Interstitial cell network volume is reduced in the terminal bowel of ageing mice

Ageing is associated with impaired neuromuscular function of the terminal gastrointestinal (GI) tract, which can result in chronic constipation, faecal impaction and incontinence. Interstitial cells of cajal (ICC) play an important role in regulation of intestinal smooth muscle contraction. However, changes in ICC volume with age in the terminal GI tract (the anal canal including the anal sphincter region and rectum) have not been studied. Here, the distribution, morphology and network volume of ICC in the terminal GI tract of 3‐ to 4‐month‐old and 26‐ to 28‐month‐old C57BL/6 mice were investigated. ICC were identified by immunofluorescence labelling of wholemount preparations with an antibody against c‐Kit. ICC network volume was measured by software‐based 3D volume rendering of confocal Z stacks. A significant reduction in ICC network volume per unit volume of muscle was measured in aged animals. No age‐associated change in ICC morphology was detected. The thickness of the circular muscle layer of the anal sphincter region and rectum increased with age, while that in the distal colon decreased. These results suggest that ageing is associated with a reduction in the network volume of ICC in the terminal GI tract, which may influence the normal function of these regions.     

Spatial localization and properties of pacemaker potentials in the canine rectoanal region

The present study investigated the spatial organization of electrical activity in the canine rectoanal region and its relationship to motility patterns. Contraction and resting membrane potential (E(m)) were measured from strips of circular muscle isolated 0.5-8 cm from the anal verge. Rapid frequency [25 cycles/min (cpm)] E(m) oscillations (MPOs, 12 mV amplitude) were present across the thickness of the internal anal sphincter (IAS; 0.5 cm) and E(m) was constant (-52 mV). Between the IAS and the proximal rectum an 18 mV gradient in E(m) developed across the muscle thickness with the submucosal edge at -70 mV and MPOs were replaced with slow waves (20 mV amplitude, 6 cpm). Slow waves were of greatest amplitude at the submucosal edge. Nifedipine (1 micro M) abolished MPOs but not slow waves. Contractile frequency changes were commensurate with the changes in pacemaker frequency. Our results suggest that changing motility patterns in the rectoanal region are associated with differences in the characteristics of pacemaker potentials as well as differences in the sites from which these potentials emanate.     

Enteric motor neurons form synaptic-like junctions with interstitial cells of Cajal in the canine gastric antrum

Morphological studies have shown synaptic-like structures between enteric nerve terminals and interstitial cells of Cajal (ICC) in mouse and guinea pig gastrointestinal tracts. Functional studies of mice lacking certain classes of ICC have also suggested that ICC mediate enteric motor neurotransmission. We have performed morphological experiments to determine the relationship between enteric nerves and ICC in the canine gastric antrum with the hypothesis that conservation of morphological features may indicate similar functional roles for ICC in mice and thicker-walled gastrointestinal organs of larger mammals. Four classes of ICC were identified based on anatomical location within the tunica muscularis. ICC in the myenteric plexus region (IC-MY) formed a network of cells that were interconnected to each other and to smooth muscle cells by gap junctions. Intramuscular interstitial cells (IC-IM) were found in muscle bundles of the circular and longitudinal layers. ICC were located along septa (IC-SEP) that separated the circular muscle into bundles and were also located along the submucosal surface of the circular muscle layer (IC-SM). Immunohistochemistry revealed close physical associations between excitatory and inhibitory nerve fibers and ICC. These contacts were synaptic-like with pre- and postjunctional electron-dense regions. Synaptic-like contacts between enteric neurons and smooth muscle cells were never observed. Innervated ICC formed gap junctions with neighboring smooth muscle cells. These data show that ICC in the canine stomach are innervated by enteric neurons and express similar structural features to innervated ICC in the murine GI tract. This morphology implies similar functional roles for ICC in this species.     

Rectoanal reflex in healthy subject

The present work was performed in order to examine the rectoanal reflex in healthy subject. Our findings suggest that the fecal continence is sustained by both the anal sphincters. The maximal anal pressure at rest is governed by activity of the internal sphincter while the external anal sphincter promptly contracts when the rectum is distended.     

