Intestinal Peristaltic Reflex Research Articles

Recreating Intestinal Peristalsis in the Petri Dish.

Here we describe a culture technique of cells dissociated from the external muscularis of the guinea pig small intestine, which allows us to maintain all the elements involved in the intestinal peristaltic reflex. After a few days in culture, these cells reorganize to form a small group of cells that permit the generation of pacemaker activity, spontaneous contractions, and the development of inhibitory and excitatory junction potentials in the petri dish, all elements involved in the peristaltic reflex. Therefore, these co-cultures are suitable to study the cellular and molecular aspects related to the development, maintenance, and modulation of motor intestinal functions.

The endocannabinoid anandamide regulates the peristaltic reflex by reducing neuro-neuronal and neuro-muscular neurotransmission in ascending myenteric reflex pathways in rats

BackgroundEndocannabinoids (EC) and the cannabinoid-1 (CB1) receptor are involved in the regulation of motility in the gastrointestinal (GI) tract. However, the underlying physiological mechanisms are not completely resolved. The purpose of this work was to study the physiological influence of the endocannabinoid anandamide, the putative endogenous CB1 active cannabinoid, and of the CB1 receptor on ascending peristaltic activity and to identify the involved neuro-neuronal, neuro-muscular and electrophysiological mechanisms. MethodsThe effects of anandamide and the CB1 receptor antagonist SR141716A were investigated on contractions of the circular smooth muscle of rat ileum and in longitudinal rat ileum segments where the ascending myenteric part of the peristaltic reflex was studied in a newly designed organ bath. Additionally intracellular recordings were performed in ileum and colon. ResultsAnandamide significantly reduced cholinergic twitch contractions of ileum smooth muscle whereas SR141716A caused an increase. Anandamide reduced the ascending peristaltic contraction by affecting neuro-neuronal and neuro-muscular neurotransmission. SR141716A showed opposite effects and all anandamide effects were antagonized by SR141716A (1μM). Anandamide reduced excitatory junction potentials (EJP) and inhibitory junction potentials (IJP), whereas intestinal slow waves were not affected. ConclusionsCB1 receptors regulate force and timing of the intestinal peristaltic reflex and these actions involve interneurons and motor-neurons. The endogenous cannabinoid anandamide mediates these effects by activation of CB1 receptors. The endogenous cannabinoid system is permanently active, suggesting the CB1 receptor being a possible target for the treatment of motility related disorders.

Agonist-biased Trafficking of Somatostatin Receptor 2A in Enteric Neurons

Somatostatin (SST) 14 and SST 28 activate somatostatin 2A receptors (SSTR2A) on enteric neurons to control gut functions. SST analogs are treatments of neuroendocrine and bleeding disorders, cancer, and diarrhea, with gastrointestinal side effects of constipation, abdominal pain, and nausea. How endogenous agonists and drugs differentially regulate neuronal SSTR2A is unexplored. We evaluated SSTR2A trafficking in murine myenteric neurons and neuroendocrine AtT-20 cells by microscopy and determined whether agonist degradation by endosomal endothelin-converting enzyme 1 (ECE-1) controls SSTR2A trafficking and association with β-arrestins, key regulators of receptors. SST-14, SST-28, and peptide analogs (octreotide, lanreotide, and vapreotide) stimulated clathrin- and dynamin-mediated internalization of SSTR2A, which colocalized with ECE-1 in endosomes and the Golgi. After incubation with SST-14, SSTR2A recycled to the plasma membrane, which required active ECE-1 and an intact Golgi. SSTR2A activated by SST-28, octreotide, lanreotide, or vapreotide was retained within the Golgi and did not recycle. Although ECE-1 rapidly degraded SST-14, SST-28 was resistant to degradation, and ECE-1 did not degrade SST analogs. SST-14 and SST-28 induced transient interactions between SSTR2A and β-arrestins that were stabilized by an ECE-1 inhibitor. Octreotide induced sustained SSTR2A/β-arrestin interactions that were not regulated by ECE-1. Thus, when activated by SST-14, SSTR2A internalizes and recycles via the Golgi, which requires ECE-1 degradation of SST-14 and receptor dissociation from β-arrestins. After activation by ECE-1-resistant SST-28 and analogs, SSTR2A remains in endosomes because of sustained β-arrestin interactions. Therapeutic SST analogs are ECE-1-resistant and retain SSTR2A in endosomes, which may explain their long-lasting actions.

