Biomedical subjects

M Neunlist

Publications and source records attributed to M Neunlist.

At least 19 recordsLinked to original sources

PubMed2006

Enteric glia inhibit intestinal epithelial cell proliferation partly through a TGF-beta1-dependent pathway.

Although recent studies have shown that enteric neurons control intestinal barrier function, the role of enteric glial cells (EGCs) in this control remains unknown. Therefore, our goal was to characterize the role of EGCs in the control of intestinal epithelial cell proliferation using an in vivo transgenic and an in vitro coculture model. Assessment of intestinal epithelial cell proliferation after ablation of EGCs in transgenic mice demonstrated a significant increase in crypt cell hyperplasia. Furthermore, mucosal glial network (assessed by immunohistochemical detection of S-100beta) is altered in colon adenocarcinoma compared with control tissue. In an in vitro coculture model of subconfluent Caco-2 cells seeded onto Transwell filters with EGCs, Caco-2 cell density and [3H]thymidine incorporation were significantly lower than in control (Caco-2 cultured alone). Flow cytometry analysis showed that EGCs had no effect on Caco-2 cell viability. EGCs induced a significant increase in Caco-2 cell surface area without any sign of cellular hypertrophy. These effects by EGCs were also seen in various transformed or nontransformed intestinal epithelial cell lines. Furthermore, TGF-beta1 mRNA was expressed, and TGF-beta1 was secreted by EGCs. Exogenously added TGF-beta1 reproduced partly the EGC-mediated effects on cell density and surface area. In addition, EGC effects on Caco-2 cell density were significantly reduced by a neutralizing TGF-beta antibody. In conclusion, EGCs have profound antiproliferative effects on intestinal epithelial cells. Functional alterations in EGCs may therefore modify intestinal barrier functions and be involved in pathologies such as cancer or inflammatory bowel diseases.

Adenocarcinoma↗

PubMed2005

Changes in enteric neurone phenotype and intestinal functions in a transgenic mouse model of enteric glia disruption.

AIMS: The influence of enteric glia on the regulation of intestinal functions is unknown. Our aim was to determine the phenotype of enteric neurones in a model of glia alterations and the putative changes in intestinal motility and permeability. METHODS: Transgenic mice expressing haemagglutinin (HA) in glia were used. Glia disruption was induced by injection of activated HA specific CD8+ T cells. Control mice consisted of non-transgenic littermates injected with activated HA specific CD8+ T cells. Immunohistochemical staining for choline acetyltransferase (ChAT), substance P (SP), vasoactive intestinal peptide (VIP), and nitric oxide synthase (NOS) was performed on jejunal submucosal plexus (SMP) and myenteric plexus (MP). Neurally induced jejunal muscle activity was characterised in vitro. Gastrointestinal transit and paracellular permeability were measured using fluorescein isothiocyanate-dextran markers. RESULTS: CD3 positive T cells infiltrates were observed in the MP of transgenic mice. In the SMP, the proportions of VIP and SP positive neurones decreased in transgenic mice compared with control mice. ChAT remained unchanged. In the MP, the proportions of ChAT and NOS positive neurones increased and decreased, respectively, in transgenic mice. In contrast, VIP and SP remained unchanged. Neurally mediated jejunal relaxation was lower in transgenic mice than in controls. This relaxation was reduced by NG-nitro-L-arginine methyl ester in control mice but not in transgenic mice. Gastrointestinal transit was delayed and intestinal permeability increased in transgenic mice compared with control mice. CONCLUSION: Glia disruption induces changes in the neurochemical coding of enteric neurones, which may partly be responsible for dysfunctions in intestinal motility and permeability.

Adoptive Transfer↗

PubMed2005

Serum from achalasia patients alters neurochemical coding in the myenteric plexus and nitric oxide mediated motor response in normal human fundus.

