Synergistic action of gastrin and ghrelin on gastric acid secretion in rats

Gastrin and ghrelin are secreted from G cells and X/A-like cells in the stomach, respectively, and respective hormones stimulate gastric acid secretion by acting through histamine and the vagus nerve. In this study, we examined the relationship between gastrin, ghrelin and gastric acid secretion in rats. Intravenous (iv) administration of 3 and 10 nmol of gastrin induced transient increases of ghrelin levels within 10 min in a dose-dependent manner. Double immunostaining for ghrelin and gastrin receptor revealed that a proportion of ghrelin cells possess gastrin receptors. Although (iv) administration of gastrin or ghrelin induced significant gastric acid secretion, simultaneous treatment with both hormones resulted in a synergistic, rather than additive, increase of gastric acid secretion. This synergistic increase was not observed in vagotomized rats.These results suggest that gastrin may directly stimulate ghrelin release from the stomach, and that both hormones may increase gastric acid secretion synergistically.     

Effect of chronic stress on gastric emptying and plasma ghrelin levels in rats

Chronic stress is associated with gastrointestinal functional diseases. Although the pathophysiology seems to be associated with gastrointestinal motility, their mechanisms remain unclear. We investigated gastric emptying and chemical mediators under conditions of continuous stress, which were produced using 8-week-old male Wistar rats kept in a cage filled with water to 2 cm height for 5 days. We examined gastric emptying by the phenol red method and chemical mediators at 4, 8, and 24 h and 3 and 5 days after initiation of stress restraint. Plasma ACTH level was significantly higher in the stress throughout the period of measurement. Continuous stress delayed gastric emptying until 24 h: peak delay was observed at 8 h, whereas gastric emptying was accelerated on days 3 and 5. Plasma noradrenalin level was significantly elevated at every time point until 24 h. Guanethidine pretreatment eliminated the delay in gastric emptying at 8 h. Active ghrelin was significantly increased on days 3 and 5 after peak (at 24 h) plasma total and desacyl ghrelin in the stress group. Number of ghrelin-immunoreactive cells and level of preproghrelin mRNA expression in the gastric body increased in parallel with plasma active ghrelin level. Pretreatment with growth hormone secretagogue receptor antagonist at 5 days partially inhibited the stress-induced acceleration of gastric emptying. Delayed gastric emptying at acute phase of continuous stress was mediated via sympathetic pathway, while acceleration at chronic phase was mediated via increased active ghrelin release from the stomach.     

Intracerebroventricular injection of ghrelin decreases wheel running activity in rats

There is an increasing interest in elucidating the molecular mechanisms by which voluntary exercise is regulated. In this study, we examined how the central nervous system regulates exercise. We used SPORTS rats, which were established in our laboratory as a highly voluntary murine exercise model. SPORTS rats showed lower levels of serum ghrelin compared with those of the parental line of Wistar rats. Intracerebroventricular and intraperitoneal injection of ghrelin decreased wheel-running activity in SPORTS rats. In addition, daily injection of the ghrelin inhibitor JMV3002 into the lateral ventricles of Wistar rats increased wheel-running activity. Co-administration of obestatin inhibited ghrelin-induced increases in food intake but did not inhibit ghrelin-induced suppression of voluntary exercise in rats. Growth hormone secretagogue receptor (GHSR) in the hypothalamus and hippocampus of SPORTS rats was not difference that in control rats. We created an arcuate nucleus destruction model by administering monosodium glutamate (MSG) to neonatal SPORTS rats. Injection of ghrelin into MSG-treated rats decreased voluntary exercise but did not increase food intake, suggesting that wheel-running activity is not controlled by the arcuate nucleus neurons that regulate feeding. These results provide new insights into the mechanism by which ghrelin regulates voluntary activity independent of arcuate nucleus neurons.     

Physiological roles revealed by ghrelin and ghrelin receptor deficient mice

Ghrelin is a hormone made in the stomach and known primarily for its growth hormone releasing and orexigenic properties. Nevertheless, ghrelin through its receptor, the GHS-R1a, has been shown to exert many roles including regulation of glucose homeostasis, memory & learning, food addiction and neuroprotection. Furthermore, ghrelin could promote overall health and longevity by acting directly in the immune system and promoting an extended antigen repertoire. The development of mice lacking either ghrelin (ghrelin−/−) or its receptor (ghsr−/−) have provided a valuable tool for determining the relevance of ghrelin and its receptor in these multiple and diverse roles. In this review, we summarize the most important findings and lessons learned from the ghrelin−/− and ghsr−/− mice.     

