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1.
Dev Dyn ; 238(10): 2641-51, 2009 Oct.
Article in English | MEDLINE | ID: mdl-19718764

ABSTRACT

Early expression of estrogen receptors (esr) and their role in regulating early expression of cyp19a1b encoding brain aromatase were examined in the brain of zebrafish. Using in toto hybridization and quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), a significant increase in the expression of esr1, esr2a, and esr2b was observed between 24 and 48 hours postfertilization (hpf). In toto hybridization demonstrated that esr2a and esr2b, but not esr1, are found in the hypothalamus. Using real-time RT-PCR, an increase in cyp19a1b mRNAs occurs between 24 and 48 hpf, indicating that expression of cyp19a1b is temporally correlated with that of esr. This increase is blocked by the pure anti-estrogen ICI182,780. Furthermore, E2 treatment of cyp19a1b-GFP (green fluorescent protein) transgenic embryos results in appearance of GFP expression in the brain as early as 25 hpf. These results indicate that basal expression of cyp19a1b expression in the brain of developing zebrafish most likely relies upon expression of esr that are fully functional before 25 hpf.


Subject(s)
Aromatase/metabolism , Brain , Embryo, Nonmammalian , Gene Expression Regulation, Developmental , Receptors, Estrogen/metabolism , Zebrafish Proteins/metabolism , Zebrafish , Animals , Animals, Genetically Modified , Aromatase/genetics , Brain/embryology , Brain/enzymology , Embryo, Nonmammalian/anatomy & histology , Embryo, Nonmammalian/enzymology , Estradiol/analogs & derivatives , Estradiol/metabolism , Estrogen Antagonists/metabolism , Fulvestrant , Protein Isoforms/genetics , Protein Isoforms/metabolism , Receptors, Estrogen/genetics , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Zebrafish/anatomy & histology , Zebrafish/embryology , Zebrafish/metabolism , Zebrafish Proteins/genetics
2.
Brain Res Bull ; 75(2-4): 274-80, 2008 Mar 18.
Article in English | MEDLINE | ID: mdl-18331884

ABSTRACT

In contrast to other vertebrates, in which the adult brain shows limited adult neurogenesis, teleost fishes exhibit an unparalleled capacity to generate new neurons as adults, suggesting that their brains present a highly permissive environment for the maintenance and proliferation of adult progenitors. Here, we examine the hypothesis that one of the factors permitting establishment of this favourable environment is estradiol. Indeed, recent data showed that radial glial cells strongly expressed one of two aromatase duplicated genes. Aromatase is the estrogen-synthesizing enzyme and this observation is of great interest, given that radial glial cells are progenitor cells capable of generating new neurons. Given the well-documented roles of estrogens on cell fate, and notably on cell proliferation, these data suggest that estradiol could be involved in maintaining and/or activating these progenitors. Examination of recent data in birds and mammals suggests that the situation in fish could well be an exaggeration of a more general mechanism implicating estrogens in neurogenesis. Indeed, there is accumulating evidence that estrogens are involved in embryonic, adult or reparative neurogenesis in other vertebrates, notably in mammals.


Subject(s)
Brain/cytology , Estrogens/biosynthesis , Fishes/anatomy & histology , Stem Cells/metabolism , Animals , Biological Evolution , Cell Proliferation , Neurons/physiology
3.
Biol Reprod ; 65(5): 1548-57, 2001 Nov.
Article in English | MEDLINE | ID: mdl-11673274

ABSTRACT

In oviparous species, in addition to a full-length estrogen receptor alpha (ER alpha), another ER alpha isoform lacking the A domain and exhibiting a ligand-independent transactivation function has been consistently reported. Although both isoforms are expressed in the liver, their respective sites of expression in other potential target tissues are unknown. In contrast to the situation in Xenopus and chicken, the two isoforms of rainbow trout (Oncorhynchus mykiss) are generated from two classes of transcripts with different 5' untranslated sequences issued from the same gene by alternative splicing and promoter usage. The aim of this study was to take advantage of the unique organization of the rainbow trout ER alpha gene to investigate the tissue distribution of these two messenger species along the reproductive axis of female trout. The S1 nuclease assay and in situ hybridization were used, with probes specific for each of the transcripts. Reverse transcription polymerase chain reaction (RT-PCR) with primers specific for each of the isoforms also was performed. The data indicated that the full-length ER alpha is expressed in liver, brain, pituitary, and ovary, whereas expression of the isoform with the truncated A domain is restricted to the liver, demonstrating a tissue-specific expression of these two ER alpha isoforms. The presence of a short liver-specific isoform in oviparous species suggests its role in the development and/or maintenance of the unique function of the liver in the vitellogenesis process.


