Biomedical subjects

G Pelletier

Publications and source records attributed to G Pelletier.

At least 721 records · Page 40Linked to original sources

PubMedDate not supplied

Localization of melanin-concentrating hormone-like immunoreactivity in the brain and pituitary of the frog Rana ridibunda.

The distribution of melanin-concentrating hormone (MCH) in the central nervous system of the frog Rana ridibunda was determined by the indirect immunofluorescence technique using antibodies against synthetic salmon MCH, generated in rabbits. The most prominent group of MCH-like containing perikarya was detected in the preoptic nucleus. Comparatively, a moderate number of cell bodies was observed in the dorsal infundibular nucleus and in the ventral thalamic area. Brightly immunofluorescent nerve bundles were found in the preoptic nucleus and in the ventral infundibular nucleus, coursing towards the internal zone of the median eminence and the pituitary stalk. An intense network of immunofluorescent fibers was localized in the neural lobe of the pituitary. The subcellular localization of MCH-like material was studied in the neurohypophysis using the immunogold technique. It was demonstrated that MCH-like material was contained in dense core vesicles (80-90 mm in diameter) within specific nerve terminals. The present findings indicate that, in amphibians, MCH-like peptide is located in specific hypothalamic neurons. Our data suggest that MCH may be released by neurohypophyseal nerve endings as a typical neurohormone.

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Endorphinic neurons are contacting the tuberoinfundibular dopaminergic neurons in the rat brain.

The anatomical relationships between endorphinic neurons and dopaminergic neurons were evaluated in the rat hypothalamus using a combination of immunocytochemistry and autoradiography. In the arcuate nucleus, endorphinic endings were seen making contacts with dopaminergic cell bodies and dendrites. No synapsis could be observed at the sites of contacts. These results strongly suggest that the endorphinic neurons are directly acting on dopaminergic neurons to modify the release of dopamine into the pituitary portal system.

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Distribution of neuropeptide Y immunoreactivity in human visual cortex and underlying white matter.

Immunocytochemical techniques have been used to study neuropeptide Y (NPY) distribution in the human visual cortex (Brodman's areas 17, 18 and 19) NPY cell bodies belong mostly to inhibitory (multipolar and bitufted) but also to excitatory (bipolar and some pyramidal) neuronal types. Their distribution is similar in the three cortical areas studied: 20 to 40% of the NPY perikarya are located in the cortical gray matter, mostly in the deep layers, while the remaining 60 to 80% are located in the underlying white matter. Immunoreactive NPY processes form a rich network of intersecting fibers throughout the entire visual cortex. A superficial plexus (layers I and II) and a deep plexus (deep layer V and layer VI) of NPY fibers are present in areas 17, 18 and 19. In area 17, an additional well developed plexus is present in layers IVb and IVc. These plexuses receive branches from long parallel fibers arising from deep cortical layers or underlying white matter and terminating in superficial layers. Local or extrinsic NPY terminals wind around vessels in the cortex as well as in the white matter, and either penetrate them or form clusters of club endings on their walls. Our results suggest a role for NPY in human visual circuitry and in cortical blood flow regulation.

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An NPY-like peptide may function as MSH-release inhibiting factor in Xenopus laevis.

This study demonstrates the presence of a rich plexus of neuropeptide Y (NPY) immunoreactive fibers in the hypothalamus and in the intermediate lobe of the pituitary of Xenopus laevis. During superfusion of neurointermediate lobe tissue, synthetic NPY induces a rapid, powerful and dose-dependent inhibition of in vitro release of MSH, endorphin and other proopiomelanocortin (POMC) derived peptides. Therefore, NPY undoubtedly is one of the growing number of neuropeptides that are likely involved in control of the amphibian MSH cells. Although a number of stimulatory neuropeptides have been found, this is the first neuropeptide to apparently function through an inhibitory mechanism. In that a 2-hr treatment with NPY did not influence POMC biosynthesis, nor processing of this prohormone to smaller peptides, we conclude that the primary action of NPY is a direct effect on the secretory process of the MSH cell.

