Effects of ghrelin administration on decreased growth hormone status in obese animals

Obesity is characterized by markedly decreased ghrelin and growth hormone (GH) secretion. Ghrelin is a GH-stimulating, stomach-derived peptide that also has orexigenic action. Ghrelin supplement may restore decreased GH secretion in obesity, but it may worsen obesity by its orexigenic action. To reveal effects of ghrelin administration on obese animals, we first examined acute GH and orexigenic responses to ghrelin in three different obese and/or diabetic mouse models: db/db mice, mice on a high-fat diet (HFD mice), and Akita mice for comparison. GH responses to ghrelin were significantly suppressed in db/db, HFD, and Akita mice. Food intake of db/db and Akita mice were basally higher, and further stimulation of food intake by ghrelin was suppressed. Pituitary GH secretagogue receptor mRNA levels in db/db and HFD mice were significantly decreased, which may partly contribute to decreased GH response to ghrelin in these mice. In Akita mice for comparison, decreased hypothalamic GH-releasing hormone (GHRH) mRNA levels may be responsible for decreased GH response, since maximum GH response to ghrelin needs GHRH. When ghrelin was injected into HFD mice with GHRH coadministrated, GH responses to ghrelin were significantly emphasized. HFD mice injected with low-dose ghrelin and GHRH for 10 days did not show weight gain. These results indicate that low-dose ghrelin and GHRH treatment may restore decreased GH secretion in obesity without worsening obesity.     

Lecithinized Cu, Zn-superoxide dismutase limits the infarct size following ischemia-reperfusion injury in rat hearts in vivo

Ghrelin is a novel gut-brain peptide that binds to the growth hormone secretagogue receptor (GHS-R), thereby functioning in the regulation of growth hormone (GH) release and food intake. Ghrelin-producing cells are most abundant in the oxyntic glands of the stomach. The regulatory mechanism that governs the biosynthesis and secretion of ghrelin has not been clarified. We report that ghrelin mRNA expression in the gastric fundus was increased, but that ghrelin peptide content decreased after a 48-h fast. Both values returned to control levels after refeeding. The ghrelin plasma concentration in the gastric vein and systemic venous blood increased after 24- and 48-h fasts. Furthermore, des-octanoylated ghrelin and n-octanoylated ghrelin were found in rat stomach, with the ratio of des-octanoylated ghrelin to n-octanoylated ghrelin markedly increased after fasting. The ghrelin mRNA level in the stomach also increased after administration of insulin and leptin. Conversely, db/db mice, which are deficient in the leptin receptor, had lower ghrelin mRNA levels than control mice. These findings suggest that this novel gastrointestinal hormone plays a role in the regulation of energy balance.     

Ghrelin--not just another stomach hormone

Growth hormone (GH) secretagogues (GHSs) are non-natural, synthetic substances that stimulate GH secretion via a G-protein-coupled receptor called the GHS-receptor (GHS-R). The natural ligand for the GHS-R has been identified recently; it is called ghrelin. Ghrelin and its receptor show a widespread distribution in the body; the greatest expression of ghrelin is in stomach endocrine cells. Administration of exogenous ghrelin has been shown to stimulate pituitary GH secretion, appetite, body growth and fat deposition. Ghrelin was probably designed to be a major anabolic hormone. Ghrelin also exerts several other activities in the stomach. The findings that ghrelin is produced in mucosal endocrine cells of the stomach and intestine, and that ghrelin is measurable in the general circulation indicate its hormonal nature. A maximal expression of ghrelin in the stomach suggests that there is a gastrointestinal hypothalamic-pituitary axis that influences GH secretion, body growth and appetite that is responsive to nutritional and caloric intakes.     