Subject(s)
Gene Expression , Oncorhynchus mykiss , Organ Specificity , Receptors, Estrogen/genetics , Animals , Autoradiography , Estrogen Receptor alpha , Female , In Situ Hybridization , Liver/chemistry , Male , Ovary/chemistry , Pituitary Gland/chemistry , Prosencephalon/chemistry , Protein Isoforms/genetics , RNA, Messenger/analysis , Reverse Transcriptase Polymerase Chain Reaction , Single-Strand Specific DNA and RNA Endonucleases
4.
J Comp Neurol ; 429(1): 144-55, 2001 Jan 01.
Article in English | MEDLINE | ID: mdl-11086295

ABSTRACT

The expression sites of three prepro-gonadotrophin-releasing hormones (GnRHs), corresponding to seabream GnRH (sbGnRH: Ser(8)-mGnRH, mammalian GnRH), salmon GnRH (sGnRH: Trp(7)Leu(8)-mGnRH), and chicken GnRH-II (cGnRH-II: His(5)Trp(7)Tyr(8)-mGnRH) forms were studied in the brain of a perciform fish, the European sea bass (Dicentrarchus labrax) by means of in situ hybridization. The riboprobes used in this study correspond to the three GnRH-associated peptide (GAP)-coding regions of the prepro-GnRH cDNAs cloned from the same species (salmon GAP: sGAP; seabream GAP: sbGAP; chicken GAP-II: cIIGAP), which show little oligonucleotide sequence identity (sGAP versus sbGAP: 42%; cIIGAP versus sbGAP: 36%; sGAP versus cIIGAP: 41%). Adjacent paraffin sections (6 mm) throughout the entire brain were treated in parallel with each of the three anti-sense probes and the corresponding sense probes, demonstrating the high specificity of the hybridization signal. The results showed that both sGAP and sbGAP mRNAs had a broader expression in the olfactory bulbs, ventral telencephalon, and preoptic region, whereas cIIGAP mRNA expression was confined to large cells of the nucleus of the medial longitudinal fascicle. In the olfactory bulbs, both the signal intensity and the number of positive cells were higher with the sGAP probe, whereas sbGAP mRNA-expressing cells were more numerous and intensely stained in the preoptic region. Additional isolated sbGAP-positive cells were detected in the ventrolateral hypothalamus. These results demonstrate a clear overlapping of sGAP- and sbGAP-expressing cells in the forebrain of the European sea bass, in contrast to previous reports in other perciforms showing a clear segregation of these two cell populations.


Subject(s)
Bass/metabolism , Brain/metabolism , Gonadotropin-Releasing Hormone/genetics , Neurons/metabolism , Protein Precursors/genetics , Animals , Bass/anatomy & histology , Brain/anatomy & histology , Chickens/anatomy & histology , Chickens/metabolism , Female , Gonadotropin-Releasing Hormone/chemistry , Gonadotropin-Releasing Hormone/metabolism , Hypothalamus/cytology , Hypothalamus/metabolism , Male , Neurons/cytology , Olfactory Bulb/cytology , Olfactory Bulb/metabolism , Protein Precursors/chemistry , Protein Precursors/metabolism , RNA, Messenger/metabolism , Salmon/anatomy & histology , Salmon/metabolism , Sea Bream/anatomy & histology , Sea Bream/metabolism , Telencephalon/cytology , Telencephalon/metabolism
5.
Biol Reprod ; 63(6): 1857-66, 2000 Dec.
Article in English | MEDLINE | ID: mdl-11090458

ABSTRACT

A full-length cDNA encoding a GnRH receptor (GnRH-R) has been obtained from the brain of rainbow trout. This cDNA encodes a protein of 386 amino acids (aa) exhibiting the typical arrangement of the G-protein-coupled receptors in seven transmembrane domains. However, a second ATG could give rise to a receptor with a 30-aa longer extracellular domain. As already shown in other fish and Xenopus, this protein possesses an intracellular domain, in contrast with its mammalian counterparts. In the case of rainbow trout, this intracellular carboxy-terminal tail consists of 58 residues. Northern blotting experiments carried out in the brain, the pituitary, and the liver only resulted in a single band of 1.9-2 kilobases in the pituitary, although reverse transcription-polymerase chain reaction amplification products were found in the brain, the pituitary, the retina, and the ovary. In situ hybridization using a probe corresponding to the full-length coding region of the receptor was performed on vitellogenic or ovulating females and allowed to detect a weak but specific signal in the proximal pars distalis of the pituitary, the preoptic region, the mediobasal hypothalamus, and the optic tectum. However, the strongest signal was consistently detected in a mesencephalic structure, the nucleus lateralis valvulae, the significance of which is presently open to speculation.


