Distribution and Neurochemistry of the Porcine Ileocaecal Valve Projecting Sensory Neurons in the Dorsal Root Ganglia and the Influence of Lipopolysaccharide from Different Serotypes of Salmonella spp. on the Chemical Coding of DRG Neurons in the Cell Cultures.

The ileocecal valve (ICV)-a sphincter muscle between small and large intestine-plays important roles in the physiology of the gastrointestinal (GI) tract, but many aspects connected with the innervation of the ICV remain unknown. Thus, the aim of this study was to investigate the localization and neurochemical characterization of neurons located in the dorsal root ganglia and supplying the ICV of the domestic pig. The results have shown that such neurons mainly located in the dorsal root ganglia (DRG) of thoracic and lumbar neuromers show the presence of substance P (SP), calcitonin gene-related peptide (CGRP), and galanin (GAL). The second part of the experiment consisted of a study on the influence of a low dose of lipopolysaccharide (LPS) from Salmonella serotypes Enteritidis Minnesota and Typhimurium on DRG neurons. It has been shown that the LPS of these serotypes in studied doses does not change the number of DRG neurons in the cell cultures, but influences the immunoreactivity to SP and GAL. The observed changes in neurochemical characterization depend on the bacterial serotype. The results show that DRG neurons take part in the innervation of the ICV and may change their neurochemical characterization under the impact of LPS, which is probably connected with direct actions of this substance on the nervous tissue and/or its pro-inflammatory activity.

New insights into the neuromuscular anatomy of the ileocecal valve.

The neuroanatomy of the ileocecal valve (ICV) is poorly understood. A better understanding of this important functional component of the gastrointestinal tract would enable surgeons to reconstruct an effective valve following surgical resection of the ICV. ICVs were examined in young pigs (N = 5) using frontal and transverse paraffin embedded and frozen sections. Hematoxylin+Eosin (H+E) staining, acetylcholinesterase (AchE), and NADPH-diaphorase (NADPH-d) histochemistry and protein gene product 9.5 (PGP 9.5) and C-kit immunohistochemistry were performed. The H+E staining revealed that the ICV consists of three muscle layers: an external circular muscle layer continuous with that of the ileal circular muscle layer, an inner circular muscle layer continuous with that of the cecal circular muscle layer, and a single longitudinal muscle layer, which appears to be secondary to a fusion of the ileal and cecal longitudinal muscle layers. The AchE, NADPH-d, and PGP 9.5 staining revealed two distinct coaxial myenteric plexuses, together with superficial and deep submucosal plexuses. The C-kit immunostaining showed a continuous myenteric ICC network within the ICV. The structure of the neuromuscular components within the ICV suggests that the valve is a result of a simple intussusception of the terminal ileum into the cecum. This knowledge may help surgeons in their future attempts at reconstructing more anatomically and functionally suitable ICVs following surgical resection of native ICVs.

Parasympathetic innervation of the initial segments of the large intestine in cats.

The locations and morphometric characteristics of efferent parasympathetic neurons in the dorsal motor nucleus of the vagus nerve and the cruciform parasympathetic nucleus of the spinal cord, innervating the area of the ileocecal sphincter and the ascending and transverse segments of the colon, were studied. Horseradish peroxidase solution was injected beneath the serous membranes of these parts of the intestine in urethane-anesthetized cats. After 48 h, animals were subjected to transcardiac perfusion with a fixative mixture and sections of the medulla oblongata and spinal cord were prepared and processed by the Mesulam method. The results showed that all these parts of the large intestine received parasympathetic innervation from neurons in the ventrolateral part of the dorsal motor nucleus, which were uniform in terms of their morphometric characteristics. The number of neurons of this group sending axons to the ileocecal area was greater than the number of neurons innervating the ascending colon. A second group of neurons, which were smaller cells, was located in the same part of the nucleus and innervated the transverse colon. The transverse colon also received innervation from neurons in the cruciform parasympathetic nucleus of the spinal cord.

New hypothesis on the pathogenesis of ileocecal intussusception.

