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Biomedical subjects

Y Tillet

Publications and source records attributed to Y Tillet.

At least 55 records · Page 3Linked to original sources
PubMed1993

Preparation of an antiserum using a fusion protein produced by a cDNA for rat aromatic L-amino acid decarboxylase.

Aromatic L-amino acid decarboxylase (AADC) decarboxylates L-DOPA and 5-hydroxytryptophan into dopamine and serotonin, respectively. Starting from a rat AADC cDNA clone isolated in our laboratory, we produced a beta-galactosidase-AADC fusion protein in E. coli. It was purified from inclusion bodies and injected into a rabbit. The antiserum identified AADC on a Western blot of extracts from rat organs as a unique 50 kDa band; it also strongly reacted by immunohistochemistry with dopaminergic and serotonergic neurons. This new beta-galactosidase-AADC fusion protein then constitutes a useful tool for producing AADC as an antigen free of contamination by mammalian proteins.

Animals↗
PubMed1993

Neuronal projections to the medial preoptic area of the sheep, with special reference to monoaminergic afferents: immunohistochemical and retrograde tract tracing studies.

The preoptic area contains most of the luteinizing hormone releasing hormone immunoreactive neurons and numerous monoaminergic afferents whose cell origins are unknown in sheep. Using tract tracing methods with a specific retrograde fluorescent tracer, fluorogold, we examined the cells of origin of afferents to the medial preoptic area in sheep. Among the retrogradely labeled neurons, immunohistochemistry for tyrosine hydroxylase, dopamine-beta-hydroxylase, phenylethanolamine N-methyltransferase, and serotonin was used to characterize catecholamine and serotonin fluorogold labeled neurons. Most of the afferents came from the ipsilateral side to the injection site. It was observed that the medial preoptic area received major inputs from the diagonal band of Broca, the lateral septum, the thalamic paraventricular nucleus, the lateral hypothalamus, the area dorsolateral to the third ventricle, the perimamillary area, the amygdala, and the ventral part of the hippocampus. Other numerous, scattered, retrogradely labeled neurons were observed in the ventral part of the preoptic area, the vascular organ of the lamina terminalis, the ventromedial part of the hypothalamus, the periventricular area, the area lateral to the interpeduncular nucleus, and the dorsal vagal complex. Noradrenergic afferents came from the complex of the locus coeruleus (A6/A7 groups) and from the ventro-lateral medulla (group A1). However, dopaminergic and adrenergic neuronal groups retrogradely labeled with fluorogold were not observed. Serotoninergic fluorogold labeled neurons belonged to the medial raphe nucleus (B8, B5) and to the serotoninergic group situated lateral to the interpeduncular nucleus (S4). In the light of these anatomical data we hypothesize that these afferents have a role in the regulation of several functions of the preoptic area, particularly those related to reproduction. Accordingly these afferents could be involved in the control of luteinizing hormone releasing hormone (LHRH) pulsatility or of preovulatory LHRH surge.

Animals↗
PubMed1992

Immunohistochemical colocalization of tyrosine hydroxylase and estradiol receptors in the sheep arcuate nucleus.

In sheep, the arcuate nucleus contains numerous tyrosine hydroxylase (TH) and estradiol receptor (rE2) immunoreactive (IR) perikarya and it has been shown previously in this species that catecholaminergic neurons can mediate the gonadal steroid action on the reproductive function. In the present study, double immunohistochemical labelling with antibodies against TH and rE2 have been used to demonstrate the presence of rE2 in TH-IR neurons in the arcuate nucleus where the distribution of TH-IR and rE2-IR neurons overlap each other. Only less than 10% of all the rE2-IR perikarya presented TH immunoreactivity. It was therefore hypothesized that either such a low number of double labelled neurons can support the effects of estradiol in this area or that the effect of this steroid was indirect. In the latter case it might be first mediated by beta-endorphin neurons which have been previously described in this nucleus.

Animals↗
PubMed1992

Serotoninergic projections from the raphe nuclei to the preoptic area in sheep as revealed by immunohistochemistry and retrograde labeling.

A retrograde tracer, fluorogold, was injected into the sheep preoptic area in order to demonstrate the origin of the serotoninergic fibers observed in this area. Within the raphe nuclei, retrogradely fluorogold-labeled neurons were observed mainly in the median raphe nucleus (B8), groups B6/B5, and in the area lateral to the nucleus interpeduncularis (group S4), but not in the dorsal raphe nucleus. About 50% of these fluorogold-containing neurons were immunostained with a specific antiserum raised against serotonin. Double-labeled neurons (serotonin-immunoreactive and fluorogold containing neurons) represented less than 20% of the whole number of serotoninergic neurons. We concluded that a few serotoninergic neurons in the median raphe nucleus and in groups B5/B6 and S4 project to the preoptic area. Moreover, these nuclei contained non-serotoninergic neurons which project to the same area. These results give new information on the serotoninergic innervation of the preoptic area in the sheep.

Animals↗
PubMed1992

Immunohistochemical demonstration of melatonin in the female mink harderian gland.

