Biogenic amines in the ponerine ant Harpegnathos saltator: serotonin and dopamine immunoreactivity in the brain

Many studies suggest a role for biogenic amines in a variety of insect behaviors including intraspecific aggression. In ants, despite a rich behavioral repertoire and prominent aggressive interactions, little is known about the potential impact of biogenic amines. This may partly be due to the general lack of information about aminergic systems in the ant brain. The present study investigates serotonergic and dopaminergic neuronal systems in the brain of the ponerine ant Harpegnathos saltator. In H. saltator, intraspecific aggression is important for the regulation of reproduction. This species, therefore, is amenable to comparative studies of aminergic neuronal effects on long-term changes in aggression. Using immunocytochemistry and confocal microscopy, we found that in the brains of sterile workers, the distributions of serotonergic and dopaminergic neuronal processes differed substantially. In addition, branching patterns of serotonergic neurons showed marked differences between males and females. Brains of workers after 3 days and 3 weeks of aggressive interactions revealed no marked differences in serotonergic and dopaminergic neurons compared to those of reproductive and non-aggressive individuals. We conclude that different levels of intraspecific aggression do not involve profound anatomical changes in serotonergic and dopaminergic neurons. Subtle changes may be masked by inter-individual variances.     

Catecholaminergic and serotoninergic brain structures in European green frogs: an autoradiographical study

Aminergic brain structures have been investigated by means of light microscopical autoradiography after injection of the tritiated catecholamines noradrenaline and dopamine and the indoleamine (or tryptamine) serotonin into the brain cavity of frogs of the Rana esculenta complex. These amines are fairly specifically taken up by catecholaminergic and serotoninergic neurons, respectively, which are located in structures like the catecholaminergic preoptic recess organ; the mixed catecholaminergic-serotoninergic paraventricular organ/nucleus infundibularis-complex and nucleus reticularis mesencephali; the telencephalic septal and striatal areas and the tectum opticum, which contain many catecholaminergic axon terminals; the habenular area, which contains serotoninergic axon terminals. The autoradiographical data on the location and the nature of these aminergic brain structures agree well with the mainly fluorescence microscopical and immunocytochemical data from the literature. The autoradiographical detection method can be combined at the light and the electron microscopical level with other histological, histochemical, or immunohistochemical techniques in one and the same preparation, and the results of the different treatments may eventually be made visible simultaneously.     

Specific cellular expression of monoamine oxidase B during early stages of quail embryogenesis

Monoamine oxidase-B (MAO-B) is one of two distinct molecular forms of MAO that in part regulate the cellular levels of biogenic amines. In order to determine whether discrete cell populations known to express aminergic properties in the vertebrate embryo also express MAO-B, the distribution of MAO-B-immunoreactive cells was examined in early developing quail embryos. Two major patterns of staining emerge. First, tissues known to express several aminergic characteristics are initially MAO-B positive at early stages of development and continue to express immunoreactivity through Zacchei stage 20, the oldest stage examined. These include cells of the sympathetic and some cranial and trunk sensory ganglia (beginning at stage 13), the pancreas (stage 14), and the brain stem raphe (stage 14). Second, other structures that contain or accumulate biogenic amines are transiently MAO-B immunopositive during early stages of development. These tissues include extraembryonic yolk sac and presumptive gut endoderm (with most intense staining between stages 8 and 13), the ventral trunk neural tube (stages 14 and 16), and the notochord (stages 8–10). MAO-B immunoreactivity disappears from these regions at different stages, and none are MAO-B positive by stage 19–20. Other structures without known aminergic properties during early development also stain; these include liver (stage 20), mesenchymal cells that surround the Wolffian duct and lung buds (stages 14 and 18), and endothelial cells surrounding the dorsal aorta (stages 14 and 20). In general, however, MAO-B appears to be distributed in embryonic tissues that can use this enzyme to regulate biogenic amine levels either transiently or during initial phenotypic expression of aminergic traits.     