Subject(s)
Oncorhynchus mykiss/genetics , Receptors, LHRH/biosynthesis , Receptors, LHRH/genetics , Amino Acid Sequence , Animals , Base Sequence , Blotting, Northern , Blotting, Southern , Cloning, Molecular , DNA, Complementary/biosynthesis , DNA, Complementary/genetics , Female , In Situ Hybridization , Molecular Sequence Data , Phylogeny , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction , Tissue Distribution
6.
Mol Endocrinol ; 14(10): 1627-48, 2000 Oct.
Article in English | MEDLINE | ID: mdl-11043578

ABSTRACT

LIM/Homeodomain (HD) proteins are classically considered as major transcriptional regulators which, in cooperation with other transcription factors, play critical roles in the developing nervous system. Among LIM/HD proteins, Islet-1 (ISL1) is the earliest known marker of motoneuron differentiation and has been extensively studied in this context. However, ISL1 expression is not restricted to developing motoneurons. In both embryonic and adult central nervous system of rodent and fish, ISL1 is found in discrete brain areas known to express the estrogen receptor (ER). These observations led us to postulate the possible involvement of ISL1 in the control of brain functions by steroid hormones. Dual immunohistochemistry for ISL1 and ER provided evidence for ISL1-ER coexpression by the same neuronal subpopulation within the rat hypothalamic arcuate nucleus. The relationship between ER and ISL1 was further analyzed at the molecular level and we could show that 1) ISL1 directly interacts in vivo and in vitro with the rat ER, as well as with various other nuclear receptors; 2) ISL1-ER interaction is mediated, at least in part, by the ligand binding domain of ER and is significantly strengthened by estradiol; 3) as a consequence, ISL1 prevents ER dimerization in solution, thus leading to a strong and specific inhibition of ER DNA binding activity; 4) ISL1, via its N-terminal LIM domains, specifically inhibits the ER-driven transcriptional activation in some promoter contexts, while ER can serve as a coactivator for ISL1 in other promoter contexts. Taken together, these data suggest that ISL1-ER cross-talk could differentially regulate the expression of ER and ISL1 target genes.


Subject(s)
Homeodomain Proteins/pharmacology , Nerve Tissue Proteins , Receptors, Estrogen/drug effects , Receptors, Estrogen/physiology , Animals , Arcuate Nucleus of Hypothalamus/chemistry , Binding Sites , Brain Chemistry , CHO Cells , Cricetinae , DNA/metabolism , Dimerization , Estradiol/pharmacology , Female , Fluorescent Antibody Technique , Homeodomain Proteins/analysis , Immunohistochemistry , LIM-Homeodomain Proteins , Neurons/chemistry , Nuclear Proteins/metabolism , Promoter Regions, Genetic , Rats , Rats, Wistar , Receptors, Estrogen/analysis , Transcription Factors , Transcriptional Activation/drug effects
7.
J Chem Neuroanat ; 19(4): 197-210, 2000 Sep.
Article in English | MEDLINE | ID: mdl-11036237

ABSTRACT

The distribution of neuropeptide Y (NPY) gene expression was mapped in the brain of the sea bass (Dicentrarchus labrax) by in situ hybridization with 35S-UTP labeled cRNA probes. Gene expression was mainly detected within the forebrain, although NPY mRNA transcripts were also localized in the tectum and tegmentum mesencephali and posterior brain. New NPY-expressing nuclei were found in the dorsal and ventral telencephalon, preoptic area, tuberal hypothalamus, synencephalon, tegmentum mesencephali and posterior brain. The profuse NPY gene expression within the main neuroendocrine areas of the teleost fish further supports a physiological role in the control of the pituitary secretion. In addition, NPY gene was expressed within the primary visual, olfactory and gustatory circuits of teleost which, subsequently, project to hypothalamic feeding center in teleost fish. Our results extend the NPY-expressing areas known in teleost species.


Subject(s)
Bass/metabolism , Brain Mapping , Brain/metabolism , Gene Expression/physiology , Neuropeptide Y/metabolism , Animals , Female , RNA, Messenger/metabolism
8.
J Comp Neurol ; 426(2): 197-208, 2000 Oct 16.
Article in English | MEDLINE | ID: mdl-10982463

ABSTRACT

Tetrapod vertebrates express three neuropeptide Y (NPY)-related peptides: NPY, peptide YY (PYY), and pancreatic polypeptide (PP). Both NPY and PYY mRNA have been localized in the brain of tetrapods whereas PP expression is restricted to the pancreas. Some teleost fish commonly produce NPY and PYY but pancreatic peptide Y (PY) instead of PP. Both NPY and PYY mRNAs are widely distributed in the brain of non-tetrapod species, but no information about PY central expression is available. In the present study, molecular riboprobes were used to study PYY and PY mRNA central distribution in the sea bass (Dicentrarchus labrax). PYY and PY gene expression was predominantly detected within the sea bass forebrain. Telencephalic PYY gene expression was restricted to the ventral part of the ventral telencephalon, and no PY expression was detected in the cerebral hemispheres. Both PYY and PY mRNAs were found within the preoptic area and lateral hypothalamus. Distinct PY or PYY mRNA cell groups were localized in the pretectal area and synencephalon or posterior tubercle, respectively. Caudally, PY gene expression was found in the medial reticular formation, whereas PYY transcripts were localized within the vagal lobe. The results demonstrate that vertebrate brain expresses three NPY-related genes and further support the hypothesis that PP and PY arose by independent gene duplications from PYY. The receptor system of the NPY family as well as gene expression within the main hypophysiotropic and feeding behavior areas suggest an involvement of both peptides in the control of food intake and pituitary secretion.