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Localization and identification of immunoreactive atrial natriuretic factor (ANF) in the frog ventricle.

The localization of ANF-like immunoreactivity in the ventricle of the frog Rana ridibunda was examined by the indirect immunofluorescence and the immunogold techniques, using an antiserum against synthetic ANF (Arg 101-Tyr 126). At the optic level, an appreciable number of positive cardiocytes was observed in the frog ventricle. Electron microscopic studies showed that all secretory granules present in ventricular cardiocytes contain immunoreactive ANF. The immunoreactive material has been characterized by Sephadex G-50 gel chromatography and reverse phase high performance liquid chromatography (RP-HPLC). After gel filtration, ANF-like immunoreactivity eluted in 3 peaks. The major immunoreactive peak corresponded to high molecular weight material, while one peak co-eluted with synthetic ANF (Arg 101-Tyr 126). Further analysis of frog ventricular extracts by RP-HPLC revealed that the low molecular weight material has the same retention time as synthetic ANF, suggesting a high degree of sequence homology between amphibian and mammalian ANF. These results indicate that in amphibians, ventricular cardiocytes synthesize a peptide immunologically and chemically related to mammalian ANF.

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NPY-like peptides occur in the nervous system and midgut of the migratory locust, Locusta migratoria and in the brain of the grey fleshfly, Sarcophaga bullata.

The distribution of the NPY-like substances in the nervous system and the midgut of the migratory locust, Locusta migratoria and in the brain of the grey fleshfly, Sarcophaga bullata was determined by immunocytochemistry using an antiserum directed against synthetic porcine NPY. The peroxidase-antiperoxidase procedure revealed that NPY immunoreactive cell bodies and nerve fibers were observed in the brain, optic lobes, corpora cardiaca, suboesophageal ganglion and ventral nerve cord of the locust and in the brain, optic lobes and suboesophageal ganglion of the fleshfly. In the locust midgut, numerous endocrine cells and nerve fibers penetrating the outer musculature contained NPY-like immunoreactivity. The concentrations of NPY immunoreactive material in acetic acid extracts of locust brain, optic lobes, thoracic ganglia, ovaries and midguts was measured using a specific radioimmunoassay technique. The dilution curves of the crude tissue extracts were parallel to the standard curve. The highest amount of NPY-like immunoreactivity was found in the locust ovary and midgut. Reverse-phase high-performance liquid chromatography (RP-HPLC) and radioimmunoassay were used to characterize the NPY-like substances in the locust brain and midgut. HPLC-analysis revealed that NPY-immunoreactivity in the locust brain eluted as three separate peaks. The major peak corresponded to a peptide less hydrophobic than synthetic porcine NPY. RP-HPLC analysis of midgut extracts revealed the presence of an additional NPY-immunoreactive peak which had a retention time similar to the porcine NPY standard. The present data show the existence of a widespread network of NPY immunoreactive neurons in the nervous system of the locust and the fleshfly. Characterization of the immunoreactive substances indicates that peptides similar but not identical to porcine NPY are present in the central nervous system and midgut of insects.

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Neuronal interactions between neuropeptide Y (NPY) and catecholaminergic systems in the rat arcuate nucleus as shown by dual immunocytochemistry.

Several recent studies have suggested interactions between catecholamine (CA) and neuropeptide Y (NPY) neuronal systems in the rat brain. In order to obtain morphological evidence for such CA/NPY interactions in the arcuate nucleus, we have used a double immunostaining procedure using an anti-tyrosine hydroxylase (TH) antiserum as a marker for catecholamine neurons and an anti-NPY antiserum. This double staining, where the first staining is silver-gold intensified, was detectable at both light and electron microscopic levels. In semi-thin sections, a substantial overlap and close proximity of TH-immunopositive neurons and NPY neuronal elements could be seen within the arcuate nucleus. At the electron microscopic level, direct appositions between TH- and NPY-immunoreactive structures could be detected. These appositions were of axosomatic, axodendritic or axoaxonic types without any synaptic membrane differentiation. Moreover, direct appositions between NPY-immunoreactive structures have also been observed. This morphological study showing appositions between TH and NPY neuronal systems suggest direct interactions between these two systems in the arcuate nucleus.

