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International audience; BackgroundConstitutional thinness (CT), a non‐malnourished underweight state with no eating disorders, is characterized by weight gain resistance to high fat diet. Data issued from muscle biopsies suggested blunted anabolic mechanisms in free‐living state. Weight and metabolic responses to protein caloric supplementation has not been yet explored in CT.MethodsA 2 week overfeeding (additional 600 kcal, 30 g protein, 72 g carbohydrate, and 21 g fat) was performed to compare two groups of CTs (12 women and 11 men) to normal‐weight controls (12 women and 10 men). Bodyweight, food intake, energy expenditure, body composition, nitrogen balance, appetite hormones profiles, and urine metabolome were monitored before and after overfeeding.ResultsBefore overfeeding, positive energy gap was found in both CT genders (309 ± 370 kcal in CT‐F and 332 ± 709 kcal in CT‐M) associated with higher relative protein intake per kilo (1.74 ± 0.32 g/kg/day in CT‐F vs. 1.16 ± 0.23 in C‐F, P < 0.0001; 1.56 ± 0.36 in CT‐M vs. 1.22 ± 0.32 in C‐M, P = 0.03), lower nitrogen (7.26 ± 2.36 g/day in CT‐F vs. 11.41 ± 3.64 in C‐F, P = 0.003; 9.70 ± 3.85 in CT‐M vs. 14.14 ± 4.19 in C‐M, P = 0.02), but higher essential amino acids urinary excretion (CT/C fold change of 1.13 for leucine and 1.14 for arginine) in free‐living conditions. After overfeeding, CTs presented an accentuated positive energy gap, still higher than in controls (675 ± 540 in CTs vs. 379 ± 427 in C, P = 0.04). Increase in lean mass was induced in both controls genders but not in CTs (a trend was noticed in CT women), despite a similar nitrogen balance after overfeeding (5.06 ± 4.33 g/day in CTs vs. 4.28 ± 3.15 in controls, P = 0.49). Higher anorectic gut hormones' tone, glucagon‐like peptide 1 and peptide tyrosine tyrosine, during test meal and higher snacking frequency were noticed before and after overfeeding in CTs.ConclusionsThe blunted muscle energy mechanism, previously described in CTs in free‐living state, is associated with basal saturated protein turn over suggested by the concordance of positive nitrogen balance and an increased urine excretion of several essential amino acids. This saturation cannot be overpassed by increasing this spontaneous high‐protein intake suggesting a resistance to lean mass gain in CT phenotype.

Introduction: While the vast majority of research investigating the role of ghrelin or its receptor, GHS-R1a, in growth, feeding, and metabolism has been conducted in male rodents, very little is known about sex differences in this system. Furthermore, the role of GHS-R1a signaling in the control of pulsatile GH secretion and its link with growth or metabolic parameters has never been characterized. Methods: We assessed the sex-specific contribution of GHS-R1a signaling in the activity of the GH/IGF-1 axis, metabolic parameters, and feeding behavior in adolescent (5–6 weeks old) or adult (10–19 weeks old) GHS-R KO (Ghsr−/−) and WT (Ghsr+/+) male and female mice. Results: Adult Ghsr−/− male and female mice displayed deficits in weight and linear growth that were correlated with reduced GH pituitary contents in males only. GHS-R1a deletion was associated with reduced meal frequency and increased meal intervals, as well as reduced hypothalamic GHRH and NPY mRNA in males, not females. In adult, GH release from Ghsr−/− mice pituitary explants ex vivo was reduced independently of the sex. However, in vivo pulsatile GH secretion decreased in adult but not adolescent Ghsr−/− females, while in males, GHS-R1a deletion was associated with reduction in pulsatile GH secretion during adolescence exclusively. In males, linear growth did not correlate with pulsatile GH secretion, but rather with ApEn, a measure that reflects irregularity of the rhythmic secretion. Fat mass, plasma leptin concentrations, or ambulatory activity did not predict differences in GH secretion. Discussion/Conclusion: These results point to a sex-dependent dimorphic effect of GHS-R1a signaling to modulate pulsatile GH secretion and meal pattern in mice with different compensatory mechanisms occurring in the hypothalamus of adult males and females after GHS-R1a deletion. Altogether, we show that GHS-R1a signaling plays a more critical role in the regulation of pulsatile GH secretion during adolescence in males and adulthood in females.

This study investigated whether sst2 gene deletion interacts with age and chronic stress exposure to produce exacerbated emotional and cognitive ageing. Middle-aged (10–12 month) sst2 knockout (sst2KO) and wild-type (WT) mice underwent an unpredictable chronic mild stress (UCMS) procedure for 6 weeks or no stress for control groups. This was followed by a battery of tests to assess emotional and cognitive functions and neuroendocrine status (CORT level). A re-evaluation was performed 6 months later (i.e. with 18-month-old mice). UCMS reproduced neuroendocrine and behavioral features of stress-related disorders such as elevated circulating CORT levels, physical deteriorations, increased anxiety- and depressive-like behaviors and working memory impairments. sst2KO mice displayed behavioral alterations which were similar to stressed WT and exhibited exacerbated changes following UCMS exposure. The evaluations performed in the older mice showed significant long-term effects of UCMS exposure. Old sst2KO mice previously exposed to UCMS exhibited spatial learning and memory accuracy impairments and high levels of anxiety-like behaviors which drastically added to the effects of normal ageing. Spatial abilities and emotionality scores (mean z-scores) measured both at the UCMS outcome and 6 months later were correlated with the initially measured CORT levels in middle-age. The present findings indicate that the deletion of the sst2 receptor gene produces chronic hypercorticosteronemia and exacerbates sensitivity to stressors which over time, have consequences on ageing brain function processes.

International audience; Age-associated cognitive impairment is a major health and social issue because of increasing aged population. Cognitive decline is not homogeneous in humans and the determinants leading to differences between subjects are not fully understood. In middle-aged healthy humans, fasting blood glucose levels in the upper normal range are associated with memory impairment and cerebral atrophy. Due to a close evolutional similarity to Man, non-human primates may be useful to investigate the relationships between glucose homeostasis, cognitive deficits and structural brain alterations. In the grey mouse lemur, $Microcebus\ murinus$, spatial memory deficits have been associated with age and cerebral atrophy but the origin of these alterations have not been clearly identified. Herein, we showed that, on 28 female grey mouse lemurs (age range 2.4-6.1 years-old), age correlated with impaired fasting blood glucose (r$_s$=0.37) but not with impaired glucose tolerance or insulin resistance. In middle-aged animals (4.1-6.1 years-old), fasting blood glucose was inversely and closely linked with spatial memory performance (r$_s$=0.56) and hippocampus (r$_s$=-0.62) or septum (r$_s$=-0.55) volumes. These findings corroborate observations in humans and further support the grey mouse lemur as a natural model to unravel mechanisms which link impaired glucose homeostasis, brain atrophy and cognitive processes.

International audience; A combination of low cerebrospinal fluid (CSF) Amyloid β1–42 (Aβ1–42) and high Total-Tau (T-Tau) and Phosphorylated-Tau (P-Tau) occurs at a prodromal stage of Alzheimer’s disease (AD) and recent findings suggest that network abnormalities and interneurons dysfunction contribute to cognitive deficits. Somatostatin (SOM) and Neuropeptide Y (NPY) are two neuropeptides which are expressed in GABAergic interneurons with different fates in AD the former only being markedly affected. The aim of this study was to analyze CSF SOM, NPY and CSF Aβ1–42; T-Tau, P-Tau relationships in 43 elderly mild cognitively impairment (MCI) participants from the Biomarker of AmyLoïd pepTide and AlZheimer’s disease Risk (BALTAZAR) cohort. In these samples, CSF SOM and CSF Aβ1–42 on the one hand, and CSF NPY and CSF T-Tau and P-Tau on the other hand are positively correlated. CSF SOM and NPY concentrations should be further investigated to determine if they can stand for early AD biomarkers.Clinical Trial Registration: www.ClinicalTrials.gov, identifier #NCT01315639.

Background Constitutional thinness (CT) is a natural state of underweight (13–17.5 kg/m2) without the presence of any eating disorders and abnormal hormonal profile, and with preserved menses in women. We previously conducted a four-week fat overfeeding study showing weight gain resistance in CT women and one of our main results was the identification of an energy gap: a positive energy balance (higher energy intake than energy expenditure). Objective This new overfeeding study is designed to confirm the energy gap and propose mechanistic hypothesis. Design A 2-week overfeeding (daily consumption of one bottle of Renutryl® Booster (600 kcal, 30 g protein, 72 g carbohydrate, 21 g fat) on top of the dietary intake) is performed to compare 15 women and men in each CT group (Body Mass Index [BMI] Results and conclusions Data inter-linking will be able to be established with results of this study. The findings could possibly open to therapeutic approaches to help CT patients to gain weight as well as provide a better understanding of energy regulation with regard to treat obesity (resistance to weight loss), a mirror image of CT (resistance to weight gain).

