.

• Mechanisms responsible for enteric neuron death include signals that initiate oxidative stress, purinergic signaling, toll like receptors, and several intracellular death cascades.

• A deeper understanding of mechanisms involved in enteric neuropathies would be beneficial to protect neurons and preserve gut function in multiple disorders.

1. INTRODUCTION

The enteric nervous system (ENS) is the largest and most complex branch of the autonomic nervous system and fulfills integrative functions to provide moment‐to‐moment command of gastrointestinal motility, fluid exchange, and immune homeostasis. The cellular units of the ENS are enteric neurons and glia, which are organized into two interconnected, gangliated plexuses with neurocircuits that control mucosal functions in the submucosal plexus and patterns of motor activity in the myenteric plexus. Intrinsic (enteric) neurocircuits interact extensively with extrinsic parasympathetic, sympathetic, and sensory neurons to coordinate homeostatic signaling along the brain–gut axis and adapt to ongoing systemic needs. Enteric neurons and glia also exhibit extensive bi‐directional interactions with gut immune cells that influence local and systemic defensive functions.

Given the central role of the ENS in coordinating gut functions, it is of little surprise that conditions in which the neural innervation supplying the GI tract is impaired produce major disturbances in normal gut activities that are long‐lasting and difficult to treat. Collectively, these conditions are referred to as enteric neuropathies and present with abnormal gut motility, secretions, pain, immune dysregulation, and disrupted signaling along the gut–brain axis due to enteric neuroplasticity, which involves functional reorganization of neuron properties and intercellular signaling. ¹ The conditions under which enteric neuropathy occurs are diverse and the mechanistic underpinnings are incompletely understood. The purpose of this brief review is to summarize the current understanding of the cell types involved, the conditions in which neuropathy occurs, and the mechanisms implicated in enteric neuropathy.

2. ENTERIC NEURON TYPES AND FUNCTIONS

Enteric neurons are diverse, and a basic understanding of major functional subtypes affected by neuropathy is integral to developing a mechanistic view of the resulting changes in gut functions. While it is not our intention to provide a comprehensive description of neuron subtypes, the following discussion provides an overview of major functional classes and characteristics. Readers are referred to excellent recent review articles for further information. ²

Enteric neurons are broadly grouped into four major functional classes: intrinsic primary afferent neurons (IPANs), motor neurons, interneurons, and intestinofugal neurons. IPANs are present in the myenteric and submucosal plexuses ³ and are multipolar cells with a Dogiel type II morphology. These neurons have at least one process that branches into the mucosa and multiple other projections that extend from the cell body to synapse with other neurons and enteric glia within the plexus. ³ IPANs are commonly considered to exhibit “sensory” functions whereby mucosal stimuli are detected and conveyed to the rest of ENS to coordinate motor responses. Primary afferent neurons are excitatory in nature and stimulate neurocircuitry by releasing transmitters such as acetylcholine and substance P. Enteric interneurons convey signals between neurons in the myenteric plexus and are primarily unipolar relay cells with Dogiel type I morphology. There are two major classes of interneurons; ascending which transmits information orally and descending which transmits information in the aboral direction. ⁴ Enteric interneuron neurochemical coding is diverse; however, ascending interneurons typically use acetylcholine and tachykinins as neurotransmitters while descending interneurons use combinations of acetylcholine, nitric oxide (NO), vasoactive intestinal peptide (VIP), and adenosine triphosphate (ATP) as neurotransmitters. ⁵ Additionally, there are mechanosensitive descending nitrergic interneurons that suppress the activity of primary afferent neurons by being sensitive to stretching in the longitudinal direction. ⁶ Motor neurons are present in both submucosal and myenteric plexuses and control glandular secretions (secretomotor), vasoactive functions (vasomotor), and contractions and relaxations of the longitudinal and circumferential smooth muscle layers. Excitatory motor neurons are primarily cholinergic and release acetylcholine to promote contractions, while inhibitory motor neurons prevent smooth muscle contraction by releasing nitric oxide and purines. ⁷ Intestinofugal neurons are the only enteric neuron subtype with projections that leave the intestine. These cells are housed in the myenteric plexus and send projection outward to postganglionic sympathetic neurons which then project back to the gut to influence GI motility, secretion, and blood flow. Intestinofugal neurons have a Dogiel type I morphology and are mechanosensitive. They are primarily activated by increased luminal volume but are also heavily influenced by input from other enteric neurons, and their main function is to integrate information from the intestine with other digestive organs. ⁸

