Gonadotropin-releasing hormone is prerequisite for the constitutive expression of pituitary annexin A5.

Annexin A5 (ANXA5), a member of the structurally related family of annexin proteins, is expressed in pituitary gonadotropes. We previously reported that ANXA5 expression is stimulated by gonadotropin-releasing hormone (GnRH). In the present study, we investigated ANXA5 expression in the anterior pituitary gland of GnRH-deficient mutant hypogonadal (hpg) mice. RT-PCR demonstrated that luteinizing hormone ß subunit (LHß) and ANXA5 mRNA levels were both lower in the pituitary gland of hpg mice than in wild-type mice. Immunohistochemistry showed that ANXA5 expression throughout the pituitary gland was very low in hpg mice, suggesting that ANXA5 is diminished in gonadotropes and also in other cell types. Subcutaneous administration of a GnRH analogue, des-gly10 (Pro9)-GnRH ethylamide (1 µg/day for 7 days), augmented the expression of LHß and ANXA5 in the pituitary gland in hpg mice. However, LHß- and ANXA5-positive cells did not show exactly matched spatial distributions. These findings suggest that GnRH is necessary for constitutive ANXA5 expression in the pituitary gland, not only in gonadotropes but also in other pituitary gland cell types. A close relationship between ANXA5 and LHß expression was confirmed. It is suggested that a significant role of ANXA5 in the physiologic secretion of LH.

Effects of castration on androgen receptors and gonadotropins in the pituitary of adult male viscachas.

The aims of the present study were to determine whether castration results in quantitative immunohistochemical changes in androgen receptors (AR), LH-immunoreactive (IR) cells and FSH-IR cells, and to analyse the colocalisation of AR and gonadotropins in the pituitary pars distalis (PD) of viscachas. Pituitaries were processed for light and electron microscopy. AR-IR, LH-IR and FSH-IR cells were detected by immunohistochemistry. In morphometric studies, the percentage of AR-IR, LH-IR, FSH-IR, LH-IR/AR-IR and FSH-IR/AR-IR cells was determined. In intact viscachas, AR were distributed throughout the PD; they were numerous at the caudal end, with intense immunostaining. LH-IR cells and FSH-IR cells were found mainly in the ventral region and at the rostral end of the PD. Approximately 45%-66% of LH-IR cells and 49%-57% of FSH-IR cells expressed AR in the different zones of the PD. In castrated viscachas, there was a significant decrease in the percentage of AR-IR, LH-IR, FSH-IR, and FSH-IR/AR-IR cells. Some pituitary cells from castrated viscachas also exhibited ultrastructural changes. These results provide morphological evidence that gonadal androgens are directly related to the immunolabelling of AR, LH and FSH. Moreover, the colocalisation of AR and FSH is most affected by castration, suggesting the existence of a subpopulation of gonadotrophs with different regulatory mechanisms for hormonal synthesis, storage and secretion.

Immunocytochemical and ultrastructural identification of adenohypophyseal cells in Ctenopharyngodon idella (Cypriniformes: Cyprinidae) during gonadal differentiation.

The adenohypophysis was studied by immunocytochemical and ultrastructural methods in juvenile grass carp (Ctenopharyngodon idella) from natural reproduction in Northern Italian rivers. The adenohypophysis included the rostral pars distalis (RPD), the proximal pars distalis (PPD) and the pars intermedia (PI), all deeply penetrated by branches of the neurohypophysis (Nh). The prolactin (PRL), adrenocorticotropic (ACTH), somatotropic (GH), thyrotropic (TSH), gonadotropic type I (GtH I) and type II (GtH II), somatolactin (SL), melanotropic (MSH) and endorphin (END) cells were identified with antisera raised against piscine and human pituitary hormones. In juveniles of 51-69 mm of total body length (TL) with undifferentiated gonads, the PRL cells, arranged in thick strands, occupied most of the RPD. The ACTH and GH cells organized in cords bordering Nh were, respectively, confined to RPD and PPD. The TSH cells were scattered among ACTH cells in RPD and among GH cells in PPD. Cells simultaneously immunoreactive to anti-follicle stimulating hormone and to anti-croaker gonadotropin were intermingled among GH and TSH cells, which were mostly in the dorsal PPD. The SL cells were detected in PI layers bordering the Nh. The MSH and END cells were intermingled in PI and, unlike what observed in other teleosts, their respective antisera did not cross-react. In individuals of 78-112 mm TL with gonads at the beginning of differentiation, the GtH II cells were detected in PPD; all other cell types increased in number. These results, supported by ultrastructural investigations, suggest that SL and GtH II cells are directly involved in gonadal differentiation in C. idella.

