Development of the enteric nervous system,smooth muscle and interstitial cells of Cajal in the human gastrointestinal tract

The generation of functional neuromuscular activity within the pre-natal gastrointestinal tract requires the coordinated development of enteric neurons and glial cells, concentric layers of smooth muscle and interstitial cells of Cajal (ICC). We investigated the genesis of these different cell types in human embryonic and fetal gut material ranging from weeks 4–14. Neural crest cells (NCC), labelled with antibodies against the neurotrophin receptor p75^NTR, entered the foregut at week 4, and migrated rostrocaudally to reach the terminal hindgut by week 7. Initially, these cells were loosely distributed throughout the gut mesenchyme but later coalesced to form ganglia along a rostrocaudal gradient of maturation; the myenteric plexus developed primarily in the foregut, then in the midgut, and finally in the hindgut. The submucosal plexus formed approximately 2–3 weeks after the myenteric plexus, arising from cells that migrated centripetally through the circular muscle layer from the myenteric region. Smooth muscle differentiation, as evidenced by the expression of -smooth muscle actin, followed NCC colonization of the gut within a few weeks. Gut smooth muscle also matured in a rostrocaudal direction, with a large band of -smooth muscle actin being present in the oesophagus at week 8 and in the hindgut by week 11. Circular muscle developed prior to longitudinal muscle in the intestine and colon. ICC emerged from the developing gut mesenchyme at week 9 to surround and closely appose the myenteric ganglia by week 11. By week 14, the intestine was invested with neural cells, longitudinal, circular and muscularis mucosae muscle layers, and an ICC network, giving the fetal gut a mature appearance.A.S.W. is funded by a PhD studentship awarded to A.J.B. by the Child Health Research Appeal Trust.     

Distribution of pituitary adenylyl cyclase activating peptide (PACAP) immunoreactivity in neurons of the guinea-pig digestive tract and their projections in the ileum and colon

Pituitary adenylyl cyclase activating peptide (PACAP) is a novel hypothalamic peptide that is widely distributed in neurons, including those of the gastrointestinal tract. In this study, a polyclonal antiserum directed against PACAP-27 was used to investigate the localisation of PACAP throughout the gut and to determine the projections of PACAP-immunoreactive (IR) neurons in the guinea-pig small and large intestines. PACAP-IR fibres were seen in the myenteric and submucous plexuses, in the longitudinal and circular muscle layers and around blood vessels of the submucosa throughout the gut. In both the small and large intestine, PACAP-IR cell bodies, most with Dogiel type-I morphology, were seen in the myenteric ganglia following colchicine treatment. Lesion studies (myotomy and myectomy operations) revealed that PACAP-IR interneurons projected anally in the ileum and colon. Myectomy operations resulted in a loss of PACAP-IR fibres in the circular muscle under the operation, whereas PACAP-IR fibres remained in the submucosa and around blood vessels. Following extrinsic denervation of the ileum, the number of PACAP-IR fibres in the submucosal ganglia and around blood vessels decreased. This suggests that a portion of PACAP-IR fibres supplying the submucosal ganglia and blood vessels have an extrinsic source. To investigate this, immunohistochemical studies were performed on sympathetic and dorsal root ganglia. Numerous reactive cells were seen in the dorsal root ganglia, but none was seen in sympathetic pre- or paravertebral ganglia.     

