Neuronal localization of pancreatic polypeptide (PP) and vasoactive intestinal peptide (VIP) immunoreactivity in the earthworm (Lumbricus terrestris)

Nerve fibres and cell bodies displaying vasoactive intestinal polypeptide (VIP) or pancreatic polypeptide (PP) immunoreactivity were demonstrated in ganglia of the earthworm (Lumbricus terrestris). VIP cell bodies were found in the most anterior ganglion of the ventral nerve cord, the subpharyngeal ganglion. Immunoreactive nerves were seen running in the center of the cord until about the 10th segment. PP cell bodies were found in the cerebral ganglion where VIP was lacking, in the subpharyngeal ganglion and in more posteriorly located ganglia of the ventral nerve cord. PP nerve fibres could be followed below the 10th segment of the cord.     

Distinct localization of FMRFamide- and Bovine Pancreatic Polypeptide-like material in the brain, retrocerebral complex and suboesophageal ganglion of the cockroach Periplaneta americana L.

One bovine pancreatic polypeptide (BPP) antiserum and two FMRFamide antisera were applied in the peroxidase-antiperoxidase (PAP) immunohistochemical technique on a complete series of sections of brains, suboesophageal ganglia (SOG), corpora cardiaca (CC) and corpora allata of Periplaneta americana L. Double immunohistochemical staining demonstrated that the same perikarya and processes were stained by both the BPP and FMRFamide antisera. This was caused by cross-reaction of the BPP and FMRFamide antisera with common antigenic determinants as was shown by a number of solid-phase absorptions. Application of a third FMRFamide antiserum, which was especially selected for its inability to react with bovine and avian pancreatic polypeptide, showed that more than half of the structures that were stained with the ‘unspecific’ BPP and FMRFamide antisera, contained material which was genuinely FMRFamide-like. This peptide material was located in cerebral neuronal structures, in the SOG, in the storage site of the CC and in numerous nerve fibres throughout the neuropile regions, which suggested a neurotransmitter/modulator as well as a neurohormonal role. The FMRFamide-like peptide was also found to be present in the same brain sites as an adipokinetic hormone-like peptide, but double labelling revealed that these two substances were never located in the same perikarya or fibres.     

FMRFamide- and adipokinetic hormone-like immunoreactivity in the nervous system of the mosquito, Aedes aegypti

As demonstrated with immunocytochemistry, specific cells and axons in the nervous system of female Aedes aegypti contain antigens immunologically related to FMRFamide (phenylalanine-methionine-arginine-phenylalanine-amide) and locust adipokinetic hormone I (AKH). In the supra-esophageal ganglion, including some medial neurosecretory cells, and in all ganglia of the ventral nerve cord, there are 100-120 cells immunoreactive to a FMRFamide antiserum. The same cells cross-react with a bovine pancreatic polypeptide antiserum, but when the latter antiserum is preabsorbed with FMRFamide, immunoreactivity is lost. However, immunoreactivity is maintained when FMRFamide antiserum is preabsorbed with pancreatic polypeptide, suggesting that the immunoreactive peptide is more closely related to FMRFamide. There are 6-12 cells in the supra- and subesophageal ganglia immunoreactive to an AKH antiserum, and some of the same cells are reactive to the FMRFamide antiserum. As well, unpaired cells in each of the abdominal ganglia are positive for both AKH and FMRFamide. Although the function of the FMRFamide- and AKH-like peptides in mosquitoes is unknown, this study, combined with previous reports on the localization of FMRFamide-like peptides in midgut endocrine cells, supports the concept of a brain-midgut neuroendocrine axis in this insect.     

