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.     

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.     

Neurocalcin-immunoreactive neurons in the petrosal ganglion innervate the taste bud

The distribution and origin of neurocalcin-immunoreactive (NC-ir) nerve fibers in the taste bud and carotid body were examined by an immunofluorescence method. In the circumvallate papilla of the tongue, NC-ir nerve fibers made subepithelial nerve plexuses and occasionally penetrated the taste bud. However, the carotid body was devoid of ir nerve fibers. In the petrosal ganglion, 32% of neurons were immunoreactive for NC. Such neurons were mostly medium-sized to large, and scattered throughout the ganglion. In the superior cervical and intralingual ganglia, numerous ir varicose fibers surrounded postsynaptic neurons. However, NC-ir could not be detected in cell bodies of these neurons. The retrograde tracing method indicated that NC-ir petrosal neurons innervated taste buds in the circumvallate papilla. NC-ir neurons may have a gustatory function in the petrosal ganglion.     

Distribution of carbonic anhydrase activity in neurons of the rat

Certain neurons of dorsal root ganglia (DRG) and some fibers of the sciatic nerve contain histochemically demonstrable carbonic anhydrase activity. Since the distribution of this enzyme throughout the nervous system has not yet been evaluated systematically, we conducted a comprehensive histochemical survey focusing particularly on structures derived from the neural crest and nonneural crest ectoderm. In the peripheral nervous system, we observed carbonic anhydrase activity in some, but not all, neurons of dorsal root, trigeminal, celiac, and myenteric ganglia as well as in glial cells throughout the CNS. Some neurons of the nodose ganglion also showed carbonic anhydrase activity. In all first order sensory ganglia that were studied, the enzyme was found only in large (50 micron or above) and medium (20-50 micron) size neurons; in the case of spinal ganglia, the reactive neurons constituted approximately 30% of the total neuronal population. Of these reactive neurons, 56% were heavily stained and 44% were moderately stained. Several possible roles for neuronal carbonic anhydrase are considered.     

Distribution and origin of serotoninergic afferents to guinea pig cochlear nucleus

The distribution of serotoninergic fibers in the guinea pig cochlear nucleus was studied with serotonin immunohistochemistry. In addition, the origin of the serotoninergic fibers was determined by combining the retrograde transport of wheat germ agglutinin-apohorseradish peroxidase (gold conjugated) with serotonin immunohistochemistry. Immunoreactivity was present in varicose and nonvaricose fibers that were unevenly distributed throughout the cochlear nucleus. The fibers were most prominent in the superficial layers of the dorsal cochlear nucleus and the anterior spherical cell area of the anteroventral cochlear nucleus. Although less prominent, serotonin-positive fibers were also present in the remaining part of the anteroventral cochlear nucleus and the posteroventral cochlear nucleus. A few positive fibers were present in the auditory nerve root and the dorsal and intermediate acoustic stiae. Double-labeled cells were found throughout the rostral- caudal extent of the serotoninergic system from the caudal linear nucleus to the nucleus raphe pallidus. However, most were confined to the dorsal (52%) and median (18%) raphe nuclei. Some serotoninergic cell groups contained retrogradely labeled cells that were not serotonin immunoreactive, indicating nonauditory afferents to cochlear nucleus containing other neurotransmitter substances. Serotonin may tonically modulate auditory processing within the cochlear nucleus as well as influence certain ascending auditory pathways. Most of the serotonin in the cochlear nucleus comes from superior raphe nuclei that also project to basal ganglia motor systems and limbic strctures. Therefore, the effect of serotonin on the cochlear nucleus may be related to level of arousal or behavioral state. © 1995 Willy-Liss, Inc.     

On the occurrence of substance P-containing fibers in sympathetic ganglia: immunohistochemical evidence.

Several sympathetic ganglia of the guinea pig, cat and rat were studied with indirect immunofluorescence technique using antibodies to substance P (SP) and dopamine-beta-hydroxylase (DBH), the latter of which represents a marker for noradrenaline containing neurons. In all ganglia studied SP-like immunoreactivity could be observed in certain nerve fibers (SP-positive fibers). Their number, localization fluorescence intensity and 'fluorescence morphology' varied considerably between the different ganglia as well as between species. In the inferior mesenteric ganglion and the coeliac-superior mesenteric ganglion complex of all species, and in particular of the guinea pig, a dense plexus of varicose SP-positive fibers was observed around the mostly DBH-positive, principal ganglion cells. In contrast no such fibers were seen in relation to the DBH-positive SIF cells of the ganglia. No SP-positive ganglion cells were observed in any of the ganglia studied. Previous biochemical and immunohistochemical studies have indicated that peripheral SP-positive fibers may represent the branches of primary sensory neurons. The rich supply of varicose, SP-positive nerve terminals, often surrounding the noradrenergic ganglion cells in a basket-like manner, may indicate that these fibers may play a functional role within the ganglia. Instead of merely passing through the ganglion they may be involved in special types of reflex arches.     