The structure of the coeliac ganglion of a teleost fish Myoxocephalus scorpius

Summary In order to compare the structure of a teleost sympathetic ganglion with those of other vertebrates, light, fluorescence histochemical and electron microscopy were carried out on the coeliac ganglion of the scorpion fish, Myoxocephalus scorpius. In common with studies on other vertebrates, fluorescence histochemistry distinguished two cell types: a) principal neurones which exhibited low levels of specific catecholamine fluorescence and comprise the majority of neurones in the ganglia, and b) smaller intensely fluorescent cells, some of which had processes tens of micrometers long.With the electron microscope, the principal cells were seen to make axodendritic and axosomatic synapses with axons containing mainly 30 nm agranular vesicles at the synaptic site while in other vertebrates usually only one or other synaptic association is present.Both the somata and the processes of intensely fluorescent cells contain 300–600 nm diameter vesicles many of which have electron dense cores. These cells are also innervated by axons containing 30 nm agranular vesicles.     

Function of Catecholamine-containing Neurones in Mammalian Central Nervous System

SEVERAL chemical substances are involved in synaptic transmission in the mammalian central nervous system^1–3. The Falck-Hillarp technique⁴ has demonstrated noradrenaline, dopamine and 5-hydroxytryptamine within nerve cell bodies and terminals^5,6 and the belief that these amines act as neurohumours is strengthened by observations that nerve fibre activation leads to their release from the terminals^7,8. Histo-chemical evidence suggests that discrete systems of neurones can be identified by their content of particular amines and it seems possible that such neurohumorally homogeneous systems have a functional as well as a chemical identity. Before the anatomical distribution of amine-containing neurones had been described, Brodie and Shore⁹ proposed that noradrenaline functions as the central neurohumour of the sympathetic and 5-hydroxytryptamine of the parasympathetic system. This suggestion has not been supported by anatomical evidence; the amine-containing neurones form systems of small diameter fibres of very diffuse terminal distribution, which do not correspond to recognized ascending or descending pathways^5,6, although amine-containing neurones in invertebrates have been identified as sensory systems¹⁰.     

Glyoxylic acid fluorescence and ultrastructural studies of neurones in the coeliac-superior mesenteric ganglion of the aged rat

Summary Sympathetic post-ganglionic neurones in the coeliac-superior mesenteric ganglion (CSMG) complex of aged (24 month) rats have been studied by glyoxylic acid-induced fluorescence and electron microscopy. Comparisons have been made with the CSMG of young adult (4 month) rats. In the aged rats the noradrenaline fluorescence of the majority of neuronal perikarya was very low or absent and few intraganglionic fluorescent varicosities were seen. Lipofuscin pigment was very prominent at the nuclear pole region of neurones and also in dendrites and axonal processes. Ultrastructural studies revealed large accumulations of residual bodies at the nuclear poles and in axons and dendritic profiles. Within the perikarya many mitochondria were distorted or swollen, the rough endoplasmic reticulum was disarranged and much dilated as were Golgi cisternae. Primary lysosomes were encountered throughout the neurone perikaryon and its axonal or dendritic processes.In contrast to the young adult CSMG, no evidence for loading of transmitter storage vesicles with an identical dose level of 5-hydroxydopamine was detected in any part of the neurones of aged rats. This might reflect an impairment of the uptake mechanisms and/or storage of noradrenaline in aged sympathetic neurones and their axonal and dendritic processes.     

The Localization of Biogenic Amines in the Echinopluteus

The distribution of biogenic amines in the young pluteus of Psammechinus miliaris Gmelin was examined by means of the histochemical fluorescence method of Falck and Hillarp. Two different systems where monoamines occur were detected. Catecholamines were mainly found in a presumably impulse propagating neuron-like system, while 5-HT (serotonin) was predominantly localized in the endoderm wall. The results are discussed in relation to the ontogeny and phylogeny of sea urchins.     

