Biomedical subjects

M Geffard

Publications and source records attributed to M Geffard.

At least 127 records · Page 7Linked to original sources

PubMed1989

Central dopamine-synthesis regulation by the calcium-calmodulin-dependent system.

The effects of the intraventricular (IVT) administration of calcium on the amount of dopamine (DA) in various regions of the mouse brain were analyzed immunohistochemically by using a microphotometry system. The DA levels in the nucleus accumbens and the lateral part of the neostriatum were increased by approximately 45% (p less than 0.01) and 25-35% (p less than 0.01), respectively, by the IVT administration of CaCl2 (10 mumol/kg). It was also found that this effect was abolished by the calmodulin antagonist, W-7 (4.2 micrograms/mouse, IVT). The brain regions in which the amount of DA was increased by calcium were areas where high levels of calmodulin and tyrosine hydroxylase are distributed. These findings suggest that the synthesis of central DA is regulated by calcium through a calmodulin-dependent system.

Animals↗

PubMed1989

Intraspinal transplants of serotonergic neurons in the adult rat.

Adult male Sprague-Dawley rats were made paraplegic by a complete transection of the spinal cord at lower thoracic level. One week later they were transplanted, below the level of the section, with a cell suspension prepared from the raphe region of 14-day embryos. After survival periods of 10 days to 1 year, the animals were sacrificed and the spinal cord processed for the immunocytochemical detection of 5-HT. Axons from grafted cells grew extensively into the grey matter of the host, and established axosomatic and axodendritic synapses in the anterior horn and intermediolateral column, similar to those of the intact animal. In addition, a group of transplanted animals was tested for sexual reflexes which are under the control of serotonin. It was found that ejaculation reflexes, which are absent in paraplegic rats, are restored in transplanted animals.

Animals↗

PubMed1989

Morphological and electrophysiological studies of human hippocampal transplants in the anterior eye chamber of athymic nude rats.

Human fetal hippocampal tissue from normal women was obtained following elective abortion in the 8th to the 11th week of gestation. The hippocampal tissue was transplanted to the anterior chamber of the eye of adult athymic nude rats, where it was allowed to develop for up to 9 months before histological and electrophysiological evaluation. The transplants were revascularized from the host iris and many grew extensively in oculo. Large neurons were present in all transplants. Immunohistochemical studies revealed glutamic acid decarboxylase-containing terminals and clusters of gamma-aminobutyric acid-positive nerve cell bodies within the transplants, as well as scattered tyrosine hydroxylase-positive and acetylcholinesterase-containing fibers. Single neurons recorded extracellularly from transplants 4-9 months in oculo showed a slow spontaneous discharge, with both complex and single action potentials. Stimulation of the transplant surface evoked a small initial wave followed by a larger and longer-lasting field potential, similar to that seen in hippocampus in situ. A conditioning-testing paradigm was used to evaluate the presence of inhibitory circuitry in the hippocampal transplants. Significant suppression of the evoked test response was seen with interstimulus intervals ranging from 20 to 500 ms. Superfusion of enkephalin (100-300 nM) or penicillin (1600 U/ml) increased slow-wave activity, as did tetanic electrical stimulation. These treatments appeared to generate ictal-like activity, which in some cases persisted as interictal spikes. Illumination of the retina also increased neuronal activity, presumably by reflex activation of cholinergic afferents from the parasympathetic innervation of the iris. Taken together, our data suggest that fragments of hippocampus from aborted first trimester human fetuses, grafted to the eye chamber of rodent hosts, develop many organotypic histological and physiological features. This preparation may provide a unique means for the study of neurobiological properties of human brain in both normal and disease states.

Animals↗

PubMed1989

Dopamine synaptic complex with pyramidal neurons in primate cerebral cortex.