Neuropeptides in insect sensory neurones: tachykinin-, FMRFamide- and allatotropin-related peptides in terminals of locust thoracic sensory afferents

Sensory afferents in the thoracic ganglia of the locust Locusta migratoria were labelled with antisera to different neuropeptides: locustatachykinins, FMRFamide and allatotropin. The locustatachykinin-immunoreactive (LTKIR) sensory fibres were derived from the legs and entered the ventral sensory neuropil of each of the thoracic ganglia via nerve 5. In the thoracic neuropil, the LTKIR sensory fibres formed a distinct plexus of terminations ventrally in the ipsilateral hemisphere. The peripheral cell bodies of the sensory neurones could not be revealed, but lesion experiments indicated that origin of the LTKIR fibres was the tarsus of each leg. Possibly the thin fibres are from tarsal chemoreceptors. Double labelling immunocytochemistry revealed that all the LTKIR sensory fibres contained colocalized FMRFamide immunoreactivity. A larger population of sensory fibres reacted with antiserum to moth (Manduca sexta) allatotropin. By means of double labelling immunocytochemistry, we could show that the LTKIR fibres constituted a subpopulation of the larger set of allatotropin-like immunoreactive fibres. Thus some sensory fibres may contain colocalized peptides related to locustatachykinins, FMRFamide-related peptide(s) and allatotropin-like peptide. A separate non-overlapping small set of sensory fibres in nerve 5 reacted with an antiserum to serotonin. Sensory fibres of the other nerves of the ventral nerve cord, including the abdominal ganglia, did not react with the peptide antisera. Since acetylcholine is the likely primary neurotransmitter of insect sensory fibres, it is possible that the peptides and serotonin are colocalized with this transmitter and serve modulatory functions in a subset of the leg afferents.     

Organization of the primary projections of the lateral line nerves in the lamprey Lampetra japonica

The lateral line sensory system of Lampetra japonica is innervated by the anterior and posterior lateral line nerves. The anterior lateral line nerve innervates all electroreceptors throughout the body and mechanoreceptors of the head. The posterior lateral line nerve innervates trunk mechanoreceptors. The anterior lateral line nerve consists of two ganglia (anterior lateral line and intracapsular) and four major peripheral branches (superficial ophthalmic, buccal, hyomandibular, and recurrent nerves). The posterior lateral line nerve has one posterior lateral line ganglion and one peripheral branch. The location and central projection patterns of the primary sensory neurons of these branches of the lateral line nerves were studied with the aid of horseradish peroxidase labeling. The ganglion cells of the buccal nerve were found in the rostral half, and those of the hyomandibular nerve were found in the caudal half of the medial part of the anterior lateral line ganglion. The lateral part of the anterior lateral line ganglion contains ganglion cells of the recurrent nerve and the superficial ophthalmic nerve. The rostral half of the intracapsular ganglion contains ganglion cells of the recurrent, hyomandibular, and buccal nerves. The ganglion cells of the posterior lateral line nerve were found in the posterior lateral line ganglion. The buccal nerve afferents terminated mainly in the lateral part of the ipsilateral mechanoreceptive medial nucleus. The peripheral part of the electroreceptive dorsal nucleus also received several afferents. The hyomandibular afferents terminated ipsilaterally in the central part of the medial nucleus and in the dorsolateral part of the dorsal nucleus. Some afferents of the hyomandibular nerve ascended and descended in the descending nucleus of the trigeminal nerve near its dorsal margin. The ventral nucleus, the primary nucleus of the VIIIth nerve, received a few fibers of the buccal and hyomandibular nerves. In the recurrent nerve, the fibers of the lateral part of the anterior lateral line ganglion terminated throughout the entire dorsal nucleus, and the fibers of the intracapsular ganglion projected to the dorsolateral part of the nucleus. The afferents of the posterior lateral line nerve terminated in the medial part of the ipsilateral medial nucleus and in the lateral part of the contralateral medial nucleus. In the cerebellar area, afferents of the anterior lateral line nerve were located laterally to those of the posterior lateral line nerve. Several fibers terminated in some branchiomotor nuclei, the cerebellar crest, and the dorsal gray near the obex level. No efferent cell bodies were found in the place where efferent neurons of the VIIIth nerve have been previously reported.     