It is worthwhile to note that each of the neuron subtypes discussed above are accompanied by enteric glia. Enteric glia are the cellular partners of enteric neurons and are associated with neuron cell bodies in submucosal and myenteric plexuses, nerve fibers in interganglionic connective tracts, and with nerve fibers that extend to the mucosa and smooth muscle. The importance of enteric glia cannot be overstated in relation to enteric neuropathy given that enteric glia regulate homeostasis, support neuronal functions and survival, and adjust neurotransmission. ⁹ Myenteric glia support motor function through bi‐directional signaling with enteric neurons that involves glial activity encoded by Gq signaling pathways and intercellular calcium responses. ¹⁰ , ¹¹ Additionally, submucosal and mucosal glia help to regulate barrier function of the intestinal mucosa and enhance secretomotor function. ¹²

3. CONDITIONS OF ENTERIC NEURON DEATH

Identifying the conditions under which enteric neurons die is central to understanding causal mechanisms and potential therapies. Enteric neurons are lost in both physiological and pathophysiological settings; however, abnormal losses caused by pathophysiology are considered neuropathy while losses during development reflect normal maturation and refinement of the network. ¹³ The following discussion summarizes several exemplary conditions in which enteric neuron death is a key feature of the pathophysiology observed in humans and animal models.

3.1. Inflammatory related enteric neuropathy

Inflammation is a potent driver of enteric neuroplasticity, which can involve both neuron gains and losses depending on the severity, stage, and nature of the inflammatory insult. Losses of up to 20% of myenteric neurons occur in several animal models of acute gastrointestinal inflammation including tri‐ or di‐nitrobenzene sulphonic acid (TNBS/DNBS) colitis in guinea pigs, ¹⁴ , ¹⁵ rats ¹⁶ , ¹⁷ , ¹⁸ , ¹⁹ and mice, ¹¹ , ²⁰ , ²¹ , ²² , ²³ , ²⁴ dextran sodium sulphate (DSS) colitis in mice, ²² , ²⁵ and IL‐10 knockout mice. ²² Nitrergic neurons are most susceptible to inflammation and are preferentially lost in several models ²² however, indiscriminate neuron loss has been reported to occur in guinea pig and rat TNBS colitis models. ¹⁴ , ¹⁸

Similar degeneration of enteric neuron cell bodies and axons has been reported in samples from humans with Crohn's disease ²⁶ and ulcerative colitis (UC). ²⁵ Neuronal apoptosis increases in the myenteric plexus during UC and myenteric apoptosis correlates with disease activity in Crohn's disease. ²⁷ However, assessments of neuron numbers in IBD are variable and appear to differ based on several factors including disease type and stage. For example, some reports indicate no change in neuron numbers in involved regions during Crohn's and a decrease in myenteric neurons in uninvolved areas ²⁸ while others report neuronal hypertrophy and hyperplasia. ²⁹ In Crohn's an increase in the size of neural fibers in the mucosa, submucosa, and myenteric plexus have been reported. ³⁰ Additionally, hypertrophy in the mucosa is more often associated in areas where the submucosa overlaps and has been demonstrated in the ileum and colon. ³⁰ Hypertrophy in Crohn's has been identified in areas of the GI tract that are severely affected by inflammation, often nearby plasma cells, lymphocytes, and mast cells with higher levels of cellular infiltrate around the fibers. ³⁰ Interestingly, this type of hypertrophy is uncommon in UC, ³⁰ although there have been signs of increased enteric glial activity due to elevated levels of glial‐neurotrophic factor in patients with inflamed. ³¹ A decrease in submucosal ganglia has been observed in experimental colitis in rats while the number of ganglia seems to remain stable in the myenteric plexus. ³² Although there were reduced numbers of neurons and signs of hyperplasia of the smooth muscle, the density of axons along the smooth muscle cells was unchanged. In contrast, significant neuronal apoptosis is observed in the myenteric plexus during DNBS colitis in mice which remains consistent following resolution of acute inflammation. ²³ Similar discrepancies are observed in the popular mouse DSS colitis model in which decreases, ²² , ²⁵ increases ³³ and no change ³⁴ , ³⁵ in neuron numbers have been observed. Why such differences occur is not entirely clear but appears to depend on the severity of the inflammatory insult and contributing factors such as the microbiota. It is also possible that differences in neuron numbers contribute to disease susceptibility in IBD and that changes in enteric neuron numbers in mouse models reflect adaptive attempts to control inflammation given that the number of enteric neurons affects the severity of inflammation in mouse models of colitis. ³⁶