In the Harderian gland of the female mink, either intact or killed after a bilateral ablation of the cervical superior ganglion, almost all of the cells of the alveoli were immunolabeled with anti-melatonin antiserum. Animals were killed during the day or during the night. The immunolabelling was observed only in the cytoplasm, while the nucleus remained unstained. Using successive dilutions of the antiserum on serial sections of the Harderian gland to qualitate the melatonin content, a circadian rhythm of melatonin immunoreactivity was observed. The intensity of immunofluorescence labelling was higher in intact animals killed during the day than in those killed during the night. These results could be explained by the inhibitory or stimulatory influence of pineal melatonin released during the night on melatonin synthesis or release in the Harderian gland, respectively. In the Harderian gland of ganglionectomized animals, the intensity of melatonin immunofluorescence was lower than in intact animals killed during the day. It is concluded that the Harderian gland might be involved in the perception of the day/night cycle and that melatonin synthesis/secretion was likely controlled by the cervical superior ganglion in this organ.

Animals↗
PubMed1992

Localization of luteinizing hormone beta-mRNA by in situ hybridization in the sheep pars tuberalis.

The localization of luteinizing hormone beta (LH beta)-mRNA was studied by in situ hybridization in the pars tuberalis of sheep using a homologous sheep double-stranded 32P- or 35S-cDNA. The labelled cDNA probe detected one mRNA sequence in the pars tuberalis by Northern blot analysis; this sequence was similar to that detected in the pituitary. In situ, the labelling of LH beta-mRNA in the horizontal and sagittal tissue sections was found throughout the pars tuberalis. This labelling was prevented by adding an excess of cold probe or treating the sections by ribonuclease before in situ hybridization. Controls showed a labelling in the pars distalis, but not in the median eminence, hypothalamus, cerebral cortex and liver sections. Double labelling by in situ hybridization followed by immunohistochemistry using a specific LH beta-antiserum indicated that the labelling of LH beta-mRNA appeared more intense in LH-containing cells that were found only in the ventral part of the pars tuberalis. These results suggest that the entire pars tuberalis is able to produce the LH beta subunit, but that the level of translation greatly varies according to the location of the cells.

Animals↗
PubMed1991

Melatonin binding sites in the brain of sheep exposed to light or pinealectomized.

The binding sites of [125I]melatonin were identified in the sheep brain using a specific and sensitive autoradiographical method. Rams were either untreated (controls) or exposed to light before slaughter or pinealectomized (px). In all animals labelling was intense in the pars tuberalis (PT) and absent in the suprachiasmatic nucleus (SCN). In light-treated and in px rams, but not in controls, we demonstrated melatonin binding sites in the intermediate and ventrolateral septum and in the stratum lacunosum of the hippocampus. Labelling was less marked in nervous tissues than in PT cells and did not seem to be different between treatments. These results indicate the presence of light-dependent melatonin binding sites in the septum and hippocampus of sheep, but rise the question of the involvement of melatonin in the SCN activity in this species.

Animals↗
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

The sheep pars tuberalis: an immunohistochemical study. Demonstration of the presence of glycoprotein and lipotropin hormones.

Using indirect immunofluorescence with fourteen different antisera raised against pituitary hormones and peptides, we characterized immunochemically the cells of the sheep pars tuberalis. The presence of LH- and FSH-containing cells, shown in previous studies, was also observed in the present investigation. In addition, we found TSH-containing cells, never observed in sheep, and beta LPH-containing cells. The latter hormone has never been found in any studied species. It appeared that a small amount of perikarya (less than 20%) were immunolabelled and, that the sheep pars tuberalis contained a majority of immunonegative cells as in the guinea-pig rabbit and rhesus monkey. This study may contribute to a better knowledge of the function of the sheep pars tuberalis.

Animals↗
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

Melatonin binding sites in the sheep pars tuberalis.

The localization of [125I]melatonin binding sites has been studied by autoradiography on frozen unfixed sections of the pituitary stalk, the suprachiasmatic area, the pineal and the pituitary glands in sheep. Dense specific labelling has been found exclusively in the pars tuberalis of the pituitary stalk but not in the part of the median eminence surrounded by the pars tuberalis. The labelling was completely excluded by a 200-fold excess of cold melatonin. No comparable labelling was found in the suprachiasmatic nucleus, the pineal and the pituitary glands. These results constitute the first report of melatonin-specific labelling in the ovine species.

Animals↗
PubMed1989

Immunohistochemical demonstration and radioimmunoassay of melatonin in the mink pineal gland.

An antiserum raised against N-amino-3-propyl melatonin bound to a protein carrier was used to visualize melatonin by immunohistochemistry and to measure melatonin concentration by radioimmunoassay in the pineal gland of intact mink females killed throughout the 24 h cycle and females killed after a bilateral ablation of the cervical superior ganglion. Melatonin immunoreactivity revealed by immunofluorescence or by the peroxidase-antiperoxidase complex was observed in the cytoplasm of presumed pinealocytes of all the females. Circadian changes in pineal melatonin content were not visualized by immunohistochemistry; furthermore, immunoreactivity was also present in the pineal gland of the ganglionectomized females. However, the melatonin content measured by radioimmunoassay was significantly higher in the pineal gland from intact females killed during the night compared with that of intact females killed during the day or of ganglionectomized females. The discrepancy between the results obtained using the two methods may arise because immunohistochemistry can detect very small amounts of melatonin.

