mRNA expression of corticotropin-releasing factor and urocortin 1 after restraint and foot shock together with alprazolam administration

Corticotropin-releasing factor (CRF) is expressed in the paraventricular nucleus of the hypothalamus (PVN), and act centrally to provoke stress-like autonomic and behavioral responses. Urocortins 1-3 are additional ligands to the CRF receptors 1 and 2. Ucn 1 neurons are primarily concentrated in the Edinger-Westphal (EW) nucleus and also have been associated with stress responses. It is also known that UCN 1 respond in different ways depending on the stressor presented. Benzodiazepines can act via the CRF peptidergic system and chronic administration of alprazolam does not interfere with CRF mRNA expression in the PVN, but significantly increase Ucn 1 mRNA expression in the EW. The aim of our study was to investigate the relationship between different stressor stimuli, foot shock (FS) and restraint (R), and the mRNA expression of CRF and Ucn 1 in the PVN and EW using alprazolam (A). We employed fos activation and in situ hybridization. Restraint group presented increased fos-ir and CRF mRNA expression in the PVN compared to FS group. The stress responses of R group were prevented by A. In the EW,fos-ir was higher in the FS group than in the R group, whereas Ucn 1 mRNA expression was higher in the R group than in the FS group. Alprazolam significantly increased fos-ir and Ucn 1 mRNA expression in both groups. Our results show that PVN and EW respond in different ways to the same stressors. Furthermore, EW of stressed animals replies in a complementary way comparing to PVN with the use of Alprazolam. (C) 2010 Elsevier Inc. All rights reserved.

Potential Role of Growth Hormone in Impairment of Insulin Signaling in Skeletal Muscle, Adipose Tissue, and Liver of Rats Chronically Treated with Arginine

We have shown that rats chronically treated with Arginine (Arg), although normoglycemic, exhibit hyperinsulinemia and decreased blood glucose disappearance rate after an insulin challenge. Attempting to investigate the processes underlying these alterations, male Wistar rats were treated with Arg (35 mg/d), in drinking water, for 4 wk. Rats were then acutely stimulated with insulin, and the soleus and extensorum digitalis longus muscles, white adipose tissue (WAT), and liver were excised for total and/or phosphorylated insulin receptor (IR), IR substrate 1/2, Akt, Janus kinase 2, signal transducer and activator of transcription (STAT) 1/3/5, and p85 alpha/55 alpha determination. Muscles and WAT were also used for plasma membrane (PM) and microsome evaluation of glucose transporter (GLUT) 4 content. Pituitary GH mRNA, GH, and liver IGF-I mRNA expression were estimated. It was shown that Arg treatment: 1) did not affect phosphotyrosine-IR, whereas it decreased phosphotyrosine-IR substrate 1/2 and phosphoserine-Akt content in all tissues studied, indicating that insulin signaling is impaired at post-receptor level; 2) decreased PM GLUT4 content in both muscles and WAT; 3) increased the pituitary GH mRNA, GH, and liver IGF-I mRNA expression, the levels of phosphotyrosine-STAT5 in both muscles, phosphotyrosine-Janus kinase 2 in extensorum digitalis longus, phosphotyrosine-STAT3 in liver, and WAT as well as total p85 alpha in soleus, indicating that GH signaling is enhanced in these tissues; and 4) increased p55 alpha total content in muscles, WAT, and liver. The present findings provide the molecular mechanisms by which insulin resistance and, by extension, reduced GLUT4 content in PM of muscles and WAT take place after chronic administration of Arg, and further suggest a putative role for GH in its genesis, considering its diabetogenic effect. (Endocrinology 150: 2080-2086, 2009)

Chronic supplementation of beta-hydroxy-beta methylbutyrate (HM beta) increases the activity of the GH/IGF-I axis and induces hyperinsulinemia in rats

Objective: Beta-hydroxy-beta-methylbutyrate (HM beta) is a metabolite of leucine widely used for improving sports performance. Although limp is recognized to promote anabolic or anti-catabolic effects on protein metabolism, the impact of its long-term use on skeletal muscle and/or genes that control the skeletal protein balance is not fully known. This study aimed to investigate whether chronic HM beta treatment affects the activity of GH/IGF-I axis and skeletal muscle IGF-I and myostatin mRNA expression. Design: Rats were treated with HK beta (320 mg/kg BW) or vehicle, by gavage, for 4 weeks, and killed by decapitation. Blood was collected for evaluation of serum insulin, glucose and IGF-I concentrations. Samples of pituitary, liver, extensor digitorum longus (EDL) and soleus muscles were collected for total RNA or protein extraction to evaluate the expression of pituitary growth hormone (GH) gene (mRNA and protein), hepatic insulin-like growth factor I (IGF-I) mRNA, skeletal muscle IGF-I and myostatin mRNA by Northern blotting/real time-PCR, or Western blotting. Results: Chronic HM beta treatment increased the content of pituitary GH mRNA and GH, hepatic IGF-I mRNA and serum IGF-I concentration. No changes were detected on skeletal muscle IGF-I and myostatin mRNA expression. However, the HIM-treated rats although normoglycemic, exhibited hyperinsulinemia. Conclusions: The data presented herein extend the body of evidence on the potential role of HM beta-treatment in stimulating GH/IGF-I axis activity. In spite of this effect, HM beta supplementation also induces an apparent insulin resistance state which might limit the beneficial aspects of the former results, at least in rats under normal nutritional status and health conditions. (C) 2010 Growth Hormone Research Society. Published by Elsevier Ltd. All rights reserved.