Aging and diets regulate the rat anterior pituitary and hypothalamic transcriptome.

Dietary interventions involving caloric restriction represent a powerful strategy to prevent or delay age-related deteriorations and diseases. Their beneficial effects have been observed in several tissues and species. This microarray study investigated the effects of aging, long-term moderate caloric restriction (LTMCR) and long-term dietary soy on the regulation of gene expression in the anterior pituitary and hypothalamus of 20-month-old Sprague-Dawley rats. In both tissues, aging regulated genes mainly involved in cell defense and repair mechanisms related to apoptosis, DNA repair, cellular stress, inflammatory and immune response. In the aging pituitary, the highest upregulated gene was the regenerating islet-derived 3ß (5.77-fold), coding for a secretory protein involved in acute stress and inflammation. A protective effect of LTMCR on age-related change of gene expression was observed for 35 pituitary genes. In addition, beneficial effects of LTMCR in the pituitary were observed on new regulated genes mainly involved in cell death and cell stress response. In the hypothalamus, the effects of LTMCR on age-related changes were modest. Finally, changing the quality of dietary protein (20% casein for soy) had a low impact on the regulation of mRNA levels in both tissues. Genes associated with the somatotroph function were also differentially expressed in the aging pituitary. Interestingly, LTMCR prevented the effect of aging on insulin-like growth factor-binding protein-3 gene. Altogether, this study proposes novel pituitary and hypothalamic molecular targets and signaling pathways to help in understanding the mechanisms involved in aging processes and LTMCR.

Hypogonadism and obesity in mice with a targeted deletion of the Nhlh2 gene.

The family of basic helix-loop-helix (bHLH) genes comprises transcription factors involved in many aspects of growth and development. We have previously described two bHLH transcription factors, Nhlh1 and Nhlh2 (originally named NSCL1 and NSCL2). The nucleotide and predicted protein sequences of Nhlh1 and Nhlh2 are homologous within their bHLH domain where there are only three conservative amino acid differences. During murine embryogenesis, Nhlh1 and Nhlh2 share an overlapping but distinct pattern of expression in the developing nervous system. To improve our understanding of the role of these genes during neurogenesis, we have generated mice containing targeted deletions of both genes and here describe our results for Nhlh2. Loss of Nhlh2 results in a disruption of the hypothalamic-pituitary axis in mice. Male Nhlh2-/- mice are microphallic, hypogonadal and infertile with alterations in circulating gonadotropins, a defect in spermatogenesis and a loss of instinctual male sexual behaviour. Female Nhlh2-/- mice reared alone are hypogonadal, but when reared in the presence of males, their ovaries and uteri develop normally and they are fertile. Both male and female homozygotes exhibit progressive adult-onset obesity. Nhlh2 is expressed in the ventral-medial and lateral hypothalamus, Rathke's pouch and in the anterior lobe of the adult pituitary. Our results support a role for Nhlh2 in the onset of puberty and the regulation of body weight metabolism.

The GnRH Antagonist Degarelix Suppresses Gonadotropin Secretion and Pituitary Sensitivity in Midgestation Sheep Fetuses.

The specific role of gonadotropin-releasing hormone (GnRH) on brain sexual differentiation remains unclear. To investigate whether gonadotropin and, in turn, testosterone (T) secretion is regulated by GnRH during the critical period for brain differentiation in sheep fetuses, we attempted to selectively suppress pituitary-testicular activation during midgestation with the long-acting GnRH antagonist degarelix. Fetuses received subcutaneous injections of the antagonist or vehicle on day 62 of gestation. After 2 to 3 weeks we examined consequences of the intervention on baseline and GnRH-stimulated plasma luteinizing hormone (LH) and T levels. In addition, we measured the effect of degarelix-treatment on messenger RNA (mRNA) expression for the pituitary gonadotropins and key gonadal steroidogenic enzymes. Baseline and GnRH-stimulated plasma LH levels were significantly suppressed in degarelix-treated male and female fetuses compared to control values. Similarly, T concentrations were suppressed in degarelix-treated males. The percentage of LHß-immunoreactive cells colocalizing c-fos was significantly reduced by degarelix treatment indicating that pituitary sensitivity was inhibited. Degarelix treatment also led to the significant suppression of mRNA expression coding for the pituitary gonadotropin subunits and for the gonadal enzymes involved in androgen synthesis. These findings demonstrate that pharmacologic inhibition of GnRH early in gestation results in suppression of LH secretion and deficits in the plasma T levels of male lamb fetuses. We conclude that GnRH signaling plays a pivotal role for regulating T exposure during the critical period of sheep gestation when the brain is masculinized. Thus, disturbance to gonadotropin secretion during this phase of gestation could have long-term consequence on adult sexual behaviors and fertility.