Subject(s)
Bass/metabolism , Brain/metabolism , Neuropeptide Y/genetics , Peptide YY/metabolism , Amino Acid Sequence , Animals , Brain/cytology , Female , In Situ Hybridization , Molecular Sequence Data , Neurons/metabolism , Neuropeptide Y/metabolism , Peptide YY/genetics , RNA, Messenger/metabolism , Tissue Distribution
9.
J Comp Neurol ; 422(4): 612-20, 2000 Jul 10.
Article in English | MEDLINE | ID: mdl-10861529

ABSTRACT

To identify brain structures potentially acting as biological clocks in rainbow trout (Oncorhynchus mykiss), the expression sites of a trout homolog of the mouse clock gene were studied and compared with that of melatonin receptors (Mel-R). For this purpose, a partial sequence of the trout clock gene, including a PAS domain, was obtained by reverse transcription-polymerase chain reaction and used to perform in situ hybridization. The highest density of clock transcripts was observed in the periventricular layer (SPV) of the optic tectum, but a weaker expression was detected in some pretectal nuclei, such as the posterior pretectal nucleus (PO) and the periventricular regions of the diencephalon. Comparison of the hybridization signal in fish sacrificed at 08:00 and 17:00 did not indicate major changes in clock expression levels. Comparison of adjacent sections alternatively treated with clock and Mel-R probes suggests that both messengers are probably expressed in the same cells in the SPV and PO. In addition, in situ hybridization with a glutamate decarboxylase 65 probe, demonstrates that cells expressing clock and Mel-R in the optic tectum are gamma-aminobutyric acid neurons. The tight overlapping between the expression of Mel-R and clock transcripts in cells of the PO and SPV suggests a functional link between these two factors. These results indicate that the optic tectum and the pretectal area of the rainbow trout are major sites of integration of the melatonin signal, express the clock gene, and may act as biological clocks to influence behavioral and endocrine responses in trout.


Subject(s)
Biological Clocks/physiology , Oncorhynchus mykiss/metabolism , Receptors, Cell Surface/metabolism , Receptors, Cytoplasmic and Nuclear/metabolism , Superior Colliculi/metabolism , Trans-Activators/metabolism , Amino Acid Sequence , Animals , Base Sequence , CLOCK Proteins , Molecular Sequence Data , Receptors, Melatonin , Trans-Activators/chemistry
10.
J Comp Neurol ; 410(2): 277-89, 1999 Jul 26.
Article in English | MEDLINE | ID: mdl-10414533

ABSTRACT

By using degenerate primers designed from glutamate decarboxylase (GAD) sequences of mammals, Xenopus and Drosophila, a 270-bp cDNA fragment was cloned by reverse transcriptase-polymerase chain reaction (RT-PCR) from cerebellum total RNA of rainbow trout. This partial cDNA shows 90% identity with mammalian GAD 65 and presents the Asn-Pro-His-Lys (NPHK) sequence corresponding to the pyridoxal-binding region of porcine DOPA decarboxylase or mammalian GAD. The distribution of GAD 65 mRNA-expressing neurons in the forebrain of the trout was studied by in situ hybridization using either digoxigenin- or 35S-labeled probes. The results demonstrate that gamma-amino butyric acid (GABA) neurons are widely distributed throughout the forebrain, with a high density in the periventricular regions. In this study, we report their precise distribution in the telencephalon and diencephalon. GAD mRNA-expressing cells were particularly abundant in the preoptic region and the mediobasal hypothalamus, two major neuroendocrine and estrogen-sensitive regions in fish. The presence of GAD mRNA-expressing neurons was observed in visually related structures such as the suprachiasmatic nucleus, the pretectal region, and the thalamus. Immunohistochemistry with antibodies directed against mouse GAD failed to demonstrate the presence of immunoreactive cell bodies, but showed a very high concentration of GAD-immunoreactive fibers in many brain regions, notably in the preoptic area, hypothalamus, and neurohypophyseal digitations of the pituitary, in particular in the proximal pars distalis. These results indicate that GABA neurons are ideally placed to modulate neuroendocrine activities at the hypothalamic and pituitary levels and to participate in the processing of sensorial information.