Interstitial cells in the primate gastrointestinal tract

Kit immunohistochemistry and confocal reconstructions have provided detailed 3-dimensional images of ICC networks throughout the gastrointestinal (GI) tract. Morphological criteria have been used to establish that different classes of ICC exist within the GI tract and physiological studies have shown that these classes have distinct physiological roles in GI motility. Structural studies have focused predominately on rodent models and less information is available on whether similar classes of ICC exist within the GI tracts of humans or non-human primates. Using Kit immunohistochemistry and confocal imaging, we examined the 3-dimensional structure of ICC throughout the GI tract of cynomolgus monkeys. Whole or flat mounts and cryostat sections were used to examine ICC networks in the lower esophageal sphincter (LES), stomach, small intestine and colon. Anti-histamine antibodies were used to distinguish ICC from mast cells in the lamina propria. Kit labeling identified complex networks of ICC populations throughout the non-human primate GI tract that have structural characteristics similar to that described for ICC populations in rodent models. ICC-MY formed anastomosing networks in the myenteric plexus region. ICC-IM were interposed between smooth muscle cells in the stomach and colon and were concentrated within the deep muscular plexus (ICC-DMP) of the intestine. ICC-SEP were found in septal regions of the antrum that separated circular muscle bundles. Spindle-shaped histamine⁺ mast cells were found in the lamina propria throughout the GI tract. Since similar sub-populations of ICC exist within the GI tract of primates and rodents and the use of rodents to study the functional roles of different classes of ICC is warranted.     

The intrinsic myenteric innervation of the hind-gut and accessory muscles of defaecation in the cat

Summary The anatomy and intrinsic innervation of the colon, rectum, internal anal sphincter, ano-coccygeus and recto-coccygeus have been studied in the cat with cholinesterase and catecholamine-fluorescence histochemical techniques. A variable pattern of intrinsic innervation by acetylcholinesterase-positive and adrenergic nerves along the length of the large bowel is described and is related to segmental variations in motor activity. A variation in the distribution of non-specific cholinesterase within the muscle layers is also described. Adrenergic nerves in proximal colon are arranged in the usual peri-ganglionic manner but there is also a rich direct adrenergic innervation of the longitudinal muscle in distal colon and rectum, and of circular muscle in lower rectum and internal anal sphincter. This distribution has not been reported in other species. Direct adrenergic innervation of muscle cells has been confirmed at ultrastructural level after treatment with 5-hydroxydopamine. Adrenergic neurones have not been detected in cat bowel. The ano- and recto-coccygeus muscles and internal anal sphincter possess a dense innervation of adrenergic and cholinesterase-positive nerves. It is suggested that the variation in intrinsic innervation along the large bowel should be considered in the interpretation of pharmacological and physiological experiments on this part of the gut.This work was supported by a grant from the King's College Hospital Voluntary Research Trust. We wish to thank Dr. J. P. Tranzer and F. Hoffman-La Roche & Co. Ltd., Basle, for the gift of 5-hydroxydopamine.We also thank Miss M. K. Egan and Mr. K. J. Davies for their technical assistance.     

Integrative control of rectoanal reflex in guinea pigs through lumbar colonic nerves

The aim of the present study was to analyze the neuromodulation of rectoanal reflex activity by lumbar sympathetic nerves in guinea pigs. The mechanical activities of the rectum were recorded with a balloon connected to a pressure transducer, and those of the internal anal sphincter (IAS) were recorded with a custom-made strain gauge force transducer. Gradual and sustained rectal distension evoked the rectoanal reflex, causing cholinergic contractions of the rectum and synchronous nitrergic relaxations of the IAS. Section of the lumbar colonic nerves enhanced both rectal contractions and IAS relaxations. Section of the 13th thoracic cord abolished both rectal contractions and IAS relaxations, but section of the lumbar colonic nerves restored them. Lumbar sympathectomy and pithing sacral cords greatly diminished these rectal contractions and IAS relaxations, but the intrinsic reflex component remained. NG-nitro-L-arginine methyl ester enhanced the intrinsic reflex-mediated contraction of the rectum and abolished reflex-mediated relaxation of the IAS and converted into cholinergic contractions. The present results indicate that the extrinsic lumbar inhibitory outflow causes marked inhibition of the rectoanal reflex via the lumbar colonic nerves.     

A qualitative and quantitative study on the enkephalinergic innervation of the pig gastrointestinal tract.

Enkephalins are involved in neural control of digestive functions such as motility, secretion, and absorption. To better understand their role in pigs, we analyzed the qualitative and quantitative distribution of enkephalin immunoreactivity (ENK-IR) in components of the intestinal wall from the esophagus to the anal sphincter. Immunohistochemical labelings were analyzed using conventional fluorescence and confocal microscopy. ENK-IR was compared with the synaptophysin immunoreactivity (SYN-IR). The results show that maximal ENK-IR levels in the entire digestive tract are reached in the myenteric plexuses and, to a lesser extent, in the external submucous plexus and the circular muscle layer. In the longitudinal muscle layer, ENK-IR was present in the esophagus, stomach, rectum, and anal sphincter, whereas it was absent from the duodenum to the distal colon. In the ENK-IR plexuses and muscle layers, more than 60% of the nerve fibers identified by SYN-IR expressed ENK-IR. No ENK-IR was observed in the internal submucous plexus and the mucosa; the latter was found to contain ENK-IR endocrine cells. These results strongly suggest that, in pigs, enkephalins play a major role in the regulatory mechanisms that underlie the neural control of digestive motility.     

