The Mechanism and Spread of Pacemaker Activity Through Myenteric Interstitial Cells of Cajal in Human Small Intestine

doi:10.1053/j.gastro.2007.02.049

Gastroenterology

Volume 132, Issue 5, May 2007, Pages 1852-1865

https://doi.org/10.1053/j.gastro.2007.02.049 Get rights and content

Background & Aims: It has been generally assumed that interstitial cells of Cajal (ICC) in the human gastrointestinal tract have similar functions to those in rodents, but no direct experimental evidence exists to date for this assumption. This is an important question because pathologists have noted decreased numbers of ICC in patients with a variety of motility disorders, and some have speculated that loss of ICC could be responsible for motor dysfunction. Our aims were to determine whether myenteric ICC (ICC-MY) in human jejunum are pacemaker cells and whether these cells actively propagate pacemaker activity. Methods: The mucosa and submucosa were removed, and strips of longitudinal muscle were peeled away to reveal the ICC-MY network. ICC networks were loaded with the Ca²⁺ indicator fluo-4, and pacemaker activity was recorded via high-speed video imaging at 36.5°C ± 0.5°C. Results: Rhythmic, biphasic Ca²⁺ transients (6.03 ± 0.33 cycles/min) occurred in Kit-positive ICC-MY. These consisted of a rapidly propagating upstroke phase that initiated a sustained plateau phase, which was associated with Ca²⁺ spikes in neighboring smooth muscle. Pacemaker activity was dependent on inositol 1,4,5-triphosphate receptor–operated stores and mitochondrial function. The upstroke phase of Ca²⁺ transients in ICC-MY appeared to result from Ca²⁺ influx through dihydropyridine-resistant Ca²⁺ channels, whereas the plateau phase was attributed to Ca²⁺ release from inositol 1,4,5-triphosphate receptor–operated Ca²⁺ stores. Conclusions: Each ICC-MY in human jejunum generates spontaneous pacemaker activity that actively propagates through the ICC network. Loss of these cells could severely disrupt the normal function of the human small intestine.

Section snippets

Human Tissue Preparation

The segments of human small intestine used in this study were obtained from the jejunum of morbid obese patients of either sex ranging in age from 26 to 58 years as surgical waste tissues during gastric bypass operations performed for morbid obesity.¹¹ The protocol was approved by the human subjects research committees at the University of Nevada and the University of California Davis. A segment of jejunum was opened along the mesenteric border, and the mucosa and submucosa were removed by

Morphologic Studies

Using Kit immunohistochemistry, we found that the ICC-MY network in the human jejunum had a complex 3-dimensional structure (Figure 1A–C and G–I). Bundles of ICC-MY were observed to run parallel both to the LM and to the CM (Figure 1A). These orthogonal bundles of ICC-MY were connected by obliquely running bundles or strands of ICC (Figure 1B, C, H, and I). In cross section, ICC-MY lined myenteric ganglia and formed a continuous network that extended down as septal ICC lining muscle bundles (

Discussion

To date, the role of ICC in human GI muscles has been speculative and based on studies of this class of cells in rodents because functional studies of human ICC have not been reported previously. Our data suggest that the mechanisms of generation and propagation of Ca²⁺ transients in human ICC-MY networks are similar to those in the murine small intestine⁶ and antrum.⁸ Imaging of multicellular networks of ICC-MY has many important advantages over electrophysiological recordings. Imaging can

References (28)

A.J. Bauer et al.
Opioids inhibit neuromuscular transmission in circular muscle of human and baboon jejunum
Gastroenterology
(1991)
M.E. Stark et al.
Nitric oxide mediates inhibitory nerve input in human and canine jejunum
Gastroenterology
(1993)
K.M. Sanders et al.
Organization and electrophysiology of interstitial cells of Cajal and smooth muscle cells in the gastrointestinal tract
J.J. Rumessen et al.
Interstitial cells in the musculature of the gastrointestinal tract: Cajal and beyond
Int Rev Cytol
(2003)
R.J. Stevens et al.
Propagation and neural regulation of calcium waves in circular and longitudinal muscle layers of guinea-pig small intestine
Gastroenterology
(2000)
S. Torihashi et al.
c-Kit immunoreactive interstitial cells in the human gastrointestinal tract
J Auton Nerv Syst
(1999)
K.M. Sanders et al.
Interstitial cells of Cajal as pacemakers in the gastrointestinal tract
Annu Rev Physiol
(2006)
Y. Hara et al.
Electrophysiology of smooth muscle of the small intestine of some mammals
J Physiol
(1986)
T.K. Smith et al.
Interaction of two electrical pacemakers in muscularis of canine proximal colon
Am J Physiol
(1987)
W.J.E.P. Lammers
Spatial and temporal coupling between slow waves and pendular contractions
Am J Physiol
(2005)

A. Bortoff

Electrical transmission of slow waves from longitudinal to circular intestinal muscle

Am J Physiol

(1965)

K.J. Park et al.