Uptake of l-Dopa by cells in the taste buds of the vallate papilla of the mouse

Summary Several precursor substances and biogenic amines were admistered intraperitoneally to mice and were examined by the histochemical formaldehyde induced fluorescence method. It was found that after treatment with l-Dopa a number of cells inside the taste buds showed fluorescence.     

Histochemistry of nitric oxide synthase in the nervous system

Summary Nitric oxide synthase, which generates the physiological messenger molecule nitric oxide, and its associated NADPH diaphorase (NADPHd) activity are distributed throughout selective neuronal populations of the central and peripheral nervous system. Considerable evidence has been accumulated to indicate that NADPHd activity labels cells lacking neuronal nitric oxide synthase, i.e., the specificity of the reaction has to be considered for the reliable detection of the enzyme in neuronal but also non-neuronal tissue. In the present review, critical aspects of nitric oxide synthase visualization in neurones, using its NADPHd activity, are discussed. Furthermore, the organization of the central and peripheral nitric oxide synthase-containing neuronal systems is described. Nitric oxide synthase is present in local cortical and striatal neurones, hypothalamic magnocellular neurones, mesopontine cholinergic neurones, cerebellar interneurones, preganglionic sympathetic and parasympathetic neurones, neurones in parasympathetic autonomic and enteric ganglia and primary viscero-afferent neurones. Finally, injury-related alterations in nitric oxide synthase activity are briefly outlined. In this respect, the histochemistry of nitric oxide synthase may represent a valuable marker for neurochemical, if not structural, alterations observed in neural diseases, regeneration and transplantation.     

The occurrence of the cell type containing a specific monoamine in the taste bud of the rabbit's foliate papila.

The fluorescence histochemical examination on biogenic amines of the rabbit's foliate papilla revealed that a specific monoamine exhibiting an yellow fluorescence was present in a certain cell type of taste buds. The fluorescence had the emission maximum at 520 mmu and faded rapidly under the influence of the UV-irradiation. The green fluorescence of adrenergic nerve had the emission maximum at 480 mmu and was fairly stable upon the UV-irradiation. The yellow fluorescence disappeared completely following reserpine treatment, while it was markedly enhanced by nialamide treatment. From the observations, it is suggested that certain taste bud cells of the foliate papilla contain a biogenic monoamine, probably 5-hydroxytryptamine (serotonin).     

Fluorescence histochemical studies of the uptake of exogenous noradrenaline and dopamine by in vitro cultured cerebrocortex explants of the rat

Cerebrocortex of the neonatal rats were cultivated (--14 days). The cultures were studied living and with histological and fluorescence histochemical methods. A differentiation of neuronal cell- and fiber elements, oligodendro glial cells and astrocytes was found. The glyoxylic acid technique to estimate biogenic monoamines (Lindvall et al. 1974) was adapted up the cultivated explants. The normal cultures have only 24 h post cultivationem a specific fluorescence granularly in small concentration of the surface of the explant and in the explant self. Incubations with noradrenaline and dopamine demonstrated a various accumulation of the exogenous transmitters in the various parts of the cultivated explants. Uptake and releasing mechanisms in the cultivated material of the cerebrocortex were discussed with respect to the results of the sympathetic ganglia in vitro.     

Dispensable,Redundant, Complementary,and Cooperative Roles of Dopamine,Octopamine, and Serotonin in Drosophila melanogaster

To investigate the regulation of Drosophila melanogaster behavior by biogenic amines, we have exploited the broad requirement of the vesicular monoamine transporter (VMAT) for the vesicular storage and exocytotic release of all monoamine neurotransmitters. We used the Drosophila VMAT (dVMAT) null mutant to globally ablate exocytotic amine release and then restored DVMAT activity in either individual or multiple aminergic systems, using transgenic rescue techniques. We find that larval survival, larval locomotion, and female fertility rely predominantly on octopaminergic circuits with little apparent input from the vesicular release of serotonin or dopamine. In contrast, male courtship and fertility can be rescued by expressing DVMAT in octopaminergic or dopaminergic neurons, suggesting potentially redundant circuits. Rescue of major aspects of adult locomotion and startle behavior required octopamine, but a complementary role was observed for serotonin. Interestingly, adult circadian behavior could not be rescued by expression of DVMAT in a single subtype of aminergic neurons, but required at least two systems, suggesting the possibility of unexpected cooperative interactions. Further experiments using this model will help determine how multiple aminergic systems may contribute to the regulation of other behaviors. Our data also highlight potential differences between behaviors regulated by standard exocytotic release and those regulated by other mechanisms.     