3.2. Drug induced enteric neuropathy

Gastrointestinal side effects are a major limitation of several classes of drugs due to their effects on peripheral neuropathy. Cardiovascular drugs, chemotherapy, antibiotics, and biologics all have detrimental effects on the peripheral nervous system. ³⁷ Statins, for example, are widely used to treat cardiovascular disease and prolonged use is associated with high rates of peripheral neuropathy due to membrane dysfunction, disruption of ubiquinone synthesis, and misuse of energy in peripheral nerves. ³⁸

Detrimental effects of chemotherapeutic drugs on the ENS are clear. Oxaliplatin is a first line chemotherapeutic drug used in the treatment of tumors resistant to first‐ and second‐generation platinum‐based agents and increases overall survival rates of patients with colorectal cancer. However, acute gastrointestinal side effects limit drug dosing and side effects persist up to 10 years following treatment. The cause of these effects appears to involve significant enteric neuron degeneration in the submucosal and myenteric plexuses. ³⁹ , ⁴⁰ , ⁴¹ Both nitrergic and cholinergic neurons are lost following treatment with oxaliplatin; however, there is a relative increase in the proportion of nitrergic neurons remaining in the myenteric and submucosal plexuses. In contrast, nitrergic neurons are initially lost at a greater rate following treatment with the chemotherapeutic drug 5‐fluorouracil in the mouse colon myenteric plexus followed by balanced losses in both excitatory and inhibitory subsets ⁴⁰ while there is a relative increase in the proportion of cholinergic myenteric neurons following treatment with irinotecan. ⁴² These observations suggest that enteric neuropathy is a consistent feature of multiple anticancer drugs but that neuron subtypes differ in their susceptibility to specific drugs. Alternatively, the antibiotic metronidazole causes axonal degeneration resulting in motor and sensory peripheral neuropathy, optic and autonomic neuropathy. ⁴³ While enteric neuropathy has not been characterized by prolonged use of statins and metronidazole, they both are implicated in autonomic neuropathy and have the potential to contribute to enteric neuropathy providing an interesting area of further study.

3.3. Infectious disease related enteric neuropathy

Acute infections are a potent driver of neuroplasticity in the gut and are a major contributor to the subsequent development of post‐infectious irritable bowel syndrome. ⁴⁴ Infections involve an inflammatory component, so it is of little surprise that enteric neuropathies also occur under these conditions. Enteric neuropathy can be induced by a range bacterial and parasitic infections including enterotoxigenic bacteria (E. coli), protozoan parasites (T. cruzi), ⁴⁵ nematode parasites (S. venezuelensis), ⁴⁶ and gram‐negative bacteria (S. enterica and Y. pseudotuberculosis). ⁴⁶ , ⁴⁷ A classic example of gastrointestinal neuropathy due to infection is digestive Chagas disease. Protozoan parasite T. cruzi infection produces decreased motility of the colon irrespective of the amount of food eaten post infection. T. cruzi infection has been shown to cause a decrease of neuronal nitric oxide synthase suggesting a decrease of nitrergic neuron activity contributing to colonic dysmotility post‐infection. ⁴⁵ Additional examples of gastrointestinal parasitic infections which can contribute to neuronal cell death include Schistosoma, Giardia, and Entamoeba, while other examples of bacterial pathogens include S. typhimurium, SpiB, and Y. pseudotuberculosis. These bacterial infections trigger long‐term impairment of the GI tract in mice through mechanisms that involve a cause preferential loss of VGLUT2+ myenteric neurons in the ileum and colon. ⁴⁷ VGLUT2+ myenteric neurons in mice appear to be descending interneurons that are activated indirectly by mechanical stretch, which could explain some of the motor defects observed following infection. ⁴⁸

