Nitric oxide synthase is co-localized with vasoactive intestinal polypeptide in postganglionic parasympathetic nerves innervating the rat vas deferens

Cross-sections of the vas deferens taken from control adult male rats showed positive histochemical reactivity to acetylcholinesterase and immunoreactivity for antibodies to protein gene product 9.5, tyrosine hydroxylase, neuropeptide Y, vasoactive intestinal polypeptide, nitric oxide synthase and calcitonin gene-related peptide. Immunoreactivity to substance P was very sparse. Histochemical reactivity to acetylcholinesterase and immunoreactivity to vasoactive intestinal polypeptide and nitric oxide synthase was concentrated in the subepithelial lamina propria and inner smooth muscle layers. Complete surgical denervation resulting from transection of the nerve arising from the pelvic ganglion which supplies the vas deferens totally abolished the immunoreactivity to all of the antibodies tested as well as the histochemical reactivity to acetylcholinesterase. In sections of the prostatic end of the vas deferens taken from rats neonatally pretreated with capsaicin, immunoreactivity to calcitonin gene-related peptide and substance P was reduced by 75 and 83%, respectively. Immunoreactivity to neuropeptide Y, vasoactive intestinal polypeptide and nitric oxide synthase was similar in tissue sections taken from capsaicin-treated rats and those taken from control tissues. Pretreatment of rats with guanethidine or 6-hydroxydopamine decreased immunoreactivity to tyrosine hydroxylase and neuropeptide Y by 60-70%, but immunoreactivity to substance P, vasoactive intestinal polypeptide and nitric oxide synthase was unchanged, while immunoreactivity to calcitonin gene-related peptide and acetylcholinesterase staining was increased by guanethidine but not by 6-hydroxydopamine treatment. Triple labelling experiments showed nitric oxide synthase, vasoactive intestinal polypeptide and acetylcholinesterase all to be co-localized in some nerve fibres. These results indicate that the nitric oxide synthase contained in the nerve fibres innervating the rat vas deferens is unaffected by pretreatment of rats with capsaicin, 6-hydroxydopamine or guanethidine but is abolished by surgical denervation, of postganglionic parasympathetic, sympathetic and sensory nerves. Therefore it appears that nitric oxide synthase is co-localized with vasoactive intestinal polypeptide in the postganglionic parasympathetic nerves which innervate the rat vas deferens.     

Behavioural topography in the striatum: differential effects of quinpirole and D-amphetamine microinjections

The structure of the nervous network and the distribution of tyrosine hydroxylase (TH)- and various neuropeptide-containing nerves were immunohistochemically studied in the glottis of the dog. The nervous network in the glottis revealed apparent regional differences in morphology. The nervous network in the cartilaginous vocal fold of the posterior glottis consisted of nerve bundles running parallel to the edge of the vocal fold. Only a small number of nerve bundles were observed in the anterior glottis, specifically in membranous vocal fold. In the subepithelial layer of the posterior glottis, a moderate number of galanin (GAL)-immunoreactive nerve fibers were observed, while only a few fibers were present in the anterior glottis. Numerous vasoactive intestinal peptide (VIP)-, GAL-, methionine-enkephalin (ENK)- and TH-immunoreactive nerve fibers were observed within and around the laryngeal submucosal seromucous gland. Many TH- and neuropeptide Y (NPY)-immunoreactive fibers were arranged around the blood vessels. In the epithelia, free nerve endings with immunoreactivity for substance P (SP) and calcitonin gene-related peptide (CGRP) was observed. Furthermore, nerve cell bodies with SP-, VIP-, GAL-, ENK-, and NPY-immunoreactivity were observed in the deep region of the submucosal layer. The results from the present study suggest that there is autonomic regulation of the glottis. Regional structural differences in the nervous network of the glottis may reflect functional differences.     

