Vasoactive Intestinal Polypeptide Immunoreactivity Research Articles

Nerve and blood vessel growth in response to grafted dermis and cultured keratinocytes.

The aim of this study was to study innervation and angiogenesis in response to grafts of dermis and cultured keratinocytes using immunohistochemical techniques. In a porcine model, fresh autologous de-epidermalized dermis and cultured autologous keratinocytes were combined using a two-stage technique, to produce keratodermal grafts. Wounds were encased within skin graft chambers that prevented the influence of the surrounding skin. As grafts contracted, a peripheral rim of granulation tissue became exposed, allowing us to compare the wound bed beneath grafts with that beneath the raw granulating surface. Grafts were studied for 6 weeks. Angiogenesis was studied using antisera to von Willebrand factor to detect endothelial cells. Nerve growth was studied using antisera to S-100, a Schwann cell marker, and to four axonal markers: protein gene product 9.5, C-flanking peptide of neuropeptide Y, calcitonin gene-related peptide, and vasoactive intestinal peptide. In kerato-dermal grafts (n = 28), organization of blood vessels and nerve growth occurred only beneath areas with epidermal cover as compared with the surrounding granulation tissue. Initially, the immunoreactivity to von Willebrand factor was high, but in areas with epidermal cover it assumed a more orderly pattern with fewer blood vessels. Innervation was first detected by S-100 immunoreactivity seen at 1 to 2 weeks, closely followed by that to protein gene product 9.5 and much later to calcitonin gene-related peptide. C-flanking peptide of neuropeptide Y and vasoactive intestinal peptide immunoreactivity were detected in the wound depth surrounding large blood vessels at 4 to 6 weeks. In control wounds that had been either grafted with de-epidermalized dermis alone (n = 10) or allowed to granulate (n = 10), persistently there was high immunoreactivity to von Willebrand factor but minimal immunoreactivity to the neural markers. In conclusion, kerato-dermal grafts become innervated, and beneath their surface there is also vascular organization to resemble normal skin. Keratinocytes themselves may influence angiogenesis and innervation, as both processes failed to occur beneath granulating areas.

Polarised innervation pattern of the mucosa of the guinea pig distal colon

A neuronal retrograde tracing method with the dye DiI in combination with the immunohistochemical detection of vasoactive intestinal polypeptide (VIP) and choline acetyltransferase (ChAT) was used to characterise the mucosal projection of neurones located in the submucosal plexus of the guinea pig distal colon. VIP and ChAT immunoreactivity labelled separate populations. The mucosa was innervated by descending (59.7±7.4%), ascending (8.7±5.0%) and circumferential (31.7±6.1%) pathways. Descending neurones had longer projections than ascending ones. Descending DiI-labelled neurones were in their vast majority VIP-positive (88.4±3.6%) and only 3.5±4.8% of them were ChAT-positive. Conversely, the ascending pathways were predominantly ChAT-positive (90.9±20.3%) and only 7.8±20.6% of them were VIP-positive. A significantly larger proportion of DiI-neurones with circumferential projection were ChAT- than VIP-positive (70.9±15.6 vs. 27.8±15.7%). The proportion of ascending, descending and circumferential ChAT-positive DiI-neurones was 23:7:70%, respectively, whereas the proportion for VIP-positive DiI-neurones was 2:84:14%, respectively. The results suggest a conservation of the submucosal innervation pattern along the guinea pig colonic mucosa but also reveal differences in the mucosal enteric innervation of other regions of the gastrointestinal tract.

Morphologic alterations of the enteric nervous system and deficiency of non-adrenergic non-cholinergic inhibitory innervation in neonatal necrotizing enterocolitis.

