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Background: Aryl hydrocarbon receptor-interacting protein (AIP) is evolutionarily conserved and expressed widely throughout the organism. Loss-of-function AIP mutations predispose to young-onset pituitary adenomas. AIP co-localizes with growth hormone in normal and tumorous somatotroph secretory vesicles. AIP protein is detectable in circulation. We aimed to investigate possible AIP and GH co-secretion, by studying serum AIP and GH levels at baseline and after GH stimulation or suppression, in GH deficiency (GHD) and in acromegaly patients. Subjects and methods: Insulin tolerance test (ITT) was performed in GHD patients (n = 13) and age-BMI-matched normal GH axis control patients (n = 31). Oral glucose tolerance test (OGTT) was performed in active acromegaly patients (n = 26) and age-BMI-matched normal GH axis control patients (n = 18). In-house immunometric assay was developed for measuring circulating AIP. Results: Serum AIP levels were in the 0.1 ng/mL range independently of gender, age or BMI. Baseline AIP did not differ between GHD and non-GHD or between acromegaly and patients with no acromegaly. There was no change in peak, trough or area under the curve during OGTT or ITT. Serum AIP did not correlate with GH during ITT or OGTT. Conclusions: Human circulating serum AIP in vivo was assessed by a novel immunometric assay. AIP levels were independent of age, sex or BMI and unaffected by hypoglycaemia or hyperglycaemia. Despite co-localization in secretory vesicles, AIP and GH did not correlate at baseline or during GH stimulation or suppression tests. A platform of reliable serum AIP measurement is established for further research of its circulatory source, role and impact.

The adipose tissue has detrimental effects on growth hormone secretion, even in the absence of obesity. The majority of previous studies have shown an inverse relationship between fat mass and the growth hormone response to several stimulation tests in adults and in children. The contribution of body fat mass on growth hormone response to provocative tests during the transition age is not known . We analyzed the GHIGF1 axis by GHRH-arginine test in 30 healthy adolescents with normal stature during the transition period from 14 to 18 years. All subjects underwent body composition analysis by dualenergy X-ray absorptiometry (DXA). We found that total body fat mass was inversely correlated with peak GH to provocative test (r = - 0.6, p = 0.004). GH deficiency was shown in 2 of our healthy patients if diagnosis was based on GH peak below 19 μg/l. Both children who failed the GHRH-arginine were overweight (BMI for age above 85th percentile). However, their GH status was normal when assessed by insulin tolerance test. Multivariate analysis demonstrated strong correlation between peak stimulated GH and measures of body adiposity, including body mass index and fat mass index, with the latter showing the most important effect on GH secretion . Fat mass index alone explained 34.5 % of the variability in peak GH. This study has shown for the first time that during the transition period, GH response to GHRH-arginine test is strongly influenced by body composition, and cutoff values appropriate for overweight and obese adolescents are needed. Supporting information available online at http:// www.thieme-connect.de/ejournals/toc/hmr. [ABSTRACT FROM AUTHOR]

The stimulatory effects of ghrelin, a 28-AA acylated peptide originally isolated from stomach, on growth hormone (GH) secretion and feeding are exclusively mediated through the growth hormone secretagogue 1a receptor (GHS-R1a), the only ghrelin receptor described so far. Several GHS-R1a agonists and antagonists have been developed to treat metabolic or nutritional disorders but their mechanisms of action in the central nervous system remain poorly understood. In the present study, we compared the activity of BIM-28163, a GHS-R1a antagonist, and of several agonists, including native ghrelin and the potent synthetic agonist, BIM-28131, to modulate food intake, GH secretion, and cFos activity in arcuate nucleus (ArcN), nucleus tractus solitarius (NTS), and area postrema (AP) in wild-type and NPY-GFP mice. BIM-28131 was as effective as ghrelin in stimulating GH secretion, but more active than ghrelin in inducing feeding. It stimulated cFos activity similarly to ghrelin in the NTS and AP but was more powerful in the ArcN, suggesting that the super-agonist activity of BIM-28131 is mostly mediated in the ArcN. BIM-28163 antagonized ghrelin-induced GH secretion but not ghrelin-induced food consumption and cFos activation, rather it stimulated food intake and cFos activity without affecting GH secretion. The level of cFos activation was dependent on the region considered: BIM-28163 was as active as ghrelin in the NTS, but less active in the ArcN and AP. All compounds also induced cFos immunoreactivity in ArcN NPY neurons but BIM-28131 was the most active. In conclusion, these data demonstrate that two peptide analogs of ghrelin, BIM-28163, and BIM-28131, are powerful stimulators of appetite in mice, acting through pathways and key brain regions involved in the control of appetite that are only partially superimposable from those activated by ghrelin. A better understanding of the molecular pathways activated by these compounds could be useful in devising future therapeutic applications, such as for cachexia and anorexia. [ABSTRACT FROM AUTHOR]