Histochemical studies on the regeneration of aminergic nerves in rat cerebral artery after superior cervical ganglionectomy

The course of regeneration of aminergic nerves in rat cerebral arteries was studied by means of histochemical methods, after uni- or bilateral cervical sympathectomy. Degeneration of aminergic nerves started on day 1 and was complete between days 3 and 7 after surgery. Between weeks 4 and 6, regenerating nerves started to appear from the proximal internal carotid artery. Regenerated aminergic nerve fibres were generally unbeaded and intensity of fluorescence was weak. The circular nerves appeared earlier than the longitudinal ones. The number of regenerating nerves reached the maximum, between months 9 and 12, at about half the normal level. AChE activity of the cerebral arteries showed no significant changes at any stage.     

The effect of toluene inhalation exposure on catecholamine contents in rat sympathetic neurons

Adult male rats were exposed to toluene in short-term exposure by inhalation for 48 h (2000 ppm, continuously), and in long-term inhalation for 3 months (1000 ppm, 8 h daily). The formaldehyde-induced fluorescence (FIF) technique for histochemical demonstration of catecholamines (CA) was used to detect changes in the catecholamine stores. The concentration of CA in the sympathetic neurons of superior cervical ganglia and adrenal medulla was measured by the FIF technique combined with microfluorimetry. The short-term toluene exposure induced a statistically significant reduction of CA contents in sympathetic neurons. After long-term exposure, no change in the CA level could be demonstrated either in sympathetic ganglion or adrenal medulla. In electron microscopic studies no clear pathological changes were detected after toluene exposure.     

Ultrastructural cytochemistry of biogenic amines in nervous tissue: methodologic improvements.

A new fixation method has been developed for localizing biogenic amines in nervous tissue. The method is a modification of the chromaffin reaction in which all fixation steps are buffered with mixtures of sodium chromate and potassium dichromate. In this way the fixation and cytochemical reaction are carried out almost simultaneously. Using the rat vas deferens as a model tissue, it was found that the preservation of electron dense (chromaffin) cores in the vesicles of adrenergic nerve terminals depended on several factors: a short primary fixation using low concentrations of aldehydes, the presence of the chromate/dichromate buffer during all fixation steps and, finally, a long incubation period in a slightly acidic (pH 6.0) solution of this buffer before postfixation in osmium tetroxide. Using this method it was possible to identify not only small and large dense-cored vesicles as storage sites for amines but also a tubular reticulum (neuronal endoplasmic reticulum), the latter especially in nerve terminals of mesenteric arteries and iris. Biogenic amines were also visualized in sympathetic ganglion cells and in the central nervous system e.g., supraependymal nerve terminals, tissues that up to now proved the most difficult in terms of amine localization. In all the tissues examined the cytochemical reaction was highly selective and present in well preserved tissue, which is a significant advance over previously available techniques. It therefore offers new opportunities for further studies on the role of biogenic amines as neurotransmitters.     

The sphincter of the efferent filament artery in teleost gills: II. Sympathetic innervation

In addition to the cholinergic innervation described in the sphincter of the efferent filament arteries (Bailly and Dunel-Erb, ′86), an aminergic component has been demonstrated by specific techniques. The Falck fluorescence technique reveals a network of nerve fibers displaying a green fluorescence characteristic of catecholamines. At the ultrastructural level two types of fibers are present, one with clear vesicles and another with densecored vesicles. Axo-axonal synaptic relationships exist between the two types. Results of 5- and 6-OHDA (hydroxydopamine) treatments confirm the presence of an aminergic component. These observations support the notion of a dual innervation: cholinergic and adrenergic of, respectively, parasympathetic and sympathetic origin. The presence of presynaptic modulation is suggested. The aminergic component could inhibit or reduce the release of acetylcholine from cholinergic nerve endings. These results suggest that the sympathetic innervation modulates the vasoconstriction effect of the parasympathetic component.     

