The distribution and origin of calcitonin gene-related peptide-containing nerve fibres in feline dental pulp

Summary The origin and distribution of calcitonin gene-related peptide (CGRP)-like immunoreactivity in feline dental pulp were studied using indirect immunofluorescence. Nerve fibres with varicosities exhibiting CGRP-like immunoreactivity were observed to enter the pulp with blood vessels. Many CGRP-containing nerve fibres were found to extend along blood vessels in the central pulp, and some of these fibres exhibited a network arrangement in the walls of dental pulp blood vessels. However, some of fibres were apparently not associated with blood vessels. Some thin, CGRP-containing nerve fibres formed a part of the nerve plexus in the subodontoblastic area and penetrated into the odontoblastic layer. In animals that had undergone transection of the inferior alveolar nerve, no CGRP-containing nerve fibres were observed. Application of a double-immunofluorescence staining technique also revealed that the distribution of CGRP-containing nerve fibres is very similar to that of substance P-containing nerve fibres.     

Neurokinin A-like immunoreactivity in feline dental pulp: its distribution,origin and coexistence with substance P-like immunoreactivity

Summary The distribution and origin of neurokinin A (NKA)-like immunoreactivity were investigated in feline dental pulp by an indirect immunofluorescence method. NKA-containing nerve fibres with varicosities, which entered the dental pulp via apical foramen, were distributed throughout this tissue. Many NKA-containing nerve fibres were localized around blood vessels, but some were observed apart therefrom. At the odontoblastic layer, thin NKA-containing nerve fibres were observed running straight toward the pulp-predentinal border between odontoblasts. After inferior alveolar nerve section, all NKA-containing nerve fibres disappeared in the dental pulp, while the removal of the superior cervial ganglion resulted in no change in the distribution of these fibres. The correlation of NKA-like immunoreactivity and substance P (SP)-like immunoreactivity was also investigated by double-immunofluorescence technique. The distribution of NKA-containing nerve fibres was very similar to that of SP-containing nerve fibres; it appeared that all NKA-containing nerve fibres contained SP.     

Monoaminergic and cholinergic nerve fibres in the human dental pulp

Summary Distribution of cholinesterase-containing nerve fibres was studied in 15 human dental pulps by the thiocholine method. Falk's fluorescent method was used to demonstrate catecholamines (8 dental pulps).Cholinesterases were localized partly in the subodontoblastic plexus sending out fine branches towards odontoblasts, and partly in the nerve fibres attached to the blood vessel walls. These fibres in contrast to those of the subodontoblastic plexus were finer and showed fine varicosities.Monoaminergic terminals were localized mainly along blood vessel walls, however, some fibres having no relation to the blood vessels were also found.Cholinesterase-containing nerve fibres in the periphery of the pulp are considered to be sensitive nerve fibres originating from n.V. Distribution of cholinesterase-containing nerve fibres and monoaminergic terminals along the blood vessel walls indicates that the blood vessels in the human dental pulp might be under both parasympathetic and sympathetic control.     

5-Hydroxytryptamine immunoreactivity is detectable in sympathetic nerve fibres in rat oral tissues

Summary The aim of this investigation was to examine if 5-hydroxytryptamine (5-HT) is detectable not only in mast cells but also in sympathetic nerve fibres in oral sites of the rat, including the periodontal ligament, pulp, palatal mucosa, and vestibular sulcus. Antibodies against 5-HT and tyrosine hydroxylase were used. Maxillae from rats were dissected free, fixed, decalcified, cut transversally, and processed for immunohistochemistry. Nerve fibres showing 5-HT-like immunoreactivity were regularly observed in the walls of the arteries and arterioles in the vestibular sulcus and the periodontal ligament. However, 5-HT-like immunoreactivity was not seen in the walls of the vessels of the palatal mucosa. Interestingly, 5-HT-like immunoreactivity coexisted with tyrosine hydroxylase-like immunoreactivity in the innervation of the periodontal ligament and the vestibular sulcus. Thus, the present study gives morphological correlate for the occurrence of effects of 5-HT derived not only from mast cells but also from sympathetic nerve fibres in oral tissues. The source of 5-HT in the nerve fibres as well as the functional implications of the observations remain to be determined.     