Dopamine (DA)-containing projections to the cerebral cortex are considered to play an important role in cognitive processes. Using a recently developed monoclonal antiserum directed against DA and an antibody directed against tyrosine hydroxylase in combination with Golgi impregnation and electron microscopy, we have observed that DA and tyrosine hydroxylase afferents establish symmetric membrane specializations with the soma, dendritic shafts, and spines of identified pyramidal cells in the prefrontal, cingulate, and motor cortex of primates. The axospinous contacts invariably formed part of a synaptic complex in which the dendritic spine of a pyramidal neuron was the target of both a DA-positive symmetric and an unlabeled asymmetric bouton. This arrangement allows direct DA modulation of the overall excitability of cortical projection neurons by altering local spine responses to excitatory inputs.

Animals↗

PubMed1989

Monoclonal anti-conjugated acetylcholine antibody and immunohistochemical applications in rat nervous system.

Acetylcholine (ACh) conjugates were injected into AKR and DBA mice over a period of 10 weeks. The polyclonal antisera were tested at various immunization times for affinity and specificity using an enzyme-linked immunosorbent assay (ELISA). The most immunoreactive compound was found to be choline-glutaryl-bovine serum albumin (or conjugated ACh). The AKR and DBA mice yielding the highest apparent affinity were killed, and the spleen cells were fused with X63 or SP2/O/Ag mouse myeloma cells. Supernatants of confluent cultures were tested for the presence of anti-conjugated ACh antibodies using the same ELISA method. The best results were obtained with the hybridomas from AKR spleen cells and X63 mouse myeloma cells. Monoclonal antibody affinity and specificity were then evaluated by a radioimmunological procedure using iodinated monoclonal anti-conjugated ACh antibody. From competition experiments, the most immunoreactive compound was choline-glutaryl-protein. The other related compounds were recognized either poorly or not at all. The high affinity and specificity of our monoclonal antibody enabled us to visualize ACh molecules on fixed rat brain sections. ACh was fixed with a mixture of nitrobenzyl alcohol and glutaraldehyde. Many ACh-immunoreactive cell bodies and fibers were seen on sections from the basal forebrain and spinal cord. Preadsorption and other immunohistochemical tests demonstrated that the ACh staining was highly specific.

Acetylcholine↗

PubMed1989

Acute action of DSP-4 on central norepinephrine axons: biochemical and immunohistochemical evidence for differential effects.

Previous immunohistochemical studies of the long-term effects of the noradrenergic neurotoxin DSP-4 have demonstrated a remarkably selective vulnerability of norepinephrine (NE) axons of the locus coeruleus (LC). NE axons originating in non-LC NE neurons appear to be largely resistant to the neurotoxic action of DSP-4. We conducted this study to evaluate the acute effects of DSP-4 on NE axons in four different brain regions: cerebral cortex, cerebellum, ventral forebrain, and hypothalamus. NE levels were determined by high-performance liquid chromatography (HPLC) 6 and 24 hr and 14 days after DSP-4 administration. NE axons in these brain regions were visualized in brain sections at 6 and 24 hr after drug treatment, using a specific antiserum to NE. HPLC assays revealed profound reductions of NE levels in cerebral cortex and cerebellum, but only minor decreases in ventral forebrain and hypothalamus. NE immunohistochemistry showed dramatic differences in the acute effects of DSP-4 on NE axon staining: nearly complete loss of staining in cortex and cerebellum, in contrast to an almost unchanged staining pattern in ventral forebrain and hypothalamus. This study demonstrates that NE immunohistochemistry is a valuable tool to assess the acute effects of DSP-4 on NE axons in different brain regions. The results provide the first direct evidence that NE axons are not uniformly acted on by DSP-4 and suggest that the acute effects of DSP-4 are restricted to LC axons.

Animals↗

PubMed1989

[Cells presenting GABA immunoreactivity in the hypothalamus of the cat].

The distribution of GABA-immunoreactive (IR) cells was studied by immunohistochemistry in conjunction with highly specific antiserum GABA in the cat hypothalamus. Colchicine pretreatment made it possible to visualize a large number of labeled cells in the medial preoptic and dorsal hypothalamic areas. In contrast, the ventromedial and anterior hypothalamic nuclei contained only a few labeled cells, and the paraventricular and supraoptic nuclei were devoid of them. A very dense network of GABA-IR presumptive terminals was seen in the ventrolateral posterior hypothalamus where labeled cells could be recognized. The possibility of an involvement of the GABAergic neuronal system in the regulation of sleep-waking cycle is discussed.