Morphology, distribution and innervation of the lateral-line receptors of the Florida gar, Lepisosteus platyrhincus

The lateral-line system of gars consists of superficial neuromasts, which are arranged in lines termed pit lines, canal neuromasts and spiracular organs, which are located within diverticula of the hyoid gill pouch. Both canal and superficial neuromasts possess polarized hair cells whose directional sensitivity parallels the long axis of their respective lines. However, the apical surfaces of canal neuromasts are larger and possess far more hair cells than do those of superficial neuromasts, but superficial neuromasts have longer kinocilia and, presumably, longer cupulae. The receptors of the lateral-line system are innervated by three pairs of cranial nerves: anterior, middle and posterior lateral-line nerves. The anterior lateral-line nerves innervate neuromasts of the supraorbital, infraorbital and preoperculo-mandibular canals as well as dorsally located anterior pit lines, cheek (horizontal, vertical and mandibular) and gular pit lines of superficial neuromasts and the spiracular organ. The middle lateral-line nerves innervate dorsally located middle pit lines and a single neuromast in each temporal canal. The posterior lateral-line nerves innervate dorsally located posterior pit lines, neuromasts of the supratemporal commissures and all remaining postotic and trunk neuromasts. The ganglion of the anterior lateral-line nerve is divided into dorsal and ventral subganglia; the single ganglion of the middle lateral line nerve has no recognizable subdivisions, and the ganglion of the posterior lateral-line nerve consists of rostral and caudal subganglia. Analysis of the roots of these nerves and review of the embryonic origin of their ganglia as well as comparisons with cranial nerves in other anamniotes suggest that the anterior and posterior lateral-line nerves of gars may represent the fusion of four to five separate lateral-line nerves at some stage in vertebrate phylogeny. Thus, with the addition of the middle lateral-line nerve, and the possible existence of a ventral lateral-line nerve of the trunk, it is possible that the earliest jawed vertebrates possessed six or even seven pairs of lateral-line nerves.     

FMRFamide immunoreactivity in the brain and pituitary of Carassius auratus (Cyprinidae, Teleostei)

Neurons immunoreactive for the molluscan cardioactive peptide FMRFamide (FMRFi) in the brain of Carassius were demonstrated immunohistochemically with the peroxidase-antiperoxidase technique (PAP) and three antisera against this tetrapeptide. FMRFi perikarya were consistently found in the ganglion of the nervus terminalis, in some scattered neurons of the nucleus entopeduncularis, in the nucleus praeopticus pars magno- and parvocellularis, the nucleus ventromedialis, the nucleus posterior periventricularis, nucleus recessus lateralis and posterior. Some weak FMRFi neurons occurred in the rostral mesencephalic tegmentum and in the region of the nucleus gustatorius secundus. Both the medullary Mauthner neurons and the cerebellar Purkinje cells were only weakly immunoreactive for FMRFamide, while a group of intensely FMRFi rhombencephalic perikarya, presumably the nucleus motorius nervi vagi, occurred subependymally next to the fourth ventricle. FMRFi fibres and nerve endings occurred in the bulbus olfactorius, in the tractus opticus, and in the central parts of the dorsal, medial and ventral telencephalon. Some FMRFi fibres traversed through the commissura anterior while others occurred close to the hypothalamic nuclei, in the medial layers of the tectum opticum, in the brain stem, in the vagal lobe and in the ventral medulla oblongata. The present results are compared to the FMRFi structures in other fish brains.     

Octopamine immunoreactive cell populations in the locust thoracic-abdominal nervous system.

We describe octopamine-immunoreactive somata and their projections in the pro- meso-, meta- and pregenital abdominal-ganglia of locusts. Immunoreactive midline somata were identified as dorsal- and ventral- unpaired median (DUM- and VUM-, respectively) neurones due to their: characteristic large size and positions of somata, primary neurites in DUM-tracts giving rise to T-junctions, and bilaterally projecting axons. In the prothoracic ganglion there are most likely 8 such cells; in the meso- and metathoracic, some 20 each; and in each individual pregenital abdominal ganglion, typically 3. All appear to project to peripheral nerves and their numbers correspond to the number of peripherally projecting DUM-cells identified to date in each ganglion. We suggest that probably all peripherally projecting DUM-cells are octopaminergic in the examined ganglia. Presumptive DUM-interneurones are not octopamine-immunoreactive, but, confirming other studies, are shown to label with an antiserum to gamma-amino butyric acid (GABA). Other octopamine-immunoreactive neurones include a pair of midline, prothoracic, anterior medial cells, not necessarily DUM-cells, and a pair of ventral lateral somata in each thoracic- and the first abdominal ganglion. The latter project intersegmentally in ventral tracts. Intersegmentally projecting octopamine-immunoreactive fibers in dorsal tracts probably arise from a prothoracic DUM-cell, which leaves through suboesophageal nerves, or descending suboesophageal DUM-cells. Thus, the octopamine-immunoreactive system of thoracic and pregenital abdominal ganglia in locust comprises all peripherally projecting DUM-cells and a plurisegmental network.     