Ghrelin is a potent orexigenic peptide hormone that acts through the growth hormone secretagogue receptor (GHSR), a G protein-coupled receptor highly expressed in the hypothalamus. In vitro studies have shown that GHSR displays a high constitutive activity, whose physiological relevance is uncertain. As GHSR gene expression in the hypothalamus is known to increase in fasting conditions, we tested the hypothesis that constitutive GHSR activity at the hypothalamic level drives the fasting-induced hyperphagia. We found that refed wild-type (WT) mice displayed a robust hyperphagia that continued for 5 days after refeeding and changed their food intake daily pattern. Fasted WT mice showed an increase in plasma ghrelin levels, as well as in GHSR expression levels and ghrelin binding sites in the hypothalamic arcuate nucleus. When fasting-refeeding responses were evaluated in ghrelin- or GHSR-deficient mice, only the latter displayed an ∼15% smaller hyperphagia, compared with WT mice. Finally, fasting-induced hyperphagia of WT mice was significantly smaller in mice centrally treated with the GHSR inverse agonist K-(D-1-Nal)-FwLL-NH2, compared with mice treated with vehicle, whereas it was unaffected in mice centrally treated with the GHSR antagonists D-Lys3-growth hormone-releasing peptide 6 or JMV2959. Taken together, genetic models and pharmacological results support the notion that constitutive GHSR activity modulates the magnitude of the compensatory hyperphagia triggered by fasting. Thus, the hypothalamic GHSR signaling system could affect the set point of daily food intake, independently of plasma ghrelin levels, in situations of negative energy balance., Instituto Multidisciplinario de Biología Celular

International audience; The neuropeptide somatostatin (SOM) is widely expressed in rodent brain and somatostatin-IRES-Cre (SOM-cre) mouse strains are increasingly used to unravel the physiology of SOM-containing neurons. However, while knock-in targeting strategy greatly improves Cre-Lox system accuracy, recent reports have shown that genomic insertion of Cre construct per se can markedly affect physiological function. We show that Cre transgene insertion into the 3'UTR of the somatostatin gene leads to the selective and massive depletion of endogenous SOM in all tested brain regions. It also strongly impacts SOM-related neuroendocrine responses in a similar manner to what has been reported for SST KO mice: increased corticosterone levels after 30-min restraint stress, decreased amplitude and regularity of ultradian growth hormone secretory patterns accompanied by changes in sexually dimorphic liver gene expression (serpina1, Cyp2b9, Cyp2a4, Cyp2d9, and Cyp7b1). In addition to demonstrating the need for examination of the consequences of Cre transgenesis, these results also reveal how this SOM-cre strain may be a useful tool in studying the functional consequences of moderate to low SOM levels as reported in neurological and psychiatric disorders.

Ghrelin is an octanoylated peptide that acts via its specific receptor, the growth hormone secretagogue receptor type 1a (GHSR-1a), and regulates a vast variety of physiological functions. It is well established that ghrelin is predominantly synthesized by a distinct population of endocrine cells located within the gastric oxyntic mucosa. In addition, some studies have reported that ghrelin could also be synthesized in some brain regions, such as the hypothalamus. However, evidences of neuronal production of ghrelin have been inconsistent and, as a consequence, it is still as a matter of debate if ghrelin can be centrally produced. Here, we provide a comprehensive review and discussion of the data supporting, or not, the notion that the mammalian central nervous system can synthetize ghrelin. We conclude that no irrefutable and reproducible evidence exists supporting the notion that ghrelin is synthetized, at physiologically relevant levels, in the central nervous system of adult mammals., Instituto Multidisciplinario de Biología Celular

International audience; Nutrition during the perinatal period programs body growth. Growth hormone (GH) secretion from the pituitary regulates body growth and is controlled by Growth Hormone Releasing Hormone (GHRH) neurons located in the arcuate nucleus of the hypothalamus. We observed that dietary restriction during the early postnatal period (i.e. lactation) in mice influences postnatal growth by permanently altering the development of the somatotropic axis in the pituitary gland. This alteration may be due to a lack of GHRH signaling during this critical developmental period. Indeed, underfed pups showed decreased insulin-like growth factor I (IGF-I) plasma levels, which are associated with lower innervation of the median eminence by GHRH axons at 10 days of age relative to normally fed pups. IGF-I preferentially stimulated axon elongation of GHRH neurons in in vitro arcuate explant cultures from 7 day-old normally fed pups. This IGF-I stimulating effect was selective since other arcuate neurons visualized concomitantly by neurofilament labeling, or AgRP immunochemistry, did not significantly respond to IGF-I stimulation. Moreover, GHRH neurons in explants from age-matched underfed pups lost the capacity to respond to IGF-I stimulation. Molecular analyses indicated that nutritional restriction was associated with impaired activation of AKT. These results highlight a role for IGF-I in axon elongation that appears to be cell selective and participates in the complex cellular mechanisms that link underfeeding during the early postna-tal period with programming of the growth trajectory.

[This corrects the article DOI: 10.1371/journal.pone.0170083.].

Chantier qualité GA; International audience; Objective: Since a reduction of the insulin/IGF-1 signaling cascade extends life span in many species and IGF-1 signaling might partly mediate the effects of caloric restriction (CR), an experimental intervention for increasing longevity, the purpose of the present study was to use quantitative trait loci (QTL) analysis, an unbiased genetic approach, to identify particular regions of the genome influencing plasma IGF-1 levels in an F2 intercross between F344 and LOU/C rats; the latter being an inbred strain of Wistar origin, considered as a model of healthy aging since it resists to age (and diet)-induced obesity. Design: F1 hybrids were obtained by crossbreeding LOU/C with F344 rats, and then F1 were bred inter se to obtain the F2 population, of which 93 males and 94 females were studied. Total plasma IGF-1 levels were determined by radioimmunoassay. A genome scan of the F2 population was made with 100 microsatellite markers) selected for their polymorphism between LOU/C and F344 strains (and by covering evenly the whole genome. Results: By simple interval mapping sex-dependent QTLs were found on chromosome 17 in males and on chromosome 18 in females. By multiple interval mapping, additional QTLs were found on chromosomes 1, 4, 5, 6, 12, 15 and 19 in males and on chromosomes 3, 5, 6, 12 and 17 in females. Only the markers D1Rat196 and D12Mgh5 were found in both males and females. The majority of QTLs corresponded to metabolic syndrome (cardiac function: n = 45 (30%), obesity/diabetes: n = 22 (15%), inflammation: n = 19 (13%) and only a limited number to body weight: n = 13 (9%), proliferation (n = 10 (7%) or ossification: n = 7 (5%). Ninety-six candidate genes were located on the different QTLs. A significant proportion of these genes are connected to IGF-1 production and receptor pathways (n = 18) or metabolic syndrome (n = 11). Conclusions: Subsequent studies are necessary to determine whether the genetic networks underscored are also involved in age-associated obesity, diabetes and inflammation as well as cardiovascular impairments.

International audience; Altered brain somatostatin functions recently appeared as key elements for the pathogenesis of stress-related neuropsychiatric disorders. The hippocampus exerts an inhibitory feedback on stress but the mechanisms involved remain unclear. We investigated herein the role of hippocampal somatostatin receptor subtypes in both stress response and behavioral emotionality using C57BL/6, wild type and sst2 or sst4 knockout mice. Inhibitory effects of hippocampal infusions of somatostatin agonists on stress-induced hypothalamo-pituitary-adrenal axis (HPA) activity were tested by monitoring peripheral blood and local hippocampus corticosterone levels, the latter by using microdialysis. Anxiolytic and antidepressant-like effects were determined in the elevated-plus maze, open field, forced swimming, and stress-sensitive beam walking tests. Hippocampal injections of somatostatin analogs and sst2 or sst4, but not sst1 or sst3 receptor agonists produced rapid and sustained inhibition of HPA axis. sst2 agonists selectively produced anxiolytic-like behaviors whereas both sst2 and sst4 agonists had antidepressant-like effects. Consistent with these findings, high corticosterone levels and anxiety were found in sst2KO mice and depressive-like behaviors observed in both sst2KO and sst4KO strains. Both hippocampal sst2 and sst4 receptors selectively inhibit stress-induced HPA axis activation but mediate anxiolytic and antidepressive effects through distinct mechanisms. Such results are to be accounted for in development of pathway-specific somatostatin receptor agents in the treatment of hypercortisolism (Cushing's disease) and stress-related neuropsychiatric disorders.