Neuropeptide Y-like immunoreactivity in perivascular nerve fibres of the guinea-pig

The distribution of perivascular nerve fibres displaying neuropeptide Y-like immunoreactivity was studied in the guinea-pig. Generally, neuropeptide Y fibres were numerous around arteries and moderate in number around veins. In the heart, immunoreactive fibres were numerous in the auricles and the atria (epi- and endocardium) whereas the ventricles had a more scarce supply. The coronary vessels were richly supplied with fibres. Around large elastic and muscular arteries the fibres formed well developed plexuses. Small arteries in the respiratory tract, the gastrointestinal tract and the genito-urinary tract received a particularly rich supply. In the liver, spleen and kidney only few perivascular fibres were seen. Since immunoreactive fibres around blood vessels disappeared upon surgical or chemical sympathectomy, and sequential immunostaining with antisera against dopamine-beta-hydroxylase (a marker for adrenergic neurons) and against neuropeptide Y revealed their co-existence, it is concluded that neuropeptide Y fibres around blood vessels are sympathetic and adrenergic.     

Peptidergic nerves in human dental pulp. An immunocytochemical study

The peptidergic innervation of human dental pulp was studied with indirect immunofluorescence and immunoperoxidase techniques. Pulpal nerve fibres displaying immunoreactivity for cholecystokinin, calcitonin gene-related peptide, C-terminal flanking peptide of neuropeptide tyrosine, leucine-enkephalin, methionine-enkephalin, neuropeptide K, neuropeptide tyrosine, peptide with N-terminal histidine and C-terminal isoleucine, somatostatin-28, substance P and vasoactive intestinal polypeptide were observed. Immunoreactive axon varicosities were detectable within radicular and coronal nerve trunks and within the nerve plexus of Raschkow in the para-odontoblastic region. Many peptidergic nerve fibres were observed in association with blood vessels of various sizes. Substance P- and calcitonin-gene-related peptide-immunoreactive axons were visible in the odontoblastic layer. The occurrence of VIP- and PHI-immunoreactive fibres lends support to the hypothesis that human tooth may be supplied by parasympathetic nerves. The immunocytochemical results here shown provide a morphological basis to previous experimental studies concerning the possible roles of neuropeptides in nociception mechanisms, control of the blood flow and modulation of the inflammatory response in dental tissues.     

Trigeminal substance P-like immunoreactive fibres in the frog olfactory mucosa

The location and distribution of nerve fibres displaying substanceP (SP) immunoreactivity were studied in the frog olfactory mucosa.Many immunoreactive nerve fibres were noted in close associationwith Bowman's glands and blood vessels in the lamina propria.In addition, such fibres were also found beneath and withinthe olfactory epithelium proper. These fibres are clearly oftrigeminal origin since SP immunoreactivity was abolished aftersection of the trigeminal nerve. Functionally, they might influencelocal blood flow, secretion of Bowman's glands and/or activityof olfactory receptor cells.     

Peptidergic nerves in human dental pulp

Summary The peptidergic innervation of human dental pulp was studied with indirect immunofluorescence and immunoperoxidase techniques. Pulpal nerve fibres displaying immunoreactivity for cholecystokinin, calcitonin gene-related peptide, C-terminal flanking peptide of neuropeptide tyrosine, leucine-enkephalin, methionine-enkephalin, neuropeptide K, neuropeptide tyrosine, peptide with N-terminal histidine and C-terminal isoleucine, somatostatin-28, substance P and vasoactive intestinal polypeptide were observed. Immunoreactive axon varicosities were detectable within radicular and coronal nerve trunks and within the nerve plexus of Raschkow in the para-odontoblastic region. Many peptidergic nerve fibres were observed in association with blood vessels of various sizes. Substance P- and calcitonin-gene-related peptide-immunoreactive axons were visible in the odontoblastic layer. The occurrence of VIP- and PHI-immunoreactive fibres lends support to the hypothesis that human tooth may be supplied by parasympathetic nerves. The immunocytochemical results here shown provide a morphological basis to previous experimental studies concerning the possible roles of neuropeptides in nociception mechanisms, control of the blood flow and modulation of the inflammatory response in dental tissues.     