Subject(s)
Glutamate Decarboxylase/genetics , Glutamate Decarboxylase/metabolism , Prosencephalon/enzymology , RNA, Messenger/metabolism , Animals , Binding Sites , Brain Stem/cytology , Brain Stem/enzymology , Cerebellum/cytology , Cerebellum/enzymology , Culture Techniques , DNA, Complementary/genetics , Dopa Decarboxylase/metabolism , Female , Gene Expression/genetics , Humans , Hypothalamus/cytology , Hypothalamus/enzymology , Immunohistochemistry , In Situ Hybridization , Neurons/enzymology , Oncorhynchus mykiss/genetics , Prosencephalon/cytology , gamma-Aminobutyric Acid/metabolism
11.
J Comp Neurol ; 409(2): 313-24, 1999 Jun 28.
Article in English | MEDLINE | ID: mdl-10379923

ABSTRACT

To better define the role of melatonin in fish, we have compared in detail the distribution of 2-[125I]iodomelatonin binding sites with gene expression for melatonin receptor subtypes in a widely studied seasonal species, the rainbow trout. Three distinct partial sequences of the melatonin receptor gene were cloned from trout genomic DNA. Two of the sequences corresponded to the Mella receptor subtype, and one corresponded to the Mellb receptor subtype. Analysis of numerous clones failed to find a sequence equivalent to the Mel1c receptor subtype. Comparison of receptor gene expression with 2-[125I]iodomelatonin binding distribution indicated dendritic transport of the receptor. Melatonin receptors were associated predominantly with visually related areas of the trout brain, such as the thalamic region, the pretectal area, and the optic tectum. The pituitary was devoid of 2-[125I]iodomelatonin binding, and melatonin receptor gene expression was not detectable. It would appear from the results of the present study that melatonin in this species is involved primarily in the processing of visual signals. How melatonin interacts with circannual rhythms of growth and reproduction is unclear, although a direct interaction between melatonin and the hypothalamo-pituitary axis is not clearly indicated.


Subject(s)
Brain Chemistry/physiology , Oncorhynchus mykiss/physiology , Receptors, Cell Surface/genetics , Receptors, Cytoplasmic and Nuclear/genetics , Animals , Autoradiography , DNA Primers , Evolution, Molecular , Gene Expression/physiology , In Situ Hybridization , Iodine Radioisotopes , Ligands , Molecular Sequence Data , Photoperiod , Phylogeny , Polymerase Chain Reaction , Protein Binding/physiology , RNA, Messenger/analysis , Radioligand Assay , Receptors, Cell Surface/analysis , Receptors, Cell Surface/metabolism , Receptors, Cytoplasmic and Nuclear/analysis , Receptors, Cytoplasmic and Nuclear/metabolism , Receptors, Melatonin , Reproduction/physiology , Sequence Homology, Amino Acid , Vision, Ocular/physiology
12.
Neuroendocrinology ; 69(4): 269-80, 1999 Apr.
Article in English | MEDLINE | ID: mdl-10207279

ABSTRACT

The potential role of the neurotransmitter gamma-aminobutyric acid (GABA) in the control of the secretion of the two pituitary fish gonadotropins (GTH-1 and GTH-2) was investigated in male and female rainbow trout (Oncorhynchus mykiss). The presence of glutamate decarboxylase-positive fibers in the neurohypophyseal digitations adjacent to the gonadotropic cells was demonstrated by means of double immunohistochemistry, providing a morphofunctional support for potential GABA-gonadotropin interactions in both sexes. In spermiating males, in vivo treatment with GABA did not affect basal gonadotropin release, but stimulated GTH-1 release when coadministered with a gonadotropin-releasing hormone analogue (GnRHa), and potentiated GnRHa-stimulated GTH-2 release. In vitro, using dispersed pituitary cells, GABA stimulated basal GTH-1 and GTH-2 secretion, in a dose-dependent manner, and potentiated salmon GnRH effect on both hormones. In mature females, GABA induced in vivo a strong elevation of plasma GTH-2 levels after 2- 6 h of injection, but had no effect in vitro. GABA treatment in vivo was also stimulatory in recrudescent females, slightly increasing plasma GTH-2 levels in both saline- and GnRHa-treated fish (GnRHa alone has no effect at this stage). Immature fish were unresponsive to GABA/GnRHa treatments but, after steroid implantation [testosterone (T) or estradiol] for 13 days, injection of GABA stimulated GTH-2 release in vivo (also GTH-1 slightly in T-implanted fish). In conclusion, GABA has an overall stimulatory action on GTH-1 and GTH-2 secretion in rainbow trout, which depends on the sex and the reproductive stage of the fish. The stimulatory action of GABA might be exerted, at least in part, directly onto the gonadotropes, as it stimulates basal and GnRH-induced GTH-1 and GTH-2 secretion from dispersed pituitary cells.