International audience; Anorexia nervosa (AN) is classically defined as a condition in which an abnormally low body weight is associated with an intense fear of gaining weight and distorted cognitions regarding weight, shape, and drive for thinness. This article reviews recent evidences from physiology, genetics, epigenetics, and brain imaging which allow to consider AN as an abnormality of reward pathways or an attempt to preserve mental homeostasis. Special emphasis is put on ghrelino-resistance and the importance of orexigenic peptides of the lateral hypothalamus, the gut microbiota and a dysimmune disorder of neuropeptide signaling. Physiological processes, secondary to underlying, and premorbid vulnerability factors—the “pondero-nutritional-feeding basements”- are also discussed.

This article reviews the main findings that emerged in the intervening years since the previous volume on hormonal control of growth in the section on the endocrine system of the Handbook of Physiology concerning the intra- and extrahypothalamic neuronal networks connecting growth hormone releasing hormone (GHRH) and somatostatin hypophysiotropic neurons and the integration between regulators of food intake/metabolism and GH release. Among these findings, the discovery of ghrelin still raises many unanswered questions. One important event was the application of deconvolution analysis to the pulsatile patterns of GH secretion in different mammalian species, including Man, according to gender, hormonal environment and ageing. Concerning this last phenomenon, a great body of evidence now supports the role of an attenuation of the GHRH/GH/Insulin-like growth factor-1 (IGF-1) axis in the control of mammalian aging.

Fasting results in the mobilization of adipose stores and the elevation of levels of free fatty acids (FFA). In humans, this process is driven by a release in GH. Little is known regarding the role of GH in modulating this process during early stages of fasting in the mouse. Confirmation of the role of GH in modulating FFA release in the fasting mouse is of particular importance given the frequent use of mouse models to study metabolic mechanisms. Here, we correlate the initial release of FFA throughout fasting in mice with pulsatile GH secretion. Observations illustrate the rapid release of FFA in response to food withdrawal. This does not correlate with a rise in GH secretion. Rather, we observed a striking loss in pulsatile secretion of GH throughout the first 6 h of fasting, suggesting that GH does not modulate the initial release of FFA in the mouse in response to fasting. This was confirmed in GH receptor knockout mice, in which we observed a robust fasting-induced rise in FFA. We further illustrate the dynamic relationship between the orexigenic and anorexigenic hormones ghrelin and leptin during fasting in the mouse. Our findings show an initial suppression of leptin and the eventual rise in circulating levels of acyl-ghrelin with fasting. However, altered acyl-ghrelin and leptin secretion occurs well after the rise in FFA and the suppression of GH secretion. Consequently, we conclude that although acyl-ghrelin and leptin may modulate the physiological response to drive food intake, these changes do not contribute to the initial loss of pulsatile GH secretion. Rather, it appears that the suppression of GH secretion in fasting may occur in response to an elevation in fasting levels of FFA or physiological stress. Observations highlight a divergent role for GH in modulating FFA release between man and mouse.

Among the factors playing a crucial role in the regulation of energy metabolism, gastro-intestinal peptides are essential signals to maintain energy homeostasis as they relay to the central nervous system the informations about the nutritional status of the body. Among these factors, preproghrelin is a unique prohormone as it encodes ghrelin, a powerful GH secretagogue and the only orexigenic signal from the gastrointestinal tract and obestatin, a proposed functional ghrelin antagonist. These preproghrelin-derived peptides may contribute to balance energy intake, metabolism and body composition by regulating the activity of the GH/IGF-1 axis and appetite. Whereas the contribution of ghrelin has been well characterized, the role of the more recently identified obestatin, in this regulatory process is still controversial. In this chapter, we describe the contribution of these different preproghrelin-derived peptides and their receptors in the regulation of GH secretion and feeding. Data obtained from pharmacological approaches, mutant models and evaluation of the hormones in animal and human models are discussed.

Ghrelin is a 28 amino acid acylated peptide originally characterised for its capacity to stimulate growth hormone secretion. Ghrelin is also an orexigenic and adipogenic hormone and is thought to be a signal to increase locomotor activity in anticipation of a scheduled meal. Although ghrelin is considered to be up-regulated during fasting, there are still conflicting data regarding the impact of starvation on ghrelin secretion. To test whether the secretory pattern of acylated ghrelin is altered during fasting, plasma levels were monitored every 20 min for 6 h in freelybehaving rats at the light ⁄ dark cycle transition, when animals initiate feeding and activity and use preferentially free fatty acids (FFA) as a source of energy. Rats were fed ad lib. or fasted at dark onset for 24, 48 or 72 h, with or without refeeding rate. The anticipatory rise in ghrelin levels, as well as home-cage activity at the onset of darkness, was significantly reduced from 48 h of fasting compared to ad lib. conditions. A delayed ghrelin peak, sensitive to renutrition, was observed in fasted animals. Although their motivation to eat appeared to be intact, rats fasted for 72 h showed the smallest compensatory refeeding rate after fasting, possibly reflecting altered gut function. Expression of agouti-related protein and neuropeptide Y, was significantly increased in 48- and 72-h fasted animals. Thus, following fasting, a blunted acylated ghrelin secretion at dark onset (i.e. a period when animals depend on FFA as a source of energy) is associated with reduced locomotor activity and refeeding and an up-regulation of anabolic neuropeptides. Such changes could be interpreted as compensatory mechanisms for helping to conserve energy under conditions where food is not available.

Ghrelin, a natural ligand of the growth hormone secretagogue receptor (GHS-R), is synthesized in the stomach but may also be expressed in lesser quantity in the hypothalamus where the GHS-R is located on growth hormone-releasing hormone (GHRH) neurons. Obestatin, a peptide derived from the same precursor as ghrelin, is able to antagonize the ghrelin-induced increase of growth hormone (GH) secretion in vivo but not from pituitary explants in vitro. Thus, the blockade of ghrelin-induced GH release by obestatin could be mediated at the hypothalamic level by the neuronal network that controls pituitary GH secretion. Ghrelin increased GHRH and decreased somatostatin (somatotropin-releasing inhibitory factor) release from hypothalamic explants, whereas obestatin only reduced the ghrelin-induced increase of GHRH release, thus indicating that the effect of ghrelin and obestatin is targeted to GHRH neurons. Patch-clamp recordings on mouse GHRH-enhanced green fluorescent protein neurons indicated that ghrelin and obestatin had no significant effects on glutamatergic synaptic transmission. Ghrelin decreased GABAergic synaptic transmission in 44% of the recorded neurons, an effect blocked in the presence of the GHS-R antagonist BIM28163, and stimulated the firing rate of 78% of GHRH neurons. Obestatin blocked the effects of ghrelin by acting on a receptor different from the GHS-R. These data suggest that: (i) ghrelin increases GHRH neuron excitability by increasing their action potential firing rate and decreasing the strength of GABA inhibitory inputs, thereby leading to an enhanced GHRH release; and (ii) obestatin counteracts ghrelin actions. Such interactions on GHRH neurons probably participate in the control of GH secretion.

Summary Constitutional thinness (CT) and anorexia nervosa (AN) are two categories of severely underweight subjects. Some appetite-regulating hormones display opposite levels in AN and CT. While levels of ghrelin, an orexigenic hormone, fit with the normal food intake in CT, the lack of efficacy of increased ghrelin levels in AN is not clear. Obestatin is a recently described peptide derived from the preproghrelin gene, reported to inhibit appetite in contrast to ghrelin. The aim of this study was to determine whether the circadian profile of obestatin, total and acylated ghrelin levels is different in CT subjects when compared with AN patients. Six-points circadian profiles of plasma obestatin, acylated ghrelin, total ghrelin and other hormonal and nutritional parameters were evaluated in four groups of young women: 10 CT, 15 restricting-type AN, 7 restored from AN and 9 control subjects. Obestatin circadian levels were significantly higher inAN (p < 0.0001) while no difference was found between CTand control subjects. Acylated and total ghrelin were found increased in AN. Acylated ghrelin/obestatin and total ghrelin/obestatin were found decreased in AN compared to CTor C subjects (p < 0.05). The percentage of acylated ghrelin was found decreased in CT group (p < 0.05). The decreased ghrelin/obestatin ratio found in AN might participate in the restraint in nutriment intake of these patients. In contrast, in CTa lower percentage of acylated over total ghrelin might be considered in the aetiology of this condition.