Sympathetic axons surround neuropeptide-negative axotomized sensory neurons

Nerve injury can lead to sympathetically dependent neuropathic pain. A possible site of sympathetic-sensory interaction is the dorsal root ganglion (DRG), where sympathetic axons form pericellular 'baskets' around a subpopulation of DRG neurons. Since these structures possibly represent functional units of sympathetic pain, we attempted to characterize the neuropeptidergic phenotype of basketed DRG neurons. We performed double-labeling immunohistochemistry for tyrosine hydroxylase and neuropeptides on DRG sections, 2 weeks following L5 spinal nerve ligation (a well-characterized animal model of sympathetic pain). We found that basketed DRG neurons typically do not contain substance P, calcitonin gene-related peptide, galanin, neuropeptide tyrosine, or vasoactive intestinal polypeptide, and we conclude that if sympathetic baskets contribute to neuropathic pain, the involvement of these neuropeptides is unimportant.     

Innervation and nitric oxide modulation of mesenteric arteries of the golden hamster

Immunohistochemical and pharmacological techniques were used to examine perivascular nerves, endothelium and the effects of inhibition of nitric oxide synthesis on responses in mesenteric arteries/perfused mesenteric arterial beds of the Golden hamster. Frequency-dependent vasoconstrictions to electrical field stimulation and dose-dependent vasoconstrictions to noradrenaline were significantly augmented by NG-nitro-L-arginine methyl ester (10(-5) M), an inhibitor of nitric oxide synthase. In preparations with tone raised with methoxamine (10 microM) dose-dependent relaxations to ATP, but not to acetylcholine, were blocked by NG-nitro-L-arginine methyl ester. In the presence of guanethidine (5 microM) to block sympathetic neurotransmission there was no neurogenic relaxation to electrical field stimulation. Furthermore, the sensory neurotoxin capsaicin (0.05-5 nmol) did not elicit relaxation. Immunohistochemical studies demonstrated dense plexuses of fibres immunoreactive for tyrosine hydroxylase and neuropeptide Y, a plexus of moderate density for calcitionin gene-related peptide and an absence of fibres immunoreactive for substance P and vasoactive intestinal polypeptide. Of particular interest is the finding that whereas sympathetic perivascular nerves and nitric oxide regulate the function of hamster mesenteric arteries, there is no apparent motor function of calcitonin gene-related peptide-containing sensory nerves.     

Prolonged increase in micturition threshold volume by anogenital afferent stimulation in the rat

Neuroendocrine components exist in the human nasal mucosa. However, the pathophysiological and neuroimmunological roles of the regulatory peptides in allergic rhinitis (AR) require further investigation. To analyse the functional morphology and quantify the tissue concentration of regulatory peptides in the nasal mucosa of AR subjects, human inferior turbinate mucosa specimens from 25 patients with AR, 20 patients with non-allergic rhinitis and 10 patients without any nasal diseases were investigated. Using immunohistochemistry and radioimmunoassays, we detected the presence, distribution and concentrations of various neuropeptides [vasoactive intestinal peptide (VIP), neuropeptide Y (NPY), substance P (SP) and calcitonin gene-related peptide (CGRP)] and general neuroendocrine markers (neuron-specific enolase and chromogranin A). Quantitative analysis of the stained fibres and cells was performed using a graphic AutoCAD program. The presence and distribution of NPY, CGRP and SP nerve fibres and neuroendocrine cells were similar among the three subject groups. AR subjects had significantly higher tissue concentrations of VIP and SP. AR subjects had increased numbers of VIP fibres which predominantly innervated vessels. Thus, VIP and SP play important neuroimmunological roles in the pathogenesis of AR.     