Spatial and temporal mapping of pacemaker activity in interstitial cells of Cajal in mouse ileum in situ

Am J Physiol

(2006)

F.R. Edwards et al.

An electrical description of the generation of slow waves in the antrum of the guinea pig

J Physiol

(2005)

G.W. Hennig et al.

Propagation of pacemaker activity in the guinea-pig antrum

J Physiol

(2004)

Cited by (88)

A multi-parameter approach to measurement of spontaneous myogenic contractions in human stomach: Utilization to assess potential modulators of myogenic contractions
2022, Pharmacological Research
Citation Excerpt :
Nevertheless, in mouse intestine similar concentrations did not inhibit myogenic contractions under conditions in which tissue penetration was proven [31] and did not reduce the occurrence of slow waves in this tissue [24] or in guinea-pig ileum [32]. Inhibition of SERCA pumps by 2-APB and CPA may explain the progressive abolition of myogenic contractions of the human stomach, also reported with similar concentrations in mouse intestine [24,31,33], an activity consistent with abolition of electrical slow waves in human [29] and mouse intestine [24,31]. In the human stomach 2-APB also initially caused a short-lived increase in muscle tone, and CPA markedly increased muscle tone and myogenic contraction amplitude prior to inhibition of myogenic contractions.
Electrical slow waves, generated by interstitial cells of Cajal (ICC), cause spontaneous contractions of human stomach. Software was developed to measure muscle tone and eleven different parameters defining these contractions in human stomach, displaying data as radar plots. A pilot study assessed the effects of potential modulators, selected from among compounds known to influence ICC activity; n = 4–7 each concentration tested/compound. Human distal stomach (corpus-antrum) muscle strips were suspended in tissue baths for measuring myogenic (non-neuronal) contractions in the presence of tetrodotoxin (10^-6 M). Initial characterization: Contractions (amplitude 4 ± 0.4mN, frequency 3 ± 0.1 min^-1, n = 49) were unchanged by ꭃ-conotoxin GVIA (10^-7 M) or indomethacin (10^-6 M) but abolished by nifedipine (10^-4 M). Carbachol (10^-7 M) increased contraction rate and amplitude; 10^-6-10^-5 M increased tone and caused large, irregular contractions. [Ca²⁺]_i modulators: Ryanodine (10^-5-10^-4 M) increased muscle tone accompanied by inhibition of myogenic contractions. Xestospongin-C (10^-6 M; IP₃ channel inhibitor) had no effects. SERCA pump inhibitors, 2-APB and cycloplazonic acid (10^-5-10^-4 M) increased tone and myogenic contraction amplitude before abolishing contractions; thapsigargin was weakly active. CaCC blockers: MONNA and CaCCinh-A01 had little-or-no effects on tone but reduced myogenic contractions; MONNA (10^-4 M) was more effective, reducing amplitude (77.8 ± 15.2%) and frequency. Ca_V3.1/3.2/3.3 channel block: Mibefradil reduced tone and myogenic contraction amplitude (pIC₅₀ 4.8 ± 0.9). Inward-rectifying K⁺-channel inhibitor: E-4031 (10^-4 M) increased contraction duration (17.4 ± 5.8%). Conclusions: (1) Measurement of multiple parameters of myogenic contractions identified subtle differences between compounds, (2) only E-4031 and CaCC blockers influenced myogenic contractions, not muscle tone, (3) studies are needed with compounds with known and/or improved selectivity/potency for human targets affecting ICC functions.
Ca<sup>2+</sup> transients in ICC-MY define the basis for the dominance of the corpus in gastric pacemaking
2021, Cell Calcium
Myenteric interstitial cells of Cajal (ICC-MY) generate and actively propagate electrical slow waves in the stomach. Slow wave generation and propagation are altered in gastric motor disorders, such as gastroparesis, and the mechanism for the gradient in slow wave frequency that facilitates proximal to distal propagation of slow waves and normal gastric peristalsis is poorly understood. Slow waves depend upon Ca²⁺-activated Cl⁻ channels (encoded by Ano1). We characterized Ca²⁺ signaling in ICC-MY in situ using mice engineered to have cell-specific expression of GCaMP6f in ICC. Ca²⁺ signaling differed in ICC-MY in corpus and antrum. Localized Ca²⁺ transients were generated from multiple firing sites and were organized into Ca²⁺ transient clusters (CTCs). Ca²⁺ transient refractory periods occurred upon cessation of CTCs, but a relatively higher frequency of Ca²⁺ transients persisted during the inter-CTC interval in corpus than in antrum ICC-MY. The onset of Ca²⁺ transients after the refractory period was associated with initiation of the next CTC. Thus, CTCs were initiated at higher frequencies in corpus than in antrum ICC-MY. Initiation and propagation of CTCs (and electrical slow waves) depends upon T-type Ca²⁺ channels, and durations of CTCs relied upon L-type Ca²⁺ channels. The durations of CTCs mirrored the durations of slow waves. CTCs and Ca²⁺ transients between CTCs resulted from release of Ca²⁺ from intracellular stores and were maintained, in part, by store-operated Ca²⁺ entry. Our data suggest that Ca²⁺ release and activation of Ano1 channels both initiate and contribute to the plateau phase of slow waves.
Interactions between the microbiota and enteric nervous system during gut-brain disorders
2021, Neuropharmacology