The amyloid precursor protein (APP) and APP-like (APLP) material, as visualized with the Mab22C11 antibody, have previously been shown to be associated with radial glia in hypothalamus, which are known to promote neurite outgrowth. By Northern blot analysis, APP 695 mRNA levels increased steadily over hypothalamic development, APP 770 mRNA was transiently expressed at 12 days postnatally, and APLP mRNA was only weakly expressed in the hypothalamus. The developmental pattern of APP moeities in mouse hypothalamus and in fetal hypothalamic neurons in culture was compared with a presenilin 2 (PS2) related protein using an antibody developed against the N-terminal part of PS2. By Western blot analysis, APP and PS2-like immunoreactivity were visualized as a 100-130 and 52 kDa bands, respectively. An APP biphasic increase was observed during hypothalamic development in vivo. APP immunoreactivity was equally detected in neuronal and glial cultures, while PS2-like material was more concentrated in neurons. A correlation between APP/APP-like and PS2-like levels was observed during development in vivo. While APP was mostly associated with membrane fractions, a significant portion of PS2-like material was also recovered from cytosolic fractions in vitro. In contrast to native PS2 in COS-transfected cells, the PS2-like material did not aggregate after heating for 90 s at 90 degrees C. These results indicate a close association between APP and PS2-like material during hypothalamic development in vivo, and suggest that neuronal and glial cultures may provide appropriate models to test their interactions.

Somatostatin receptors were visualized by [125I]-Tyr0-DTrp8-somatostatin radioautography on 35% of arcuate neurons containing proopiomelanocortin (POMC) mRNA, as identified by in situ hybridization using a [35S] labelled riboprobe on 5 microm-thick consecutive sections. Furthermore, double immunohistochemical staining revealed contacts of beta-endorphin or alpha-MSH containing fibres with a majority of somatostatin perikarya in the anterior hypothalamic periventricular nucleus. Taken together, these data indicate that hypothalamic somatostatin and POMC neurons are interconnected. The results are discussed in term of intrahypothalamic control of GH secretion.

Energy homeostasis is achieved by the integration of peripheral metabolic signals by the neural circuits involving specific hypothalamic nuclei and brain stem regions. These neural circuits mediate many of the effects of the adipocyte-derived hormone leptin and gut-derived hormone ghrelin. The former is strongly anorexigenic while the latter is the only orexigenic agent active when administered by a peripheral route. Abnormal regulation of these 2 antagonistic regulatory peptides in patients with schizophrenia could play a role in the impairment in the regulation of food intake and energy balance. This bibliographical analysis aims to compare 27 prospective and cross-sectional studies published on circulating leptin and ghrelin levels during acute and chronic administration of antipsychotics treatment, especially atypical ones. Fasting morning leptin levels of schizophrenic patients increase rapidly in the first 2 weeks after atypical antipsychotic (AAP) treatment (mostly olanzapine and clozapine) and remain somehow elevated after that period up to several months. On the contrary, conventional antipsychotics (such as haloperidol) do not interfere with leptin levels. In contrast to leptin, fasting morning ghrelin levels decrease during the first few weeks after the beginning of AAPs treatment while they increase in the longer run. Surprisingly, body weight gain and correlations between the variation of these 2 peptides and adiposity and metabolism-related parameters such as the body mass index and abdominal perimeter were not systematically considered. Finally, an objective evaluation of feeding behavior during antipsychotic treatment remains to be determined.

Somatostatin is abundantly expressed in mammalian brain. The peptide binds with high affinity to six somatostatin receptors, sst1, sst2A and B, sst3 to 5, all belonging to the G-protein-coupled receptor family. Recent advances in the neuroanatomy of somatostatin neurons and cellular distribution of sst receptors shed light on their functional roles in the neuronal network. Beside their initially described neuroendocrine role, somatostatin systems subserve neuromodulatory roles in the brain, influencing motor activity, sleep, sensory processes and cognitive functions, and are altered in brain diseases like affective disorders, epilepsia and Alzheimer's disease.

In adults, the adipocyte-derived hormone, leptin, regulates food intake and body weight principally via the hypothalamic arcuate nucleus (ARC). During early postnatal development, leptin functions to promote the outgrowth of neuronal projections from the ARC, whereas a selective insensitivity to the effects of leptin on food intake appears to exist. To investigate the mechanisms underlying the inability of leptin to regulate food intake during early development, leptin signaling was analyzed both in vitro using primary cultures of rat embryonic ARC neurones and in vivo by challenging early postnatal rats with leptin. In neuronal cultures, despite the presence of key components of the leptin signaling pathway, no detectable activation of either signal transducer and activator of transcription 3 or the MAPK pathways by leptin was detected. However, leptin down-regulated mRNA levels of proopiomelanocortin and neuropeptide Y and decreased somatostatin secretion. Leptin challenge in vivo at postnatal d (P) 7, P14, P21, and P28 revealed that, in contrast to adult and P28 rats, mRNA levels of neuropeptide Y, proopiomelanocortin, agouti-related peptide and cocaine- and amphetamine-regulated transcript were largely unaffected at P7, P14, and P21. Furthermore, leptin stimulation increased the suppressor of cytokine signaling-3 mRNA levels at P14, P21, and P28 in several hypothalamic nuclei but not at P7, indicating that selective leptin insensitivity in the hypothalamus is coupled to developmental shifts in leptin receptor signaling. Thus, the present study defines the onset of leptin sensitivity in the regulation of energy homeostasis in the developing hypothalamus.

The secretion of growth hormone (GH) is inhibited by hypothalamic somatostatin (SRIF) in somatotropes through five subtypes of the somatostatin receptor (SSTR1–SSTR5). We aimed to characterize the subtype(s) of SSTRs involved in the Ca2+current reduction in GH3 somatotrope cells using specific SSTR subtype agonists. We used nystatin-perforated patch clamp to record voltage-gated Ca2+currents, using a holding potential of −80 mV in the presence of K+and Na+channel blockers. We first established the presence of T-, L-, N-, and P/Q-type Ca2+currents in GH3 cells using a variety of channel blockers (Ni+, nifedipine, ω-conotoxin GVIA, and ω-agatoxin IVA). SRIF (200 nM) reduced L- and N-type but not T- or P/Q-type currents in GH3 cells. A range of concentrations of each specific SSTR agonist was tested on Ca2+currents to find the maximal effective concentration. Activation of SSTR2 with 10−7and 10−8M L-797,976 decreased the voltage-gated Ca2+current and abolished any further decrease by SRIF. SSTR1, SSTR3, SSTR4, and SSTR5 agonists at 10−7M did not modify the voltage-gated Ca2+current and did not affect the Ca2+current response to SRIF. These results indicate that SSTR2 is involved mainly in regulating voltage-gated Ca2+currents by SRIF, which contributes to the decrease in intracellular Ca2+concentration and GH secretion by SRIF.

Growth hormone (GH) secretion decreases spontaneously during lifespan, and the resulting GH deficiency participates in aging-related morbidity. This deficiency appears to involve a defect in the activity of hypothalamic GH-releasing hormone (GHRH) neurons. Here, we investigated this hypothesis, as well as the underlying mechanisms, in identified GHRH neurons from adult ( approximately 13 weeks old) and aged ( approximately 100 weeks old) transgenic GHRH-green fluorescent protein mice, using morphological, biochemical and electrophysiological methods. Surprisingly, the spontaneous action potential frequency was similar in adult and aged GHRH neurons studied in brain slices. This was explained by a lack of change in the intrinsic excitability, and simultaneous increases in both stimulatory glutamatergic- and inhibitory GABAergic-synaptic currents of aged GHRH neurons. Aging did not decrease GHRH and enhanced green fluorescent protein contents, GHRH neuronal number or GHRH-fibre distribution, but we found a striking enlargement of GHRH-positive axons, suggesting neuropeptide accumulation. Unlike in adults, autophagic vacuoles were evident in aged GHRH-axonal profiles using electron microscopy. Thus, GHRH neurons are involved in aging of the GH axis. Aging had a subtle effect at the nerve terminal level in GHRH neurons, contrasting with the view that neuronal aging is accompanied by more widespread damage.

In the present work, we took advantage of a recently described model of GHRH-enhanced green fluorescent protein (eGFP) transgenic mice to evaluate the extent of co-localization of GHRH neurons with galanin (GAL), neurotensin (NT) and tyrosine hydroxylase (TH) in 3- and 8-month-old male and female mice. The total number of GHRH-eGFP neurons along the rostro-caudal axis of the arcuate nucleus did not differ according to gender or age. GAL-immunoreactivity was present in 40–44% of 3-month-old GHRH-eGFP neurons in male and female arcuate nucleus, respectively, but only 25–22% in 8-month-old mice. TH immunoreactivity occurred in 36–35% of GHRH-eGFP neurons in male and female arcuate nucleus from 3-month-old mice and these proportions increased to 40 and 45% in 8-month-old mice. NT immunoreactivity was present in 14 and 24% of GHRH-eGFP neurons in male and female arcuate nucleus from 3-month-old mice up to 28 and 26% in 8-month-old mice. Thus, co-localization of peptides and enzyme in GHRH-eGFP neurons displays a sexual dimorphism at 3-month of age for NT, and at 8-month for TH, while the total number of GHRH-eGFP neurons does not exhibit gender difference at either age. In summary, it appears that changes in co-localized (and presumably co-released) peptides, rather than GHRH per se , may contribute to the changes in sexually dimorphic GH secretion with aging in the mouse.