BACKGROUND AND AIMS: Achalasia is a disease of unknown aetiology. An immune mechanism has been suggested on the basis of previous morphological observations. The objective of this study was to test whether the serum of achalasia patients could reproduce the phenotype and functional changes that occur with disease progression in an ex vivo human model. METHODS: Specimens of normal human fundus were maintained in culture in the presence of serum from patients with achalasia, gastro-oesophageal reflux disease (GORD), or healthy subjects (controls). Immunohistochemical detection of choline acetyltransferase (ChAT), neurone specific enolase (NSE), vasoactive intestinal polypeptide (VIP), nitric oxide synthase (NOS), and substance P was carried out in whole mounts of gastric fundus myenteric plexus. In addition, the effects of achalasia serum on electrical field stimulation (EFS) induced contractions were measured in circular muscle preparations. RESULTS: Serum from achalasia patients did not affect the number of myenteric neurones. Tissues incubated with serum from achalasia patients showed a decrease in the proportion of NOS (-26% of NSE positive neurones; p=0.016) and VIP (-54%; p=0.09) neurones, and a concomitant increase in ChAT neurones (+16%; p<0.001) compared with controls. In contrast, GORD serum did not modify the phenotype of myenteric neurones. Area under the curve of EFS induced relaxations (abolished by N-nitro-L-arginine methyl ester) was significantly decreased following incubation with serum from achalasia patients compared with controls (-7.6 (2.6) v -14.5 (5.0); p=0.036). CONCLUSIONS: Serum from achalasia patients can induce phenotypic and functional changes which reproduce the characteristics of the disease. Further identification of putative seric factors and mechanisms involved could lead to the development of novel diagnostic and/or therapeutic strategies in achalasia.

Adult↗

PubMed2004

Effect of LPS on basal and induced apo E secretion by 25-OH chol and 9cRA in differentiated CaCo-2.

The infection and inflammation process is associated with disturbances in lipid and lipoprotein metabolism. The apolipoprotein E (apo E) plays an important role in the lipoprotein metabolism and has been linked to inflammatory disease such as atherosclerosis and Alzheimer disease. An anti-inflammatory effect has also been suggested. The heterodimer nuclear receptor Liver-X-Receptor(alpha)/Retinoid-X-Receptor (LXR(alpha)/RXR) is considered to be a transcription factor for apo E. The aim of this study was to determine whether lipopolysaccharide (LPS) (principal component of the outer membrane Gram-negative bacteria) has an effect on apo E secretion by intestinal mucosa cells, using the Caco-2 cell line. Differentiated Caco-2 cells grown on filter inserts were incubated apically with LPS and/or 25-hydroxycholesterol (25-OH chol) and 9 cis retinoic acid (9cRA), ligands of LXR and RXR, respectively. The apical and basolateral media were separately collected. Apo E was detected by specific antibodies after protein separation by Two-dimensional nondenaturing gradient gel electrophoresis and apo E secreted in the cell culture media was measured by enzyme linked immunosorbent assay (ELISA). Apo E mRNA was analyzed by reverse transcription-polymerase chain reaction (RT-PCR). LXR(alpha) and RXR mass was analyzed by Western Blot. We demonstrate here that CaCo-2 cells secrete apo E, by either apical or basolateral sides, associated with a high-density like lipoprotein, with a stoke's diameter comprised between 7.10 and 8.16 nm. We show that only apical secretion is decreased by LPS in a dose and time dependent manner. This is associated with a decrease in apo E gene expression contrasting with an increase of Il-8, a chemokine factor. Moreover, we demonstrate that only basolateral apo E secretion by CaCo-2 is significantly increased by 25-OH chol and 9cRA while apical secretion remains unchanged. LPS does not decrease the 25-OH chol and 9cRA mediated apo E secretion in basolateral compartment, while apical secretion is diminished under these circumstances. Our results provide evidence for the polarized secretion of apo E by intestinal epithelium. They also demonstrate that apo E secretion by CaCo-2 cell line is decreased by LPS through an LXR(alpha)/RXR independent signaling pathway.

Alitretinoin↗

PubMed2004

The human enteric nervous system.

Decades of work in animal models have demonstrated that the enteric nervous system (ENS) plays a key role in controlling gut functions. Recent advances made it possible to extend such studies to the ENS of man in health and even in disease. Such studies have already provided new insights into the pathophysiology of inflammatory and possibly functional bowel diseases. Studies on human ENS revealed both important similarities and differences between the ENS of man and of experimental animals. Here we summarize the current state of knowledge of the electrophysiology and neurochemistry of the human ENS, including relevant reflex mediated functions in the human gut. Additionally, we review disease associated changes in human ENS properties. Finally, we highlight some research areas that hold special promise in advancing our understanding of the human ENS.

Animals↗

PubMed2003

Toxin B of Clostridium difficile activates human VIP submucosal neurons, in part via an IL-1beta-dependent pathway.