Calcitonin gene-related peptide (CGRP) immunoreactivity in the neuroendocrine Merkel cells and nerve fibres of pig and human skin

Summary The presence of calcitonin gene-related peptide (CGRP) in the skin of pig snout and human fingertip was investigated using immunohistochemical techniques. CGRP immunoreactivity was found in Merkel cells and nerve fibres of both species. In pig snout skin, Merkel cells containing CGRP were seen forming clusters at the tips of rete ridge epidermis and in the external root sheath of sinus hair follicles (vibrissae). Human Merkel cells immunostained for CGRP were found isolated or forming small groups in the basal layer of glandular epidermal ridges. In all cases, immunoreactivity was more intense on the side of the Merkel cell facing the associated nerve terminal (which was never positive for CGRP). This part of the Merkel cell has the greatest density of dense-cored granules, suggesting that CGRP must be stored in these granules. Nerve, bundles containing CGRP-immunoreactive fibres were found at dermal and hypodermal level, and blood vessels were often surrounded by CGRP nerve fibres. In pig snout skin some nerve fibres containing CGRP penetrated the epidermis and terminated as free endings, and in the human fingertip a small number of CGRP-immunoreactive nerve fibres were seen in Meissner's corpuscles.     

Calcitonin gene-related peptide (CGRP)-immunoreactive sensory and motor nerves of the mammalian palate

The distribution of calcitonin gene-related peptide (CGRP)-immunoreactive nerve fibres in the palate of rat, cat and monkey was studied using immunocytochemistry and radioimmunoassay. CGRP-containing nerve fibres were found, in all species studied, to form a rich plexus in the subepithelial and submucous layers, around excretory ducts and blood vessels. A small number of CGRP-containing nerve fibres penetrated the epithelium of the hard and soft palate, and terminated as free endings. Some CGRP-containing nerve fibres were found in the vicinity of the mucous glands. CGRP-immunoreactive motor end plates were seen in the striated muscle (tensor veli palatini) of the soft palate. Following capsaicin treatment a small depletion in CGRP-immunoreactive nerve fibres in the rat palate epithelium was noted. In contrast, CGRP immunoreactive fibres forming rich plexuses in other layers of the palate, including motor end plates, were not affected. The extractable CGRP showed no significant depletion (normal animals [n = 10] 21.7 +/- 2.4 pmol/g compared with capsaicin-treated animals [n = 10] 17.5 +/- 1.8 pmol CGRP/g wet weight). The reduction in the number of visible immunoreactive nerves following capsaicin application tends to confirm the sensory character of the CGRP-containing nerve fibres terminating in the epithelium of the hard and soft palate. The capsaicin insensitive CGRP-immunoreactive nerve fibres may thus have a predominantly motor function.(ABSTRACT TRUNCATED AT 250 WORDS)     

Protein gene product (PGP) 9.5 immunoreactivity in nerve fibres and pinealocytes of guinea-pig pineal gland: interrelationship with tyrosine-hydroxylase- and neuropeptide-Y-immunoreactive nerve fibres