Human and clinical perspectives on leptin

The neurochemical coding of neurones located in ganglia of the nerve trunk accompanying the chicken ureter was analysed and quantified using NADPH-diaphorase reactivity and immunohistochemistry against tyrosine hydroxylase (TH), nitric oxide synthase (NOS), calbindin (CAL), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), somatostatin (SOM), substance P (SP) and calcitonin gene-related peptide (CGRP) in untreated or colchicine-treated preparation. Almost all neurones were either positive for TH (38%) or for SOM (60%). Only 4% of the neurones were both TH- and SOM-positive and 3% of the neurones exhibited neither TH nor SOM immunoreactivity. The relative numbers of NPY-, NOS-, CAL- and VIP-positive neurones were 57%, 28%, 14% and 7%, respectively. No SP- or CGRP-positive neurones were observed. All NADPH-diaphorase-positive neurones expressed NOS immunoreactivity. Only in some TH-positive neurones was NPY and/or NOS found. Four major subpopulations were found in the ureteric ganglia. The SOM-positive neurones were subdivided into SOM/NPY/NOS- (28% of all neurones), SOM/NPY- (18%) and SOM/CAL/NPY-positive neurones (14%). A subpopulation of these peptid- ergic neurones also contained VIP. About 35% of the neurones contained TH only. Neurones of all subpopulations (72% of the neurones), except most of the CAL-positive neurones, were encircled by dense plexus of varicose SP/CGRP-positive, presumably sensory nerve fibres. Dense plexus of VIP-positive fibres were observed around 89% of the neurones. The chemical coding of the neuronal subpopulations identified in the ganglia accompanying the chicken ureter resembled that observed in the ganglia of Remak's nerve but was remarkably different from that of the autonomic neurones described in mammalian species.     

Hirschsprung's disease--immunohistochemical findings

Hirschsprung's disease (HSCR) is characterized by a non-propulsive distal intestinal segment (usually colon) leading to a functional obstruction. An absence of ganglia in the affected segment explains the synonymous term "aganglionosis coli". The lack of peristalsis is partly due to a deficient intestinal smooth muscle relaxation based on an absence of non-adrenergic, non-cholinergic (NANC) inhibitory innervation. Morphological studies using conventional microscopy, immunohistochemistry and immunochemistry against general neuronal markers and neuropeptides have been used to characterize the disturbed NANC innervation in HSCR. An increased cholinergic and adrenergic innervation is registered in the aganglionic segment in spite of the lack of neuronal cell bodies: Neuropeptides like vasoactive intestinal peptide (VIP), pituitary adenylate cyclase-activating polypeptide (PACAP), gastrin-releasing peptide (GRP), calcitonin gene-related peptide (CGRP), substance P (SP), enkephalins and galanin immunoreactive nerve fibres are all reduced in number in the aganglionic segment. In contrast, neuropeptide Y (NPY)-containing nerve fibres are increased in number in the diseased segment, probably reflecting the adrenergic hyperinnervation. General neuronal markers including chromogranins have been used to map the neuronal network in the HSCR intestine and also to investigate the endocrine cell system in the intestinal mucosa. Nitric oxide is a potent component of the NANC inhibitory innervation and its synthesizing enzyme, nitric oxide synthase (NOS), is shown to be almost absent in the neuronal system in aganglionic intestine.     

Sequence diversity of SIV(Mne) Nef in vivo and in vitro

The immune system and nervous system are intimately related. In addition to neuroendocrine mechanisms, neuropeptides have a variety of effects on immune cells and are responsible at least in part for neurogenic inflammation. The presence of neuropeptides in the skin has been well documented. The influence of neuropeptides on Langerhans cells is the focus of this paper. The physical presence and effects of calcitonin gene-related peptide on Langerhans cells is emphasized. Discussion also includes the putative inflammatory and immunologic roles of vasoactive intestinal peptide, substance P, neurotensin, neuropeptide Y, and somatostatin in the skin.     

Location, immunohistochemical features, and spinal connections of autonomic neurons innervating the rat seminal vesicles

