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Pancreatic neuroendocrine tumours (islet-cell tumours) are rare and usually sporadic, but they may be associated with complex familial endocrine cancer syndromes. Recognized types of pancreatic neuroendocrine tumours are those that are non-functioning (often advanced at diagnosis and presenting with mass effects due to the absence of symptoms attributable to hormone hypersecretion), insulinoma (the most frequent type), and others including gastrinoma, VIPoma, and glucagonoma. The following should be considered in addition to the symptomatic treatments: surgical resection—the only curative treatment, but not possible in many cases; tyrosine kinase inhibitors which inhibit specific kinases involved in tumour cell proliferation, growth, and angiogenesis; mammalian Target of Rapamycin (mTOR) inhibitors; peptide-receptor radionuclide therapy (radiolabelled somatostatin analogues).

Carcinoid syndrome is a paraneoplastic syndrome caused by the release of 5-hydroxytryptamine (5-HT or serotonin) and other vasoactive substances from neuroendocrine tumours (NETs), typically those arising from the duodenum, jejunum, ileum, and also from bronchial NETs. Characteristic clinical features, which typically arise when tumours have metastasized to the liver, are flushing and secretory diarrhoea, and occasionally wheezing. Carcinoid crisis is an acute and life-threatening manifestation with sustained flushing, hyperdynamic shock, and acute kidney injury. Carcinoid heart disease typically manifests in the right side of the heart with valvular insufficiency and heart failure. Diagnosis is made by a combination of the characteristic clinical syndrome, biochemical markers such as 5-hydroxyindoleacetic acid (a metabolite of 5-HT), histopathological examination of tissue from tumour deposits, and imaging with conventional cross-sectional modalities as well as somatostatin receptor scintigraphy. Treatment is most often directed at control of symptoms, with the standard of care for control of the carcinoid syndrome being a somatostatin analogue. Symptomatic therapies are used to palliate diarrhoea. Niacin supplements should be given to forestall the development of pellagra. Multiple modalities for treatment of NETs exist, which should be directed by a multidisciplinary team. Their general prognosis is good with median overall survival of 9.3 years, but the presence of carcinoid syndrome cuts this to 5.0 years, and less in the presence of heart disease.

ObjectiveRoux-en-Y gastric bypass (RYGB) surgery is currently the most effective treatment for diabetes and obesity. An increasingly recognized and highly disabling complication of RYGB is postprandial hypoglycaemia (PPH). The pathophysiology of PPH remains unclear with multiple mechanisms suggested including nesidioblastosis, altered insulin clearance and increased glucagon-like peptide-1 (GLP-1) secretion. Whilst many PPH patients respond to dietary modification, some have severely disabling symptoms. Multiple treatments are proposed, including dietary modification, GLP-1 antagonism, GLP-1 analogues and even surgical reversal, with none showing a more decided advantage over the others. A greater understanding of the pathophysiology of PPH could guide the development of new therapeutic strategies.MethodsWe studied a cohort of PPH patients at the Imperial Weight Center. We performed continuous glucose monitoring to characterize their altered glycaemic variability. We also performed a mixed meal test (MMT) and measured gut hormone concentrations.ResultsWe found increased glycaemic variability in our cohort of PPH patients, specifically a higher mean amplitude glucose excursion (MAGE) score of 4.9. We observed significantly greater and earlier increases in insulin, GLP-1 and glucagon in patients who had hypoglycaemia in response to an MMT (MMT Hypo) relative to those that did not (MMT Non-Hypo). No significant differences in oxyntomodulin, GIP or peptide YY secretion were seen between these two groups.ConclusionAn early peak in GLP-1 and glucagon may together trigger an exaggerated insulinotropic response to eating and consequent hypoglycaemia in patients with PPH.

Aim: The performance of glucagon and GLP-1 immunoassays is often poor, but few sensitive LC–MS/MS methods exist as alternatives. Experimental: A multiplexed LC–MS/MS method using a 2D extraction technique was developed. Results: The method was established for the quantitation of endogenous glucagon (LLOQ: 15 pg/ml) and dosed GLP-1 (LLOQ: 25 pg/ml) in human plasma, and is the first such method avoiding immunoenrichment. Specificity of endogenous glucagon quantitation was assured using a novel approach with a supercharging mobile phase additive to access a sensitive qualifier SRM. Endogenous glucagon concentrations were within the expected range, and showed good reproducibility after extended sample storage. A cross-validation against established immunoassays using physiological study samples demonstrated some similarities between methods. Conclusion: The LC–MS/MS method offers a viable alternative to immunoassays for quantitation of endogenous glucagon, dosed glucagon and/or dosed GLP-1.

STUDY QUESTION Are novel circulating placental markers prokineticin-1 (PK-1), soluble fms-like tyrosine kinase-1 (sFlt-1), soluble endoglin (sEng) and placental growth factor (PlGF) associated with late miscarriage in asymptomatic first trimester pregnant women? SUMMARY ANSWER Increased serum sFlt-1 or PlGF, but not sEng or PK-1, were significantly associated with reduced miscarriage risk after adjustment for age, BMI, gestational age, smoking and blood pressure. WHAT IS KNOWN ALREADY Abnormal placental development is observed in two-thirds of miscarriages. Identifying women at high risk of late miscarriage could help diagnose potentially treatable causes of miscarriage such as infection, thrombosis or immunological disease. Recently, the circulating placental markers PK-1, sFlt-1, sEng and PlGF have been identified; however, it is not known if circulating levels of these markers are associated with late miscarriage. STUDY DESIGN, SIZE, DURATION A single-centre observational cohort study with prospectively collected data was carried out at a tertiary care centre 2010-2012, in 993 asymptomatic pregnant women. Plasma PK-1, and serum sEng, sFlt-1 and PlGF were measured once in each patient during the antenatal booking visit, and pregnancy outcome was monitored prospectively. Less than 1% of patients were lost to follow-up. Multiples of median (MOM) levels were calculated to adjust for gestational age. PARTICIPANTS/MATERIALS, SETTING, METHODS Nine-hundred and ninety-three asymptomatic pregnant women attending antenatal clinic for a routine booking antenatal appointment were recruited to the study, of whom 12 were lost to follow-up and excluded from analysis. Of the cohort, 50 of the remaining 981 women suffered late miscarriage. MAIN RESULTS AND THE ROLE OF CHANCE Gestation-adjusted sEng, sFlt-1 and PlGF levels were 11% (P

