Subepithelial Network Research Articles

Expression of vasoactive intestinal peptide, calcitonin gene-related peptide, substance P, and intermediate neurofilaments in nasal mucosal nerve fibers of horses without nasal disease.

To determine the distribution of nerve fibers containing calcitonin gene-related peptide (CGRP), substance P (SP), vasoactive intestinal peptide (VIP), and intermediate neurofilaments in nasal mucosa of horses. 6 horses without evidence of nasal disease. Full-thickness nasal tissue specimens were obtained from the rostral portion of the nasal septum at necropsy, and fluorescence immunohistochemistry was performed to assess mucosal distribution of nerve fibers. Nerve fibers with CGRP-like immunoreactivity (CGRP-Li) formed a dense subepithelial network, and a large number of fibers were found coursing between epithelial cells. Fibers with CGRP-Li were also associated with blood vessels and mucous glands. Fibers with SP-like immunoreactivity (SP-Li) had a similar distribution and density. In contrast, there were few fibers with VIP-like immunoreactivity. Fibers containing intermediate neurofilaments were prominent and appeared as large nerve fiber bundles mainly adjacent to the nasal septum but also close to mucous glands and within the lamina propria. Intermediate neurofilaments were also identified in single nerve fibers at all sites, but the density of fibers with intermediate neurofilaments did not match that of fibers with CGRP- or SP-Li. The density and distribution of nerve fibers containing SP- or CGRP-Li in nasal mucosa of horses was similar to that reported for other species. However, expression of VIP in nerve fibers was low. Antibodies against intermediate neurofilaments identified many nerve fibers in nasal mucosa of horses but did not appear to identify small diameter fibers expressing SP or VIP.

Physiological role of the secondary circulatory system in fish

Most studies of the secondary circulation or the “secondary vascular system” (Vogel 1981; Vogel and Claviez 1981) have concentrated on descriptions of morphology, rather than function, among various species (see Steffensen and Lomholt 1992). The scarce available evidence at this time characterizes the secondary vascular system in fishes as being a pool of blood which is low in pressure, low in red cell content, absent from the muscle tissues, associated with both the inner and outer surfaces of body including the gills, and of a volume that approaches or exceeds that of the primary circulation. Vogel (1985a) demonstrated the occurrence of an extremely dense capillary network of the secondary system over the scales of trout and tilapia. Satchell (1991) pointed out that similar vascularization on the scales had been observed for both freshwater and seawater teleosts in old anatomical literatures (Jakubowski 1960a,b, 1989; Kaczmarski 1966; Tyszkiewicz 1969), while it is not always clear whether such capillary networks belong to either the primary or secondary vascular system. A similar subepithelial vascular network has also been observed in the Atlantic hagfish Myxine glutinosa (Lametschwandtner et al. 1989). Vogel (1985a) also stated that secondary capillary networks are distributed in the mucous membrane of the mouth and the peritoneum, and possibly in the intestinal wall. The central venous sinus (CVS) which is located within the primary lamella of the gill is currently considered to be part of the secondary vascular system (Vogel 1985a).

An immunohistochemical study of the ontogeny of the neuroendocrine system in the chicken oesophagus.

The ontogenesis and distribution of serotonin-, chromogranin A-, chromogranin B-, galanin-, neurotensin-, bombesin- and neuropeptide Y-immunoreactive elements were studied in the chicken oesophagus during pre- and post-hatching life. Unlike positive nerve elements, that were present in pre- and post-hatching life, positive endocrine cells were observed only during embryonic life in the oesophageal epithelium. The first endocrine cells, immunoreactive for serotonin and chromogranins, appeared on day 12, in the cervical and thoracic portions of the oesophagus. At the same age, but only in its distal portion, a few bombesin- and neurotensin-immunoreactive cells also appeared. The number of the endocrine cells progressively increased, reaching a maximum on day 15. They then decreased, with a cranio-caudal progression, until they disappeared a few days after hatching. Almost all the serotonin-immunoreactive cells but only a subpopulation of bombesin- and neurotensin-immunoreactive cells colocalized chromogranins. About half of this subpopulation also colocalized serotonin. All these cells reacted positively with Grimelius argyrophile stain. The mucosa of the crop never contained positive endocrine cells. Positive nervous elements appeared first in the wall of the terminal oesophagus and only one or two days later in the proximal oesophagus including the crop. Nervous elements immunoreactive for galanin first appeared from days 6 to 7, for neurotensin from days 7 to 8, for neuropeptide Y from 13 to 15 and for bombesin from 15 to 18. At day 15 galanin-immunoreactive ganglionic cells and fibres occupied both the myenteric and submucous plexus and galanin-positive nerve fibres could be seen throughout the oesophageal wall from the adventitia to a thin subepithelial network. Neurotensin- and neuropeptide Y-immunopositive ganglionic cells and fibres, by contrast, invariably occupied the muscular and submucous layers. Scattered bombesin-immunoreactive ganglionic cells were observed only in the myenteric plexus. The number of positive nerve elements progressively increased until some weeks after birth. Density and intensity were always much higher for galanin and neurotensin than for neuropeptide Y and bombesin.