This study investigated whether toxin B of Clostridium difficile can activate human submucosal neurons and the involved pathways. Isolated segments of human colon were placed in organ culture for 3 h in the presence of toxin B or IL-1beta. Whole mounts of internal submucosal plexus were stained with antibodies against c-Fos, neuron-specific enolase (NSE), vasoactive intestinal polypeptide (VIP), and substance P (SP). The membrane potential (Vm) response of submucosal neurons to local application of toxin B and IL-1beta was determined by a multisite optical recording technique. Toxin B (0.1 to 10 ng/ml) increased the proportion of c-Fos-positive neurons dose dependently compared with the control. In the presence of toxin B (10 ng/ml), most c-Fos-positive neurons were immunoreactive for VIP (79.8 +/- 22.5%) but only 19.4 +/- 14.0% for SP. Toxin B induced a rapid rise in IL-1beta mRNA level and a sixfold increase in IL-1beta protein in supernatant after 3 h of incubation. c-Fos expression induced by toxin B was reduced dose dependently by IL-1 receptor antagonist (0.1-10 ng/ml). IL-1beta significantly increased c-Fos expression in submucosal neurons compared with the control (34.2 +/- 10.1 vs. 5.1 +/- 1.3% of NSE neurons). Microejection of toxin B had no effect on the Vm of enteric neurons. Evidence of a direct excitatory effect of IL-1beta on Vm was detected in a minority of enteric neurons. Therefore, toxin B of C. difficile activates VIP-positive submucosal neurons, at least in part, via an indirect IL-1beta-dependent pathway.

Aged↗

PubMed2003

Neurochemical coding of myenteric neurones in the human gastric fundus.

The major functions of the stomach are under the control of the enteric nervous system (ENS), but the neuronal circuits involved in this control are largely unknown in humans. Enteric neurones can be characterized by their neuromediator or marker content, i.e. by neurochemical coding. The purpose of this study was to characterize the presence and co-localization of neurotransmitters in myenteric neurones of the human gastric fundus. Choline acetyltransferase (ChAT), neurone-specific enolase (NSE), vasoactive intestinal polypeptide (VIP), nitric oxide synthase (NOS), substance P (SP) were detected by immunohistochemical methods in whole mounts of gastric fundus myenteric plexus (seven patients). Antibodies against ChAT and NOS labelled the majority of myenteric neurones identified by NSE (57.2 +/- 5.6% and 40.8 +/- 4.5%, respectively; mean +/- SD). The proportions of VIP- and SP-immunoreactive neurones were significantly smaller, constituting 19.6 +/- 6.9% and 16.0 +/- 3.7%, respectively. Co-localization studies revealed five major populations representing over 75% of the myenteric neurones: ChAT/-, 30.1 +/- 6.1%; NOS/-, 24.2 +/- 4.4%; ChAT/SP/-, 8.3 +/- 3.1%; NOS/VIP/-, 7.2 +/- 6.0%; ChAT/VIP/-, 4.9 +/- 2.6. Some similarities are apparent in the neurochemical coding of myenteric neurones in the stomach and intestine of humans, and between the stomach of humans and animals, but striking differences exist. The precise functional role of the neurochemically identified classes of neurones remains to be determined.

Aged↗

PubMed2003

Human submucosal neurones regulate intestinal epithelial cell proliferation: evidence from a novel co-culture model.

The role of the human enteric nervous system (ENS) in the control of the intestinal epithelium organization and proliferation is unknown. To address this issue, we developed a novel co-culture model, consisting of human submucosa containing the submucosal plexus and a human colonic epithelial monolayer. After 3 days in basal conditions (i.e. in absence of neuronal activation) epithelium disorganization and proliferation occurred. In contrast, electrical activation of submucosal neurones maintained monolayer organization and decreased cell proliferation. These effects were blocked by tetrodotoxin and a vasoactive intestinal peptide (VIP) receptor antagonist, and reproduced by VIP. In conclusion, our study suggests that the human ENS is involved in the control of epithelial cell proliferation.

Aged↗

PubMed2003

Changes in chemical coding of myenteric neurones in ulcerative colitis.