[Hypoplasia adrenal congenita of anencephalic type: two cases with pituitary abnormalities and review of literature]. / L'hypoplasie surrénale congénitale de type anencéphalique: deux cas avec anomalies hypophysaires, sans mutation de DAX-1 et SF-1 et revue de la littérature.

Hypoplasia adrenal congenita is an extremely uncommon disease of early onset. This condition can be lethal in the absence of treatment. Some forms are due to the congenital adrenal hypoplasia of anencephalic type whose origin is even unknown. Here, we present two cases of congenital adrenal hypoplasia of anencephalic type with pituitary abnormalities. The two male newborns died because adrenal insufficiency in the neonatal period. The adrenal glands were hypoplastic with a histological structure of anencephalic type Immunocytochemical study of the pituitary revealed an absence of the gonadotrophs. No mutation of DAX 1 and SF-1 was found.

Localization of an endocannabinoid system in the hypophysial pars tuberalis and pars distalis of man.

The hypophysial pars tuberalis (PT) acts as an important interface between neuroendocrine brain centers (hypothalamus, pineal organ) and the pars distalis (PD) of the hypophysis. Recently, we have identified an endocannabinoid system in the PT of hamsters and provided evidence that 2-arachidonoylglycerol is a messenger molecule that appears to play an essential role in seasonal reproduction and prolactin release by acting on the cannabinoid receptors in the PD. We now demonstrate the enzymes involved in endocannabinoid synthesis and degradation, namely sn-1-selective diacylglycerol lipase α, N-acylphosphatidylethanolamine-specific phospholipase D, and monoacylglycerol lipase, in the PT of man by means of immunohistochemistry. High-performance liquid chromatography coupled with tandem mass spectrometry revealed 2-arachidonoylglycerol and other endocannabinoids in the human PT. Furthermore, we detected the expression of the cannabinoid receptor 1 (CB1), a primary receptor for endocannabinoids, in the PD. Double-immunofluorescence staining for CB1 and various hypophysial hormones or S-100, a marker for folliculostellate (FS) cells, revealed that CB1 immunoreactivity was mainly localized to corticotrophs and FS-cells. A limited number of lactotrophs and somatotrophs also showed CB1 immunoreactivity, which was however absent from gonadotrophs and thyrotrophs. Our data thus indicate that the human PT comprises an endocannabinoid system, and that corticotrophs and FS-cells are the main target cells for endocannabinoids. The functional significance of this newly discovered pathway remains to be elucidated in man; it might be related to the control of stress responses and/or reflect a remnant seasonal control of hypophysial hormonal secretion.

Localization of three types of arginine vasotocin receptors in the brain and pituitary of the newt Cynops pyrrhogaster.

The distribution of three types of arginine vasotocin (AVT) receptors in the brain and pituitary of the newt Cynops pyrrhogaster, namely, the V1a-, V2-, and V3/V1b-type receptors, was studied by means of in situ hybridization and immunohistochemistry. mRNA signals and immunoreactive cells for the V1a-type receptor were observed in the telencephalon (mitral layer of the olfactory bulb, dorsal and medial pallium, lateral and medial amygdala, bed nucleus of the decussation of the fasciculus telencephali, bed nucleus of the stria terminalis), diencephalon (anterior preoptic area, magnocellular preoptic nucleus, suprachiasmatic nucleus, ventral thalamus, dorsal and ventral hypothalamic nucleus), mesencephalon (tegmentum, interpeduncular nucleus), and medulla oblongata (median reticular formation, nucleus motorius tegmenti). Cells expressing the V2-type receptor were found in the telencephalon (medial pallium, lateral and medial amygdala, bed nucleus of the decussation of the fasciculus telencephali), and mesencephalon (tegmentum trigemini and facialis). In the paraphysis (possibly the main site of cerebrospinal fluid production), only V2-type receptor mRNA signal and immunoreactivity were detected. V3/V1b-type receptor mRNA was expressed in the diencephalon (dorsal hypothalamic nucleus, nucleus tuberculi posterioris), mesencephalon (tegmentum, interpeduncular nucleus), and medulla oblongata (raphe nucleus), whereas V3/V1b-type-receptor-like immunoreactivity was scarcely detectable in the entire brain. The V3/V1b-type receptor was predominantly expressed in the anterior pituitary. V3/V1b-type receptor and proopiomelanocortin mRNAs were co-localized in the distal lobe of the pituitary. This is the first report of the distribution of three types of AVT receptor in the brain and pituitary of non-mammalian vertebrates.