Although damage to intramural nervous tissue should be expected in neonatal necrotizing enterocolitis (NNEC), as the pathology of NNEC is defined by substantial destruction of the bowel wall, only a few studies have considered its implication. Thus, the aim of the study has been to examine morphological alterations of the enteric nervous system (ENS) in intestinal segments affected by NNEC. Immunohistochemical methods allowed the demonstration of both neuronal and glial elements and the assessment of an altered localization of non-adrenergic non-cholinergic (NANC) inhibitory mediators within the intramural plexuses. Intestinal segments from patients with NNEC (n = 8) and control subjects (n = 3) were obtained and submitted to immunohistochemical examination incubating with antibodies against protein gene product (PGP) 9.5, protein S-100, vasoactive intestinal polypeptide (VIP) and nitric oxide synthase (NOS). The most severe damage of nervous tissue was found within the plexus mucosus and plexus submucosus internus. The ganglionated plexuses showed a loss ot both glial and nerve cells with various stages of cell deterioration and the formation of central lesions within the myenteric ganglia. The observed neuropathologic changes correspond to the group of acquired segmental hypoganglionosis. Specimens from patients with NNEC were also characterized by an absence of immunoreactive VIP and NOS in the plexus submucosus and within the circular muscle layer. The deficiency in NANC inhibitory innervation may contribute to the formation of functional obstructions following acute NNEC. Furthermore, it is likely that the neuropathological lesions induced in early stages of NNEC may result in dysfunctional intestinal motility facilitating intraluminal bacterial overgrowth and translocation, and therefore, possibly promote the self-perpetuating pathophysiologic cycle culminating in progressive NNEC. As an additional finding, two patients with NNEC showed typical features of intestinal neuronal dysplasia (IND). The association of NNEC and IND is reviewed in the literature and possible causalties are discussed.

N-Methyl- d-aspartate receptors are expressed by intrinsic neurons of rat larynx and esophagus

Overactivation of N-methyl- d-aspartate receptors (NMDAR), a mechanism of central neurotoxicity, has recently been shown to increase airway responsiveness in rat lungs. NMDAR have not been localized in the airways, but neurons of the myenteric plexus in the rat express mRNA for NMDAR. Furthermore, a population of glutamate-containing cell bodies in the nucleus ambiguus projects to the rat larynx. On this basis, we hypothesized that some postganglionic parasympathetic neurons of the larynx, trachea and esophagus may express NMDAR. Sections of rat larynx, trachea and esophagus were immunocytochemically labeled for NMDAR subtype 2B using a specific antibody. NMDAR immunoreactivity was observed in cell bodies of individual neurons located in the submucosa and on the external surface of skeletal muscle in the larynx and also in neurons of the esophageal plexus. All NMDAR-positive nerve cell bodies also contained immunoreactivity for vasoactive intestinal peptide (VIP) and some were immunoreactive for nitric oxide synthase (NOS). None of the cell bodies of the tracheal plexus contained NMDAR immunoreactivity. The findings demonstrate that NMDAR are expressed in neurons of the rat larynx and esophagus and their activation may be associate with VIP or NO release.

Modular distribution of vasoactive intestinal polypeptide in the rat barrel cortex: changes induced by neonatal removal of vibrissae

The distribution of vasoactive intestinal polypeptide-immunoreactive neuronal structures in the barrel cortex (posteromedial barrel subfield) of adult rats was analysed after unilateral removal of the vibrissal follicles of row C in neonatal rats. The hypothesis was tested whether the distribution of vasoactive intestinal polypeptide-immunoreactive structures depends on the normal anatomical organization of the specific sensory input. After three months survival the distribution of the vasoactive intestinal polypeptide-immunoreactive structures was morphometrically evaluated. This approach revealed alterations in the contralateral posteromedial barrel subfield, where the disappearance of barrel row C and a substantial increase in size mainly of barrel row D, but also of other rows could be detected. Increase in row D included both barrels and the interspace (septal segments between barrels in one row). As vasoactive intestinal polypeptide immunoreactivity of the barrel field was found previously to be localized in synaptic boutons involved in symmetric synapses, our present findings suggest that (i) the interspace is enriched in inhibitory vasoactive intestinal polypeptide-immunoreactive synapses as opposed to the excitatory thalamocortical input reaching the barrel hollow, (ii) the spatial distribution of the vasoactive intestinal polypeptide system in the barrel cortex is closely associated with the neuronal organization of the sensory input and reacts with a considerable plasticity to lesion-induced changes of the input, and (iii) the compensatory barrel hypertrophy in a row neighbouring the deafferented row involves an increasing number of vasoactive intestinal polypeptide-immunoreactive synapses per barrel.