Occurrence and distribution of substance P receptors in the cerebral blood vessels of the rat

The distribution of immunoreactivity to the receptor for substance P was examined in the cerebral blood vessels of the rat. Substance P immunoreactivity has been demonstrated in the nerve fibers of the cerebral blood vessels. Recently, the production of substance P receptor specific antibody has enabled the detection of localization of the substance P receptor in the central nervous system. In this study, we examined the existence of nerve fibers with substance P receptor immunoreactivity in the cerebral blood vessels and the cranial ganglia innervating the cerebral blood vessels. Sprague–Dawley rats were perfused with fixative and the pial arteries and the cranial ganglia known to innervate the cerebral blood vessels, i.e., trigeminal, sphenopalatine, internal carotid, otic and superior cervical ganglia, were dissected. All specimens were incubated with anti-substance P receptor IgG, then stained by the avidin–biotin–peroxidase complex method. Numerous nerve fibers with varicosities forming plexuses, with substance P receptor immunoreactivity were observed on the walls of the major extracerebral arteries forming the circle of Willis and its branches. Substance P receptor immunoreactivity was also detected in the endothelium of the cerebral arteries. Substance P receptor immunoreactivity was positive in many neurons of the sphenopalatine ganglion, otic ganglion, trigeminal ganglion, superior cervical ganglion and internal carotid ganglion. The present study demonstrated the existence of nerve fibers with substance P receptor immunoreactivity in the cerebral blood vessels and the cranial ganglia that innervate the cerebral blood vessels. These findings are important in understanding the responsiveness of the cerebral blood vessels to substance P.     

Distribution of modulatory inputs to the stomatogastric ganglion of the crab, Cancer borealis.

The pyloric and gastric mill neural networks in the crustacean stomatogastric ganglion receive modulatory inputs from more anteriorly located ganglia via the stomatogastric nerve. In this study we employed biocytin backfilling and immunostaining, as well as electron microscopy, to determine the origin of these inputs in the crab, Cancer borealis. Fiber counts from electron micrographs of sections through the stomatogastric nerve showed that this nerve contains 55-60 medium to large diameter fibers (1-13 microns). These fibers were individually wrapped by several layers of membrane, presumably glial in origin. There was also a single cluster of jointly wrapped, small diameter (< 1 micron) fibers that may originate from peripheral sensory somata. Biocytin backfills revealed that approximately two thirds of the individually wrapped fibers in this nerve originate from somata in the other three ganglia of the stomatogastric nervous system, including the paired commissural ganglia and the single oesophageal ganglion. There were approximately 20 biocytin-labeled somata in each commissural ganglion and 3 somata in the oesophageal ganglion. An additional ten somata were localized to the stomatogastric ganglion itself. This accounts for nearly all of the medium to large diameter fibers in the stomatogastric nerve. We used double-labeling with backfills and immunocytochemistry to determine that there are two proctolin-immunoreactive neurons and four FMRFamide-like immunoreactive neurons among the biocytin-labeled neurons in each commissural ganglion. Both peptides modulate neural network activity in the stomatogastric ganglion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Localization of GABA-like immunoreactivity in the central nervous system of Aplysia californica.

Gamma-aminobutyric acid (GABA) is present in the central nervous system of Aplysia californica (Gastropoda, Opisthobranchia) where its role as a neurotransmitter is supported by pharmacological, biochemical, and anatomical investigations. In this study, the distribution of GABA-immunoreactive (GABAi) neurons and fiber systems in Aplysia was examined by using wholemount immunohistochemistry and nerve backfill methods. GABAi neurons were located in the buccal, cerebral, and pedal ganglia. Major commissural fiber systems were present in each of these ganglia, whereas more limited fiber systems were observed in the ganglionic connectives. Some of the interganglionic fibers were found to originate from two unpaired GABAi neurons, one in the buccal ganglion and one in the right pedal ganglion, each of which exhibited bilateral projections. No GABAi fibers were found in the nerves that innervate peripheral sensory, motor, or visceral organs. Although GABAi cells were not observed in the pleural or abdominal ganglia, these ganglia did receive limited projections of GABAi fibers originating from neurons in the pedal ganglia. The distribution of GABAi neurons suggests that this transmitter system may be primarily involved in coordinating certain bilateral central pattern generator (CPG) systems related to feeding and locomotion. In addition, the presence of specific interganglionic GABAi projections also suggests a role in the regulation or coordination of circuits that produce components of complex behaviors.     