Reproductive axis function and gonadotropin microheterogeneity in a male rat model of diet-induced obesity.

Obesity causes complex metabolic and endocrine changes that may lead to adverse outcomes, including hypogonadism. We herein studied the reproductive axis function in male rats under a high-fat diet and analyzed the impact of changes in glycosylation of pituitary LH on the bioactivity of this gonadotropin. Rats were fed with a diet enriched in saturated fat (20% of total calories) and euthanized on days 90 or 180 of diet. Long-term (180 days), high-fat feeding rats exhibited a metabolic profile compatible with insulin resistance and metabolic syndrome; they concomitantly showed decreased intrapituitary and serum LH concentrations, low serum testosterone levels, and elevated serum 17beta-estradiol concentrations. A fall in biological to immunological ratio of intrapituitary LH was detected in 180 days control diet-treated rats but not in high-fat-fed animals, as assessed by a homologous in vitro bioassay. Chromatofocusing of pituitary extracts yielded multiple LH charge isoforms; a trend towards decreased abundance of more basic isoforms (pH 9.99-9.0) was apparent in rats fed with the control diet for 180 days but not in those that were fed the diet enriched in saturated fat. It is concluded that long-term high-fat feeding alters the function of the pituitary-testicular axis, resulting in hypogonadotropic hypogonadism. The alterations in LH function found in these animals might be subserved by changes in hypothalamic GnRH output and/or sustained gonadotrope exposure to an altered sex steroid hormone milieu, representing a distinctly different regulatory mechanism whereby the pituitary attempts to counterbalance the effects of long-term obesity on reproductive function.

Associations between early life stress and anterior pituitary gland volume development during late childhood.

Early Life Stress (ELS) is thought to influence Hypothalamic-Pituitary-Adrenal-Axis (HPAA) functioning, contributing to an increased risk for psychopathology through dysregulation of biological stress responses. Research exploring relationships between ELS and HPAA functioning has largely focused on its key hormonal output, cortisol. However, findings have been inconsistent, potentially due to cortisol's distinctive diurnal patterns and dynamic nature complicating its accurate measurement. Thus, this study explored the link between ELS and a more stable, structural component of the HPAA, specifically, anterior pituitary gland volume (PGV) in a community sample of children (N = 129, 68 female). PGV was traced from Magnetic Resonance Imaging brain scans across two time-points at ages 8 (baseline) and 10 years (follow-up). ELS exposure was assessed at baseline through parent-report questionnaires and maternal affective behavior observed in mother-child interaction tasks. ELS variables were reduced to a 5-factor structure using exploratory factor analysis - Uninvolved Parenting, Negative Affective Parenting, Neglect, Trauma, and Dysfunctional Discipline. Direct and sex-moderated associations between ELS and PGV were explored using regression and linear mixed models analyses. PGV-mediated associations between ELS and internalizing symptoms were also investigated. Childhood Neglect was significantly associated with greater baseline anterior PGV, that was stable over the follow-up period. This effect was found in the whole sample, and in males, specifically. No mediation effects were found. Results suggest that neglect may play a unique role in HPAA neurodevelopment; however, it is important that future research extends into adolescence to more clearly characterize these neurodevelopmental associations and any subsequent psychopathological outcomes.

Prohormone processing in the trans-Golgi network: endoproteolytic cleavage of prosomatostatin and formation of nascent secretory vesicles in permeabilized cells.

