Growth Hormone Secretagogue Research Articles

The Links of Ghrelin to Incretins, Insulin, Glucagon, and Leptin After Bariatric Surgery.

Type 2 diabetes mellitus (T2DM) is one of the most prominent and socially significant problems. The present study aimed to identify the mechanisms of interaction of critical regulators of carbohydrate metabolism using bioinformatics and experimental methods and to assess their influence on the development of T2DM. We conducted an in silico search for the relationship of hormones and adipokines and performed functional annotation of the receptors for ghrelin and incretins. Hormones and adipokines were assessed in the plasma of obese patients with and without T2DM as well as after laparoscopic sleeve gastrectomy (LSG) and Roux-en-Y gastric bypass (RYGB) surgeries. Incretin- and ghrelin-associated functions and metabolic processes were discovered. Low ghrelin levels were observed in obese patients without T2DM compared with healthy volunteers and the other groups. The highest ghrelin levels were observed in obese patients with T2DM. This defense mechanism against insulin resistance could be realized through the receptors G-protein-coupled receptor (GPCR), growth hormone secretagogue receptor (GHSR), and growth hormone-releasing hormone receptor (GHRHR). These receptors are associated with proliferative, inflammatory, and neurohumoral signaling pathways and regulate responses to nutrient intake. Signaling through the GPCR class unites ghrelin, glucagon, glucose-dependent insulinotropic polypeptide (GIP), and glucagon-like peptide (GLP)-1. Ghrelin impairs carbohydrate and lipid metabolism in obese patients. Ghrelin is associated with elevated plasma levels of insulin, glucagon, and leptin. Specific activation of receptors and modulation by posttranslational modifications of ghrelin can control IR’s development in obesity, which is a promising area for research.

Contribution of growth hormone secretagogue receptor (GHSR) signaling in the ventral tegmental area (VTA) to the regulation of social motivation in male mice

Most psychiatric disorders are characterized by deficits in the ability to interact socially with others. Ghrelin, a hormone normally associated with the regulation of glucose utilization and appetite, is also implicated in the modulation of motivated behaviors including those associated with food and sex rewards. Here we hypothesized that deficits in ghrelin receptor (growth hormone secretagogue receptor; GHSR) signaling are also associated with deficits in social motivation in male mice. To test this hypothesis, we compared social motivation in male mice lacking GHSR or mice treated with the GHSR antagonist JMV2959 with that of WT or vehicle-treated mice. GHSR signaling in dopamine cells of the ventral tegmental area (VTA) has been implicated in the control of sexual behavior, thus we further hypothesized that GHSR signaling in the VTA is important for social motivation. Thus, we conducted studies where we delivered JMV2959 to block GHSR in the VTA of mice, and studies where we rescued the expression of GHSR in the VTA of GHSR knockout (KO) mice. Mice lacking GHSR or injected with JMV2959 peripherally for 3 consecutive days displayed lower social motivation as reflected by a longer latency to approach a novel conspecific and shorter interaction time compared to WT or vehicle-treated controls. Furthermore, intra-VTA infusion of JMV2959 resulted in longer latencies to approach a novel conspecific, whereas GHSR KO mice with partial rescue of the GHSR showed decreased latencies to begin a novel social interaction. Together, these data suggest that GHSR in the VTA facilitate social approach in male mice, and GHSR-signaling deficits within the VTA result in reduced motivation to interact socially.

