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M Geffard

Publications and source records attributed to M Geffard.

At least 163 records · Page 9Linked to original sources
PubMed1987

Central GABAergic innervation of the pituitary in goldfish: a radioautographic and immunocytochemical study at the electron microscope level.

The GABAergic innervation of the goldfish pituitary was studied at the light and electron microscope levels by means of radioautography after in vitro incubation in tritiated gamma-aminobutyric acid (GABA) and immunocytochemistry using antibodies against GABA. Following incubation of pituitary fragments in a medium containing tritiated GABA, a selective uptake of the tracer was observed within the digitations of the neurohypophysis. Silver grain clusters were also observed in the adenohypophyseal tissue. At the electron microscope level, this uptake was found to correspond to nerve endings containing small clear and dense-core vesicles. These labeled profiles were located mainly in neurohypophyseal digitations in close apposition with the basement membrane separating the neurohypophysis from the adenohypophysis. However, they were also encountered in direct contact with most adenohypophyseal cell types in the different lobes. These results were confirmed by immunocytochemical data demonstrating the presence of numerous GABA immunoreactive fibers in both anterior and neurointermediate lobes. They were found either in the digitations of the neurohypophysis or in the adenohypophysis in direct contact with the glandular cells with a distribution and an ultrastructural aspect similar to those observed by radioautography. These data demonstrate that the pituitary of teleosts receives a massive GABAergic innervation. Although physiological data providing a functional significance for such an innervation are lacking, the present study suggests that, as already documented in mammals, GABA may be involved in the neuroendocrine regulation of pituitary functions in teleosts.

Animals↗
PubMed1987

Ultrastructural localization of GABA in the supraoptic nucleus and neural lobe.

Antibodies directed against the neurotransmitter gamma-aminobutyric acid (GABA) enabled the ultrastructural localization of GABA in conventional glutaraldehyde fixed and osmium postfixed material of the rat supraoptic nucleus and neural lobe. GABA was visualized using immunogold postembedding staining in axonal profiles that terminate on dendrites, axons or cell bodies throughout the supraoptic nucleus. The optimum ultrastructural preservation made possible the visualization of GABA terminals, also in the neural lobe. Here GABA axons were found to terminate synaptically on pituicytes and axonal profiles containing large dense core vesicles. These results emphasize, from an anatomical point of view, the potency of GABA to influence, as a transmitter, the release of vasopressin and oxytocin, both at the level of the cell body and of the neural lobe.

Animals↗
PubMed1987

Anatomical distribution and ultrastructural organization of the GABAergic system in the rat spinal cord. An immunocytochemical study using anti-GABA antibodies.

gamma-Aminobutyric acid (GABA)-containing elements have been studied by light and electron microscopy in the rat spinal cord, using immunocytochemistry with anti-GABA antibodies. Light microscopy showed immunoreactive somata localized principally in laminae I-III, and occasionally in the deeper laminae of the dorsal horn and in the ventral horn. Small somata were also observed around the central canal. Punctate GABA-immunoreactive profiles were particularly concentrated in laminae I-III, and moderately abundant in the deeper laminae and in the ventral horn where they were observed surrounding the unlabelled motoneurons. At the ultrastructural level, the punctate profiles corresponded to GABA-containing axonal varicosities or small dendrites. GABA-immunoreactive varicosities were presynaptic to labelled or unlabelled dendrites and cell bodies. Some unlabelled terminals presynaptic to unlabelled dendrites received symmetrical synaptic contacts from GABA-immunoreactive terminals. These results confirm data obtained with L-glutamate decarboxylase immunocytochemistry, and support the role of GABA in pre- and postsynaptic inhibition in the spinal cord, respectively via axoaxonal and axosomatic or axodendritic synapses.

Animals↗
PubMed1987

Immunocytochemical localization of dopamine in the prefrontal cortex of the rat at the light and electron microscopical level.

In the present study the dopaminergic innervation of the prefrontal cortex was studied by means of a recently developed anti-dopamine serum. This method can demonstrate endogenous dopamine in a specific way, and offers the opportunity to study the distribution of dopaminergic fibres in the cortex in detail in counterstained sections. Furthermore, dopaminergic nerve endings can be visualized at the electron microscopic level. Light microscopic observations demonstrated that the highest density of dopaminergic fibres in the frontal cortex is found in the prefrontal cortex and the infralimbic cortex. Within the prefrontal cortex, a good correlation is found between regional differences in distribution of dopaminergic fibres and the cytoarchitectonic parcellation of this part of the cortex. Outside the prefrontal cortex dopaminergic fibres were observed in adjacent frontal areas, the cortex surrounding the entire rhinal sulcus and the retrosplenial cortex. Electron microscopic observations demonstrated dopaminergic terminals through all cortical layers. The majority of dopaminergic terminals in the prefrontal cortex from synaptic contacts with dendritic processes. The synaptic profiles were usually symmetric and were characterized by the presence of many clear vesicles and an occasional dense-core vesicle.