人胎大肠氮能神经元发育的研究

By using histochemical methed of NADPH-diaphorse, the development of the nitrergic neurons in the large intestine of human fetus were studied. The results showed: At the fifth month of gestation, weak positive reaction of nitric oxide synthase (NOS) appeared in part of the round cells of intermuscular ganglia. The round cells differentiated into the nitrergic nerve cells. At the sixth month, the bodies of nitrergic neurons were obviously enlarged, the processes of which were lengthened. The nitrergic nerve fibers were seen in the muscle layer, the submucosa and the base of the intestinal gland. The growth and development of nitrergic neurons reached its peak at the seventh month. The number of intermuscular ganglionic cells was increased. The density of nitrergic nerve fibers was increased in the inner circular muscle layer, and have bead-like structures. At the eighth to tenth month, the staining intensity of nitrergic neurons was increased. The myenteric plexus was densely distributed with nitrergic nerve cell bodies, whereas the submucosa and the inner circular muscle layers contained only a few neurons. The nitrergic nerve fibers were observed in all layer of large intestine, the density of the distribution of nitrergic nerve fibers was by far the highest in the inner circular muscle layer, less in the submucosa and outer longitudinal muscle layer, and only a few were found in the mucous layer. To our knowledge, it is the first time that the development of nitrergic neurons in the large intestine of human fetus was demonstrated.     

人胎大肠氮能神经元发育的研究

用NADPH-d组织化学法对人胎大肠氮能神经元的发育进行了观察.结果表明第5个月胎龄时,肌间神经节处圆形细胞中部分细胞出现一氧化氮合酶(NOS)阳性反应,并分化成氮能神经细胞.第6个月胎龄时,氮能神经元胞体增大,突起伸长,在肌层、粘膜下层和肠腺基部出现氮能神经纤维分布.第7个月胎龄时,氮能神经元生长发育达到高峰,肌间神经节细胞数目增多,环肌层神经纤维分布密度增加,膨体结构明显.第8-10个月胎龄时,氮能神经元染色强度加深,其胞体分布以肌间神经节最多,粘膜下层和内环肌层较少.氮能神经纤维的分布密度以内环肌层最高,粘膜下层和外纵肌层次之,粘膜层较低.本研究揭示了大肠氮能神经元发育的变化规律.     

Three-dimensional structures of c-Kit-positive cellular networks in the guinea pig small intestine and colon

Cryosections and whole-mount preparations of the guinea pig small intestine and colon were single or double immunolabeled using the anti-c-Kit and protein gene product 9.5 antibodies. Immunolabeled specimens were observed under a confocal laser scanning microscope. The main findings of the present study are: (1) the distribution and profiles of three-dimensional structures of c-Kit-positive cellular networks in the small intestine and colon, and (2) the anatomical relations of c-Kit-positive cells to the enteric nerves in the layers. In the small intestine, c-Kit-positive cellular networks were observed at levels of the deep muscular plexus and myenteric plexus. The c-Kit-positive cellular networks ran along or overlay the nerve fibers at the deep muscular plexus, while they showed the reticular structures intermingled with the nerve elements at the myenteric plexus. In the colon, c-Kit-positive cellular networks were observed at levels of the submuscular plexus and myenteric plexus, and were further identified within the circular and longitudinal muscle layers as well as in the subserosal layer. In the circular muscle layer, c-Kit-positive cells surrounded the associated nerve fibers and extended several long processes toward the adjacent c-Kit-positive cells. The c-Kit-positive cellular networks within the longitudinal muscle layer as well as in the subserosal layer were not associated with the nerve fibers. In the layers of the intestinal wall with c-Kit-positive cells, the cellular networks of the interstitial cells were identified in ultrastructure. The characteristic profiles of c-Kit-positive cellular networks provide a morphological basis upon which to investigate the mechanisms regulating intestinal movement. Received: 14 July 1998 / Accepted: 2 September 1998     

胚胎小肠Cajal细胞的发育研究

目的研究人胚胎小肠cajal细胞的发育变化规律。方法采用全层铺片结合切片的免疫细胞化学技术。结果Cajal细胞呈酪氨酸激酶受体(Kit)和波形蛋白(vinlentin)免疫反应阳性。在胚胎发育早期,cajal细胞较少,为单层,稀疏分布于肌间神经丛周围,细胞为梭形,可见两个短而小的突起,未见分支;随着胎龄的增加,Cajal细胞数量增多,胞体增大,突起伸长,并出现分支。此时,肌间神经丛周围的Cajal细胞出现两层,其长轴彼此垂直,分别平行于环行肌和纵行肌。与此同时环行肌层内亦可见少许Cajal细胞;出生前,肌间神经丛部位的Cajal细胞接近成熟,两层细胞的突起进一步增多、伸长,彼此间形成与成人相似的完整的细胞网络。此时深肌丛附近亦可见少量Cajal细胞。结论人的小肠Cajal细胞发育有一定的时间顺序,即肌间神经丛周围最先出现,肌内次之,深肌丛较晚,出生前肌间神经丛周围的Cajal细胞已经接近成熟。这种发育演变若发生异常,可能导致某些胃肠动力障碍性疾病。     