Interestingly, there is also evidence suggesting that viral infections, such as Sars‐CoV‐2, ⁴⁹ human polymavirus or John Cunningham virus (JC), ⁵⁰ and human immunodeficiency virus type 1 (HIV‐1), may be implicated in enteric neuropathy. GI symptoms are common in cases of severe COVID‐19 illness ⁵¹ and the digestive tract is a potential entry point for Sars‐CoV‐2. ⁵² , ⁵³ Data obtained from brain organoids show that central nervous system neurons die following exposure to Sars‐CoV‐2. ⁵⁴ Endoplasmic reticulum stress has been shown to increase during Sars‐CoV‐2 infection of enterocytes. ⁵⁵ Consequently, this increases DAMPs further increasing the release of VIP, and inhibiting water absorption leading to diarrhea. Whether enteric neurons are also susceptible to death following Sars‐CoV‐2 inflection remains unknown but is an important consideration of long‐term effects following the pandemic. Interestingly, HIV‐1 transactivating factor protein (Tat) has been identified as one of the major pathogenic contributors to HIV associated diarrhea in patients. ⁵⁶ Tat effects the function ENS by altering the intracellular calcium concentration of enterocytes and has been shown to increase neuronal excitability of cultured enteric neurons. ⁵⁷ , ⁵⁸ Additionally, Tat has been to shown to stimulate the release of proinflammatory cytokines via the activation of enteric glia through the over expression of S100B and GFAP which cause an increase of in proinflammatory factors such as iNOS protein and NO contributing to the ongoing diarrhea experienced by HIV patients. ⁵⁹ Other viruses which cause GI disruption are norovirus, rotavirus, and adenovirus to list a few.

3.4. Metabolic related enteric neuropathy

Metabolic diseases can be inherited or acquired and encompass several conditions that feature disrupted metabolism; the most common of which are Type 2 diabetes and obesity. Diabetes is characterized by dysregulated blood glucose levels due to destruction of pancreatic islet cells (Type 1 diabetes) or a loss of insulin sensitivity (Type 2 diabetes). Subsequent effects of abnormal blood glucose involve inflammation and damage to peripheral neurons over time. Diabetic neuropathy is most well‐known for affecting the extremities such as the hands and feet; however, diabetic neuropathy also extends into the ENS and contributes to a high incidence of GI symptoms such as constipation and diarrhea. Evidence from animal models suggests that enteric neurons are susceptible to death during both Type 1 and Type 2 diabetes, as well as obesity; however, the specific mechanisms and types of neurons affected may vary. Myenteric neurons are lost in a region‐specific manner in rat models of Type 1 diabetes with nitrergic neurons being most affected in the colon and peptidergic (CGRP+) neurons exhibiting susceptibility in the ileum ⁶⁰ while myenteric nitrergic neurons are vulnerable to degeneration in models of type 2 diabetes. ⁶¹ , ⁶² Type 2 diabetes involves low‐grade inflammation in the intestine characterized by increases in proinflammatory cytokines including TNF‐a, IL‐1b, and IL‐6 ⁶³ , ⁶⁴ and decreases in IL‐18 and IL‐22 expression which are typically involved in intestinal barrier function and defense against pathogens. ⁶⁵ , ⁶⁶ The proinflammatory environment, weakened barrier, and compromised gut defenses are considered important contributors to mechanisms of enteric neuron death in type 2 diabetes. This is further supported by human studies showing a decrease in diabetic ganglion size in the colon due to increased apoptosis and loss of peripherin, nNos, NPY, and ChAT neurons. ⁶⁷ These, and mechanisms involving increased oxidative stress, will be discussed further later in the review.