This light-microscopic (LM) immunohistochemical study has evaluated the presence and distribution of the pan-neural and neuroendocrine marker protein gene product (PGP) 9.5 in pinealocytes and nerve fibres of guinea-pig pineal gland. The pattern of PGP 9.5-immunoreactive (ir) nerve fibres has been compared with that of fibres staining for tyrosine hydroxylase (TH) or neuropeptide Y (NPY). The vast majority of pinealocytes stained for PGP 9.5, although with variable intensity. PGP 9.5 immunoreactivity was localized in pinealocytic cell bodies and processes. Double-immunofluorescence revealed that PGP 9.5 immunoreactivity was absent from glial cells identified with a monoclonal antibody against glial fibrillary acidic protein (GFAP), PGP 9.5 immunoreactivity was also present in a large number of nerve fibres and varicosities distributed throughout the pineal gland. The number of TH-ir and NPY-ir nerve fibres was lower compared with those containing PGP 9.5 immunoreactivity. All fibres staining for NPY also stained for TH. NPY-ir nerve fibres were found to be much more numerous than previously reported for this species. The double-immunofluorescence analysis indicated that almost all TH-ir nerve fibres of the pineal gland contained PGP 9.5 immunoreactivity. However, few PGP 9.5-ir nerve fibres, located in the periphery and the central part of the gland, were TH-negative. A large number of PGP 9.5-ir fibres was concentrated in the pineal stalk. In contrast, TH-ir and NPY-ir nerve fibres were rare in this part of the pineal gland. Our data provide evidence that immunohistochemistry for PGP 9.5 may be a useful tool further to differentiate central and peripheral origins of pineal innervation. Furthermore, the staining of pinealocytes for PGP 9.5 may be exploited to study the three-dimensional morphology and the architecture of pinealocytes and their processes under various experimental conditions.     

Calcitonin gene-related peptide (CGRP) immunoreactivity in the neuroendocrine Merkel cells and nerve fibres of pig and human skin

The presence of calcitonin gene-related peptide (CGRP) in the skin of pig snout and human fingertip was investigated using immunohistochemical techniques. CGRP immunoreactivity was found in Merkel cells and nerve fibres of both species. In pig snout skin, Merkel cells containing CGRP were seen forming clusters at the tips of rete ridge epidermis and in the external root sheath of sinus hair follicles (vibrissae). Human Merkel cells immunostained for CGRP were found isolated or forming small groups in the basal layer of glandular epidermal ridges. In all cases, immunoreactivity was more intense on the side of the Merkel cell facing the associated nerve terminal (which was never positive for CGRP). This part of the Merkel cell has the greatest density of dense-cored granules, suggesting that CGRP must be stored in these granules. Nerve bundles containing CGRP-immunoreactive fibres were found at dermal and hypodermal level, and blood vessels were often surrounded by CGRP nerve fibres. In pig snout skin some nerve fibres containing CGRP penetrated the epidermis and terminated as free endings, and in the human fingertip a small number of CGRP-immunoreactive nerve fibres were seen in Meissner's corpuscles.     

Substance P-containing nerve fibres in large peripheral blood vessels of the rat

Substance P-immunoreactive nerve fibres were localized by the indirect immunohistochemical method in the adventitia and the adventitial-medial border of large peripheral arteries and veins of the rat. Arteries showed a richer substance P-containing innervation than veins. The superior mesenteric artery was densely innervated, whereas no substance P-containing fibres were found around the carotid artery. Substance P produced a vasoconstriction of the veins, but was basically without effect on arteries, although with the carotid artery a dose-dependent relaxation was observed. The absence of a correlation between the degree of innervation of the blood vessels and their responsiveness to exogenous substance P suggests that there nerves do not subserve a vasomotor function. The depletion of substance P immunoreactivity from nerves in arteries and veins by capsaicin suggest that substance P-containing vascular nerves are primarily sensory in nature.     

The sympathetic and parasympathetic nature of neuropeptide Y-immunoreactive nerve fibres in the major salivary glands of the rat

Summary The distribution and origin of neuropeptide Y in the major salivary glands of the rat was studied by indirect immunofluorescence technique. Numerous nerve fibres immunoreactive for the peptide were seen in the parotid and sublingual glands. Most of the fibres were located around blood vessels and salivary acini. In the submandibular gland the number of immunoreactive nerve fibres around the acini was lower in comparison with that in the parotid and sublingual glands. Some immunoreactive nerve fibres were also found around or along intra- and interlobular ducts in all major salivary glands.A large number of the neuropeptide-containing neuronal cell bodies and nerve fibres were detected in the sympathetic superior cervical ganglion. Sympathetic postganglionic nerve trunks of this ganglion contained numerous immunoreactive nerve fibres as well. A subpopulation of the neuronal cell bodies in the submandibular ganglion were immunoreactive to neuropeptide Y.Both uni- and bilateral superior cervical ganglionectomies caused a significant decrease in the number of immunoreactive nerve fibres around the blood vessels in all the major salivary glands. However, these denervations did not affect the density of nerve fibres around the acini and ducts. On the contrary, unilateral parasympathetic denervation by sectioning the auriculotemporal nerve reduced the fibres around the secretory acini in the parotid gland remarkably, while only a minor reduction in the density of immunoreactive fibres associated with the blood vessels of the gland was detected. Unilateral electrocoagulation of the trigeminal nerve branches caused no detectable change in the density of immunoreactive nerve fibres in any of the major salivary glands.On the basis of the present findings it is concluded that neuropeptide Y-reactive nerve fibres present in all major salivary glands around the blood vessels seem to be mainly sympathetic, whereas those around the acini and ducts seems to be of parasympathetic origin.     

