Interstitial cells of Cajal and purinergic signalling

doi:10.1016/S1566-0702(02)00005-X

Autonomic Neuroscience

Volume 97, Issue 1, 18 April 2002, Pages 68-72

https://doi.org/10.1016/S1566-0702(02)00005-X Get rights and content

Abstract

Interstitial cells of Cajal (ICC) in the guinea pig intestine, identified by the tyrosine kinase receptor, c-Kit, have been shown with immunohistochemistry to express nucleotide P2X₂ and P2X₅ receptors. P2X₅ receptors have also been demonstrated on interstitial cells in the mouse ileum. It is speculated that release of ATP from enteric nerves, enteric glial cells or from contracting smooth muscle may provide a feedback mechanism for pacemaker activity in the intestine.

Introduction

Interstitial cells of Cajal (ICC) are a special class of cells dispersed in the muscle and nerve plexuses of the gastrointestinal tract Cajal, 1893, Komuro et al., 1996. These cells are of mesenchymal origin and play major roles in gastrointestinal motility (see Ward and Sanders, 2001). They are now well established as the pacemaker cells for the spontaneous contractions of the smooth muscle coats Thuneberg, 1982, Huizinga, 1999. ICC are also responsible for the active propagation of electrical slow waves (Dickens et al., 2001) and have been claimed to mediate motor inputs from enteric nerves (Ward and Sanders, 2001). NK₁ receptors for substance P (Portbury et al., 1996), muscarinic M₂ and M₃ receptors for acetylcholine (Epperson et al., 2000), and somatostatin 2A receptors (Sternini et al., 1997) have been identified on ICCs.

In the present study, we describe the expression of P2X₂ and/or P2X₅ receptors on ICCs, identified with the tyrosine kinase receptor, c-Kit (Maeda et al., 1992), in the guinea pig and mouse ileum and discuss the possibility that release of ATP from enteric nerves, enteric glial cells or during spontaneous contractions of smooth muscle provides a feedback mechanism for pacemaker activity in the gut.

Section snippets

Animals

Breeding, maintenance, and killing of the animals used in this study followed the principles of good laboratory animal care and experimentation in compliance with the UK national law and regulations. Experiments were carried out on male guinea pigs at 2 weeks and 1 year and on mature male mice. Animals were killed by a rising concentration of CO₂ and death was confirmed by cervical dislocation. Subsequently, animals were perfused, through the heart, with 4% formaldehyde (in 0.1 M phosphate

Results

ICCs are classified into four groups based on their morphology and distribution within the different layers of the gastrointestinal tract (Komuro et al., 1996). In this study, we focussed on the type of ICC population that is located at the level of the myenteric plexus (ICC-MP). The ICC-MP branches out to form a network within the plane of the MP that is closely associated with neurons and adjacent smooth muscle cells.

When wholemounts of guinea pig tissue were incubated with the P2X

Discussion

Unambiguous identification of ICC is now possible. In early papers, supravital methylene blue staining was used Cajal, 1893, Thuneberg, 1982. However, this staining was usually incomplete such that uptake of dye was restricted to patches of the ICC network, although a modification was introduced that claimed ‘nearly selective’ staining of the entire ICC network by brief exposure of the intestine to lysolecithin prior to vital methylene blue staining (Mikkelsen et al., 1988). The Champy–Maillet