With the use of retrograde tracing techniques, selective spinal nerve transections, and immunohistochemistry to label noradrenergic and peptidergic pathways, this study has for the first time defined in detail the autonomic innervation to the rat seminal vesicles. The majority of this innervation originates from the bilateral major pelvic ganglia, whereas very few neurons are located in the accessory, inferior mesenteric, or paravertebral chain ganglia. Neuropeptide Y was the most abundant marker, followed by tyrosine hydroxylase (an enzyme involved in noradrenaline synthesis), and then vasoactive intestinal peptide. Sympathetic axons with tyrosine hydroxylase and neuropeptide Y supplied vascular and nonvascular smooth muscle whereas parasympathetic, cholinergic neuropeptide Y terminals were associated with the glandular epithelium. In contrast, vasoactive intestinal peptide was found only in cholinergic neurons, which may have either parasympathetic or sympathetic spinal connections. The latter were far more prevalent, demonstrating a substantial sympathetic cholinergic innervation to the seminal vesicles. Vasoactive intestinal peptide axons were associated with the glandular epithelia, as well as vascular and nonvascular smooth muscle. Axons associated with the secretory epithelia may regulate secretion or perhaps provide trophic support. Finally, acute damage to preganglionic sacral and lumbar nerves caused a transient increase in glandular weight.     

Dendritic morphology of presumptive vasoconstrictor and pilomotor neurons and their relations with neuropeptide-containing preganglionic fibres in lumbar sympathetic ganglia of guinea-pigs

We have used intracellular dye-filling combined with multiple-labelling immunofluorescence to examine the dendritic morphology of neurons and their relations with neuropeptide-containing preganglionic terminals in the lumbar sympathetic chain of guinea-pigs. Presumptive vasoconstrictor neurons with immunoreactivity for both tyrosine hydroxylase and neuropeptide Y dendritic fields that were significantly smaller, on average, than those of presumptive pilomotor neurons containing immunoreactivity to tyrosine hydroxylase but not to neuropeptide Y. However, there was considerable variation in the sizes of the dendritic fields of the vasoconstrictor neurons. Preganglionic nerve terminals containing immunoreactivity to calcitonin gene-related peptide, but not to substance P, only surrounded cell bodies of vasoconstrictor neurons containing immunoreactivity to tyrosine hydroxylase and neuropeptide Y. In most cases, the neuropeptide-containing preganglionic terminals were not associated closely with the distal dendrites of these neurons. Few neuropeptide-containing terminals were associated closely with either the cell bodies or dendrites of the pilomotor neurons. These results show that there is a considerable range in the size of dendritic trees of sympathetic final motor neurons. Some of this variation is related to the pathways within which the neurons lie, so that presumptive pilomotor neurons generally are larger than presumptive vasoconstrictor neurons. The marked variation in size of vasoconstrictor neurons raises the possibility that there may be a size dependent recruitment of these neurons, similar to that seen in pools of spinal motor neurons. The distribution of the peptide-containing preganglionic endings suggests that they would act predominantly at the cell body and proximal dendrites of the final motor neurons.     

Spectral and interferometrical study of the interaction of haemin with glutathione

Neurones in the ureterovesical ganglion complex provide autonomic innervation to the pelvic ureter, the ureterovesical junction and the bladder trigone. We examined the distribution and peptide co-expression pattern of nitric oxide synthase (NOS) in the human ureterovesical ganglia by combining NADPH-diaphorase histochemistry with immunoreactivity for vasoactive intestinal peptide (VIP), neuropeptide Y (NPY), and calcitonin gene-related peptide (CGRP). Less than 20% of nerve cells in the large ganglia of the ureterovesical complex were stained for NOS activity. In elderly individuals, ganglion cells regularly exhibited conspicuous morphological alterations suggestive of degenerative changes. Most of the NOS-positive cell bodies costained for VIP-immunoreactivity. A minority of NOS-expressing cells also reacted for NPY-immunoreactivity. CGRP-immunoreactivity was present in varicose terminal-like nerve fibres which were found to encircle NOS-containing perikarya. Occasionally, NOS-positive somata were surrounded by plexiform axon terminals which immunostained for VIP or NPY. We conclude that the passage of urine across the ureterovesical junction is under relaxatory control of a local nitric oxide/VIP(NPY) pathway which may be modulated by preganglionic efferent and/or primary afferent input.     