PURPOSE: Ileocecal intussusception is a relatively common surgical emergency in infants and young children. The etiology of intussusception is not clearly understood. Nitric oxide (NO) is a major inhibitory neurotransmitter in the enteric nervous system, which causes relaxation of the smooth muscles. In a lipopolysaccharide-induced experimental model of intussusception, altered intestinal motility is shown to be the result of increased NO released from various inflammatory mediators, which in turn leads to increased incidence of intussusception. The aim of this study was to examine the age-related changes in the nitrergic innervation of the ileocecal valve (ICV) to gain insights into the pathogenesis of intussusception. METHOD: Whole-mount preparations of the myenteric plexus from the ileum, ICV, and proximal colon were stained using NADPH diaphorase histochemistry in newborn piglets (n = 3), 4-week-old (n = 3), 12-week-old (n = 3), and adult pigs (n = 3). Using light microscopy, the number of ganglia per square centimeter, the number of cells per ganglion, and the number of ganglion cells per square centimeter were determined. RESULTS: There were striking regional and age-related differences in nitrergic innervation of myenteric plexus. Density of nitrergic neurons was significantly higher in the ICV than in the terminal ileum and proximal large bowel in the young animals (P < .001). CONCLUSION: These findings suggest that the inflammatory reactions that usually precede intussusception may cause overproduction of NO by the nitrergically hyperinnervated ICV causing relaxation of the ICV and thereby facilitating ileocecal intussusception.

Disposition of muscularis propria and nerve elements in the lip of the ileocaecal valve.

The presence of a valve and the concept of a functional sphincter at the ileocaecal junction have been debated in the literature. Twenty four specimens of ileocaecal organ complex including 10cm of ileum and colon were obtained from cadavers from the Departments of Anatomy and Pathology, PGIMER, Chandigarh. 5 micron thick longitudinal sections from the upper lip of the ileocaecal valve were stained with Hematoxylin and Eosin and Silver stain. Sections of the ileum and colon from 5 ileocaecal organ complexes were also prepared and similarly stained. All the sections were viewed under the light microscope. The muscle bundles constituting the muscularis propria in the ileocaecal valve were arranged in two rows and were tapering in size from the base to the apex. The disposition of the muscle fibres was not uniform in all the specimens. The muscle fibers could be traced up to the upper, middle and lower third of the valve in 8 (33.3%), 14 (58.3%) and 2 (8.3%) specimens respectively. Silver stain of these sections revealed large numbers of nerve elements distributed within the submucosa and muscularis propria. The density of nerve elements in the ileocaecal valve was similar in all the sections irrespective of the extent of the muscularis propria and was comparable to the density of nerve elements in the muscularis propria and submucosa of the ileum and colon. The present study shows that the ileocaecal valve has a well developed muscularis mucosa and a proportionate component of the nerve elements.

Neurogenic pathways mediating ascending and descending reflexes at the porcine ileocolonic junction.

We studied the pharmacology of the neural pathways mediating the responses of ileo- and coloileo-colonic junction (ICJ) to regional distension in ten anaesthetized pigs. Using manometric pullthroughs and a sleeve sensor, we found the ICJ demonstrated sustained tone that was resistant to tetrodotoxin. Ileal distension decreased ICJ pressure by 22.2 ¿ 10.1% (11.9 ¿ 2.7-10.1 ¿ 2.6 mmHg; P=0.002) and colonic distension augmented ICJ pressure by 23.5 ¿ 8.6% (12.8 ¿ 1.5-15.6 ¿ 2.1 mmHg; P=0.02). Bethanecol and Nw-nitro-L-arginine methyl ester (L-NAME) increased ICJ pressure (P=0.002, P=0.01, respectively). Sodium nitroprusside and isoproterenol reduced ICJ pressure (P=0.004, P=0.02, respectively). In the presence of L-NAME, the early inhibitory ileo-ICJ response was abolished, while early and late inhibitory responses were abolished by further addition of propranolol but not by the addition of hexamethonium, atropine, prazosin or yohimbine. The excitatory colo-ICJ response was replaced by inhibition in the presence of L-NAME. We concluded that: (1) the porcine ICJ displays myogenic tone which is influenced by excitatory muscarinic and inhibitory nitrergic and beta adrenergic pathways (2) an inhibitory ileo-sphincteric reflex mediated by nitrergic and beta adrenergic postganglionic neural pathways (3) both excitatory and inhibitory neurogenic colo-sphincteric reflexes exist, and the excitatory pathway involves nitrergic neurotransmission.