Animals↗
PubMed1987

Monoclonal antibodies against glutaraldehyde-conjugated dopamine.

Four mice were immunized with dopamine (DA)-glutaraldehyde (G)--protein conjugates over a period of 8-10 weeks. Polyclonal antisera, obtained at various intervals, were tested using an enzyme-linked immunosorbent assay (ELISA). All had anti-conjugated DA antibodies. As soon as good antibody affinity was detected between 10(-10) and 10(-6) M, the mouse yielding the highest apparent affinity was killed, and the spleen was dissected out. Hybridomas were obtained from spleen cells fused with SP2/O/Ag myeloma cells. Supernatant culture media of hybridomas were tested for the presence of anti-conjugated DA antibodies with the ELISA method. Selected hybridomas giving good antibody affinity and specificity were then cloned by the limiting dilution technique. The resulting supernatant culture media were again tested by ELISA. Clones that gave a high antibody affinity (10(-10)-10(-8)M) for G-conjugated DA were used for histochemical localization of DA in rat brain. G-fixed rat brains were sectioned from the telencephalon to the mesencephalon, reduced with sodium borohydride, and prepared for peroxidase-antiperoxidase immunocytochemistry using supernatant (diluted 1:100) or ascites fluid (diluted 1:50,000). Dense networks of very fine fibers were observed in the striatum, septum, and cortex. Numerous immunoreactive cell bodies were found in the ventral tegmental area, the substantia nigra, the hypothalamus, and the dorsal raphe. The ELISA tests and adsorption controls suggested that the monoclonal antibody allowed highly specific detection of DA in tissues.

Aldehydes↗
PubMed1987

Specific antisera against the catecholamines: L-3,4-dihydroxyphenylalanine, dopamine, noradrenaline, and octopamine tested by an enzyme-linked immunosorbent assay.

Antisera were raised against L-3,4-dihydroxyphenylalanine (L-DOPA), dopamine (DA), noradrenaline (NA), and octopamine (OA). This was achieved by coupling each molecule to bovine serum albumin or human serum albumin using glutaraldehyde. The conjugated aromatic amines were kept in a reducing medium containing sodium metabisulfite. Antiserum specificity was tested using an enzyme-linked immunosorbent assay method for catecholamines. Competition experiments were done between the immunogen coated on the well plates and each catecholamine, either in the free state or in conjugated form, previously incubated with an antiserum. In each case, the nonconjugated compound was poorly recognized. The nonreduced conjugates of L-DOPA and DA were well recognized, whereas those of NA and OA were poorly immunoreactive. The cross-reactivity ratios established in the competition experiments allowed the specificity of the immune response to be defined. In each case, it was found to be high. The results suggest that the antibodies of L-DOPA and DA antisera recognize preferentially the catechol moiety, whereas for the anti-NA and anti-OA antibodies, the lateral chain is important.

Animals↗
PubMed1987

Anti-acetylcholine antibodies and the pathogenesis of myasthenia gravis.

Using an ELISA system, antibodies recognizing conjugated acetylcholine (ACh) were detected in sera of patients suffering from myasthenia gravis. The mean antibody level was three times higher in sera from myasthenic than from control patients. No correlation was found between anti-ACh antibody levels and anti-ACh receptor (AChR) titer. Also, the anti-ACh antibody titers were independent of sex and age of patients. Competition experiments demonstrated that the most immunoreactive compounds were choline-glutaryl-bovine serum albumin (BSA) and choline-succinyl-BSA. Antibodies present in the sera of myasthenic patients recognized an antigenic determinant mimicking conjugated ACh. The antibody affinity and specificity were sufficiently high for the detection of ACh in locust brain.

Acetylcholine↗
PubMed1987

An improved immunocytochemical method for subcellular localization of serotonin in rat enterochromaffin cells.