Aims To investigate, for a given energy expenditure (EE) rise, the differential effects of glucagon infusion and cold exposure on brown adipose tissue (BAT) activation in humans. Methods Indirect calorimetry and supraclavicular thermography was performed in 11 healthy male volunteers before and after: cold exposure; glucagon infusion (at 23 °C); and vehicle infusion (at 23 °C). All volunteers underwent 18F-fluorodeoxyglucose (18F-FDG) positron emission tomography (PET)/CT scanning with cold exposure. Subjects with cold-induced BAT activation on 18F-FDG PET/CT (n = 8) underwent a randomly allocated second 18F-FDG PET/CT scan (at 23 °C), either with glucagon infusion (n = 4) or vehicle infusion (n = 4). Results We observed that EE increased by 14% after cold exposure and by 15% after glucagon infusion (50 ng/kg/min; p < 0.05 vs control for both). Cold exposure produced an increase in neck temperature (+0.44 °C; p < 0.001 vs control), but glucagon infusion did not alter neck temperature. In subjects with a cold-induced increase in the metabolic activity of supraclavicular BAT on 18F-FDG PET/CT, a significant rise in the metabolic activity of BAT after glucagon infusion was not detected. Cold exposure increased sympathetic activation, as measured by circulating norepinephrine levels, but glucagon infusion did not. Conclusions Glucagon increases EE by a similar magnitude compared with cold activation, but independently of BAT thermogenesis. This finding is of importance for the development of safe treatments for obesity through upregulation of EE.

The prevalence of obesity is steadily rising and has huge health and financial implications for society. Weight gain is due to an imbalance between dietary intake and energy expenditure and research has focused on trying to understand the complex pathways involved in controlling these aspects. This review highlights the key areas of research in the hypothalamic control of appetite. The hypothalamus consists of several nuclei that integrate peripheral signals, such as adiposity and caloric intake, to regulate important pathways within the CNS controlling food intake. The best characterized pathways are the orexigenic neuropeptide Y/Agouti-related protein and the anorexigenic pro-opiomelanocortin/cocaine- and amphetamine-related transcript neurons in the arcuate nucleus of the hypothalamus. These project from the arcuate nucleus to other key hypothalamic nuclei, such as the paraventricular, dorsomedial, ventromedial and lateral hypothalamic nuclei. There are also projections to and from the brainstem, cortical areas and reward pathways, all of which influence food intake. The challenge at present is to understand the complexity of these pathways and try to find ways of modulating them in order to find potential therapeutic targets.

Introduction Glucagon like-peptide 1 receptor (GLP-1R) agonists diminish appetite and may contribute to the weight loss in inflammatory bowel disease (IBD). Objectives The aim of this study was to determine, for the first time, the expression of GLP-1R by colon nerve fibres in patients with IBD, and functional effects of its agonists in cultured rat and human sensory neurons. Methods GLP-1R and other nerve markers were studied by immunohistochemistry in colon biopsies from patients with IBD (n = 16) and controls (n = 8), human dorsal root ganglia (DRG) tissue, and in GLP-1R transfected HEK293 cells. The morphological effects of incretin hormones oxyntomodulin, exendin-4 and glucagon were studied on neurite extension in cultured DRG neurons, and their functional effects on capsaicin and ATP signalling, using calcium imaging. Results Significantly increased numbers of colonic mucosal nerve fibres were observed in IBD biopsies expressing GLP-1R (p = 0.0013), the pan-neuronal marker PGP9.5 (p = 0.0008), and sensory neuropeptide CGRP (p = 0.0014). An increase of GLP-1R positive nerve fibres in IBD colon was confirmed with a different antibody to GLP-1R (p = 0.016). GLP-1R immunostaining was intensely positive in small and medium-sized neurons in human DRG, and in human and rat DRG cultured neurons. Co-localization of GLP-1R expression with neuronal markers in colon and DRG confirmed the neural expression of GLP-1R, and antibody specificity was confirmed in HEK293 cells transfected with the GLP-1R. Treatment with oxyntomodulin, exendin-4 and GLP-1 increased neurite length in cultured neurons compared with controls, but did not stimulate calcium influx directly, or affect capsaicin responses. However, exendin-4 significantly enhanced ATP responses in human DRG neurons. Conclusion Our results show that increased GLP-1R innervation in IBD bowel could mediate enhanced visceral afferent signalling, and provide a peripheral target for therapeutic intervention. The differential effect of GLP-1R agonists on capsaicin and ATP responses in neurons suggest they may not affect pain mechanisms mediated by the capsaicin receptor TRPV1, but may enhance the effects of purinergic agonists.