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Localization of binding sites for ANF in the rat mammary gland.

The localization of ANF binding sites in the mammary gland has been performed in virgin and lactating rats using an in vitro autoradiographic procedure performed on slide-mounted frozen sections. The radioligand used was rat [125I]ANF (99-126). It was demonstrated that specific ANF binding sites were present in the secretory cells and also to a lesser extent in acinar lumen. These results suggest that ANF could play some role in the production of fluid during lactation.

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Modulation of hippocampal LHRH receptors by sex steroids in the rat.

The modulation of brain LHRH receptors by sex steroids was assessed in the female rat hippocampus using in vitro autoradiography and iodinated [D-Ser (TBU)6, des-Gly-NH2(10)]LHRH ethylamide as the radioligand. As evaluated by optical densitometry, the density of hippocampal LHRH receptors was increased by castration. In castrated animals, estradiol administration produced a small decrease in receptor concentration, whereas the concomitant administration of progesterone and estradiol as well as the injection of dihydrotestosterone induced a marked decrease in receptor density. Progesterone administration did not produce any significant change in receptor concentration. These results indicate that brain LHRH receptors can be modulated by sex steroids. Interestingly, this modulation of hippocampal LHRH receptors is similar to that previously observed in pituitary gland.

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Localization and characterization of the N-terminal fragment of atrial natriuretic factor (ANF) precursor in the frog heart.

The localization of the N-terminal fragment of the atrial natriuretic factor (ANF) precursor in the heart of the frog Rana ridibunda was examined by the indirect immunofluorescence and the immunogold techniques using an antiserum directed against synthetic rat ANF (Asp11-Ala37). At the optic level, positive material was found in most atrial myocytes. Staining of consecutive sections of frog heart with antibodies against N-terminal and C-terminal regions of the proANF molecule showed that both peptides are contained in the same cardiocytes. In the rat atrium, antibodies against the N-terminal ANF region induced a more intense labeling than in the frog atrium. Electron microscopic studies indicated that all secretory granules present in frog atrial cardiocytes contain N-terminal ANF-like immunoreactive material. The positive material localized in frog atrium was characterized by gel filtration and radioimmunological detection. Serial dilutions of frog atrial extracts exhibited displacement curves which were parallel to that obtained with synthetic human ANF (Asn1-Asp30). Sephadex G-50 gel chromatography of the immunoreactive material showed that the N-terminal ANF-like immunoreactivity eluted in a single peak corresponding to high molecular weight material. These results indicate that the N-terminal fragment of frog proANF is immunologically and biochemically related to the homologous mammalian peptide.

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Immunoelectron microscopic localization of thyrotropin-releasing hormone (TRH) precursor in the rat raphe nuclei.

Using antisera to two pro-thyrotropin-releasing hormone (pro-TRH)-derived cryptic peptides, we have studies by immunocytochemistry the ultrastructural localization of pro-TRH in the rat raphe nuclei. The same results were obtained with both antisera. Immunostaining was found in cell bodies, dendrites and endings. In cell bodies, the reaction product was restricted to Golgi saccules and dense core vesicles which were very few in number. In dendrites, the staining was rather diffuse without any association with specific organelles. These results suggest that the Golgi apparatus might be involved in pro-TRH processing.

Amino Acid Sequence↗

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Increased brain content of the endogenous benzodiazepine receptor ligand, octadecaneuropeptide (ODN), following portacaval anastomosis in the rat.

