Immunohistochemical evidence for the presence of calcium-binding proteins in enteric neurons

Summary Immunoreactivity for vitamin D-dependent calcium-binding protein (CaBP) has been localized in nerve cell bodies and nerve fibres in the gastrointestinal tracts of guinea-pig, rat and man. CaBP immunoreactivity was found in a high proportion of nerve cell bodies of the myenteric plexus, particularly in the small intestine. It was also found in submucous neurons of the small and large intestines. Immunoreactive nerve fibres were numerous in the myenteric ganglia, and were also common in the submucous ganglia and in the intestinal mucosa. Immunoreactive fibres were rare in the circular and longitudinal muscle coats. In the myenteric ganglia of the guinea-pig small intestine the immunoreactivity is restricted to one class of nerve cell bodies, type-II neurons of Dogiel, which display calcium action potentials in their cell bodies. These neurons were also immunoreactive with antibodies to spot 35 protein, a calcium-binding protein from the cerebellum. From the distribution of their terminals and the electrophysiological properties of these neurons it is suggested they might be sensory neurons, or perhaps interneurons. The discovery of CaBP in restricted sub-groups of enteric neurons may provide an important key for the analysis of their functions.     

Neuropeptide Y-like immunoreactivity in intramural ganglia of the newborn guinea pig urinary bladder

Immunoreactive neuropeptide Y (NPY) was demonstrated in neuronal elements in the urinary bladder wall of the newborn guinea pig. Numerous intramural ganglia were found lying among the smooth muscle bundles and in the submucosa, and NPY-like immunoreactive nerve cell bodies were demonstrated within all of these ganglia. Nerve fibres containing NPY were also richly distributed in the detrusor muscle, submucosa and around blood vessels. In dissociated cell cultures from newborn guinea pig detrusor muscle, a subpopulation (70-85%) of both mononucleate and binucleate intramural neurones was shown to contain NPY-like immunoreactivity. A low percentage (1-6%) of the intramural bladder neurones contained dopamine-beta-hydroxylase. In conclusion, while some NPY-containing nerve fibres in the wall of the bladder are of sympathetic origin, especially those supplying blood vessels, the results of this present study establish that many of these NPY-containing nerve fibres originate from non-adrenergic cell bodies within the intramural bladder ganglia.     

Origin and development of VIP and substance P containing neurons in the embryonic avian gut

The development of substance P (SP) and VIP containing structures of the quail and chick guts was studied by immunocytochemistry. The appearance of VIP and substance P nerves follows a rostrocaudal pattern from day 9 in the quail and day 10 in the chick embryo. Immunoreactive fibres are first visible in the oesophagus and at 12 days they extend over the whole length of the intestine. VIP and substance P ganglionic cells are first localized in the foregut (day 9 for VIP containing neurons and day 13 for SP ones) and observed in the mid- and hind-gut just before hatching. Transplantation on the chorioallantoic membrane (CAM) of fragments of various parts of the digestive tract were carried out to see whether in such circumstances the pattern of VIP and SP containing nerves was comparable to normal. The explants contained numerous SP and VIP immunofluorescent nerve fibres. In addition, cell bodies with VIP and SP immunoreactivity appeared brightly fluorescent in the enteric ganglia of the graft showing that these peptidergic nerve cells belong to the intrinsic innervation of the gut.     

Origin and development of VIP and substance P containing neurons in the embryonic avian gut

Summary The development of substance P (SP) and VIP containing structures of the quail and chick guts was studied by immunocytochemistry. The appearance of VIP and substance P nerves follows a rostrocaudal pattern from day 9 in the quail and day 10 in the chick embryo. Immunoreactive fibres are first visible in the oesophagus and at 12 days they extend over the whole length of the intestine. VIP and substance P ganglionic cells are first localized in the foregut (day 9 for VIP containing neurons and day 13 for SP ones) and observed in the mid- and hind-gut just before hatching. Transplantation on the chorioallantoic membrane (CAM) of fragments of various parts of the digestive tract were carried out to see whether in such circumstances the pattern of VIP and SP containing nerves was comparable to normal. The explants contained numerous SP and VIP immunofluorescent nerve fibres. In addition, cell bodies with VIP and SP immunoreactivity appeared brightly fluorescent in the enteric ganglia of the graft showing that these peptidergic nerve cells belong to the intrinsic innervation of the gut.     