Effects of gastrin-releasing peptide (GRP) on the mechanical activity of the human ileocaecal region in vitro.

Mechanical activity was recorded in muscle preparations isolated from the human ileocaecal region. Gastrin-releasing peptide (GRP, 10(-9)-10(-7) mol L-1) produced two types of response in the different muscle layers. Longitudinally cut strips showed a concentration-dependent increase in the rhythmic activity, whereas the circularly orientated layers generally reacted with a small decrease in tone. These effects could not be influenced by blockade of adrenergic or cholinergic receptors or nerve blockade with tetrodotoxin (TTX). Application of pentagastrin did not mimic the action of GRP. These findings suggest a direct action of GRP on smooth muscle via distinct receptors which have already been demonstrated to exist in human gastrointestinal tract. The opposite effects on circular and longitudinal strips might indicate a modulatory role of GRP in the control of ileocolonic transit.

[Distribution and localization of nitric oxide containing neural elements in the digestive tract]. / Nitrogén-monoxid-tartalmú idegelemek megoszlása és lokalizációja a tápcsatornában.

Nitric oxide free radical is a presumed neurotransmitter of the gastrointestinal tract. It can play an important role in the relaxation of the smooth muscles. We used nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry and the localization and morphology of the NADPH-d positive neural elements of the different areas were compared in the cat. NADPH-d positive neurons could be found mainly in the myenteric plexus but some of them were located in the submucous plexus and in the inner circular muscle layer as well as around the blood vessels. The greatest number of the positive neuronal cell bodies could be seen in the myenteric plexuses of the sphincter regions. Stained nerve fibers were seen mainly in the inner circular muscle layer. These results suggest that the positive neurons of the sphincter regions can have an important role in the relaxation. NADPH-d positive neuronal elements of the submucous plexus were located around blood vessels and can regulate the blood flow and secretion of the glands or it is also possible that they belong to the sensory neurons.

Innervation of the rat gastrointestinal sphincters: changes during development and aging.

The effect of age on the adrenergic and peptidergic innervation of the lower oesophageal, pyloric and ileocaecal sphincters of the rat was investigated using immunohistochemical techniques. The distribution of nerve fibres containing the neuronal protein, growth associated protein-43, was also studied to determine the integrity of the enteric nervous system during development and aging. The four age groups examined were 2-3 days, 6 weeks, 3 months and 25 months old rats. Using protein gene product 9.5 antibody (a non-specific general neuronal marker), it was revealed that the myenteric ganglia in all sphincter regions were compactly arranged and were smaller in size at neonatal stage getting more spaced out and larger in size with age. There was no obvious change in the structure of the neutral elements with age. In the lower oesophageal sphincter, calcitonin gene-related peptide- and substance P-like immunoreactive nerve fibres showed notable changes in density and fluorescence intensity with age, decreasing and increasing, respectively, with no obvious change in vasoactive intestinal polypeptide- and growth-associated protein-like immunoreactivity. A slight increase in dopamine-beta-hydroxylase-like immunoreactivity was seen in old age. In the pyloric sphincter, there was an increase in calcitonin gene-related peptide- and substance P-like immunoreactivity with a less notable increase in dopamine-beta-hydroxylase-like immunoreactivity. A decrease in vasoactive intestinal polypeptide- and growth-associated protein-43-like immunoreactivity in the circular muscle of the sphincter was seen in old age. In the ileocaecal sphincter there was a marked increase in growth associated protein-43-, vasoactive intestinal polypeptide-, dopamine-beta-hydroxylase and substance P-like immunoreactivity. There was a decrease in the density of calcitonin gene-related peptide-like immuno-reactive nerve fibres in old age. In summary, two main conclusions can be drawn from the results of the present study. First, there was an age-related differential change in the density of immunoreactive nerve fibres containing various neuroactive substances. This indicates a level of plasticity of the various enteric nerve types and may reflect the degree of importance of the different neurotrasmitters in the physiological activities of the specific sphincter.(ABSTRACT TRUNCATED AT 400 WORDS)

Localization of nitric oxide synthase in canine ileocolonic and pyloric sphincters.