Abstract: Background: The use of growth hormone (GH) in deficient adults has already been demonstrated to result in several benefits regarding metabolic parameters, body composition and quality of life. Due to the similarities between GH deficiency in adults and the aging process, the concept of somatopause has emerged. Objectives: Correlate the GH secretion profile in healthy men older than 50 years with anthropometric parameters and exercise capacity. Patients and methods: Twenty-nine healthy male were selected, with a mean age of 57.9±4.2 years (range 50–66). After hospital admission, body mass index (BMI), body composition (fat mass) and abdominal circumference, 24-h GH profile, GH peak and basal IGF-I were evaluated, and all the patients underwent a treadmill stress testing to estimate exercise capacity with the Bruce protocol, with evaluation of the maximum oxygen peak, maximum heart rate and METs. All the results are shown as mean±Std deviation: BMI −26.5±4.9kg/m2, percent fat mass −27.1±6.2%, abdominal circumference −92.1±10.1cm, 24h GH profile −0.3±0.2ng/dl, peak GH −2.5±2.0ng/dl, IGF-I −202.4±72.4ng/dl, maximum oxygen peak −31.9±6.8L, maximum heart rate – 161.4±7.5 bpm and METs – 9.1±1.9. After regression analysis using the GH secretion profile (mean GH in 24h, spontaneous peak GH and basal IGF-I) as dependent variable, no correlations were found between these and the other evaluated parameters. [Copyright &y& Elsevier]

The presence of ghrelin and its receptor, growth hormone (GH) secretagogue receptor, in the hypothalamus and pituitary, and its ability to stimulate GH release in vivo and in vitro, strongly support a significant role for this peptide in the control of somatotroph function. We previously demonstrated that ghrelin elicits GH secretion directly in somatotrophs by activating two major signalling cascades, which involve inositol phosphate and cAMP. In as much as nitric oxide (NO) and its mediator cGMP have been recently shown to contribute substantially to the response of somatotrophs to key regulatory hormones, including GH-releasing hormone, somatostatin and leptin, we investigated the possible role of this signalling pathway in ghrelin-induced GH release in vitro. Accordingly, cultures of pituitary cells from prepuberal female pigs were challenged with ghrelin (10−8 m, 30 min) in the absence or presence of activators or blockers of key steps of the NO synthase (NOS)/NO/guanylate cyclase (GC)/cGMP route and GH secretion was measured. Two distinct activators of the NO route, S-nitroso- N-acetylpenicillamine (SNAP) (5 × 10−4 m) andl-arginine methyl ester hydrochloride (l-AME) (10−3 m), comparably stimulated GH secretion when applied alone. The presence ofl-AME enhanced ghrelin-stimulated GH secretion, whereas SNAP did not alter its effect. Conversely, two different NOS/NO pathway inhibitors, Nw-nitro-l-arginine methyl ester hydrochloride (10−5 m) or haemoglobin (20 μg/ml), similarly blocked ghrelin-induced (but not basal) GH release, thus indicating that NO contributes critically to ghrelin action in somatotrophs. Moreover, incubation with a permeable cGMP analogue, 8-Br-cGMP (10−8 m) stimulated GH secretion, but did not modify the stimulatory action of ghrelin, suggesting that cGMP could mediate the action of NO. Indeed, inhibition of GC by 10 μm LY-53,583 did not alter basal GH secretion but abolished the GH-releasing action of ghrelin. Taken together, our results provide novel evidence indicating that ghrelin requires activation of the NOS/NO route, and its subsequent GC/cGMP signal transduction pathway, as necessary steps to induce GH secretion from somatotrophs. [ABSTRACT FROM AUTHOR]