Modular and functionally-different descending recto-anal motor pathways in a rat model

We evaluated the motor responses in recto-anal preparations obtained from rats, in terms of the excitation displayed by modules of nerve networks and descending distally directed pathways, when subjected to the mechanographic on-line technique, a partitioned organ bath, electrical stimulation (EFS, 0.8 ms, 5 Hz) and distension. EFS elicited modular contractions, which increased in amplitude distally, in circular muscle rings isolated from the proximal, middle or distal rectum. The modular responses of the internal anal sphincter or anal canal were relaxation or contraction, respectively. The application of EFS to the distal rectum induced a descending contractile response in the anal canal (5.24±0.34 mN), while distension by balloon evoked a descending response consisting of contraction (1.72±0.20 mN) followed by relaxation (3.42±0.24 mN). The responses were sensitive to tetrodotoxin. Atropine considerably depressed the contractions in all preparations. Whether or not atropine was present, L-NNA increased the excitatory responses, while L-arginine decreased the contractions and extended the relaxation of internal anal sphincter and anal canal. The results suggest that excitatory neurotransmission(s) expressed in the distal rectum dominate modular nerve networks. Functionally-different descending pathways are involved in the motor activity of the anal canal. Stimulatory cholinergic pathways are dependent on the electrically-induced excitation, and inhibitory nitrergic pathways are sensitive to distension of rectal wall.     

Unique distribution of interstitial cells of Cajal in the feline pylorus

The feline gastrointestinal (GI) tract is an important model for GI physiology but no immunohistochemical assessment of interstitial cells of Cajal (ICC) has been performed because of the lack of suitable antibodies. The aim of the present study was to investigate the various types of ICC and associated nerve structures in the pyloric sphincter region, by using immunohistochemistry and electron microscopy to complement functional studies. In the sphincter, ICC associated with Auerbach’s plexus (ICC-AP) were markedly decreased within a region of 6–8 mm in length, thereby forming an interruption in this network of ICC-AP, which is otherwise continuous from corpus to distal ileum. In contrast, intramuscular ICC (ICC-IM) were abundant within the pylorus, especially at the inner edge of the circular muscle adjacent to the submucosa. Similar distribution patterns of nerves positive for vesicular acetylcholine transporter (VAChT), nitric oxide synthase (NOS) and substance P (SP) were encountered. Quantification showed a significantly higher number of ICC-IM and the various types of nerves in the pylorus compared with the circular muscle layers in the adjacent antrum and duodenum. Electron-microscopic studies demonstrated that ICC-IM were closely associated with enteric nerves through synapse-like junctions and with smooth muscle cells through gap junctions. Thus, for the first time, immunohistochemical studies have been successful in documenting the unique distribution of ICC in the feline pylorus. A lack of ICC-AP guarantees the distinct properties of antral and duodenal pacemaker activities. ICC-IM are associated with enteric nerves, which are concentrated in the inner portion of the circular muscle layer, being part of a unique innervation pattern of the sphincter. This study was supported by operating grants from the Canadian Institutes of Health Research (to J.D.H. and N.E.D.) and from the Canadian Association of Gastroenterology (to L.W.C.L.).     

C-kit-immunopositive interstitial cells of Cajal in human embryonal and fetal oesophagus

Interstitial cells of Cajal (ICC) are morphologically and functionally intercalated between the elements of the enteric nervous system and the smooth muscle cells (SMCs) in the musculature of the digestive tract. Kit immunohistochemistry reliably identifies the location of these cells and provides information on changes in ICC distribution and density. Human oesophagus specimens (7 embryos, 23 fetuses at 7-27 weeks gestational age; both sexes) were exposed to Kit antibodies to determine ICC differentiation. Enteric plexuses were examined immunohistochemically by using anti-neuron-specific enolase, whereas the differentiation of SMCs was studied with antibodies against α-smooth-muscle actin and desmin. By week 7, c-kit-immunopositive cells were present along the entire oesophagus in the form of an uninterrupted layer around the myenteric plexus (MP) elements. From the beginning of the 3rd month, the number of ICC progressively decreased around the MP ganglia but increased within the muscle layers. Concomitantly, differences in the number and distribution of ICC were established in the various portions of the oesophagus: specifically, ICC were abundant in the lower portion, less numerous in the middle region and rare in the upper part. By the 5th month of development, the relationship as found in later developmental stages had been established: C-kit IR ICC were present within the circular muscle layer, within the longitudinal layer and in the connective septa surrounding the muscle bundles but were completely missing around the MP ganglia.     