A NEW ELECTROPHYSIOLOGICAL TOOL TO INVESTIGATE THE SPATIAL NEURONAL PROJECTIONS WITHIN THE MYENTERIC ASCENDING REFLEX OF THE MOUSE COLON

1. The intestinal peristaltic reflex is regulated by local microcircuits that, upon activation, result in an oral contraction and anal relaxation of the circular muscle. This contractile response is associated with typical electrophysiological changes in membrane potential resulting from excitatory and inhibitory myenteric pathways. 2. The aim of the present study was to investigate the influence of local electrical stimulation (ES; single pulses, 15 V, 0.3 msec duration) on the ascending gastrointestinal electrophysiological potentials of the mouse colon using a novel 12-channel stimulation electrode in a newly designed model of the ascending myenteric pathways with simultaneous intracellular recording. 3. Local myenteric reflex responses in the proximal colon were initiated by ES (12 bipolar stimulation electrodes (SE) 0.7 mm apart from each other) and excitatory and inhibitory junction potentials (EJP and IJP, respectively) were recorded from circular smooth muscle cells with intracellular recording techniques. In vivo colonic propulsion was determined by measuring the time to expulsion of a 3 mm glass bead inserted 2.5 cm into the distal colon of mouse. 4. Under basal conditions, circular smooth muscle cells displayed a stable membrane potential (-56.7 +/- 6.9 mV; n = 13). Electrical stimulation elicited a tetrodotoxin (3 micromol/L)-sensitive, neuronal-induced EJP (cholinergic; atropine (1 micromol/L) sensitive) and a biphasic IJP. Both the EJP and IJP showed characteristic responses dependent on the distance between stimulation and recording sites. The EJP could be recorded over long distances, resulting in a maximal EJP amplitude at a distance of 10 mm distance (represented by stimulation electrodes (SE) number 6/7) and a maximal projection distance of 18-20 mm. Both components of IJP were maximal during direct stimulation (at SE1; stimulation at the recording site) and gradually decreased to SE6/7 (10 mm). At distances greater than 10 mm apart, ES did not produce IJP. The ganglionic blocker hexamethonium (10-100 micromol/L) concentration dependently abolished all inhibitory junction potentials at distances greater than 10 mm and significantly reduced the amplitude of EJP for the first 10 mm. Colonic propulsion was decreased by hexamethonium (40 mg/kg) and atropine (0.7 mg/kg). 5. Neuronal circuits of the ascending myenteric reflex functionally project distances ranging up to 18-20 mm. Our newly designed setup allows simultaneous electrophysiological investigations of neuronal microcircuitry within the myenteric plexus over short and long distances and enables conclusions to be drawn regarding neuroneuronal and neuromuscular transmission.

ORL-1 Receptor Mediates the Action of Nociceptin on Ascending Myenteric Reflex Pathways in Rats