Evidence suggests that endogenous benzodiazepine receptor ligands such as diazepam binding inhibitor (DBI) and its metabolite octadecaneuropeptide (ODN) may be implicated in the pathogenesis of hepatic encephalopathy. Using an immunocytochemical technique and an antibody of high specific activity to synthetic ODN, we studied the effects of portacaval anastomosis (PCA) on ODN distribution in rat brain. Four weeks after PCA, ODN immunolabeling was increased in several brain regions including cerebral cortex, hippocampus, hypothalamus and thalamus. Increased ODN immunolabeling was confined to nonneuronal elements such as astrocytes and ependymal cells. Neuropathological evaluation of brain following PCA reveals astrocytic rather than neuronal changes. These results are consistent with a role for endogenous neuropeptide ligands for astrocytic benzodiazepine receptors in the pathogenesis of hepatic encephalopathy.

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Dihydrotestosterone (DHT) regulation of insulin-like growth factor II mRNA in neonatal rats.

Insulin-like growth factors (IGFs) are well known as peptide mitogens and important growth factors in fetal as well as in early postnatal development. In particular, IGF II is strongly expressed during fetal life and in neonatal animals. Very little is known about the regulation of IGF II gene expression. In order to study in detail the regulation of IGF II mRNA levels in the liver by the potent nonaromatizable androgen dihydrotestosterone (DHT), we have used quantitative in situ hybridization to detect the mRNA encoding for this growth factor. Pups were separated into 4 groups and injected twice a day immediately after birth with 3 different doses of DHT: 0.1 mg DHT/day, 0.25 mg DHT/day, 0.5 mg DHT/day for 4 and 7 days, and the control groups were injected with the vehicle alone. Animals were perfused with 4% paraformaldehyde and sections from the liver, heart, kidneys and brain were cut with a cryostat. A [35S]-labeled cDNA probe was used to detect IGF II mRNA levels. After hybridization, sections were autoradiographed with X-ray films and then coated with liquid photographic emulsion. Densitometric measurement revealed that, at 4 days of age, IGF II mRNA levels were lower in DHT-treated rats than in control animals. No statistically significant differences in IGF II mRNA levels were observed among the three groups treated with the different doses of DHT, thus revealing that even the lowest dose of DHT (0.1 mg/day) used was sufficient to inhibit IGF II gene expression in neonatal rats. Moreover, at 7 days of age, DHT-treated rats showed the same levels of IGF II mRNA as those observed in rats treated with DHT for 4 days. These results suggest that DHT may play an important role in the regulation of IGF II gene expression in the rat liver during the neonatal period.

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Characterization of endozepine-related peptides in the central nervous system and in peripheral tissues of the rat.

Endozepines represent a novel family of regulatory peptides that have been isolated by their ability to displace benzodiazepines from their binding sites. All endozepines derive from an 86 amino acid precursor polypeptide called diazepam binding inhibitor (DBI), which generates, through proteolytic cleavage, several biologically active endozepines. The aim of the present study was to compare the molecular forms of endozepines present in different regions of the rat brain and in various peripheral organs using an antiserum raised against the central (biologically active) region of DBI. Combination of HPLC analysis and RIA detection revealed the existence of two major forms (peaks I and II) of endozepine-immunoreactive peptides. The retention times of the two peaks (36 and 39 min, respectively) were identical in all tissues or organs tested. Western blotting analysis of cerebral cortex extracts confirmed the existence of two immunoreactive species with apparent molecular weights 4000 and 6000 Da, which respectively correspond to peaks I and II. Tryptic digestion of peaks I and II generated a single immunoreactive peptide that coeluted with the synthetic octadecaneuropeptide ODN [DBI(33-50)]. These results show that, in different parts of the brain and in various peripheral organs, DBI is rapidly processed to generate two peptides of apparent molecular weight of 4000 and 6000 Da, which both possess the biologically active determinant of endozepines.

