Catecholamine biosynthetic enzyme expression in neurological and psychiatric disorders.
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Alternative Splicing↗
Biomedical subjects
S Dumas
Publications and source records attributed to S Dumas.
[Convulsions after renal angiography and angioplasty].
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Aged↗
Plasticity of tyrosine hydroxylase gene expression in the rat nucleus tractus solitarius after ventilatory acclimatization to hypoxia.
The aim of this study was to define the influence of long-term hypoxia on gene expression of tyrosine hydroxylase (TH) in the rat nucleus tractus solitarius (NTS). Animals were exposed to normobaric hypoxia (10% O2 in nitrogen) for 2 weeks. At this time, the hypoxia-induced hyperventilation reached a plateau, indicating ventilatory acclimatization. In horizontal brainstem sections, hypoxia-induced changes in TH protein and TH mRNA were assessed by immunocytochemistry and in-situ hybridization, respectively. Long-term hypoxia increased TH mRNA levels seen as both an increase in the number of grains per cell and an extension of the labeled area. The highest degree of labeling was found selectively located in caudal NTS. Hypoxia also enhanced TH immunoreactivity in the caudal NTS but this labeling extended more rostrally than that of TH mRNA. The data suggest that there is an hypoxia-induced plasticity of gene expression at the gene level in the NTS, which is associated with ventilatory acclimatization. The hypoxia model described in this study may serve as a framework for future regulatory studies.
Animals↗
A novel rat tyrosine hydroxylase mRNA species generated by alternative splicing.
Tyrosine hydroxylase (TH) catalyzes the first and rate-limiting step in the biosynthesis of catecholamines. Among the various mechanisms implicated in the regulation of TH activity, alternative splicing of TH primary transcript has been described as a characteristic of higher primates and Drosophila. We investigated whether there is such a regulatory mechanism in the rat. Reverse transcriptase-PCR experiments were performed with RNA from PC12 cells. A new TH mRNA species was evidenced, resulting from the use of an alternative donor site in exon 2. RNase protection assays and in situ hybridization experiments detected this mRNA species in the adrenal medulla but not in the main catecholaminergic nuclei of the CNS. The corresponding putative protein lacks 33 amino acids in the N-terminal regulatory domain. A recombinant protein was produced in E. coli. Its in vitro specific activity was similar to that of the previously identified TH protein.
Adrenal Medulla↗
New species of human tyrosine hydroxylase mRNA are produced in variable amounts in adrenal medulla and are overexpressed in progressive supranuclear palsy.
Alternative splicing of human tyrosine hydroxylase (TH) pre-mRNA produces four mRNAs leading to four different TH isoforms and is thought to have important regulatory functions. We show that the diversity of TH mRNAs is greater than previously described in the autonomous nervous system: New splice junctions corresponding to the skipping of exon 3 were identified by amplification of cDNA synthesized from pheochromocytoma RNA. In all cases the reading frame was maintained. These species were assayed by RNase protection experiments; their abundance (4-6%) was comparable to that of the previously identified human TH-3 and -4 species in normal adrenal medulla. However, higher levels (11-34%) of these species were found in adrenal medullas of patients suffering from progressive supranuclear palsy. Whether such changes are specific to the disease or the consequences of the stress associated with this severe neurodegeneration remains to be established.
Adrenal Medulla↗
HRRI: a high resolution radioimager for fast, direct quantification in in situ hybridization experiments.
We present a high-speed, high-resolution beta imager. It has been developed to be used in in situ hybridization experiments, either instead of or in complement with autoradiographic film and emulsions that are currently used for these experiments. It allows the user to locate and perform quantitative analyses of (3H-, 14C-, 35S-, 32P-, 125I-) labeled molecules with a 15-microns spatial resolution on a 1.2 cm2 area. We have combined recent techniques (specific scintillator thin sheets and intensified charge-coupled device [CCD]) so that this imager offers a wide dynamic range and real-time acquisition. Several biological applications will be discussed.