Nociceptin is the endogenous agonist of the "orphan" opioid receptor-1 (ORL-1). We investigated whether activation of the ORL-1 receptor influences smooth muscle contractility and enteric neurotransmission within ascending myenteric reflex pathways of rats. Reverse transcriptase polymerase chain reaction was performed to evaluate the presence of ORL-1 receptors. The ascending part of the ascending myenteric reflex in rats was studied in ileal segments using a 3-chambered organ bath. Intracellular recordings were performed to evaluate pharmacologic effects on excitatory and inhibitory junction potentials (EJP; IJP). Single- and double-labeling immunohistochemistry was used to examine the distribution of ORL-1 within the intestinal wall. ORL-1 expression and immunoreactivity was found in the large majority of myenteric neurons. In addition to the cholinergic myenteric neurons, all nitrergic myenteric neurons expressed the ORL-1 receptor. Nociceptin significantly reduced cholinergic twitch contractions, an effect that was reversed by the ORL-1 receptor antagonist [Nphe(1)]nociceptin(1-13)NH(2). Neither nociceptin nor [Nphe(1)]nociceptin(1-13)NH(2) had a direct influence on smooth muscle contractility. Nociceptin significantly reduced ascending myenteric reflex contractions and prolonged the latency from stimulation to contraction. Both effects were antagonized by [Nphe(1)]nociceptin(1-13)NH(2). Intracellular recordings demonstrated that nociceptin reduces the cholinergically mediated EJP and the nitrergic phase of IJP in a concentration-dependent manner, effects that were reversible in presence of [Nphe(1)]nociceptin(1-13)NH(2). We conclude that activation of ORL-1 receptors on myenteric neurons reduce excitatory and inhibitory neurotransmission within the gastrointestinal tract. This is accompanied by a reduction of the small intestinal peristaltic reflex response. These effects might be used pharmacologically.

Intestinal Peristaltic Reflex (IPR) is Regulated by Mast Cells and IL-9

Intestinal Peristaltic Reflex (IPR) is Regulated by Mast Cells and IL-9

Desensitization of the peristaltic reflex induced by mucosal stimulation with the selective 5-HT4agonist tegaserod

The intestinal peristaltic reflex induced by mucosal stimulation is mediated by mucosal release of serotonin (5-HT), which acts on 5-HT(4) receptors located on CGRP-containing afferent nerve terminals. Exposure of the colonic mucosa to the 5-HT(4) receptor agonist tegaserod in the range of 1 nM to 10 muM elicits a peristaltic reflex and stimulates colonic propulsion. The present study was designed to identify the 5-HT(4) receptor subtype mediating the reflex and determine whether functionally effective concentrations of tegaserod desensitize the reflex induced by mucosal stimulation. Exposure of rat colonic mucosa to tegaserod in the range of 5 nM to 5 muM for 5 or 10 min caused rapid time- and concentration-dependent desensitization of the peristaltic reflex induced by mucosal stroking, consistent with the operation of a rapidly desensitizing 5-HT(4b) receptor subtype. Desensitization was accompanied by a decrease in CGRP release. The rate of recovery of peristaltic response depended on the desensitizing concentration of tegaserod: ascending contraction and descending relaxation recovered within 15 min after 5-50 nM tegaserod, 30 min after 0.5 muM, and 60 min after 5 muM. Neither CGRP release nor the peristaltic reflex induced by muscle stretch was affected by 5-HT(4) receptor desensitization, providing further evidence that 5-HT does not mediate the reflex induced by muscle stretch. These results suggest in cases of increased 5-HT availability or prolonged exposure, such as colitis, that it is likely the peristaltic reflex will be blunted.

Gastrin-releasing peptide is a modulatory neurotransmitter of the descending phase of the peristaltic reflex.

The physiological role of gastrin-releasing peptide (GRP) and of its cognate receptors in regulating the intestinal peristaltic reflex was examined in a three-compartment flat-sheet preparation of rat colon. Mucosal stimulation applied to the central compartment at high, but not low levels of intensity, induced GRP release in the caudad compartment where descending relaxation was measured, but not into the ascending compartment where ascending contraction was measured or into the central compartment where the stimuli were applied. The selective GRP (BB(2)) receptor antagonist, [D-Phe(6),des-Met(14)]bombesin(6-14), inhibited descending relaxation and VIP release in the caudad compartment induced by high but not by low levels of stimulation applied to the mucosa in the central compartment. The selective neuromedin B (BB(1)) receptor antagonist, BIM-23127, had no effect on descending relaxation or VIP release. Neither the BB(1) nor the BB(2) antagonist had any effect on ascending contraction or substance P release in the orad compartment. Consistent with the effects of the antagonists on the peristaltic reflex, the BB(2) antagonist but not the BB(1) antagonist decreased the velocity of propulsion of artificial fecal pellets through isolated segments of guinea pig distal colon. The results indicate that GRP is selectively released from myenteric neurons in descending pathways during the peristaltic reflex and that it acts via BB(2) receptors to augment the descending phase of the peristaltic reflex and propulsion.