3.5. Autoimmune disorders

Autoimmune disorders represent a different mode of enteric neuropathy driven by antibodies that target enteric neurons. The most characterized example of this scenario occurs in paraneoplastic syndromes in which autoantibodies against the neuronal RNA‐binding protein HuC/D produce gut dysmotility through mechanisms that involve enteric neuron death. ⁶⁸ , ⁶⁹ Patients with paraneoplastic dysmotility often have high titers of circulating antineuronal antibodies in their serum. Among these, anti‐HuC/D antibodies are of particular importance because these antibodies recognize a group of proteins expressed by both neurons and neoplastic cells. In vitro work with anti‐Hu antibodies isolated from patients with small lung carcinoma and from commercial sources showed that anti‐HuD antibodies induce apoptosis which is preceded by immediate spike discharge in myenteric neurons. ⁷⁰ , ⁷¹ Similar neurotoxic outcomes have been observed in enteric neurons exposed to patient serum containing antibodies against gonadotropin‐releasing hormone (GnRH), which are often found in patients presenting with dysmotility following treatment with GnRH analogues. ⁷² Interestingly, commercial GnRH antibodies did not affect neuronal viability.

Anti‐enteric neuronal antibodies are also prevalent in irritable bowel syndrome and are thought to contribute to neuronal death and/or damage underlying symptoms. ⁷³ While the exact nature of the antigens remains unknown, immunolabeling suggests extensive overlap with Hu + enteric neurons. Likewise, autoantibodies present in serum from individuals with multiple sclerosis (MS) target cells of the ENS and anti‐ENS antibodies are thought to contribute to gastrointestinal symptoms such as constipation. ⁷⁴ Corroborating evidence from the encephalomyelitis (EAE) mouse model of MS shows increased gastric emptying and delayed whole gastrointestinal transit with decreased colonic motility. ⁷⁴ Interestingly, B‐cell deficiency was protective against colonic dysmotility in this model, supporting the role for anti‐ENS antibodies in the GI phenotype. While the antigen for anti‐ENS antibodies in MS is unknown, it is possible that similar antigens as those associated with autoantibodies in the CNS are present in the ENS or that unique GI antibodies such as those identified in MS patients that target mucosal antigens are responsible. ⁷⁵ Enteric neuron subtype susceptibility to MS is unclear as the mouse EAE model exhibits a decrease in the proportion of ChAT+ myenteric neurons while human data from MS patients suggest the opposite. ⁷⁶

3.6. Physiological aging

The gut has been said to lose its mind with age based on extensive age‐related neurodegeneration observed in animals and humans. ⁷⁷ Age‐related enteric neurodegeneration aging often manifests as constipation, but can also contribute to dysphagia, reflux, and incontinence. Several reports indicate that up to half of all myenteric neurons are lost in old age in mice, ⁷⁸ , ⁷⁹ rats, ⁸⁰ , ⁸¹ guinea pigs, ⁸² and humans. ⁸³ , ⁸⁴ , ⁸⁵ , ⁸⁶ However, the consistency of large magnitude age‐related enteric neurodegeneration is unclear as only minimal losses are observed in some conditions. ⁸⁷ One possible explanation for this discrepancy is that estimates may be exaggerated when age‐related changes in gut dimensions are not also taken into consideration. ⁸⁸ Such changes can produce a decrease in neuron density while not affecting the total number of enteric neurons. ⁸⁹ However, in either scenario, aging creates a mismatch between target tissue and the population of neurons capable of exerting control, ⁹⁰ which could contribute to gastrointestinal dysfunction in older adults.