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Enteric glial cells immunoreactive for P2X7 receptor are affected in the ileum following ischemia and reperfusion
2019, Acta Histochemica
Citation Excerpt :
The results showed that 85% of P2X7 receptor immunoreactive neurons were immunoreactive for Hu, and 16% of the cells positive for the P2X7 receptor are non-neuronal (Hu negative cells). Studies have been demonstrated presence of P2X2 receptor in non-neuronal cells such as intersticial cells of Cajal (ICC) (Burnstock and Lavin, 2002) and P2X7 receptor in the enteric glial cells (Vanderwinden et al., 2003). Given that purinergic P2X7 receptor could be expressed in other cell types, this study investigated the presence of these receptors in EGCs.
The aim of this study was to analyze the effect of ischemia and reperfusion injury (IS) on enteric glial cells (EGCs) and neurons immunoreactive for the P2X7 receptor. Intestinal ischemia was induced by obstructing blood flow in the ileal vessels for 35 min. Afterwards, the vessels were reperfused for 14 days. Tissues were prepared for immunohistochemical labeling of P2X7 receptor, HuC/D (Hu) (pan-neuronal marker) and S100β (glial marker); HuC/D (Hu) and glial fibrillary acidic protein (GFAP, glial marker)/DAPI (nuclear marker); or S100β and GFAP/DAPI. Qualitative and quantitative analyses of colocalization, density, profile area and cell proliferation were performed via fluorescence and confocal laser scanning microscopy. The quantitative analyses revealed that a) neurons and EGCs were immunoreactive for P2X7 receptor; b) the P2X7 receptor immunoreactive cells and Hu immunoreactive neurons were reduced after 0 h and 14 days of reperfusion; c) the S100β and GFAP immunoreactive EGCs were increased; d) the profile area of S100β immunoreactive EGCs was increased by IS; e) few GFAP immunoreactive proliferated at 14 days of reperfusion; f) distinct populations of glial cells can be discerned: S100β⁺/GFAP⁺ cells, S100β⁺/GFAP^– cells and S100β^–/GFAP ⁺ cells; g) histological analysis revealed less alterations in the epithelium cells in the IS groups and h) myeloperoxidase reaction revealed increased of the neutrophils in the lamina propria in the IS groups. This study showed that IS is associated with significant neuronal loss, increase of glial cells and altered purinergic receptor expression and that these changes may contribute to intestinal disorders.
Effects of San-Huang-Xie-Xin-tang, a traditional Chinese prescription for clearing away heat and toxin, on the pacemaker activities of interstitial cells of Cajal from the murine small intestine
2014, Journal of Ethnopharmacology
San-Huang-Xie-Xin-Tang (SHXXT) is a traditional Chinese medicinal formula composed of Coptidis rhizoma (Coptis chinesis Franch), Scutellariae radix (Scutellaria baicalensis Georgi), and Rhei rhizoma (Rheum officinale Baill) and is widely used in Eastern Asia, especially to ameliorate the symptoms of gastrointestinal (GI) disorders related to gastritis, gastric bleeding, peptic ulcers, and abnormal GI motility
Interstitial cells of Cajal (ICCs) are pacemaker cells in the GI tract that generate rhythmic oscillations in membrane potentials known as slow waves. Because GI disorders, especially abnormal GI motility, are major lifelong problems, the authors investigated the effects of SHXXT on mouse small intestine ICCs, and sought to identify the receptors and the action mechanisms involved.
Enzymatic digestions were used to dissociate ICCs from small intestines, and the whole-cell patch-clamp configuration was used to record potentials generated by cultured ICCs.
SHXXT produced membrane depolarization in current-clamp mode, and Y25130 (a 5-HT₃ receptor antagonist) and RS39604 (a 5-HT₄ receptor antagonist) blocked SHXXT-induced membrane depolarizations, whereas SB269970 (a 5-HT₇ receptor antagonist) did not. However, during external Ca²⁺ free conditions or in the presence of thapsigargin, SHXXT did not exhibit membrane depolarization. Furthermore, the application of flufenamic acid (a nonselective cation channel (NSCC) blocker) or DIDS (a chloride channel blocker) abolished pacemaker potential generation and blocked SHXXT-induced membrane depolarizations. In addition, SHXXT-induced membrane depolarizations, which are dependent on G-protein, in ICCs were blocked by PD 98059 (a p42/44 mitogen-activated protein kinase (MAPK) inhibitor), SB203580 (a p38 MAPK inhibitor), and by a c-jun NH2-terminal kinase (JNK) II inhibitor. Regarding the components of SHXXT, Coptidis rhizome and Rhei rhizoma modulated ICC pacemaking activity, whereas Scutellariae radix did not.
SHXXT modulates pacemaker potentials via 5-HT₃ and 5-HT₄ receptor-mediated pathways, external Ca²⁺ influx, and Ca²⁺ release from internal stores. Furthermore, NSCCs and Cl- channels play important roles in the regulation of pacemaking activity in a MAPK dependent manner in ICCs. The regulation of pacemaking activity by SHXXT may be due to the activity of Coptidis rhizome and Rhei rhizome. The study shows SHXXT can modulate the pacemaking activity of ICCs in the GI tract, and thus, suggests SHXXT has potential pharmacological relevance for the treatment of GI motility disorders.
Expression of β<inf>2</inf> adrenoceptors within enteric neurons of the horse ileum
2013, Research in Veterinary Science
Citation Excerpt :
The ICCs regulate visceral smooth muscle activity by producing electrical slow waves, generated by the periodical activation of spontaneous inward currents (pacemaker currents). Several receptors have been identified on ICCs such as receptors for neurokinin 1 and 3 (NK1 and NK3) (Sternini et al., 1995; Grady et al., 1996; Portbury et al., 1996; Vannucchi et al., 1997), somatostatin 2A (Sternini et al., 1997), muscarinic (M2 and M3) (Epperson et al., 2000), VIP1 (Epperson et al., 2000), and purinergic P2X (Burnstock and Lavin, 2002). The evidence of β2-IR on horse ICCs suggests that catecholamines may regulate smooth muscle activity not only on neurons but also on pacemaker cells.