Unusual orthologs shed new light on the binding mechanism of ghrelin to its receptor GHSR1a

The gastric peptide ghrelin has important functions in energy metabolism and cellular homeostasis by activating growth hormone secretagogue receptor type 1a (GHSR1a). The N-terminal residues of ghrelin orthologs from all vertebrates are quite conserved; however, in orthologs from Cavia porcellus and Phyllostomus discolor, Ser2 and Leu5 are replaced by a smaller Ala and a positively charged Arg, respectively. In the present study, we first demonstrated that the hydrophobic Leu5 is essential for the function of human ghrelin, because Ala replacement caused an approximately 100-fold decrease in activity. However, replacement of Leu5 by an Arg residue caused much less disruption; further replacement of Ser2 by Ala almost restored full activity, although the [S2A] mutation itself showed slight detriments, implying that the positively charged Arg5 in the [S2A,L5R] mutant might form alternative interactions with certain receptor residues to compensate for the loss of the essential Leu5. To identify the responsible receptor residues, we screened GHSR1a mutants in which all conserved negatively charged residues in the extracellular regions and all aromatic residues in the ligand-binding pocket were mutated separately. According to the decrease in selectivity of the mutant receptors towards [S2A,L5R]ghrelin, we deduced that the positively charged Arg5 of the ghrelin mutant primarily interacts with the essential aromatic Phe286 at the extracellular end of the sixth transmembrane domain of GHSR1a by forming cation−π and π−π interactions. The present study provided new insights into the binding mechanism of ghrelin with its receptor, and thus would facilitate the design of novel ligands for GHSR1a.

Potential Antioxidative, Anti-inflammatory and Immunomodulatory Effects of Ghrelin, an Endogenous Peptide from the Stomach in SARS-CoV2 Infection.

The current COVID-19 pandemic is one of the most devastating events in recent history. The respiratory effects of this disease include acute respiratory distress syndrome, systemic inflammation, cytokine storm, and pulmonary fibrosis. Ghrelin, an endogenous ligand for the growth hormone secretagogue receptor, is a peptide hormone secreted mainly by the stomach. Interestingly, ghrelin possesses promising antioxidant, anti-and inflammatory effects, making it an attractive agent to reduce the complications of the SARS-CoV-2. In addition, ghrelin exerts a wide range of immunomodulatory and anti-inflammatory effects and can mitigate the uncontrolled cytokine production responsible for acute lung injury by upregulating PPARγ and down-regulating NF-κB expression. Ghrelin has also been reported to enhance Nrf2 expression in inflammatory conditions which led to the suppression of oxidative stress. The current opinion summarizes the evidence for the possible pharmacological benefits of ghrelin in the therapeutic management of SARS-CoV-2 infection.

β Cell GHS-R Regulates Insulin Secretion and Sensitivity.

Growth hormone secretagogue receptor (GHS-R) is widely known to regulate food intake and adiposity, but its role in glucose homeostasis is unclear. In this study, we investigated the expression of GHS-R in mouse pancreatic islets and its role in glycemic regulation. We used Ghsr-IRES-tauGFP mice, with Green Fluorescent Protein (GFP) as a surrogate for GHS-R, to demonstrate the GFP co-localization with insulin and glucagon expression in pancreatic islets, confirming GHS-R expression in β and α cells. We then generated β-cell-specific GHSR-deleted mice with MIP-Cre/ERT and validated that GHS-R suppression was restricted to the pancreatic islets. MIP-Cre/ERT;Ghsrf/f mice showed normal energy homeostasis with similar body weight, body composition, and indirect calorimetry profile. Interestingly, MIP-Cre/ERT;Ghsrf/f mice exhibited an impressive phenotype in glucose homeostasis. Compared to controls, MIP-Cre/ERT;Ghsrf/f mice showed lower fasting blood glucose and insulin; reduced first-phase insulin secretion during a glucose tolerance test (GTT) and glucose-stimulated insulin secretion (GSIS) test in vivo. The isolated pancreatic islets of MIP-Cre/ERT;Ghsrf/f mice also showed reduced insulin secretion during GSIS ex vivo. Further, MIP-Cre/ERT;Ghsrf/f mice exhibited improved insulin sensitivity during insulin tolerance tests (ITT). Overall, our results confirmed GHS-R expression in pancreatic β and α cells; GHS-R cell-autonomously regulated GSIS and modulated systemic insulin sensitivity. In conclusion, β cell GHS-R was an important regulator of glucose homeostasis, and GHS-R antagonists may have therapeutic potential for Type 2 Diabetes.