Dopamine in the lobster Homarus gammarus. I. Comparative analysis of dopamine and tyrosine hydroxylase immunoreactivites in the nervous system of the juvenile

As a catecholamine, depamine belogs to a class of molecules that have multiple transmitter and hhormonal functions in vertebrate and invertebrate nervous systems. However, in the lobster, where many central beurons have been identified and the peripheral innervation pattern is well known, the distribution of dopamine-containing neurons has not been examined in detail. Therefore, immunocytochemical methods were used to identify neurons likey to contain dopamine and tyrosine hydroxylase in the central nervous system of the juvenile lobster Homarus gammarus.Approximately 100 neuronal somata stain for the catecholamine and/or its synthetic enzyme in the brain and ventral nerve cord. The systems of neurons labeled with dopamine and tyrosine hydroxylase natibodies have the following characteristic: (1) the two systems are nearly identical; (2) every segmental ganglion contains at least one pair of labeled neurons; (3)the positions and numbers of cell bodies labeled with each antiserum are similar in the various segmental ganglia; (4) six labeled neurons are anatomically identified; two interneurons from the brain project within the ventral cord to reach the last abdominal ganglion, two neurons from the commissural ganglia are presumably neurosecretory neurons, and two anterior unpaired medial abdominal neurons project to the hindgut muscles; and (5) nocell bodies are labeled in the stomatogastric ganglion, but fibers and terminals in the neuropil are stained. The remarkably small numbers of labeled neurons and the presence of very large labeled somata with far-reaching projections are distinctive features consistent with other modulatory aminergic systems in both vertebrates and invertebrates. © 1994 Wiley-Liss, Inc.     

Immunoreactivity in Limulus. II. Studies of serotoninlike immunoreactivity, endogenous serotonin, and serotonin synthesis in the brain and lateral eye

Serotoninlike immunoreactivity was examined by the fluorescein-isothiocyanate-labeled secondary antibody technique in the lateral eye and brain of Limulus. Endogenous serotonin was measured with high-performance liquid chromatography and electrochemical detection. The synthesis of [3H]serotonin from [3H]tryptophan was measured in the presence and absence of reserpine. Fibers with serotoninlike immunoreactivity were found in the proximal stalks of the corpora pedunculata, in the neuropil of the central body, in the neuropils of the visual centers (lamina, medulla, and ocellar ganglion), in the optic tract that connects the ocellar ganglion with the posterior medial medulla, and in the central neuropil of the brain. Immunoreactive somata were found in four groups in the brain. Up to 50 somata were scattered through each side of the dorsal medial group that lies centered on the dorsal surface within the curve of the central body. These neurons innervate the central body neuropil and send processes into the central neuropil. Three or four reactive somata formed the ventral pole of each medullar group. These may provide the innervation of the proximal stalk of the corpora pedunculata. Five to ten reactive neurons were observed anteriorly in the ventral posterior lateral group #2 on each side that send processes into the central neuropil. Ten to 15 reactive somata were found on either side of the midline in the dorsal anterior part of the ventral medial group that contribute processes to the central neuropil. The remainder of the brain was not immunoreactive. No immunoreactive fibers or somata were found in the lateral eye or in the lateral optic nerve. Serotoninlike and substance P-like immunoreactivities were not found to be colocalized anywhere in the brain. Significant amounts of endogenous serotonin were detected in the lamina and medulla whose neuropils are rich in immunoreactive fibers and in the central body and dorsal medial group that are also rich in immunoreactive somata and fibers. No endogenous serotonin was detected in either the lateral eye or the lateral optic nerve. The lamina, medulla, and central body and dorsal medial group also synthesized and stored [3H]serotonin from [3H]tryptophan. It is likely that serotonin is a neurotransmitter in the brain, but not in the lateral eye of the horseshoe crab. In particular, it appears that serotoninergic neurons may play a role in central visual processing.     