Scanning electron microscopy of the muscle coat of the guinea-pig small intestine

Summary We have studied the layers of the muscular coat of the guinea-pig small intestine after enzymatic and chemical removal of extracellular connective tissue. The cells of the longitudinal muscle layer are wider, have rougher surfaces, more finger-like processes and more complex terminations, but fewer intercellular junctions than cells in the circular muscle layer. A special layer of wide, flat cells with a dense innervation exists at the inner margin of the circular muscle layer, facing the submucosa. The ganglia of the myenteric and submucosal plexuses are covered by a smooth basal lamina, a delicate feltwork of collagen fibrils, and innumerable connective tissue cells. The neuronal and glial cell processes at the surface of ganglia form an interlocking mosaic, which is loosely packed in newborn and young animals, but becomes tightly packed in adults. The arrangement of glial cells becomes progressively looser along finer nerve bundles. Single varicose nerve fibres are rarely exposed, but multiaxonal bundles are common. Fibroblast-like cells of characteristic shape and orientation are found in the serosa; around nerve ganglia; in the intermuscular connective tissue layer and in the circular muscle, where they bridge nerve bundles and muscle cells; at the submucosal face of the special, flattened inner circular muscle layer; and in the submucosa. Some of these fibroblast like cells correspond to interstitial cells of Cajal. Other structures readily visualized by scanning electron microscopy are blood and lymphatic vessels and their periendothelial cells. The relationship of cellular elements to connective tissue was studied with three different preparative procedures: (1) freeze-cracked specimens of intact, undigested intestine; (2) stretch preparations of longitudinal muscle with adhering myenteric plexus; (3) sheets of submucosal collagen bundles from which all cellular elements had been removed by prolonged detergent extraction.     

An immunohistochemical study of the innervation of the large intestine of the toad (Bufo marinus)

The distribution of intrinsic enteric neurons and extrinsic autonomic and sensory neurons in the large intestine of the toad, Bufo marinus, was examined using immunohistochemistry and glyoxylic acid-induced fluoresecence. Three populations of extrinsic nerves were found: unipolar neurons with morphology and location typical of parasympathetic postganglionic neurons containing immunoreactivity to galanin, somatostatin and 5-hydroxytryptamine were present in longitudinally running nerve trunks in the posterior large intestine and projected to the muscle layers and myenteric plexus throughout the large intestine. Sympathetic adrenergic fibres supplied a dense innervation to the circular muscle layer, myenteric plexus and blood vessels. Axons containing colocalized calcitonin gene-related peptide immunoractivity and substance P immunoreactivity distributed to all layers of the large intestine and are thought to be axons of primary afferent neurons. Five populations of enteric neurons were found. These contained immunoreactivity to vasoactive intestinal peptide, which distributed to all layers of the large intestine; galanin/vasoactive intestinal peptide, which projected to the submucosa and mucosa; calcitonin gene-related peptide/vasoactive intestinal peptide, which supplied the circular muscle, submucosa and mucosa; galanin, which projected to the submucosa and mucosa; and enkephalin, which supplied the circular muscle layer.     