Mapping of serotonin-like immunoreactivity in the ventral nerve cord of crayfish

Whole-mount immunohistochemical technique using antibody to serotonin (5-HT) had been used to map presumptive serotoninergic structures in the ventral abdominal and thoracic nerve cord of crayfish Procambarus clarkii. 5-HT immunoreactivity was detected in more than 30 cell bodies, numerous fibers and peripheral nerve endings of root plexus and neuropilar regions. Immunoreactive fibers are arranged in 3 pairs of rostrocaudal bundles. The median (MFB) and the lateral fiber bundles run longitudinally through the entire thoracic and abdominal nerve cord (first thoracic T1 to sixth abdominal A6 ganglia). The central (CFB) fiber bundles extend only from the subesophageal to the fourth thoracic ganglia. In the 4 anterior thoracic ganglia (T1-T4), two lateroposterior cell bodies send their major processes in the ipsilateral MFB. In the fifth thoracic (T5) and first abdominal (A1) ganglia, the pattern of reactive structures is similar. Two large anterior cells which send a single prominent process to join the ipsilateral MFB and 4 smaller posterior cells. In other abdominal ventral ganglia, immunoreactive structures are smaller and less labeled. Cell bodies are displayed in two kinds of arrangement giving the appearance of two distinct homogeneous groups of ganglia: an anterior group (A2-A3-A4) that contained two pairs of small neurons and a posterior group (A5-A6) that contained only a large unpaired medial neuron. These results were discussed in relation to the serotonin-like immunoreactivity pattern previously described in lobster by Beltz and Krawitz.     

Coexistence of bovine pancreatic polypeptide-like immunoreactivity and catecholamine in neurons of the ventral aminergic pathway of the rat brain

The coexistence of bovine pancreatic polypeptide (BPP) within the cell bodies and axons of noradrenergic neurons of the rat brain was studied. Adjacent hindbrain sections stained by the indirect immunofluorescence technique for either BPP or dopamine-beta-hydroxylase (DBH) revealed A1 and A2 cell bodies containing both antigens. Following bilateral knife cuts of the ventral noradrenergic bundle (VB), the localization of BPP, DBH and catecholamine within the VB and terminal fields of the hypothalamus was determined. An axoplasmic buildup of BPP immunofluorescent material was observed caudal to the knifecut together in the same axonal fields as catecholamine and DBH-containing axons. In the hypothalamus there was a marked decrease in the number of catecholamine-containing nerve fibers. However, there was no obvious decrease in the number of BPP fibers. The influence of high intraventricular doses of 6-hydroxydopamine on BPP and DBH-containing nerves in the hypothalamus was studied. While there was a marked reduction in DBH immunoreactive fibers, BPP fibers appeared relatively unchanged. These results suggest that BPP coexists within some hindbrain catecholamine neurons and their axons. It is suggested that BPP and catecholamine coexist in terminal fields within the hypothalamus. Failure to reveal a decrease in the BPP-fibers in these fields suggests that sprouting of arcuate nucleus derived non-aminergic BPP-containing fibers has occurred.     