Age‐related neurodegeneration occurs in both excitatory and inhibitory subsets of enteric neurons but seem to preferentially affect excitatory cells in rats ⁸¹ , ⁹¹ and guinea pigs. ⁹² Nitrergic neurons are comparatively spared during the aging process. ⁸⁰ Whether neuron subtypes display similar vulnerability in aging humans is not clear. Mechanisms involved in age‐related enteric neurodegeneration involve an increase in oxidative stress, inflammation, and an accumulation of unrepaired DNA damage, which will be discussed further below. Interestingly, caloric restriction limits age‐related enteric neuron losses and has the potential to preserve gut function in old age. ⁹³

4. MECHANISMS OF ENTERIC NEURON DEATH

The contexts under which enteric neurons die are seemingly diverse in the examples illustrated above. Despite this, several key mechanisms have emerged that contribute to neuron death in multiple conditions (Figure 1, Table 1). Much of this overlap is due to the underlying inflammatory nature of disease processes and the subsequent effects on the ENS. The following sections provide a brief summary of several mechanisms that play central roles in enteric neuropathies.

FIGURE 1.

Summary of pathophysiological conditions causing enteric neuropathy with the main mechanisms discussed included. Not all mechanisms may be occurring in one cell at the same time.

TABLE 1.

The above table provides an overview of the conditions, mechanisms, and neuron types described in further detail in the main text.

Condition	Potential Mechanisms	Type of neuron affected	Citation
Drug induced	Oxidative stress	Nitrergic neurons	37, 94
Metabolic disorder	Oxidative stress, Toll like receptors	Inhibitory neurons	67, 95
Autoimmune disease	Auto‐antibodies	Non‐specific	74, 76
Aging	Oxidative stress, Inflammation	Excitatory and inhibitory neurons	21, 77, 96
Inflammation	Oxidative stress, Toll like receptors, apoptosis, necroptosis, pyroptosis	Non‐specific	96, 97
Infectious disease	Toll like receptor, pyroptosis	VGLUT2+ neurons, descending interneurons	44, 45, 47, 98, 99, 100

4.1. Oxidative stress

Oxidative stress has received broad attention as a mechanism involved in enteric neuron death and is hypothesized to contribute to nitrergic neuron susceptibility in several contexts. By synthesizing NO, nitrergic neurons exist in a precarious pro‐oxidative balance that is thought to reduce the threshold for reactive oxygen species (ROS) to cause cellular damage during inflammation. ⁹⁶ Increased levels of ROS are prominent during inflammatory responses associated with neuropathies of inflammatory diseases, metabolic disorders, aging, infections, hypoxia, and drugs. ROS bursts include the hydroxyl radical, hydrogen peroxide, hydrochlorous acid, nitric oxide, and peroxynitrite which are all crucial to immune responses. ¹⁰¹ ROS can also produce detrimental effects including cell death when produced in excess through enzymes such as NOX, inducible nitric oxide synthase, and myeloperoxidase. ¹⁶ , ²¹ ROS can cause direct damage to neurons or impact mechanisms in surrounding cells, such as enteric glia, which have detrimental effects on neuron survival during inflammation. ²¹ Hypoxic conditions during IBD may also have an active role in creating a pro‐inflammatory environment leading to increased rates of cell death by inducing acidification and increasing ROS. ¹⁰² Oxygen sensitive prolyl hydroxylases control the activity of HIF1 (hypoxia inducing factors), HIF2, and NF‐kB which coordinate an adaptive response during hypoxic conditions causing stress in the cellular environment. ¹⁰³ ROS may also be elevated when endogenous antioxidant defenses are overwhelmed or defective. Enteric neurons and glia produce elements of antioxidant defense pathways mediated by glutathione and impairing glutathione synthesis produces enteric neuron death in vitro. ²⁰ These observations suggest an ongoing need for neuroprotection against oxidative stress which may be even more critical during disease. This is supported by evidence showing that enteric neuron death is lessened by antioxidants in diabetes, ¹⁰⁴ , ¹⁰⁵ aging, ⁸¹ and in models of chemotherapy induced neuropathy. ⁴⁰ , ⁴¹ As noted above, neuron subtype susceptibility to anticancer drugs is complex and this complexity appears to involve mechanisms of oxidative stress and excitotoxicity. Nitrergic neuron susceptibility in these conditions is induced through muscarinic receptors and subsequent excitatory feedback loop that cause aberrant PKC activity and increase NF‐kB mediated neuroinflammatory activity. ⁹⁴ Antioxidants also have complex roles during inflammation and inhibiting glutathione synthesis in vivo has neuroprotective effects during acute colitis that may involve limiting immune cell infiltration. ²⁰