Background: Familial isolated growth hormone deficiency (IGHD) is a disorder with about 5–30% of patients having affected relatives. Among those familial types, IGHD type II is an autosomal dominant form of short stature, associated in some families with mutations that result in missplicing to produce del32–71-GH, a GH peptide which cannot fold properly. The mechanism by which this mutant GH may alter the controlled secretory pathway and therefore suppress the secretion of the normal 22-kDa GH product of the normal allele is not known in detail. Previous studies have shown variance depending on cell type, transfection technique used, as well as on the method of analysis performed. Aim: The aim of our study was to analyse and compare the subcellular distribution/localization of del32–71-GH or wild-type (wt)-GH (22-kDa GH), each stably transfected into AtT-20, a mouse pituitary cell line endogenously producing ACTH, employed as the internal control for secretion assessment. Methods: Co-localizationof wt- and del32–71 mutant GH form was studied by quantitative confocal microscopy analysis. Using the immunofluorescent technique, cells were double stained for GH plus one of the following organelles: endoplasmic reticulum (ER anti-Grp94), Golgi (anti-βCOP) or secretory granules (anti-Rab3a). In addition, GH secretion and cell viability were analysed in detail. Results/Conclusions: Our results show that in AtT-20 neuroendocrine cells, in comparison to the wt-GH, the del32–71-GH has a major impact on the secretory pathway not only affecting GH but also other peptides such as ACTH. The del32–71-GH is still present at the secretory vesicles’ level, albeit in reduced quantity when compared to wt-GH but, importantly, was secretion-deficient. Furthermore, while focusing on cell viability an additional finding presented that the various splice site mutations, even though leading eventually to the same end product, namely del32–71-GH, have different and specific consequences on cell viability and proliferation rate. Copyright © 2006 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Abstract: We assessed the inhibitory effects of butyrate on the growth hormone (GH) secretion in order to investigate the cellular mechanisms in rat somatotrophs. Isolated anterior pituitary cells were cultured in DMEM for several hours, either in the presence (1, 3, or 10mM) or absence of butyrate, and then stimulated with 10−7 M GHRH for 30min, in the presence of butyrate at the concentrations used for the previous culture. The increase in GHRH-induced GH release was significantly reduced in a time-dependent and concentration-dependent manner in the cells previously cultured with butyrate. GH content (the sum of GH released into the medium induced by GHRH stimulation and the GH remaining in the cells after stimulation) was reduced by the culture of cells in the presence of butyrate, which was also inversely dependent on the concentrations used for the culture. Simultaneous addition of an L-type Ca2+ channel blocker, nifedipine (10pM), to the medium during 10−9 M GHRH stimulation significantly reduced the stimulated GH release, which was further significantly decreased by a simultaneous addition of 10mM butyrate. Butyrate blunted the GHRH (10−9 M)-induced increase in cellular cyclic AMP and calcium ion concentrations, the activity of protein kinases (A and C), and GHmRNA expression. The expression of mRNA for GPR 41 and 43, known as receptors for short-chain fatty acids, was confirmed in the anterior pituitary cells. These findings suggest that butyrate inhibits GHRH-induced GH release as well as GH production, and the cellular inhibitory actions of butyrate occur in diverse cellular signaling pathways of rat somatotrophs. [Copyright &y& Elsevier]

There is increasing evidence that nitric oxide (NO) produced by NO synthase (NOS), and their signalling partners, guanylyl cyclase and cGMP, play a relevant role in growth hormone (GH) secretion from somatotrophs. We previously demonstrated that both GH-releasing hormone (GHRH; 10−8 M) and low concentrations of somatostatin (10−15 M) stimulate pig GH release in vitro, whereas a high somatostatin concentration (10−7 M) inhibits GHRH-induced GH secretion. To ascertain the possible contribution of the NOS-NO and guanylyl cyclase-cGMP routes to these responses, cultures of pituitary cells from prepubertal female pigs were treated (30 min) with GHRH (10−8 M) or somatostatin (10−7 or 10−15 M) in the absence or presence of activators or blockers of key steps of these signalling cascades, and GH release was measured. Two distinct activators of NO route, SNAP (5 × 10−4 M) or L-AME (10−3 M), similarly stimulated GH release when applied alone (with this effect being blocked by 10−7 M somatostatin), but did not alter the stimulatory effect of GHRH or 10−15 M somatostatin. Conversely, two NO pathway inhibitors, NAME (10−5 M) or haemoglobin (20 µg/ml) similarly blocked GHRH- or 10−15 M somatostatin-stimulated GH release. 8-Br-cGMP (10−8 to 10−4 M) strongly stimulated GH release, suggesting that cGMP may function as a subsequent step in the NO pathway in this system. Interestingly, 10−7 M somatostatin did not inhibit the stimulatory effect of 8-Br-cGMP. Moreover, although 8-Br-cGMP did not modify the effect of GHRH, it enhanced GH release stimulated by 10−15 M somatostatin. Accordingly, a specific guanylyl cyclase inhibitor, LY-83, 583 (10−5 M) did not alter 10−15 M somatostatin-induced GH release, whereas it blocked GHRH-induced GH secretion. These results demonstrate for the first time that the NOS/NO signalling pathway contributes critically to the stimulatory effects of both GHRH and low-concentration somatostatin on GH release, and that, conversely, the subsequent guanylyl cyclase/cGMP step only mediates GHRH- and not low-concentration somatostatin-induced GH secretion from somatotrophs. [ABSTRACT FROM AUTHOR]