Vasoactive intestinal polypeptide (VIP)--immunoreactive nerve fibres in the mammary gland of the pig

Immunohistochemical studies have been performed to investigate the coexistence of VIP with dopamine-beta-hydroxylase (D(beta)H), vesicular acetylcholine transporter (VAChT), somatostatin (SOM) or neuropeptyd Y (NPY) within nerve fibres supplying the immature mammary gland in the pig. Generally, a moderate number of the VIP-immunoreactive (VIP-IR) nerve fibres were located in the nipple and parenchyma of the gland. VIP-IR fibres surrounded smooth muscle cells (SMC), blood vessels (BV) and lactiferous ducts (LD). Double-labelling immunohistochemistry revealed that some of VIP-IR nerve fibres also contained immunoreactivity to D(beta)H. VIP/D(beta)H-IR nerves were associated with BV and SMC and single fibres were observed around the LD in both nipple and parenchyma of the gland. VIP/VAChT-IR nerve fibres were not observed. The majority of VIP-IR fibres associated with SMC were also SOM-IR. Less numerous VIP/SOM-IR fibres supplied the BV and were located around the LD of the gland. A small number of VIP-IR nerves also displayed immunoreactivity to NPY. VIP/NPY-IR nerve fibres supplied the BV of the gland.     

Blood vessels in liver metastases from both sarcoma and carcinoma lack perivascular innervation and smooth muscle cells

Hepatic arterial infusion (HAI) chemotherapy as treatment for human colorectal liver metastases is promising, but not entirely satisfactory. Improved drug delivery during HAI may be achieved by manipulating the different control mechanisms of normal versus tumour blood vessels. The peptidergic/aminergic innervation of vessels in normal liver and in two animal models of liver metastasis (Lister Hooded rat with syngeneic MC28 sarcoma; athymic (nude) rat with human HT29 carcinoma) was investigated to assess the suitability of these models for future pharmacological studies. Normal liver and metastases were studied immunohistochemically for the presence of protein gene product 9.5 (PGP), neuropeptide Y (NPY), tyrosine hydroxylase (TH), calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP) and substance P (SP). Perivascular innervation was also examined by transmission electron microscopy. In Lister rat normal livers, perivascular immunoreactive nerve fibres containing PGP, NPY, TH, CGRP and SP were observed around the interlobular blood vessels near the hilum and in the portal tracts. The highest density was seen for PGP, followed in decreasing order, by NPY, TH, CGRP and SP. VIP-immunoreactive nerves were absent. No immunoreactive nerves were observed in the hepatic lobule. In athymic rat livers, the pattern of innervation was similar, except that SP immunoreactivity was more sparse. No perivascular immunoreactive nerves were observed in either MC28 or HT29 tumours. Electron microscopy confirmed the absence of perivascular nerves. Smooth muscle cells were not observed in tumour blood vessel walls. These results are comparable with previous observations on human liver metastases and suggest that the animal models may be suitable for pharmacological studies on vascular manipulation of HAI chemotherapy.     

Partial and complete sciatic nerve injuries induce similar increases of neuropeptide Y and vasoactive intestinal peptide immunoreactivities in primary sensory neurons and their central projections