STUDY QUESTION How potently does the novel hypothalamic stimulator of reproduction, kisspeptin, increase gonadotrophin secretion when compared with GnRH in healthy men? SUMMARY ANSWER At the doses tested, intravenous administration of either of two major kisspeptin isoforms, kisspeptin-10 and -54, was associated with similar levels of gonadotrophin secretion in healthy men; however, GnRH was more potent when compared with either kisspeptin isoform. WHAT IS KNOWN ALREADY Kisspeptin-10 and -54 are naturally occurring hormones in the kisspeptin peptide family which potently stimulates endogenous GnRH secretion from the hypothalamus, so have the potential to treat patients with reproductive disorders. Rodent studies suggest that kisspeptin-54 is more potent when compared with kisspepitn-10; however, their effects have not previously been directly compared in humans, or compared with direct pituitary stimulation of gonadotrophin secretion using GnRH. STUDY DESIGN, SIZE AND DURATION A single-blinded placebo controlled physiological study was performed from January to December 2013. Local ethical approval was granted, and five participants were recruited to each dosing group. PARTICIPANTS/MATERIALS, SETTING, METHODS Healthy men were administered vehicle, kisspeptin-10, kisspeptin-54 and GnRH intravenously for 3 h on different study days. Each hormone was administered at 0.1, 0.3 and 1.0 nmol/kg/h doses (n = 5 subjects per group). Regular blood sampling was conducted throughout the study to measure LH and FSH. Study visits were conducted at least a week apart. MAIN RESULTS AND THE ROLE OF CHANCE Serum LH and FSH levels were ∼3-fold higher during GnRH infusion when compared with kisspeptin-10 and ∼2-fold higher when compared with kisspeptin-54 [mean area under the curve serum LH during infusion (in hours times international units per litre, h.IU/l): 10.81 ± 1.73, 1.0 nmol/kg/h kisspeptin-10; 14.43 ± 1.27, 1.0 nmol/kg/h kisspeptin-54; 34.06 ± 5.18, 1.0 nmol/kg/h GnRH, P < 0.001 versus kisspeptin-10, P < 0.01 versus kisspeptin-54]. LIMITATIONS, REASONS FOR CAUTION This study had a small sample size. WIDER IMPLICATIONS OF THE FINDINGS Kisspeptin offers a novel means of stimulating the reproductive axis. Our data suggest that kisspeptin stimulates gonadotrophin secretion less potently when compared with GnRH; however, kisspeptin may stimulate gonadotrophins in a more physiological manner when compared with current therapies. Kisspeptin is emerging as a future therapeutic agent, so it is important to establish which kisspeptin hormones could be used to treat patients with infertility. Results of this study suggest that either isoform has similar effects on reproductive hormone secretion in healthy men when administered intravenously. STUDY FUNDING/COMPETING INTERESTS This work is funded by grants from the MRC and NIHR and is supported by the NIHR Imperial Biomedical Research Centre Funding Scheme. C.N.J. is supported by an NIHR Clinical Lectureship. A.A. is supported by Wellcome Trust Research Training Fellowships. A.N.C. is supported by Wellcome Trust Translational Medicine Training Fellowship. W.S.D. is supported by an NIHR Career Development Fellowship.

The brain relies on a constant supply of glucose, its primary fuel, for optimal function. A taste-independent mechanism within the CNS that promotes glucose delivery to the brain has been postulated to maintain glucose homeostasis; however, evidence for such a mechanism is lacking. Here, we determined that glucokinase activity within the hypothalamic arcuate nucleus is involved in regulation of dietary glucose intake. In fasted rats, glucokinase activity was specifically increased in the arcuate nucleus but not other regions of the hypothalamus. Moreover, pharmacologic and genetic activation of glucokinase in the arcuate nucleus of rodent models increased glucose ingestion, while decreased arcuate nucleus glucokinase activity reduced glucose intake. Pharmacologic targeting of potential downstream glucokinase effectors revealed that ATP-sensitive potassium channel and P/Q calcium channel activity are required for glucokinase-mediated glucose intake. Additionally, altered glucokinase activity affected release of the orexigenic neurotransmitter neuropeptide Y in response to glucose. Together, our results suggest that glucokinase activity in the arcuate nucleus specifically regulates glucose intake and that appetite for glucose is an important driver of overall food intake. Arcuate nucleus glucokinase activation may represent a CNS mechanism that underlies the oft-described phenomena of the "sweet tooth" and carbohydrate craving.

The obesity pandemic presents a significant burden, both in terms of healthcare and economic outcomes, and current medical therapies are inadequate to deal with this challenge. Bariatric surgery is currently the only therapy available for obesity which results in long-term, sustained weight loss. The favourable effects of this surgery are thought, at least in part, to be mediated via the changes of gut hormones such as GLP-1, PYY, PP and oxyntomodulin seen following the procedure. These hormones have subsequently become attractive novel targets for the development of obesity therapies. Here, we review the development of these gut peptides as current and emerging therapies in the treatment of obesity.

Objective: To investigate whether gastric bypass induces a higher activity of brown adipose tissue and greater levels of the brown adipose tissue-specific protein uncoupling protein-1 (UCP-1) in rats. Methods: Gastric bypass rats and sham-operated controls (each n = 8) underwent whole body 1H-MR spectroscopy for analysis of body composition and 18F-fluorodeoxyglucose positron emission tomography combined with computed tomography (18F-FDG PET/CT) imaging for measurement of the metabolic activity of brown adipose tissue. Brown adipose tissue was harvested and weighed, and UCP-1 mRNA content was measured by Northern Blot technique. Results: Gastric bypass rats had a significantly lower percentage of whole body adipose tissue mass compared to sham-operated rats (p = 0.001). There was no difference in brown adipose tissue activity between the two groups (standardised uptake value sham 2.81 ± 0.58 vs. bypass 2.56 ± 0.46 ; p = 0.73). Furthermore, there was no difference in the UCP-1 mRNA content of brown adipose tissue between the two groups (sham 49.5 ± 13.2 vs. bypass 43.7 ± 13.1; p = 0.77). Conclusion: Gastric bypass does not increase the activity of brown adipose tissue in rats suggesting that other mechanisms are involved to explain the increased energy expenditure after bypass surgery. Our results cannot justify the radiation dose of 18F-FDG PET/CT studies in humans to determine potential changes in brown adipose tissue after gastric bypass surgery.

The global prevalence of type 2 diabetes mellitus and impaired glucose metabolism continues to rise in conjunction with the pandemic of obesity. The metabolic Roux-en-Y gastric bypass operation offers the successful resolution of diabetes in addition to sustained weight loss and excellent long-term outcomes in morbidly obese individuals. The procedure consists of the physiological BRAVE effects: (i) Bile flow alteration; (ii) Reduction of gastric size; (iii) Anatomical gut rearrangement and altered flow of nutrients; (iv) Vagal manipulation and (v) Enteric gut hormone modulation. This operation provides anti-diabetic effects through decreasing insulin resistance and increasing the efficiency of insulin secretion. These metabolic outcomes are achieved through weight-independent and weight-dependent mechanisms. These include the foregut, midgut and hindgut mechanisms, decreased inflammation, fat, adipokine and bile metabolism, metabolic modulation, shifts in gut microbial composition and intestinal gluconeogenesis. In a small minority of patients, gastric bypass results in hyperinsulinaemic hypoglycaemia that may lead to nesidioblastosis (pancreatic beta-cell hypertrophy with islet hyperplasia). Elucidating the precise metabolic mechanisms of diabetes resolution and hyperinsulinaemia after surgery can lead to improved operations and disease-specific procedures including 'diabetes surgery'. It can also improve our understanding of diabetes pathogenesis that may provide novel strategies for the management of metabolic syndrome and impaired glucose metabolism.