Microvascular anatomy of the nose.

The microvasculature of the nose consists of: 1) A dense subepithelial network of capillaries, with fenestrations between the endothelial cells. This network provides nutrients to the epithelium and glands, and allows passage of water into the lumen for evaporation and air-conditioning. 2) A system of capacitance vessels or sinuses, which when they distend, block the nasal lumen, and when they empty, open the nasal passages. Changes in their volume will affect the filtering and air-conditioning functions of the nose. 3) Arteriovenous anastomoses which allow rapid passage of blood through the mucosa. They are probably important in air-conditioning, and in the countercurrent mechanisms that tend to keep the brain cool in a hot dry climate. The anatomical interrelationships between these different systems is not well understood, nor is their differential control in terms of actions of mediators and nerves. In neurogenic inflammation sensory nerves are excited and release local mediators such as substance P via axon reflexes. These sensory neuropeptides will cause vasodilatation, vascular congestion and extravasation of liquid from the postcapillary venules, with resultant oedema and exudate. They may also cause secretion from the submucosal glands.

Microvascular adaptation to transmucosal implants. A scanning electron microscopic study in the rat.

The profuse gingival microvasculature surrounding natural teeth has a unique architecture that contributes to the biologic seal and the anti-infective defense in the sulcular region. This study tested the hypothesis that this characteristic vascular arrangement is not replicated when gingival tissues adapt to a transmucosal implant. In 20 4-week-old Wistar rats, the upper first molar was extracted bilaterally and the gingiva at this site excised. Four weeks later, a 5-mm-long titanium screw was implanted at the extraction site on one side. After another 4 weeks, the rats were injected with liquid plastic resin through the carotid arteries and vascular casts prepared for scanning electron microscopy. The results showed that within 4 weeks after implant insertion, the microvasculature of the regenerated soft tissues immediately surrounding the implant had formed a characteristic arrangement that consisted of a dense subepithelial network as well as an orderly array of vertically oriented capillary loops. The density and diameter of the capillaries varied within a wide range, possibly related to the extent of healing and/or inflammatory conditions. On the contralateral, edentulous side, the capillaries formed a flattened meshwork showing a conspicuous absence of capillary loops. This study indicates that, when a transmucosal implant is inserted in an edentulous area, the mucosal microvasculature adapts and forms an arrangement very similar to that surrounding natural teeth.

Immunohistochemical studies of the neuroanatomy of the human turbinates: innervation pattern of seromucous glands

Seromucous glands are among the main components of human nasal mucosa. To control the different physiological functions of these glands, a dense nerval network is necessary. The aim of this study was to demonstrate the general innervation of the seromucous glands in nasal mucosa. Tissue samples of inferior human turbinates were fixed and embedded in paraffin wax or frozen. Serial sections were performed and incubated with antibodies either to neuron-specific enolase (NSE) or S-100 protein. The ABC method was employed to demonstrate the immunacomplexes. The sections were counterstained with haemotoxylin. Either NSE and S-100 protein-immunoreactive nerve fibres were found around the acini, ducts and in the connective tissue of the glands. Furthermore, a dense network of fine nerve fibres was detected in the submucosal region. The localisation of neurons in nasal glands confirms the direct nerval control of the diverse glandular functions. Additionally, the sensitive subepithelial network of fine nerve fibres might be involved in the regulation of glandular secretion.

Independent control of mucosal and total airway blood flow during hypoxemia

In the larger airways, the blood circulation forms a subepithelial (mucosal) and outer (peribronchial) microvascular network. This raises the possibility that blood flow in these two networks is regulated independently. We used hypoxemia as a stimulus to induce changes in tracheal mucosal blood flow normalized for systemic arterial pressure (Qtr n) measured with an inert soluble gas technique and total bronchial blood flow (Qbr) and normalized Qbr (Qbrn) measured with an electromagnetic flow probe in anesthetized sheep. Fifteen minutes of hypoxemia [PO2 40 +/- 7 (SD) Torr] decreased mean Qtr n from 1.1 +/- 0.4 to 0.8 +/- 0.4 ml.min-1.mmHg-1.10(2) (-27%; P less than 0.05; n = 7) and increased mean Qbr n from 12.1 +/- 3.2 to 17.1 +/- 5.4 ml.min-1.mmHg-1.10(2) (+41%; P less than 0.05; n = 6). The rise in Qbr correlated with cardiac output (r = 0.68; P less than 0.05). Phentolamine pretreatment (0.1 mg/kg iv) blunted the hypoxemia-related decrease of mean Qtr n (-8%; P = NS). Tyramine (2.5 mg) applied locally to the trachea decreased mean Qtr n significantly after 30 and 45 min by 31 and 19%, respectively (P less than 0.05). 6-Hydroxydopamine (0.2 mg 4 times for 1 h locally applied) prevented the hypoxemia-induced as well as local tyramine-induced decrease in mean Qtr n (0 and 0%).(ABSTRACT TRUNCATED AT 250 WORDS)

Villus contraction aids repair of intestinal epithelium after injury.