Role of the AMP-activated protein kinase (AMPK) signaling pathway in the orexigenic effects of endogenous ghrelin

Ghrelin, released from the stomach, stimulates food intake through activation of the ghrelin receptor (GHS-R) located on neuropeptide Y (NPY)/agouti-related peptide (AgRP) neurons in the hypothalamus. A role for the energy sensor AMP-activated protein kinase (AMPK) and its downstream effector uncoupling protein 2 (UCP2) in the stimulatory effect of exogenous ghrelin on NPY/AgRP expression and food intake has been suggested. This study aimed to investigate whether a rise in endogenous ghrelin levels is able to influence hypothalamic AMPK activity, pACC, UCP2 and NPY/AgRP expression through activation of GHS-R. An increase in endogenous ghrelin levels was established by fasting (24h) or by induction of streptozotocin(STZ)-diabetes (15 days) in GHS-R(+/+) and GHS-R(-/-) mice. GHS-R(+/+) mice showed a significant increase in AgRP and NPY mRNA expression after fasting, which was not observed in GHS-R(-/-) mice. Fasting did not affect AMPK activity nor ACC phosphorylation in both genotypes and increased UCP2 mRNA expression. The hyperghrelinemia associated with STZ-induced diabetes was accompanied by an increased NPY and AgRP expression in GHS-R(+/+) but not in GHS-R(-/-) mice. AMPK activity and UCP2 expression in GHS-R(+/+) mice after induction of diabetes were decreased to a similar extent in both genotypes. Exogenous ghrelin administration tended to decrease hypothalamic AMPK activity. In conclusion, an increase in endogenous ghrelin levels triggered by fasting or STZ-induced diabetes stimulates the expression of AgRP and NPY via interaction with the GHS-R. The changes in AMPK activity, pACC and UCP2 occur independently from GHS-R suggesting that they do not play a major role in the orexigenic effect of endogenous ghrelin.     

Therapeutic applications of ghrelin to cachexia utilizing its appetite-stimulating effect

Ghrelin, which is a natural ligand for the growth hormone (GH)-secretagogue receptor (GHS-R), stimulates food intake in both animals and humans. Ghrelin is the only circulating hormone known to stimulate appetite in humans. Ghrelin also stimulates GH secretion and inhibits the production of anorectic proinflammatory cytokines. As GH is an anabolic hormone, protein stores are spared at the expense of fat during conditions of caloric restriction. Thus, ghrelin exhibits anti-cachectic actions via both GH-dependent and -independent mechanisms. Several studies are evaluating the efficacy of ghrelin in the treatment of cachexia caused by a variety of diseases, including congestive heart failure, chronic obstructive pulmonary disease, cancer, and end-stage renal disease. These studies will hopefully lead to the development of novel therapeutic applications for ghrelin in the future. This review summarizes the recent advances in this area of research.     

Contractile effects of ghrelin-related peptides on the chicken gastrointestinal tract in vitro

Ghrelin is an endogenous ligand for growth hormone secretagogue receptor (GHS-R), and it stimulates growth hormone (GH) release, food intake and gastrointestinal motility in mammals. Ghrelin has also been identified in the chicken, but this peptide inhibits food intake in the chicken. We examined the effects of ghrelin and related peptides on contractility of the isolated chicken gastrointestinal tract in vitro. Among ghrelin-related peptides examined (1 microM of rat ghrelin, human ghrelin, chicken ghrelin and growth hormone releasing peptide-6 (GHRP-6)), only chicken ghrelin was effective on contraction of the chicken gastrointestinal tract. Des-acyl chicken ghrelin was ineffective, suggesting that octanoylation at Ser3 residue of chicken ghrelin was essential for inducing the contraction. Amplitude of chicken ghrelin-induced contraction was region-specific: highest in the crop and colon, moderate in the esophagus and proventriculus, and weak in the small intestine. The contractile response to chicken ghrelin in the crop was not affected by tetrodotoxin (TTX), but that in the proventriculus was decreased by TTX and atropine to the same extents. D-Lys3-GHRP-6 (a GHS-R antagonist) caused a transient contraction and inhibited the effect of chicken ghrelin without affecting the high-K+-induced contraction. Chicken ghrelin potentiated electrical field stimulation-induced cholinergic contraction without affecting the responsiveness to bath-applied carbachol in the proventriculus. The location of GHS-R differs in the crop (smooth muscle) and proventriculus (smooth muscle and enteric neurons). These results indicate that ghrelin has contractile activity on gastrointestinal tract in the chicken in vitro, and the effect was region-specific. The action would be mediated through the GHS-R, which is highly sensitive to chicken ghrelin.     