International audience; BACKGROUND: Insulin-like growth factor-I (IGF-I) serum level decreases with age, and this decrease may underlie hemoglobin (Hb) decrease. The objective of the study was to assess the relationship between IGF-I and insulin-like growth factor binding protein-3 (IGFBP-3) serum levels and Hb, after adjustment especially for major nutritional factors in an elderly population because IGF-I system depends on nutritional state, often impaired in the elderly.METHODS: Hemoglobin concentration was tested for 672 participants evaluated during an outpatient geriatric assessment. IGF-I and IGFBP-3 serum levels were assessed by Enzyme Linked Immunosorbent Assay. The molar ratio of IGF-I/IGFBP-3 that reflects the bioavailable IGF-I was calculated. Levels of IGF-I and IGFBP-3 were plotted against quartiles of Hb. Final linear models for IGF-I, IGFBP-3 and ratio molar included factors that could modify the Hb level.RESULTS: Mean age (SD) of the sample was 78.0 (8.5) years old and 32% were men. After adjustment for age and sex, IGF-I serum level, IGFBP-3 serum level and molar ratio significantly increased with increasing quartiles of Hb. After adjustment for age, gender, diabetes, albumin, pre-albumin, renal function, total cholesterol, angiotensin converting enzyme inhibitors and angiotensin II receptor blockers consumption, C-Reactive Protein, Hb was significantly associated and with IGF-I level (p = .002) and molar ratio (p = .02).CONCLUSIONS: IGF-I serum level and IGF-I/IGFBP-3 molar ratio were associated with Hb in an elderly population, independently of nutritional biological parameters. Thus, the association between the IGF-I system and Hb merits further investigation to determine whether interventions that modulate circulating IGF-I or IGF-I/BP3 ratio might preserve Hb in the elderly.

We have added the following correction to the Editorial article, now including a reference to Kern et al. “Apo-ghrelin receptor (apo-GHSR1a) regulates dopamine signaling in the brain.” Four articles address the complex interactions between metabolism and neuropsychiatric symptoms. The first one focuses on biological differences between restrictive anorexia nervosa and constitutional thinness, a controversial concept to describe young girls who follow a normal diet and differ from restrictive anorexia nervosa on a number of endocrine parameters (10). At the opposite of the spectrum, the second one reviews the role of inflammatory processes in the neuropsychiatric comorbidity associated with obesity (11). The third one summarizes the fascinating link through ghrelin peptides between appetite/reward/growth hormone axis and psychiatric disorders (12). Finally, the last one proposes a molecular mechanism through allosteric interactions between dopamine/DRD2 and GHSR1 receptors for controlling appetite and the uncontrollable hyperphagia associated with Prader–Willi syndrome (13). Addition of reference: 13. Kern A, Grande C, Smith RG. Apo-ghrelin receptor (apo-GHSR1a) regulates dopamine signaling in the brain. Front Endocrinol (2014) 5:129. doi:10.3389/fendo.2014.00129

In the last few decades, a number of reciprocal associations between endocrine system dysfunctions and neurological or psychiatric manifestations have appeared, and only in the minority of the cases this link has been fully elucidated (1). Unfortunately, the complexity, and sometimes also the severity, of the clinical issues and the limited availability of reliable experimental models for the study of the endocrine system–nervous system cross-interactions do not help the fast achievement of scientific progresses. Isolated neurological or psychiatric symptoms as well as more complex disorders can occur in the setting of most endocrine diseases. Such manifestations can be the direct result of hyper- or hyposecretion of hormones from the endocrine glands or may occur as events secondary to the pathogenic mechanisms of the endocrinopathy, like in the case of autoimmunity affecting both endocrine glands and the brain (2). Moreover, the medical or surgical treatment of endocrine diseases can sometimes determine the occurrence of neurological or psychiatric syndromes. Some genetic aberrations can also lead to conditions affecting both the endocrine and the nervous system with a variety of possible manifestations. On the other side, many psychiatric or neurological diseases, including but not limiting to conditions, which affect the hypothalamus or the pituitary gland, may impair the physiological endocrine functions and demonstrate once again the intricate complexity of the psychoneuroendocrine connection. Covering all basic, translational, and clinical aspects in a single Research Topic on the “Neurological and psychiatric disorders in endocrine diseases” cannot be realistically feasible. The scope is simply raising a number of interesting points from experts in the field to bring a contribution to the knowledge and maybe direct future research in this very intriguing area. Thus, we have collected reviews and original research articles, which may speak to a wide audience, including neurologists, psychiatrists, endocrinologists, and basic scientists in neuroscience, endocrinology, and metabolism. In the clinical area, an interesting study assesses the plasma levels of the muscle-derived hormone irisin in anorexic women, without finding any significant correlation between irisin level and physical exercise (3). Another fascinating study analyzes the complexity of psychiatric and personality disorders in women with polycystic ovary syndrome, highlighting the issue of psychological distress in this patient group (4). A comprehensive picture of gender dysphoria, a psychiatric condition, which requires endocrine management, in Ireland is presented (5). In a short review, the most relevant reproductive, neurodevelopmental, and genetic aspects of hypogonadotropic hypogonadal syndromes are outlined (6). A German study indicates that psychopathology significantly predicts quality of life in patients with acromegaly and suggests that depressive symptoms and anxiety, being modifiable factors, may represent relevant targets for a broad treatment intervention in acromegalic patients (7). In a concise review, the psychiatric alterations, the neurocognitive impairment, and the altered quality of life affecting at some extent the majority of the patients with Cushing’s syndrome are summarized, and the authors highlight that resolution of hypercortisolism, a challenging and non-granted achievement, does not always lead to the complete remission of the neuropsychiatric changes or restore the quality of life (8). With an incursion in the surgical field, the complications potentially affecting the laryngeal nerves during thyroid surgery are extensively reviewed, suggesting that still the incidence of laryngeal nerves damage secondary to thyroid surgery cannot be suppressed but may be maintained in a low range with thorough surgical techniques and the use of intraoperative neuromonitoring (9). Three articles address the complex interactions between metabolism and neuropsychiatric symptoms. The first one focuses on biological differences between restrictive anorexia nervosa and constitutional thinness, a controversial concept to describe young girls who follow a normal diet and differ from restrictive anorexia nervosa on a number of endocrine parameters (10). At the opposite of the spectrum, the second one reviews the role of inflammatory processes in the neuropsychiatric comorbidity associated with obesity (11). Finally, the last one summarizes the fascinating link through ghrelin peptides between appetite/reward/growth hormone axis and psychiatric disorders (12).

Ghrelin, a gut hormone originating from the post-translational cleavage of preproghrelin, is the endogenous ligand of growth hormone secretagogue receptor 1a (GHS-R1a). Within the growth hormone (GH) axis, the biological activity of ghrelin requires octanoylation by ghrelin-O-acyltransferase (GOAT), conferring selective binding to the GHS-R1a receptor via acylated ghrelin. Complete loss of preproghrelin-derived signalling (through deletion of the Ghrl gene) contributes to a decline in peak GH release; however, the selective contribution of endogenous acyl-ghrelin to pulsatile GH release remains to be established. We assessed the pulsatile release of GH in ad lib. fed male germline goat(-/-) mice, extending measures to include mRNA for key hypothalamic regulators of GH release, and peripheral factors that are modulated relative to GH release. The amount of GH released was reduced in young goat(-/-) mice compared to age-matched wild-type mice, whereas pulse frequency and irregularity increased. Altered GH release did not coincide with alterations in hypothalamic Ghrh, Srif, Npy or Ghsr mRNA expression, or pituitary GH content, suggesting that loss of Goat does not compromise canonical mechanisms that contribute to pituitary GH production and release. Although loss of Goat resulted in an irregular pattern of GH release (characterised by an increase in the number of GH pulses observed during extended secretory events), this did not contribute to a change in the expression of sexually dimorphic GH-dependent liver genes. Of interest, circulating levels of insulin-like growth factor (IGF)-1 were elevated in goat(-/-) mice. This rise in circulating levels of IGF-1 was correlated with an increase in GH pulse frequency, suggesting that sustained or increased IGF-1 release in goat(-/-) mice may occur in response to altered GH release patterning. Our observations demonstrate that germline loss of Goat alters GH release and patterning. Although the biological relevance of altered GH secretory patterning remains unclear, we propose that this may contribute to sustained IGF-1 release and growth in goat(-/-) mice.