Relationship between Orexigenic Peptide Ghrelin Signal, Gender Difference and Disease.

Growth hormone secretagogue receptor 1a (GHS-R1a), which is one of the G protein-coupled receptors (GPCRs), is involved in various physiological actions such as energy consumption, growth hormone secretion promoting action, and cardiovascular protective action. The ligand was searched for as an orphan receptor for a while, but the ligand was found to be acylated ghrelin (ghrelin) discovered by Kangawa and Kojima et al. in 1999. Recently, it has also been reported that dysregulation of GHS-R1a mediates reduced feeding in various diseases. On the other hand, since the physiological effects of ghrelin have been studied exclusively in male mice, few studies have been conducted on gender differences in ghrelin reactivity. In this review, we describe (1) the characteristics of GHS-R1a, (2) the role of ghrelin in hypophagia due to stress or anticancer drugs, and (3) the gender differences in the physiological effects of GHS-R1a and the influence of stress on it.

GHS-R in brown fat potentiates differential thermogenic responses under metabolic and thermal stresses.

In response to cold or diet, fatty acids are dissipated into heat through uncoupling protein 1 (UCP1) in brown adipose tissue (BAT). This process is termed non-shivering thermogenesis, which is important for body temperature maintenance and contributes to obesity pathogenesis. Thermogenic enhancement has been considered a promising anti-obesity strategy. Ghrelin and its receptor Growth Hormone Secretagogue Receptor (GHS-R) have critical roles in energy intake, nutrient sensing, and lipid metabolism. We previously reported that global Ghsr-knockout mice have increased energy expenditure due to enhanced thermogenesis. To determine the site of action for GHS-R mediated thermogenesis, we generated brown adipocyte-specific Ghsr knockout mice (UCP1-CreER/Ghsrf/f) and assessed thermogenic responses under regular diet (RD) fed homeostatic metabolic state or high-fat diet (HFD) fed metabolically-impaired obese state, under normal or cold housing environment. Under a RD-feeding, UCP1-CreER/Ghsrf/f mice showed increased body fat and a slightly elevated core body temperature under cold but not under normal temperature. Consistently, the expression of thermogenic genes in BAT of RD-fed UCP1-CreER/Ghsrf/f mice was increased in reposes to cold. Under HFD feeding, HFD-fed UCP1-CreER/Ghsrf/f mice showed no difference in body fat or body temperature under either normal or cold exposure. Interestingly, the expression of thermogenic genes in BAT of HFD-fed UCP1-CreER/Ghsrf/f mice was upregulated under normal temperature but downregulated under cold exposure. Overall, our data show that GHS-R has cell-autonomous effect in brown adipocytes, and GHS-R regulates BAT thermogenic activity in a temperature- and metabolic state-dependent manner. The thermogenic effect of GHS-R in BAT is more pronounced in cold environment and differentially variable based on metabolic state; under cold exposure, GHS-R inhibition in BAT activates thermogenesis under homeostatic state but suppresses thermogenesis under obese state. Our finding collectively suggests that GHS-R in BAT, acting as a “metabolic thermostat”, differentially regulates thermogenesis in response to different metabolic and thermal stimuli.

Genetic deletion of the ghrelin receptor (GHSR) impairs growth and blunts endocrine response to fasting in Ghsr-IRES-Cre mice