Many peptide hormones are synthesized as larger precursors which undergo endoproteolytic cleavage at paired basic residues to generate a bioactive molecule. Morphological evidence from several laboratories has implicated either the TGN or immature secretory granules as the site of prohormone cleavage. To identify the site where prohormone cleavage is initiated, we have used retrovirally infected rat anterior pituitary GH3 cells which express high levels of prosomatostatin (proSRIF) (Stoller, T. J., and D. Shields. J. Cell Biol. 1988. 107:2087-2095). By incubating these cells at 20 degrees C, a temperature that prevents exit from the Golgi apparatus, proSRIF accumulated quantitatively in the TGN and no proteolytic processing was evident; processing resumed upon shifting the cells back to 37 degrees C. After the 20 degrees C block, the cells were mechanically permeabilized and pro-SRIF processing determined. Cleavage of proSRIF to the mature hormone was approximately 35-50% efficient, required incubation at 37 degrees C and ATP hydrolysis, but was independent of GTP or cytosol. The in vitro ATP-dependent proSRIF processing was inhibited by inclusion of chloroquine, a weak base, CCCP, a protonophore, or by preincubating the permeabilized cells with low concentrations of N-ethylmaleimide, an inhibitor of vacuolar-type ATP-dependent proton pumps. These data suggest that: (a) proSRIF cleavage is initiated in the TGN, and (b) this reaction requires an acidic pH which is facilitated by a Golgi-associated vacuolar-type ATPase. A characteristic feature of polypeptide hormone-producing cells is their ability to store the mature hormone in dense core secretory granules. To investigate the mechanism of protein sorting to secretory granules, the budding of nascent secretory vesicles from the TGN was determined. No vesicle formation occurred at 20 degrees C; in contrast, at 37 degrees C, the budding of secretory vesicles was approximately 40% efficient and was dependent on ATP, GTP, and cytosolic factors. Vesicle formation was inhibited by GTP gamma S suggesting a role for GTP-binding proteins in this process. Vesicle budding was dependent on cytosolic factors that were tightly membrane associated and could be removed only by treating the permeabilized cells with high salt. After high salt treatment, vesicle formation was dependent on added cytosol or the dialyzed salt extract. The formation of nascent secretory vesicles contrasts with prosomatostatin processing which required only ATP for efficient cleavage. Our results demonstrate that prohormone cleavage which is initiated in the TGN, precedes vesicle formation and that processing can be uncoupled from the generation of nascent secretory vesicles.

Substance P-immunoreactive cells in the ovine pars tuberalis.

The pars tuberalis (PT) is a distinct subdivision of the anterior pituitary gland that plays a central role in regulating seasonal prolactin release. In sheep, there is compelling evidence that seasonal changes in light, transformed into a melatonin signal, are interpreted by the PT to modulate the release of a factor which affects prolactin release. The identity of this factor(s) is unknown but has been preemptively called 'tuberalin'. In the present study, we report on an initial immunocytochemical investigation where we have identified that many ovine PT cells are immunoreactive for the tachykinin substance P (SP). Few cells in the pars distalis immunoreact for SP. The SP-immunoreactive cells did not colocalize with beta-luteinizing hormone. RT-PCR confirmed the presence of preprotachykinin A mRNA in the PT. We hypothesize that SP, and possibly other preprotachykinin A-derived tachykinins, may play a role in the seasonal regulation of prolactin secretion in sheep.

Immunohistochemical distribution of regulatory peptides in the human fetal adenohypophysis.

We have studied here the cellular distribution of several regulatory peptides in hormone-producing cells of the human pituitary during the fetal period. Immunohistochemistry was used to show the expression of several regulatory peptides, namely Angiotensin-II, Neurotensin and Galanin, at successive gestational stages and their co-localization with hormones in the human fetal adenohypophysis. Somatotrophs, gonadotrophs and thyrotrophs were differentiated earliest. At gestational week 9, Angiotensin-II immunoreactivity was co-localized only with growth hormone immunoreactivity in somatotrophs, one of the first hormone-producing cells to differentiate. This co-localization remained until week 37. Neurotensin immunoreactivity was present in gonadotrophs and thyrotrophs in week 23, after FSH and TSH hormone differentiation. Galanin immunoreactivity was present in all hormone-producing cell types except corticotrophs. The different pro-opiomelanocortin-derived peptides were detected at different stages of gestation and adrenocorticotrophic hormone immunoreaction was the last to be detected. Our results show an interesting relationship between regulatory peptides and hormones during human fetal development, which could imply that these peptides play a regulatory role in the development of pituitary function.