Aim: Relaxin is a polypeptide hormone involved in pregnancy and lactation. It is mainly secreted by the corpus luteum and placenta, but is expressed in a number of other tissues, including heart and brain. Within the brain, relaxin is expressed in the olfactory and limbic systems, the cortex and the hypothalamic arcuate nucleus (ARC). Its cognate receptor, relaxin family peptide receptor 1 (RXFP1), is also widely expressed in the brain, including the hypothalamic ARC and paraventricular nucleus (PVN), areas important in appetite regulation. The aim of this study was to investigate whether relaxin influences food intake through central hypothalamic circuits. Methods: The human form of relaxin, human relaxin-2 (H2) was administered centrally and peripherally to male Wistar rats and food intake measured. Behaviour was also assessed. Results: Intracerebroventricular (ICV) administration of H2 significantly decreased 1-h food intake in the early dark phase [2.95 ± 0.45 g (saline) vs. 0.95 ± 0.18 g (180 pmol H2), p < 0.001]. ICV administration of H2 decreased feeding behaviour and increased grooming and headdown behaviour. Intraparaventricular injections of H2 significantly decreased 1-h food intake in the early dark phase [3.13 ± 0.35 g (saline) vs. 1.35 ± 0.33 g (18 pmol H2), p < 0.01, 1.61 ± 0.31 g (180 pmol H2), p < 0.05 and 1.23 ± 0.32 g (540 pmol H2), p < 0.001]. Intraperitoneal (IP) administration of H2 significantly decreased 1-h food intake in the early dark phase [4.63 ± 0.46 g (vehicle) vs. 3.08 ± 0.15 g (66 nmol H2), p < 0.01, 3.00 ± 0.17 g (200 nmol H2), p < 0.01 and 2.26 ± 0.36 g (660 nmol H2), p < 0.001]. Conclusions: Central and peripheral administration of H2 reduces the food intake in rats. This effect may be mediated via the PVN and/or other brain regions.

BACKGROUND AND PURPOSE Alarin is a recently discovered member of the galanin peptide family encoded by a splice variant of galanin-like peptide (GALP) mRNA. Galanin and GALP regulate energy homeostasis and reproduction. We therefore investigated the effects of alarin on food intake and gonadotrophin release. EXPERIMENTAL APPROACH Alarin was administered into the third cerebral ventricle (i.c.v.) of rats, and food intake or circulating hormone levels were measured. The effect of alarin on the hypothalamo–pituitary–gonadal axis was investigated in vitro using hypothalamic and anterior pituitary explants, and immortalized cell lines. Receptor binding assays were used to determine whether alarin binds to galanin receptors. KEY RESULTS The i.c.v. administration of alarin (30 nmol) to ad libitum fed male rats significantly increased acute food intake to 500%, and plasma luteinizing hormone (LH) levels to 170% of responses to saline. In vitro, 100 nM alarin stimulated neuropeptide Y (NPY) and gonadotrophin-releasing hormone (GnRH) release from hypothalamic explants from male rats, and 1000 nM alarin increased GnRH release from GT1-7 cells. In vivo, pretreatment with the GnRH receptor antagonist cetrorelix prevented the increase in plasma LH levels observed following i.c.v. alarin administration. Receptor binding studies confirmed alarin did not bind to any known galanin receptor, or compete with radiolabelled galanin for hypothalamic binding sites. CONCLUSIONS AND IMPLICATIONS These results suggest alarin is a novel orexigenic peptide, and that it increases circulating LH levels via hypothalamic GnRH. Further work is required to identify the receptor(s) mediating the biological effects of alarin.

To determine the efficacy of a long-acting oxyntomodulin (OXM) analogue, OXM6421, in inhibiting food intake and decreasing body weight in lean and diet-induced obese (DIO) rodents.The glucagon-like peptide-1 (GLP-1) receptor binding affinity and efficacy, sensitivity to enzymatic degradation in vitro and persistence in the circulation after peripheral administration were investigated for OXM6421 and compared with native OXM. The chronic effect of OXM6421 on food intake, body weight and energy expenditure was examined in lean rats, and its anti-obesity potential was evaluated in DIO mice.OXM6421 showed enhanced GLP-1 receptor binding affinity and cyclic adenosine monophosphate (cAMP) stimulation, and higher resistance to enzymatic degradation by dipeptidyl peptidase IV (DPP-IV) and neutral endopeptidase (NEP) compared with native OXM. OXM6421 persisted longer in the circulation than OXM after peripheral administration. Acute administration of OXM6421 potently inhibited food intake in lean rodents, with cumulative effects lasting up to 24 h. In lean rats, daily subcutaneous (s.c.) administration of OXM6421 caused greater weight loss than the pair-fed animals, and a higher rate of oxygen consumption than both the pair-fed and the saline controls. In DIO mice, continuous s.c. infusion of OXM6421 resulted in a significant weight loss, accompanied by an improvement in glucose homeostasis and an increase in circulating adiponectin levels. Once-daily s.c. administration of OXM6421 for 21 days caused sustained weight loss in DIO mice.OXM6421 induces negative energy balance in both lean and obese rodents, suggesting that long-acting OXM analogues may represent a potential therapy for obesity.