Physiological role of neuropeptide Y in the regulation of the ascending phase of the peristaltic reflex.

The physiological role of neuropeptide Y (NPY) and of specific NPY receptors in regulating the intestinal peristaltic reflex was examined in three-compartment flat-sheet preparations of rat colon. Graded muscle stretch or mucosal stimulation applied to the central compartment inhibited NPY release in the orad compartment where ascending contraction was measured. NPY and the Y1-receptor agonist [Leu31, Pro34]NPY inhibited, whereas the selective Y1-receptor antagonist BIBP 3226 augmented ascending contraction and substance P (SP) release in the orad compartment induced by muscle stretch or mucosal stimulation. Neither agonist nor antagonist had any effect on descending relaxation or VIP release in the caudad compartment. The Y2-receptor agonist NPY13-36 and antagonist BIIE 0246 had no effect on peptide release or mechanical response. The results indicate that suppression of a tonic inhibitory influence of NPY neurons on excitatory neurotransmitter release contributes substantially to the orad contractile phase of the peristaltic reflex. The effect of NPY on neurotransmitter release is mediated by Y1 receptors.

Reciprocal activity of longitudinal and circular muscle during intestinal peristaltic reflex.

A two-compartment, flat-sheet preparation of rat colon was devised, which enabled exclusive measurement of longitudinal muscle activity during the ascending and descending phases of the peristaltic reflex. A previous study using longitudinal muscle strips revealed the operation of an integrated neuronal circuit consisting of somatostatin, opioid, and VIP/pituitary adenylate cyclase-activating peptide (PACAP)/nitric oxide synthase (NOS) interneurons coupled to cholinergic/tachykinin motor neurons innervating longitudinal muscle strips that could lead to descending contraction and ascending relaxation of this muscle layer. Previous studies in peristaltic preparations have also shown that an increase in somatostatin release during the descending phase causes a decrease in Met-enkephalin release and suppression of the inhibitory effect of Met-enkephalin on VIP/PACAP/NOS motor neurons innervating circular muscle and a distinct set of VIP/PACAP/NOS interneurons. The present study showed that in contrast to circular muscle, longitudinal muscle contracted during the descending phase and relaxed during the ascending phase. Somatostatin antiserum inhibited descending contraction and augmented ascending relaxation of longitudinal muscle, whereas naloxone had the opposite effect. VIP and PACAP antagonists inhibited descending contraction of longitudinal muscle and augmented ascending relaxation. Atropine and tachykinin antagonists inhibited descending contraction of longitudinal muscle. As shown in earlier studies, the same antagonists and antisera produced opposite effects on circular muscle. We conclude that longitudinal muscle contracts and relaxes in reverse fashion to circular muscle during the peristaltic reflex. Longitudinal muscle activity is regulated by excitatory VIP/PACAP/NOS interneurons coupled to cholinergic/tachykinin motor neurons innervating longitudinal muscle.

Fundic balloon distension stimulates antral and duodenal motility in man.

Distension of the intestine triggers the peristaltic reflex, which consists of orad contraction and aborad relaxation. Whether a similar response occurs in the human stomach is unclear. Our aim was to investigate the antral and duodenal motor response(s) to mechanical distension of the proximal stomach. In six healthy volunteers, a large compliant balloon was placed in the proximal stomach. Alongside this a water-perfused manometry probe with six sensors was placed to measure the antral and duodenal motility. Pressure activity was assessed before and during balloon distension. In five of six subjects, balloon distension triggered a salvo of antral pressure waves within 3-5 min, some of which propagated into the duodenum. The amplitude of waves was higher (P < 0.05) at the antrum than at the duodenum. The area under the curve of pressure waves was higher (P < 0.05) at the antrum than at the duodenum. In conclusion, distension of the proximal stomach, at or below the threshold for perception, evokes phasic motor activity in the antrum and duodenum. Thus, the gastric response to distension differs from that observed during the intestinal peristaltic reflex.

