Interactions between neurotensin and GnRH neurons in the positive feedback control of GnRH/LH secretion in the mouse.

In female mammals, increased ovarian estradiol (E(2)) secretion triggers GnRH release from neurons in the basal forebrain, which drives LH secretion from the pituitary and subsequently induces ovulation. However, the neural circuits that activate this preovulatory GnRH/LH surge remain unidentified. Neurotensin is expressed in neurons of the anteroventral periventricular nucleus (AVPV), a region thought to be critical for generating the preovulatory GnRH/LH surge.

Expression and activation of N-methyl-D-aspartate receptor subunit-1 receptor subunits in gonadotrophin-releasing hormone neurones of young and middle-aged mice during the luteinising hormone surge.

Glutamate is an important excitatory neurotransmitter that stimulates the release of gonadotrophin-releasing hormone (GnRH) and participates in the generation of the luteinising hormone (LH) surge. To determine the mechanisms of action of glutamate and possible changes in the glutamatergic input to GnRH neurones during reproductive ageing, we measured the expression and activation of the mandatory N-methyl-D-aspartate receptor subunit-1 (NMDAR1) in GnRH neurones of young and middle-aged mice prior to and during a steroid-induced LH surge. The results show that, in young animals, approximately 55% of all GnRH neurones contain immunoreactive NMDAR1 protein and this percentage does not change during the day of the LH surge.

Microwave-assisted antigen retrieval and incubation with cox-2 antibody of archival paraffin-embedded human oligodendroglioma and astrocytomas.

Immunohistochemistry is an important tool that is often used for the diagnosis of pathologies; however, the length of time required to process the tissue is relatively long. Furthermore, the quality and sensitivity of immunohistochemical staining is affected by formalin fixation which results in variable loss of antigenicity, known as masking effect. Here we assess the effect of microwave irradiation on the incubation time required to obtain high quality immunohistochemical staining for cox-2 using archival formalin-fixed, paraffin-embedded human oligodendrogliomas and astrocytomas.

Expression of AMPA receptor subunits (GluR1-GluR4) in gonadotrophin-releasing hormone neurones of young and middle-aged persistently oestrous rats during the steroid-induced luteinising hormone surge.

Glutamate provides excitatory input to gonadotrophin-releasing hormone (GnRH) neurones and elicits a response indicative of AMPA receptors. To determine if and which AMPA subunits are expressed by GnRH neurones, we conducted triple-label immunohistochemistry and confocal analyses on tissue obtained at 08.00, 12.

Localization of kainate receptor subunit GluR5-immunoreactive cells in the rat hypothalamus.

Glutamate is the major excitatory neurotransmitter in the hypothalamus, which exerts its effects by activating ion channel-forming (ionotropic) or G-protein-coupled (metabotropic) receptors. Kainate-preferring glutamate receptor subunits (GluR5, GluR6, GluR7, KA1, and KA2) form one of the three ionotropic receptor families. In the present study, we analyzed the distribution of GluR5 subunit protein in the rat hypothalamus with immunohistochemistry.

Identification of neurones in the female rat hypothalamus that express oestrogen receptor-alpha and vesicular glutamate transporter-2.

Oestrogen exerts its effects in the brain by binding to and activating two members of the nuclear receptor family, oestrogen receptor (ER)-alpha and ER-beta. Evidence suggests that oestrogen-receptive neurones participate in the generation of reproductive behaviours and that they convey the oestrogen message to gonadotropin-releasing hormone (GnRH) neurones. The aim of the present study was to identify the neurochemical phenotype of a subset of oestrogen receptor-expressing neurones.

Distribution of vesicular glutamate transporter-2 messenger ribonucleic Acid and protein in the septum-hypothalamus of the rat.

The excitatory neurotransmitter glutamate is involved in the control of most, perhaps all, neuroendocrine systems, yet the sites of glutamatergic neurons and their processes are unknown. Here, we used in situ hybridization and immunohistochemistry for the neuron-specific vesicular glutamate transporter-2 (VGLUT2) to identify the neurons in female rats that synthesize the neurotransmitter glutamate as well as their projections throughout the septum-hypothalamus. The results show that glutamatergic neurons are present in the septum-diagonal band complex and throughout the hypothalamus.

Expression of estrogen receptor-alpha and cFos in norepinephrine and epinephrine neurons of young and middle-aged rats during the steroid-induced luteinizing hormone surge.

Norepinephrine (NE) and epinephrine are important stimulators of GnRH release during the preovulatory surge in female rats. Previous studies have shown that the catecholaminergic neurons are sensitive to estradiol and that NE release in the hypothalamus is decreased in middle-aged rats at the time when the estrous cycles become irregular and later cease to exist. The aims of the present study were to determine whether the NE and epinephrine neurons continue to express estrogen receptor (ER)-alpha in middle-aged rats; temporal expression of ER-alpha and cFos changes with age during the steroid-induced surge; and tyrosine hydroxylase (TH), dopamine-beta-hydroxylase (DBH), and phenylethanol-N-methyltransferase mRNA content in catecholaminergic neurons of the brain stem changes during the surge with age.