The origin and distribution of adrenergic nerve fibres in the guinea-pig colon

Summary A detailed study of the origin and distribution of sympathetic fibres in the distal colon of the guinea-pig has been made using the fluorescent histochemical method for localizing catecholamines. The extrinsic adrenergic fibres of the colonie sympathetic nerves follow the inferior mesenteric artery and its branches to the colon. Some of the extrinsic adrenergic fibres are associated with the parasympathetic fibres of the pelvic nerves near the colon. Complete adrenergic denervation follows the removal of the inferior mesenteric ganglion or the destruction of the nerves running with the inferior mesenteric artery.No fluorescent fibres, other than those associated with blood vessels, were observed in air-dried stretch preparations of the isolated longitudinal muscle. However, a substantial number of varicose, terminal fibres, not associated with blood vessels, were observed in the circular muscle. Some varicose fibres, apart from those associated with ganglion cells, were observed in the myenteric plexus. These fibres were seen in the bundles of nerves running between the nodes of the plexus and also as single fibres which branched from the plexus to end in areas free of ganglion cells.Three plexuses of adrenergic nerve fibres have been distinguished in the submucosa: a dense plexus of terminal fibres innervating both the veins and arteries; a plexus consisting of innervated nodes of ganglion cells, connected by bundles of fluorescent and non-fluorescent nerves; and a plexus of varicose and non-varicose fibres, which is not associated with ganglion cells. Some groups of ganglion cells in the submucosa were without adrenergic innervation.A plexus of varicose fibres forms a meshwork in the lamina propria of the mucosa. The muscularis mucosae is sparsely innervated. Most of the blood vessels in the mucosa are not associated with adrenergic fibres.     

Noradrenergic and peptidergic innervation of the mammary gland in the immature pig

The presence and pattern of coexistence of some biologically active substances in nerve fibres supplying the mammary gland in the immature pig were studied using immunohistochemical methods. The substances studied included: protein gene product 9.5 (PGP), tyrosine hydroxylase (TH), somatostatin (SOM), neuropeptide Y (NPY), galanin (GAL), calcitonin gene-related peptide (CGRP) and substance P (SP). The mammary gland was found to be richly supplied by PGP-immunoreactive (PGP-IR) nerve fibres that surrounded blood vessels, bundles of smooth muscle cells and lactiferous ducts. The vast majority of these nerves also displayed immunoreactivity to TH. Immunoreactivity to SOM was observed in a moderate number of nerve fibres which were associated with smooth muscles of the nipple and blood vessels. Immunoreactivity to NPY occurred in many nerve fibres associated with blood vessels and in single nerves supplying smooth muscle cells. Solitary GAL-IR axons supplied mostly blood vessels. Many CGRP-IR nerve fibres were associated with both blood vessels and smooth muscles. SP-IR nerve fibres richly supplied blood vessels only. The colocalization study revealed that SOM, NPY and GAL partly colocalized with TH in nerve fibres supplying the porcine mammary gland.     