Animals↗

PubMed1989

[Autoantibodies directed against membrane phospholipid in serum in patients with malignant tumors].

Modifications of membrane lipids in the levels of fatty acids and phospholipids are associated with malignant tumors. In order to evaluate if these changes are recognized by the immune system, we have attempted to assay the possible presence of autoantibodies directed against the following lipids: phosphatidylinositol (PI), phosphatidylserine, phosphatidylcholine, phosphatidylethanolamine, gangliosides, galactocerebrosides, sphingomyelin, sphingosin, and cardiolipin, in the sera of patients with malignant tumors (n = 324) and from controls [healthy subjects (n = 20) and patients suffering of other diseases (n = 60)]. Using an adaptated immunoenzymatic assay (ELISA method), a highly significant difference (p less than 0.001) was found between the mean absorbances read on the cancer and control group for only one lipid, the PI. Whatsmore, these auto anti-PI were found in all sera (diluted 15,000 times) of patients with malignant tumors, whatever type, grade, or organ localization defined. These data indicated that the immune system recognized the PI antigenic modifications which appear to be linked to the cell transformation. This PI-immunological binding may have a predictive value as we have recently noticed in animals bearing chemically-induced malignant tumors.

Autoantibodies↗

PubMed1989

Critical review on quantitative autoradiography of D1 and D2 dopaminergic receptors in the striatum of the mammalian brain: differential localization and plastic changes after pharmacological manipulation and dopaminergic input disruption.

Major technical progress in the development of computer-based image analysis systems has made possible the entry of autoradiographic and immunohistochemical techniques into a new era where quantification via densitometry and morphometry has become easily accessible. In this context, quantitative biochemical data can be adapted to anatomical and histological resolution. This adaptation is most efficient in the neuroscience fields because of the huge importance of cellular communication via neuronal networks in the nervous system. Therefore, any experimental approach to the brain which considers the brain as a 'black box' appears now as very crude. In fact, subtle heterogeneity in the distribution of biochemical markers can now be demonstrated, as illustrated here by the use of quantitative autoradiography of D1 and D2 dopaminergic receptors in the striatum of the mammalian brain. Also, local adaptive changes resulting from chronic blockade of the dopaminergic input can be detected after repeated treatments with dopaminergic antagonists selective for D1 or D2 receptors or with surgical lesioning of the dopaminergic nigrostriatal pathway. The resulting plastic changes are unevenly distributed throughout the striatal target organ and vary according to the mode of suppressing the dopaminergic flow: direct destruction of the dopaminergic pathway or selective pharmacological manipulation without physical elimination of the dopaminergic cells themselves. All these results are discussed and reviewed in light of the most recent reports in this field.

Animals↗

PubMed1989

Initial GABAergic expression in embryonic amphibian neuroblasts after neural induction.

At the late gastrula-early neurula stage some embryonic neuroblasts from neural plate and neural fold present apparently as a consequence of neural induction, the capability to develop in vitro into different neuronal subpopulations (cholinergic, dopaminergic, noradrenergic, somatostatinergic and some other peptidergic subpopulations without ongoing influences from the chordamesoderm (Duprat et al., 1987). Using the same in vitro model system, the aim of the present work was to delineate the abilities of these neuroblasts to develop GABAergic traits. The initial appearance and development of GABAergic phenotype has been quantitated by assaying the activity of glutamic acid decarboxylase (GAD). GAD activity was undetectable at the early gastrula stage (stage 8a) and was slightly measurable at the early neurula stage (stage 14- onset of the culture). It increased subsequently over the next 14 days in vitro. The temporal pattern of appearance and development of GAD activity in culture was in agreement with that observed in vivo. Immunocytochemical studies showed that GABA-like immunoreactivity was expressed in vitro in a subpopulation of neurons. Thus the developmental program for GAD expression and GABA phenotype maturation is acquired at least in some neuronal precursors. These data together with previously reported results on the expression of cholinergic, catecholaminergic and peptidergic phenotypes demonstrate that different neuronal subpopulations emerge near the end of gastrulation i.e. immediately after neural induction. The embryonic origin of this neuroblast heterogeneity remains to be determined.