Characterization of a polyclonal antibody to human pituitary tumor transforming gene 1 (PTTG1) protein.

Pituitary tumor transforming gene 1 (PTTG1), recently cloned from human testis, is a potent oncogene that is expressed in most tumors. However, assessment of its potential value as a prognostic marker is dependent on the development of a suitable antibody. We have developed a rabbit polyclonal antibody, SK601, that is highly specific for the PTTG1 gene product using recombinant PTTG1 protein (24 kD) containing an N-terminal His(6) tag as the immunogen.

Presence of luteinizing hormone-releasing hormone fragments in the rhesus monkey forebrain.

Previously, we have shown that two types of luteinizing hormone-releasing hormone (LHRH) -like neurons, "early" and "late" cells, were discernible in the forebrain of rhesus monkey fetuses by using antiserum GF-6, which cross-reacts with several forms of LHRH. The "late" cells that arose from the olfactory placode of monkey fetuses at embryonic days (E) 32-E36, are bona fide LHRH neurons. The "early" cells were found in the forebrain at E32-E34 and settled in the extrahypothalamic area.

Neural signals that regulate GnRH neurones directly during the oestrous cycle.

GnRH, produced by a loose network of neurones in the basal forebrain, is the primary brain signal responsible for the release of LH and FSH from the anterior pituitary gland. The ovarian steroid hormone oestradiol feeds back at both the central nervous system and the anterior pituitary to regulate the patterns of release of GnRH and the gonadotrophins. Although recent evidence indicates that oestradiol may act directly on some GnRH neurones through classical genomic mechanisms, data from published studies have demonstrated that neurotransmission of afferent neuronal systems that are receptive to oestradiol is necessary to drive reproductive cyclicity.

Expression of multidrug resistance-associated protein 2 in small intestine from pregnant and postpartum rats.

We analyzed the expression of multidrug resistance-associated protein 2 (mrp2) in the small intestine of control female rats and in rats during late pregnancy (19-20 days of pregnancy) and lactation (2-4, 10-14, and 21 days after delivery). Western blot analysis was performed on brush-border membranes prepared from different regions of the small intestine. Expression of mrp2 was maximal in the proximal segments for all experimental groups, was preserved in pregnant rats, and increased by 100% in postpartum rats by late lactation with respect to control animals.

Distribution of ionotropic glutamate receptor subunit mRNAs in the rat hypothalamus.

The excitatory amino acid neurotransmitter glutamate participates in the control of most (and possibly all) neuroendocrine systems in the hypothalamus. This control is exerted by binding to two classes of membrane receptors, the ionotropic and metabotropic receptor families, which differ in their structure and mechanisms of signal transduction. To gain a better understanding about the precise sites of action of glutamate and the subunit compositions of the receptors involved in the glutamatergic neurotransmission in the hypothalamus and septum, in situ hybridization was used with 35S-labeled cRNA probes for the different ionotropic receptor subunits, including glutamate receptor subunits 1-4 (GluR1-GluR4), kainate-2, GluR5-GluR7, N-methyl-D-aspartate (NMDA) receptor 1 (NMDAR1), and NMDAR2A-NMDAR2D.

Expression of estrogen receptor-alpha and c-Fos in adrenergic neurons of the female rat during the steroid-induced LH surge.

Epinephrine is an important neurotransmitter that is synthesized in relatively few neurons of the medullary regions C1-C3. Epinephrine is involved, among others in the control of most neuroendocrine systems, such as corticotropin releasing hormone-, gonadotropin releasing hormone- and oxytocin/vasopressin-containing neurons as part of complex feedback loop systems that often include interactions with the gonadal or adrenal steroid hormones. In order to determine if the interactions between gonadal steroid hormones with the adrenergic neurons are direct or involve steroid-receptive interneurons that in turn innervate the adrenergic neurons, dual immunohistochemistry was applied to identify if estrogen receptor-alpha (ERalpha) protein was expressed by adrenergic, phenylethanolamine-N-methyl transferase (PNMT)-positive neurons and if estradiol can activate these neurons as determined by the transient expression of the transcription factor c-Fos.

Expression and localization of multidrug resistant protein mrp2 in rat small intestine.

The expression of multidrug resistance-associated protein isoform 2 (mrp2), the ATP-dependent export pump that mediates the transport of glucuronic acid-, glutathione-, and sulfate-conjugated derivatives, was studied in rat small intestine. The small intestine was divided into nine equal segments, and mrp2 content was analyzed in homogenate and brush border membrane preparations by Western analysis. mrp2 protein was present mainly in brush border membrane of the proximal segments and gradually decreased from jejunum to the distal ileum.

Localization of serotonin(5A) receptors in discrete regions of the circadian timing system in the Syrian hamster.