The cross-ply arrangement of collagen fibres in the submucosa of the mammalian small intestine

Summary Whole-mount preparations of the submucosa were made from the small intestine of rats, guinea-pigs, rabbits and sheep. In the distended intestine the collagen fibres ran straight and approximately parallel to the serosal surface. They formed a characteristic lattice, with two arrays of fibres running diagonally in a clockwise and an anticlockwise direction, and making an angle of 50°–55° with the longitudinal axis of the intestine. This collagenfibre lattice was flexible and changed with the movements of the intestinal wall; when the radial distension predominated, the angle between collagen fibres of the submucosa and longitudinal axis of the intestine increased to 60°–65°, and when the longitudinal distension predominated the angle decreased to about 30°.     

Evidence that myenteric neurons of the gastric corpus project to both the mucosa and the external muscle: myectomy operations on the canine stomach

Summary The distribution of nerve cell bodies and fibres in the canine stomach was investigated using antibodies to the general neuronal marker, neuron-specific enolase. Prominent ganglia containing many reactive nerve cells were found in the myenteric plexus of the gastric corpus and antrum. Nerve cells were absent from the submucosa of the corpus and were extremely rare in the antrum. Renoval of areas of longitudinal muscle and myenteric plexus from the corpus (myectomy), with 7 days allowed for axon degeneration, resulted in the loss of fibres reactive for galanin, gastrin-releasing peptide, substance P and vasoactive intestinal peptide from both the circular muscle and mucosa in the area covered by the lesion. Combined vagotomy and sympathetic denervation did not significantly affect these fibres, but did cause fibres reactive for calcitonin gene-related peptide to degenerate. It is concluded that the myenteric plexus of the gastric corpus, like the myenteric plexus of the small intestine and colon, is the source of nerve fibres innervating the circular muscle, but, in contrast to other regions of the gastrointestinal tract, myenteric ganglia, not submucous ganglia, are the major, or sole, source of the intrinsic innervation of the mucosa.     

Neuroendocrine system of the digestive tract in Rhamdia quelen juvenile: an immunohistochemical study

In this work, an immunohistochemical study was performed to determine the distribution and relative frequencies of some neuromodulators of the digestive tract of silver catfish (Rhamdia quelen). The digestive tract of silver catfish was divided into six portions; the oesophagus, stomach, intestine (ascendant, descendant and convoluted segments), and rectum. Immunohistochemical method using a pool of specific antisera against-gastrin, -cholecystokinin-8, -leu-enkephalin, -neuropeptide Y, -calcitonin gene-related peptide (CGRP), and -vasoactive intestinal peptide (VIP) was employed. Immunoreactivity to all antisera was identified in neuroendocrine cells (NECs) localized in the gut epithelium, although no reaction was observed in the oesophagus or stomach. The morphology of NECs immunopositive to each antibody was similar. They were slender in shape, with basally located nucleus, and their main axis perpendicular to the basement membrane. The number of NECs immunoreactive to all antisera was higher in the ascendant and descendant intestine, exhibiting a decreasing trend toward distal segments of the gut. In addition, immunoreactivity to CGRP and VIP was observed in the myenteric plexus and nerve fibers distributed in the mucosal, submucosal and muscular layers. The higher number of immunopositive NECs in the ascendant and descendant intestine may indicate the primary role of these segments in the control of food intake by means of orexigenic and anorexigenic peripheral signals.     

Somatostatin in human enteric nerves

Summary Somatostatin-immunoreactive nerves and endocrine cells were localized by use of immunohistochemistry in human stomach, small and large intestine. The nature of the immunoreactivity in acid extracts of separated layers of intestine was determined with separation by high pressure liquid chromatography followed by detection with radioimmunoassay; authentic somatostatin-14 was found in the external musculature, which contains nerves, and in the submucosa and mucosa, which contain both nerve fibres and endocrine cells.The distribution of somatostatin nerves in the gastric antrum, duodenum, jejunum, ileum, ascending and sigmoid colon, and rectum is described. In the intestine many positive perikarya and fine varicose fibres were seen. Mucosal fibres formed a sub-epithelial plexus and a looser network in the lamina propria; this nerve supply was less dense in the large intestine. Submucous ganglia contained positive perikarya and terminals; many terminals formed pericellular baskets, mainly around non-reactive cells. A small number of nerve fibres were associated with submucosal blood vessels. The innervation of the circular and longitudinal muscle was sparse. Positive nerve terminals were seen in the myenteric plexus, although fewer than in the submucous ganglia; positive perikarya were scarce in myenteric ganglia. Somatostatin-immunoreactive nerves were found in the muscle layers and myenteric plexus of the gastric antrum, but were not detected in the antral mucosa and all layers of the gastric body.The distribution of human enteric somatostatin nerves is compared to that in small laboratory animals, and possible roles for these nerves are discussed.     