Distribution of neuropeptides in the porcine stellate ganglion

The localization and distribution of neuropeptides including neuropeptide Y (NPY), [Met⁵]enkephalin-Arg⁶-Gly⁷-Leu⁸ (MEAGL), vasoactive intestinal polypeptide (VIP), calcitonin gene-related peptide (CGRP), substance P and somatostatin (SOM) were analyzed in the stellate ganglion of the pig by use of the indirect immunofluorescence technique. NPY, MEAGL, SOM, VIP and CGRP immunoreactivities were found to exist in subpopulations of neuronal cell bodies of the stellate ganglion. A population of the small intensely fluorescent (SIF) cells showed MEAGL immunoreactivity. In addition, the presence of NPY-, MEAGL-, CGRP-, SP-, SOM- and VIP-immunoreactive nerve fibers and axonal varicosities were observed in the stellate ganglion. The localization and pattern of distribution of these peptides in the porcine stellate ganglion were compared with studies carried out on stellate ganglia of other mammalian species.     

Nerves containing substance P,vasoactive intestinal polypeptide,enkephalin or somatostatin in the guinea-pig taenia coli

Summary The guinea-pig taenia coli is rich in peptide-containing nerves. Nerve fibres containing substance P (SP), vasoactive intestinal peptide (VIP), or enkephalin, were numerous in the smooth muscle while somatostatin fibres were very few. Nerve fibres displaying SP or VIP immunoreactivity were numerous in the myenteric plexus. Enkephalin nerve fibres were fairly numerous in the plexus while somatostatin nerve fibres were sparse. Nerve cell bodies containing immunoreactive SP or VIP were regularly seen in the plexus. Delicate varicose elements of the different types of nerve fibres were found to ramify around nerve cell bodies in a manner suggestive of innervation.In the electron microscope the various peptide-storing nerve fibres (i.e., elements containing SP, VIP or enkephalin) were found to contain a varying number of fairly large, electron-opaque vesicles in the varicose swellings. These vesicles represent the storage site of the neuropeptides.The isolated taenia coli responded to electrical nerve stimulation with a contraction. After cholinergic and adrenergic blockade the contractile response was replaced by a relaxation followed by a contraction upon cessation of stimulation. SP contracted the taenia while VIP caused a relaxation. The enkephalins raised the resting tension slightly while somatostatin had no effect. These observations are compatible with a role for SP as an excitatory neurotransmitter and for VIP as an inhibitory one, and with the view that both SP neurones and VIP neurones act as motor neurones. In preparations contracted by SP the electrically induced contractions were reduced in amplitude while the electrically induced relaxations seen after adrenergic and cholinergic blockade were enhanced in amplitude. In preparations relaxed by VIP there was an increased contractile response to electrical stimulation, while in the atropine + guanethidine-treated preparation the electrically induce relaxations were reduced in amplitude. The enkephalins reduced the contractile response to electrical stimulation, while somatostatin induced a very small reduction in the amplitude of such responses. These observations suggest that SP neurones and VIP neurones may play additional roles as interneurones. Somatostatin neurones probably act as interneurones. Enkephalin-containing fibres may serve to modify the release of transmitter from other nerves in the smooth muscle, perhaps through axo-axonal arrangements. Alternatively, the enkephalin nerve fibres in the smooth muscle are afferent elements involved in mediating sensory impulses to the myenteric plexus.     

Origin and distribution of neuropeptide Y-, vasoactive intestinal polypeptide- and substance P-containing nerve fibers in the urinary bladder of the rat

Summary The origin and distribution in the urinary bladder of nerve fibers containing neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP) and substance P (SP) were investigated in rats. Experimental procedures comprised preganglionic decentralization or postganglionic denervation of the bladder and also chemical sympathectomy as well as capsaicin treatment of newborn rats.Nerve fibers containing NPY were richly distributed in the detrusor muscle and also in the pelvic ganglia. Numerous NPY-containing nerve cell bodies were found in pelvic ganglia. A rich occurrence of VIP fibers and a more sparse distribution of SP-containing fibers were also found in the bladder as well as a relatively rich representation of VIP- containing nerve cell bodies in the pelvic ganglia. After decentralization the intensity of VIP and NPY immunofluorescence increased in nerve cell bodies of the pelvic ganglia and in nerve fibers in the wall of the bladder. Postganglionic denervation, on the other hand, eliminated all peptides examined in the bladder wall. After postganglionic denervation the situation in the ganglia was approximately the same as after decentralization. Chemical sympathectomy (6-OHDA) did not seem to change significantly the frequency and distribution of VIP-, SP- and NPY-fibers in the muscle layer of the bladder or in the pelvic ganglia, while the NPY-containing nerve fibers in the submucosal layer and around blood vessels of the bladder disappeared. Adrenergic nerve fibers in the wall of the bladder (visualized by histofluorescence) were markedly reduced in number after administration of 6-OHDA. Capsaicin-treatment of newborn rats caused a loss of SP-fibers in the wall of the bladder, supporting the view that these fibers are sensory in nature in the urinary bladder. Although it cannot be entirely excluded that NPY-containing fibers in the wall of the bladder are adrenergic, the present results suggest that the NPY-fibers as well as the VIP-fibers of the bladder wall originate mainly in non-adrenergic cell bodies of the pelvic ganglia. However, perivascular NPY-containing nerve fibers are adrenergic in nature.     

