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J Thibault

Publications and source records attributed to J Thibault.

At least 91 records · Page 5Linked to original sources
PubMed1990

Presence of dopamine-immunoreactive cell bodies in the catecholaminergic group A15 of the sheep brain.

Antisera were raised in rabbits against dopamine or noradrenaline conjugated to thyroglobulin with glutaraldehyde. These antisera, tested in enzyme linked immunosorbent assay and immunohistochemistry specifically recognized their homologous antigens. With the aid of anti-tyrosine hydroxylase, anti-aromatic aminoacid decarboxylase, anti-dopamine-beta-hydroxylase, anti-dopamine, and anti-noradrenaline antisera, immunohistochemical reactions were performed on glutaraldehyde fixed sections of sheep diencephalon in order to determine the presence of dopamine in the catecholaminergic group A15. Perikarya of this nucleus were stained with anti-tyrosine hydroxylase, anti-aromatic aminoacid decarboxylase and anti-dopamine, but not with anti-dopamine-beta-hydroxylase or anti-noradrenaline. Both of these latter antisera stained fibers within this area. So as recently found in the rat, we could conclude that dopamine is present in group A15 of the sheep.

Animals↗
PubMed1990

Topographical relationships between catecholamine- and neuropeptide-containing fibers in the median eminence of the newt, Triturus alpestris. An ultrastructural immunocytochemical study.

Dopaminergic and peptidergic nerve fibers were simultaneously demonstrated with a double-labeling technique at the ultrastructural level. The first antibody, raised against tyrosine hydroxylase, was applied during the preembedding phase and visualized with the peroxidase method. The second antibody, raised against one of the peptides met-enkephalin, somatostatin or gonadotropin-releasing hormone (GnRH), was applied to the ultrathin sections and visualized with gold-labeled goat anti-rabbit IgG. The fibers of both categories were present in the zona externa of the median eminence, frequently contacting the basal lamina of the portal vessels. In addition, topographical relationships between different types of nerve fibers were observed in the perivascular areas, although there were no morphological signs of synaptic specializations. Using serial sections, it could be established that one GnRH-fiber contacted both a dopaminergic fiber and a fiber immunoreactive for met-enkephalin. The observations support earlier physiological data concerning the regulation of the hypothalamo-hypophyseal axis, with special emphasis on the release of neurohormones in the median eminence of the newt.

Animals↗
PubMed1990

Influence of initial collagen and cellular concentrations on the final surface area of dermal and skin equivalents: a Box-Behnken analysis.

Our laboratory has been involved in finding optimal conditions for producing dermal and skin equivalents. As an original approach, a Box-Behnken experimental design was used to study the effects of the initial collagen and fibroblast concentrations and the initial gel thickness on the contraction of dermal and skin equivalents. The final surface area of dermal equivalent varied significantly with the initial concentration of collagen and fibroblast, whereas the initial thickness of gel had no appreciable effect on the contraction of the dermal equivalent. When keratinocytes were grown on these dermal equivalents they produced a very severe contraction, to an extent that all skin equivalents had a similar final surface area. This severe contraction was independent of collagen and fibroblast concentrations. Models for the prediction of the final percentage contraction of dermal and skin equivalents as a function of the initial concentration of collagen, the logarithm of fibroblast concentration, and the initial gel thickness were obtained and analyzed. Keratinocytes grown at the lowest seeding density did not contract the equivalents. However, histologic analysis has shown an incomplete coverage by these cells of the equivalents. The extensive contraction of the skin equivalent presenting adequate morphology is a major drawback toward its clinical utilization for burn wound coverage.

Cells, Cultured↗
PubMed1990

Characterization of DOPA decarboxylase mRNA in rat pheochromocytoma.

Total poly (A+) RNA has been extracted from rat pheochromocytoma and translated in vitro by means of a reticulocyte lysate system. We show that two antisera, prepared against pig kidney DOPA decarboxylase (DDC) or rat pheochromocytoma DDC, immunoprecipitate an in vitro synthetized 50 kDa polypeptide identified as DDC by competition experiments with pure DDC. The proportion of specific mRNA has been calculated and represents 0.05% of total poly A+ mRNA. Its size has been established by electrophoresis in methylmercuric hydroxide containing agarose gel, corresponding to a 2.2 kb length mRNA.

Animals↗
PubMed1990

Preparation of a human DOPA decarboxylase cDNA probe by PCR and its assignment to chromosome 7.