Different distribution of S-100 protein and glial fibrillary acidic protein (GFAP) immunoreactive cells and their relations with nitrergic neurons in the human fetal small intestine.

The appearance, distribution and some histochemical features of non-neuronal cells (NN cells) associated with the myenteric plexus of human fetal small intestine have been studied by means of S-100 protein and GFAP immunocytochemistry between the 10th and 17th week of gestation. In addition, double labelling immunocytochemistry using an antibody raised against a constitutive isoform of nitric oxide synthase (bNOS) in combination with an S-100 protein antibody was applied to investigate the morphological relations between NN cells and nitrergic neurons in the developing gut wall. Cells with immunoreactivity for both glial-specific proteins are already present in the 10th week of gestation. While cells with S-100 protein immunoreactivity are located within the circular muscle layer as well as in the myenteric, and submucous plexuses, cells with GFAP immunopositivity are mainly restricted to the side of the myenteric plexus adjacent to the longitudinal muscle layer. In contrast to the dense network formed by S-100 protein immunopositive structures the GFAP immunopositive cells appear singly and do not connect into a network. Double-labelling immunocytochemistry reveals nitrergic fibers (NOS-IR) in close relation to the S-100 protein immunoreactive glial network. NOS-IR varicosities are in close association with the surface of those cells both in the circular muscle layer (CM) and in the tertiary plexus. It is concluded that two populations of NN cells with different locations and different immunohistochemical characters appear and develop together with the enteric ganglia in the human fetal intestine. The close morphological relation of NOS-IR fibers with S-100 protein immunopositive cellular network indicate a functional relationship between S-100 protein immunopositive cells and the nitrergic nerves during the early development of human enteric nervous system (ENS).     

Projections of neurons with neuromedin U-like immunoreactivity in the small intestine of the guinea-pig

Summary Neuromedin U immunoreactivity was located histochemically in the guinea-pig small intestine. Projections of immunoreactive neurons were determined by analysing patterns of degeneration following nerve lesions. The co-localization of neuromedin U immunoreactivity with immunoreactivity for substance P, neuropeptide Y, vasoactive intestinal peptide and calbindin was also investigated. Neuromedin U immunoreactivity was found in nerve cells in the myenteric and submucous plexuses and in nerve fibres in these ganglionated plexuses, around submucous arterioles and in the mucosa. Reactive fibres did not supply the muscle layers. Most reactive nerve cells in the myenteric ganglia had Dogiel type-II morphology and in many there was co-localization of calbindin, although some Dogiel type-II neuromedin U neurons were calbindin negative. Lesion studies suggest that these myenteric neurons project circumferentially to local myenteric ganglia. Projections from myenteric neurons also run anally in the myenteric plexus, while other projections extend to submucous ganglia, and still further projections run from the intestine to provide terminals in the coeliac ganglia. In the submucous ganglia neuromedin U was co-localized in three populations of nerve cells: (i) those with vasoactive intestinal peptide immunoreactivity, (ii) neurons containing neuropeptide Y, and (iii) neurons containing substance P. Each of these populations sends nerve fibres to the mucosa. Neuromedin U immunoreactivity is thus located in a variety of neurons serving different functions in the intestine and therefore probably does not have a single role in intestinal physiology.     