Neurosecretory neurons and their projections to the serotonin neurohemal system of the cockroach Periplaneta americana (L.), and identification of mandibular and maxillary motor neurons associated with this system

The neuroanatomy of a serotonin neurohemal system in the head of Periplaneta americana was studied by means of immunohistochemistry, cobalt backfilling, transmission electron microscopy, and nerve transection. This neurohemal system is supplied by bilateral groups of two or three neurons whose somata are located ventrally in the subesophageal ganglion, near the root of each mandibular nerve. Axons of these serotoninergic neurons extend into all of the nerves of the mouth parts but reach most of these nerves by a very circuitous route. Initially the axons extend from the subesophageal ganglion, through the ipsilateral mandibular nerve trunk, and into the third branch of the mandibular nerve. From here the axons extend into the second branch of the maxillary nerve by way of a link nerve, and then they project retrogradely to reenter the subesophageal ganglion. In the ganglion, branches of these axons extend into the labial nerves, and the axons run dorsally through the subesophageal ganglion, circumesophgeal connectives, and tritocerebrum to reach the labral nerves. In the nerves of the mouth parts the serotoninergic axons give rise to numerous secondary branches that form an extensive neurohemal system at the surface of these nerves. The relatively large surface and cephalic location of this system probably indicate that the timely release of relatively large amounts of serotonin plays an important role in the physiology of feeding in this insect. The somata, neurites, and dendritic fields of the serotonin neurohemal neurons and those of the motor neurons of the mandibular abductor muscle occur together, and some of the mandibular abductor motor neurons also stain for serotonin. In order to distinguish clearly between these neurohemal and motor neurons, the anatomy of the mandibular abductor motor neurons has also been determined. Similarly, in the course of this study it has been necessary to work out the anatomy of the motor neurons of the maxillary retractor and cardo rotator muscles in order to distinguish them from the serotoninergic neurons. A nonserotoninergic peripheral neuron is associated with the serotonin neurohemal system, and its soma is located on the mandibular-maxillary link nerve. This link nerve neuron appears to be neurosecretory.     

PACAP-containing intrapineal nerve fibers originate predominantly in the trigeminal ganglion: a combined retrograde tracing- and immunohistochemical study of the rat

Pituitary adenylate-cyclase activating polypeptide (PACAP) is a neuropeptide originally isolated from the hypothalamus and located in many neuronal systems in both the central and peripheral nervous system. PACAP is also found in nerve fibers innervating the pineal gland, where it stimulates the secretion of the pineal hormone, melatonin, by binding to specific PACAP-receptors located on the cell membrane of the pinealocyte. In this study we have investigated the origin of PACAP-containing nerve fibers innervating the rat pineal gland by combined retrograde tracing with Fluorogold and immunohistochemistry for PACAP. A solution of 2% Fluorogold was injected iontophoretically into the superficial pineal gland of Wistar rats, and the animals were allowed to survive for 1 week. After perfusion fixation of the rats, the location of the tracer was investigated in the brain and the sphenopalatine, otic, and trigeminal ganglia. The tracer was found in all the investigated ganglia. However, colocalization with PACAP was predominantly found in the trigeminal ganglion and only occasionally in the sphenopalatine and otic ganglia. Due to the stimulatory function of PACAP on pineal melatonin secretion, the PACAP-containing neurons of this ganglion could be considered a subset of the parasympathetic nervous system. The presence of neurons with a parasympathetic function in a ganglion that has been considered a purely sensory ganglion, is a new concept in neuroanatomy.     

Development of segmental differences in the pressure mechanosensory neurons of the leech Haementeria ghilianii

Using a monoclonal antibody specific for the pressure mechanosensory neurons (P cells) of the leech Haementeria ghilianii, we have examined the segmental differences between P cells in the adult nerve cord, as well as the development of these differences during embryogenesis. The standard segmental ganglion contains two pairs of P cells of about the same size and staining intensity. The sex ganglia appear to be missing the P cells that normally innervate ventral skin, and ganglia 20 and 21 have much smaller ventral P cells than most segments. The pattern of P cells in the head and tail ganglia also differs slightly from that of the standard ganglia. During embryogenesis, when the neurons are first stained by the antibody, there are two pairs of P cells of equal size in each segmental ganglion. Obvious segmental differences arise subsequently, modifying an initially identical set of cells.     