The reverse transcription of mRNA of a human pheochromocytoma using an oligonucleotide complementary to rat DOPA decarboxylase (DDC) sequence as a primer, gave rise to a single strand cDNA. This resultant cDNA permitted by PCR amplification the preparation and cloning in PUC 19 of a human DDC probe of 747 base pairs. The choice of primer was dictated by the presence of tryptophan codons in the DOPA decarboxylase sequence which were conserved in different species throughout evolution. The presence of mismatches on the primers was not an obstacle to a specific amplification and the probe sequence was found identical to the human DDC cDNA sequence. This probe, labelled by nick translation detected on Southern blot of human-rodent hybrids, bands on human DNA after EcoRI, BamHI or HindIII digestion. The results with EcoRI were explicit and drove to the conclusion that DDC is located on chromosome 7. Five bands were obtained on human DNA digested with EcoRI, indicating that either numerous introns could interrupt the coding sequence of DDC gene or duplicated sequences could be present in chromosome 7.

Base Sequence↗
PubMed1989

Catecholamine-containing neurons in the sheep brainstem and diencephalon: immunohistochemical study with tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DBH) antibodies.

The present study describes the distribution and morphological characteristics of neurons and nerve fibers containing the catecholamine-synthesizing enzymes, tyrosine hydroxylase and dopamine-beta-hydroxylase, in the sheep brainstem and diencephalon on the basis of immunohistochemical procedures. Neurons and fibers were considered to be dopaminergic if they showed anti-tyrosine hydroxylase immunoreactivity, without corresponding anti-dopamine-beta-hydroxylase immunoreactivity. The structures labeled with both antisera were considered noradrenergic or adrenergic. The distribution of catecholaminergic neurons corresponds to that described by other authors with similar methods in the rat and in primates. The noradrenergic neurons belong to cell groups A1 to A7 and the dopaminergic neurons to cell groups A8 to A15. In almost all studied areas, the catecholaminergic innervation is similar to that observed in the other species. However, the central catecholaminergic systems of the sheep showed some specific characteristics: (1) groups A3 and A4, described in the rat, were not found, (2) group A14 contains fewer neurons than in the rat, (3) group A15 does not contain a dorsal but only a ventral portion, (4) there is a larger dispersion of neurons within each group, especially A6 and A7, than in rodents, and (5) there is a larger noradrenergic innervation of the catecholaminergic groups than in the other species.

Adrenergic Fibers↗
PubMed1989

Ontogenesis of tyrosine hydroxylase-immunopositive structures in the rat hypothalamus. An atlas of neuronal cell bodies.

The development of the catecholaminergic system in the hypothalamus and in the septal region was studied in rats from the 12th fetal day until the 9th postnatal day. Catecholaminergic structures were visualized with pre-embedding immunocytochemistry using antiserum to tyrosine hydroxylase. An intensification of diaminobenzidine product with silver and gold was additionally applied to make the immunocytochemical technique more sensitive. In this paper only the data on the appearance and distribution of the tyrosine hydroxylase-immunopositive neurons (cell bodies) are presented, whereas the catecholaminergic innervation of the hypothalamus with the tyrosine hydroxylase-immunopositive fibers is the topic of an accompanying paper. Sparse tyrosine hydroxylase-immunopositive neurons were first observed in the anlage of the hypothalamus and septal region on the 13th fetal day. Their number increased progressively with age and by the 15th fetal day they already gave rise to a large dorsal accumulation. From the 18th fetal day on, tyrosine hydroxylase immunopositive neurons began to occupy their definitive positions, mainly concentrating within the hypothalamus: in the zona incerta, periventricular and arcuate nuclei. To a lesser extent, they were concentrated in the medial preoptic area, suprachiasmatic, supraoptic, paraventricular, dorsomedial, and anterior hypothalamic nuclei. The data on the distribution of the tyrosine hydroxylase-immunopositive neurons both in the hypothalamus and in the septal region during ontogenesis are summarized in the precise atlas. Primarily small bi- and unipolar catecholaminergic neurons first observed in the youngest fetuses undergo cytodifferentiation during ontogenesis, giving rise to at least two different populations localized ventrally, mainly in the arcuate nucleus, and dorsally, in the zona incerta. The neurons of the former population remain similar to those of the youngest fetuses, whereas the neurons of the latter increase significantly in size, forming several long, highly ramified processes.

Aging↗
PubMed1989

Ontogenesis of tyrosine hydroxylase-immunopositive structures in the rat hypothalamus. Fiber pathways and terminal fields.