Kisspeptin signaling via the kisspeptin receptor G-protein-coupled receptor-54 plays a fundamental role in the onset of puberty and the regulation of mammalian reproduction. In this immunocytochemical study we addressed the (i) topography, (ii) sexual dimorphism, (iii) relationship to gonadotropin-releasing hormone (GnRH) neurons and (iv) neurokinin B content of kisspeptin-immunoreactive hypothalamic neurons in human autopsy samples. In females, kisspeptin-immunoreactive axons formed a dense periventricular plexus and profusely innervated capillary vessels in the infundibular stalk. Most immunolabeled somata occurred in the infundibular nucleus. Many cells were also embedded in the periventricular fiber plexus. Rostrally, they formed a prominent periventricular cell mass (magnocellular paraventricular nucleus). Robust sex differences were noticed in that fibers and somata were significantly less numerous in male individuals. In dual-immunolabeled specimens, fine kisspeptin-immunoreactive axon varicosities formed axo-somatic, axo-dendritic and axo-axonal contacts with GnRH neurons. Dual-immunofluorescent studies established that 77% of kisspeptin-immunoreactive cells in the infundibular nucleus synthesize the tachykinin peptide neurokinin B, which is known to play crucial role in human fertility; 56 and 17% of kisspeptin fibers in the infundibular and periventricular nuclei, respectively, contained neurokinin B immunoreactivity. Site-specific co-localization patterns implied that kisspeptin neurons in the infundibular nucleus and elsewhere contributed differentially to these plexuses. This study describes the distribution and robust sexual dimorphism of kisspeptin-immunoreactive elements in human hypothalami, reveals neuronal contacts between kisspeptin-immunoreactive fibers and GnRH cells, and demonstrates co-synthesis of kisspeptins and neurokinin B in the infundibular nucleus. The neuroanatomical information will contribute to our understanding of central mechanisms whereby kisspeptins regulate human fertility.

Background and purpose: The kisspeptins are critical regulators of reproduction and a therapeutic target for reproductive disease. Intracerebroventricular (i.c.v.) or peripheral injection of kisspeptin potently stimulates the hypothalamic-pituitary gonadal (HPG) axis via gonadotrophin-releasing hormone (GnRH). However, little is known regarding the effects of kisspeptin administration on testicular function. We investigated the mechanism(s) of kisspeptin-induced testicular degeneration in the rat. Experimental approach: Kisspeptin-54 (50 nmol·day−1) was continuously administered subcutaneously (6 h to 3 days) to male Wistar rats and reproductive hormones and testicular histology analysed. We also investigated the effects of a single subcutaneous injection of 0.5, 5 or 50 nmol kisspeptin-54. In order to determine whether the testicular degeneration observed is peripherally or centrally mediated, we investigated effects of i.c.v. injections of 5 nmol kisspeptin-54 and pre-administered a GnRH-receptor antagonist (cetrorelix) to rats peripherally treated with kisspeptin-54. Key results: Continuous subcutaneous administration of kisspeptin-54 caused testicular degeneration after only 12 h, when gonadotrophins were still markedly raised, suggesting that the degeneration is independent of the desensitization of the HPG axis to kisspeptin-54. Furthermore, a single subcutaneous injection of kisspeptin-54 caused dose-dependent testicular degeneration. Continuous kisspeptin-54 administration is thus not required to cause testicular degeneration. Pretreatment with cetrorelix blocked kisspeptin-induced testicular degeneration, and a single i.c.v. injection of kisspeptin-54 caused testicular degeneration, suggesting it is GnRH-mediated. Conclusions and implications: Kisspeptin-induced testicular degeneration appears to be centrally mediated, and result from acute hyper-stimulation of the HPG axis. Doses must be carefully considered if kisspeptin is to be used therapeutically. Mandarin translation of abstract

SUMMARY Galanin was infused intravenously into eight healthy volunteers at a dose of 40 pmol kg−1 min−1 for 1 h to investigate the pharmacological effects of this peptide on the postprandial sialagogical response in man. Galanin significantly increased the salivary volume and the saliva output of sodium, chloride and bicarbonate compared to control saline infusion, but had no effect on the output of potassium and α-amylase. An increase in salivary pH was also observed. The increase in salivary volume may indicate a physiological role of galanin in the control of salivary secretion.

SMS 201-995 is an octapeptide analogue of somatostatin. The effect of a single subcutaneous (s.c.) injection of 50 micrograms SMS 201-995 on post-prandial intermediary metabolism was investigated in normal subjects. In spite of a long-lasting post-prandial suppression of insulin secretion, there were no significant changes in the plasma concentration of alanine, glycerol, 3-OH-butyrate or lactate. However, SMS 201-995 impairs carbohydrate tolerance, probably due to inhibition of insulin secretion. Basal and post-prandial plasma concentrations of the gut regulatory peptides pancreatic glucagon, motilin, pancreatic polypeptide, gastric inhibitory polypeptide, enteroglucagon, gastrin and peptide YY were suppressed up to 5 hours after subcutaneous administration of a single dose of SMS 201-995.

SUMMARY Therapy for diarrhoea associated with the carcinoid syndrome is often unsatisfactory. In an open study ICS 205-930 (Sandoz Limited), a novel 5HT3-antagonist, controlled diarrhoea in five of six patients studied. This drug may be a useful advance in the symptomatic treatment of the carcinoid syndrome.

SUMMARY Although enterochromaffin-like (ECL) cells form the major endocrine cell population of the non-antral stomach, they have been largely overlooked in the study of gastric disease. In the human, their product and functions are unknown, but they are associated with histamine secretion in rodents. The cells are controlled by neural and hormonal factors, the most significant of the latter being gastrin. Interest in ECL cells has been stimulated by the observation that hyperplasia of these cells, sometimes leading to formation of gastric carcinoid tumours, occurs in conditions of persistent hypergastrinaemia — for example, in response to the achlorhydria of individuals with pernicious anaemia. The advent of new highly potent inhibitors of gastric acid secretion is allowing more information to be obtained on the physiology and functions of the ECL cell. However, there is clearly a great deal more to be discovered about this enigmatic endocrine cell type.

Plasma samples of seven patients with gut and pancreatic endocrine tumours who have been on long-term treatment with a long-acting somatostatin analogue (SMS 201-995) were investigated for endogenous antibodies to the peptide by incubation with radiolabelled SMS 201-995. The duration of treatment with the somatostatin analogue was between 9 and 26 months and the dose from 100 to 300 micrograms day-1. In none of the patients could antibodies to SMS be detected. The effect of this somatostatin analogue is unlikely to be impaired by formation of endogenous antibodies, even after long-term treatment.