The activity of the gastrointestinal tract is regulated through the activation of adrenergic receptors (ARs). Since data concerning the distribution of ARs in the horse intestine is virtually absent, we investigated the distribution of β₂-AR in the horse ileum using double-immunofluorescence. The β₂-AR-immunoreactivity (IR) was observed in most (95%) neurons located in submucosal plexus (SMP) and in few (8%) neurons of the myenteric plexus (MP). Tyrosine hydroxylase (TH)-IR fibers were observed close to neurons expressing β2-AR-IR. Since β₂-AR is virtually expressed in most neurons located in the horse SMP and in a lower percentage of neurons in the MP, it is reasonable to retain that this adrenergic receptor could regulate the activity of both secretomotor neurons and motor neurons innervating muscle layers and blood vessels. The high density of TH-IR fibers near β₂-AR-IR enteric neurons indicates that the excitability of these cells could be directly modulated by the sympathetic system.
The role of purinergic pathways in the pathophysiology of gut diseases: Pharmacological modulation and potential therapeutic applications
2013, Pharmacology and Therapeutics
Citation Excerpt :
At the same time, these findings deserve further investigations, since the receptor subtypes involved in the regulatory actions of purinergic nucleotides on ICC remain undetermined. Of note, the TTX-insensitive excitatory effect has been postulated to result from P2X2 and P2X5 receptors, which have been found to be expressed in ICC of mouse ileum (Burnstock & Lavin, 2002). Moreover, the neural nitrergic inhibitory component is likely to be regulated by P2X3 receptors, based on the observation that these receptors are expressed on NOS-immunoreactive neurons of murine ileum (van Nassauw et al., 2002).
Gut homeostasis results from complex neuro-immune interactions aimed at triggering stereotypical and specific programs of coordinated mucosal secretion and powerful motor propulsion. A prominent role in the regulation of this highly integrated network, comprising a variety of immune/inflammatory cells and the enteric nervous system, is played by purinergic mediators. The cells of the digestive tract are literally plunged into a “biological sea” of functionally active nucleotides and nucleosides, which carry out the critical task of driving regulatory interventions on cellular functions through the activation of P1 and P2 receptors.
Intensive research efforts are being made to achieve an integrated view of the purinergic system, since it is emerging that the various components of purinergic pathways (i.e., enzymes, transporters, mediators and receptors) are mutually linked entities, deputed to finely modulating the magnitude and the duration of purinergic signaling, and that alterations occurring in this balanced network could be intimately involved in the pathophysiology of several gut disorders. This review article intends to provide a critical appraisal of current knowledge on the purinergic system role in the regulation of gastrointestinal functions, considering these pathways as a whole integrated network, which is capable of finely controlling the levels of bioactive nucleotides and nucleosides in the biophase of their respective receptors. Special attention is paid to the mechanisms through which alterations in the various compartments of the purinergic system could contribute to the pathophysiology of gut disorders, and to the possibility of counteracting such dysfunctions by means of pharmacological interventions on purinergic molecular targets.
Evidence for Ca<sup>2+</sup>-regulated ATP release in gastrointestinal stromal tumors
2013, Experimental Cell Research
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ICC express purinoceptors that may function as a feedback mechanism for pacemaker activity in the intestine under normal physiological conditions [50]. In the tumor microenvironment increased levels of extracellular ATP have been ascribed a potential role for tumor growth, as an important potential source of the immunosuppressive agent adenosine [38,50]. In conclusion, we demonstrate for the first time an experimental system where GIST cells have, like ICCs, a functional intracellular Ca2+ signal transduction pathway.
Gastrointestinal stromal tumors (GISTs) are thought to originate from the electrically active pacemaker cells of the gastrointestinal tract. Despite the presence of synaptic-like vesicles and proteins involved in cell secretion it remains unclear whether GIST cells possess regulated release mechanisms. The GIST tumor cell line GIST882 was used as a model cell system, and stimulus-release coupling was investigated by confocal microscopy of cytoplasmic free Ca²⁺ concentration ([Ca²⁺]_i), flow cytometry, and luminometric measurements of extracellular ATP. We demonstrate that GIST cells have an intact intracellular Ca²⁺-signaling pathway that regulates ATP release. Cell viability and cell membrane integrity was preserved, excluding ATP leakage due to cell death and suggesting active ATP release. The stimulus-secretion signal transduction is at least partly dependent on Ca²⁺ influx since exclusion of extracellular Ca²⁺ diminishes the ATP release. We conclude that measurements of ATP release in GISTs may be a useful tool for dissecting the signal transduction pathway, mapping exocytotic components, and possibly for the development and evaluation of drugs. Additionally, release of ATP from GISTs may have importance for tumor tissue homeostasis and immune surveillance escape.
Roles of substance P and ATP in the subepithelial fibroblasts of rat intestinal villi
2013, International Review of Cell and Molecular Biology
The ingestion of food and water induces chemical and mechanical signals that trigger peristaltic reflexes and also villous movement in the gut. In the intestinal villi, subepithelial fibroblasts under the epithelium form contractile cellular networks and closely contact to the varicosities of substance P and nonsubstance P afferent neurons. Subepithelial fibroblasts of the duodenal villi possess purinergic receptor P2Y₁ and tachykinin receptor NK1. ATP and substance P induce increase in intracellular Ca^2 + and cell contraction in subepithelial fibroblasts. They are highly mechanosensitive and release ATP by mechanical stimuli. Released ATP spreads to form an ATP “cloud” with nearly 1 μM concentration and activates the surroundings via P2Y₁ and afferent neurons via P2X receptors. These findings suggest that villous subepithelial fibroblasts and afferent neurons interact via ATP and substance P. This mutual interaction may play important roles in the signal transduction of mechano reflex pathways including a coordinate villous movement and also in the maturation of the structure and function of the intestinal villi.