The central effect of beta-endorphin and naloxone on the expression of GnRH Gene and GnRH receptor (GnRH-R) gene in the hypothalamus, and on GnRH-R gene in the anterior pituitary gland in follicular phase ewes.

The effect of prolonged intermittent infusion of beta-endorphin or naloxone into the third cerebral ventricle in ewes during the follicular phase of the estrous cycle on the expression of GnRH gene and GnRH-R gene in the hypothalamus and GnRH-R gene in the anterior pituitary gland was examined by Real time-PCR. Activation of micro opioid receptors decreased GnRH mRNA levels in the hypothalamus and led to complex changes in GnRH-R mRNA: an increase of GnRH-R mRNA in the preoptic area, no change in the anterior hypothalamus and decrease in the ventromedial hypothalamus and stalk/median eminence. In beta-endorphin treated ewes the levels of GnRH-R mRNA in the anterior pituitary gland also decreased significantly. These complex changes in the levels of GnRH mRNA and GnRH-R mRNA were reflected in the decrease of LH secretion. Blockade of micro opioid receptors affected neither GnRH mRNA and GnRH-R mRNA nor LH levels secretion. These results indicate that beta-endorphin displays a suppressive effect on the expression of the GnRH gene in the hypothalamus and GnRH-R gene in the anterior pituitary gland, but affects GnRH-R gene expression in a specific manner in the various parts of hypothalamus; altogether these events lead to the decrease in GnRH/LH secretion.

Effect of Early Calf-Hood Nutrition on the Transcriptional Regulation of the Hypothalamic-Pituitary-Testicular axis in Holstein-Friesian Bull Calves.

The aim of this study was to investigate the effect of early calf-hood nutrition on the transcriptomic profile of the arcuate nucleus of the hypothalamus, anterior pituitary and testes in Holstein-Friesian bulls. Holstein-Friesian bull calves with a mean (±S.D.) age and bodyweight of 19 (±8.2) days and 47.5 (±5.3) kg, respectively, were offered a high (n = 10) or low (n = 10) plane of nutrition in order to achieve an overall growth rate of 1.2 and 0.5 kg/day. At 126 (±3) days of age, calves were euthanized, hypothalamus (arcuate region), anterior pituitary and testicular parenchyma samples were harvested and RNAseq analysis was performed. There were 0, 49 and 1,346 genes differentially expressed in the arcuate nucleus, anterior pituitary and testicular tissue of bull calves on the low relative to the high plane of nutrition, respectively (P < 0.05; False Discovery Rate <0.05). Cell cycle processes in the anterior pituitary were down regulated in the low relative to the high plane of nutrition; there was no differential expression of genes related to reproductive processes. Gene expression involved in cholesterol and androgen biosynthesis in the testes were down regulated in animals on the low plane of nutrition. This study provides insight into the effect of early life plane of nutrition on the regulation of the HPT axis.

The influence of 17beta-estradiol on annexin 1 expression in the anterior pituitary of the female rat and in a folliculo-stellate cell line.

Annexin 1 (ANXA1) is a Ca2+- and phospholipid-binding protein that plays an important role as a mediator of glucocorticoid action in the host-defence and neuroendocrine systems. Sex differences in hypothalamo-pituitary-adrenal (HPA) axis activity are well documented and a number of studies have demonstrated that gonadal steroids act as regulators of HPA activity. The aim of this study was to investigate the effect of ovariectomy and 17beta-estradiol replacement, and estrous cycle stage, on anterior pituitary ANXA1 content. The amount of anterior pituitary ANXA1 determined by western blotting varied with estrous cycle stage with a peak at estrus declining to a trough at proestrus. Ovariectomy resulted in a significant (P<0 x 05) decrease in anterior pituitary ANXA1 content. Administration of 17beta-estradiol (1 microg/100 g) significantly (P<0 x 01) increased anterior pituitary ANXA1 expression in the ovariectomized animals. In contrast, there was no change in pituitary ANXA1 content in response to 17beta-estradiol in adrenalectomized and adrenalectomized/ovariectomized rats. Treatment of TtT/GF cells, a folliculo-stellate cell line, with 17beta-estradiol (1 x 8-180 nM) increased ANXA1 mRNA expression and increased the amount of ANXA1 protein externalized in response to a dexamethasone stimulus. These results indicate that 17beta-estradiol stimulates ANXA1 expression in the anterior pituitary and in vivo an adrenal factor contributes to the mechanism of action.

Muscular dystrophy-related quantitative and chemical changes in adenohypophysis GH-cells in golden retrievers.