ObjectiveThe orexigenic hormone ghrelin exerts its physiological effects by binding to and activating the growth hormone secretagogue receptor (GHSR). The recent development of a Ghsr-IRES-Cre knock-in mouse line has enabled to genetically access GHSR-expressing neurons. Inserting a Cre construct using a knock-in strategy, even when following an upstream internal ribosome entry site (IRES) can, however, interfere with expression of a targeted gene, with consequences for the phenotype emerging. This study aimed to phenotype, both physically and metabolically, heterozygous and homozygous Ghsr-IRES-Cre mice, with a view to discovering the extent to which the ghrelin signalling system remains functional in these mice. MethodsWe assessed feeding and arcuate nucleus (Arc) Fos activation in wild-type, heterozygous and homozygous Ghsr-IRES-Cre mice in response to peripherally-administered ghrelin. We also characterised their developmental and growth phenotypes, as well as their metabolic responses upon an overnight fast. ResultsInsertion of the IRES-Cre cassette into the 3′-untranslated region of the Ghsr gene led to a gene-dosage GHSR depletion in the Arc. Whereas heterozygotes remained ghrelin-responsive and more closely resembled wild-types, ghrelin had reduced orexigenic efficacy and failed to induce Arc Fos expression in homozygous littermates. Homozygotes had a lower body weight accompanied by a shorter body length, less fat tissue content, altered bone parameters, and lower insulin-like growth factor-1 levels compared to wild-type and heterozygous littermates. Moreover, both heterozygous and homozygous Ghsr-IRES-Cre mice lacked the usual fasting-induced rise in growth hormone (GH) and displayed an exaggerated drop in blood glucose and insulin compared to wild-types. Unexpectedly, fasting acyl-ghrelin levels were allele-dependently increased. ConclusionsOur data suggest that (i) heterozygous but not homozygous Ghsr-IRES-Cre mice retain the usual responsiveness to administered ghrelin, (ii) the impact of fasting on GH release and glucose homeostasis is altered even when only one copy of the Ghsr gene is non-functional (as in heterozygous Ghsr-IRES-Cre mice) and (iii) homozygous Ghsr-IRES-Cre mice exhibit growth retardation. Of the many transgenic models of suppressed ghrelin signalling, Ghsr-IRES-Cre mice emerge as best representing the full breadth of the expected phenotype with respect to body weight, growth, and metabolic parameters.

Effect of Growth Hormone Secretagogue Receptor Deletion on Growth, Pulsatile Growth Hormone Secretion, and Meal Pattern in Male and Female Mice

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.

Ghrelin Receptors Enhance Fat Taste Responsiveness in Female Mice.

Ghrelin is a major appetite-stimulating neuropeptide found in circulation. While its role in increasing food intake is well known, its role in affecting taste perception, if any, remains unclear. In this study, we investigated the role of the growth hormone secretagogue receptor’s (GHS-R; a ghrelin receptor) activity in the peripheral taste system using feeding studies and conditioned taste aversion assays by comparing wild-type and GHS-R-knockout models. Using transgenic mice expressing enhanced green fluorescent protein (GFP), we demonstrated GHS-R expression in the taste system in relation phospholipase C ß2 isotype (PLCβ2; type II taste cell marker)- and glutamate decarboxylase type 67 (GAD67; type III taste cell marker)-expressing cells using immunohistochemistry. We observed high levels of co-localization between PLCβ2 and GHS-R within the taste system, while GHS-R rarely co-localized in GAD67-expressing cells. Additionally, following 6 weeks of 60% high-fat diet, female Ghsr−/− mice exhibited reduced responsiveness to linoleic acid (LA) compared to their wild-type (WT) counterparts, while no such differences were observed in male Ghsr−/− and WT mice. Overall, our results are consistent with the interpretation that ghrelin in the taste system is involved in the complex sensing and recognition of fat compounds. Ghrelin-GHS-R signaling may play a critical role in the recognition of fatty acids in female mice, and this differential regulation may contribute to their distinct ingestive behaviors.

The ups and downs of growth hormone secretagogue receptor signaling.