Animals↗
An efficient method to detect calcitonin mRNA in normal and neoplastic rat C-cells (medullary thyroid carcinoma) by in situ hybridization using a digoxigenin-labeled synthetic oligodeoxyribonucleotide probe.
We report here an efficient and rapid method for the specific detection of calcitonin in tumor C-cells of medullary thyroid carcinoma (MTC). This occasionally aggressive tumor arises from the endocrine thyroid C-cells. Its principal marker is calcitonin, the predominant C-cell secretion, which is detected in patients and in our animal model by radioimmunoassay of the plasma, as well as by immunohistochemistry of thyroid tissues. Although calcitonin is easily detectable in normal C-cells, its content is greatly reduced in tumor cells owing to the disappearance of the secretory granules that store the mature peptide. This finding suggests cell dedifferentiation correlated with an increasing aggressivity of the tumor. We therefore developed a rapid detection of calcitonin mRNA by in situ hybridization on routine paraffin sections, using a synthetic oligodeoxyribonucleotide probe labeled with digoxigenin-dUTP. The reaction was detected with an anti-digoxigenin antibody conjugated with alkaline phosphatase, and the enzyme catalyzed the appearance of a dark blue color. The signal was exclusively restricted to the normal, hyperplastic, and tumor C-cells. It was specific, as increasing concentrations of the unlabeled oligonucleotide led to progressive disappearance of the reaction. Its sensitivity was slightly diminished as compared with corresponding frozen sections, but the intensity of the signal was quite acceptable. High levels of calcitonin mRNA were found in all normal and hyperplastic C-cells. They were increased in most of the tumor MTC cells, which did not correlate with the amount of intracellular peptide stores but explained the abnormally high basal levels of circulating calcitonin of the tumor-bearing rats. ISH is therefore of greater value than ICC for an early anatomopathological detection of this tumor. Our data show that the tumor cells are not "dedifferentiated." They only lack the granular compartment storing the mature peptide before exocytosis, but CT biosynthesis and the rest of the secretory process seem to be complete. Our results suggest that factors expressed in malignant C-cells affect basic cell mechanisms involved in the storage of the mature calcitonin, rather than the expression of the CALC gene.
Animals↗
Behavioral sensitization induced by psychostimulants or stress: search for a molecular basis and evidence for a CRF-dependent phenomenon.
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Amphetamine↗
Tyrosine hydroxylase gene expression in human ventral mesencephalon: detection of tyrosine hydroxylase messenger RNA in neurites.
The distribution of both tyrosine hydroxylase (TH) antigen and messenger RNA was investigated in the ventral mesencephalon from human brain using a 35S-labeled human TH cDNA probe and a polyclonal TH antibody. A hybridization signal was detected not only in the perikaryon but also in the neurites, most probably dendrites, of immunoreactive neurons. In cell bodies, the level of TH mRNA varies, revealing a regional heterogeneity. In the substantia nigra of patients with Parkinson's disease the number of labeled cells was markedly reduced as compared to controls.
Aged↗
Neurons containing messenger RNA encoding glutamate decarboxylase in rat hypothalamus demonstrated by in situ hybridization, with special emphasis on cell groups in medial preoptic area, anterior hypothalamic area and dorsomedial hypothalamic nucleus.