Serotonin-like immunoreactivity (5-HT-LI) has been localized at the ultrastructural level in enterochromaffin (EC) cells of rat gastrointestinal tract. Ultra-thin sections of tissues embedded in epoxy resin were incubated with 5-HT antisera and antibody binding sites were visualized with protein A-gold. Three different antisera were compared and were shown to require different fixation regimens for optimal preservation of 5-HT-LI. For one antiserum, tissues fixed in glutaraldehyde and osmium tetroxide could be used to demonstrate 5-HT-LI in EC cells. Immunocytochemical localization of 5-HT can thus be performed with good ultrastructural preservation of tissues. Quantitative evaluation of the intracellular distribution of 5-HT-LI was performed on EC cells from antrum, duodenum, and proximal colon, fixed in glutaraldehyde only. In all three locations, the majority of the gold particles (90%) in EC cells were localized over the dense core of the secretory granules, while a minor fraction (10%) were localized in parts of the cytoplasm devoid of granules. In EC cells fixed in glutaraldehyde and post-fixed in osmium tetroxide, 5-HT-LI was reduced by about 85%, although intracellular distribution was essentially the same as in cells fixed in glutaraldehyde alone. The results indicate that 5-HT in EC cells is stored mainly in secretory granules, with a small fraction of 5-HT being localized outside the granules.

Animals↗
PubMed1987

An ultrastructural study of GABA-immunoreactive neurons and terminals in the septum of the rat.

The fine structure and types of contact made by GABAergic elements in the septal nuclei were studied at the electronmicroscopic level by means of peroxidase immunocytochemistry, using anti-GABA antibodies. Observations were made on normal and colchicine-injected rats. GABA-immunoreactivity was distributed within somata, dendrites, axonal varicosities and terminals, and myelinated axons. The peroxidase reaction product was diffuse in the cytoplasm; cytoplasmic organelles were generally devoid of immunoreactivity, while showing a strong reaction on the outer surface of their membrane. GABA-immunoreactive (GABA-I) neurons were small (10 microns on average) to medium (20 microns) in size, with round or multipolar cell bodies. Additionally, labeled large (30 microns) cells were observed within the myelinated fibers of the medial septal nucleus after intraseptal administration of colchicine. No difference in the ultrastructural features and distribution of the immunoreactivity of the 2 kinds of cell was noticed, except for a higher number of synaptic contacts on large neurons of the medial septum. GABA-I cells of the medial and lateral nuclei received synapses on their soma and dendrites, made by both immunonegative and GABA-I terminals. Nonimmunoreactive boutons contacting GABA-I cell bodies were of 2 types: those containing small, clear synaptic vesicles and those that additionally contained large dense vesicles. Synaptic vesicles of GABA-I boutons were rarely labeled internally, but showed varying electron densities. Synapses made by GABA-I boutons on GABA-I or unlabeled somata and dendrites were always of symmetrical type. Synapses made by non-GABA-I boutons on GABA-I cells were either symmetrical or asymmetrical.

Animals↗
PubMed1987

Suppression of oscillatory activity in crustacean pyloric neurons: implication of GABAergic inputs.

Generation of rhythmic pyloric motor output in the crustacean stomatogastric ganglion results from synaptic connections and cellular properties of a 14-cell network of pyloric neurons. These cellular properties are under the influences of modulatory inputs, which act, for the most part, in an activating mode, i.e., they enhance the bursting properties of the pyloric neurons and/or their ability to express their regenerative properties. Here we attempt to demonstrate that the pyloric motor output is also under the control of suppressive afferent inputs that are able to stop the pyloric rhythm in a long-lasting manner. Immunohistochemistry, using GABA antibodies, indicates that GABAergic-like fibers are present in both the stomatogastric ganglion and its afferent nerve. Bath-applied GABA suppresses spontaneous pyloric rhythmic activity. This is due to an inability of the pyloric pacemakers to express their bursting properties. The suppressive effect of GABA is blocked by picrotoxin and mimicked by muscimol. Isolating the pyloric neurons from all descending spiking influences with tetrodotoxin demonstrates that exogenously applied GABA acts directly on the pyloric neurons. To confirm the existence of a physiological suppressive system for the pyloric motor pattern, we show that the stimulation of an afferent nerve, known to contain GABA-like fibers, also causes the cessation of rhythmic activity and the inability of the pyloric neurons to express their bursting properties.

Animals↗
PubMed1986

Membrane labeling of the cortex of 'aging' rats by anti-fatty acid antibodies.

The cortex of 'aging' rat brain exhibits no significant difference in the lipid and fatty acid composition in comparison with that of control rat brain, except for a lowered value of the cholesterol/phospholipids ratio. However, an antiserum raised against oleic acid, specifically labels neurones of the cortex of the 'aging' rat mainly within layers IV and V. An electron microscopical study revealed that immunoreactivity was associated with cytoplasmic vesicular inclusions (lipofuscin) and with membranes. Thus, these anti-fatty acid antibodies may help in the evaluation of local modifications of membranes which are not predictable on the basis of biochemical analysis of lipids.

Aging↗
PubMed1986

Immunological similarities between an experimental autoimmune myasthenia gravis model and human myasthenia gravis.