Immunocytochemical localization of substance P in the neurohypophysis and hypothalamus of the mouse compared with the distribution of other neuropeptides

Summary Substance-P immunoreactivity has been located in semithin sections of mouse hypothalami and pituitaries and compared with the distribution of other hypothalamic peptides. In the mouse, nerve fibres and terminals reacting with antibodies against substance P (SP) were detected both in the external zone of the median eminence (ME) and in the neural lobe of the pituitary. Immunoreactive SP (ISP) axons of the ME did not react with antibodies against other peptides, i.e. arginine-vasopressin (AVP), oxytocin (OT), somatostatin and enkephalin, and were also negative with an antibody to serotonin. In the neural lobe, SP immunostaining occurred in AVP but not in OT axons. In the hypothalamus, ISP axons were widespread but conspicuously lacking in areas containing AVP neurones, i. e. in the suprachiasmatic nucleus and the clusters of AVP cells in the SO and PV nuclei. In contrast, multiple ISP endings were seen in contact with OT neurones. Immunoreactive cell bodies, only detected after colchicine treatment, belonged to two distinct classes of neurones: 1) single AVP neurones of the SO and PV nuclei; 2) specific (staining only for SP) neurones, scattered or grouped in different areas of the hypothalamus, not showing relationship with any circumscribed nucleus. These results reinforce the opinion that SP can be released as a neurohormone into the vascular portal system and can directly affect the pars distalis. The presence of immunoreactive SP in the neural lobe, which has not been reported in species other than the mouse, may have a different physiological significance.     

Cholinergic, nitrergic and peptidergic (Substance P- and CGRP-utilizing) innervation of the horse intestine. A histochemical and immunohistochemical study

The small and large intestine of adult horses were histochemically and immunohistochemically investigated in order to evidence components of the intramural nervous system. The general structural organization of the intramural nervous system was examined by using Nissl-thionin staining as well as the anti-neurofilament 200 (NF200) immunoreaction, which demonstrated the presence of neurons in the submucous as well as myenteric plexuses. The additional presence of subserosal ganglia was shown in the large intestine. Acetylcholinesterase (AChEase) activity was observed in both the submucous and myenteric plexuses. Localization of acetylcholine-utilizing neurons was also evidenced by immunohistochemical reactions for choline acetyltransferase (ChAT) and vesicular acetylcholine transporter (VAChT). With both histochemistry and immunohistochemistry possible cholinergic nerve fibres were detected in the inner musculature. The two possible cholinergic co-mediators Calcitonin Gene-Related Peptide (CGRP) and Substance P (SP) have been investigated by an immunohistochemical approach. CGRP immunoreactivity was detected in roundish nerve cell bodies as well as in nerve fibres of the submucous plexus, whereas SP immunoreactivity was evidenced in nerve fibres of the tunica mucosa, in nerve cell bodies and fibres of the submucous plexus and in nerve fibres of the myenteric plexus. NADPH-diaphorase reactivity, which is linked to the synthesis and release of nitric oxide, was detected in nerve cell bodies and nerve fibres of both the submucous and myenteric plexuses as well as in a subserosal localization of the large intestine. The nitrergic components were confirmed by the anti-NOS (nitric oxide synthase) immunoreaction. Results are compared with those of other mammals and related to the complex intestinal horse physiology and pathophysiology.     