Effects of leptin replacement on hypothalamic-pituitary growth hormone axis function and circulating ghrelin levels in ob/ob mice

Leptin-deficient obese mice (ob/ob) have decreased circulating growth hormone (GH) and pituitary GH and ghrelin receptor (GHS-R) mRNA levels, whereas hypothalamic GH-releasing hormone (GHRH) and somatostatin (SST) expression do not differ from lean controls. Given the fact that GH is suppressed in diet-induced obesity (a state of hyperleptinemia), it remains to be determined whether the absence of leptin contributes to changes in the GH axis of ob/ob mice. Therefore, to study the impact of leptin replacement on the hypothalamic-pituitary GH axis of ob/ob mice, leptin was infused for 7 days (sc), resulting in circulating leptin levels that were similar to wild-type controls (approximately 1 ng/ml). Leptin treatment reduced food intake, body weight, and circulating insulin while elevating circulating n-octanoyl ghrelin concentrations. Leptin treatment did not alter hypothalamic GHRH, SST, or GHS-R mRNA levels compared with vehicle-treated controls. However, leptin significantly increased pituitary GH and GHRH-R expression and tended to enhance circulating GH levels, but this latter effect did not reach statistical significance. In vitro, leptin (1 ng/ml, 24 h) did not affect pituitary GH, GHRH-R, or GHS-R mRNA but did enhance GH release. The in vivo effects of leptin on circulating hormone and pituitary mRNA levels were not replicated by pair feeding ob/ob mice to match the food intake of leptin-treated mice. However, leptin did prevent the fall in hypothalamic GHRH mRNA and circulating IGF-I levels observed in pair-fed mice. These results demonstrate that leptin replacement has positive effects on multiple levels of GH axis function in ob/ob mice.     

Ghrelin secretion is not reduced by increased fat mass during diet-induced obesity

Ghrelin is a stomach hormone that stimulates growth hormone (GH) secretion, adiposity, and food intake. Gastric ghrelin production and secretion are regulated by caloric intake; ghrelin secretion increases during fasting, decreases with refeeding, and is reduced by diet-induced obesity. The aim of the present study was to test the hypotheses that 1) an increase in body adiposity will play an inhibitory role in the reduction of gastric ghrelin synthesis and secretion during chronic ingestion of a high-fat (HF) diet and 2) chronic ingestion of an HF diet will suppress the rise in circulating ghrelin levels in response to acute fasting. Adult male Sprague-Dawley rats were fed a standard AIN-76A (approximately 5-12% of calories from fat) or an HF (approximately 45% of calories from fat) diet. The effect of increased adiposity on gastric ghrelin homeostasis was assessed by comparison of stomach ghrelin production and plasma ghrelin levels in obese and nonobese rats fed the HF diet. HF diet-fed, nonobese rats were generated by administration of triiodothyronine to lower body fat accumulation. Our findings indicate that an increased fat mass per se does not exert an inhibitory effect on ghrelin homeostasis during ingestion of the HF diet. Additionally, the magnitude of change in plasma ghrelin in response to fasting was not blunted, indicating that a presumed, endogenous signal for activation of ingestive behavior remains intact, despite excess stored calories in HF-fed rats.     

Importance of melanin-concentrating hormone receptor for the acute effects of ghrelin

The hypothalamic peptide melanin-concentrating hormone (MCH) and the gastric hormone ghrelin take part in the regulation of energy homeostasis and stimulate food intake. In the present study, ghrelin was administered centrally to MCH-receptor knockout (MCHr KO) mice. MCHr KO mice and wild type (WT) controls both consumed more food when treated with ghrelin. After ghrelin administration, the serum levels of insulin increased only in WT mice whereas the serum levels of corticosterone increased both in WT and MCHr KO mice. The level of growth hormone (GH) mRNA in the pituitary gland was markedly increased in response to ghrelin injection in the WT mice but was unaffected in the MCHr KO mice. The different ghrelin responses could not be explained by a difference in growth hormone secretagogue receptor expression between MCHr KO and WT mice in the pituitary or hypothalamus. In summary, the MCHr is not required for ghrelin induced feeding. However, the MCHr does play a role for the effect of ghrelin on GH expression in the pituitary and serum insulin levels.     