Evaluation of the total number of myenteric neurons in the developing chicken gut using cuprolinic blue histochemical staining and neurofilament immunocytochemistry

The aim of this study was to find an improved method with which to stain the entire population of myenteric neurons in the different segments of the developing chicken intestine. Histochemical staining with cuprolinic blue (quinolinic phthalocyanine) and immunostaining against neurofilament (NF) were performed on whole mounts prepared from intestinal segments of embryonic (day 19 of incubation) and hatched (1, 2, 4 and 7 days after hatching) chickens. Double labelling was performed to evaluate to what extent the two markers visualise the same nerve cell population. Cuprolinic blue stained neuronal somata highly selectively, whereas processes and glia cells were poorly labelled. The cuprolinic blue-positive neurons were uniform in shape. NF immunostaining revealed a morphologically highly variable neuron population. Double labelling with cuprolinic blue and NF resulted in an intensification of both stainings, allowing an accurate morphological classification of NF-stained myenteric neurons. Data obtained from the counting of cuprolinic blue-positive neurons were subjected to two-way ANOVA and the Tukey probe. The densities of ganglia and neurons were found to decrease, and the mean number of neurons per myenteric ganglion to increase, with different dynamics along the longitudinal axis of the gut during the examined time span. The variances in the number of NF-positive neurons were not homogeneous, and the data were therefore not suitable for ANOVA. Accordingly, only semiquantitative conclusions could be drawn.     

Role of the "little brain" in the gut in water and electrolyte homeostasis

The enteric nervous system plays a key role in maintenance of body fluid homeostasis by regulating the transport of ions by the intestinal epithelium. The epithelial cells normally absorb large volumes of fluid and ions daily, but tonically active submucosal neurons continuously suppress ion transport and limit the absorptive capacity of the intestine. Specialized nerve endings detect chemical, osmotic, or thermal alterations of the luminal contents or mechanical activity of the gut wall and encode this information as action potentials that propagate along nerve processes to the ganglia. Information transfer within the ganglia occurs at nicotinic cholinergic or other synapses. Ion transport is altered when neurotransmitters released from motor neurons interact with receptors on epithelial cells to initiate stimulus-response coupling. The signals that transduce changes in epithelial ion transport are largely unknown, except for acetylcholine, but may include vasoactive intestinal peptide or other peptides. These trigger changes in intracellular messengers that influence the state of ionic channels in the epithelial cells and thereby inhibit absorptive processes or stimulate secretory mechanisms. When conservation of salt and water is necessary, command signals from the central nervous system, and perhaps from the myenteric ganglia, will shut down the synaptic circuits in the submucosal ganglia and enhance the absorptive capacity of the bowel.     

The source of the nerve fibres forming the deep muscular and circular muscle plexuses in the small intestine of the guinea-pig

Summary A quantitative ultrastructural study was made of the neuntes forming the deep muscular and circular muscle plexuses of the guinea-pig small intestine following microsurgical lesions designed to interrupt intrinsic and extrinsic nerve pathways within the intestinal wall. Removal of a collar of longitudinal muscle with attached myenteric plexus from the circumference of a segment of small intestine resulted in the subsequent disappearance of 99.3% of neurites in the underlying circular muscle. The few surviving neurites in the deep muscular plexus and circular muscle disappeared completely from lesioned segments that were, in addition, extrinsically denervated surgically. These results indicate that the majority of nerve fibres in the deep muscular and circular muscle plexuses of the guinea-pig small intestine is intrinsic to the intestine and originates from nerve cell bodies located in the overlying myenteric plexus. At the light-microscopic level, nerve bundles were traced from the myenteric plexus to the circular muscle.     

