Hypothalamic transcriptional expression of the kisspeptin system and sex steroid receptors differs among polycystic ovary syndrome rat models with different endocrine phenotypes

OBJECTIVES: Polycystic ovary syndrome is a heterogeneous endocrine disorder that affects reproductive-age women. The mechanisms underlying the endocrine heterogeneity and neuroendocrinology of polycystic ovary syndrome are still unclear. In this study, we investigated the expression of the kisspeptin system and gonadotropin-releasing hormone pulse regulators in the hypothalamus as well as factors related to luteinizing hormone secretion in the pituitary of polycystic ovary syndrome rat models induced by testosterone or estradiol. METHODS: A single injection of testosterone propionate (1.25 mg) (n=10) or estradiol benzoate (0.5 mg) (n=10) was administered to female rats at 2 days of age to induce experimental polycystic ovary syndrome. Controls were injected with a vehicle (n=10). Animals were euthanized at 90-94 days of age, and the hypothalamus and pituitary gland were used for gene expression analysis. RESULTS: Rats exposed to testosterone exhibited increased transcriptional expression of the androgen receptor and estrogen receptor-β and reduced expression of kisspeptin in the hypothalamus. However, rats exposed to estradiol did not show any significant changes in hormone levels relative to controls but exhibited hypothalamic downregulation of kisspeptin, tachykinin 3 and estrogen receptor-α genes and upregulation of the gene that encodes the kisspeptin receptor. CONCLUSIONS: Testosterone- and estradiol-exposed rats with different endocrine phenotypes showed differential transcriptional expression of members of the kisspeptin system and sex steroid receptors in the hypothalamus. These differences might account for the different endocrine phenotypes found in testosterone- and estradiol-induced polycystic ovary syndrome rats.

Glial cell activity is maintained during prolonged inflammatory challenge in rats

We evaluated the expression of glial fibrillary acidic protein (GFAP), glutamine synthetase (GS), ionized calcium binding adaptor protein-1 (Iba-1), and ferritin in rats after single or repeated lipopolysaccharide (LPS) treatment, which is known to induce endotoxin tolerance and glial activation. Male Wistar rats (200-250 g) received ip injections of LPS (100 µg/kg) or saline for 6 days: 6 saline (N = 5), 5 saline + 1 LPS (N = 6) and 6 LPS (N = 6). After the sixth injection, the rats were perfused and the brains were collected for immunohistochemistry. After a single LPS dose, the number of GFAP-positive cells increased in the hypothalamic arcuate nucleus (ARC; 1 LPS: 35.6 ± 1.4 vs control: 23.1 ± 2.5) and hippocampus (1 LPS: 165.0 ± 3.0 vs control: 137.5 ± 2.5), and interestingly, 6 LPS injections further increased GFAP expression in these regions (ARC = 52.5 ± 4.3; hippocampus = 182.2 ± 4.1). We found a higher GS expression only in the hippocampus of the 6 LPS injections group (56.6 ± 0.8 vs 46.7 ± 1.9). Ferritin-positive cells increased similarly in the hippocampus of rats treated with a single (49.2 ± 1.7 vs 28.1 ± 1.9) or repeated (47.6 ± 1.1 vs 28.1 ± 1.9) LPS dose. Single LPS enhanced Iba-1 in the paraventricular nucleus (PVN: 92.8 ± 4.1 vs 65.2 ± 2.2) and hippocampus (99.4 ± 4.4 vs 73.8 ± 2.1), but had no effect in the retrochiasmatic nucleus (RCA) and ARC. Interestingly, 6 LPS increased the Iba-1 expression in these hypothalamic and hippocampal regions (RCA: 57.8 ± 4.6 vs 36.6 ± 2.2; ARC: 62.4 ± 6.0 vs 37.0 ± 2.2; PVN: 100.7 ± 4.4 vs 65.2 ± 2.2; hippocampus: 123.0 ± 3.8 vs 73.8 ± 2.1). The results suggest that repeated LPS treatment stimulates the expression of glial activation markers, protecting neuronal activity during prolonged inflammatory challenges.