Increased defecation during stress or after 5-hydroxytryptophan: selective inhibition by the 5-HT(4) receptor antagonist, SB-207266.

5-HT(4) receptor antagonism prevents the ability of exogenous 5-HT or 5-HTP to sensitize the intestinal peristaltic reflex and increase the rate of defecation, generally without affecting non-stimulated intestinal function. In this study we confirmed the ability of the selective 5-HT(4) receptor antagonist SB-207266 1 - 1000 microg kg(-1) p.o., to prevent the increase in defecation evoked over a 60 min period by 5-HTP 10 mg kg(-1) s.c. in conscious mice, in the absence of an apparent constipating action. The role of endogenous 5-HT in the mechanisms of increased defecation and/or diarrhoea was then investigated in conscious, fed rats. This was evoked by 180 min exposure to restraint stress, which increased both the number and mean weight of formed, faecal pellets excreted over the entire time period. SB-207266 1 - 1000 microg kg(-1) p.o. (dosed 30 min before restraint) did not affect the increase in defecation evoked during the first 60 min of restraint stress, but significantly and dose-dependently reduced or prevented the increased defecation during the remaining 120 min of the experiment; this action occurred in the absence of an apparent constipating action of SB-207266. In fasted rats exposed to restraint stress, watery diarrhoea developed and although there was a tendency for SB-207266 1 - 1000 microg kg(-1) p.o. (dosed 30 min before restraint) to reduce the incidence of diarrhoea, this inhibition was not complete. We conclude that selective 5-HT(4) receptor antagonism prevents disruptions in defecation behaviours caused by exogenous or endogenous enteric 5-HT and that this activity is not accompanied by a concomitant suppression of activity (constipation-like) within the intestine itself.

5-HT released by mucosal stimuli initiates peristalsis by activating 5-HT4/5-HT1p receptors on sensory CGRP neurons.

The intestinal peristaltic reflex can be elicited by mucosal stimulation or circular muscle stretch. Muscle stretch activates extrinsic, whereas mucosal stimulation activates intrinsic calcitonin gene-related peptide (CGRP)-containing sensory neurons. The present study examined the role of 5-hydroxytryptamine (5-HT) in sensory transmission. A three-compartment preparation of rat colon was used that enables separate measurement of sensory transmitters and modulators. Mucosal stimuli (2-8 brush strokes) caused concurrent increase in 5-HT and CGRP release in proportion to the intensity of stimulation. Release of both 5-HT and CGRP occurred exclusively into the central compartment where the stimuli were applied. Exogenous 5-HT caused a concentration-dependent release of CGRP. Release of CGRP induced by exogenous 5-HT or mucosal stimulation was inhibited by selective 5-HT4 and 5-HT1p antagonists but was not affected by 5-HT1A, 5-HT2, and 5-HT3 antagonists. Ascending contraction and descending relaxation of circular muscle measured in the peripheral orad and caudad compartments, respectively, were also selectively inhibited by 5-HT4 and 5-HT1p antagonists added to the central but not peripheral compartments. In contrast, muscle stretch elicited CGRP but not 5-HT release; the ascending contraction and descending relaxation components of the peristaltic reflex induced by muscle stretch were not affected by 5-HT antagonists. We conclude that 5-HT released by mucosal stimulation initiates the peristaltic reflex by activating 5-HT4/5-HT1p receptors on sensory CGRP-containing neurons.