BACKGROUND: Morphological and functional changes in the enteric nervous system (ENS) have been reported in inflammatory bowel diseases but it is still uncertain whether neurochemical coding of myenteric neurones is altered in ulcerative colitis (UC). AIMS: In this study we investigated transmitter co-localisation in myenteric neurones of normal colon and the colon of patients with UC. METHODS: Choline acetyltransferase (ChAT), neurone specific enolase (NSE), vasoactive intestinal peptide (VIP), and substance P (SP) were detected by immunohistochemical methods in whole mounts of colonic myenteric plexus of UC patients (n=10) and controls (n=8). RESULTS: The proportion of ChAT positive and VIP positive neurones relative to the NSE population did not differ in inflamed (33.3% and 9.3%, respectively) and non-inflamed segments (33.6% and 9.7%) of UC colon compared with controls (35.0% and 6.9%). The proportion of SP positive neurones was significantly larger in both inflamed (15.5%) and non-inflamed (20.3%) segments than in controls (5.9%). Analysis of changes in subpopulations showed that 26.9% of neurones were only ChAT positive in controls but that the proportion was significantly smaller in inflamed (18.8%) and non-inflamed (15.8%) areas of UC. The proportions of neurones containing ChAT and SP were significantly higher in inflamed (11.8%) and non-inflamed (13.9%) areas than in controls (5.0%). CONCLUSION: Remodelling of myenteric neurones in UC involves a shift from mainly cholinergic to more SP positive innervation. This effect may constitute part of the neuronal basis for the motility disturbances observed in UC.

Adult↗

PubMed2001

Glycine activates myenteric neurones in adult guinea-pigs.

1. We studied the effects of glycine on myenteric neurones and muscle activity in the colon and stomach of adult guinea-pigs. 2. Intracellular recordings revealed that myenteric neurones responded to local microejection of glycine (1 mM) with a fast, transient membrane potential depolarisation (57 % of 191 colonic neurones and 26 % of 50 gastric neurones). Most glycine-sensitive neurones had ascending projections and were choline acetyltransferase immunoreactive. Glycine preferentially activated neurones with a late afterhyperpolarisation (AH-neurones) and tonic spiking neurones with fast synaptic inputs (tonic S-neurones) but less frequently phasic S-neurones and inexcitable (non-spiking) neurones. The depolarisation had a reversal potential at -19 +/- 13 mV, which was increased by 18 +/- 10 % upon lowering extracellular chloride concentration and decreased by 38 +/- 14 % in furosemide (frusemide, 2 mM). 3. Strychnine (300 nM) reversibly abolished the glycine-induced depolarisation and the Cl(-) channel blocker picrotoxin (100 microM) reduced the amplitude of the depolarisation by 55 +/- 5 %. The glycine effect was a postsynaptic response because it was not changed after nerve blockade with tetrodotoxin (1 microM) or blockade of synaptic transmission in reduced extracellular [Ca(2+)]. The effect was specific since the response was not changed by the nicotinic antagonists hexamethonium (200 microM) and mecamylamine (100 microM), the GABA(A) receptor antagonist bicuculline (10 microM), the NMDA antagonist MK-801 (20 microM) or the 5-HT(3) antagonist ICS 205930 (1 microM). 4. Glycine (1 mM) induced a tetrodotoxin- and strychnine-sensitive contractile response in the colon; the contractile response in the stomach was tetrodotoxin insensitive. 5. Glycine activated myenteric neurones in the adult enteric nervous system through strychnine-sensitive mechanisms. The glycine-evoked depolarisation was caused by Cl(-) efflux and the maintenance of relatively high intracellular chloride concentrations involved furosemide-sensitive cation-chloride co-transporters.

Animals↗

PubMed2001

Neural components of distension-evoked secretory responses in the guinea-pig distal colon.