T3 acutely increases GH mRNA translation rate and GH secretion in hypothyroid rats

Cytoskeleton controls the stability of transcripts, by mechanisms that involve mRNAs and eEF1A attachment to it. Besides, it plays a key role in protein synthesis and secretion, which seems to be impaired in somatotrophs of hypothyroid rats, whose cytoskeleton is disarranged. This study investigated the: eEF1A and GH mRNA binding to cytoskeleton plus GH mRNA translation rate and GH secretion, in sham-operated and thyroidectomized rats treated with T3 or saline, and killed 30 min thereafter. Thyroidectomy reduced: (a) pituitary F-actin content, and eEF1A plus GH mRNA binding to it; (b) GH mRNA recruitment to polysome; and (c) liver IGF-1 mRNA expression, indicating that GH mRNA stability and translation rate, as well as GH secretion were impaired. T3 acutely reversed all these changes, which points toward a nongenomic action of T3 on cytoskeleton rearrangement, which might contribute to the increase on GH mRNA translation rate and GH secretion. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Reduced maximal oxygen consumption and overproduction of proinflammatory cytokines in athletes

Objective: It was the aim of this study to evaluate whether chronic pain in athletes is related to performance, measured by the maximum oxygen consumption and production of hormones and cytokines. Methods: Fifty-five athletes with a mean age of 31.9 +/- 4.2 years engaged in regular competition and showing no symptoms of acute inflammation, particularly fever, were studied. They were divided into 2 subgroups according to the occurrence of pain. Plasma concentrations of adrenaline, noradrenaline, cortisol, prolactin, growth hormone and dopamine were measured by radioimmunoassay, and the production of the cytokines interleukin (IL)-1, IL-2, IL-4, IL-6, tumor necrosis factor-alpha, interferon-alpha and prostaglandin E-2 by whole-blood culture. Maximal oxygen consumption was determined during an incremental treadmill test. Results: There was no change in the concentration of stress hormones, but the athletes with chronic pain showed a reduction in maximum oxygen consumption (22%) and total consumption at the anaerobic threshold (25%), as well as increased cytokine production. Increases of 2.7-, 8.1-, 1.7- and 3.7-fold were observed for IL-1, IL-2, tumor necrosis factor-alpha and interferon-alpha, respectively. Conclusions: Our data show that athletes with chronic pain have enhanced production of proinflammatory cytokines and lipid mediators and reduced performance in the ergospirometric test. Copyright (c) 2008 S. Karger AG, Basel.

Neutral Aminopeptidase and Dipeptidyl Peptidase IV Activities in Plasma of Monosodium Glutamate Obese and Food-deprived Rats

Biometric parameters, glycemia and activity levels of plasma neutral aminopeptidase (APN) and dipeptidyl peptidase IV (DPPIV) were measured in monosodium glutamate obese and food-deprived rats (MSG-FD), to analyze the involvement of these enzymes in such situations. Plasma APN was distinguished as sensitive (PSA) (K(m) = 7.8 x 10(-5) mol/l) and predominantly insensitive (APM) (K(m) = 21.6 x 10(-5) mol/l) to puromycin, whereas DPPIV was sensitive (DPPIV-DS) (K(m) = 0.24 x 10(-5) mol/l) and predominantly insensitive (DPPIV-DI) (K(m) = 7.04 x 10(-5) mol/l) to diprotin A. Although unchanged in the MSG and food-deprived animals, APM activity levels were closely correlated with body mass, Lee index, and mass of retroperitoneal fat pad in the food deprived, but not in the MSG animals. DPPIV-DI activity levels decreased by 33% and were correlated with body mass, Lee index, and mass of periepididymal fat pad in the food-deprived MSG rats. These data suggest that APM and DPPIV-DI are respectively related to the downregulation of somatostatin in food-deprived rats, and to the recovery of energy balance in MSG obese rats during food deprivation.