The growth hormone secretagogue receptor (GHSR) has emerged as one of the most fascinating molecules from the perspective of neuroendocrine control. GHSR is mainly expressed in the pituitary and the brain, and plays key roles regulating not only growth hormone secretion but also food intake, adiposity, body weight, glucose homeostasis and other complex functions. Quite atypically, GHSR signaling displays a basal constitutive activity that can be up- or downregulated by two digestive system-derived hormones: the octanoylated-peptide ghrelin and the liver-expressed antimicrobial peptide 2 (LEAP2), which was recently recognized as an endogenous GHSR ligand. The existence of two ligands with contrary actions indicates that GHSR activity can be tightly regulated and that the receptor displays the capability to integrate such opposing inputs in order to provide a balanced intracellular signal. This article provides a summary of the current understanding of the biology of ghrelin, LEAP2 and GHSR and discusses the reconceptualization of the cellular and physiological implications of the ligand-regulated GHSR signaling, based on the latest findings.

Development of a ghrelin receptor inverse agonist for positron emission tomography.

Imaging of Ghrelin receptors in vivo provides unique potential to gain deeper understanding on Ghrelin and its receptors in health and disease, in particular, in cancer. Ghrelin, an octanoylated 28-mer peptide hormone activates the constitutively active growth hormone secretagogue receptor type 1a (GHS-R1a) with nanomolar activity. We developed novel compounds, derived from the potent inverse agonist K-(D-1-Nal)-FwLL-NH2 but structurally varied by lysine conjugation with 1,4,7-triazacyclononane,1-glutaric acid-4,7-acetic acid (NODAGA), palmitic acid and/or diethylene glycol (PEG2) to allow radiolabeling and improve pharmacokinetics, respectively. All compounds were tested for receptor binding, potency and efficacy in vitro, for biodistribution and -kinetics in rats and in preclinical prostate cancer models on mice. Radiolabeling with Cu-64 and Ga-68 was successfully achieved. The Cu-64- or Ga-68-NODAGA-NH-K-K-(D-1-NaI)-F-w-L-L-NH2 radiotracer were specifically accumulated by the GHS-R1a in xenotransplanted human prostate tumor models (PC-3, DU-145) in mice. The tumors were clearly delineated by PET. The radiotracer uptake was also partially blocked by K-(D-1-Nal)-FwLL-NH2 in stomach and thyroid. The presence of the GHS-R1a was also confirmed by immunohistology. In the arterial rat blood plasma, only the original compounds were found. The Cu-64 or Ga-68-NODAGA-NH-K-K-(D-1-NaI)-F-w-L-L-NH2 radiolabeled inverse agonists turned out to be potent and safe. Due to their easy synthesis, high affinity, medium potency, metabolic stability, and the suitable pharmacokinetic profiles, they are excellent tools for imaging and quantitation of GHS-R1a expression in normal and cancer tissues by PET. These compounds can be used as novel biomarkers of the Ghrelin system in precision medicine.

Cannabinoid-Induced Conditioned Place Preference, Intravenous Self-Administration, and Behavioral Stimulation Influenced by Ghrelin Receptor Antagonism in Rats.

Cannabis/cannabinoids are widely used for recreational and therapy purposes, but their risks are largely disregarded. However, cannabinoid-associated use disorders and dependence are alarmingly increasing and an effective treatment is lacking. Recently, the growth hormone secretagogue receptor (GHSR1A) antagonism was proposed as a promising mechanism for drug addiction therapy. However, the role of GHS-R1A and its endogenous ligand ghrelin in cannabinoid abuse remains unclear. Therefore, the aim of our study was to investigate whether the GHS-R1A antagonist JMV2959 could reduce the tetrahydrocannabinol (THC)-induced conditioned place preference (CPP) and behavioral stimulation, the WIN55,212-2 intravenous self-administration (IVSA), and the tendency to relapse. Following an ongoing WIN55,212-2 self-administration, JMV2959 3 mg/kg was administered intraperitoneally 20 min before three consequent daily 120-min IVSA sessions under a fixed ratio FR1, which significantly reduced the number of the active lever-pressing, the number of infusions, and the cannabinoid intake. Pretreatment with JMV2959 suggested reduction of the WIN55,212-2-seeking/relapse-like behavior tested in rats on the twelfth day of the forced abstinence period. On the contrary, pretreatment with ghrelin significantly increased the cannabinoid IVSA as well as enhanced the relapse-like behavior. Co-administration of ghrelin with JMV2959 abolished/reduced the significant efficacy of the GHS-R1A antagonist in the cannabinoid IVSA. Pretreatment with JMV2959 significantly and dose-dependently reduced the manifestation of THC-induced CPP. The THC-CPP development was reduced after the simultaneous administration of JMV2959 with THC during conditioning. JMV2959 also significantly reduced the THC-induced behavioral stimulation in the LABORAS cage. Our findings suggest that GHS-R1A importantly participates in the rewarding/reinforcing effects of cannabinoids.