Prognosis in patients with neuroendocrine tumors (NETs) is often poor, frequently reflecting delayed diagnosis. Hence, accurate and practical NET markers are needed. Cocaine- and amphetamine-regulated transcript (CART) peptide is a potential novel NET marker.Circulating levels of CART peptide and the established NET markers chromogranin A (CgA) and chromogranin B (CgB) were measured using RIA in 353 patients with NET (normal renal function) and in controls. Clinical data were collected retrospectively.The comparative and combined utility of CART, CgA, and CgB for diagnosis and assessment of disease progression was measured in different NET subtypes.CgA and CgB in combination improved diagnostic accuracy in patients with gut NETs, nongastroenteropancreatic NETs, and NETs with an unknown primary origin compared with each biomarker alone. Measuring CART did not further improve diagnosis in these NET subtypes. For pancreatic NETs, CgB was superior to CgA and CART in detecting stable disease (P.007), whereas CgA and CART in combination were most effective in identifying progressive disease. In phaeochromocytomas/paragangliomas (PCC/PGL), CART was the most useful biomarker for identifying stable (P.001) and progressive (P = .001) disease. Consistent with this, plasma CART decreased following PCC/PGL tumor resection, remaining low in all patients in remission, but increasing in those with progressive disease.CART is a useful marker for identifying progressive pancreatic NETs. CART is superior to CgA and CgB in detecting stable and progressive PCC/PGLs, and may have a role as a surveillance marker for PCC/PGL patients.

This review provides a summary of currently available pharmaceutical therapies for the treatment of obesity, along with an overview of the pipeline of products currently in development, and the key mechanisms on which the major development candidates are based. In particular, the recent increase in understanding of the role of gut peptides in energy homeostasis is highlighted as a promising source of potential future obesity therapies.

Oxyntomodulin has recently been found to decrease body-weight in obese humans and may be a potential anti-obesity therapy.To determine whether oxyntomodulin alters energy expenditure, in addition to reducing energy intake, in 'free-living' overweight and obese volunteers.Randomized double-blind controlled cross-over trial.Community and hospital-based.Fifteen healthy overweight and obese men and women (age: 23-49 years, BMI: 25.1-39.0 kg/m(2)). All volunteers completed the study protocol.Four-day subcutaneous self-administration of pre-prandial oxyntomodulin, three times daily. Participants were advised to maintain their normal dietary and exercise regimen.(1) Energy expenditure, measured by indirect calorimetry and combined heart rate and movement monitoring; (2) energy intake, measured during a study meal.Oxyntomodulin administration reduced energy intake at the study meal by 128+/-29 kcal (P=0.0006) or 17.3+/-5.5% (P=0.0071), with no change in meal palatability. Oxyntomodulin did not alter resting energy expenditure; but increased activity-related energy expenditure by 143+/-109 kcal/day or 26.2+/-9.9% (P=0.0221); total energy expenditure by 9.4+/-4.8% (P=0.0454) and physical activity level by 9.5+/-4.6% (P=0.0495). A reduction in body weight of 0.5+/-0.2% was observed during the oxyntomodulin administration period (P=0.0232).Oxyntomodulin increases energy expenditure while reducing energy intake resulting in negative energy balance. This data supports the role of oxyntomodulin as a potential anti-obesity therapy.

BackgroundBariatric surgery is the most effective treatment for achieving long-term weight loss in morbidly obese patients. This study investigated prospective changes in gut hormones and metabolic indices after Roux-en-ϒ gastric bypass (RYGB).MethodsSix patients were seen before, and at 1, 3 and 6 months after operation. Blood was collected after a 12-h fast and at regular intervals after a mixed 420-kcal meal. Hormonal responses were determined, and comparisons between basal levels and areas under the curve were made. Visual analogue scores were used to assess satiety, hunger and nausea.ResultsMean body mass index decreased from 48·3 kg/m2 before surgery to 36·4 kg/m2 6 months after RYGB. This was accompanied by a decrease in fasting leptin (P < 0·001) and insulin (P = 0·021) levels. At 1, 3 and 6 months after operation, progressively increasing peptide YY (P < 0·001), enteroglucagon (P = 0·045) and glucagon-like peptide 1 (P = 0·042) responses were observed. There was no change in fasting ghrelin levels (P = 0·144). Postprandial satiety was significantly increased by 1 month after surgery and this was maintained until the end of the study (P < 0·001).ConclusionRYGB resulted in substantial weight loss with enhanced postprandial satiety, a sustained weight plateau, and proportionate reduction in fasting insulin and leptin levels. Lack of the expected increase in appetite and food intake as components of a counter-regulatory response may be explained by gut adaptation and the consequent graded rise in the levels of gut hormones that promote satiety.

Background: Published LC–MS/MS methods are not sensitive enough to quantify endogenous levels of glucagon. Results: An ultra high performance liquid chromatography-MS/MS (SRM) method for the quantitation of endogenous levels glucagon was successfully developed and qualified. A novel 2D extraction procedure was used to reduce matrix suppression, background noise and interferences. Glucagon levels in samples from healthy volunteers were found to agree with radioimmunoassay (RIA) derived literature values. Bland–Altman analysis showed a concentration-dependent positive bias of the LC/MS–MS assay versus an RIA. Both assays produced similar pharmacokinetic profiles, both of which were feasible considering the nature of the study. Conclusion: Our method is the first peer reviewed LC–MS/MS method for the quantitation of endogenous levels of glucagon, and offers a viable alternative to RIA-based approaches.