Estradiol-induced hypophagia is associated with the differential mRNA expression of hypothalamic neuropeptides

Estradiol participates in the control of energy homeostasis, as demonstrated by an increase in food intake and in body weight gain after ovariectomy in rats. In the present study, female Wistar rats (200-230 g, N = 5-15 per group), with free access to chow, were individually housed in metabolic cages. We investigated food intake, body weight, plasma leptin levels, measured by specific radioimmunoassay, and the hypothalamic mRNA expression of orexigenic and anorexigenic neuropeptides, determined by real-time PCR, in ovariectomized rats with (OVX+E) and without (OVX) estradiol cypionate treatment (10 µg/kg body weight, sc, for 8 days). Hormonal and mRNA expression were determined at pre-feeding and 4 h after food intake. OVX+E rats showed lower food intake, less body weight gain and lower plasma leptin levels. In the OVX+E group, we also observed a reduction of neuropeptide Y (NPY), agouti-related protein (AgRP) and cocaine- and amphetamine-regulated transcript (CART) mRNA expression in the arcuate nucleus and a decrease in orexin A in the lateral hypothalamic area (LHA). There was an increase in leptin receptor (LepRb), melanocortin-4 receptor (MC4-R), CART, and mainly corticotropin-releasing hormone (CRH) mRNA in the paraventricular nucleus and LepRb and CART mRNA in the LHA. These data show that hypophagia induced by estradiol treatment is associated with reduced hypothalamic expression of orexigenic peptides such as NPY, AgRP and orexin A, and increased expression of the anorexigenic mediators MC4-R, LepRb and CRH. In conclusion, estradiol decreases food intake, and this effect seems to be mediated by peripheral factors such as leptin and the differential mRNA expression of neuropeptides in the hypothalamus.

The impact of antipsychotic drugs on food intake and body weight and on leptin levels in blood and hypothalamic ob-r leptin receptor expression in wistar rats

OBJECTIVES: The aim of our study was to investigate the impact of typical and atypical antipsychotic drugs on leptin concentration in blood and changes in the receptor expression in the hypothalamus of male Wistar rats. METHODS: From the age of 13 to 18 weeks, three groups of 20 animals were fed an average dose of 3.5 + 0.03 mg/ kg body weight (BW) haloperidol; 30.6 + 0.22 mg/kg BW clozapine; or 14.9 + 0.13 mg/kg BW ziprasidone in ground food pellets containing 15 percent fat. Twenty control animals received no drugs. Blood samples were taken at week 14, 16, and 19. Locomotor activity and exploratory behavior were measured using the alcove test at weeks 15 and 17. The expression of the hypothalamic leptin receptor in rat brains was determined by using a Western blot. RESULTS: Rats medicated with haloperidol and ziprasidone showed a significantly decreased percentage weight gain and food consumption. We observed no differences in the alcove test, but locomotor activity was significantly reduced in the haloperidol group. Except for rats in the clozapine and ziprasidone groups, after 2 weeks of drug application, we found no changes in the leptin blood concentrations among the four groups or animals within each group. Moreover, we did not find specific differences in hypothalamic leptin receptor expression among the groups. CONCLUSION: We concluded that in male Wistar rats during this treatment period, the tested drugs did not act directly on the leptin regulatory system. We recommend further studies using long-term treatment of different rat strains.

Effect of equine hypothalamic extract on hormone secretion by the adenohypophysis of rats

The effect of Equine Hypothalamic Extract (EHE) on pituitary weight and secretion of TSH, FSH-LH and ACTH was studied in the rat. The pituitary respponse to EHE was assessed by measuring (131) I uptake by the thyroid and by weight changes of the pituituray glands, thyroids, adrenals, ovaries and uteri. (131) I uptake of the thyroid and weights of the pituitary, thyroid, adrenal and uterus increased in the treated rats, whereas ovarian weights were similar to control groups. These findings indicate that the EHE containes hypophysiotropic peptides which can stimulate the secretion of pituitary hormones in the rat.

Cellular localization of atrial natriuretic peptide and tissue kallikrein in the human hypothalamus

In the kidney, renal atrial natriuretic peptide (ANP) is considered to play an important role wter and salt homeostasis. Immunoreactive ANP in the brain of lower invertebrates, such as the rat, has been shown to be localizaed in the hypothalamus and septum. Several studies have investigated the possibility of a regulatory system in the brain similar to that of the kidney. Since neuronal function is acutely sensitive to disturbances of the intracranial water and salt balance we have attempted to immunolocalize ANP-containing cells in the normal human hypothalamus, using a polyclonal antiserum specific to ANP. Also, we have observed tissue kallikrein (TK), using a polylonal antiserum specific to TK, in the same areas as ANP. A regulatory role for TK on prolactin has been suggested as the rationale for the co-localization of these two hormones in human prolactinomas. Therefore, it could be suggested that TK plays a similar role in the processing of precursor ANP in the brain. It is contemplated to examine the status of these peptides in patients with cerebral oeodema