The distribution of neurons containing NAD-PH-diaphorase (NADPH-d) activity and nitric oxide synthase-like immunoreactivity (NOS-LI) in the canine pyloric and ileocolonic sphincters was studied. Cells within the myenteric and submucosal ganglia were positive for NADPH-d. These cells generally had the morphology of Dogiel type-I enteric neurons, however, there was some diversity in the morphology of NADPH-d-positive neurons in the myenteric plexus of the pylorus. Intramuscular ganglia were observed in both sphincters, and NADPH-d was found in a sub-population of neurons within these ganglia. Dual staining with an antiserum raised against nitric oxide synthase (NOS) demonstrated that almost all cells with NOS-LI were also NADPH-d positive. Varicose fibers within ganglia and within the circular and longitudinal muscle layers also possed NOS-LI and NADPH-d activity. Dual staining with anti-VIP antibodies showed that some of the NADPH-d-positive cells in the myenteric and submucosal ganglia also contained VIP-LI, but all VIP-LI-positive cells did not express NADPH-d activity. These data are consistent with recent physiological studies suggesting that nitric oxide serves as an inhibitory neurotransmitter in the pyloric and ileocolonic sphincters. The data also suggest that VIP is expressed in a sub-population of NADPH-d-positive neurons and may therefore act as a co-transmitter in enteric inhibitory neurotransmission to these specialized muscular regions.

Distribution of VIP-immunoreactive nerve cells and fibers in the human ileocecal region.

VIP-containing nerve cells and fibers in the human ileocecal region (pre-junctional ileum, ileocecal and cecocolonic junctions, post-junctional cecum and colon) have been evaluated by immunocytochemistry. A high density of VIP-positive neurons and nerve fibers was found in all layers of the ileum. At all colonic levels examined and at both junctions, the percentage of VIP-containing cells was higher in the submucous plexus than in the myenteric plexus. At both junctions, the muscle wall was devoid of, and the myenteric plexus extremely poor in VIP-positive nerve fibers and cells. These data suggest that motility of these junctions is not--or only to a minor extent--regulated in man by VIP-containing nerves, at variance with other gut sphincteric areas.

Stimulated pressure response of the ileocolonic junctional zone and its use as a continence mechanism in a canine model.

Mechanisms for maintaining passive continence in the efferent limb of urinary diversions include compression of tissue, peristalsis, equilibration of pressure and use of valves. Motor activity and pressure in the ileum, ileocecal valve (ICV) and the colon were evaluated in dogs. Spontaneous activity and pressure were compared with stimulated pressure response and activity. Stimulation was performed at the pelvic nerve and the small nerves in the mesenterium, as well as direct neurostimulation of the bowel. Resting pressure at the ICV was 12.7 +/- 0.4 cmH2O rising to 26.4 +/- 2.2 cmH2O during spontaneous depolarization. Stimulation of the pelvic nerve resulted in increased colonic motor activity with unchanged pressure. Electric stimulation of small mesenterical nerves to the ICV increased pressure in the ICV to 35.0 +/- 4.1 cmH2O, while direct myoelectric stimulation of the ICV zone increased the intraluminal pressure to 75.0 +/- 3.2 cmH2O. Termination of the electric stimulation was followed by a slow decrease of pressure to the resting level over a period of 30-45 s. Maintaining continence at the ICV with long-term constant or intermittent stimulation seems feasible.

Control of the human ileocaecal junction: an in vitro analysis of adrenergic and non-adrenergic non-cholinergic mechanisms.