Partial nerve injury is more likely to cause neuropathic pain than complete nerve injury. We have compared the changes in neuropeptide expression in primary sensory neurons which follow complete and partial injuries to determine if these might be involved. Since more neurons are damaged by complete injury, we expected that complete sciatic nerve injury would simply cause greater increases in neuropeptide Y and vasoactive intestinal peptide than partial injury. We examined neuropeptide Y and vasoactive intestinal peptide immunoreactivities in L4 and L5 dorsal root ganglia, the dorsal horn of L4-L5 spinal cord, and the gracile nuclei of rats killed 14 days after unilateral complete sciatic nerve transection, partial sciatic nerve transection and chronic constriction injury of the sciatic nerves. In all three groups of rats, neuropeptide Y- and vasoactive intestinal peptide-immunoreactive neurons were increased in the ipsilateral L4 and L5 dorsal root ganglion when compared with the contralateral side. Most neuropeptide Y-immunoreactive neurons were of medium and large size, but a few were small. Neuropeptide Y-immunoreactive axonal fibers were increased from laminae I to IV, and vasoactive intestinal peptide-immunoreactive axonal fibers were increased in laminae I and II, of the ipsilateral dorsal horn of L4-L5 spinal cord. The increases of neuropeptide Y and vasoactive intestinal peptide immunoreactivities in the dorsal horn were similar among the three groups. However, only after constriction injury were some vasoactive intestinal peptide-immunoreactive neurons seen in the deeper laminae of the ipsilateral dorsal horn. Robust neuropeptide Y-immunoreactive axonal fibers and some neuropeptide Y-immunoreactive cells were seen in the ipsilateral gracile nuclei of all three groups of animals, but neuropeptide Y-immunoreactive cells were more prominent after constriction injury. Contrary to our expectations, partial and complete sciatic nerve injuries induced similar increases in neuropeptide Y and vasoactive intestinal peptide in lumbar dorsal root ganglion neurons and their central projections in the dorsal horn and the gracile nuclei two weeks after injury. Some neurons whose axons were spared by partial injury may also increase neuropeptide Y or vasoactive intestinal peptide expression. Altered neuropeptide release from these functional sensory neurons may play a role in neuropathic pain.     

A role for HEM2 in cadmium tolerance

The innervation of the knee joint synovial membrane of the guinea pig, i.e., the synoviocyte layer, the subjacent connective tissue and the connective tissue region beneath, was analyzed with immunohistofluorescence and electron microscopy. A screening of the innervation with antibodies against the general axon marker -- protein gene product (PGP) 9,5 -- revealed the presence of nerve fibers distributed in various regions of the knee joint synovial membrane. Confirming previous studies, some of these nerve fibers stained with antibodies to tyrosine hydroxylase (TH), neuropeptide Y (NPY), substance P (SP), calcitonin gene-related peptide (CGRP), and vasoactive intestinal polypeptide (VIP). In addition, dynorphin (DYN)-containing fibers were detected, which have not been reported previously in normal joints. In general, the immunoreactive fibers were observed close to the synoviocytes and at blood vessels. Fibers with colocalization of NPY- and TH-like immunoreactivities (LIs), as well as of DYN- and TH-LIs were demonstrated. In the electron microscope, bundles of unmyelinated fibers as well as single fibers were found in the connective tissue region below the synoviocytes. Varicose parts of the nerve fibers contained mainly small, clear vesicles. Small and large dense-cored vesicles were also seen, but less frequently. Denser portions of the plasma membranes of some axons were observed in these regions, facing the extracellular space. Myelinated fibers were also observed in some nerve bundles. These findings emphasize the complex innervation of the synovial membrane, with nerve fibers containing a host of neuroactive substances. Altogether, these fibers are probably involved in many functions such as vasoregulation and control of synovial secretion in addition to being a source of mediators in joint inflammation.     

Genome-wide analysis of integral membrane proteins from eubacterial, archaean, and eukaryotic organisms

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.     

Stimulation of retinal pigment epithelial cell growth by neuropeptides in vitro

PURPOSE: The proliferation of many cell types are regulated by cytokines and neuropeptides by autocrine and paracrine mechanisms. Retinal pigment epithelial (RPE) cells are also regulated by cytokines. But RPE cells are very close to the neural retina which has some neuropeptides. The present study was to investigate the effects of neuropeptides on the growth of RPE cells. METHODS: RPE cells were obtained from the eyes of 11 day old chick embryos and cultured in Dulbecco's modified Eagle's culture medium containing 10% fetal calf serum. The growth of RPE cells was evaluated by [3H]-thymidine uptake. RESULTS: Substance P, beta-endorphin and calcitonin gene-related peptide markedly stimulated the growth of RPE cells. The effects of methionine-enkephalin, somatostatin and vasoactive intestinal peptide were intermediate. The strongest effects of substance P, beta-endorphin and calcitonin gene-related peptide were observed at 10(-6) to 10(-7) M. The stimulation of RPE cells with beta-endorphin was inhibited by naloxone, suggesting that the stimulation with beta-endorphin is mediated by an opioid receptor. beta-endorphin and substance P induced RPE cell growth stimulating activity. Leucine-enkephalin and neuropeptide Y did not affect the growth of RPE cells. CONCLUSIONS: These results suggest that neuropeptides play an important role in the regulation of RPE cell growth.     