The distribution and origin of VIP in the spinal cord of six mammalian species

The distribution of VIP-immunoreactivity was studied in the spinal cord and dorsal root ganglia of 6 mammalian species. Immunoreactive fibres and cell bodies were most apparent in the dorsal horn, dorsolateral funiculus, intermediolateral cell columns and the area around the central canal. The distribution of VIP immunoreactivity was similar in all species studied, mouse, rat, guinea pig, cat, horse and the marmoset monkey. There were fewer VIP fibres in the dorsal horn of cervical and thoracic segments than in lumbosacral segments. Using radioimmunoassay this gradient increase was quantitatively most marked in the sacral spinal cord of the cat. In dorsal root ganglia few nerve cell bodies but numerous fibres were present. A dual origin for VIP in the spinal cord is suggested: (A) Extrinsic, from dorsal root afferent fibres since immunoreactivity was decreased in dorsally rhizotomized animals (cats and rats) and in capsaicin pretreated rats (microinjection of dorsal root ganglia). (B) From local cell bodies intrinsic to the spinal cord which became visible after colchicine pretreatment of rats.     

Evidence for 5-hydroxytryptamine in neurones in the gut of the toad,Bufo marinus

Summary The stomach, small intestine and large intestine of the toad, Bufo marinus, were processed for formaldehyde-induced fluorescence histochemistry. After extrinsic denervation or pretreatment with 6-hydroxydopamine to remove catecholamine fluorescence, yellow fluorescence typical of 5-hydroxytryptamine was observed in neurones in the small intestine only. The cell bodies and their processes were confined to the myenteric plexus. Additional pretreatment with 5-hydroxytryptamine enhanced the fluorescence of neurones in the small intestine and revealed yellowfluorescent nerve fibres, but not cell bodies, in the longitudinal and circular muscle layers and myenteric plexus of the large intestine. No fluorescent neurones were observed in the stomach. Following reserpine treatment, which removed native yellow fluorescence in the small intestine, exposure to 5-hydroxytryptophan produced yellow fluorescence in axons in both small and large intestine; exposure to tryptophan never restored fluorescence. The neurotoxin, 5,7-dihydroxytryptamine had no effect on the distribution of yellow-fluorescent neurones in the small and large intestine. No 5-HT-containing mast cells were present in either the small or large intestine. Thin layer chromatography with three different mobile phases showed a 5-hydroxytryptamine-like compound in extracts of mucosa-free small and large intestine but not of stomach.     

Distribution of subgroups of noradrenaline neurons in the coeliac ganglion of the guinea-pig

Summary The distributions within the coeliac ganglion of different chemically coded subgroups of noradrenaline neurons, and the relationships between these neurons and nerve fibres projecting to the ganglion from the intestine, have been assessed quantitatively by use of an immunohistochemical double-staining method. Noradrenaline (NA) neurons made up 99% of all cell bodies. Of these, 21% were also reactive for somatostatin (NA/SOM neurons), 53% were also reactive for NPY (NA/NPY neurons), and 26% were not reactive for either peptide. NA neurons without reactivity for any of the peptides whose localization was tested have been designated NA/-. A small percentage, about 1%, of neurons were reactive for both NPY and SOM. The three major types of NA neurons were arranged in clumps or ribbons throughout the ganglia, with a tendency for NA/SOM neurons to be medial and NA/NPY neurons to be lateral in the ganglia. A small group of neurons (<1%) encoded with dynorphin, NPY and vasoactive intestinal peptide (VIP) was encountered. VIP-immunoreactive nerve terminals, projecting to the ganglion from cell bodies in the intestine, ended around NA/SOM and NA/neurons but not around NA/NPY neurons. Thus, the VIP axons from the intestine end selectively around neurons that modify intestinal function (NA/SOM and NA/-neurons) but not around neurons, the terminals of which supply blood vessels (NA/NPY neurons).     

Neuronal subpopulations in autonomic ganglia associated with the chicken ureter: an immunohistochemical study

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.     