The concentric structure of the developing gut is regulated by Sonic hedgehog derived from endodermal epithelium

The embryonic gut of vertebrates consists of endodermal epithelium, surrounding mesenchyme derived from splanchnic mesoderm and enteric neuronal components derived from neural crest cells. During gut organogenesis, the mesenchyme differentiates into distinct concentric layers around the endodermal epithelium forming the lamina propria, muscularis mucosae, submucosa and lamina muscularis (the smooth muscle layer). The smooth muscle layer and enteric plexus are formed at the outermost part of the gut, always some distance away from the epithelium. How this topographical organization of gut mesenchyme is established is largely unknown. Here we show the following: (1) Endodermal epithelium inhibits differentiation of smooth muscle and enteric neurons in adjacent mesenchyme. (2) Endodermal epithelium activates expression of patched and BMP4 in adjacent non-smooth muscle mesenchyme, which later differentiates into the lamina propria and submucosa. (3) Sonic hedgehog (Shh) is expressed in endodermal epithelium and disruption of Shh-signaling by cyclopamine induces differentiation of smooth muscle and a large number of neurons even in the area adjacent to epithelium. (4) Shh can mimic the effect of endodermal epithelium on the concentric stratification of the gut. Taken together, these data suggest that endoderm-derived Shh is responsible for the patterning across the radial axis of the gut through induction of inner components and inhibition of outer components, such as smooth muscle and enteric neurons.     

Distribution and projections of neurons with immunoreactivity for both gastrin-releasing peptide and bombesin in the guinea-pig small intestine

Summary Bombesin-like and gastrin-releasing peptide (GRP)-like immunoreactivities were localized in nerves of the guinea-pig small intestine and celiac ganglion with the use of antibodies raised against the synthetic peptides. The anti-bombesin serum (preincubated to avoid cross reactivity with substance P) and the anti-GRP serum revealed the same population of neurons. Preincubation of the antibombesin serum with bombesin abolished the immunoreactivity in nerves while absorption of the anti-GRP serum with either bombesin or the 14–27 C-terminal of GRP only reduced the immunoreactivity. The immunoreactivity was abolished by incubation with GRP 1–27.Immunoreactive nerves were found in the myenteric plexus, circular muscle, submucous plexus and in the celiac ganglion. Faintly reactive nerve cell bodies were found in the myenteric ganglia (3.2% of all neurons) but not in submucous ganglia. After all ascending and descending pathways in the myenteric plexus had been cut, reactive terminals disappeared in the myenteric plexus, circular muscle (including the deep muscular plexus) and the submucous plexus on the anal side. After the mesenteric nerves were cut no changes were observed in the intestinal wall but the reactive fibres in celiac ganglia disappeared. It is deduced that GRP/bombesin-immunoreactive nerve cell bodies in myenteric ganglia project from the myenteric plexus to other myenteric ganglia situated further anally (average length 12 mm), anally to the circular muscle (average length 9 mm), anally to submucous ganglia (average length 13 mm) and external to the intestine to the celiac ganglia.It is concluded that the GRP/bombesin-reactive neurons in the intestinal wall represent a distinct population of enteric neurons likely to be involved in controlling motility and in the coordination of other intestinal functions.     

Immunohistochemical study of 5-HT-containing neurons in the teleost intestine: relationship to the presence of enterochromaffin cells

Summary The formaldehyde-induced fluorescence technique had shown 5-hydroxytryptamine-containing enteric neurons in the intestine of the teleost Platycephalus bassensis, but did not reveal such neurons in the intestine of Tetractenos glaber or Anguilla australis. Re-examination of these animals with 5-hydroxytryptamine immunohistochemistry showed immunoreactive enteric neurons in the intestine of all three teleost species. The 5-hydroxytryptamine-containing enteric neurons showed essentially the same morphology in all species examined: the somata were situated in the myenteric plexus, extending down into the circular muscle layer, but none were found in the submucosa; processes were found in the myenteric plexus, the circular muscle layer and the lamina propria. It was concluded that the neurons may innervate the muscle layers or the mucosal epithelium, but were unlikely to be interneurons. In a range of teleosts, enterochromaffin cells were found in the intestine of only those species in which the formaldehyde technique did not visualize neuronal 5-hydroxytryptamine. Available evidence suggests that, in vertebrates, 5-HT-containing enterochromaffin cells are lacking only where there is an innervation of the gut mucosa by nerve fibres containing high concentrations of 5-HT.