Animals↗

PubMed1988

L-dopa-immunoreactive neurons in the rat hypothalamic tuberal region.

The presence of L-DOPA-immunoreactivity is reported for the first time in the rat hypothalamic tuberal region. L-DOPA-immunoreactive neurons were demonstrated to be present in the ventrolateral part of the arcuate nucleus and periarcuate region just dorsal to the ventral surface of the brain (VLAR/PA). Weakly L-DOPA-immunostained neurons were found in the dorsomedial part of the arcuate nucleus and its neighboring periventricular nucleus (DMAR/PV). In contrast, dopamine (DA)-immunoreactive neurons were detected only in the DMAR/PV. These findings suggest that L-DOPA exists not only as a precursor of DA in neurons of the DMAR/PV, but also as an end-product in cells of the VLAR/PA.

Animals↗

PubMed1988

Endogenous L-dopa, its immunoreactivity in neurons of midbrain and its projection fields in the cat.

L-DOPA (L-3,4-dihydroxyphenylalanine) immunoreactivity was demonstrated in neurons of the cat ventral midbrain and its projection areas, using an immunohistochemical method in conjunction with a newly developed highly specific anti-L-DOPA serum. L-DOPA-immunoreactive (IR) neurons were found in the substantia nigra, retrorubral area and ventral tegmental area of Tsai. L-DOPA-labeled fibers and terminals were hardly detectable in the nigrostriatal pathway and in the caudate nucleus which showed very intense dopamine-immunoreactivity. In contrast, many short labeled processes were detectable in the central amygdala and, although very few in number, in the entorhinal cortex.

Animals↗

PubMed1988

Comparative topography of dopamine- and tyrosine hydroxylase-immunoreactive neurons in the rat arcuate nucleus.

The distribution of dopamine (DA)-immunoreactive (IR) cells is described in the rat arcuate nucleus of the hypothalamus and its adjacent areas and compared with that of tyrosine hydroxylase (TH)-IR cells. Small DA-IR cells were seen to be aggregated mainly in the dorsomedial part of the nucleus, but were hardly detectable in its ventrolateral portion and neighbouring periarcuate region which showed many larger TH-IR cells. This study reveals, for the first time, the differences in the respective topography of those neurons which actually contain detectable DA and those which contain TH, the initial synthesizing enzyme of catecholamine.

Animals↗

PubMed1988

Role of astroglial cell clones in the survival and differentiation of cerebellar embryonic neurons.

To investigate the role of astrocytes in the survival and differentiation of cerebellar neurons during development, we have used astroglial cell clones, derived from 8-day postnatal cerebellar explants and which might be the in vitro equivalents of the 3 main types of cerebellar astrocytes, the Golgi epithelial cells and their Bergmann processes, the velate protoplasmic and the fibrous astrocytes (F. Alliot and B. Pessac, Brain Res., 306 (1984) 283-291). Nearly all single cells, dissociated from 15-day embryonic mouse cerebella and seeded at low density, adhered to layers of each of the cerebellar astroglial cell clones as well as to other glial lines or artificial substrates. However, the cerebellar embryonic neurons survived well only on monolayers of either the 'Golgi-Bergmann'-like or the 'velate protoplasmic'-like clones. On these layers, 60-80% of the neurons were still present after 5 days of co-culture, while only less than 5% survived on the other types of substrates. The differentiation pattern of the neurons surviving on the 'Golgi-Bergmann' and the 'velate protoplasmic' astroglial clones was studied with markers of postmitotic granule cells, the major neuronal population in adult cerebellum. The velate protoplasmic-like clone was the only one able to support the coordinate acquisition by most surviving neurons of the phenotypic characteristics of granule cells, i.e. a distinct morphology, a specific epitope binding the monoclonal antibody 7-8 D2 and immunoreactivity to glutamate. These data show a broad heterogeneity in the capacity of astroglial cell clones to support embryonic cerebellar neurons. In addition, they indicate that neuronal survival per se is not sufficient for the acquisition of a differentiated neuronal phenotype.