1. Using a Ussing chamber and neuronal retrograde tracing with 1,1'-didodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) we characterized the afferent and efferent neuronal pathways which mediated distension-evoked secretion in the guinea-pig distal colon. 2. Acute capsaicin application (10 microM) to the serosal site of the Ussing chamber evoked a secretory response which was blocked by tetrodotoxin (1 microM), the combined application of the NK1 and NK3 receptor antagonists CP-99,994-1 and SR 142801 (1 microM), and by combined application of atropine (10 microM) and the VIP receptor antagonist VIP(6-28) (10 microM). Functional desensitization of extrinsic primary afferents by long-term application of capsaicin significantly diminished distension-evoked secretion by 46 %. 3. After functional desensitization by capsaicin, serosal application of gadolinium (100 microM) inhibited the distension-evoked chloride secretion by 54 %; the L-type Ca(2+) channel blocker nifedipine (1 microM) and the 5-HT(1P) receptor antagonist renzapride (1 microM) had no effect. The combination of atropine and VIP(6-28) or the combination of NK1 and NK3 receptor antagonists almost abolished distension-evoked secretion. 4. The secretory response evoked by electrical field stimulation, carbachol (1 microM) or VIP (1 microM) was not attenuated by gadolinium. Field stimulation-evoked chloride secretion was not affected by blockade of NK1 and NK3 receptors. 5. Twelve per cent of DiI-labelled submucosal neurones with projections to the mucosa were immunoreactive for choline acetyltransferase, substance P and calbindin and very probably represented intrinsic primary afferent neurones. 6. Distension-evoked chloride secretion was mediated by capsaicin-sensitive extrinsic primary afferents and by stretch-sensitive intrinsic primary afferent neurones. Both the extrinsic and intrinsic afferents converge on common efferent pathways. These pathways consist of VIPergic and cholinergic secretomotor neurones that are activated via NK1 and NK3 receptors.

Afferent Pathways↗

PubMed2001

Projections of excitatory and inhibitory motor neurones to the circular and longitudinal muscle of the guinea pig colon.

The aim of this study was to identify myenteric pathways to the circular and longitudinal muscle of the guinea pig proximal colon. To identify excitatory and inhibitory muscle motoneurones, we applied the neuronal retrograde tracer DiI onto the circular or longitudinal muscle layer and performed additional immunohistochemistry for nitric oxide synthase (NOS) and choline acetyltransferase (ChAT). On average 166 +/- 81 circular muscle motoneurones (CMMN) and 100 +/- 74 longitudinal muscle motoneurones (LMMN) were labelled by DiI tracing. Myenteric pathways innervating the muscle were either ascending (DiI-labelled neurones with oral projections) or descending (DiI-labelled neurones with anal projections). The circular muscle was preferentially innervated by ascending pathways (66.0 +/- 9.1%). Most ascending CMMN were ChAT-positive (87.2 +/- 8.5%), whereas descending CMMN were mainly NOS-positive (82.3 +/- 14.6%). Most ascending (62.2 +/- 11.1%) and descending (82.0 +/- 12.5%) CMMN had circumferential projection preferences (circumferential projections were longer than projections along the longitudinal gut axis). In contrast to the polarised projections to the circular muscle, the longitudinal muscle was equally innervated by ascending (46.2 +/- 15.1%) and descending (53.9 +/- 15.1%) neurones. Ascending and descending pathways to the longitudinal muscle consisted predominantly of ChAT-positive neurones (98.1 +/- 1.9% and 68.0 +/- 8.5%, respectively), and both pathways had prominent longitudinal projection preferences. Only 25.5% of the descending LMMN were NOS-positive. In conclusion, the circular muscle in the proximal colon is innervated by descending inhibitory (NOS-positive neurones) and ascending excitatory (ChAT-positive neurones) pathways. In contrast, the longitudinal muscle is primarily innervated by ascending and descending excitatory motoneurones, and only a small proportion of the descending pathway consisted of inhibitory motoneurones.

Animals↗

PubMed2001

Neurotransmitter coding of enteric neurones in the submucous plexus is changed in non-inflamed rectum of patients with Crohn's disease.

Knowledge of the neurochemical coding of submucosal neurones in the human gut is important to assess neuronal changes under pathological conditions. We therefore investigated transmitter colocalization patterns in rectal submucosal neurones in normal tissue (n=11) and in noninflamed tissue of Crohn's disease (CD) patients (n=17). Neurone-specific enolase (NSE), choline acetyltransferase (ChAT), vasoactive intestinal polypeptide (VIP), substance P (SP), nitric oxide synthase (NOS) and calcitonin gene-related peptide (CGRP) were detected immunohistochemically in whole-mount preparations from rectal biopsies. The neuronal marker NSE revealed no differences in the number of cells per ganglion (controls 5.0; CD 5.1). Four cell populations with distinct neurochemical codes were identified. The sizes of the populations ChAT/VIP (58% vs. 55%), ChAT/SP (8% vs. 8%), and ChAT/- (22% vs. 22%) were similar in control and CD. The population VIP/- was significantly increased in CD (12% vs. 2% in controls). Unlike in controls, all NOS neurones colocalized ChAT in CD. Thickened CGRP-fibres occurred in CD. We identified neurochemically distinct populations in the human submucous plexus. The increase in the VIP/- population, extensive colocalization of ChAT and NOS and hypertrophied CGRP fibres indicated adaptive changes in the enteric nervous system in noninflamed rectum of CD patients.