Previous deafferentation studies have suggested that most hypothalamic GABAergic innervation originates from neurons within the hypothalamus. We have investigated the distribution of GABAergic cell groups in the rat hypothalamus by means of the in situ hybridization technique, using a cDNA probe for messenger RNA encoding glutamate decarboxylase. Several major GABAergic cell groups were demonstrated, including cells of the tuberomammillary nucleus, arcuate nucleus, suprachiasmatic nucleus, medial preoptic area, anterior hypothalamic area, the dorsomedial hypothalamic nucleus, perifornical area, and lateral hypothalamic area. The most prominent glutamate decarboxylase mRNA-containing cell groups were located in the medial preoptic area, anterior hypothalamic area and dorsomedial hypothalamic nucleus, and were composed of small- to medium-sized neurons. Compared to previously well-characterized GABAergic cell groups in the tuberomammillary nucleus, reticular thalamic nucleus, and non-pyramidal cells of cerebral cortex, the cells of these GABAergic groups demonstrated only weak cDNA labelling, indicating that they contain lower levels of glutamate decarboxylase mRNA. Several types of control experiments supported the specificity of this cDNA labelling, and the GABAergic nature of these cell populations was further supported by detection of glutamate decarboxylase and GABA immunoreactivity. Abundance of GABAergic cells in many hypothalamic nuclei indicates that GABA represents quantitatively the most important transmitter of hypothalamic neurons, and may be involved in neuroendocrine and autonomic regulatory functions.
Animals↗
Decreased tyrosine hydroxylase messenger RNA in the surviving dopamine neurons of the substantia nigra in Parkinson's disease: an in situ hybridization study.
Parkinson's disease is associated with a massive loss in melanized dopaminergic neurons in the substantia nigra resulting in a severe striatal dopaminergic denervation. The hyperactivity which develops in the remaining striatal dopaminergic terminals may be related to an increased rate of tyrosine hydroxylation. This could be related to changes in the level of expression of the gene coding for tyrosine hydroxylase. Thus, the detection of tyrosine hydroxylase messenger RNA was looked for at cellular levels by in situ hybridization histochemistry. Image analysis shows that the hybridization signal was significantly reduced in the surviving neurons when compared to control. The subnormal tyrosine hydroxylase messenger RNA content may express a change in level of tyrosine hydroxylase gene transcription, possibly in relation to sustained suffering of the neurons still present at late stages of the disease.
Aged↗
Open trial of a calcium antagonist, nimodipine, in acute mania.
Following previous works showing an action of calcium channel blockers in mania, this trial explores the clinical efficiency of another anticalcic agent, nimodipine, which has been selected for its specific action on the CNS. Our design has been a 7-day controlled open trial. We included six inpatients diagnosed according to DSM III-R criteria as having mild to acute mania. Evaluations have been made by clinical scales, electroencephalography, electrocardiography, and extensive biological blood tests. No other medication than nimodipine was allowed excepted i.m. droperidol in case of severe excitement. We used nimodipine at a dose of 360 mg/day. The results showed good clinical efficiency in each of our six inpatients, with an important and significant improvement of scales scores. We noticed an early and significant action on mood without important sedative effect. The treatment has been well tolerated, without biological effects. Further studies are needed, but nimodipine could be an alternative treatment to the neuroleptics with a different type of action, less sedative and more specific on mood.
Acute Disease↗
Localization of copper-zinc superoxide dismutase mRNA in human hippocampus by in situ hybridization.
The cellular localization of copper-zinc superoxide dismutase (CuZn SOD) mRNA was determined in the human hippocampus by in situ hybridization with a 35S-labelled DNA probe complementary to human CuZn SOD mRNA. A positive hybridization signal was detected in pyramidal cell layers CA1-CA4 of Ammon's horn (CA), pyramidal cells of subiculum and in the granule cells of the dentate gyrus. The fact that CuZn SOD gene expression is important in neurones which are preferentially vulnerable in neurodegenerative processes such as Alzheimer's disease, suggests a role played by oxygen free radicals in the mechanism of nerve cell death.
Aged↗
Expression of carbonic anhydrase II gene in early brain cells as revealed by in situ hybridization and immunohistochemistry.