Lesion-induced transneuronal plasticity of the cholinergic innervation in the adult rat entorhinal cortex.

The present experiments were designed to determine the effect that lesions of the basal forebrain cholinergic system exert on cholinergic interneurons within the entorhinal cortex (EC) in the rat. Unilateral infusion of 192 IgG-saporin into the nucleus of the horizontal diagonal band of Broca (HDB) decreased the number of ipsilateral choline acetyltransferase immunoreactive (ChAT-ir) neurons by 54%. Two-four weeks after the lesion, the ipsilateral EC exhibited a moderate but significant loss of ChAT-ir fibres and interneurons. Adjacent sections revealed a parallel loss of vasoactive intestinal polypeptide (VIP) immunoreactivity. Cell counts in the cingulate cortex were unaffected, suggesting that this effect was indeed specific to the main target area for HDB neurons. Ibotenic acid lesions also induced a significant 36% decrease in the number of cholinergic neurons in the ipsilateral HDB, and disappearance of ChAT terminals in the EC, whereas the number of ChAT-ir neurons in the EC was unchanged. Since ibotenic acid affects all cells and not only cholinergic ones, our results suggest that the specific degeneration of cholinergic neurons in the HDB after 192 IgG-saporin treatment could be inducing transsynaptic effects on their targets. Injections of 192 IgG-saporin directly into the EC also lesioned the cholinergic projection from the HDB, but had no effect on the intrinsic population. Eight weeks after immunolesion, the number of interneurons immunoreactive for ChAT and VIP in the EC had returned to normal values, and persisted for as long as 6 months after the lesion. By contrast, ChAT-ir neurons in the HDB were permanently lost. Our results suggest that the transient down-regulation of the cholinergic phenotype in entorhinal cortex interneurons could be a manifestation of activity-dependent plasticity, and that the loss of cholinergic innervation from the basal forebrain could be responsible for these effects through an imbalance of inputs. We hypothesize that the recovery of the phenotypic expression of entorhinal interneurons could be due to a recovery in their innervation, perhaps from sprouting axons in the same fields, belonging to surviving cholinergic neurons in the basal forebrain.

Nitrergic innervation of the cerebral arterial tree in the bent-winged bat (mammalia: Microchiroptera)

The distribution and origin of cerebrovascular nitrergic nerves were studied immunohistochemically and histochemically in the bent-winged bat. The supply of nitric oxide synthase (NOS)-immunoreactive (IR) and nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd)-positive nerves to the bat major cerebral arteries differs from the general mammalian pattern in that it is preferential for the vertebrobasilar system (VBS) as opposed to the internal carotid system. Interestingly, a few nerve cells with bright NOS immunofluorescence and intense NADPHd activity were localized in the walls of the vertebral artery (VA) and basilar artery (BA) from many individual bats. Cerebral perivascular NOS-IR nerves were generally immunoreactive for vasoactive intestinal polypeptide (VIP). NOS-IR neurons intrinsic to the BA and VA expressed variable degrees of VIP immunoreactivity and showed no acetylcholinesterase (AChE) activity. Most cell bodies of the microganglia (MG) in the carotid canal and tympanic cavity, and those of the cranial and cervical facial ganglia, showed both NOS and VIP immunoreactivities and were stained intensely for NADPHd. From these and other findings, it is suggested that, in the bent-winged bat at least, the BA and VA of the cerebral arterial tree are frequently dually innervated by two neurochemically defined nitrergic neurons, the cranial parasympathetic VIP-IR and AChE-positive neurons, which are derived mainly from the MG via the internal carotid artery, and the intrinsic neurons, either IR or immunonegative for VIP but negative for AChE, which form an outflow tract from some caudally located ganglia projecting to the VBS via the VA.

Nitrergic innervation of the cerebral arterial tree in the bent-winged bat (mammalia: Microchiroptera).