Ghrelin in growth and development

Exogenous administration of ghrelin increases caloric intake and stimulates growth hormone (GH) secretion, two effects that are mediated through binding of ghrelin to the GH secretagogue receptor (GHS-R). In addition, ghrelin is thought to inhibit adipogenesis by GHS-R-independent mechanisms. In adults, ghrelin is mainly produced by the stomach. In contrast, in the fetal and early postnatal period, ghrelin gene expression is abundant in the pancreas but not in the stomach. While knockout animal studies demonstrate that ghrelin is not required for perinatal development under normal nutritional conditions, the characteristics of ghrelin metabolism during fetal development suggest that ghrelin could contribute to the programming of mechanisms involved in energy balance, such as beta-cell maturation, orexigenic pathways and adipogenesis. In humans, ghrelin concentrations progressively decrease during childhood and adolescence, as well as with advancing puberty. In adolescents, similar to adults, ghrelin concentrations are inversely related to body mass index and to circulating insulin. One notable exception is the presence of elevated ghrelin concentrations in subjects with Prader-Willi syndrome, raising the possibility that ghrelin could be part of the etiology of excess food intake in this condition. These data raise a number of fascinating questions on the potential physiologic role of this hormone during growth and development.     

Growth hormone secretagogues and ghrelin: an update on physiology and clinical relevance

The pulsatile release of growth hormone (GH) by the anterior pituitary is stimulated by small synthetic molecules termed GH secretagogues (GHS). The receptor for GHS (GHS-R) belongs to the family of G-protein-coupled receptors. An endogenous specific ligand of 28 amino acids has recently been purified from rat stomach, it has been termed 'ghrelin'. Ghrelin demonstrates potent and reproducible GH-releasing activity, as well as significant prolactin-, ACTH- and cortisol-releasing activity. However, its major physiological relevance may relate to energy homeostasis. Peripheral daily administration of ghrelin caused weight gain by reducing fat utilization in mice and rats. In man, intravenous ghrelin was shown to stimulate food intake. The pathophysiological role and the potential clinical use of ghrelin are reviewed.     

Ghrelin stimulates gastric emptying but is without effect on acid secretion and gastric endocrine cells

Ghrelin, a recently discovered peptide hormone, is produced by endocrine cells in the stomach, the so-called A-like cells. Ghrelin binds to the growth hormone (GH) secretagogue receptor and releases GH. It is claimed to be orexigenic and to control gastric acid secretion and gastric motility. In this study, we examined the effects of ghrelin, des-Gln¹⁴-ghrelin, des-octanoyl ghrelin, ghrelin-18, -10 and -5 (and motilin) on gastric emptying in mice and on gastric acid secretion in chronic fistula rats and pylorus-ligated rats. We also examined whether ghrelin affected the activity of the predominant gastric endocrine cell populations, G cells, ECL cells and D cells. Ghrelin and des-Gln¹⁴-ghrelin stimulated gastric emptying in a dose-dependent manner while des-octanoyl ghrelin and motilin were without effect. The C-terminally truncated ghrelin fragments were effective but much less potent than ghrelin itself. Ghrelin, des-Gln¹⁴-ghrelin and des-octanoyl ghrelin neither stimulated nor inhibited gastric acid secretion, and ghrelin, finally, did not affect secretion from either G cells, ECL cells or D cells.     

Ghrelin: a metabolic signal affecting the reproductive system

Ghrelin, an acylated 28 amino acid gastric peptide, was isolated from the stomach as an endogenous ligand for growth hormone (GH) secretagogue receptor in 1999. Circulating ghrelin is mainly produced by specific cells in the stomach's oxyntic glands. Ghrelin potently stimulates GH release and food intake and exhibits diverse effects, including ones on glucose metabolism and on secretion and motility of the gastrointestinal tract. Besides these effects on food intake and energy homeostasis, ghrelin is also involved in controlling reproductive functions, and a role for it as a novel regulator of the hypothalamic-pituitary gonadal axis is clearly emerging.We review recent ghrelin research with emphasis on its roles in the reproductive axis.     