Peptide YY3-36 (PYY(3-36)), glucagon-like peptide-1 (GLP-1), oxyntomodulin and cholecystokinin (CCK) are gastrointestinal-derived hormones that are released postprandially in proportion to the amount of calories ingested. All significantly reduce food intake following peripheral administration to rodents. We have investigated the effect of handling, exposure to a novel environment or to environmental enrichment on the anorectic effect of these gut hormones.Results suggest that the transfer of a rat into a novel environment (cage change) inhibits the anorectic response to peripherally administered PYY(3-36) and oxyntomodulin (1 h food intake reduction (% saline control): PYY/home cage 82.3 +/- 5.9%, P0.05; PYY/clean cage 103.4 +/- 9.7%; oxyntomodulin/home cage 71.6 +/- 12.1%, P0.05; oxyntomodulin/clean cage 103.0 +/- 8.5%) and attenuates the anorectic response to GLP-1 and CCK (1 h food intake reduction (% saline control): GLP-1/home cage 68.8 +/- 6.4%, P0.01; GLP-1/clean cage 80.0 +/- 9.3%; CCK/home cage 49.8 +/- 6.2%, P0.001; CCK/clean cage 69.4 +/- 10.6%, P0.05). We have also observed that exposure to a novel environment does not alter anorectic effect of peripherally administered melanocortin 3/4 receptor agonist, melanotan II (MTII) (1 h food intake reduction (% saline control): MTII/home cage 32.0 +/- 6.3%, P0.001; MTII/clean cage 24.8 +/- 4.2%, P0.001). The attenuation in food intake observed following exposure to a novel environment can be attributed, in part, to a significant reduction in the food intake of the saline treated animals. In a further study, the anorectic effect of peripherally administered PYY(3-36) is attenuated in unhandled rats (88 +/- 4.2% saline control, P = ns) or rats exposed to environmental enrichment (103.3 +/- 9.7% saline control, P = ns), but not in animals that were handled extensively prior to the study (80.1 +/- 7.3% saline control, P0.05).These studies highlight the importance of handling, acclimatisation and habituation of rodents to experimental conditions prior to investigating the ability of gut hormones to alter food intake.

Abnormalities of the melanocortin system produce obesity and increased linear growth. While the obesity phenotype is well characterised, the mechanism responsible for increased linear growth is unclear. The somatotrophic axis was studied in the obese agouti (A(y)/a) mouse as a model of a perturbed melanocortin system.Adult obese A(y)/a mice were compared to age- and sex-matched wild-type (WT) controls. Weight and body length (nose-anus) were recorded. Plasma growth hormone (GH), insulin-like growth factor-I (IGFI), insulin and leptin were measured using radioimmunoassay. Since ghrelin is a potent GH secretagogue, plasma ghrelin, stomach ghrelin peptide and stomach ghrelin mRNA expression were studied. Hypothalamic periventricular (PeVN) somatostatin neurones and arcuate (Arc) neuropeptide Y (NPY) neurones inhibit the growth axis, whereas Arc growth hormone-releasing hormone (GHRH) neurones are stimulatory. Therefore, specific hypothalamic expression of somatostatin, NPY and GHRH was measured using quantitative in situ hybridisation.Obese A(y)/a mice were significantly heavier and longer than WT controls. Plasma IGFI concentrations were 30% greater in obese A(y)/a mice. Obese A(y) /a mice were hyperinsulinaemic and hyperleptinaemic, yet plasma ghrelin, and stomach ghrelin peptide and mRNA were significantly reduced. In obese A(y)/a mice, PeVN somatostatin and Arc NPY mRNA expression were reduced by 50% compared to WT controls, whereas Arc GHRH mRNA expression was unchanged.Increased body length in adult obese A(y)/a mice may result from reduced Arc NPY and PeVN somatostatin mRNA expression, which in turn, may increase plasma IGFI concentrations and upregulate the somatotrophic axis.

Patients with gastric or esophageal surgery and transection of the vagus nerve may suffer from appetite and weight loss but without dysphagia or mechanical obstruction to eating. The gastric hormone ghrelin stimulates food intake and GH release in rodents and man. However, rodents with vagotomy are not sensitive to the feeding effects of ghrelin.The objective of the study was to determine whether humans with vagotomy are sensitive to ghrelin.The design was a double-blind, randomized, placebo-controlled trial.This was a hospital-based study.Six men and one woman who all had a previous complete truncal vagotomy with lower esophageal or gastric surgery entered and completed the study.Each patient received 120-min infusions of saline, 1 pmol/kg.min ghrelin, and 5 pmol/kg.min ghrelin on 3 separate days. After 90 min, a buffet meal was served.Energy intake at the buffet meal was measured.Ghrelin-stimulated GH release in a dose-dependent manner was measured, confirming bioactivity. However, no change in energy intake was observed with either dose of ghrelin [energy intake (kilojoules): saline 2805 +/- 812; ghrelin 1 pmol/kg.min, 2486 +/- 767; ghrelin 5 pmol/kg.min, 2382 +/- 543; P = not significant].Ghrelin is unlikely to be an effective appetite-stimulatory treatment for patients with vagotomy and esophageal or gastric surgery. Our results suggest that an intact vagus nerve may be required for exogenous ghrelin to increase appetite and food intake in man.

To investigate whether effects on food intake are seen in obese subjects receiving exogenous administration of ghrelin.Randomised, double-blind, placebo-controlled study of intravenous ghrelin at doses 1 pmol/kg/min and 5 pmol/kg/min.In all, 12 healthy lean subjects (mean body mass index (BMI) 20.5+/-0.17 kg/m(2)) and 12 healthy overweight and obese subjects (mean BMI 31.9+/-1.02 kg/m(2)).Food intake, appetite and palatability of food, ghrelin and other obesity-related hormones, growth hormone.Low-dose infusion of ghrelin increased ad libitum energy intake at a buffet meal in the obese group only (mean increase 36.6+/-9.4%, P0.01.) High-dose ghrelin infusion increased energy intake in both groups (mean increase 20.1+/-10.6% in the lean and 70.1+/-15.5% in the obese, P0.01 in both cases.) Ghrelin infusion increased palatability of food in the obese group.Ghrelin increases food intake in obese as well as lean subjects. Obese people are sensitive to the appetite-stimulating effects of ghrelin and inhibition of circulating ghrelin may be a useful therapeutic target in the treatment of obesity.