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Neuroanatomical connections between corticotropin-releasing factor (CRF) and somatostatin (SRIF) nerve endings and thyrotropin-releasing hormone (TRH) neurons in the paraventricular nucleus of rat hypothalamus.

In order to investigate the possible involvement of corticotropin-releasing factor (CRF) and somatostatin (SRIF) on thyrotropin-releasing hormone (TRH) neuronal cell activity in the rat hypothalamic paraventricular nucleus, we have proceeded to the simultaneous localization of CRF or SRIF and TRH. For this purpose, we used a dual immunostaining procedure that employed antibodies to CRF and SRIF and peroxidase-labeled goat anti-rabbit IgG as a first sequence, and antibodies to a cryptic fragment (Phe178-Glu199) of pro-TRH (to label TRH neurons) and alkaline phosphatase-labeled goat anti-rabbit IgG as the second sequence. A rich innervation of the paraventricular nucleus by immunoreactive CRF and SRIF fibers was observed. A large number of CRF and SRIF nerve endings were seen intimate anatomic proximity and often appeared to surround TRH-containing cell bodies. These results strongly suggest that TRH neurons might be regulated by both CRF and SRIF. These interactions might be the neuroanatomical basis for the already observed inhibitory effects of CRF and SRIF on TRH release.

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[Epidermal growth factor: structure, location, phosphorylation and regulation of its receptor].

Epidermal growth factor (EGF) is a Mr 6045 polypeptide first characterized for its ability to stimulate mitogenesis in epidermal and epithelial cells. The first step in the action of the growth factor is its binding to specific, high affinity membrane receptors. These receptors have been studied in a number of tissues and cell culture lines. The level of EGF receptors is modulated by many agents. EGF down-regulates its receptor. In addition, the number of EGF receptors is decreased by other growth factors (platelet-derived growth factor; transforming growth factor), by many tumor promoters and by viral transformation. Several hormones also can regulate EGF binding in its target tissues.

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How to characterize meiotic functions in plants?

Our understanding of plant meiosis is rapidly increasing thanks to the model Arabidopsis thaliana. Here we present the results of a screening for meiotic mutants carried out with a library containing 30,719 T-DNA insertion lines. An average of one mutant per 1000 lines was recovered. Several phenotypic classes could be distinguished and are presented. In parallel, 39 proteins known to be involved in meiosis in non-plant organisms were chosen and a search was performed for homologue sequences in the completed Arabidopsis thaliana genome. Approximately 30% of the meiotic related sequences showed similarities with one or several Arabidopsis putative genes. The relevance of forward versus reverse genetics in order to characterize meiotic functions is discussed.

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Survival after a first episode of spontaneous bacterial peritonitis. Prognosis of potential candidates for orthotopic liver transplantation.

To determine the potential role of orthotopic liver transplantation (OLT) in cirrhotic patients surviving a first episode of spontaneous bacterial peritonitis (SBP), medical records of 79 patients presenting with a first episode of SBP were reviewed. Of these patients, 37 were selected as potential candidates for OLT using the following criteria: absence of hepatocellular carcinoma; no severe organ failure other than the liver; age < or = 66 years; and survival after SBP > 60 days. Survival time was calculated from the day of SBP diagnosis. Prognostic value of clinical, biological and bacteriological data recorded at the time of SBP was determined using univariate and multivariate analysis (Cox's regression model). Survival rate of the potential candidates for OLT at 3 months, 1 year and 2 years was 94, 46 and 30% respectively. Serum creatinine value (P = 0.001) and Pugh score (P = 0.005) were independently correlated with death. The 1 year survival rate was 80% for the 11 patients with a Pugh score < 10, and 26% for the 26 patients with a Pugh score > or = 10. Our results suggest that after SBP, OLT should be considered in patients with severe liver disease. Survival of patients with a moderate liver disease (i.e. Pugh score < 10) might be relatively high.

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