Serotonin-immunoreactive neurons and endogenous serotonin in the opisthosomal ventral nerve cord of the horseshoe crab,Limulus polyphemus

It has been suggested that serotonin serves as a neurotransmitter in the horseshoe crab, Limulus polyphemus. While some studies of identified groups of central neurons have been conducted, little is known concerning the neuronal organization in Limulus central ganglia. This study was undertaken to determine the localization of serotoninergic neurons in the opisthosomal ventral nerve cord of Limulus and to construct a basis for further comparative biochemical and pharmacological studies of the specific function of these neurons. Endogenous serotonin was, detected in the ventral nerve cord (chain of abdominal ganglia) by high performance liquid chromatography and electrochemical detection (HPLC-EC). Endogenous serotonin was quantified in the 9th through 13th ganglia, anterior (hemal) nerves, posterior (branchial) nerves, and connectives. The serotonin content in the abdominal ganglia was significantly reduced when the ganglia were incubated for 24 hours in Leibovitz's (L-15) medium containing reserpine or 5, 7-dihydroxytryptamine (5, 7-DHT), neurotoxins that block the uptake of serotonin into storage vesicles. The distribution of serotonin-immunoreactive neurons in the ventral nerve cord was determined by indirect immunocytochemistry. Treatment of the chain of ganglia with an anti-serotonin antiserum followed by treatment with a fluorescent-labeled antiserum raised against the primary antibody demonstrated specific staining in each, ganglion, the ganglionic roots, and connectives. Clusters of serotonin immunoreactive neurons were observed anteriolaterally and posteriorly in each ganglion. Processes from dense fiber bundles extended from these clusters of neurons to the central region of each ganglion. These results demonstrate that serotonin-immunoreactive neurons are present in the opisthosomal ventral nerve cord of the horseshoe crab and that serotonin may function as a neurotransmitter. © 1994 Wiley-Liss, Inc.     

Neuroanatomy of the visual afferents in the horseshoe crab (Limulus polyphemus)

The central nervous system of Limulus consists of a circumesophageal ring of fused ganglia and a paired ventral nerve cord. The anterior portion, the protocerebrum, receives sensory inputs including visual information. Three optic nerves, one each from the lateral eye, median ocellus, and ventral eye enter each side of the protocerebrum. The central connections of each optic nerve were determined by staining cut nerve trunks with cobalt chloride. The lateral optic nerve innervates the lamina, medulla, optic tract, ventral central body, and ocellar ganglion. The branching patterns of single axons, probably those of eccentric cells in the lateral eye retina, were observed. Single, large-diameter axons in the lateral optic nerve ramify at seven loci including sites in each of the structures innervated by the lateral optic nerve as a whole. The median optic nerve innervates the ocellar ganglion, central body, optic tract, and medulla. Three types of branching patterns were observed for single, large-diameter fibers in the median optic nerve. One type bypasses the ocellar ganglion and innervates the central body. A second type passes through the ocellar ganglion and optic tract without branching and innervates the posterior medulla. A third type innervates the ocellar ganglion, ventral central body, optic tract, and medulla. The ventral optic nerve is composed of large-diameter axons of ventral photoreceptors. Each axon enters the ganglion cell layer of the medulla and branches over a planar area less than 150 μm in diameter. We also observed that axons from mechanoreceptors on the anterior carapace innervate the posterior neuropil of the medulla, and that about 5% of the neurons in the medullar ganglion cell layer send axons to the ipsilateral circumesophageal connective.     

Co-localization of FLRF- and vasopressin-like immunoreactivity in a single pair of sexually dimorphic neurones in the nervous system of the locust

The distribution of Phe-Leu-Arg-Phe (FLRF)-like immunoreactivity is described in the brain and in the ganglia of the ventral nerve cord of the locust Schistocerca gregaria. A single homologous pair of immunoreactive cell bodies occurs ventrally and medially in the suboesophageal ganglion. Each cell sends a process dorsally which bifurcates into anteriorly and posteriorly running neurites. The single anterior neurite passes along the circumoesophageal connectives to the brain where it ascends in a posterior running tract, giving off branches to innervate the tritocerebral neuropile and ending in an extensive network of highly varicose immunoreactive processes in the protocerebral neuropile. No processes are seen in the optic lobes or associated with the structured neuropiles of the muschroom bodies. The single posterior neurite from each cell passes into the suboesophageal-prothoracic connectives. It runs in the lateral dorsal tract of each ganglion in the ventral nerve cord as a highly varicose process and in each ganglion gives rise to an ipsilateral network of varicose processes in the dorsal and lateral neuropiles. In the seventh and terminal abdominal ganglia the innervation pattern exhibits sexual dimorphism. Vasopressin-like immunoreactivity is co-localized in the same pair of suboesophageal neurones and their processes. A similar pair of ventral median neurones stains with both antibodies in the suboesophageal ganglion of another species of locust, Locusta migratoria. Although the basic distribution pattern of immunoreactive processes is similar in both species there are also marked species differences in the pattern.     