4.2. Purines

Purines such as ATP are key mediators of inflammation that are generated by immune cells, commensal bacteria, and tissue damage. Increased local extracellular ATP levels are associated with bouts of intestinal inflammation and controlling purine levels reduces gut pathology associated with inflammatory bowel disease and mouse models of colitis. ¹⁰⁶ Some of these benefits are due to limiting enteric neuron death in the myenteric plexus. Enteric neurons are susceptible to death induced by high extracellular purine levels through mechanisms that involve neuronal purinergic P2X₇ receptors. ²² P2X₇ receptors are expressed by enteric neurons in mice, ²² rats, ¹⁸ guinea pigs, ¹⁰⁷ and humans ²² and exhibit unique characteristics among P2 purine receptors including a low affinity for ATP, little desensitization, and the ability to change channel properties in response to the number of binding sites occupied by ATP. ¹⁰⁸ These properties enable sustained activation in the presence of pathologically high concentrations of ATP. P2X₇ receptors are necessary in mechanisms that drive enteric neuron death during inflammation as demonstrated by the neuroprotective effects of eliminating P2X₇ signaling either by using selective antagonists or by ablating the P2rx7 gene. ²² , ²⁴ P2X₇‐mediated enteric neuron death involves intercellular signaling between neurons in the myenteric plexus and the surrounding enteric glia. ²¹ Neuronal pannexin‐1 channels are activated downstream of neuronal P2X₇ receptors and function to release purines that stimulate activity among the surrounding glia. Glia then promote neuron death by releasing transmitters through channels composed of connexin‐43. Interestingly, single cell RNA sequencing data suggest that P2X₇ receptors are enriched in nitrergic neurons, which could explain their susceptibility to death during acute inflammation. ¹⁰⁹ In contrast, pannexin‐1 gene expression is enriched in excitatory neurons. These data suggest an intercellular signaling cascade that involves pannexin‐1‐expressing excitatory neurons such as IPANs that signal to glia, which then promote P2X₇‐dependent cell death in nitrergic neurons.

4.3. Toll like receptors

Toll like receptors (TLRs) are pattern recognition receptors that play critical roles in innate immunity and host responses to microorganisms. TLR‐mediated interactions between the intestinal microbiota and the ENS can be either beneficial or detrimental depending on the context. Enteric neurons and glia express several TLR subtypes including TLR2, TLR3, TLR4, and TLR7 ⁹⁸ , ¹¹⁰ , ¹¹¹ have emerged as particularly important in maintaining enteric neuron survival. Mice lacking either TLR2 or TLR4 exhibit an overall loss of enteric neurons that is primarily driven by a decrease in nitrergic neurons. ¹¹⁰ , ¹¹² Likewise, TLR4 plays an important role in maintaining ENS homeostasis and dysbiosis can produce enteric neuron losses in a TLR4‐dependent manner. ¹¹³ However, exaggerated TLR4 signaling in enteric glia contributes to the pathophysiology of necrotizing enterocolitis, ⁹⁸ possibly by promoting proinflammatory glial signaling. ⁹⁷ TLR4‐mediated host–microbe interactions become particularly relevant in diabetes and obesity when intestinal barrier function is weakened. Diabetes also disrupts the normal proximal to distal gradient of myenteric TLR4 expression from small bowel to colon, ⁹⁵ which could contribute to altered neuron survival by decreasing neuronal NF‐kB activation and increasing apoptosis. ¹¹²