The innervation of the hypothalamus and septal region by catecholaminergic fibers was studied in rats from the 12th fetal day until the 9th postnatal day. Catecholaminergic fibers were visualized with preembedding immunocytochemistry using antibodies to tyrosine hydroxylase. An intensification of diaminobenzidine product with silver and gold was additionally applied to increase the sensitivity and resolution power of the routine immunocytochemical technique. It has been demonstrated that, from the 13th fetal day, the hypothalamus and the septal region receive catecholaminergic fibers either belonging to the hypothalamic neurons or coming with the medial forebrain bundle from the outside of the hypothalamus. As the development of the hypothalamus proceeds, these fibers form the extensive networks within some neurosecretory centers either containing (the zona incerta, periventricular nucleus, etc.) or almost lacking (suprachiasmatic and paraventricular nuclei) the catecholaminergic neurons. In the former case, they terminate on the processes or perikarya of catecholaminergic neurons, while in the latter case their varicosities surround the immunonegative presumptive neurons in a basket-like manner. Moreover, from the 18th fetal day catecholaminergic fibers penetrate between the ependymal cells towards the 3rd ventricle and the primary capillary plexus of the hypophysial portal circulation, apparently providing the release of catecholamines to the cerebrospinal fluid and portal blood, respectively. The data obtained in this study are considered as the morphological basis for the involvement of the hypothalamic catecholamines in neuroendocrine regulations during ontogenesis.

Aging↗
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↗
PubMed1988

Role of the N-terminus of rat pheochromocytoma tyrosine hydroxylase in the regulation of the enzyme's activity.

Activation of rat pheochromocytoma tyrosine hydroxylase by limited tryptic proteolysis was investigated. The modifications produced upon the enzyme's structure were analyzed with the use of sodium dodecyl sulfate/polyacrylamide gel electrophoresis and tyrosine hydroxylase activity was measured all through the digestion. During the proteolysis the activity of tyrosine hydroxylase was elevated threefold at the same time as a 56-kDa tryptic fragment was formed. When the enzyme was phosphorylated, at its N-terminal region, by a kinase copurified with tyrosine hydroxylase, the major 56-kDa species did not appear to be phosphorylated on the autoradiograph, suggesting that it was derived from the native subunit by cleavage of the N-terminal of the protein. The reactivity of the 2/40/15 anti-(tyrosine hydroxylase) monoclonal antibody with the N-terminal of tyrosine hydroxylase was also investigated, using the Western-blot technique. This antibody reacted with the 62-kDa hydroxylase subunit but not with the 60-kDa tryptic fragment; the amino acid sequences of these two species showed that the 60-kDa fragment lacked the first 16 N-terminal amino acids of the native molecule. These results suggest that the N-terminal region of tyrosine hydroxylase is apparently responsible for an inhibition of the hydroxylase activity and that the first N-terminal amino acids of the hydroxylase are necessary for the recognition of the enzyme by its antibody.

Amino Acid Sequence↗
PubMed1988

Differential distribution of biogenic amines in the hypoglossal nucleus of the rat.

The distribution of biogenic amines in the rat hypoglossal nucleus (nXII) was investigated with immunocytochemical methods using antibodies to tyrosine hydroxylase (TH) as a marker for catecholamines, and to 5-hydroxytryptamine (5-HT), the principal indoleamine, at the light microscopic level. TH and 5-HT immunoreactivity were found throughout all regions of nXII. Although the innervations overlapped, clearly different patterns of distribution were observed. TH immunoreactivity was localized primarily in the ventromedial quadrant of the caudal half of nXII and appeared largely as perisomatic-like profiles. In contrast, 5-HT immunoreactivity was greatest dorsally along the caudal half of nXII, although secondary foci of staining were evident ventrolaterally and, to a lesser extent, ventromedially. A perisomatic-like pattern of termination was observed for 5-HT in both dorsal and ventral regions of nXII. Since ventral and dorsal districts of nXII contain motoneurons that innervate protrusor and retrusor tongue muscles, respectively, we propose that the overlapping, yet differential distributions of catecholamines and indoleamines are important in controlling the relationships between functionally related groups of nXII motoneurons. These findings are discussed in relation to oro-lingual motor dysfunction.

Animals↗
PubMed1988

Topographic immunocytochemical mapping of monoamine oxidase-A, monoamine oxidase-B and tyrosine hydroxylase in human post mortem brain stem.

Immunocytochemical demonstration of monoamine oxidase-A, monoamine oxidase-B and tyrosine hydroxylase was performed in the human brain stem using monoclonal antibodies to monoamine oxidase-A and monoamine oxidase-B and polyclonal antibodies to tyrosine hydroxylase. In most of the brain areas examined, except the serotonergic dorsal nucleus of raphe, the noradrenergic locus coeruleus and the dorsal efferent nucleus of vagus, tyrosine hydroxylase-positive neurons were in greater number than monoamine oxidase-A-stained or monoamine oxidase-B-stained neurons. The dorsal nucleus of raphe showed no tyrosine hydroxylase immunoreactivity, but reacted positively to serotonin- and monoamine oxidase-B antibodies, while monoamine oxidase-A staining was moderate. In none of the investigated brain areas did neurons exclusively react with monoamine oxidase-B antibodies without expressing monoamine oxidase-A in a few neurons, while in some areas neurons expressed both monoamine oxidase-A and tyrosine hydroxylase (locus coeruleus; dorsal efferent nucleus of vagus). The oculomotor nucleus stained only with monoamine oxidase-A antibodies, substantia nigra neurons reacted only with tyrosine hydroxylase antibodies. Glial staining in most of the brain areas examined seemed, with slight differences, to have the same intensity with monoamine oxidase-A and monoamine oxidase-B antibodies used. No glial staining was obtained with tyrosine hydroxylase antibodies.