Endogenous serotonin and serotonergic drugs influence many aspects of circadian rhythms, including phase shifts, onset of locomotor activity, and period length and integrity of rhythms during exposure to constant light. The receptor subtype(s) mediating all of these circadian effects of serotonin has (have) not been identified. Immunoreactivity for the serotonin(5A) (5-HT(5A)) receptor has recently been identified in the rat suprachiasmatic nucleus (SCN).

Expression of ionotropic glutamate receptor subunit mRNAs in the hypothalamic paraventricular nucleus of the rat.

The hypopthalamic paraventricular nucleus (PVN) coordinates multiple aspects of homeostatic regulation, including pituitary-adrenocortical function, cardiovascular tone, metabolic balance, fluid/electrolyte status, parturition and lactation. In all cases, a substantial component of this function is controlled by glutamate neurotransmission. In this study, the authors performed a high-resolution in situ hybridization analysis of ionotropic glutamate receptor subunit expression in the PVN and its immediate surround.

Evidence that members of the TGFbeta superfamily play a role in regulation of the GnRH neuroendocrine axis: expression of a type I serine-threonine kinase receptor for TGRbeta and activin in GnRH neurones and hypothalamic areas of the female rat.

The present study was designed to determine whether transforming growth factor (TGF)beta and/or activin participate in the regulation of the gonadotropin releasing hormone (GnRH) neuroendocrine axis in vivo. Single-label in situ hybridization histochemistry was used to determine the anatomical distribution of a TGFbeta and activin type I receptor (B1) mRNA, in the adult female rat hypothalamic areas that are known to be important sites for the regulation of reproduction. Dual-label in situ hybridization histochemistry was performed to determine whether B1 mRNA was expressed in GnRH neurones.

Growth-associated protein-43 messenger ribonucleic acid expression in gonadotropin-releasing hormone neurons during the rat estrous cycle.

We have shown previously at the ultrastructural level that morphological changes occur in the external zone of the median eminence allowing certain GnRH nerve terminals to contact the pericapillary space on the day of proestrus. The present study was designed to determine whether the intrinsic determinant of neuronal outgrowth, growth-associated protein-43 (GAP-43), was expressed in GnRH neurons of adult female rats, and whether its expression varied throughout the estrous cycle. To accomplish this, we perfusion-fixed groups of adult female rats at 0800 and 1600 h on diestrous day 2 (diestrous II), at 0800 h and 1600 h on proestrus, and at 0800 and 1600 h on estrus (n = 4 rats/group) and used double labeling in situ hybridization and quantification to compare the levels of GAP-43 messenger RNA (mRNA) in cells coexpressing GnRH mRNA.

Kainate receptor subunit-positive gonadotropin-releasing hormone neurons express c-Fos during the steroid-induced luteinizing hormone surge in the female rat.

During the preovulatory and estradiol-progesterone-induced GnRH-LH surge, a subpopulation of GnRH neurons transiently expresses the transcription factor c-fos, which is a useful marker of cell activation. To further characterize this subpopulation of GnRH neurons, multiple immunohistochemical procedures were applied to visualize GnRH, c-Fos, KA2, GluR5, GluR6, and GluR7 receptor subunits during different phases of the estrogen-progesterone-induced LH surge. The results show that the LH surge begins at 1400 h and peaks at 1600 h before returning to baseline late in the evening.

Evidence for expression of galanin receptor Gal-R1 mRNA in certain gonadotropin releasing hormone neurones of the rostral preoptic area.

Previous studies have shown that galanin plays an important role in the regulation of gonadotropin releasing hormone (GnRH) release. At present, it is not known if this role is exerted by direct or indirect interactions between galanin producing neurones and GnRH neurones. The objective of this study was to determine whether or not GnRH neurones could express galanin receptor Gal-R1 mRNA.

Molecular cloning and characterization of the tumor transforming gene (TUTR1): a novel gene in human tumorigenesis.

We cloned and sequenced the cDNA of a potent tumor transforming gene (TUTR1) from human testis and determined its primary structure. The TUTR1 cDNA is composed of 656 nucleotides and encodes a novel protein of 202 amino acids. The predicted TUTR1 protein is extremely hydrophilic and contains two proline-rich motifs at its C-terminus.

Galanin within the normal and hyperplastic anterior pituitary gland: localization, secretion, and functional analysis in normal and human growth hormone-releasing hormone transgenic mice.

Studies evaluating estrogen-induced anterior pituitary tumors revealed a strong direct correlation between expression of the peptide galanin and tumor growth. To evaluate further the potential roles of galanin in the hyperplastic pituitary, we used a model of estrogen-independent anterior pituitary tumor formation, the male human GH-releasing hormone (hGHRH) transgenic mouse. Pituitaries of hGHRH transgenic mice are characterized by a hyperplasia of somatotrophs and contain markedly elevated levels of galanin.