Duchenne muscular dystrophy (DMD) is a recessive X-linked lethal condition which affects a boy in every 3300 births. It is caused by the absence of dystrophin, a protein occurring especially within the musculoskeletal system and in neurons in specific regions of the central nervous system (CNS). Growth hormone (GH) inhibition is believed to decrease the severity of DMD and could perhaps be used in its treatment. However, the underlying pathological mechanism is not known. The golden retriever muscular dystrophy dog (GRMD) represents an animal model in the study of DMD. In this paper we investigated the morphological aspects of the adenohypophysis as well as the total number and size of GH-granulated cells using design-based stereological methods in a limited number of dystrophic and healthy golden retrievers. GH-cells were larger (32.4%) in dystrophic dogs than in healthy animals (p=0.01) and they occupied a larger portion (62.5%) of the adenohypophysis volume (p=0.01) without changes in either adenohypophysis volume (p=0.893) or total number of GH-granulated cells (p=0.869). With regard to ultrastructure, granulated cells possessed double-layer electron-dense granules which were evenly distributed in the cytosol. Furthermore, these granules in dystrophic animals occupied a larger proportion of GH-granulated cell volume (66.9%; p=0.008) as well as of all GH-cells in the whole pars distalis of adenohypophysis (77.3%; p=0.035), albeit IGF-1 serum concentration was lower in severe cases. This suggests difficulties in the GH secretion that might possibly be associated to dystrophin absence. In contrast to earlier reports, our data suggest that a lower IGF-1 concentration may be more related to a severe, as opposed to a benign, clinical form of muscular dystrophy.

A glycoprotein hormone expressed in corticotrophs exhibits unique binding properties on thyroid-stimulating hormone receptor.

Corticotroph-derived glycoprotein hormone (CGH), also referred to as thyrostimulin, is a noncovalent heterodimer of glycoprotein hormone alpha 2 (GPHA2) and glycoprotein hormone beta 5 (GPHB5). Here, we demonstrate that both subunits of CGH are expressed in the corticotroph cells of the human anterior pituitary, as well as in skin, retina, and testis. CGH activates the TSH receptor (TSHR); (125)I-CGH binding to cells expressing TSHR is saturable, specific, and of high affinity. In competition studies, unlabeled CGH is a potent competitor for (125)I-TSH binding, whereas unlabeled TSH does not compete for (125)I-CGH binding. Binding and competition analyses are consistent with the presence of two binding sites on the TSHR transfected baby hamster kidney cells, one that can interact with either TSH or CGH, and another that binds CGH alone. Transgenic overexpression of GPHB5 in mice produces elevations in serum T(4) levels, reductions in body weight, and proptosis. However, neither transgenic overexpression of GPHA2 nor deletion of GPHB5 produces an overt phenotype in mice. In vivo administration of CGH to mice produces a dose-dependent hyperthyroid phenotype including elevation of T(4) and hypertrophy of cells within the inner adrenal cortex. However, the distinctive expression patterns and binding characteristics of CGH suggest that it has endogenous biological roles that are discrete from those of TSH.

Breed differences in clearance of porcine FSH in hypophysectomized rats.

Extracts of anterior pituitary (AP) glands were infused i.v. into hypophysectomized male rats followed by sequential sampling of blood for 120 min. Determination of follicle-stimulating hormone (FSH) concentrations established that FSH from Chinese Meishan males decreased in the circulation of rats more slowly than FSH in extracts of AP from crossbred occidental pigs (P<0.003). Additionally, FSH from AP extracts of castrated males disappeared somewhat more slowly (P<0.06) than FSH from extracts of boars. Evaluation of FSH by bioassay and radioimmunoassay yielded similar concentrations in AP from Meishan and crossbred boars. Serum testosterone concentrations increased with time through 90 min after infusion of AP, but the rate of increase of testosterone was not related to amount of luteinizing hormone (LH) that was administered indicating LH receptor saturation. Unexpectedly, the rate of increase in testosterone was more rapid with AP extracts from boars than with extracts from castrated males. Observations from the current study imply structural alterations of FSH in the AP of Meishan males relative to crossbred males allowing sustained concentrations in the circulation, and this FSH possesses similar activation of the FSH receptor. The amount of LH in the AP extracts saturated the LH receptors of the hypophysectomized male rats, but some factor in extracts of boars differed from those of castrated males.