A Natural Variant of Obestatin,Q90L,Inhibits Ghrelin's Action on Food Intake and GH Secretion and Targets NPY and GHRH Neurons in Mice

Background

Ghrelin and obestatin are two gut-derived peptides originating from the same ghrelin/obestatin prepropeptide gene (GHRL). While ghrelin stimulates growth hormone (GH) secretion and food intake and inhibits γ-aminobutyric-acid synaptic transmission onto GHRH (Growth Hormone Releasing Hormone) neurons, obestatin blocks these effects. In Humans, GHRL gene polymorphisms have been associated with pathologies linked to an unbalanced energy homeostasis. We hypothesized that one polymorphism located in the obestatin sequence (Q to L substitution in position 90 of the ghrelin/obestatin prepropeptide, rs4684677) may impact on the function of obestatin. In the present study, we tested the activity of native and Q90L obestatin to modulate ghrelin-induced food intake, GH secretion, cFos activity in GHRH and Neuropeptide Y (NPY) neurons and γ-aminobutyric-acid activity onto GHRH neurons.

Methodology/Principal findings

Food intake, GH secretion and electrophysiological recordings were assessed in C57BL/6 mice. cFos activity was measured in NPY-Renilla-GFP and GHRH-eGFP mice. Mice received saline, ghrelin or ghrelin combined to native or Q90L obestatin (30 nmol each) in the early light phase. Ghrelin stimulation of food intake and GH secretion varied considerably among individual mice with 59–77% eliciting a robust response. In these high-responders, ghrelin-induced food intake and GH secretion were reduced equally by native and Q90L obestatin. In contrast to in vivo observations, Q90L was slightly more efficient than native obestatin in inhibiting ghrelin-induced cFos activation within the hypothalamic arcuate nucleus and the nucleus tractus solitarius of the brainstem. After ghrelin injection, 26% of NPY neurons in the arcuate nucleus expressed cFos protein and this number was significantly reduced by co-administration of Q90L obestatin. Q90L was also more potent that native obestatin in reducing ghrelin-induced inhibition of γ-aminobutyric-acid synaptic transmission onto GHRH neurons.

Conclusions/Significance

These data support the hypothesis that Q90L obestatin partially blocks ghrelin-induced food intake and GH secretion by acting through NPY and GHRH neurons.     

Ghrelin,a novel growth hormone-releasing peptide,in the treatment of chronic heart failure

Ghrelin is a novel growth hormone (GH)-releasing peptide, isolated from the stomach, which has been identified as an endogenous ligand for growth-hormone secretagogues receptor (GHS-R). This peptide also causes a positive energy balance by stimulating food intake and inducing adiposity through growth hormone-independent mechanisms. In addition, ghrelin has some cardiovascular effects, as indicated by the presence of its receptor in blood vessels and the cardiac ventricles. In vitro, ghrelin inhibits apoptosis of cardiomyocytes and endothelial cells. In humans, infusion of ghrelin decreases systemic vascular resistance and increases cardiac output in patients with heart failure. Repeated administration of ghrelin improves cardiac structure and function and attenuates the development of cardiac cachexia in rats with heart failure. These results suggest that ghrelin has cardiovascular effects and regulates energy metabolism through GH-dependent and -independent mechanisms. Thus, administration of ghrelin may be a new therapeutic strategy for the treatment of severe chronic heart failure (CHF).     

Evidence that growth hormone exerts a feedback effect on stomach ghrelin production and secretion

Ghrelin is a recently discovered stomach hormone that stimulates pituitary growth hormone (GH) secretion potently. The purpose of these experiments was to test the hypothesis that a stomach-ghrelin-pituitary-GH axis exists in which either an elevation or reduction in systemic GH levels will exert a negative or positive feedback action, respectively, on stomach ghrelin homeostasis. In rats, GH administration decreased stomach ghrelin mRNA levels and plasma ghrelin levels significantly. In GH-releasing hormone (GHRH) transgenic mice, GHRH overexpression decreased stomach ghrelin peptide levels when compared with control mice. In aged rats (25 months) stomach ghrelin mRNA and peptide levels and plasma ghrelin levels were decreased when compared with young rats (5 months). Because GH secretion is reduced in aged rats, the elevated stomach ghrelin production and secretion may reflect a decreased GH feedback on stomach ghrelin, homeostasis, and secretion. Together, these findings suggest that endogenous pituitary GH exerts a feedback action on stomach ghrelin homeostasis and support the hypothesis that a stomach-ghrelin-pituitary GH axis exists.