The distribution and origin of cerebrovascular nitrergic nerves were studied immunohistochemically and histochemically in the bent-winged bat. The supply of nitric oxide synthase (NOS)-immunoreactive (IR) and nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd)-positive nerves to the bat major cerebral arteries differs from the general mammalian pattern in that it is preferential for the vertebrobasilar system (VBS) as opposed to the internal carotid system. Interestingly, a few nerve cells with bright NOS immunofluorescence and intense NADPHd activity were localized in the walls of the vertebral artery (VA) and basilar artery (BA) from many individual bats. Cerebral perivascular NOS-IR nerves were generally immunoreactive for vasoactive intestinal polypeptide (VIP). NOS-IR neurons intrinsic to the BA and VA expressed variable degrees of VIP immunoreactivity and showed no acetylcholinesterase (AChE) activity. Most cell bodies of the microganglia (MG) in the carotid canal and tympanic cavity, and those of the cranial and cervical facial ganglia, showed both NOS and VIP immunoreactivities and were stained intensely for NADPHd. From these and other findings, it is suggested that, in the bent-winged bat at least, the BA and VA of the cerebral arterial tree are frequently dually innervated by two neurochemically defined nitrergic neurons, the cranial parasympathetic VIP-IR and AChE-positive neurons, which are derived mainly from the MG via the internal carotid artery, and the intrinsic neurons, either IR or immunonegative for VIP but negative for AChE, which form an outflow tract from some caudally located ganglia projecting to the VBS via the VA.

Transgene expression in rat fetal brain grafts is maintained for 7 months after ex vivo adenoviral vector-mediated gene transfer

Fetal brain tissue fragments containing the suprachiasmatic nucleus were infected with an adenoviral vector containing the marker gene LacZ encoding for β-galactosidase, and subsequently cultured or transplanted in the third ventricle of SCN-lesioned adult Wistar rats. In previous studies we optimized the infection procedure and characterized the immunological response directed against the viral vector in this model. The present study reports on β-gal expression for at least 7 months in neuronal and glial cells. Maturation of the transplanted fetal SCN with respect to immunoreactivity for vasoactive intestinal polypeptide and C-terminal propressophysin was not hampered by the viral infection.

The augmentation of pituitary adenylate cyclase-activating polypeptide (PACAP) in streptozotocin-induced diabetic rats

Pituitary adenylate cyclase activating polypeptide (PACAP), which was isolated from ovine hypothalamic extract, has been shown to have a physiological role in the regulation of insulin or islet functions. In streptozotocin (STZ)-induced diabetic rats, we examined the content of PACAP immunoreactivity and gene expression of three specific receptors. Four weeks after administration of STZ (50 mg/kg), plasma glucose levels increased 3.3-fold, and plasma insulin levels decreased to one-tenth as compared with the control. The content of PACAP immunoreactivity in the pancreas potently increased by 30%, but the content of vasoactive intestinal polypeptide (VIP) immunoreactivity was not changed. In the other tissues, the content of PACAP immunoreactivity did not significantly change except in the hypothalamus, which showed a 10% increment. In the expression level of PACAP/VIP receptors, semi-quantitative RT-PCR analysis revealed that VIP1/PACAP receptor mRNA significantly increased as compared with the other two types of receptors in the pancreas of STZ-induced diabetic rats. These findings suggest that PACAP and VIP1/PACAP receptor might be involved in the pathophysiology of diabetes mellitus.

Increase of 5HT and VIP immunoreactivity within the hamster ( Mesocricetus auratus) SCN during chronic MAOI treatment

The effects of chronic treatment with the monoamine oxidase inhibitor (MAOI), clorgyline (CLG; 2 mg/kg per day) on serotonin (5HT) and vasoactive intestinal peptide (VIP) immunoreactivity (IR) within the hamster suprachiasmatic nucleus (SCN) were examined. Optical densities of 5HT IR and VIP IR were increased by MAOI treatment. VIP IR was increased in both the ventrolateral and dorsal regions of the SCN, suggesting that VIP content was increased within both perikarya and terminals of VIP neurons. The results suggest that previously described effects of MAOIs on the mammalian circadian system may be mediated in part, by their effects on serotonergic input to VIP neurons within the SCN.