Monoamine-containing neurons in the aplysia brain

The localization of monoamine-containing neurons in the CNS of Aplysia depilans has been studied by fluorescent histochemistry (the glyoxylic acid condensation method) and microspectrofluorimetry. Yellow fluorescent nerve cells and fibers show the emission maximum at 515–520 nm which corresponds to that of serotonin fluorophore in a model system. Green fluorescent nerve cells have the emission maximum at 485 nm which corresponds to that of catecholamine. Central catecholamine-containing neurons were found in cerebral, buccal, pedal and unpaired abdominal ganglia. The majority of them were revealed in cerebral ganglia (about 40). Serotonin-containing neurons are abundant in cerebral and pedal ganglia. More than 30 serotonin-containing nerve cells were localized in cerebral ganglia. In the right pedal ganglion approximately 100 neurons were revealed; in the left one about 150. In the abdominal ganglion all nerve cells of this chemical type (except one) are located in the right hemiganglion. The results are summarized in corresponding schemes.     

Choline acetyltransferase immunoreactive sympathetic ganglion cells in a teleost,Stephanolepis cirrhifer

The present study showed neurons immunoreactive for choline acetyltransferase (ChAT) in the cranial sympathetic ganglia lying close to the trigeminal-facial nerve complex of the filefish. In these ganglia, less than 1% of ganglion cells were positive for choline acetyltransferase. Choline acetyltransferase-positive neurons were significantly larger than the randomly sampled neurons in this ganglion. The majority of choline acetyltransferase-positive neurons were negative for tyrosine hydroxylase, but many of them were positive for galanin (GAL). Some neurons were positive for both choline acetyltransferase and tyrosine hydroxylase, but these neurons were rarely immunoreactive for dopamine beta hydroxylase, suggesting that they are not adrenergic. In the cranial sympathetic ganglia and the celiac ganglia, many nerve fibers immunoreactive for galanin were seen, and varicose terminals were in contact selectively with neurons negative for both choline acetyltransferase and tyrosine hydroxylase, but not with those positive for choline acetyltransferase or tyrosine hydroxylase. Nerve fibers immunoreactive for choline acetyltransferase were found to be present in contact with the deep layer of chromatophores, which was observed only in the labial region. These results suggest that cholinergic postganglionic neurons are present in the filefish cranial sympathetic ganglia, and that they also contain galanin. As few cholinergic sympathetic neurons express tyrosine hydroxylase and none express dopamine beta hydroxylase, they are unlikely to synthesize noradrenaline or adrenaline.     

Distribution and partial characterization of FMRFamide-like peptides in the stomatogastric nervous systems of the rock crab, Cancer borealis, and the spiny lobster, Panulirus interruptus

The distribution of FMRFamide-like peptides was studied in the complete stomatogastric nervous system [the paired commissural ganglia, single oesophageal ganglion, and the single stomatogastric ganglion (STG)] of two decapod crustacean species, the spiny lobster Panulirus interruptus and the rock crab Cancer borealis, by using immunocytochemical techniques. Antiserum 231 from the O'Donohue laboratory and antiserum 671C (described here) gave essentially the same staining patterns. In the commissural ganglia of both species there were ten to 20 stained neurons and dense neuropilar staining. The oesophageal ganglion of the crab had four stained neurons. Lucifer Yellow backfills followed by immunostaining showed that the two larger stained neurons of the oesophageal ganglion sent processes into the inferior ventricular nerve. The two smaller neurons sent processes into the inferior oesophageal nerves. The oesophageal ganglion of the lobster had two stained neurons that sent processes into the inferior ventricular nerve as well. None of the somata of the STG stained in either species, but in both species stained fibers were seen in the stomatogastric nerve that entered the STGs and ramified profusely throughout the neuropil. In some preparations of the crab, a stained fiber was visible in the dorsal ventricular nerve. The amounts of the FMRFamide-like peptides found in all regions of the nervous system of P. interruptus and C. borealis were determined by radioimmune assay (RIA). Column chromatography and high-performance liquid chromatography suggest that, in both species, much if not all of the RIA-assayable material is accountable for by peptides that are larger and more hydrophobic than FMRFamide.     