A mouse carbonic anhydrase (CA II) complementary(c) DNA probe was used for in situ hybridization on mouse brain cultured cells in order to follow CA II gene expression during brain development. An improved method was established using biotinated probes that resulted in a high sensitivity and an absence of background; this method could be combined with immunohistochemistry. Hypothalamic cells of embryonic day (ED) 12-14 mice were cultured for various periods. Chronologic appearance of CA II messenger(m)RNA and protein was studied. The CA II gene transcripts are detectable as early as ED 12-13, although the protein they encode is not detectable until ED 17-18. Gene expression is restricted to 0.1% of the total population. Northern blot analysis confirmed the presence of CA II transcripts in embryonic hypothalamus. At postnatal stage, the majority of glial cells express both the CA II mRNA and the protein. Our results favour the early appearance of a glial lineage in a precise area of the developing CNS. The precocity of CA II gene transcription makes in situ hybridization an invaluable approach in defining the onset of nerve cell lineages during embryonic development.
Animals↗
Differential control of tryptophan hydroxylase expression in raphe and in pineal gland: evidence for a role of translation efficiency.
In a previous study, we characterized two tryptophan hydroxylase mRNAs (TPH mRNAs) in the pineal gland. However, we failed to detect these species in the raphe by Northern blot experiments. Here, we report by S1 nuclease analysis and in situ hybridization that these two TPH mRNAs, as well as a third species, are expressed both in pineal gland and in raphe. In both tissues, the three mRNAs are transcribed predominantly from the same promoter. Strikingly, from the results of S1 maping analysis, it was observed that the total level of TPH mRNA per tissue is at least 150 times lower in the raphe than in the pineal gland. In contrast, TPH antigen as quantified by immunoblot experiments is about threefold more abundant per raphe than per pineal gland. TPH mRNA from one raphe and one pineal gland yield in vitro about the same amount of TPH antigen, suggesting that the discrepancy in the ratios of TPH mRNA and TPH antigen between the raphe and the pineal gland results, at least in part, from a difference in the translation efficiency of TPH mRNAs in the two structures.
Animals↗
Analysis of the 5' region of the human tyrosine hydroxylase gene: combinatorial patterns of exon splicing generate multiple regulated tyrosine hydroxylase isoforms.
A single human gene has been described to encode multiple tyrosine hydroxylase (TH) mRNAs. The study of this variation has been extended by S1 mapping experiments and by analysis of the 5' region of the TH gene. Four different mRNAs were found to originate solely from alternative splicing of two exons. Comparison of the 5' flanking regions of human and rat genes discloses several highly conserved segments, likely to play an important role in the regulation of TH gene expression.
Base Sequence↗
Molecular cloning, expression and in situ hybridization of rat brain glutamic acid decarboxylase messenger RNA.
A cDNA library was generated in the expression vector lambda GT11 from rat brain poly(A)+ RNAs and screened with a GAD antiserum. Two clones reacted positively. One of them was shown to express a GAD activity which was specifically trapped on anti-GAD immunogel and was inhibited by gamma-acetylenic-GABA. Blot hybridization analysis of RNAs from rat brain revealed a single 4 kilobases band. Preliminary in situ hybridizations showed numerous cells labelled by the GAD probe such as the Purkinje and stellate cells in the cerebellar cortex and the cells of the reticular thalamic nucleus.
Animals↗
Molecular genetics of catecholamines as an approach to the biochemistry of manic-depression.
Manic depressive illness has been clearly established to exhibit a strong genetic component and is therefore amenable to linkage analysis using random DNA markers. In view of the catecholamine hypothesis of this disorder, the gene encoding tyrosine hydroxylase (TH) the limiting enzyme in catecholamines is a good candidate to investigate. This gene has been localized to chromosome 11 in close linkage with Harvey-ras-1. The various transcriptional and post-transcriptional mechanisms that modulate short and long-term TH activity are discussed. Human tyrosine hydroxylase is coded by at least three distinct mRNAs derived from a single gene. This variation has clear functional consequences and could represent a novel mode of regulating catecholamines levels in normal and pathological neurons.
Bipolar Disorder↗