Animals↗

PubMed1988

Rapid growth of host afferents into fetal thalamic transplants.

Fetal cell suspension grafts grow and differentiate when implanted into adult rat CNS areas previously neuron-depleted using an excitotoxin. There is some controversy in the literature concerning the timetable of establishment and possible extent of host-graft connections in these experimental conditions. The present study was undertaken to analyze the development of adult host monoaminergic afferents into a transplant formed by fetal thalamic neurons in the previously excitotoxically lesioned thalamus. It is demonstrated that both norepinephrin- and serotonin-immunoreactive fibers are present in the transplant as soon as 8 days after grafting. At those times, immunoreactive fibers exhibit morphological characteristics typically associated with immature stages. After longer survival time, up to 4 weeks after grafting, immunoreactive fibers are numerous in the transplant and exhibit morphological features comparable to those observed in the adult thalamus. These results demonstrate the rapid ingrowth of some fiber systems of the adult host into the transplant and suggest that grafted fetal cells can be functionally integrated into the host circuitry as soon as a few weeks after grafting.

Animals↗

PubMed1988

Visualization of L-dihydroxyphenylalanine in rat brain by using specific antibodies.

L-Dihydroxyphenylalanine (L-DOPA) was conjugated to different protein carriers with glutaraldehyde (G). During the synthesis of the catecholamine conjugates, precautions were taken in order to preserve the structure of L-DOPA. Reduced and non-reduced conjugates were injected to rabbits according to a specific immunization protocol. Anti-L-DOPA antibody affinity and specificity were evaluated by using ELISA tests. The most immunoreactive compounds were the non-reduced conjugate, L-DOPA = G = BSA and the reduced one, L-DOPA-G-BSA. The other conjugated catecholamines were poorly recognized or not at all. These antisera enabled us to specifically visualize the precursor of the catecholaminergic neurotransmitters which are: dopamine, noradrenaline and adrenaline in the G-fixed rat brains.

Animals↗

PubMed1988

Demonstration of high specificity antibodies against phosphatidylserine.

Anti-phosphatidylserine antibodies were raised in rabbits immunized with phosphatidylserine-polyacrylamide gels and with phosphatidylserine-cytochrome c vesicles. A solid-phase immunoenzymatic assay (ELISA) was developed to study the immune serum. The optimal conditions were defined and the technique used to obtain quantitative results. The anti-phosphatidylserine serum diluted to 1/1000 was tested against several phospholipids and was found to be highly specific to phosphatidylserine. This represents the first experimental demonstration of the specificity of antibodies raised against phosphatidylserine.

Animals↗

PubMed1988

Immunocytochemical detection of GABAergic nerve cells in the human temporal cortex using a direct gamma-aminobutyric acid antiserum.

Recently, an immunocytochemical method using glutaraldehyde fixation and an antiserum developed against a GABA--glutaraldehyde--protein conjugate has permitted direct visualization of GABAergic structures in the brains of perfused animals. This paper reports a successful use of this technique on human temporal cortex fixed by immersion. The cerebral tissue was obtained from patients operated for focal epilepsy. GABA-positive somata, fibres and terminals are observed in all layers of the temporal cortex. Terminals are particularly abundant in the superficial portion of layer I and in layers II, III and IV. Dense plexuses of fibres are located in layers II, III, IV and VI and in the underlying white matter. Somata are found in all cortical layers and in the underlying white matter; they are round, oval, fusiform or triangular and exhibit a multipolar, bitufted or bipolar dendritic pattern. This technique for the visualization of GABAergic structures in the human brain may allow a better understanding of the pathogeny of epilepsy in which the GABAergic transmission has been implicated.

Humans↗