Development of c‐kit immunopositive interstitial cells of Cajal in the human stomach

Interstitial cells of Cajal (ICC) include several types of specialized cells within the musculature of the gastrointestinal tract (GIT). Some types of ICC act as pacemakers in the GIT musculature, whereas others are implicated in the modulation of enteric neurotransmission. Kit immunohistochemistry reliably identifies the location of these cells and provides information on changes in ICC distribution and density. Human stomach specimens were obtained from 7 embryos and 28 foetuses without gastrointestinal disorders. The specimens were 7–27 weeks of gestational age, and both sexes are represented in the sample. The specimens were exposed to anti‐c‐kit antibodies to investigate ICC differentiation. Enteric plexuses were immunohistochemically examined by using anti‐neuron specific enolase and the differentiation of smooth muscle cells (SMC) was studied with anti‐α smooth muscle actin and anti‐desmin antibodies. By week 7, c‐kit‐immunopositive precursors formed a layer in the outer stomach wall around myenteric plexus elements. Between 9 and 11 weeks some of these precursors differentiated into ICC. ICC at the myenteric plexus level differentiated first, followed by those within the muscle layer: between SMC, at the circular and longitudinal layers, and within connective tissue septa enveloping muscle bundles. In the fourth month, all subtypes of c‐kit‐immunoreactivity ICC which are necessary for the generation of slow waves and their transfer to SMC have been developed. These results may help elucidate the origin of ICC and the aetiology and pathogenesis of stomach motility disorders in neonates and young children that are associated with absence or decreased number of these cells.     

Organization and development of pacemaker of gastrointestinal tract

Rhythmical depolarization and automatic contraction of smooth muscle of the gastrointestinal tract are consequences of pacemaker activity of c-Kit-immunoreactive cells of mesenchymal origin--interstitial Cajal cells (CC) that have a peculiar mechanism of intracellular Ca2+ exchange controlled by mitochondria. The intermuscular layer cells (ICC-MY) generate pacemaker potentials. They produce depolarization that is enhanced by unitary potentials evoked by the intermuscular population--ICC-IM. Summation of unitary potentials in time of the pacemaker ones leads to creation of the second potentials of slow waves--plateau-potentials. Due to the presence of synapse-like structures, ICC serve messengers of transmission of signals of the enteral nervous system to muscle. Long processes and tight intercellular contacts similar to the cleft ones provide transduction and coordination of excitation in the intestinal musculature. Electrical rhythmicity appears in the enteric musculature at the prenatal period in parallel with structural and functional ICC maturation, but formation of mature rhythm parameters occurs in postnatal ontogenesis. Features of similarity and differences in organization of control of heart and the gastrointestinal tract musculature by pacemakers are considered.     

日本沼虾消化道形态和组织学特点

应用石蜡切片和扫描电镜技术对日本沼虾消化道进行了研究。结果表明,食道壁向腔内形成四个纵突,食道上皮由单层柱状细胞构成,上皮下的结缔组织中具有放射肌和皮肤腺,环肌层近于连续。食道和胃连结处的管腔背方具食道瓣,胃内具胃磨、滤器和滤沟等结构,胃的组织学结构中除无皮肤腺分布外与食道相似。中肠较长,约占整个消化道的７１７％,具一对中肠前盲囊。中肠上皮细胞大致有两种类型,基膜着色深,环肌层连续,纵肌成束分散排列。后肠为一短管,管腔呈迷路状,其中部为一球形膨大的直肠。后肠的组织学结构与前肠相似。     