Circular muscle strips from the human ileocaecal junction developed myogenic tone (sustained resting tension) in vitro and showed an obvious component of relaxation in response to electrical field stimulation. These features are considered typical of sphincteric muscle because circular muscle strips from terminal ileum 1-2 cm proximal to the junction and from the caecum or ascending colon 1-2 cm distal to the junction developed little or no tone and showed predominantly contractile responses to electrical-field stimulation. In 18 out of 29 specimens, junctional strips showed excitatory responses to adrenaline or noradrenaline. The relaxation of junctional strips was non-adrenergic non-cholinergic (NANC) in nature. Nitric oxide appears to be involved in the NANC relaxation.

[Excitatory beta-adrenoreceptors on enteral cholinergic interneurons of the large intestine and ileocecal sphincter]. / Vozbuzhdaiushchie beta-adrenoretseptory na énteral'nykh kholinérgicheskikh interneironakh tolstoi kishki i ileotsekal'nogo sfinktera.

Acute experiments were performed on the isolated intestinal loop, vascularly perfused with arterial blood by means of the constant flow perfusion pump. Contractile activity of the ileocecal sphincter and proximal parts of the large intestine was estimated by the maximal isometric tension and total (integrated) contractile activity. Isoprenaline (1-2 mg) induced contractile responses of the colonic segment and ileocecal sphincter. These responses were abolished or dramatically diminished by the blockade of beta-adrenoceptors, muscarinic, and nicotinic cholinergic receptors. Data obtained support the idea, that the large intestine and ileocecal sphincter have excitatory beta-adrenoceptors localized on cholinergic interneurones of the enteric nervous system.

[Projection of the celiac plexus to the afferent centers demonstrated by the horseradish peroxidase retrograde transport method]. / Proektsii chrevnogo spleteniia na afferentnye tsentry, ustano- vlennye metodom retrogradnogo transporta peroksidazy khrena.

Using the horseradish peroxidase (HP) retrograde method for ascertaining connections of the celiac plexus of the white rat with various afferent centers, HP-marked neurocytes have been revealed in caudal nodes of the vagus nerves and in spinal nodes at the Th4-L2 level. Negative results have been obtained at investigation of the cervical spinal nodes and intramural nodes in the cat ileocecal part. Similar data are obtained, when connections of the celiac plexus with the same area of the cat gut are investigated. Therefore, the problem on interrelations of the celiac plexus with the proper afferent centers of the diaphragmatic+ nerve and the second type cells of Dogel in the cat intramural ganglia is still disputable.

Role of enteric plexuses in the competency of telescoped intestinal valves.

The mechanism of preventing intestinal regurgitation by means of a telescoped intestinal valve has been investigated in this study. Ten telescoped intestinal valves were uniformly created in dogs. A tolerance test to barium reflux was performed before the animals were sacrificed 16 weeks postoperatively, which yielded various barium tolerance values. Statistical analysis revealed that the differences in barium tolerance values in the uniformly constructed valves were closely related to the proportion of the enteric plexuses which contained ganglion cells with normal shapes (r = 0.97, p less than 0.001) and their intestinal segment coverage (r = 0.94, p less than 0.001), but no correlation was found between barium tolerance and the thickness of the valvular musculature (r = 0.04) or the length of the artificial valve (r = 0.39). The results in this study suggest that the intact enteric plexuses play an important role in modulating the competency of the telescoped intestinal valve.

Changes in the responsiveness of the cat ileocecal sphincter to adrenergic agents during the postnatal period.

The changes in the responsiveness of the cat ileocecal sphincter (ICS) to adrenergic agents during the postnatal period were studied by the effect of noradrenaline (applied cumulatively: 0.01-100 mumol) on the mechanical activity:smooth muscle tone and phasic contractions. Noradrenaline contracted the ICS smooth muscle through alpha 1-adrenergic receptors. These receptors were differentiated in the first postnatal days but continue to develop functionally even after 60 days postnatal. An age-determined decrease in the EC50 values of noradrenaline, inducing tonic contractions, was established. Noradrenaline inhibited the phasic contractions of ICS. This effect increased after beta-adrenergic receptor blockers, and disappeared after blocking of alpha 1-adrenergic receptors, which suggests the existence of a postjunctional inhibitory adrenergic receptor of alpha 1 type. The beta-adrenergic receptors in the ICS smooth muscle are not differentiated until 15 days postnatal. They developed functionally towards 30 days postnatal.