Sensory and autonomic innervation of the facet joint in the rat lumbar spine

The presence of sensory and autonomic nerves in the synovial membrane of the lumbar facet joint in rats was investigated by immunohistochemistry. Substance P and calcitonin gene-related peptide immunoreactivities, representing sensory nerves, were observed as varicose fibers in the synoviocyte layer. The fibers were predominantly nonvascular. The autonomic innervation was identified by the presence of neuropeptide Y- and tyrosine hydroxylase-positive fibers. Most of these fibers were found adjacent to or within blood vessel walls. Immunoreactivity to vasoactive intestinal polypeptide was seen in varicose nerve terminals in the synoviocyte layer, mostly unrelated to blood vessels. There is accumulating evidence of an involvement of both the sensory and sympathetic nervous systems in inflammatory joint disease. The neuropeptides now identified in lumbar facet joints may prove to play a significant role in the pathogenesis of low-back pain.     

Peptide-containing neurons remain unaffected after intestinal autotransplantation: an experimental study in the piglet

The gut is richly supplied with peptide-containing nervous elements. In the present immunocytochemical study the origin, occurrence and topographical distribution of nerves containing vasoactive intestinal polypeptide (VIP), enkephalin, substance P (SP), somatostatin, neuropeptide Y (NPY), calcitonin gene-related peptide (CGRP), gastrin-releasing peptide (GRP) and galanin were investigated in the porcine small intestine. In order to study the origin (extrinsic or intrinsic) of the nerve fibers, specimens from autotransplanted and extrinsically denervated jejunum were examined. Furthermore, possible changes in the distribution of intrinsic neurons after extrinsic denervation were studied. In the control jejunum each nerve fiber population had its own characteristic topographic distribution. There was no overt difference in distribution pattern of peptide-containing nerve fibers and cell bodies between the transplanted and the control segment except that NPY-, SP- and CGRP-containing nerve fibers disappeared around blood vessels. Thus VIP-, somatostatin-, GRP-, enkephalin- and galanin-containing nerve fibers were visibly unchanged in the transplanted segment. The results support the view that the peptide-containing nerve fibers are mainly intrinsic in origin except the NPY-, SP- or CGRP-containing perivascular nerve fibers which are extrinsic to the gut wall. In addition, the results of the present study suggest that transplantation and extrinsic denervation have no major effect on the distribution pattern of the intrinsic neuronal systems.     

Nitric oxide in the stress axis

Calcitonin gene-related peptide in sensory primary afferent neurons has an excitatory effect on postsynaptic neurons and potentiates the effect of substance P in the rat spinal dorsal horn. It has been established that calcitonin gene-related peptide expression in dorsal root ganglion neurons is depressed, and the effect of calcitonin gene-related peptide on dorsal horn neurons is attenuated, following peripheral nerve injury. We report here that a subpopulation of injured dorsal root ganglion neurons show increased expression of calcitonin gene-related peptide. Using in situ hybridization and the retrograde tracer, FluoroGold, we detected an increased number of medium- to large-sized rat dorsal root ganglion neurons projecting to the gracile nucleus that expressed alpha-calcitonin gene-related peptide messenger RNA following spinal nerve transection. Immunohistochemistry revealed a significant increase in calcitonin gene-related peptide immunoreactivity in the gracile nucleus and in laminae III-IV of the spinal dorsal horn. These results indicate that a subpopulation of dorsal root ganglion neurons express alpha-calcitonin gene-related peptide messenger RNA in response to peripheral nerve injury, and transport this peptide to the gracile nucleus and to laminae III-IV of the spinal dorsal horn. The increase of the excitatory neuropeptide, calcitonin gene-related peptide, in sites of primary afferent termination may affect the excitability of postsynaptic neurons, and have a role in neuronal plasticity following peripheral nerve injury.