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Short communicationInterstitial cells of Cajal and purinergic signalling

Abstract

Introduction

Section snippets

Animals

Results

Discussion

J. Biol. Chem.

Neuroscience

J. Auton. Nerv. Syst.

Prog. Neurobiol.

J. Urol.

J. Biol. Chem.

Neurosci. Lett.

ATP in brain function

Adrenergic and possible nonadrenergic sources of adenosine 5′-triphosphate release from nerve varicosities isolated from ileal myenteric plexus

J. Pharmacol. Exp. Ther.

Interstitial cells of Cajal in the guinea-pig gastrointestinal tract as revealed by c-Kit immunohistochemistry

Cell Tissue Res.

Co-transmission. The fifth Heymans memorial lecture—Ghent, February 17, 1990

Arch. Int. Pharmacodyn. Ther.

Sur les gangliones et plexus nerveux de l'intestin

C. R. Soc. Biol. Paris

Selective knockout of intramuscular interstitial cells reveals their role in the generation of slow waves in mouse stomach

J. Physiol.

Molecular markers expressed in cultured and freshly isolated interstitial cells of Cajal

Am. J. Physiol. Cell Physiol.

Localisation of P2X5 and P2X7 receptors by immunohistochemistry in rat stratified squamous epithelia

Cell Tissue Res.

P2X receptors in the rat duodenal villus

Cell Tissue Res.

Short communication
Interstitial cells of Cajal and purinergic signalling

Localisation of P2X₅ and P2X₇ receptors by immunohistochemistry in rat stratified squamous epithelia