Subject(s)
Gonadotropins, Pituitary/metabolism , Oncorhynchus mykiss/physiology , Pituitary Gland/metabolism , gamma-Aminobutyric Acid/physiology , Animals , Cells, Cultured , Drug Implants , Estradiol/administration & dosage , Estradiol/pharmacology , Female , Glutamate Decarboxylase/analysis , Gonadotropin-Releasing Hormone/analogs & derivatives , Gonadotropin-Releasing Hormone/pharmacology , Immunohistochemistry , Male , Pituitary Gland/drug effects , Pituitary Gland, Posterior/enzymology , Testosterone/administration & dosage , Testosterone/pharmacology , Vigabatrin/pharmacology , gamma-Aminobutyric Acid/administration & dosage , gamma-Aminobutyric Acid/pharmacology
13.
Biol Reprod ; 60(3): 642-50, 1999 Mar.
Article in English | MEDLINE | ID: mdl-10026111

ABSTRACT

In order to identify the nature of the glucocorticoid receptor (GR)-expressing neurons and pituitary cells that potentially mediate the negative effects of stress on reproductive performance, double immunohistochemical stainings were performed in the brain and pituitary of the rainbow trout (Oncorhynchus mykiss). To avoid possible cross-reactions during the double staining studies, combinations of primary antibodies raised in different species were used, and we report here the generation of an antibody raised in guinea pig against the rainbow trout glucocorticoid receptor (rtGR). The results obtained in vitellogenic females showed that GnRH-positive neurons in the caudal telencephalon/anterior preoptic region consistently exhibited rtGR immunoreactivity. Similarly, in the anterior ventral preoptic region, a group of tyrosine hydroxylase-positive neurons, known for inhibiting gonadotropin (GTH)-2 secretion during vitellogenesis, was consistently shown to strongly express GR. Finally, we show that a large majority of the GTH-1 (FSH-like) and GTH-2 (LH-like) cells of the pituitary exhibit rtGR immunoreactivity. These results indicate that cortisol may affect the neuroendocrine control of the reproductive process of the rainbow trout at multiple sites.


Subject(s)
Immunohistochemistry , Neurons/chemistry , Oncorhynchus mykiss/metabolism , Pituitary Gland/chemistry , Receptors, Glucocorticoid/analysis , Animals , Female , Gonadotropins, Pituitary/analysis , Preoptic Area/chemistry , Reproduction/physiology , Tyrosine 3-Monooxygenase/analysis , Vitellogenesis
14.
Article in English | MEDLINE | ID: mdl-9826997

ABSTRACT

The full length cDNA encoding a rainbow trout glucocorticoid receptor (rtGR) has been obtained from rainbow trout liver and intestine libraries. Northern blot analysis showed that the corresponding messengers are detected in the brain of trout with a size 7.5 kb similar to the size of rtGR mRNA in other target tissues. The distribution of the rtGR mRNA and protein was studied in the forebrain of the trout by means of both in situ hybridization and immunohistochemistry and compared with that of the oestrogen receptor (rtER). The GR and ER mRNAs and proteins were detected with a strong overlapping mainly in the: (a) preoptic region; (b) mediobasal hypothalamus; and (c) anterior pituitary, confirming their implication in the neuroendocrine control of pituitary functions. In both diencephalon and pituitary, the peptidergic phenotype of some neuron or cell categories expressing either type of receptors could be determined by double staining. Furthermore, double staining studies have demonstrated colocalization of the two receptors in the same neurons or pituitary cells. The rtER and rtGR were found to be co-expressed in the dopaminergic neurons inhibiting GTH2 secretion and in pituitary cells of the anterior lobe--notably the gonadotrophs. Given that the promoter of the ER gene contains several potential glucocorticoid-responsive elements (GRE) and that cortisol inhibits the oestradiol-stimulated ER expression in the liver, the possibility exists for modulation of ER gene expression by GR in the hypothalamo-pituitary complex. This could explain some of the well documented effects of stress on the reproductive performance in salmonids.


Subject(s)
Hydrocortisone/physiology , Oncorhynchus mykiss/physiology , Receptors, Glucocorticoid/physiology , Reproduction/physiology , Animals , Humans , Receptors, Glucocorticoid/drug effects
15.
J Comp Neurol ; 401(3): 395-410, 1998 Nov 23.
Article in English | MEDLINE | ID: mdl-9811116

ABSTRACT

The distribution of glucocorticoid receptor-expressing cells was studied in the forebrain of the rainbow trout by means of antibodies produced against a fusion protein made of the NH2-terminal fragment of the rainbow trout glucocorticoid receptor fused in frame with glutathione-S-transferase. The results indicate that glucocorticoid receptor-expressing cells are located in many brain regions from the telencephalon to the spinal cord, with the highest density in the neuroendocrine component of the brain, the preoptic region and the mediobasal hypothalamus, and in the periventricular zone of the optic tectum. In virtually all cases, the labeling was located in the nucleus of the cells, although on very rare occasions, a slight labeling of the cytoplasm was detected. Concerning the preoptic region, the most striking feature was the high density of glucocorticoid receptors in the magnocellular preoptic nucleus, known to contain corticotrophin-releasing factor (CRF)-, vasotocin-, and isotocin-expressing cells. Colocalization experiments showed that 100% of the CRF-immunoreactive neurons in the preoptic nucleus express glucocorticoid receptors. In the mediobasal hypothalamus, the highest expression was found in the nucleus lateralis tuberis and parts of the nucleus recessus lateralis. Concerning the pituitary, the glucocorticoid receptor was consistently found in the rostral pars distalis, with the exception of the prolactin cells, and in the proximal pars distalis, which in trout contains thyrotrophs, gonadotrophs, and somatotrophs. In the hindbrain, expression of glucocorticoid receptors were localized mainly in the periventricular regions.