Cholinergic regulation of VIP gene expression in human neuroblastoma cells

The effects of carbamylcholine (CCh) on the gene expression of the neuropeptide vasoactive intestinal polypeptide (VIP) were studied using two human neuroblastoma cell lines, NB-1 and BE(2)M17. CCh caused a fast increase in VIP mRNA level in both cell lines which was followed by an increase in VIP immunoreactivity. The time-course of the induction of both mRNA and peptide differed, however, between the two cell lines. No morphological changes of the cells were observed during 6 days of stimulation. The effect was mediated by the muscarinic class of acetylcholine receptors, since it could be totally abolished by atropine. Since CCh caused an accumulation of inositol-1,4,5-triphosphate, it is likely that muscarinic receptor subtype M1, M3 or M5 is involved. Experiments with the translational inhibitor, cycloheximide, showed that CCh mediated a direct effect on the VIP gene expression. By combining gel permeation chromatography with radioimmunoassays using antisera specific for the various VIP-precursor products, immunoreactive peaks eluting as the synthetic peptides were found in both cell lines. In addition, earlier eluting peaks which could represent partially processed or extended VIP forms were found. After CCh induction the concentration of all preproVIP-derived products increased, and there was a tendency towards a shift to more fully processed VIP. The findings give new evidence for a direct regulation of VIP gene expression in human neuronal cells by cholinergic agents.

Innervation of the gallbladder: structure, neurochemical coding, and physiological properties of guinea pig gallbladder ganglia.

The muscle and epithelial tissues of the gallbladder are regulated by a ganglionated plexus that lies within the wall of the organ. Although these ganglia are derived from the same set of precursor neural crest cells that colonize the gut, they exhibit structural, neurochemical and physiological characteristics that are distinct from the myenteric and submucous plexuses of the enteric nervous system. Structurally, the ganglionated plexus of the guinea pig gallbladder is comprised of small clusters of neurons that are located in the outer wall of the organ, between the serosa and underlying smooth muscle. The ganglia are encapsulated by a shell of fibroblasts and a basal lamina, and are devoid of collagen. Gallbladder neurons are rather simple in structure, consisting of a soma, a few short dendritic processes and one or two long axons. Results reported here indicate that all gallbladder neurons are probably cholinergic since they all express immunoreactivity for choline acetyltransferase. The majority of these neurons also express substance P, neuropeptide Y, and somatostatin, and a small remaining population of neurons express vasoactive intestinal peptide (VIP) immunoreactivity and NADPH-diaphorase enzymatic activity. We report here that NADPH-diaphorase activity, nitric oxide synthase immunoreactivity, and VIP immunoreactivity are expressed by the same neurons in the gallbladder. Physiological studies indicate that the ganglia of the gallbladder are the site of action of the following neurohumoral inputs: 1) all neurons receive nicotinic input from vagal preganglionic fibers; 2) norepinephrine released from sympathetic postganglionic fibers acts presynaptically on vagal terminals within gallbladder ganglia to decrease the release of acetylcholine from vagal terminals; 3) substance P and calcitonin gene-related peptide, which are co-expressed in sensory fibers, cause prolonged depolarizations of gallbladder neurons that resemble slow EPSPs; and 4) cholecystokinin (CCK) acts presynaptically within gallbladder ganglia to increase the release of acetylcholine from vagal terminals. Results reported here indicate that hormonal CCK can readily access gallbladder ganglia, since there is no evidence for a blood-ganglionic barrier in the gallbladder. Taken together, these results indicate that gallbladder ganglia are not simple relay stations, but rather sites of complex modulatory interactions that ultimately influence the functions of muscle and epithelial cells in the organ.

Somatostatin 2A receptor is expressed by enteric neurons, and by interstitial cells of Cajal and enterochromaffin-like cells of the gastrointestinal tract