Development and distribution of serotonin in the central nervous system of Manduca sexta during embryogenesis. II. The ventral ganglia

The distribution of serotonin (5-HT) immunoreactive cells and their projections was mapped in the ganglia of the ventral nerve cord of the tobacco hornworm, Manduca sexta, during embryonic development, using an antiserum to 5-HT. Immunoreactive cells were first seen at 60% development. By 75% of embryonic development, a total of 94 immunoreactive cells were found in the ventral ganglia, including the suboesophageal ganglion. This number decreased to 80 neurons by 100% of development. About 50% of these cells were arranged in the abdominal ganglia and the rest were located in both the suboesophageal and thoracic ganglia. The suboesophageal ganglion exhibited immunoreactive segmental interneurons in areas corresponding to the mandibular, maxillary and labial neuromeres. Two pairs of immunoreactive interneurons were also observed to occur bilaterally in each of the thoracic and abdominal ganglia, with the exception of the prothoracic ganglion. This ganglion contained three pairs of bilaterally arranged immunoreactive neurons as early as 60% of embryonic development. Serotonin immunoreactivity was also found in a number of efferent neurons in the mandibular and labial neuromeres of the suboesophageal ganglion and in the prothoracic, mesothoracic and posterior abdominal ganglia. The occurrence of 5-HT in these efferent neurons suggests an involvement of serotonin in fore- and hindgut function via its effect on the visceral muscles. Immunoreactive lateral longitudinal fibers extended along the entire length of the ventral nerve cord together with dense segmental arborizations. The latter had regressed by the time the embryo was fully developed. This regression of the arborizations in the ganglia at the end of embryonic development indicates that a reorganization of 5-HT innervation occurs to support new larval functions. The time of appearance of 5-HT immunoreactive cells and fibers suggests that serotonin may play a role in the development of the ventral nerve cord.     

Bovine pancreatic polypeptide-like immunoreactive nerves in the rat major cerebral arteries

Immunofluorescence histochemical observation of the major cerebral arteries of the rat demonstrated the presence of an extensive plexus of bovine pancreatic polypeptide (BPP) containing nerves. The density of the nerves were similar to those previously observed in the adrenergic plexus. The density of BPP-immunoreactive fibers markedly decreased following bilateral superior cervical ganglionectomy. It is concluded that BPP-containing fibers of the cerebral arteries emanate mostly from the superior cervical ganglion and that BPP and norepinephrine coexist in the adrenergic nerves.     

The central connections of the anterior lateral line nerve of Gnathonemus petersii

The central connections of the lateral line nerve ganglia, the eithth nerve ganglia and the fused fifth-seventh ganglion of Ganathonemus petersii have been studied with silver degeneration techniques. The anterior (NLLa) and posterior (NLLp) lateral line nerves have a topographically organised projection upon the posterior lateral line lobe. NLLa, representing the head region, distributes to the rostral half of the posterior lobe, while NLLp, representing the trunk, distributes to the caudal half of the posterior lobe. The lateral line nerves also end in the anterior nucleus of the anterior lobe. There is some overlap within the middle third of the anterior nucleus, although NLLp tends to have a more caudal distribution than NLLa. N VIII terminates within n. tangentialis and n. octavius; there appears to be little or no overlap between VIII and lateral line nerve territories. The V–VIIth ganglion projects to entirely different parts of the brainstem. Terminal areas of V–VII are the sensory nucleus of the vagus, the nucleus of the descending trigeminus, and the funicular nuclei.     