Acetylcholinesterase release from crude mitochondrial fractions and superfused synaptosomes of rat hypothalamus / Liberatión de acetilcolinesterarasa de la fracción mitocondrial cruda y de sinaptosomas superfundidos de hipotalamo de rata

Crude mitochondrial fractions of rat hypothalamus, incubated in oxygenated Krebs solution (37ºC), released acetylcholinesterase (AChE) (increased of enzyme activity) into the medium, upon depolarization by 50 mM K+;this effect was absent in Ca²+ free medium. Superfused-synaptosomes obtained from subfractionated crude mitochondrial fractions of rat hypothalamus also released AChE, in a calcium-dependent fashion, when dependent fashion, when depolarized by 50 mM K+ or 50 µM veratridine. The veratridine-releasing effect was antagonized by 2 mM tetrodotoxin. Lactic dehydrogenase activity did not change in the medium with the introduction of depolarizing substances

Effect of progesterone on the hypothalamic cAMP concentration in estradiol: primed rats

El efecto de progesterona sobre la concentración de 3', 5' monofosfato cíclico de adenosina (cAMP) fue estudiado en el hipotálamo de ratas ovariectomizadas tratadas con estradiol. La inyección de progesterona (2mg/rata) en ratas ovariectomizadas, 3 días luego de la dosis de estradiol, produjo a la 4 h un aumento en el contenido hipotálamo del nucleótido: mientras que bajos niveles de cAMP se encontraron 24h después de la inyección de progesterona, lo cual sugiere una respuesta bifásica de la misma. La administración de dietilditiocarbamato, un bloqueante de la síntesis noradrenérgica, no previno el aumento de cAMP hipotalámico inducido por progesterona luego de 4 h de inyectada, el cual fue bloqueado por la administración de fenoxibenzamina, un a-adrenobloqueante, o propranolol, un ß-adrenobloqueante, suguiriendo la participación de receptores a y ß adrenérgicos. Puesto que no se obdervan cambios en la concentración hipotalámica de cAMP en los animales sacrificados por la tarde, se puede inferir que los efectos de progesterona sobre este parámetro podrían depender de la hora del día en que el esteroide fue administrado

Evidence for catecholaminergic control of *-melanotropin (*-MSH) content in hypothalamic areas

Se investigó una posible regulación catecolaminérgica sobre el alfa-MSH hipotalámico, en ratas machos mediante experimentación "in vivo". La hormona se dosó por radioinmunoensayo en tres regiones hipotalámicas: hipotálamo medial basal (HMB), área preóptica hipotalámica (APO) e hipotálamo dorso-lateral (HDL). Alfa-metil-para-tirosina (300mg/Kg), un inhibidor de la tirosina-hidroxilasa, incrementó el contenido de alfa-MSH en HMB y APO a las 22:00 h. Dietilditiocarbamato (600mg/Kg), que inhibe a la dopamina-b-hidroxilasa, como también 2-3-dicloro-metil-bencil-amida (25mg/Kg), que actua inhibiendo la enzima fenil-etanol-amina transferasa, incrementan el contenido de alfa-MSH en las áreas mencionadas. El antagonista de receptores alfa-adrenérgicos fenoxibenzamina (15mg/Kg) y el antagonista específico de receptores alfa-1, prazosín (1.0mg/Kg) producen el mismo efecto. Ninguna de las drogas experimentadas modificó el contenido de alfa-MSH en el HDL. Haloperidol (1.2mg/Kg), un antagonista de receptores dopaminérgicos, propanolol (6.0mg/Kg) y yohimbina (10mg/Kg) antagonista ß-adrenérgico (no selectivo) y alfa-2 adrenérgico, respectivamente no afectaron el contenido de alfa-MSH en ninguna de las áreas hipotalámicas. Estos resultados indican que el sistema catecolaminérgico estaría involucrado en el control del alfa-MSH hipotalámico derivado de la pro-opiomelanocortina a través de un receptor de tipo alfa-adrenérgico. Los datos sugieren que el mecanismo de control de los dos sistemas hipotámicos productores de alfa-MSH son diferentes