Development of a Predictive Enrichment Marker for the Oral GH Secretagogue LUM-201 in Pediatric Growth Hormone Deficiency.

ContextWe hypothesize, based on the degree of residual hypothalamic-pituitary function, that some, but not all, children with growth hormone deficiency (GHD) may have beneficial growth responses to the orally administered growth hormone (GH) secretagogue LUM-201.ObjectiveTo determine if pretreatment testing can identify predictive enrichment markers (PEM) for subjects with adequate residual function who are responsive to LUM-201.MethodsWe performed an analysis of a completed, randomized, placebo-controlled trial of LUM-201, a GH secretagogue receptor agonist, in which all randomized subjects had pretreatment testing. This international multicenter study conducted in pediatric endocrinology clinics included 68 naïve-to-treatment, prepubertal children with established diagnoses of GHD. Outcome measures included the sensitivity, specificity, and predictive accuracy of potential markers to predict 6-month growth responses to oral LUM-201 and daily rhGH.ResultsTwo PEM were identified for use in defining PEM-positive status: (1) baseline insulin-like growth factor I (IGF-I) concentration >30 ng/mL and (2) peak GH response of ≥5 ng/mL upon administration of single-dose LUM-201. PEM-positive status enriches a population for better growth responses to LUM-201. PEM-negative status enriches a population for better growth responses to rhGH.ConclusionCombined, the peak GH response to single-dose LUM-201 and the baseline IGF-I concentration are effective PEMs for 6-month growth responses to LUM-201 and rhGH in prepubertal children with GHD.

Corroboration of Height Velocity Prediction Markers for rhGH With an Oral GH Secretagogue Treatment in Children With GHD.

ContextRecombinant human growth hormone (rhGH) is approved for treatment of pediatric growth hormone deficiency (GHD), with greatest growth responses observed in those with severe GHD. Orally administered GH secretagogues (GHS) may be useful treatment in patients with moderate GHD. Distinguishing children with severe vs moderate GHD could identify children who would be better treated with rhGH or GHS.ObjectivesEvaluate baseline insulin-like growth factor-I (IGF-I) and stimulated peak GH response as predictors of 12-month height velocity (HV) in children with GHD.DesignData on children with GHD were analyzed in a legacy data base (GeNeSIS data).Participants514 naïve to rhGH-treatment, prepubertal children with idiopathic isolated GHD for whom stimulated GH, baseline serum IGF-I, and first-year HV during rhGH treatment data are available.Outcome MeasuresChildren with severe or moderate GHD were categorized based on GH and IGF-I data and evaluated based on baseline auxologic and hormone profiles and first-year growth response to rhGH.ResultsCohorts of severe and moderate GHD were 81/514 (15.8%) and 433/514 (84.2%). Cohorts differed significantly with regard to indicators of GHD [eg, baseline height SD score (SDS), height SDS minus target height SDS, HV, HV SDS, and change in height SDS during rhGH treatment]. Multiple regression analysis showed IGF-I and stimulated GH were significant predictors of HV independent of other known variables. Expected first-year HV in moderate GHD was 8.3 cm/y.ConclusionsThe combination of peak GH to GH stimulation testing and baseline IGF-I concentration are predictive enrichment markers for annualized HV responses to rhGH therapy.