Serotoninlike immunoreactivity in the central and peripheral nervous system of the scale worm Harmothoe imbricata (Polychaeta)

The distribution of serotoninergic neurons in the nervous system of the scale worm Harmothoe imbricata was visualized in the anterior half of the body by the peroxidase-antiperoxidase (PAP) immunohistochemical method with a specific antiserotonin antibody. Immunoreactive neuronal somata were localized in discrete ganglion cell masses of the dorsally situated cerebral ganglion and in segmental ganglia of the ventral nerve cord. They also make up the majority of neurons present in the parapodial ganglia. Large and small varicose fibers stained in the neuropile of all the above-mentioned ganglia but also in interganglionic connectives and segmental nerves. On the basis of soma size and location and of fiber distribution, the reactive neurons were identified as primarily interneuronal with a few motoneurons and presumptive afferent neurons. The presence of a motor component was substantiated by observations of several reactive varicose fibers spread over longitudinal muscle layers of the trunk. In addition, neurites of the subepidermal nerve plexus and enterochromaffinlike cells of the gut epithelium reacted with the serotonin antibody. It is concluded that serotoninergic pathways are ubiquitous elements in the organization of the central and peripheral nervous system of this polychaete. The significance of these findings in relation to other annelid groups and to the physiological role of serotonin is discussed.     

Retrograde tracing of nerve fibers to the rat middle cerebral artery with true blue: colocalization with different peptides

The origin of nerve fibers to the rat middle cerebral artery was studied by retrograde tracing with the fluorescent tracer True Blue (TB) in combination with immunocytochemistry to known perivascular peptides. Application of TB to the middle cerebral artery labeled nerve cell bodies in the ipsilateral superior cervical ganglion, the otic ganglion, the sphenopalatine ganglion, the trigeminal ganglion, and the cervical dorsal root ganglion at level C2. A few labeled nerve cell bodies were seen in contralateral ganglia. Judging from the number and intensity of the labeling, the superior cervical ganglion and the trigeminal ganglion and dorsal root ganglion at level C2 contributed most to the innervation. A moderate number of nerve cell bodies were labeled in the sphenopalatine and otic ganglia. The TB-labeled nerve cell bodies were further examined for the presence of neuropeptides. For that purpose antibodies raised against neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP), substance P (SP) and calcitonin gene-related peptide (CGRP) were used. A considerable portion of the TB-labeled nerve cell bodies in the superior cervical ganglion contained NPY. About half of the labeled nerve cell bodies in the sphenopalatine and otic ganglia contained VIP. In the trigeminal ganglion and in the dorsal root ganglion at level C2, one-third of the TB-labeled nerve cell bodies were CGRP-immunoreactive, while only few nerve cell bodies contained SP. The study provides direct evidence for the origin of cerebrovascular peptidergic nerve fibers and demonstrates that not only ipsilateral but also contralateral ganglia contribute to the innervation of the cerebral circulation.     

Calcitonin gene-related peptide-immunoreactive nerves in the tongue, epiglottis and pharynx of the rat: occurrence, distribution and origin

The occurrence, distribution and nature of calcitonin gene-related peptide (CGRP) in the tongue, epiglottis and pharynx of the rat was investigated by immunocytochemistry and radioimmunoassay. Numerous CGRP-containing nerves were found to innervate and terminate freely within the epithelium of the tongue, epiglottis and pharynx. Immunoreactive fibres were also found in the muscle layer and around blood vessels in the tongue, and in motor end plates in the muscle of the epiglottis and pharynx. Section of the trigeminal nerve induced a marked reduction in the number of immunoreactive nerves in the anterior portion of the tongue, whereas glossopharyngeal denervation results in a depletion of CGRP immunoreactivity in the posterior portion of the tongue. Immunoreactive nerves of the epiglottis and pharynx were depleted only after superior laryngeal nerve section. A subpopulation of labelled primary sensory neurones were observed in trigeminal and glossopharyngeal ganglia following injection of True blue retrograde tracer in the tongue. Most of the labelled cells were also immunoreactive for CGRP. Following systemic treatment with capsaicin, a loss of intra- and subepithelial CGRP-immunoreactive nerves was observed in all investigated tissues, while immunoreactive motor end plates remained unchanged.