Somatostatin exerts multiple effects by activating distinct G protein-coupled receptors. Here we report the cellular sites of expression of the somatostatin subtype 2A (sst2A) receptor in the rat enteric nervous system by using a C-terminus-specific, affinity-purified antiserum and immunohistochemistry. Antibody specificity was confirmed by the cell surface staining of human embryonic kidney 293 cells expressing the sst2A receptor, the lack of staining of cells expressing the somatostatin subtype 2B receptor, and the abolition of staining by preincubating the antiserum with the C-terminus peptide used for immunization, SSt2A(361-369). The SSt2A receptor antibody recognized a broad 80 kDa band on Western blots of membranes prepared from cells transfected with sst2A receptor cDNA; following receptor membrane deglycosylation, the antibody detected an additional 40 kDa band. In the enteric nervous system, the sst2A antibody primarily stained neurons of the myenteric and submucosal plexuses, and abundant fibers distributed to the muscle, mucosa, and vasculature. Immunoreactive staining was also observed in non-neuronal cells, including presumed interstitial cells of Cajal of the intestine and enterochromaffin-like cells of the stomach. Fibers expressing sst2A receptor immunoreactivity were often in close proximity to D cells of the gastric and intestinal mucosa. Colocalization of somatostatin and sst2A receptor immunoreactivities was not observed in endocrine cells nor in enteric neurons. Double-label immunohistochemistry revealed colocalization of sst2A and vasoactive intestinal peptide immunoreactivities in enteric neurons. The multiple types of cells expressing the sst2A receptor, including enteric neurons and non-neuronal structures, in addition to the relationship between somatostatin and sst2A receptor elements, provide evidence that the sst2A receptor mediates somatostatin effects in the gastrointestinal tract via neuronal and paracrine pathways.

Selective innervation of different target tissues in guinea-pig cranial exocrine glands by sub-populations of parasympathetic and sympathetic neurons

This study has used multiple-labelling immunohistochemistry and quantitative analysis to examine the projections of subpopulations of parasympathetic and sympathetic neurons to different vascular and secretory structures in five cranial exocrine glands of guinea-pigs. Multiple subpopulations of parasympathetic axons, identified by immunoreactivity (IR) for various combinations of peptides, innervated arteries, arterioles, ducts and acini in sublingual, submandibular, parotid, lacrimal and zygomatic glands, although axons were absent from ducts in the parotid gland. Most parasympathetic axons contained IR for vasoactive intestinal peptide (VIP) and neuropeptide Y (NPY), with or without enkephalin (Enk). The proportion of parasympathetic axons that contained Enk-IR varied greatly between target tissues and glands: Enk-IR was more common in axons supplying secretory ducts, acini and arterioles than in axons innervating more proximal arteries; Enk-IR was less common in axons supplying the lacrimal gland than axons supplying the submandibular, lacrimal and zygomatic glands. Sympathetic axons with IR for tyrosine hydroxylase (TH) innervated arterial vessels in all glands, but innervated secretory structures only in the salivary glands. Sympathetic axons supplying proximal arterial segments often contained NPY-IR and sometimes also contained IR for dynorphin. Dynorphin-IR was more common in axons in the parotid, lacrimal and zygomatic glands than in the sublingual and submandibular glands. In contrast, axons supplying arterioles, ducts and acini lacked peptide IR. These results indicate that neuronal pathways regulating proximal arteries in cranial exocrine glands are different from the neuronal pathways regulating arterioles and acini, and may be different from neurons projecting to proximal secretory ducts. Furthermore, the peptides enkephalin, NPY and dynorphin are likely to make variable contributions to autonomic neurotransmission in different arterial segments and in different cranial exocrine glands.

Five inhibitory transmitters coexist in pelvic autonomic vasodilator neurons.

Here we describe the localization of a potent vasodilator, calcitonin gene-related peptide (CGRP), in pelvic autonomic neurons containing four other inhibitory transmitters: vasoactive intestinal peptide (VIP), neuropeptide Y, nitric oxide and acetylcholine. These neurons mediate endothelium-independent vasodilation by releasing nitric oxide and one or more neuropeptides. Sixty percent of nerve cell bodies in guinea-pig paracervical ganglia with immunoreactivity (IR) for VIP, choline acetyltransferase (ChAT) and nitric oxide synthase (NOS), also contained IR for CGRP. Furthermore, many VIP-IR varicose nerve terminals at the adventitia-medial junction of the guinea-pig uterine artery contained IR for CGRP, ChAT and NOS. Both alpha-hCGRP and beta-hCGRP were potent dilators of the uterine artery (pD2 values 8.1, 8.3, respectively), but 1 microM hCGRP(8-37) did not antagonize dilations produced by either agonist. Dilations produced by alpha-hCGRP were unaffected by removal of the endothelium. Taken together with results of our previous studies, we propose that CGRP can contribute directly to autonomic vasodilation, possibly via CGRP2 receptors on smooth muscle cells, and that CGRP is the fifth inhibitory transmitter co-existing in pelvic vasodilator neurons.