The Ghrelin/Growth Hormone Secretagogue Receptor System Is Involved in the Rapid and Sustained Antidepressant-Like Effect of Paeoniflorin.

Major depressive disorder (MDD) is a debilitating mental illness affecting people worldwide. Although significant progress has been made in the development of therapeutic agents to treat this condition, fewer than half of all patients respond to currently available antidepressants, highlighting the urgent need for the development of new classes of antidepressant drugs. Here, we found that paeoniflorin (PF) produced rapid and sustained antidepressant-like effects in multiple mouse models of depression, including the forced swimming test and exposure to chronic mild stress (CMS). Moreover, PF decreased the bodyweight of mice without affecting food intake and glucose homeostasis, and also reduced the plasma levels of total ghrelin and the expression of ghrelin O-acyltransferase in the stomach; however, the plasma levels of ghrelin and the ghrelin/total ghrelin ratio were unaffected. Furthermore, PF significantly increased the expression of growth hormone secretagogue receptor 1 alpha (GHSR1α, encoded by the Ghsr gene) in the intestine, whereas the levels of GHSR1α in the brain were only marginally downregulated following subchronic PF treatment. Finally, the genetic deletion of Ghsr attenuated the antidepressant-like effects of PF in mice exposed to CMS. These results suggested that increased GHSR1α expression in the intestine mediates the antidepressant-like effects of PF. Understanding peripheral ghrelin/GHSR signaling may provide new insights for the screening of antidepressant drugs that produce fast-acting and sustained effects.

Gut peptides and the microbiome: focus on ghrelin.

Purpose of reviewIn this review, we present recent insights into the role of the gut microbiota on gastrointestinal (GI) peptide secretion and signalling, with a focus on the orexigenic hormone, ghrelin.Recent findingsEvidence is accumulating suggesting that secretion of GI peptides is modulated by commensal bacteria present in our GI tract. Recent data shows that the gut microbiome impacts on ghrelinergic signalling through its metabolites, at the level of the ghrelin receptor (growth hormone secretagogue receptor) and highlights concomitant changes in circulating ghrelin levels with specific gut microbiota changes. However, the mechanisms by which the gut microbiota interacts with gut peptide secretion and signalling, including ghrelin, are still largely unknown.SummaryThe gut microbiota may directly or indirectly influence secretion of the orexigenic hormone, ghrelin, similar to the modulation of satiety inducing GI hormones. Although data demonstrating a role of the microbiota on ghrelinergic signalling is starting to emerge, future mechanistic studies are needed to understand the full impact of the microbiota-ghrelin axis on metabolism and central-regulated homeostatic and non-homeostatic controls of food intake.

Functional and Neurochemical Identification of Ghrelin Receptor (GHSR)-Expressing Cells of the Lateral Parabrachial Nucleus in Mice.

The lateral parabrachial nucleus (lPBN), located in the pons, is a well-recognized anorexigenic center harboring, amongst others, the calcitonin gene-related peptide (CGRP)-expressing neurons that play a key role. The receptor for the orexigenic hormone ghrelin (the growth hormone secretagogue receptor, GHSR) is also abundantly expressed in the lPBN and ghrelin delivery to this site has recently been shown to increase food intake and alter food choice. Here we sought to explore whether GHSR-expressing cells in the lPBN (GHSRlPBN cells) contribute to feeding control, food choice and body weight gain in mice offered an obesogenic diet, involving studies in which GHSRlPBN cells were silenced. We also explored the neurochemical identity of GHSRlPBN cells. To silence GHSRlPBN cells, Ghsr-IRES-Cre male mice were bilaterally injected intra-lPBN with a Cre-dependent viral vector expressing tetanus toxin-light chain. Unlike control wild-type littermates that significantly increased in body weight on the obesogenic diet (i.e., high-fat high-sugar free choice diet comprising chow, lard and 9% sucrose solution), the heterozygous mice with silenced GHSRlPBN cells were resistant to diet-induced weight gain with significantly lower food intake and fat weight. The lean phenotype appeared to result from a decreased food intake compared to controls and caloric efficiency was unaltered. Additionally, silencing the GHSRlPBN cells altered food choice, significantly reducing palatable food consumption. RNAscope and immunohistochemical studies of the lPBN revealed considerable co-expression of GHSR with glutamate and pituitary adenylate cyclase-activating peptide (PACAP), and much less with neurotensin, substance P and CGRP. Thus, the GHSRlPBN cells are important for diet-induced weight gain and adiposity, as well as in the regulation of food intake and food choice. Most GHSRlPBN cells were found to be glutamatergic and the majority (76%) do not belong to the well-characterized anorexigenic CGRP cell population.