Adolescent↗

PubMed1999

Characteristics of mucosally projecting myenteric neurones in the guinea-pig proximal colon.

1. Using retrograde tracing with 1,1'-didodecyl-3,3,3', 3'-tetramethylindocarbocyanine perchlorate (DiI) in combination with electrophysiological and immunohistochemical techniques we determined the properties of the putative intrinsic primary afferent myenteric neurones with mucosal projections in the guinea-pig proximal colon. 2. Eighty-four out of eighty-five DiI-labelled myenteric neurones were AH neurones with a late after-hyperpolarization. Thirty-three per cent of them exhibited atropine- and tetrodotoxin-resistant spontaneously occurring hyperpolarizing potentials (SHPs) during which the membrane resistance and excitability decreased. 3. DiI-labelled AH neurones had multipolar Dogiel type II morphology, primarily of the dendritic type. Sixty-one per cent of the neurones were immunoreactive for choline acetyltransferase (ChAT) and calbindin (Calb) and 23 % were ChAT positive but Calb negative. 4. DiI-labelled neurones did not receive fast excitatory postsynaptic potentials but 94 % (34/36) received slow excitatory postsynaptic potentials (sEPSPs). The neurokinin-3 (NK-3) agonist (MePhe7)-NKB but not the NK-1 agonist [(SAR9,Met(O2)11]-SP mimicked this response. The NK-3 receptor antagonist SR 142801 (1 microM) significantly decreased the amplitude and duration of the sEPSPs; the NK-1 receptor antagonist CP-99,994 (1 microM) was ineffective. Atropine (0.5 microM) increased the duration but not the amplitude of the sEPSPs. 5. Microejection of 100 mM sodium butyrate onto the neurones induced in 90 % of the DiI-labelled neurones a transient depolarization associated with an increased excitability. In neurones with SHPs sodium butyrate evoked, additionally, a late onset hyperpolarization. Perfusion of 0.1-10 mM sodium butyrate induced a dose-dependent increase in neuronal excitability. Sodium butyrate was ineffective when applied directly onto the mucosa. 6. Mucosally projecting myenteric neurones of the colon are multipolar AH neurones with NK-3-mediated slow EPSPs and somal butyrate sensitivity.

Animals↗

PubMed1999

Different tachykinin receptors mediate chloride secretion in the distal colon through activation of submucosal neurones.

We investigated the role of tachykinin receptor subtypes on secretory responses in the guinea-pig distal colon using Ussing chamber experiments and intracellular recordings from submucosal neurones. Choline acetyltransferase (ChAT) and vasoactive intestinal polypeptide (VIP) were demonstrated in submucosal neurones by immunohistochemistry. In Ussing chamber experiments substance P (SP), the NK1-receptor agonist [SAR9,Met(O2)11]-SP and the NK-3-receptor agonist (MePhe7)-NKB increased dose-dependently short-circuit currents. The NK-2-receptor agonist (betaAla8)-NKA(4-10) had no effect. Responses to 1-100 nM SP, [(SAR9,Met(O2)11]-SP and (MePhe7)-NKB were tetrodotoxin-sensitive but hexamethonium-insensitive. While (MePhe7)-NKB-responses were atropine-sensitive at all concentrations, the atropine sensitivity of the secretory responses to SP and [SAR9,Met(O2)11]-SP dramatically decreased with increasing concentrations. [SAR9,Met(O2)11]-SP and (MePhe7)-NKB effects were blocked by the selective NK-I and NK-3 antagonists CP-99,994-1 (1 microM) and SR 142801 (1 microM), respectively. Combination of both antagonists blocked the SP-response. SR 142801 also suppressed the response to [SAR9,Met(O2)11]-SP. Desensitization with [SAR9,Met(O2)11]-SP significantly decreased (MePhe7)-NKB-responses but not vice versa. In intracellular recordings 90% of submucosal neurones were activated by both ISAR9,Met(O2)11]-SP and (MePhe7)-NKB as indicated by membrane depolarisation and enhanced spike discharge. These effects were tetrodotoxin-resistant and potentiated by atropine. NK-1-and NK-3-mediated responses occurred equally in ChAT-positive and in VIP-positive neurones. The results suggest the importance of NK-1- and NK-3-receptors on cholinergic and non-cholinergic submucosal neurones for secretory processes in the guinea-pig distal colon.