Neurotransmitters in the nervous system of Macoma balthica (Bivalvia)

The distribution of histamine-, octopamine-, gamma-aminobutyric acid- (GABA) and taurine-like immunoreactivity in the bivalve mollusc Macoma balthica was studied immunocytochemically with antisera produced in rabbits. Histamine levels in the ganglia and whole animals were also measured by high-performance liquid chromatography using a postcolumn derivatization method. Immunoreactivity for these substances, except for taurine, is found in the central nervous system of this species. The most extensive neuronal system is revealed with the antiserum against histamine. All the main ganglia contain histamine-immunoreactive cell bodies, and a dense network of nerve fibers is seen in the ganglia and nerve roots. Histamine-immunoreactive nerve fibers project to the mantle edge, lips and oesophagus. The basal part of the inhalant siphon is rich in histamine-immunoreactive fibers. Unlike histamine, octopamine- and GABA-like immunoreactivities are restricted to the central nervous system. Taurine-like immunoreactivity is not found in the nervous system of this species. In the nervous system, histamine-immunoreactive cell bodies and fibers are more numerous than those that are octopamine- and GABA-immunoreactive. The distribution of these substance in the ganglia is different. GABA-immunoreactive cells are typically smaller than most of the histamine- and octopamine-immunoreactive cells. Most GABA- and octopamine-immunoreactive cells and fibers are located in the pedal ganglion. Histamine is distributed more evenly in the ganglia and nerve roots. The biochemical measurements of histamine correlate well with the immunohistochemical findings and confirm the predominant location of the amine in the nervous tissue. These results suggest that histamine is more widespread than some other putative transmitters, and support the concept that histamine may have an important role in many physiological processes in molluscs. © 1993 Wiley-Liss, Inc.     

Distribution of immunoreactive somatostatin (ISRIF) in the nervous system of the squid, Loligo pealei

Somatostatin (SRIF) is a neuropeptide with a widespread distribution in the mammalian CNS. In the present study we have examined the distribution of immunoreactive-like SRIF (ISRIF)-containing elements in the nervous system of the cephalopod mollusk Loligo pealei, or the Woods Hole squid. ISRIF was localized by light immunocytochemistry in sections of the squid-optic lobe, circumesophageal ganglia-and in stellate ganglion. In the optic lobe, ISRIF neurons were found in the internal granule cell layer and medulla and immunoreactive fibers were seen throughout the lobe and in the optic tract but were absent from the optic nerve, i.e., the projection between the retina and optic lobe. In the supraesophageal complex, ISRIF neurons were found in all lobes, but primarily in the vertical, subvertical, and frontal. In the subesophageal ganglion, ISRIF neurons were seen mainly following unilateral pallial nerve lesions; these neurons were primarily small-to-medium sized. ISRIF fibers were seen in many of the nerves exiting from the brain and in nerves extending between the sub- and supra-esophageal ganglia. In the stellate ganglion, ISRIF was present in many neurons as well as in a plexus of fibers within the ganglion; the peptide was absent from the second-order fibers and the giant axon. The data suggest that a molecule immunologically similar to vertebrate SRIF may be a major transmitter/modulator in this invertebrate. These results provide a foundation for further studies to evaluate the role of this molecule.     

Tachykinin-related neuropeptides in the central nervous system of the snail Helix pomatia: an immunocytochemical study

The distribution of neurons reacting with an antibody raised against an insect neuropeptide, locustatachykinin I, was investigated in the CNS of the snail Helix pomatia. The localization of the neurons was compared with that of the substance P-like immunoreactive (SPLI) neurons in the different ganglia. Altogether, there are 800–1000 locustatachykinin-like immunoreactive (LomTKLI) neurons in the Helix CNS, occurring with an overwhelming dominancy (83.5%) in the cerebral ganglia. Within the cerebral ganglia, the majority of LomTKLI neurons were localized in the procerebrum. The number of SPLI neurons was high; 2000 SPLI nerve cells were found in the Helix CNS. The majority (44.5%) of SPLI neurons was also found in the cerebral ganglia and they were also concentrated in the procerebrum. The neuropils of all ganglia were densely innervated by both LomTKLI and SPLI fibers except the medullary mass of the procerebrum where only SPLI elements form an extremely dense innervation. In addition to the neuropil processes, LomTKLI neurons sent axon processes to the peripheral nerves. SPLI fibers also formed a dense network of varicose fibers in the connective tissue sheath around the ganglia where they innervated the blood vessel walls too. Immunolabeling on alternating cryostat sections revealed that LomTKLI and SPLI neurons are localized near each other in most cases; co-localization of the two immunoreactive materials could be seen in a very small number of neurons of the pedal and pleural ganglia. The present results show that the Helix CNS possesses distinct neuronal populations using different tachykinin-related peptides. It is suggested that the differential distribution of these neuropeptides also implies a diversity in their central and peripheral functions.