Ontogenesis of the retinohypothalamic tract, vasoactive intestinal polypeptide- and peptide histidine isoleucine-containing neurons and melatonin binding in the hypothalamus of the mink.

The development of the retinohypothalamic tract (RHT) labeled with cholera toxin and of the vasoactive intestinal polypeptide (VIP) and peptide histidine isoleucine (PHI) systems was studied in the hypothalamus of neonatal mink by using immunohistochemistry. Retinal fibers were observed in the suprachiasmatic nucleus (SCN) from birth and were adult-like by day 14. VIP and PHI immunoreactivity was also present from birth. Melatonin binding was studied by autoradiography using [125I]melatonin as a ligand. A specific binding was detected in near-term fetal and neonate brains in the olfactory epithelium, various thalamic nuclei, the pineal gland, and the pars tuberalis of the pituitary gland, but never in the SCN. These results are discussed in the context of the potential role of daylight cycles and/or melatonin in entraining circadian rhythms in neonate mink.

The polarity of neurochemically defined myenteric neurons in the human colon

BACKGROUND & AIMS: Functional classes of enteric neurons in small mammals can be determined by neurochemical coding and axonal projections. The aim of this study was to describe the projections of vasoactive intestinal peptide (VIP)-, tachykinin (TK)-, and calretinin- immunoreactive (IR) interneurons and circular muscle motor neurons in human colon.METHODS: Human colonic myenteric neurons were retrogradely filled with 1,1'-didodecyl 3,3,3',3'-indocarbocyanine perchlorate (DII) from either the myenteric plexus or the circular muscle layers in organotypic culture preparations. The preparations were then labeled with antisera, and the position and immunoreactivity of DII-filled neurons was recorded.RESULTS: Nine percent of circular muscle motor neurons were labeled with TK immunoreactivity and these projected orally; 22% were VIP-IR and projected anally. Fifty-one percent of myenteric neurons that projected anally to the myenteric ganglia were VIP-IR and none were TK-IR. In contrast, 23% of neurons projecting orally were TK-IR but only 2% were VIP-IR. Calretinin immunoreactivity was present in 23% of neurons projecting anally and 3% projecting orally within the myenteric plexus. Six percent of circular muscle motor neurons had Dogiel type II morphology, and 50% of these were TK- IR.CONCLUSIONS: Circular muscle motor neurons and myenteric interneurons with TK, VIP, and calretinin immunoreactivity in the human colon have distinct projections and polarity.(Gastroenterology 1997 Aug;113(2):497-506)

Relation between cholecystokinin and antral innervation in the control of gastric emptying in the rat.

BACKGROUND: Antral motility and the hormone cholecystokinin (CCK) are major determinants of the rate of gastric emptying. The relation between CCK and antral neurons in regulating gastric emptying is uncertain....

Morphological relations between haem oxygenases, NO-synthase and VIP in the canine and feline gastrointestinal tracts

Carbon monoxide (CO), produced by haem oxygenase (HO), has been suggested as a messenger molecule in the central and peripheral nervous systems. In the present study, we have investigated the occurrence of the two isoforms of HO, HO-2 and HO-1 in the canine and feline gastrointestinal tracts, including the small and large intestine and the gastrointestinal sphincters. An abundance of nerve cell bodies that contained immunoreactivity for HO-2 was found in the submucosal and myenteric plexuses. HO-2 immunoreactivity was frequently co-localized with nitric oxide synthase (NOS) or vasoactive intestinal peptide (VIP) immunoreactivities and was also observed in some nerve fibres, certain non-neuronal cells dispersed among smooth muscle bundles, and in vascular endothelium. The antiserum against HO-1 revealed immunoreactivity in nerve cell bodies in the enteric plexuses, in nerve fibres and in non-neuronal cells in the smooth muscle layers. Some of the nerve structures were also NOS- or VIP-immunoreactive. These results demonstrate the presence of HO isoenzymes in nerves and other structures of the canine and feline gastrointestinal tracts and support the view that CO may have a role as a messenger molecule in the enteric nervous system.