The induction of experimental autoimmune myasthenia gravis (EAMG) in rabbits after immunization with an acetylcholine (ACh) conjugate was found to possess immunological similarities with human myasthenia gravis. Anti-ACh antibodies, present in human sera, recognized the antigenic determinant, glutarylcholine, used to raise anti-ACh antibodies in rabbits. Identification of anti-anti-ACh antibodies in MG patients enabled us to test for recognition of the anti-ACh antibodies present in rabbit sera. The reverse, the recognition of rabbit auto-anti-anti-ACh antibodies by human anti-ACh antibodies was also tested and found to be specific.

Acetylcholine↗
PubMed1986

Glycine neurons in the brain and spinal cord. Antibody production and immunocytochemical localization.

Antibodies were raised against glycine and they were specific for immunocytochemistry. Obtained from rabbits immunized with glycine conjugated to glutaryled protein-carriers, antisera were then purified by adsorption on the various glutaraldehyde-conjugated protein-carriers. Using a modified ELISA method, their specificity was determined in competition experiments between conjugated glycine and either non-conjugated glycine or other conjugated amino acids or derivatives, preincubated with anti-glycine antibodies. Calculated at half-displacement, the resulting cross-reactivity ratios showed conjugated glycine to be the best recognized compound. By revealing the presence of the majority of the glycine-containing cell bodies in the brainstem and spinal cord, immunocytochemical applications of glycine antibodies confirmed their use as specific tools for a better understanding of the role of glycine in the central nervous system.

Animals↗
PubMed1986

One perfusion mixture for immunocytochemical detection of noradrenaline, dopamine, serotonin and acetylcholine in the same rat brain.

A perfusion mixture composed of glutaraldehyde and allyl-alcohol at pH 12 is compatible for the fixation and immunocytochemical detection of noradrenaline, dopamine, serotonin and acetylcholine in the same rodent central nervous system. A technical description of this procedure is provided to facilitate the replication of this immunocytochemical approach with these antibodies.

Acetylcholine↗
PubMed1986

Antibodies to histamine. Specificity studies and radioimmunological assay.

Antibodies against conjugated histamine were raised in rabbits. This amine was coupled to different protein carriers by a bifunctional agent, hexamethylene diisocyanate. The specificity of the antibodies was determined with radioimmunological tests in equilibrium dialysis using an iodinated ligand: 125I-labelled histamine-hexamethylene diisocyanate-glycyl-tyrosine. The latter mimicked the antigenic determinant present in immunogens. Competition experiments were established between the radiolabelled ligand and conjugated histamine, conjugated analogs or unconjugated histamine. Cross-reactivity ratios and affinity constants were calculated from displacement curves, thereby allowing the antibody site to be characterized. The antibodies were found to be highly specific and were used for the assay of histamine in biological samples. For this, polystyrene beads coated with purified antiserum were used to establish a simple and reproducible test.

Animals↗
PubMed1986

Reinnervation of the nucleus accumbens and frontal cortex of the rat by dopaminergic grafts and effects on hoarding behavior.

Embryonic dopaminergic neurons were implanted in the form of a cellular suspension in the nucleus accumbens previously deprived of its dopaminergic innervation by a local injection of 6-hydroxydopamine. The graft provided a dopaminergic reinnervation to the nucleus accumbens, the anteromedial striatum, the anteromedial frontal cortex and also, in some cases, of the septum. The pattern of reinnervation was specific for each structure and similar to the innervation provided by mesocorticolimbic dopaminergic neurons to these same structures in the normal animal. The graft restored the locomotor stimulatory action of amphetamine which was abolished in the lesioned controls. Hoarding behavior, which was disrupted following the lesion, was not reinstated by the graft alone. However, if the grafted neurons were stimulated by a small dose (0.2 mg/kg, i.p.) of (+)-amphetamine, hoarding reappeared in the grafted animals, while the same dose of amphetamine had no effect in the lesioned controls.

Animals↗
PubMed1986

Transplantation of dissociated foetal serotonin neurons into the transected spinal cord of adult rats.

Twenty adult rats underwent a complete section of the spinal cord at the lower thoracic level. One week later, 15 of them received a cell suspension obtained from raphe nuclei of 14-day-old foetuses into the distal fragment of the spinal cord. They were sacrificed after survival periods of 10-60 days, and vibratome sections of the spinal cord were processed for immunocytochemical detection of serotonin (5-HT). The control, non-transplanted animals showed a total absence of 5-HT immunoreactivity below the section, whereas the transplanted rats showed many immunoreactive 5-HT perikarya in the graft region, some at a distance of up to 10 mm, and a progressive innervation of the whole grey matter extending at least over 20 mm from the graft site.

Animals↗