4.4. Modes of neuron death

Cell death primarily occurs through passive and inflammatory mechanisms referred to as necrosis or regulated pathways including apoptosis, necroptosis, and pyroptosis. Apoptosis is generally considered immunologically silent while pyroptosis and necroptosis produce proinflammatory signals that affect the surrounding environment. ¹¹⁴ Enteric neuron death may occur through mechanisms that resemble apoptosis or pyroptosis depending on the context. Inflammation‐driven neuron death in colitis shares some similarities with pyroptosis pathways including ASC activation and pannexin‐1 membrane channels. ²² Alternative pathways of pyroptosis mediated by NLRP6 and caspase 11 are also involved in enteric neuron death during inflammation triggered Salmonella infection. ⁴⁷ Similar caspase‐11 dependent mechanisms contribute to myenteric nitrergic neuron death promoted by consuming a western diet. ¹¹⁵ Necroptosis pathways in enteric neurons are less defined but could contribute to neuron death in response to double stranded DNA derived from cell stress, viruses, or bacteria given their expression of cGAS/STING mechanisms. ¹¹⁶ The relevance of this, and other pathways of cell death is specific diseases will be an important consideration for future work.

5. IS NEURON DEATH AN ISSUE OF SURVIVAL OR REPLACEMENT?

Enteric neuropathy is typically considered from the perspective of pro‐death signaling mechanisms that cause a progressive loss of enteric neurons. Alternatively, one could view neuropathy a defect in neuron replacement given recent evidence suggesting extensive ongoing neurogenesis in the ENS in health ¹¹⁷ and following acute inflammation. ³³ While the extent of enteric neurogenesis at steady state is debated, is it clearly stimulated by perturbations to the gut epithelium and microbiome. Depleting the gut microbiota with antibiotics causes an indiscriminate loss of neurons in the myenteric and submucosal plexuses that is reversed following recolonization by stimulating enteric neurogenesis. ¹¹³ Microbe‐derived factors including lipopolysaccharide and short fatty acid chains appear to play important roles in stimulating enteric neurogenesis as does serotonin acting through 5‐HT4 receptors. ¹¹⁸ Furthermore, ongoing interactions with neurosupportive muscularis macrophages sustain enteric neuron survival in adult mice and disrupting neuron‐macrophage crosstalk results in a loss of enteric neurons. ¹¹⁹ These observations suggest that enteric neuropathy may feature elements of both cell death and defective cell replacement and/or maintenance. Determining the relative contributions of each will be important to consider in future work.

6. CONCLUSIONS

Enteric neuropathy contributes to devastating gut disorders that are difficult to treat. However, new insight into the mechanisms involved could identify potential points of therapeutic intervention. For example, 5HT receptor pathways could protect against enteric neuropathy or stimulate neurogenesis in certain contexts. ¹¹⁸ Likewise, modulating muscularis macrophages could enhance neuron protection in vulnerable states such as GI infections through B2‐AR signaling constraining infection induced inflammation and cell death. ⁴⁷ Antioxidants could offer broad enteric neuroprotection, as seen with honokiol a naturally occurring activator of Sirtuin‐3 that is neuroprotective during palmitate and LPS induced cell death. ⁵⁵ Similar mechanisms may be involved in the beneficial actions of caloric restriction which protects enteric neurons in part by reducing oxidative stress. ⁹³ A deeper understanding the mechanisms involved in enteric neuropathies is needed to identify further, and more effective routes of neuroprotection. Strategies that protect enteric neurons in vulnerable populations and those that stimulate neural repair and neurogenesis could offer a new frontier in treating common gut disorders.

AUTHOR CONTRIBUTIONS

Draft composed by JRJ, edited by BDG, and both authors contributed to synthesizing the final manuscript.

FUNDING INFORMATION

This work was supported by grants R01DK103723 and R01DK120862 to BDG from the National Institute of Diabetes and Digestive and Kidney Diseases of the National Institutes of Health. The content is solely the responsibility of the Authors and does not necessarily represent the official views of the National Institutes of Health.

CONFLICT OF INTEREST STATEMENT

The authors have no financial, professional or personal conflicts that are relevant to the manuscript.

Jamka JR, Gulbransen BD. Mechanisms of enteric neuropathy in diverse contexts of gastrointestinal dysfunction. Neurogastroenterology & Motility. 2025;37:e14870. doi: 10.1111/nmo.14870

DATA AVAILABILITY STATEMENT

Data sharing not applicable to this article as no datasets were generated or analysed during the current study.