Cocaine- and amphetamine-regulated transcript (CART) mRNA and peptides are abundant in the adenohypophysis, but their role in pituitary function has not yet been elucidated. CART peptides were recently shown to colocalise with luteinising hormone (LH) or prolactin in rat anterior pituitary, and contradictory results concerning the peptide effects on pituitary hormonal secretions were obtained in vitro from pituitary cell cultures. Thus, we reinvestigated the expression of CART mRNA within the pituitary. Immunohistochemistry for pituitary hormones was performed on sections from adult male Wistar rats followed by in situ hybridisation using CART mRNA antisense 35S-labelled probes. The most represented CART-expressing cells were lactotrophs (42 +/- 1% of CART cells) and gonadotrophs (32 +/- 3%), followed by thyrotrophs (10 +/- 2%), corticotrophs (7 +/- 2%) and somatotrophs (6 +/- 1%). In the pars tuberalis, CART mRNA was easily detectable in gonadotrophs and lactotrophs and, to a lesser extent, in corticotrophs and thyrotrophs. CART peptide was quickly and potently released from perifused pituitary by depolarisation (K+ 30 mM for 15 min; 465 +/- 37% over basal release, n = 5). Gonadotrophin-releasing hormone and thyrotrophin-releasing hormone (0.1 microM) were also active to a lesser extent (138 +/- 11% and 71 +/- 17, n = 7, respectively). CART (0.1 microM) did not modify basal LH or prolactin release but selectively inhibited K+-induced LH release without affecting K+-induced prolactin secretion. Pituitary CART mRNA and content were sex dependent and varied during the oestrous cycle, being lower in dioestrous 2. Pituitary CART content also varied widely amongst rat strains being five to six-fold higher in Wistar and Fischer rats compared to Brown Norway and Lou C rats. Ageing differentially affected pituitary CART mRNA and content, resulting in a marked decrease in Lou C and an increase in Wistar and Sprague-Dawley rats. Taken together, these results suggest that pituitary CART expression is dependent of the sex steroid environment and may be physiologically involved in LH secretion.

Five somatostatin receptors (sst) have been cloned and mRNAs for the first four (sst1–4) are expressed in many brain regions. In the present work, we compared the distribution of the non-selective ligand [125I]-Tyr0-DTrp8-SRIF14 by autoradiography in 24 brain regions and pituitary in wild type, sst1- to sst4- or SRIF-gene invalidated (KO) mice. [125I]-Tyr0-DTrp8-SRIF14 binding was not significantly modified in sst1 KO mouse brain with the noticeable exception of the substantia nigra and only moderately decreased in pituitary. For sst2 KO mice, a general decrease (>75%) was observed in most regions, with the noticeable exception of the olfactory bulb and CA1 field of the hippocampus. SST3 KO brain displayed a decrease in binding in the external plexiform layer of the olfactory bulb only (−54%). For sst4 KO mice, [125I]-Tyr0-DTrp8-SRIF14 binding levels in the external plexiform (−35%) and glomerular (−39%) layers of the olfactory bulb as well as the hippocampus CA1 field (−68%) were significantly decreased. In SRIF KO mice, a significant increase in binding levels was observed in olfactory bulb, anterior olfactory nucleus, frontal cortex upper layers, lateral septum, CA1 field, zona incerta and lateral hypothalamus, substantia nigra, periaqueductal grey and parabrachial nucleus. Competition with selective ligands (CH275, octreotide or L-779,976, L-796,778, L-803,087, and octreotide or L-817,778, for sst1–5 receptors, respectively) was in accordance with these findings. Moreover, octreotide was still able to compete on residual [125I]-Tyr0-DTrp8-SRIF14 binding sites in sst2 KO pituitary. It is concluded that most [125I]-Tyr0-DTrp8-SRIF14 binding sites in mouse brain and pituitary belong to the sst2 subtype but for the olfactory bulb (sst3 and sst4 receptors), the CA1 of the hippocampus (sst4 receptors) and the pituitary (sst5 and sst1 receptors) in which other subtypes are also expressed. The overall increase in [125I]-Tyr0-DTrp8-SRIF14 binding in SRIF KO mice indicates that SRIF receptors, mostly from the sst2 subtype, are regulated by the endogenous ligand(s).

A cDNA membrane array displaying 1183 probes was used to detect hypothalamic and pituitary changes in gene expression accompanying ageing and age-associated pituitary macroadenomas. Four groups of male Sprague-Dawley rats (3-, 15-, 24-month-old and 24-month-old with prolactinoma) were compared in two independent hybridizations. cDNA array data were confirmed and completed by comparative reverse transcriptase-polymerase chain reaction on selected genes. The expression of 454 and 116 mRNAs was detected in hypothalamus and pituitary, respectively. Growth hormone (GH) mRNA alone represented 85% of total gene expression in the gland of young rats, and other pituitary hormone transcripts 2.8%, while melanin-concentrating hormone (MCH) mRNA, the most expressed neuropeptide transcript involved in neuroendocrine regulation, accounted for only 0.8% of total hypothalamic transcripts. The proportion of genes modified in the hypothalamus and pituitary was rather modest: 1.5% and 5.2%, respectively, for ageing per se, and 1.1% and 5.2% for age-associated macroprolactinomas. Among pituitary specific RNAs, GH mRNA expression was notably decreased with age. At the hypothalamic level, expression of genes directly involved in GH regulation, such as somatostatin and growth hormone-releasing hormone, was not altered, while neuropeptide transcripts involved in feeding behaviour [orexin/hypocretin, MCH, pro-opiomelanocortin (POMC), cocaine- and amphetamine-regulated transcript (CART)] were significantly altered. In addition, a few ubiquitous transcripts (hnRNP-K, PFKm, CCND 2, calponin and set) were differently affected in both tissues. Modifications in hypothalamic orexigenic (orexin, MCH) and anorexigenic (POMC, CART) gene expression are in keeping with an age-associated decrease in energy consumption but a higher one in the presence of macroprolactinomas.

The physiological actions of somatostatin-14 (SRIF: somatotrophin release inhibitory factor) receptor subtypes (sst(1)-sst(5)), which are endogenously expressed in growth cells (GC cells), have not yet been elucidated, although there is evidence that sst(2) receptors are negatively coupled to cytosolic free Ca(2+) concentration ([Ca(2+)](i)) and adenosine 3,5'-cyclic monophosphate (cAMP) accumulation. In addition, both sst(1) and sst(2) receptors are negatively coupled to growth hormone (GH) secretion in GC cells. Here we report on studies concerning the expression, the pharmacology and the functional role of native SRIF receptors in GC cells with the use of five nonpeptidyl agonists, highly selective for each of the SRIF receptors. Radioligand binding studies show that sst(2) and sst(5) receptors are present at different relative densities, while the presence of sst(3) and sst(4) receptors appears to be negligible. The absence of sst(1) receptor binding was unexpected in view of sst(1) receptor functional effects on GH secretion. This suggests very efficient receptor-effector coupling of a low-density population of sst(1) receptors. Functionally, only sst(2) receptors are coupled to the inhibition of [Ca(2+)](i) and cAMP accumulation and the selective activation of sst(5) receptors facilitates the stimulation of adenylyl cyclase activity through G(i/o) proteins. This effect was not observed when sst(2) and sst(5) receptors were simultaneously activated, suggesting that there is a functional interaction between sst(2) and sst(5) receptors. In addition, sst(1), sst(2) and sst(5) receptor activation inhibits GH release, further indicating that SRIF can modulate GH secretion in GC cells through mechanisms both dependent and independent on [Ca(2+)](i) and cAMP-dependent pathways. The present data suggest SRIF-mediated functional effects in GC cells to be very diverse and provides compelling arguments to propose that multiple native SRIF receptors expressed in the same cells are not simply redundant, but contribute to marked signalling diversity.