Action Potentials↗

PubMed1999

Multisite optical recording of excitability in the enteric nervous system.

A multisite optical recording technique consisting of an array of 464 photodiodes was used to measure dynamic changes in transmembrane potentials (Vm) of guinea-pig and mouse enteric neurones stained with the voltage-sensitive dye Di-8-ANEPPS. Optical recordings of Vm changes in enteric neurones which were evoked by depolarizing current pulses or synaptic activation mirrored the Vm changes measured intracellularly in the same neurone. Action potentials had fractional change in fluorescence of -0.09 +/- 0.06% and their peak to peak noise level was 20 +/- 14% of the action potential amplitude. Optical recordings after electrical stimulation of interganglionic nerve strands revealed slow EPSPs, nicotinergic supra- and subthreshold fast EPSPs as well as propagation of action potentials along interganglionic strands. Local application of acetylcholine onto a single ganglion induced reproducibly and dose dependently action potential discharge demonstrating the feasibility of neuropharmacological studies. The optical mapping made it possible to record action potentials simultaneously in a large number of neurones with high spatiotemporal resolution that is unattainable by conventional techniques. This technique presents a powerful tool to study excitability spread within enteric circuits and to assess differential activation of enteric populations in response to a number of stimuli which modulate neuronal activity directly or through synaptic mechanisms.

Acetylcholine↗

PubMed1999

The enteric nervous system: region and target specific projections and neurochemical codes.

The goal of this report is to summarise the current knowledge on the projection pathways of enteric neurones innervating the muscle and mucosa in different regions of the gut. Combination of neuronal tracing, immunohistochemical and electrophysiological methods has allowed researchers to gain insight into the enteric hardwiring of specific target tissue in the gut. A polarised innervation pattern of the circular muscle was demonstrated for the stomach fundus/corpus and the ileum with descending pathways being primarily nitrergic while ascending pathways were primarily cholinergic. This characteristic hardwiring is thought to set in part the functional basis for peristalsis. A similar polarised innervation pathway was found for the enteric innervation of the mucosa in the stomach and large intestine but not in the small intestine. In both the stomach (myenteric neurones) and in the proximal and distal colon (submucosal neurones), ascending pathways to the mucosa are primarily cholinergic while descending pathways are primarily non-cholinergic. In the colon, results suggest that activation of both pathways induces a cross potentiation of cholinergic and vasoactive intestinal polypeptidergic mediated secretion. Furthermore, a large population of myenteric neurone s projecting to the mucosa in the small and large intestine are probably intrinsic primary afferent neurones sensitive to mechanical as well as chemical stimuli.

Animals↗

PubMed1998

Polarised innervation pattern of the mucosa of the guinea pig distal colon.

A neuronal retrograde tracing method with the dye DiI in combination with the immunohistochemical detection of vasoactive intestinal polypeptide (VIP) and choline acetyltransferase (ChAT) was used to characterise the mucosal projection of neurones located in the submucosal plexus of the guinea pig distal colon. VIP and ChAT immunoreactivity labelled separate populations. The mucosa was innervated by descending (59.7 +/- 7.4%), ascending (8.7 +/- 5.0%) and circumferential (31.7 +/- 6.1%) pathways. Descending neurones had longer projections than ascending ones. Descending DiI-labelled neurones were in their vast majority VIP-positive (88.4 +/- 3.6%) and only 3.5 +/- 4.8% of them were ChAT-positive. Conversely, the ascending pathways were predominantly ChAT-positive (90.9 +/- 20.3%) and only 7.8 +/- 20.6% of them were VIP-positive. A significantly larger proportion of DiI-neurones with circumferential projection were ChAT- than VIP-positive (70.9 +/- 15.6 vs. 27.8 +/- 15.7%). The proportion of ascending, descending and circumferential ChAT-positive DiI-neurones was 23:7:70%, respectively, whereas the proportion for VIP-positive DiI-neurones was 2:84:14%, respectively. The results suggest a conservation of the submucosal innervation pattern along the guinea pig colonic mucosa but also reveal differences in the mucosal enteric innervation of other regions of the gastrointestinal tract.

Animals↗