Oxytocin is expressed throughout the human gastrointestinal tract

BACKGROUND/AIM: Several studies have described that oxytocin exerts stimulatory or inhibitory effects on gut functions. Recently, mRNA for oxytocin and its receptor was found throughout the entire human gastrointestinal (GI) tract. The aim of this study was to examine the cellular localization and distribution of the corresponding proteins. MATERIAL AND METHODS: Full-thickness biopsies from 24 patients, covering the entire GI tract, were collected during operations at the Department of Surgery in Malm? and Lund. The biopsies were taken from non_affected margins. The biopsies were fixed by immersion, rinsed in buffered sucrose, and kept frozen at 70 degrees C. Indirect immunofluorescence with primary antibodies to oxytocin and its receptor was used. RESULTS: Oxytocin was expressed in nerve cell bodies and nerve fibres in the myenteric and submucous ganglia all along the GI tract. Immunoreactive nerve cell bodies in myenteric ganglia predominated in the proximal (antrum and duodenum) and distal gut, while those in the submucous ganglia were more numerous in the ileum and colon. The oxytocin receptor was not detectable by two different antibodies in any tissue in the GI tract. CONCLUSION: Oxytocin is expressed in the myenteric and submucous ganglia and nerve fibres along the entire human GI tract. The role for oxytocin in the physiology and pathophysiology of the bowel remains to be settled.     

Nitric oxide synthase in the gastrointestinal tract of the estuarine crocodile, Crocodylus porosus

The aim of this study was to investigate the distribution of nitric oxide synthase (NOS)-containing nerve cells in the gastrointestinal tract of a reptile and to compare it with the pattern in other vertebrate classes. In the estuarine crocodile, Crocodylus porosus, NOS-positive nerve cell bodies and fibres were found in all regions of the gut examined. Most myenteric microganglia contained one or several NOS-immunoreactive neurons together with unlabelled neurons. The majority of the neurons were multipolar, ranging from 10 to 25 microns in diameter. Both the circular and the longitudinal muscle layers were innervated by NOS-immunoreactive nerve fibres, which mostly ran parallel to the muscle fibres. In addition, small blood vessels in the submucosa and on the serosal surface of the gut were innervated by NOS-immunoreactive fibres. Double labelling with antisera to NOS and vasoactive intestinal peptide (VIP) revealed three neuronal subpopulations. A small proportion of the NOS-immunoreactive cells also contained immunoreactivity to VIP while a majority of the VIP-immunoreactive cells were NOS immunoreactive. There were more nerve fibres showing VIP immunoreactivity than fibres with NOS immunoreactivity, although most of the latter also contained immunoreactivity to VIP. VIP-immunoreactive fibres often surrounded the NOS-immunoreactive nerve cells. These results suggest that neuronally released nitric oxide is likely to be involved in the control of gastrointestinal motility in the crocodile as in most other vertebrate species.     

Co-existence and origin of peptidergic and adrenergic nerves in the guinea pig uterus

Summary The occurrence and distribution of peptidergic nerves in the guinea pig uterus was studied by means of immunocytochemistry using numerous neuropeptide antisera. Neuropeptide Y (NPY)-immunoreactive (IR) nerves were the most abundant, whereas substance P (SP)-, calcitonine gene-related peptide (CGRP)-, and neurokinin A (NKA)-IR nerves were less frequent, and peptide histidine isoleucine (PHI)-IR nerves were the most sparse. Chemical sympathectomy by means of 6-hydroxydopamine, and capsaicin treatment revealed the division of the peptidergic nerves into three separate populations: (1) NPY-IR nerves, which co-existed with adrenergic nerves, (2) SP-, CGRP-and NKA-IR nerves, which mutually co-existed, and (3) PHI-IR nerves. Parallel-running adrenergic/NPY-IR and SP-IR nerves could be found with very similar although not completely identical morphological appearance. Paracervical ganglia contained neurotensin-and dynorphin A-IR cell bodies in addition to cell bodies with immunoreactivities similar to those in prevertebral ganglia. Combined retrograde tracing with True blue and immunocytochemistry showed that the adrenergic and NPY-IR uterine nerves originate in paracervical and prevertebral ganglia. In the prevertebral ganglia the cellular origin was the same for adrenergic and NPY-IR nerves. In contrast, SP-, CGRP-,and NKA-IR nerves originated in dorsal root ganglia. At full-term pregnancy all the neuropeptide immunoreactivities had vanished, probably reflecting a fetus-induced general nerve degeneration.     