For the last 20 years, researchers have focused their intention on the impact of gut microbiota in healthy and pathological conditions. This year (2021), more than 25,000 articles can be retrieved from PubMed with the keywords “gut microbiota and physiology”, showing the constant progress and impact of gut microbes in scientific life. As a result, numerous therapeutic perspectives have been proposed to modulate the gut microbiota composition and/or bioactive factors released from microbes to restore our body functions. Currently, the gut is considered a primary site for the development of pathologies that modify brain functions such as neurodegenerative (Parkinson's, Alzheimer's, etc.) and metabolic (type 2 diabetes, obesity, etc.) disorders. Deciphering the mode of interaction between microbiota and the brain is a real original option to prevent (and maybe treat in the future) the establishment of gut-brain pathologies. The objective of this review is to describe recent scientific elements that explore the communication between gut microbiota and the brain by focusing our interest on the enteric nervous system (ENS) as an intermediate partner. The ENS, which is known as the “second brain”, could be under the direct or indirect influence of the gut microbiota and its released factors (short-chain fatty acids, neurotransmitters, gaseous factors, etc.). Thus, in addition to their actions on tissue (adipose tissue, liver, brain, etc.), microbes can have an impact on local ENS activity. This potential modification of ENS function has global repercussions in the whole body via the gut-brain axis and represents a new therapeutic strategy.
This article is part of the special Issue on ‘Cross Talk between Periphery and the Brain’.
A high throughput machine-learning driven analysis of Ca<sup>2+</sup> spatio-temporal maps
2020, Cell Calcium
High-resolution Ca²⁺ imaging to study cellular Ca²⁺ behaviors has led to the creation of large datasets with a profound need for standardized and accurate analysis. To analyze these datasets, spatio-temporal maps (STMaps) that allow for 2D visualization of Ca²⁺ signals as a function of time and space are often used. Methods of STMap analysis rely on a highly arduous process of user defined segmentation and event-based data retrieval. These methods are often time consuming, lack accuracy, and are extremely variable between users. We designed a novel automated machine-learning based plugin for the analysis of Ca²⁺ STMaps (STMapAuto). The plugin includes optimized tools for Ca²⁺ signal preprocessing, automated segmentation, and automated extraction of key Ca²⁺ event information such as duration, spatial spread, frequency, propagation angle, and intensity in a variety of cell types including the Interstitial cells of Cajal (ICC). The plugin is fully implemented in Fiji and able to accurately detect and expeditiously quantify Ca²⁺ transient parameters from ICC. The plugin’s speed of analysis of large-datasets was 197-fold faster than the commonly used single pixel-line method of analysis. The automated machine-learning based plugin described dramatically reduces opportunities for user error and provides a consistent method to allow high-throughput analysis of STMap datasets.
Antroduodenal manometry for the evaluation of patients with suspected gastroparesis
2020, Gastroparesis: Pathophysiology, Clinical Presentation, Diagnosis and Treatment
The symptom complex of chronic nausea, vomiting and often abdominal pain is labeled gastroparesis. Many of these patients, but not all, have an associated delay in gastric emptying. The symptoms and the delay in gastric emptying may be associated with disturbed contraction pattern in the upper gut including the stomach, pylorus and duodenum. Antroduodenal manometry allows categorization of these abnormalities leading to targeted therapy. The abnormal motility may be localized or a reflection of disturbed motility throughout the gastrointestinal tract as in pseudoobstruction. Antroduodenal manometry is classically performed by placing a long tube into the intestines, but the development of a wireless motility capsule provides an alternative less invasive method. Administration of a macrolide or octreotide stimulates contractions specifically in the stomach or the intestine providing directed therapy. Elevated pyloric pressure or decreased pyloric compliance can be treated with endoscopic pyloromyotomy. Antroduodenal manometry has provided the information for more directed therapy of patients with gastroparesis.
Norepinephrine Has Dual Effects on Human Colonic Contractions Through Distinct Subtypes of Alpha 1 Adrenoceptors
2020, Cellular and Molecular Gastroenterology and Hepatology
Colonic musculature contain smooth muscle cells (SMC), interstitial cells of Cajal (ICC), and platelet-derived growth factor receptor α⁺ cells (PDGFRα⁺ cells), which are electrically coupled and operate together as the SIP syncytium. PDGFRα⁺ cells have enriched expression of small conductance Ca²⁺-activated K⁺ (SK) channels. Purinergic enteric neural input activates SK channels in PDGFRα⁺ cells, hyperpolarizes SMC, and inhibits colonic contractions. Recently we discovered that PDGFRα⁺ cells in mouse colon have enriched expression of α1A adrenoceptors (ARs), which coupled to activation of SK channels and inhibited colonic motility, and α1A ARs were principal targets for sympathetic regulation of colonic motility. Here we investigated whether PDGFRα⁺ cells in human colon express α1A ARs and share the roles as targets for sympathetic regulation of colonic motility.
Isometric tension recording, intracellular recording, and Ca²⁺ imaging were performed on muscles of the human colon. Responses to α1 ARs agonists or electric field stimulation with AR antagonists and neuroleptic reagents were studied.
Exogenous or endogenous norepinephrine released from nerve fibers inhibited colonic contractions through binding to α1A ARs or enhanced colonic contractions by acting on α1D ARs. Inhibitory responses were blocked by apamin, an antagonist of SK channels. Phenylephrine, α1 AR agonists, or norepinephrine increased intracellular [Ca²⁺] in PDGFRα⁺ cells, but not in ICC, and hyperpolarized SMCs by binding to α1 ARs expressed by PDGFRα⁺ cells.
Human colonic contractions are inhibited by α1A ARs expressed in PDGFRα⁺ cells and activated by α1D ARs expressed in SMC.