Rôle de la substance P dans le contrôle nerveux de la motricité digestive

Substance P is a 11 amino-acids peptide which belongs to the tachykinins, a family of peptide which induces a rapid contraction of the smooth muscle of the digestive tract. The occurrence of substance P has been demonstrated by immunohistochemical and radioimmunological techniques in most parts of the central and peripheral nervous system. Substance P exerts on the smooth muscle of all the areas of the digestive tract a strong excitatory effect which is either direct or relayed by the cholinergic intramural neurones. Numerous electrophysiological, pharmacological and immunohistochemical data lead to the conclusion that substance P is released by intrinsic neurones of the digestive tract or by extrinsic nerves (vagus and splanchnic nerves, etc...). This release is enhanced by acetylcholine, cholecystokinin, serotonin and neurotensin, it is reduced by opioid peptides and noradrenaline. Substance P participates in the intestinal peristaltic reflex by the activation of the smooth muscle cells of the intestine, either directly or through the activation of the cholinergic intrinsic neurones. Substance P is also involved in the genesis of a non-cholinergic ascending excitatory activity likely occurring during vomiting. Lastly, substance P participates in the reflex contraction of the lower oesophageal sphincter following acidification of the distal part of the oesophagus.

Identification of neurotransmitters regulating intestinal peristaltic reflex in humans

The components of the intestinal peristaltic reflex in humans were examined and the neurotransmitters responsible for them identified for the first time in isolated flat sheet segments of intestine. Increasing radial stretch to the caudad end elicited increasing ascending contraction only, whereas increasing radial stretch to the orad end elicited increasing descending relaxation only. Both components were abolished by hexamethonium, implying the participation of cholinergic interneurons in each component. Atropine inhibited ascending contraction only, abolishing the response to low grades of stretch and partially inhibiting the response to high grades of stretch (69% ± 17%, p < 0.01). The substance P antagonist [D-Pro2, D-Trp7,9]substance P partially inhibited ascending contraction induced by high grades of stretch only (40% ± 12%, p < 0.02). The vasoactive intestinal peptide antagonist [4-Cl-D-Phe6, Leu17]vasoactive intestinal peptide inhibited descending relaxation, abolishing the response to low grades of stretch and partially inhibiting the response to high grades of stretch (40% ± 4%, p < 0.001). Release of vasoactive intestinal peptide increased significantly by 91% during descending relaxation only, whereas release of both substance P and substance K increased significantly by 107% during ascending contraction only, supporting the participation of vasoactive intestinal peptide motor neurons in descending relaxation and tachykinin motor neurons as well as cholinergic motor neurons in ascending contraction. The components of the human peristaltic reflex and transmitters regulating them were identical to those found in rat and guinea pig intestine.

Tachykinins as transmitters of ascending contractile component of the peristaltic reflex

The role of tachykinin neurons of the myenteric plexus in the regulation of the intestinal peristaltic reflex was examined in rat colonic segments that enable separate measurement of ascending contraction and descending relaxation. Release of immunoreactive substance P (SP) and substance K (SK) increased (68% for SP; 70% for SK) during ascending contraction and decreased (37% for SP; 13% for SK) during descending relaxation. Exogenous SP and SK augmented ascending contraction and inhibited descending relaxation. Neither SP antiserum nor SK antiserum had any effect on descending relaxation. The antisera had no effect also on ascending contraction induced by low grades of stretch (2 g), but they inhibited significantly contraction induced by high grades (6-10 g) of stretch (52 +/- 8% inhibition with SP antiserum and 48 +/- 8% with SK antiserum). A combination of both antisera abolished ascending contraction induced by high grades of stretch. Atropine inhibited only partially ascending contraction at high grades of stretch. The pattern of inhibition by both antisera and by atropine implied that ascending contraction induced by high grades of stretch was mediated by tachykinin neurons acting directly on smooth muscle cells as well as indirectly via release of acetylcholine from cholinergic motor neurons.

A substance, from wheat gluten, which inhibits the intestinal peristaltic reflex

A substance which inhibits the peristaltic reflex of small intestine has been isolated from extracts of wheat gluten by chromatography and electophoresis. The nature of the substance has been investigated and it has been found to contain adenosine. The results show that adenosine is responsible for the pharmacological activity.