Protein gene product 9.5-immunoreactive nerve fibres and cells in human skin

Summary Sections of human skin were processed according to the indirect immunofluorescence technique with a rabbit antiserum against human protein gene product 9.5 (PGP 9.5). Immunoreactivity was detected in intraepidermal and dermal nerve fibres and cells. The intraepidermal nerves were varicose or smooth with different diameters, running as single processes or branched, straight or bent, projecting in various directions and terminating in the stratum basale, spinosum or granulosum. The density of the intraepidermal nerves varied between the different skin areas investigated. PGP 9.5-containing axons of the lower dermis were found in large bundles. They separated into smaller axon bundles within the upper dermis, entering this portion of the skin perpendicular to the surface. Then they branched into fibres mainly arranged parallel to the epidermal-dermal junctional zone. However, the fibres en route to the epidermis traversed the upper dermis more or less perpendicularly. Furthermore, immunoreactive dermal nerve fibres were found in the Meissner corpuscles, the arrector pili muscles, hair follicles, around the eccrine and apocrine sweat glands and around certain blood vessels. Such fibres were also observed around most subcutaneous blood vessels, sometimes heavily innervating these structures. Numerous weakly-to-strongly PGP 9.5-immunoreactive cells were found both in the epidermis and in the dermis.     

Corticotropin-releasing factor is contained within perikarya and nerve fibres of rat duodenum

Immunofluorescence microscopic studies revealed a corticotropin-releasing factor (CRF) staining within both myenteric plexus perikarya and nerve fibres of the rat duodenum. A CRF-immunofluorescence could be visualized also within nerve fibres close associated with myenteric and submucous blood vessels. Even the lamina propria contained CRF-immunoreactive nerve fibres, which were obviously often localized near the basal lamina.     

Regional difference in the distribution of vasoactive intestinal polypeptide-immunoreactive nerve fibres along the uterus and between myometrial muscle layers in the rat

To investigate a possible regional variation of the vasoactive intestinal polypeptide innervation in the uterus of the cyclic rat, the distribution of vasoactive intestinal polypeptide-containing nerve fibres from the cervix to the oviduct end of the uterine horns was studied using immunohistochemistry. Immunoreactive nerve fibres were most concentrated in the cervix, where they formed a dense plexus in association with the musculature and surrounding blood vessels. In the uterus, a clear regional distribution of the vasoactive intestinal polypeptide innervation was observed. Numerous vascular and non-vascular immunoreactive nerve fibres were present in the lower part of the uterine horns, whereas they were sparse in the median region and absent at the oviduct end. Moreover, non-vascular peptide innervation was mostly concentrated in the circular layer of the myometrium and also occurred in the endometrium. Only a very few immunoreactive nerve fibres were presen t in the longitudinal muscle layer. No change in the peptide innervation pattern was observed during the different stages of the sexual cycle. The marked regional distribution of the peptide innervation in the rat uterus suggests that the regulatory effects of the peptide occur mainly in the lower part of the organ and principally affect the circular muscle layer in the myometrium.     

Cholinesterase-containing nerve fibres on blood vessels in lungs of some laboratory mammals

Summary Distribution of cholinesterase-containing nerve fibres on blood vessels in the lungs of rats, mice, rabbits, guinea pigs, cats, dogs, and rhesus monkeys has been studied with the thiocholine method modified by Coupland and Holmes and with the direct-coloring method according to Karnovsky and Roots. The presence of acetylcholinesterase positive nerve fibres on the pulmonary arteries has been established only in four species of the studied animals, namely the cat, the dog, the rhesus monkey and the rabbit. These nerves form d distinct plexus on the border between the media and adventitia. In the thick walled perihilar branches of the pulmonary artery of the rabbit the nerve fibres penetrate—as a rule—into the outer half of the media. Despite many times repeated experiments and careful investigations no nerve fibres have been found on the intrapulmonary branches of the blood vessels in the lungs of rats, mice, and guinea pigs. In the walls of the pulmonary veins, with the exception of the pulmonary veins in the rat and the mouse, cholinesterase fibres have been identified only seldom. Acetylcholinesterase nerve fibres in medio-adventitial localization have also been found in the walls of bronchial arterioles, this being the case in all the species under study. Bronchial veins do not exhibit any nerve fibres. The distribution of the acetylcholinesterase-containing nerve fibres on the pulmonary arteries is different from species to species.