Detection of the synthetic peptide ipamorelin in dried blood spots by means of UHPLC-HRMS

Growth hormone-releasing peptides (GHRPs) are synthetic peptides that stimulate the release of growth hormones from the hypothalamus and pituitary gland. Among these, ipamorelin is a highly selective, potent and effective pentapeptide belonging to the class of growth hormone secretagogues (GHS). Because of clinical monitoring and the potential abuse by athletes in amateur and elite sports, detection methods able to identify GHRPs in biological samples are increasingly needed. As an alternative to conventional procedures, testing of Dried Blood Spots (DBS) is gaining increasing attention, since its low volume sampling is easy and scarcely invasive. In this study, an ultra high pressure liquid chromatography–high resolution mass spectrometry method was developed and validated for the screening of ipamorelin in DBS. Ipamorelin was extracted from DBS by liquid extraction and ultrasonic treatment, then detected by the acquisition of high-resolution fragmentation spectra. The method proved satisfactory in terms of sensitivity, specificity, linearity, recovery, precision and ion suppression/enhancement effects. The mass error of the precursor and fragment ions was lower than 5 ppm. Since ipamorelin was successfully detected in DBS with a detection limit of 2.5 ng/mL, the combination of DBS and high resolution mass spectrometry proved to be a suitable and effective method to detect small peptides in blood at low levels. This approach looks promising for targeted and untargeted analysis in clinical monitoring and sport drug testing, especially when unknown synthetic peptides represent the target analytes.

Natural ghrelin in advanced cancer patients with cachexia, a case series

BackgroundNatural ghrelin, a peptide growth hormone secretagogue, has a therapeutic potential in cachexia. We designed a dose‐finding trial of subcutaneous natural ghrelin to improve nutritional intake (NI) in advanced cancer patients.MethodsAdvanced cancer patients with cachexia management (symptom management, physiotherapy, nutritional, and psychosocial support) started with ghrelin at 32 μg/kg body weight, followed by 50% dose increases. Patients self‐injected ghrelin twice daily for 4 days followed by a wash‐out period. After reaching the primary endpoint, maximal NI (minimal dose for maximal NI), a maintenance period followed during which patients injected 10 doses of ghrelin per week. Safety parameters, NI, and cachexia outcomes (symptoms, narratives, muscle mass, and strength) were measured over 6 weeks.ResultsTen patients with metastatic solid tumours were included, and six (100% male, mean age 61.8 ± 8.5 SD) received ghrelin. Minimal dose for maximal NI was reached in four patients. Three patients reached the end‐of study visit. Ghrelin was well tolerated with variable results on appetite and eating‐related symptoms but a positive effect in the narratives. Mean Functional Assessment of Appetite & Cachexia Therapy score was 6.8 points lower at final measurement compared with baseline, t(5) = 5.98, P < .01. Muscle mass was stable in two patients and increased in one patient, and muscle strength was stable in three patients. Subjective tolerability was high. Patients showed a fluctuating trajectory, and median survival was 88 days (51–412 days).ConclusionsGhrelin was safe in advanced patients with cancer cachexia without dose‐limiting toxicity and well tolerated. The intervention was very complex, and the number of patients included was small. There was a positive effect on nutritional intake and patient narratives.