Either in children or in adults, arginine (ARG) alone and combined with GHRH (GHRH + ARG) are reliable tests for the diagnosis of GH deficiency. The procedures of these tests generally include GH measurement every 15 min from baseline up to 90–120 min. Aim of our study was to verify if the procedure of these tests could be usefully shortened in clinical practice. To this goal we have studied 173 normally growing children and adolescents (C, 117 M and 56 F, age: 11.3 ± 0.4 yr.) and 125 young and middle aged normal adults (A, 68 M and 57 F, age: 30.0 ± 0.6 yr.). ARG alone test was performed by 81 C and 33 A (0.5 g/kg arginine, iv, from 0 to +30 min, up to a maximum of 30 g) while GHRH (1 μg/kg iv bolus at 0 min) + ARG test was performed by 92 C and 92 A. After ARG alone, taking into account data from +15 to +105 min, GH values above the 3rd centile limit of arbitrary cut-off (7 or 10 μg/1 in C and 5 μg/1 in A) occurred in 85% or 64% and 94% subjects, respectively. After GHRH + ARG test, taking into account only data at +30, +45, +60 min GH values above the 3rd centile limit (20 μg/1 in C and 16.5 μg/1 in A) occurred in 99% of subjects in both groups. Taking into account only these 3 timing points, the percentage of GH peak above the third centile limits after ARG alone was never higher than 60% in C and 85% in A. In conclusion, this study shows that single GHRH + arginine test can be reliably performed in a shortened procedure which makes easier the clinical practice and further reduces costs. [ABSTRACT FROM AUTHOR]

Cis-unsaturated free fatty acids (FFA) like oleic acid are strong blockers of both basal and stimulated GH secretion in vivo by acting directly on the somatotroph cell. Several lines of evidence suggest that this inhibitory action is the result of a perturbation of the function of several plasma membrane integral proteins. It has been reported recently that cis-FFA are able to block several steps in the inositolphosphates/phospholipase C/Ca2+ (InsPs/PLC/Ca2+) signal transduction pathway triggered by the activation of the TRH receptor. In this paper we present evidence showing that the inhibition of growth hormone (GH) and prolactin (PRL) secretion by cis-FFA in vitro is also exerted at several different levels on the cAMP-protein kinase A (cAMP/PKA) pathway triggered by the stimulation of the vasoactive intestinal peptide (VIP) receptor in pituitary clonal cells. By means of a sequential analysis of signal transduction events, we observed that cis-unsaturated FFA: (1) reduce the activity of adenylate cyclase; (2) perturb the activity of protein kinase A; (3) suppress the VIP-triggered Ca2+ influx, and (4) do not perturb VIP binding or the homologous desensitization of the VIP receptor. [ABSTRACT FROM AUTHOR]

Ghrelin, a recently discovered 28-aa peptide, stimulates GH release through a mechanism involving PLC- and cAMP-related signaling pathways. Recently, nitric oxide (NO) and its mediator, cGMP, have been shown to be required for the response of somatotropes to various regulators (GHRH, somatostatin, leptin). Here, we explore the possible role of the NO synthase (NOS)/NO/guanylate cyclase (GC)/cGMP signaling pathway in ghrelininduced GH release from cultured pig somatotropes using blockers or activators of this route. [ABSTRACT FROM AUTHOR]

Abstract: A chronic treatment with a cholinesterase inhibitor, donepezil (0.085mg/kg/h for 30 days) increases significantly the number and amplitude of growth hormone (GH) pulses in 3- and 24-month-old rats without modifying nadir GH values. This treatment does not reduce age-related alterations in sleep/wake cycle but it increases immobility-related high-voltage spindles (HVS) in old animals. These data suggest that cholinergic mechanisms involved in age-related alterations in GH and sleep regulation are different. [Copyright &y& Elsevier]