View all citing articles on Scopus

: Supported by National Institute of Diabetes and Digestive and Kidney Diseases grant RO1 DK45713 (to T.K.S.) and program project grant DK41315 (to K.M.S.). Imaging was performed in a Core laboratory supported by P20 RR-18751.

: The authors have no conflicts of interest to disclose.

¹: H.-T.L. and G.W.H. contributed equally to this work. H.-T.L. performed most of the experimental work, and G.W.H. developed the algorithms for data analysis.

View full text

Basic–alimentary tractThe Mechanism and Spread of Pacemaker Activity Through Myenteric Interstitial Cells of Cajal in Human Small Intestine

Section snippets

Human Tissue Preparation

Morphologic Studies

Discussion

Gastroenterology

Gastroenterology

Int Rev Cytol

Gastroenterology

J Auton Nerv Syst

Interstitial cells of Cajal as pacemakers in the gastrointestinal tract

Annu Rev Physiol

Electrophysiology of smooth muscle of the small intestine of some mammals

J Physiol

Interaction of two electrical pacemakers in muscularis of canine proximal colon

Am J Physiol

Spatial and temporal coupling between slow waves and pendular contractions

Am J Physiol

Electrical transmission of slow waves from longitudinal to circular intestinal muscle

Am J Physiol

Spatial and temporal mapping of pacemaker activity in interstitial cells of Cajal in mouse ileum in situ

Am J Physiol

An electrical description of the generation of slow waves in the antrum of the guinea pig

J Physiol

Propagation of pacemaker activity in the guinea-pig antrum

J Physiol

Basic–alimentary tract
The Mechanism and Spread of Pacemaker Activity Through Myenteric Interstitial Cells of Cajal in Human Small Intestine