Subject(s)
Oncorhynchus mykiss/metabolism , Prosencephalon/metabolism , Receptors, Glucocorticoid/metabolism , Animals , Immunohistochemistry , Medical Illustration , Neurons/metabolism , Prosencephalon/cytology , Tissue Distribution
16.
Neuroendocrinology ; 63(2): 156-65, 1996 Feb.
Article in English | MEDLINE | ID: mdl-9053780

ABSTRACT

A double immunocytochemical procedure, with two different chromogens, was used to compare the respective distribution of estrogen receptor-immunoreactive cells and tyrosine hydroxylase-immunoreactive neurons on the same sections of the preoptic region of adult female rainbow trout (Oncorhynchus mykiss). Estrogen receptor-immunoreactive cells were observed in the anterior preoptic region surrounding the preoptic recess and its large lateral extensions. Tyrosine hydroxylase-immunoreactive cells were consistently detected in the ventral and ventrolateral walls of the preoptic recess, in an area that was named nucleus preopticus pars anteroventralis. Dopamine immunohistochemistry and Dil retrograde transport studies indicated that part of these catecholaminergic neurons are dopaminergic and could project to the pituitary. Double staining studies showed consistently that most estrogen receptor-positive cells located ventral to the large extensions of the preoptic recess are also tyrosine hydroxylase-positive, indicating that this region is a major target for estradiol feedback. The results are discussed in relation to the role of the nucleus preopticus pars anteroventralis in mediating the negative feedback actions of estradiol on the secretion of gonadotrophin (GTH2) secretion. A hypothesis is drawn in order to explain the synchronizing role of estradiol at the time of ovulation in rainbow trout.


Subject(s)
Neurons/metabolism , Oncorhynchus mykiss/physiology , Preoptic Area/metabolism , Receptors, Estrogen/biosynthesis , Tyrosine 3-Monooxygenase/metabolism , Animals , Antibody Specificity , Carbocyanines , Dopamine/metabolism , Female , Immunohistochemistry , Neurons/enzymology , Preoptic Area/cytology , Preoptic Area/enzymology
17.
J Comp Neurol ; 363(3): 461-74, 1995 Dec 18.
Article in English | MEDLINE | ID: mdl-8847411

ABSTRACT

A double immunocytochemical procedure, with two different chromogens, was used to compare the respective distributions of estrogen receptor-immunoreactive cells and gonadotrophin-releasing hormone-immunoreactive neurons on the same sections of the brains of adult male and female rainbow trout (Oncorhynchus mykiss). Estrogen receptor-immunoreactive cells were observed in the ventral and lateral telencephalon, the preoptic region, the mediobasal hypothalamus, and the ventromedial thalamic nucleus. Gonadotrophin-releasing hormone-immunoreactive perikarya were detected in the olfactory bulbs, the ventral telencephalon, the preoptic area, and the mediobasal hypothalamus. Double-staining studies showed that, although some estrogen receptor-positive cells were in close proximity to gonadotrophin-releasing hormone-immunoreactive perikarya, careful examination of 550 gonadotrophin-releasing hormone-positive cells from five adult females and two adult males failed to demonstrate any evidence that gonadotrophin-releasing hormone neurons coexpress estrogen receptor in the brain of the rainbow trout. The present study provides, for the first time in teleosts, morphological evidence that gonadotrophin-releasing hormone neurons do not represent major direct targets for estradiol, suggesting that the positive feedback effects of estradiol onto the gonadotrophin-releasing hormone system are likely to be conveyed via other cell populations.


Subject(s)
Gonadotropin-Releasing Hormone/metabolism , Neurons/metabolism , Oncorhynchus mykiss/metabolism , Receptors, Estrogen/biosynthesis , Animals , Autoradiography , Brain/cytology , Brain Chemistry/physiology , Female , Immunohistochemistry , In Situ Hybridization , Male
18.
J Comp Neurol ; 350(1): 63-74, 1994 Dec 01.
Article in English | MEDLINE | ID: mdl-7532194