Animals↗
PubMed1989

Role of hypothalamic catecholamines in the regulation of luteinizing hormone and prolactin secretion in the ewe during seasonal anestrus.

Separate studies with ewes have shown that catecholamines play an inhibitory role in the control of LH secretion during anestrus, and that there are structures in the lateral retrochiasmatic area (L-RCh), which could be involved in the regulation of gonadotrophin secretion. These observations have led to the hypothesis that the catecholaminergic structures in the L-RCh mediate the inhibition of pulsatile LH secretion by estradiol in the anestrous ewe. We tested this hypothesis by injecting 6-hydroxydopamine (6OH-DA) into the L-RCh of ovariectomized ewes during the anestrous season, and comparing the secretion of LH and prolactin in these animals with that in sham (injected with vehicle) and control (no injection) animals, in the presence and absence of exogenous estradiol. Finally, the effectiveness of the toxin was assessed by immunocytochemical techniques. When the ewes were treated with estradiol, LH pulse frequency was significantly lower in the controls (mean 1.1 pulses/4 h) and shams (0 pulses/4 h) than in the ewes treated with 6OH-DA (3.1 pulses/4 h). When the estradiol implants were removed, the frequencies increased to 5.1 pulses/4 h for the controls and 5.7 pulses/4 h in the ewes treated with 6OH-DA. These were not significantly different. Plasma prolactin levels were significantly reduced by 6OH-DA treatment. The 6OH-DA ewes recovered their response to estradiol by 14 weeks after the injection. The anatomical study at the end of the experiment revealed a difference between treated and control ewes of only 15% in the numbers of dopaminergic cells in the L-RCh.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Ectopic prolactinoma within the sphenoid sinus.

A case of an ectopic prolactin-producing tumor located within the sphenoid sinus is reported. It was discovered in a girl who experienced secondary amenorrhea without galactorrhea. We discuss the different diagnoses proposed before immunocytochemical staining results were available. This case presented unusual radiological, surgical, and histopathological aspects.

Adolescent↗
PubMed1989

Non-involvement of the accessory olfactory system in the LH response of anoestrous ewes to male odour.

In anoestrous ewes, male chemosignals elicit rapid increases in luteinizing hormone (LH) secretion that can ultimately lead to ovulation. To assess the possible involvement of the accessory (vomeronasal) olfactory system in the mediation of those chemical cues, we destroyed this pathway by vomeronasal organ electrocauterization (Exp. I) and vomeronasal nerve section (Exp. II). Neither of these lesions inhibited the LH response of ewes to the odour of the male. These results suggest that the vomeronasal system is not necessary to mediate the neuroendocrine response of the ewe to the male odour. As both surgical methods spared the main olfactory system but destroyed the vomeronasal system, it is likely that the main olfactory system is involved in the LH response to chemical stimulation in sexually experienced ewes.

Anestrus↗
PubMed1988

Adrenergic neurons in sheep brain demonstrated by immunohistochemistry with antibodies to phenylethanolamine N-methyltransferase (PNMT) and dopamine-beta-hydroxylase (DBH): absence of the C1 cell group in the sheep brain.

Using immunohistochemistry with specific antisera against dopamine-beta-hydroxylase and phenylethanolamine N-methyltransferase, we present the first description of the adrenergic structures of the sheep medulla oblongata. Dopamine-beta-hydroxylase-immunoreactive perikarya in the sheep brain are localized as described in the rodents (A1 and A2 groups) but their distribution is characterized by only one phenylethanolamine N-methyltransferase immunoreactive cell body group, found in the nucleus tractus solitarius. This group corresponds to the C2 group previously described in the rat, but neither group C1 nor group C3 are found in the sheep with our method. Compared with rodents or primates, this animal presents a different pattern of central adrenergic innervation and could be an alternative model to study the central role of adrenaline in various physiological functions as different as swallowing or reproduction.

Adrenergic Fibers↗
PubMed1988

Early ontogeny of serotonin-immunoreactivity in the sheep brain. An immunohistochemical study.

Using immunohistochemistry with specific antiserotonin anti-sera, the ontogeny of serotonergic neurons was studied in the foetal sheep brain. Serotonergic-immunoreactive perikarya first appeared rostrally on day 25 of pregnancy, in the medio-ventral part of the mesencephalic flexure, and caudally, on day 28, in the medio-ventral part of the cervical flexure. The development of this system is very rapid, because on day 40 of gestation, all serotonergic nuclei present in the adult were visible. Compared with other species such as rodents or primates, serotonin appears early in the sheep nervous system, and the development of the serotonergic system is even more rapid. Serotonergic immunoreactivity was seen in some cell bodies in the growing adenohypophysis between days 40 and 50. This phenomenon has not been observed in other species. Because serotonin appears very early and is present in growing areas of the nervous system, it could play a trophic role in the development and maturation of the sheep central nervous system, as has been described previously in other species.

Animals↗