Aged↗
PubMed1988

[Immunocytochemical study of catecholaminergic neurons in grafted mesencephalon transplanted into the hypothalamus of the rat].

Mesencephalic fragments from 14 day old embryonic rat brain were transplanted into the third ventricle of adult rats neonatally treated with monosodium glutamate. From two to twelve months after grafting, the implanted tissue was still present in the ventricle and contained TH immunoreactive neurons which displayed a normal appearance at ultrastructural level. While endogenous TH containing neurons were still present in dopaminergic regions of the recipient hypothalamus, grafted mesencephalic fragments could survive and develop. They contained TH immunopositive most probably dopaminergic neurons which are able, in some cases, to innervate the host brain. This model should be of interest in the study of neuroendocrine functions of dopaminergic neurons.

Animals↗
PubMed1987

Rat pheochromocytoma tyrosine hydroxylase is phosphorylated on serine 40 by an associated protein kinase.

Tyrosine hydroxylase, a key enzyme in the biosynthesis of catecholamines, was previously shown to be phosphorylated on four distinct serine residues in PC12 cell cultures, each one being specific for the kinase system involved (McTigue, M., Cremins, J., and Halegoua, S. (1985) J. Biol. Chem. 260, 9047-9056). A cAMP- and Ca2+-independent protein kinase was found to be associated with tyrosine hydroxylase purified from rat pheochromocytoma tumor. The use of this activity and the availability of a large amount of purified tyrosine hydroxylase allowed identification of the site phosphorylated by this kinase activity. A peptide of 1.5 kDa (about 12 residues long), carrying the phosphorylation site, was released from 32P-labeled tyrosine hydroxylase by limited proteolysis with trypsin. This peptide was isolated from trypsinized tyrosine hydroxylase by sequential gel filtration and ion exchange chromatographies. Analysis by thin layer chromatography of an acid hydrolysate of the peptide revealed that it contained phosphoserine. The sequence determination of the peptide showed that it corresponded to the residues 38-45 in the tyrosine hydroxylase primary structure (Arg-Gln-Ser(P)-Leu-Ile-Glu-Asp-Ala). Thus, the associated kinase phosphorylated Ser-40, one of the phosphorylation sites for the cAMP-dependent protein kinase also found in rat pheochromocytoma tumors. These results are compared to those recently appearing in a report by Campbell et al. (Campbell, D. G., Hardie, D. G., and Vulliet, P. R. (1986) J. Biol. Chem. 261, 10489-10492).

Amino Acid Sequence↗
PubMed1987

Early ontogeny of catecholaminergic structures in the sheep brain. Immunohistochemical study.

The localization of tyrosine hydroxylase was studied in the brain of sheep foetus during early ontogeny using immunohistochemistry. The first immunoreactive neurons appeared very early since they were found on day 30 of pregnancy in the medioventral part of the mesencephalic flexure. The distribution of the different catecholaminergic groups of neurons was similar to the adult's after 75 days of pregnancy. The latest groups to appear was the A12 group. Comparison of the development of the sheep foetus with rodents or primates, more commonly studied, is difficult because of its different development. It seems, however, that catecholaminergic structures appear earlier in sheep and rodents than in human. Considering the early appearance of these transmitters in the central nervous system, their role on brain development has to be studied in the future.

Animals↗
PubMed1987

Immunocytochemical demonstration of the presence of catecholamine and serotonin neurons in the sheep olfactory bulb.

The catecholamine and serotonin innervation of the sheep olfactory bulb was studied using immunocytochemistry. Specific antisera raised against tyrosine hydroxylase, dopamine beta-hydroxylase, phenylethanolamine N-methyl transferase and serotonin were used. Tyrosine hydroxylase-positive cell bodies were present in all cell layers except in the anterior olfactory nucleus, the greatest number being found in the glomerular layer. Neither dopamine beta-hydroxylase-positive nor serotonin-positive cell bodies were observed. Dopamine beta-hydroxylase-positive fibers were widely distributed in the granule cell layer but less widely in other layers. The glomerular layer contained the greatest distribution of serotonergic positive fibers, but such fibers were also visualized in other cell layers. No phenylethanolamine N-methyl transferase-positive structures were found in this investigation.

Animals↗