Effect of Denzimol, a new anticonvulsant drug, on rat cerebellum. Morphological and histochemical patterns of Purkinje cell layer

The effects of Denzimol, a new anticonvulsant drug, were analyzed in the Purkinje cell layer of rat cerebellum. No modifications in the general histochemical and ultrastructural patterns were induced by chronic administration of the drug. A slight increase in the storage of lipopigments in the cytoplasm of Purkinje neurons was found. No effects on secondary fluorescence (indicative of changes in biogenic amines) were found while an increase in specific SSADH activity, a degradative enzyme in GABA metabolism was present. Some observed histological and ultrastructural alterations (presence of "balloned" Purkinje neurons showing, in particular, enlarged cisternae of the endoplasmic reticulum) may be a stress effect probably due to the gastric tube insertion by which the drug was administered.     

Biogenic amines and the control of neuromuscular signaling in schistosomes

Biogenic amines are small cationic monoamines that function broadly as neurotransmitters and/or neuromodulators in every animal phylum. They include such ubiquitous substances as serotonin, dopamine and invertebrate-specific phenolamines (tyramine, octopamine), among others. Biogenic amines are important neuroactive agents in all the flatworms, including blood flukes of the genus Schistosoma, the etiological agents of human schistosomiasis. A large body of evidence spanning nearly five decades identifies biogenic amines as major modulators of neuromuscular function in schistosomes, controlling movement, attachment to the host and other fundamental behaviors. Recent advances in schistosome genomics have made it possible to dissect the molecular mechanisms responsible for these effects and to identify the proteins involved. These efforts have already provided important new information about the mode of action of amine transmitters in the parasite. Moreover, these advances are continuing, as the field moves into a post-genomics era, and new molecular tools for gene and protein analysis are becoming available. Here, we review the current status of this research and discuss future prospects. In particular, we focus our attention on the receptors that mediate biogenic amine activity, their structural characteristics, functional properties and "druggability" potential. One of the themes that will emerge from this discussion is that schistosomes have a rich diversity of aminergic receptors, many of which share little sequence homology with those of the human host, making them ideally suited for selective drug targeting. Strategies for the characterization of these important parasite proteins will be discussed.     

Schistosoma mansoni: a new parenchymal cell in cercariae

A new type of cell has been identified in cercariae of Schistosoma mansoni. The perikarya (cell bodies) of these cells were located in the body (midsegment), in an area oral to the acetabulum (ventral sucker). Cytoplasmic processes extending from the perikarya ramified throughout the parenchyma of the anterior organ (oral sucker), body, and tail segments by following the path of the nerve processes from the neuropile. The perikarya of these cells had heterochromatic nuclei and a predominance of particulate material and granules (240-360 nm) in their cytoplasm. Aggregates of granules (240-360 nm) and associated vesicles (34 nm) were scattered throughout the cytoplasmic processes of the cells and formed distinct varicosed areas. These processes often connected to the tegument in the midsegment (body) of the cercariae. The granules and associated vesicles reacted (became electron dense) with fixatives reported to be detectors of biogenic amines: The glutaraldehyde/osmium tetroxide fixation procedure rendered the granules electron dense while the glutaraldehyde/chromate/osmium tetroxide fixation procedure rendered the granules and the associated vesicles electron dense. The chromate solution of the latter procedure was responsible for the electron density of the associated vesicles. The morphology of these cells (their long ramifying cytoplasmic processes) and their reaction to chromium suggests that they are probably biogenic aminergic sensory cells.