Forebrain projections to brainstem nuclei involved in the control of mandibular movements in rats

Mandibular movements occur through the triggering of trigeminal motoneurons. Aberrant movements by orofacial muscles are characteristic of orofacial motor disorders, such as nocturnal bruxism (clenching or grinding of the dentition during sleep). Previous studies have suggested that autonomic changes occur during bruxism episodes. Although it is known that emotional responses increase jaw movement, the brain pathways linking forebrain limbic nuclei and the trigeminal motor nucleus remain unclear. Here we show that neurons in the lateral hypothalamic area, in the central nucleus of the amygdala, and in the parasubthalamic nucleus, project to the trigeminal motor nucleus or to reticular regions around the motor nucleus (Regio h) and in the mesencephalic trigeminal nucleus. We observed orexin co-expression in neurons projecting from the lateral hypothalamic area to the trigeminal motor nucleus. In the central nucleus of the amygdala, neurons projecting to the trigeminal motor nucleus are innervated by corticotrophin-releasing factor immunoreactive fibers. We also observed that the mesencephalic trigeminal nucleus receives dense innervation from orexin and corticotrophin-releasing factor immunoreactive fibers. Therefore, forebrain nuclei related to autonomic control and stress responses might influence the activity of trigeminal motor neurons and consequently play a role in the physiopathology of nocturnal bruxism.

Immunocytochemical identification of adenohypophyseal cells in the pirarucu (Arapaima gigas), an Amazonian basal teleost

The adenohypophysis (AH) of juvenile pirarucu (Arapaima gigas), a representative species of the Osteoglossomorpha (bonytongue fishes, one of the oldest living groups of the teleosts), was studied using histochemical and immunocytochemical methods. The AH is comprised of the pars distalis (PD), without a clear distinction between rostral pars distalis (RPD) and proximal pars distalis (PPD), and the pars intermedia (PI). The neurohypophysis (NH) is positioned on top of the PD and penetrates and branches into the PI. In the most rostral dorsal portion of the PD, adrenocorticotropic cells and fusiform gonadotropic cells were found. In the central PD, scarce prolactin-producing cells and growth-hormone-producing cells were located mainly in the dorsal part, whereas round gonadotropic cells were abundant in the ventral portion of this region. Human thyrotropin immunoreactive cells were not found in the entire AH. In the PI, melanotropic, some adrenocorticotropic, and somatolactin-producing cells were located intermingled surrounding the neurohypophyseal branches. Our results showed that the A. gigas pituitary has some basal characteristics between the ancient Actinopterygii and the more derived teleosts.

Dark-induced hormone changes coincide with the resumption of light-inhibited shoot growth in Catasetum fimbriatum (Orchidaceae)

Catasetum fimbriatum plants cultivated in the absence of light exhibit continuous shoot growth leading to the formation of nodes and internodes. On the other hand, when these plants are incubated in the presence of light, shoot longitudinal growth is inhibited and pseudobulbs develop just below the shoot apical meristem. These facts provide evidence of a possible influence of light on mitotic cell division in the shoot apex as well as on pseudobulb initiation. The effects of light and dark on the interruption and/or maintenance of shoot apex mitotic activity and the subsequent formation of pseudobulbs in the sub-meristematic regions were investigated by means of histological and hormonal studies. The interruption of shoot apex development occurred around the 150th d of light incubation and seems to have resulted from the establishment of a strong storage sink in the region of the future pseudobulb, in detriment to the continuous activity of the shoot apical meristem. The reduced total cytokinins/IAA ratio in the apex, mainly due to high levels of IAA, could be a key factor in the interruption of cell divisions. Transfer to the dark brings about the resumption of shoot apex development of plants through the re-entrance of cells in the cell cycle which coincides with a significant increase in the total cytokinins/IAA ratio. (C) 2009 Elsevier GmbH. All rights reserved.

The importance of hormone receptor analysis in osteosarcoma cells growth submitted to treatment with estrogen in association with thyroid hormone

Bone tumor incidence in women peaks at age 50-60, coinciding with the menopause. That estrogen (E2) and triiodothyronine (T3) interact in bone metabolism has been well established. However, few data on the action of these hormones are available. Our purpose was to determine the role of E2 and T3 in the expression of bone activity markers, namely alkaline phosphatase (AP) and receptor activator of nuclear factor kappa B ligand (RANKL). Two osteosarcoma cell lines: MG-63 (which has both estrogen (ER) and thyroid hormone (TR) receptors) and SaOs-29 (ER receptors only) were treated with infraphysiological E2 associated with T3 at infraphysiological, physiological, and supraphysiological concentrations. Real-time RT-PCR was used for expression analysis. Our results show that, in MG-63 cells, infraphysiological E2 associated with supraphysiological T3 increases AP expression and decreases RANKL expression, while infraphysiological E2 associated with either physiological or supraphysiological T3 decreases both AP and RANKL expression. On the other hand, in SaOs-2 cells, the same hormone combinations had no significant effect on the markers` expression. Thus, the analysis of hormone receptors was shown to be crucial for the assessment of tumor potential growth in the face of hormonal changes. Special care should be provided to patients with T3 and E2 hormone receptors that may increase tumor growth. Copyright (C) 2007 John Wiley & Sons, Ltd.