A highly reproducible in vitro model of intestinal epithelial injury in guinea pig ileum was used to study the structural and functional events that accompany rapid epithelial repair. This model is characterized by denudation of the villus tip followed by rapid restitution of the epithelial barrier. Using standard electrophysiological and quantitative morphometric techniques, we found that immediately after injury the number of cells lost exceeded the number of empty cell positions on the denuded basement membrane by 100%. Concurrently, cytoplasmic processes of subepithelial myofibroblasts contained condensations of microfilaments that were not apparent in controls. Additionally, villus height was diminished immediately after injury, and progressively decreased during the restitution period. In tissues that were depleted of ATP using the uncoupler dinitrophenol and in tissues functionally denervated by tetrodotoxin, villus shortening after injury was significantly reduced. Denervation also retarded functionally and structurally defined reestablishment of epithelial barrier function. These data suggest that intestinal epithelial repair is aided by energy-dependent, neurally mediated villus contraction. We speculate that the subepithelial network of myofibroblasts is responsible for this process, which effectively minimizes the denuded surface area to be reepithelialized.

Histologic Study of the Growth of the Subglottis after Interruption of the Circular Structure of the Cricoid

In a previous study in growing rabbits it was demonstrated that after resection of a ventral part of the cricoid the lumen of the subglottic airway developed normal size during further growth up to the adult stage. Histologic investigations of these cases suggest that a subepithelial network of elastic fibres, ventrally suspended to the thyroid and the tracheal rings, ensures the airway lumen and prevents the development of a subglottic stenosis. Resection of a ventral part of the cricoid including the adjacent soft tissue lining of the airway causes an irreversible loss of the supporting elastic layer and the formation of scar tissue resulting in a stenosis. It was concluded that the larynx and trachea can be considered as an inner, elastic tube (= conus elasticus) lined with epithelium, suspended to an outer, segmented cartilaginous tube. At the level of the subglottis an intact conus elasticus is of more importance for the normal development of the airway lumen in young rabbits than an intact ventral half of the cricoid ring.

Neovascularity of a tracheal prosthesis/tissue complex

Permanent bioincorporation of a microporous tracheal prosthesis will require a stable blood supply to connective tissue supporting an epithelial surface. In experience with over 80 tracheal implants in dogs, we have observed that end-on ingrowth and epithelialization does not occur in the absence of lateral ingrowth, epithelialization is marked by the appearance of a subepithelial network of vessels, and this process must be well advanced by 6 to 8 weeks for long-term stability. These observations were extended by using microangiography to delineate the blood supply of the prosthesis/tissue complex. Six implants of bioelectric polyurethane with 10% gentamicin (3 cm length, 2 cm diameter, 1 to 1.25 mm wall thickness, 60 to 120 mu micropore diameter) were interposed in the dog thoracic trachea and wrapped with an omental pedicle. The aorta was perfused with a barium suspension at elective sacrifice between 10 weeks and 21 months. Radiographs of specimens were correlated with bronchoscopic, gross, and histopathological findings. Neovascularity of the prosthesis/tissue complex can be described in three categories: outer capsule, prosthetic wall, and inner lining. Outer capsule vessels were oriented circumferentially immediately adjacent to the prosthetic wall. They resembled arteries up to 75 mu diameter on microscopy and appeared to develop from the omentum with connections developing to the bronchial circulation. Prosthetic wall vessels up to 75 mu with thin muscular walls were noted to traverse the porous prosthetic wall. The inner lining had a network of subepithelial vessels that connected to the lamina propria vasculature of the native trachea across the anastomoses with vessels up to 120 mu in diameter. We conclude that the omentum provides an immediate blood supply and a base for early connective tissue ingrowth. Epithelialization occurs as early as 3 weeks on the favorable bed, accompanied by vascular connections to the existing lamina propria tracheal vessels. This dual organization of blood supply with connections across the prosthetic wall is probably important to long-term stability of healing.