Neuropeptide Y (NPY) is the most potent stimulant of feeding when administered by intracerebroventricular injection. Despite this, there is conflicting evidence as to its importance in the regulation of daily food intake and energy balance. It has been suggested that whilst it is important in the response to starvation it has little role in the regulation of daily food intake. To investigate the role of NPY in the regulation of food intake, anti-sense cRNA to NPY was expressed in the arcuate nucleus of adult male rats. The anti-sense NPY (AS-NPY) construct was initially tested in vitro and there was a decrease of approximately 50% in NPY release from anti-sense treated cells compared to controls (16.3 +/- 2.0 fmol/L [AS-NPY] vs 37.3 +/- 7.7 fmol/L [control], mean +/- SEM p < 0.05). NPY release from hypothalamic explants from anti-sense injected animals was decreased by over 50% compared to those from controls at both 15 and 20 days after AAV injection (15 days 42% +/- 6.5% [AS-NPY] vs 100% +/- 36% [control], 20 days 41% +/- 6% [AS-NPY] vs 100% +/- 27% [control] mean+/-SEM, p < 0.05). In a study lasting for 50 days, weight gain was significantly lower in anti-sense injected animals from day 16 (day 16: 6.25 +/- 1.10 g [AS-NPY] vs 9.42 +/- 0.65 g [control] mean +/- SEM, p < 0.05) and remained so until the end of the study when they had gained approximately 40% less weight than controls (day 50: 52.0 +/- 9.6 g [AS-NPY] vs 82.0 +/- 6.3 g [control] mean +/- SEM, p < 0.01). Cumulative food intake was significantly lower in the anti-sense injected animals from day 23 (day 23: 225.8 +/- 1.9 g [AS-NPY] vs 250.6 +/- 8.7 g [control], mean +/- SEM, p < 0.05) and remained so until the end of the study (day 50: 834.5 +/- 14.8 g [AS-NPY] vs 926.0 +/- 31.7 g [control], mean +/- SEM, p < 0.05). Similarly mean daily food intake was also reduced in the anti-sense injected animals (days 7-14: 24.9 +/- 0.4 g/day [AS-NPY] vs 27.2 +/- 0.4 g/day [control], mean +/- SEM, p < 0.01). These data are supportive of a role for NPY in the regulation of daily food intake as well as in response to starvation.

Kisspeptin is the peptide product of the KiSS-1 gene and the endogenous agonist for the GPR54 receptor. Recent evidence suggests the kisspeptin/GPR54 system is a key regulator of the reproductive system. We examined the effect of intracerebroventricular (i.c.v.) and peripheral administration of the active kisspeptin fragment, kisspeptin-10, on circulating gonadotrophins and total testosterone levels in adult male rats. The effect of kisspeptin-10 in vitro on the release of hypothalamic peptides from hypothalamic explants and gonadotrophins from anterior pituitary fragments was also determined. The i.c.v. administration of kisspeptin-10 dose-dependently increased plasma luteinizing hormone (LH) and increased plasma follicle stimulating hormone (FSH) and total testosterone at 60 min postinjection. In a separate study investigating the time course of this response, i.c.v. administered kisspeptin-10 (3 nmol) significantly increased plasma LH at 10, 20 and 60 min, FSH at 60 min and total testosterone at 20 and 60 min postinjection. Kisspeptin-10 stimulated the release of luteinizing hormone-releasing hormone (LHRH) from in vitro hypothalamic explants. Peripheral administration of kisspeptin-10 increased plasma LH, FSH and total testosterone. However, doses of 100-1000 nM kisspeptin-10 did not influence LH or FSH release from pituitary fragments in vitro. Kisspeptin therefore potently stimulates the hypothalamic-pituitary-gonadal axis. These effects are likely to be mediated via the hypothalamic LHRH system.

Signals generated by the gastrointestinal tract are able to regulate appetite and influence body weight. Ghrelin is an orexigenic peptide produced by the stomach. Satiety signals derived from the intestine and pancreas include peptide YY, pancreatic polypeptide, glucagon-like peptide 1, oxyntomodulin, and cholecystokinin. Signals from the gut and adipose tissue are integrated in the central nervous system to provide energy homeostasis. Knowledge of the body's control of appetite is important because we strive to combat obesity in man.

Plasma osteocalcin, a marker of osteoblastic activity, is reduced in starvation, malnutrition, and anorexia nervosa, resulting in low bone turnover osteoporosis. Contradictory findings about the role of leptin as a link between nutritional status and bone physiology have been reported. We demonstrate that leptin-deficient ob/ob and leptin-resistant db/db male mice have increased plasma osteocalcin, and that in male ob/ob mice osteocalcin is not decreased by starvation, unlike control mice. Intraperitoneal leptin administration increased plasma osteocalcin in male ob/ob mice, and prevented its fall during 24 h fasting and 5 days of food restriction in normal male mice. This effect may be mediated via actions on the hypothalamic–pituitary–testicular or –growth hormone axes, or a direct action on osteoblasts. These studies support the hypothesis that the fall in leptin during starvation and weight loss is responsible for the associated reduction in osteoblast activity, and suggest a role for leptin in regulating bone turnover.

The adipocyte hormone, leptin, acts via the central nervous system to modulate glucose metabolism by skeletal muscle, but the direct effects of leptin on glucose metabolism by skeletal muscle are unclear. In this study, we have examined effects of leptin on glucose uptake by cultured L6 muscle cells assessed with the non-metabolised glucose analogue 2-deoxy-D-glucose. Reverse-transcriptase polymerase chain reaction (RT-PCR) analysis of RNA showed that L6 muscle cells express a short isoform of the leptin receptor (ObRa), but not the long isoform (ObRb). In the absence of added insulin, incubation of L6 muscle cells with murine leptin (10( -11)-10( -8) M) for 10 min and 1 h increased glucose uptake by 15 % - 23 %. This effect of leptin was lost by 4 h. Leptin (10( -10) - 10( -9) M) initially (after 10 min) suppressed insulin-stimulated glucose uptake by 14 - 16 %, but had no effect in the longer term. Leptin-stimulated glucose uptake was inhibited by the phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin, but not by the janus kinase-2 (JAK-2) inhibitor tyrphostin AG490. The results suggest that leptin can act directly on L6 muscle cellsvia a short leptin receptor isoform to acutely stimulate basal (but not insulin-stimulated) glucose uptake via a PI3K-dependent pathway.