A neurochemical characterisation of the golden hamster myenteric plexus

The neurochemical composition of nerve fibres and cell bodies in the myenteric plexus of the proventriculus, stomach and small and large intestines of the golden hamster was investigated by using immunohistochemical and histochemical techniques. In addition, the procedures for localising nitric-oxide-utilising neurones by histochemical (NADPH-diaphorase) and immunohistochemical (nitric oxide synthase) methods were compared. The co-localisation of vasoactive intestinal polypeptide and nitric oxide synthase in the myenteric plexus of all regions of the gut was also assessed. The results demonstrated the presence of nerve fibres and nerve cell bodies immunoreactive to protein gene product, vasoactive intestinal polypeptide, substance P, calcitonin gene-related peptide, tyrosine hydroxylase, 5-hydroxytryptamine and nitric oxide synthase in all regions of the gastrointestinal tract examined. The pattern of distribution of immunoreactive nerve fibres and nerve cell bodies containing the above markers was found to vary in different regions of the gut. Myenteric neurones and nerve fibres containing immunoreactivity to nitric oxide synthase and NADPH-diaphorase reactivity, however, were shown to have an identical distribution throughout the gut. In contrast to some studies on the guinea-pig and rat, the co-existence of vasoactive intestinal polypeptide and nitric oxide synthase was seen in only a small population of myenteric neurones.     

Distribution of peptide-containing neurons and endocrine cells in the rabbit gastrointestinal tract,with particular reference to the mucosa

Summary The distribution patterns of peptide-containing neurons and endocrine cells were mapped in sections of oesophagus, stomach, small intestine and large intestine of the rabbit, by use of standard immunohistochemical techniques. Whole mounts of separated layers of ileum were similarly examined. Antibodies raised against vasoactive intestinal peptide (VIP), substance P (SP), somatostatin (SOM), neuropeptide Y (NPY), enkephalins (ENK) and gastrin-releasing peptide (GRP) were used, and for each of these antisera distinct populations of immunoreactive (IR) nerve fibres were observed. Endocrine cells were labelled by the SP, SOM or NPY antisera in some regions.VIP-IR nerve fibres were common in each layer throughout the gastrointestinal tract. With the exception of the oesophagus, GRP-IR nerve fibres also occurred in each layer of the gastrointestinal tract; they formed a particularly rich network in the mucosa of the stomach and small intestine. Fewer nerve fibres containing NPY-IR or SOM-IR were seen in all areas. SOM-IR nerve fibres were very scarce in the circular and longitudinal muscle layers of each area and were absent from the gastric mucosa. The SP-IR innervation of the external musculature and ganglionated plexuses in most regions was rather extensive, whereas the mucosa was only very sparsely innervated. ENK-IR nerve fibres were extremely rare or absent from the mucosa of all areas, although immunoreactive nerve fibres were found in other layers.These studies illustrate the differences in distribution patterns of peptide-containing nerve fibres and endocrine cells along the gastrointestinal tract of the rabbit and also show that there are some marked differences in these patterns, in comparison with other mammalian species.     

The distribution and colocalization of neuropeptides and catecholamines in nerves supplying the gall bladder of the toad,Bufo marinus

The indirect immunofluorescence technique was used to determine the distribution of peptide-containing axons in the gall bladder of the cane toad, Bufo marinus. In addition, the adrenergic innervation of the gall bladder was examined by use of immunoreactivity to the catecholamine-synthesizing enzyme, tyrosine hydroxylase, and glyoxylic acid-induced fluorescence. On the basis of peptide coexistence, two intrinsic populations of neurones and their projecting fibres could be distinguished substance P neurones and vasoactive intestine peptide neurones. Neither of these two types of neurones contained any other colocalized neuropeptides. Four populations of nerve fibres arising from cell bodies outside the gall bladder were identified: nerves containing colocalized galanin, somatostatin and vasoactive intestinal peptide; nerves containing colocalized calcitonin gene-related peptide and substance P; adrenergic nerves containing neuropeptide Y; and nerves containing only adrenaline.     

Distribution of GABA-like immunoreactive neurons in the slug Limax maximus

Summary Immunohistochemical techniques were used to study the distribution of gamma-amino butyric acid (GABA)-like immunoreactive neurons in the nervous system of the slug Limax maximus. Approximately 170 GABA-like immunoreactive cell bodies were found in the central nervous system. These were located in the cerebral, buccal and pedal ganglia. Most GABA-like immunoreactive neurons had small cell bodies, which were aggregated into discrete clusters within the cerebral and pedal ganglia. Three pairs of longer, uniquely identifiable, GABA-like immunoreactive cells were found in the cerebral ganglion. GABA-like immunoreactive nerve fibres were also found in all of the central ganglia but were absent from peripheral nerves. These results suggest that GABA acts as a central neurotransmitter in the slug. The possible roles of GABA-ergic neurotransmission in the slug are discussed.