ABSTRACT

Galanin was purified from an extract of the stomach of the rainbow trout, Oncorhynchus mykiss, and its primary structure was established as Gly-Trp-Thr-Leu-Asn-Ser- Ala-Gly-Tyr-Leu10-Leu-Gly-Pro-His-Gly-Ile-Asp-Gly-His-Arg20- Thr-Leu-Ser-Asp- Lys-His-Gly-Leu-Ala. Trout galanin shows six amino acid substitutions compared with pig galanin, but the N-terminal region (residues 1-14) has been fully conserved. The distribution of galanin-immunoreactive (GAL-IR) structures in the trout brain and pituitary was studied via immunohistochemistry. GAL-IR cell bodies were observed only in the caudal telencephalon, the preoptic region, and the mediobasal hypothalamus. GAL-IR fibers, however, are widely distributed throughout the brain, with a much lower density in the midbrain and posterior brain than in the tel- and diencephalon. Particularly dense innervation of the mediobasal hypothalamus, the ventral and supracommissuralis parts of the caudal telencephalon, and the region above and below the anterior commissure was observed. A heavy innervation of the pituitary was consistently detected. GAL-IR fibers were present in neurohypophyseal digitations of both the anterior and intermediate lobes with highest density in the region of the proximal pars distalis, where growth hormone and gonadotropic cells are located. Fibers were also seen in digitations of the rostral pars distalis, in particular between the prolactin follicles. The distribution of GAL-IR neurons in the central nervous system and pituitary of the trout suggests that the peptide may exercise an important role in the regulation of neuroendocrine functions, particularly those related to reproduction.


Subject(s)
Brain/metabolism , Oncorhynchus mykiss/metabolism , Peptides/chemistry , Peptides/metabolism , Pituitary Gland/metabolism , Amino Acid Sequence , Animals , Brain/cytology , Brain/ultrastructure , Female , Galanin , Immunohistochemistry , Molecular Sequence Data , Nerve Fibers/metabolism , Neurons/metabolism , Neuropeptides/metabolism , Pituitary Gland/cytology , Pituitary Gland/ultrastructure , Tissue Distribution
19.
J Neuroendocrinol ; 6(5): 573-83, 1994 Oct.
Article in English | MEDLINE | ID: mdl-7827628

ABSTRACT

Using antibodies against the hormone binding domain of the trout estrogen receptor (ER), the distribution of ER-immunoreactive (ER-IR) cells was studied in the brain of maturing diploid and triploid female rainbow trout using a streptavidin-biotin-peroxidase method followed by a nickel-intensified diaminobenzidine reaction. This technique resulted in an excellent signal/background ratio allowing unambiguous identification of positive cells. In all animals, ER-IR cells were consistently located in three brain regions, the ventral telencephalon, the anterior ventral preoptic region, and the mediobasal hypothalamus. About 250 ER-IR cells were observed in the ventral and dorsal parts of the ventral telencephalon. In the anterior nucleus preopticus periventricularis, about 2400 ER-IR cells were observed surrounding the preoptic recess. In the posterior hypothalamus, approximately 2700 ER-IR cells were located in the anterior, posterior and inferior divisions of the nucleus lateralis tuberis and in the nucleus saccus vasculosus. In these regions cell nuclei exhibiting different densities of staining were observed and absolutely no labeling of cytoplasmic processes was detected. These results are in partial agreement with those obtained either after injection of tritiated-estradiol in other teleots species or in situ hybridization of ER mRNAs in trout. In particular, no immunoreactivity was observed in the thalamic region nor in the nucleus posterioris periventricularis. These data indicate that target cells for estradiol are essentially located in brain regions involved in the neuroendocrine control of pituitary functions and having direct connections with the hypophysis.


Subject(s)
Brain Chemistry , Oncorhynchus mykiss , Receptors, Estrogen/analysis , Animals , Brain/ultrastructure , Cell Nucleus/chemistry , Female , Hypothalamus, Middle/chemistry , Immunohistochemistry , Preoptic Area/chemistry , Telencephalon/chemistry , Tissue Distribution
20.
J Comp Neurol ; 348(4): 511-30, 1994 Oct 22.
Article in English | MEDLINE | ID: mdl-7836560

ABSTRACT

The distribution of neurotensin (NT) was studied in the brain of three species belonging to the three major classes of cold-blooded vertebrates: teleost fishes (Carassius auratus), anuran amphibians (Hyla meridionalis), and reptiles (Gallotia galloti; Lacertidae). By using antibodies directed against synthetic bovine NT in the three species, immunoreactive cell bodies were discovered mostly in the telencephalon and diencephalon, in particular at the level of the preoptic region the mediobasal hypothalamus, and the thalamus. In the frog and the lizard, additional immunoreactive (ir) structures were observed in the optic tectum and the tegmentum of the mesencephalon. In the goldfish pituitary, an extensive innervation was consistently observed at the level of the rostral pars distalis, whereas in both frog and lizard, positive fibers were only detected in the external layer of the median eminence. In the three species there is a striking overlap between the distribution of the NT-ir cell bodies and that of the target cells for sexual steroids. The results are discussed in relation with those reported in birds and mammals, and with the possible interactions among NT, sexual steroids, and the neuroendocrine control of pituitary hormone release, in particular prolactin and gonadotrophin.


Subject(s)
Amphibians/metabolism , Brain Chemistry/physiology , Goldfish/metabolism , Neuropeptides/analysis , Neurotensin/analysis , Reptiles/metabolism , Amino Acid Sequence , Animals , Biological Evolution , Molecular Sequence Data
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