Numerous physiological studies as well as in situ hybridization and PCR experiments concur in reporting a role for the sst2A receptor in transducing somatostatin (SRIF) actions in the rat hypothalamus. However, the distribution of this receptor protein is not known within this structure. Regional and cellular localization of the sst2A receptor was therefore examined in the rat hypothalamus using highly sensitive immunohistochemical techniques. In close correspondence with the distribution of SRIF-immunoreactive fibers, numerous hypothalamic areas displayed sst2A receptor immunoreactivity. Receptor labeling was, however, diffusely distributed over the tissue, and few immunopositive cells were apparent. Unraveling the distribution of receptor-expressing cells was achieved through acute in vivo agonist stimulation and subsequent receptor internalization. At the cellular level, double-immunolabeling experiments with synaptophysin and microtubule-associated protein 2 demonstrated that sst2A receptors were predominantly internalized in perikarya and dendrites. Double-labeling experiments with SRIF revealed that 93% of arcuate, but only 18% of periventricular, SRIF-positive neurons expressed internalized receptors. Taken together, these results demonstrate for the first time that the sst2A receptor protein is widely, but selectively, distributed in the hypothalamus, and that postsynaptic sst2A auto- and heteroreceptors are well poised to play an important role in the somatostatinergic regulation of hypothalamic endocrine and metabolic processes.

A somatostatin deficit occurs in the cerebral cortex of Alzheimer's disease patients without a major loss in somatostatin-containing neurons. This deficit could be related to a reduction in the rate of proteolytic processing of peptide precursors. Since the two proprotein convertases (PC)1 and PC2 are responsible for the processing of neuropeptide precursors directed to the regulated secretory pathway, we examined whether they are involved first in the proteolytic processing of prosomatostatin in mouse and human brain and secondly in somatostatin defect associated with Alzheimer's disease. By size exclusion chromatography, the cleavage of prosomatostatin to somatostatin-14 is almost totally abolished in the cortex of PC2 null mice, while the proportions of prosomatostatin and somatostatin-28 are increased. By immunohistochemistry, PC1 and PC2 were localized in many neuronal elements in human frontal and temporal cortex. The convertases levels were quantified by Western blot, as well as the protein 7B2 which is required for the production of active PC2. No significant change in PC1 levels was observed in Alzheimer's disease. In contrast, a marked decrease in the ratio of the PC2 precursor to the total enzymatic pool was observed in the frontal cortex of Alzheimer patients. This decrease coincides with an increase in the binding protein 7B2. However, the content and enzymatic activity of the PC2 mature form were similar in Alzheimer patients and controls. Therefore, the cortical somatostatin defect is not due to convertase alteration occuring during Alzheimer's disease. Further studies will be needed to assess the mechanisms involved in somatostatin deficiency in Alzheimer's disease.

We have investigated the role of somatostatin receptor subtypes sst 2 and sst 4 in limbic seizures and glutamate-mediated neurotransmission in mouse hippocampus. As compared to wild-type littermates, homozygous mice lacking sst 2 receptors showed a 52% reduction in EEG ictal activity induced by intrahippocampal injection of 30 ng kainic acid (P < 0.05). The number of behavioural tonic-clonic seizures was reduced by 50% (P < 0.01) and the time to onset of seizures was doubled on average (P < 0.05). Seizure-associated neurodegeneration was found in the injected hippocampus (CA1, CA3 and hilar interneurons) and sporadically in the ipsilateral latero-dorsal thalamus. This occurred to a similar extent in wild-type and sst 2 knock-out mice. Intrahippocampal injection of three selective sst 2 receptor agonists in wild-type mice (Octreotide, BIM 23120 and L-779976, 1.5-6.0 nmol) did not affect kainate seizures while the same compounds significantly reduced seizures in rats. L-803087 (5 nmol), a selective sst4 receptor agonist, doubled seizure activity in wild-type mice on average. Interestingly, this effect was blocked by 3 nmol octreotide. It was determined, in both radioligand binding and cAMP accumulation, that octreotide had no direct agonist or antagonist action at mouse sst4 receptors expressed in CCl39 cells, up to micromolar concentrations. In hippocampal slices from wild-type mice, octreotide (2 μM) did not modify AMPA-mediated synaptic responses while facilitation occurred with L-803087 (2 μM). Similarly to what was observed in seizures, the effect of L-803087 was reduced by octreotide. In hippocampal slices from sst 2 knock-out mice, both octreotide and L-803087 were ineffective on synaptic responses. Our findings show that, unlike in rats, sst 2 receptors in mice do not mediate anticonvulsant effects. Moreover, stimulation of sst 4 receptors in the hippocampus of wild-type mice induced excitatory effects which appeared to depend on the presence of sst 2 subtypes, suggesting these receptors are functionally coupled.

The vasopressin V3 receptor (V3) is specifically expressed in pituitary corticotropes and mediates the stimulatory effect of vasopressin on adrenocorticotropic hormone (ACTH) release. The V3 gene is overexpressed in corticotrope pituitary tumours compared to normal pituitaries. We hypothesized that V3 overexpression might induce changes in corticotrope function and alter the regulation of the hypothalamic-pituitary-adrenal axis. Thus, we generated transgenic mice (POMV3) expressing the human V3 receptor in the pituitary under the control of rat pro-opiomelanocortin (POMC) promoter sequences. The transgene was efficiently transcribed and vasopressin binding was increased in both corticotropes and melanotropes. In-vitro ACTH release and inositol phosphate formation were unchanged in POMV3 pituitaries, but the responses to vasopressin were significatively increased. In vivo, basal circulating concentrations of ACTH in POMV3 mice were similar to those of controls but corticosterone concentrations were moderately increased. In addition, the levels of POMC mRNA in the transgenic pituitaries were comparable to those of control mice. Finally, POMV3 mice responded with a similar maximal increase of ACTH and corticosterone to a 20-min acute restraint stress. Together, these results show that hypophyseal V3 overexpression led to increased basal concentrations of corticosterone and suggest that the negative glucocorticoid feedback may be altered at the pituitary level.

The expression of the five somatostatin receptor subtypes, sst 1 - 5 was compared on tissue containing glial tumours (glioblastomas or oligodendrogliomas), medulloblastomas, and on normal human cortex. By semiquantitativereverse transcription coupled to polymerase chain reaction, the receptor expression profiles were high in cortex and in tissue containing oligodendrogliomas. It was moderate in medulloblastomas. Tissue containing glioblastomas displayed lower expression of somatostatin receptor subtypes, sst 1 and sst 3 being mostly expressed. By 1 2 5 I-Tyr 0 DTrp 8 somatostatin-14 or 1 2 5 I-Leu 8 DTrp 2 2 Tyr 2 5 somatostatin-28 autoradiography combined with synaptophysin immunohistochemistry, it was possible to differentiate between isolated tumoral cell component infiltrating the cerebral parenchyma (cortex or white matter) and tumoral tissue (without residual parenchyma) in glioblastomas or oligodendrogliomas. Glial tumoral tissue per se presented few somatostatin receptors. By contrast, medulloblastoma tumoral cells exhibited numerous octreotide sensitive somatostatin receptors, sst 2 immunocytochemistry demonstrated immunostaining of neuronal cells and neuropile; sst 2 and sst 3 immunostaining was identified on glioblastoma proliferating vessels endothelial cells and on medulloblastomas tumoral cells. Faint sst 2 immunostaining among glial tumoral cells was due to microglia, while glioma cells did not significantly stain. In summary, medulloblastoma tumoral cells express sst 2 /sst 3 receptors at a high level while glioma cells do not. In gliomas, sst expression is restricted to endothelial cells on proliferating vessels (displaying both sst 2 and sst 3 receptors), including parenchyma and reactive microglia (only sst 2 ). The differential expression of sst 2 /sst 3 receptors on gliomas and medulloblastomas has implications for the therapy of these tumours.

Ghrelin, an endogenous ligand for the GHS receptor, stimulates GH secretion and gastrointestinal motility and has orexigenic effects. In this study, the relationships between ghrelin, GH secretion, feeding behavior, and sleep-wake patterns were investigated in adult male rats. The half-life of exogenous ghrelin (10 microg i.v.) in plasma was about 30 min. Repeated administration of ghrelin at 3- to 4-h intervals (one during lights-on and two during lights-off periods) increased GH release and feeding activity, and decreased rapid eye movement sleep duration. Endogenous plasma ghrelin levels exhibited pulsatile variations that were smaller and less regular compared with those of GH. No significant correlation between GH and ghrelin circulating levels was found, although mean interpeak intervals and pulse frequencies were close for the two hormones. In contrast, ghrelin pulse variations were correlated with food intake episodes in the lights off period, and plasma ghrelin concentrations decreased by 26% in the 20 min following the end of the food intake periods. A positive correlation between ghrelin levels and active wake was found during the first 3 h of the dark period only. In conclusion, ghrelin, in addition to affecting GH secretion, gastrointestinal motility, and feeding activity, also modifies sleep-wake patterns. However, a direct action of ghrelin per se or the indirect effects of feeding (and all of its attendant metabolic sequelae) on sleep cannot be differentiated. Moreover, ghrelin secretion is pulsatile and directly related to feeding behavior only.