Telocytes express PDGFRα in the human gastrointestinal tract

Telocytes (TC), a cell population located in the connective tissue of many organs of humans and laboratory mammals, are characterized by a small cell body and extremely long and thin processes. Different TC subpopulations share unique ultrastructural features, but express different markers. In the gastrointestinal (GI) tract, cells with features of TC were seen to be CD34‐positive/c‐kit‐negative and several roles have been proposed for them. Other interstitial cell types with regulatory roles described in the gut are the c‐kit‐positive/CD34‐negative/platelet‐derived growth factor receptor α (PDGFRα)‐negative interstitial cells of Cajal (ICC) and the PDGFRα‐positive/c‐kit‐negative fibroblast‐like cells (FLC). As TC display the same features and locations of the PDGFRα‐positive cells, we investigated whether TC and PDGFRα‐positive cells could be the same cell type. PDGFRα/CD34, PDGFRα/c‐kit and CD34/c‐kit double immunolabelling was performed in full‐thickness specimens from human oesophagus, stomach and small and large intestines. All TC in the mucosa, submucosa and muscle coat were PDGFRα/CD34‐positive. TC formed a three‐dimensional network in the submucosa and in the interstitium between muscle layers, and an almost continuous layer at the submucosal borders of muscularis mucosae and circular muscle layer. Moreover, TC encircled muscle bundles, nerve structures, blood vessels, funds of gastric glands and intestinal crypts. Some TC were located within the muscle bundles, displaying the same location of ICC and running intermingled with them. ICC were c‐kit‐positive and CD34/PDGFRα‐negative. In conclusion, in the human GI tract the TC are PDGFRα‐positive and, therefore, might correspond to the FLC. We also hypothesize that in human gut, there are different TC subpopulations probably playing region‐specific roles.     

Organization and development of pacemaker of the gastrointestinal tract

Rhythmical depolarization and automatic contractions of smooth musculature of the gastrointestinal tract are a consequence of pacemaker activity of c-Kit-immunoreactive cells of mesenchymal origin—interstitial Cajal cells (ICC) that have a peculiar mechanism of intercellular Ca²⁺ balance, which is controlled by mitochondria. Intermuscular layer cells (ICC-MY) generate pacemaker potentials. Their induced depolarization is enhanced by unitary potentials generated by intracellular population—ICC-IM. Summation of unitary potentials in the tact of the pacemaker ones leads to creation of the second potential of slow waves—plateau potentials. Due to the presence of synapse-like structures, ICC serve messenger of transmission of the enteral nervous system onto the muscle. Long processes and close intercellular contacts similar to tight junction provide conductance and coordination of excitation in the intestinal musculature. Electrical rhythmicity appears in the intestinal muscle at the prenatal development period in parallel with the structural and functional ICC maturation, but establishment of mature rhythm parameters occurs in early postnatal ontogenesis. Features of similarity and difference in organization of control by pacemakers of the heart and musculature of the gastrointestinal tract are discussed.     

Function of the anal sphincters following anterior resections of rectum

The purpose of the present work is to evaluate the function of the anal sphincter following anterior resections of the rectum. Our data have shown that the anorectal reflex did not depend on the presence of the rectal mucosa. The external anal sphincter contraction involves a reflex which is initiated by stretch receptors in the levator ani muscle. The internal anal sphincter relaxation is likely a local reflex involving nerve pathways not yet completely defined.     

Caja1间质细胞与慢性假性结肠肠梗阻关系的研究进展

Cajal间质细胞（interstitial cells of cajal,ICC）主要分布在胃肠道平滑肌细胞与神经纤维之间,是一类特殊的间质细胞,它是胃肠运动的起搏细胞,具有产生、传导慢波,调节胃肠道平滑肌运动的功能。而慢性假性肠梗阻是由于胃肠神经抑制,毒素刺激或肠壁平滑肌本身病变,导致的肠壁肌肉运动功能减弱,临床上具有肠梗阻的症状和体征,但无肠内外机械性肠梗阻因素存在,故又称动力性肠梗阻。按病程有急性和慢性之分,麻痹性肠梗阻和痉挛性肠梗阻属于急性假性肠梗阻,深入研究Caja1间质细胞,对进一步认识胃肠运动的生理及胃肠动力疾病的发生机制有重要意义。     

Interstitial cells of Cajal and electrical activity in ganglionic and aganglionic colons of mice

An antibody directed against Kit protein was used to investigate the distribution of interstitial cells of Cajal (ICC) within the murine colon. The ICC density was greatest in the proximal colon and decreased along its length. The distribution of the different classes of ICC in the aganglionic colons of lethal spotted (ls/ls) mice was found to be similar in age-matched wild-type controls. There were marked differences in the electrical activities of the colons from ls/ls mutants compared with wild-type controls. In ls/ls aganglionic colons, the circular muscle was electrically quiescent compared with the spontaneous spiking electrical activity of wild-type tissues. In ls/ls aganglionic colons, postjunctional neural responses were greatly affected. Inhibitory junction potentials were absent or excitatory junction potentials inhibited by atropine were observed. In conclusion, the distribution of ICC in the ganglionic and aganglionic regions of the colons from ls/ls mutants appeared similar to that of wild-type controls. The electrical activity and neural responses of the circular layer are significantly different in aganglionic segments of ls/ls mutants.