The origins, pathways and terminations of neurons with VIP-like immunoreactivity in the guinea-pig small intestine

We have analyzed changes in the distributions of terminals with vasoactive intestinal polypeptide (VIP)-like immunoreactivity, and accumulations in severed processes, that occur after lesions of intrinsic and extrinsic nerve pathways of the guinea-pig small intestine. The observations indicate that enteric vasoactive intestinal polypeptide immunoreactive neurons have the following projections. Nerve cell bodies in the myenteric plexus provide varicose processes to the underlying circular muscle; the majority of these pathways, if they extend at all in the anal or oral directions, do so for distances of less than 1 mm. Nerve cell bodies of the myenteric plexus also project anally to provide terminals to other myenteric ganglia. The lengths of the majority of these projections are between 2 and 10 mm, with an average length of about 6 mm. Processes of myenteric neurons also run anally in the myenteric plexus and then penetrate the circular muscle to provide varicose processes in the submucous ganglia at distances of up to 15 mm, the average length being 9–12 mm. In addition, there is an intestinofugal projection of myenteric neurons whose processes end around nerve cell bodies of the coeliac ganglia. A similar projection from the colon supplies the inferior mesenteric ganglia. The nerve cell bodies in submucous ganglia give rise to a subepithelial network of fibres in the mucosa and also supply terminals to submucous arterioles. It is concluded that vasoactive intestinal polypeptide is contained in neurons of a number of intrinsic nerve pathways, influencing motility, blood flow and mucosal transport. The myenteric neurons that project to prevertebral sympathetic ganglia may be involved in intestino-intestinal reflexes.

Neurohistological observations on the oesophageal innervation of rabbit.

The vegetative, motor, and sensitive innervation of the oesophagus of rabbit was studied by means of several neurohistological techniques. A great deal of vegetative and somatic nervous formations were found and described within the various segments of the organ; namely at level of the cervical, thoracic, prediaphragmatic, and abdominal oesophageal tracts. In particular, isolated and grouped ganglion cells, interstitial and associative neurons, free nervous terminations, and an amyelinated subepithelial network sending delicate fibrils to the basal layers of the impending epithelium were described. The vegetative nervous component is organized into an extramural oesophageal plexus, and into an intramural one. The numerous motor endplates lying on the striated muscle fibres show different forms and several other structural pecularities. The sensitive terminations are represented by simple and non-capsulated Ruffini's corpuscles contained within the submucous connective tissue. The possible functional correlations of these morphological findings are discussed.

THE CERIANTHAR1AN NERVOUS SYSTEM

Evidence supporting the presence of nerve cells in the column and tentacles of Pacliycerianthiis is described. G. F. Gwilliam has shown that electrical stimuli can be transmitted to the ectodermal muscle by the intact epithelium and subepithelial network of the ectoderm of the column. In these preparations the ectodermal muscle, mesogloea, and endoderm were cut. In contrast, preparations in which the ectodermal muscle has been left intact and the epithelium and subepithelial network cut do not show such transmission. The author and Gwilliam have independently used different silver stain methods to demonstrate large cells, piobably nerve cells, with cell bodies in the base of the epithelium and with fibers running into the subepithelial network. The author has found similar bipolar cells in the tentacles and column by using maceration techniques. These cells are compared with other cell types found in the tentacles.

Nerve plexuses in the oral mucosa

Oral nerve plexuses were studied in the mucosa of the tongue, hard palate, cheek and gingiva of rabbits, cats and monkeys. Whole thickness methylene blue preparations, frozen sections of tissue impregnated with silver and photographic reconstructions were used to assess the topographical arrangement and density of nervous plexuses. In the various regions a coarse submucous plexus and a much finer subepithelial network were identified, and these were associated with sensory nerve terminations. Plexus formations were found to have greatest density in the dorsum of the tongue, hard palate and gingiva, particularly in the anterior part of the mouth. Myelinated somatic fibres predominated in the plexuses but, from a consideration of nerve fibre diameters, it was concluded that autonomic fibres contribute to the formation of oral nerve networks. The findings lend further support to the view that sensory discrimination largely depends on a complex pattern of nerve impulses reaching higher centres.

Cholinesterases in the tongue of the potto (Perodicticus Potto)

AbstractThe tongue has three circumvallate papillae, arranged in a triangle ahead of the foramen caecum. Gustatory buds are found only on the walls of the vallate papillae and at the surface of the fungiform papillae. The gustatory cells contain acetylcholinesterase; the sustentacular cells contain no such enzymes. A subepithelial network of nerve fibers, containing acetyl‐and butyrylcholinesterases is parallel to the surface of the tongue and extends into the connective tissue cores of all papillae. Numerous pleomorphic end organs are found below all four types of papillae. These abound in acetyl‐and butyrylcholinesterases. The dorsal subepithelial blood vessels contain butyrylcholinesterase, and small amounts of acetyl‐cholinesterase. The ventral aspect of the tongue has fewer nerve fibers and end organs than the dorsal surface does. The nerve fibers to the serous glands of von Ebner, and particularly those to the mucous glands contain considerable acetylcholinesterase.