The peptide hormone/neurotransmitter somatostatin (somatotropin release inhibiting factor; SRIF) and its receptors (sst(1)-sst(5)) appear to regulate many physiological functions in the CNS. Semi-quantitative analysis of the densities of mRNA expression for sst(1-5) receptors and SRIF receptor binding sites were established in sst(2) receptor knock-out (KO) mice. Patterns of sst(1-5) receptor mRNA expression were largely conserved for sst(1,3,4) and sst(5) selective oligonucleotide probes; whereas sst(2) signals were completely absent in KO mouse brain. Autoradiographic analysis demonstrated [(125)I]LTT SRIF(28), [(125)I]CGP 23996 (two radioligands known to label all five recombinant SRIF receptors) and [(125)I]Tyr(3)-octreotide (sst(2) and sst(5) receptor selective) binding in wild type (WT) mouse brain sections; yet no specific binding of [(125)I]Tyr(3)-octreotide in KO mice. In contrast, [(125)I]LTT SRIF(28) and [(125)I]CGP 23996 binding was still present in a number of brain areas in KO mice, although to a lesser degree than in those regions where [(125)I]Tyr(3)-octreotide binding was found, in WT animals. The present data suggest first, that both sst(2) receptor protein and mRNA were completely absent in the brain of these KO animals. Second, there was little evidence of compensatory regulation, at the mRNA level, of the other SRIF receptors as a consequence of the sst(2) KO. Third, the absence of any [(125)I]Tyr(3)-octreotide binding, in KO mice, suggests that this particular ligand is selective for the sst(2) receptor subtype (under the conditions utilised); or that sst(5) receptors are only marginally expressed in brain. Fourth, there were regions where the binding of [(125)I]LTT SRIF(28) and [(125)I]CGP 23996 were moderately affected by the sst(2) KO, suggesting that additional SRIF receptors may well contribute to the binding of the aforementioned radioligands. Finally, since the relative distribution of these two ligands were not entirely superimposable, it suggests that their respective selectivity profiles towards the different SRIF receptor subtypes in situ are not identical.

Psychiatric disorders are often associated with metabolic and hormonal alterations, including obesity, diabetes, metabolic syndrome as well as modifications in several biological rhythms including appetite, stress, sleep–wake cycles, and secretion of their corresponding endocrine regulators. Among the gastrointestinal hormones that regulate appetite and adapt the metabolism in response to nutritional, hedonic, and emotional dysfunctions, at the interface between endocrine, metabolic, and psychiatric disorders, ghrelin plays a unique role as the only one increasing appetite. The secretion of ghrelin is altered in several psychiatric disorders (anorexia, schizophrenia) as well as in metabolic disorders (obesity) and in animal models in response to emotional triggers (psychological stress …) but the relationship between these modifications and the physiopathology of psychiatric disorders remains unclear. Recently, a large literature showed that this key metabolic/endocrine regulator is involved in stress and reward-oriented behaviors and regulates anxiety and mood. In addition, preproghrelin is a complex prohormone but the roles of the other ghrelin-derived peptides, thought to act as functional ghrelin antagonists, are largely unknown. Altered ghrelin secretion and/or signaling in psychiatric diseases are thought to participate in altered appetite, hedonic response and reward. Whether this can contribute to the mechanism responsible for the development of the disease or can help to minimize some symptoms associated with these psychiatric disorders is discussed in the present review. We will thus describe (1) the biological actions of ghrelin and ghrelin-derived peptides on food and drugs reward, anxiety and depression, and the physiological consequences of ghrelin invalidation on these parameters, (2) how ghrelin and ghrelin-derived peptides are regulated in animal models of psychiatric diseases and in human psychiatric disorders in relation with the GH axis.

Ghrelin is a gut hormone processed from the proghrelin peptide acting as the endogenous ligand of the GH secretagogue receptor 1a. The regulatory role of endogenous ghrelin on pulsatile GH secretion and linear growth had to be established. The aim of the present study was to delineate the endogenous actions of preproghrelin on peripheral and central components of the GH axis. Accordingly, the ultradian pattern of GH secretion was measured in young and old preproghrelin-deficient males. Blood samples were collected by tail bleeding every 10 minutes over a period of 6 hours. Analysis of the GH pulsatile pattern by deconvolution showed that GH was secreted in an ultradian manner in all genotypes, with major secretory peaks occurring at about 3-hour intervals. In older mice, the peak number was reduced and secretion was less irregular compared with younger animals. Remarkably, in young Ghrl−/− mice, the amplitude of GH secretory bursts was significantly reduced. In older mice, however, genotype differences were less significant. Changes in GH pulsatility in young Ghrl−/− mice were associated with a tendency for reduced GH pituitary contents and plasma IGF-I concentrations, but with only a minor impact on linear growth. In Ghrl+/− mice, despite reduced Acyl ghrelin to des-acyl ghrelin ratio, GH secretion was not impaired. Ghrelin deficiency was not associated with a reduction in hypothalamic GHRH content or altered response to GHRH stimulation. Therefore, reduction in GHRH production and/or sensitivity do not primarily account for the altered GH pulsatile secretion of young Ghrl−/− mice. Instead, GHRH expression was elevated in young but not old Ghrl−/− mice, suggesting that differential compensatory responses resulting from the absence of endogenous ghrelin is occurring according to age. These results show that endogenous ghrelin is a regulator of GH pulse amplitude in growing mice but does not significantly modulate linear growth.

Supplementary information available for this article at http://www.nature.com/nutd/journal/v4/n7/suppinfo/nutd201417s1.html; BACKGROUND: Contrasting with obesity, constitutional thinness (CT) is a rare condition of natural low bodyweight. CT exhibits preserved menstruation in females, no biological marker of undernutrition, no eating disorders but a bodyweight gain desire. Anorexigenic hormonal profile with high peptide tyrosine tyrosine (PYY) was shown in circadian profile. CT could be considered as the opposite of obesity, where some patients appear to resist diet-induced bodyweight loss.OBJECTIVE: The objective of this study was to evaluate appetite regulatory hormones in CTs in an inverse paradigm of diet-induced weight loss.METHODS: A 4-week fat overfeeding (2640 kJ excess) was performed to compare eight CT women (body mass index (BMI) < 17.5 kg m(-2)) to eight female controls (BMI 18.5-25 k gm(-2)). Appetite regulatory hormones profile after test meal, food intake, bodyweight, body composition, energy expenditure and urine metabolomics profiles were monitored before and after overfeeding.RESULTS: After overfeeding, fasting total and acylated ghrelin were significantly lower in CTs than in controls (P = 0.01 and 0.03, respectively). After overfeeding, peptide tyrosine tyrosine (PYY) and glucagon-like-peptide 1 both presented earlier (T15 min vs T30 min) and higher post-meal responses (incremental area under the curve) in CTs compared with controls. CTs failed to increase bodyweight (+0.22 +/- 0.18 kg, P = 0.26 vs baseline), contrasting with controls (+0.72 +/- 0.26 kg, P = 0.03 vs baseline, P = 0.01 vs CTs). Resting energy expenditure increased in CTs only (P = 0.031 vs baseline). After overfeeding, a significant negative difference between total energy expenditure and food intake was noticed in CTs only (-2754 +/- 720 kJ, P = 0.01).CONCLUSION: CTs showed specific adaptation to fat overfeeding: overall increase in anorexigenic hormonal profile, enhanced post prandial GLP-1 and PYY and inverse to controls changes in urine metabolomics. Overfeeding revealed a paradoxical positive energy balance contemporary to a lack of bodyweight gain, suggesting yet unknown specific energy expenditure pathways in CTs.

To determine whether cellular compartmentalization of somatostatin receptors can be regulated in vivo, we examined the immunocytochemical distribution of the sst2A receptor (sst2AR) after stereotaxical injections of somatostatin analogs into the rat parietal cortex. Whereas CH-275, a sst1R agonist, failed to induce changes in the diffuse sst2AR immunostaining pattern characteristic of control animals, somatodendritic profiles displaying intracytoplasmic immunoreactive granules became apparent short-term after injection of either somatostatin or the sst2R agonist octreotide. Confocal microscopy revealed that 90% of sst2AR-immunoreactive endosome-like organelles displayed transferrin receptor immunoreactivity. At the electron microscopic level, the percentage of sst2AR immunoparticles dramatically decreased at the plasmalemma of perikarya and dendrites after octreotide injection. Conversely, it significantly increased in endosomes-like organelles. These results demonstrate that sst2ARs undergo, in vivo, rapid and massive internalization into the endocytic recycling compartment in response to acute agonist stimulation and provide important clues toward elucidating somatostatin receptor signaling in the mammalian brain.