Exercise in rats does not alter hypothalamic AMP-activated protein kinase activity

Recent studies have demonstrated that AMP-activated protein kinase (AMPK) in the hypothalamus is involved in the regulation of food intake. Because exercise is known to influence appetite and cause substrate depletion, it may also influence AMPK in the hypothalamus. Male rats that either rested or ran for 30 or 60 min on a treadmill (22 m/min, 10% slope) were sacrificed immediately after exercise or after 60 min recovery either in the fasted state or after oral gavage with glucose (3g/kg body weight). Exercise decreased muscle and liver glycogen substantially. Hypothalamic total or alpha2-associated AMPK activity and phosphorylation state of the AMPK substrate acetyl-CoA carboxylase were not changed significantly immediately following treadmill running or during fed or fasted recovery. Plasma ghrelin increased (P<0.05) by 40% during exercise whereas the concentration of PYY was unchanged. In recovery, glucose feeding increased plasma glucose and insulin concentrations whereas ghrelin and PYY decreased to (ghrelin) or below (PPY) resting levels. It is concluded that 1h of strenuous exercise in rats does not elicit significant changes in hypothalamic AMPK activity despite an increase in plasma ghrelin. Thus, changes in energy metabolism during or after exercise are likely not coordinated by changes in hypothalamic AMPK activity.     

Effect of chronic treatments with ghrelin on milk secretion in lactating rats

Ghrelin, a novel acylated peptide and endogenous ligand for growth hormone (GH) secretagogue receptor, was originally isolated from rat and human stomachs. In addition to its GH-releasing activity, ghrelin plays an important role in many physiological functions, including food intake, gastric acid secretion, neonatal development, and so on. In this study, the effect of daily treatment with ghrelin on milk production was investigated in lactating rats and the development of the pups was monitored. Daily subcutaneous injection of ghrelin into nursing dams for 8 days from parturition caused a significant increase in milk yield and litter weight gain. When litters nursed by ghrelin-treated and saline-treated dams were interchanged on day 4 of lactation, the growth curves were reversed. Daily injections of ghrelin also increased plasma GH levels. Northern blot analysis revealed that daily injection of ghrelin significantly increased mammary casein mRNA expression. In addition, RT-PCR analysis showed that a ghrelin receptor was present in the mammary glands of lactating rats. These results suggest that ghrelin may play an important role in milk production in lactating dams.     

Effect of peripherally administered ghrelin on gastric emptying and acid secretion in the rat

Ghrelin is a gut peptide that is secreted from the stomach and stimulates food intake. There are ghrelin receptors throughout the gut and intracerebroventricular ghrelin has been shown to increase gastric acid secretion. The aim of the present study was to examine the effects of peripherally administered ghrelin on gastric emptying of a non-nutrient and nutrient liquid, as well as, basal and pentagastrin-stimulated gastric acid secretion in awake rats. In addition, gastric contractility was studied in vitro. Rats equipped with a gastric fistula were subjected to an intravenous infusion of ghrelin (10-500 pmol kg(-1) min(-1)) during saline or pentagastrin (90 pmol kg(-1) min(-1)) infusion. After administration of polyethylene glycol (PEG) 4000 with 51Cr as radioactive marker, or a liquid nutrient with (51)Cr, gastric retention was measured after a 20-min infusion of ghrelin (500 pmol kg(-1) min(-1)). In vitro isometric contractions of segments of rat gastric fundus were studied (10(-9) to 10(-6) M). Ghrelin had no effect on basal acid secretion, but at 500 pmol kg(-1) min(-1) ghrelin significantly decreased pentagastrin-stimulated acid secretion. Ghrelin had no effect on gastric emptying of the nutrient liquid, but significantly increased gastric emptying of the non-nutrient liquid. Ghrelin contracted fundus muscle strips dose-dependently (pD2 of 6.93+/-0.7). Ghrelin IV decreased plasma orexin A concentrations and increased plasma somatostatin concentrations. Plasma gastrin concentrations were unchanged during ghrelin infusion. Thus, ghrelin seems to not only effect food intake but also gastric motor and secretory function indicating a multifunctional role for ghrelin in energy homeostasis.     

Effects of ghrelin on gastric distention sensitive neurons in the arcuate nucleus of hypothalamus and gastric motility in diabetic rats

This study was performed to observe the effects of ghrelin on the activity of gastric distention (GD) sensitive neurons in the arcuate nucleus of hypothalamus (Arc) and on gastric motility in vivo in streptozocin (STZ) induced diabetes mellitus (DM) rats. Electrophysiological results showed that ghrelin could excite GD-excitatory (GD-E) neurons and inhibit GD-inhibitory (GD-I) neurons in the Arc. However, fewer GD-E neurons were excited by ghrelin and the excitatory effect of ghrelin on GD-E neurons was much weaker in DM rats. Gastric motility research in vivo showed that microinjection of ghrelin into the Arc could significantly promote gastric motility and it showed a dose-dependent manner. The effect of ghrelin promoting gastric motility in DM rats was weaker than that in normal rats. The effects induced by ghrelin could be blocked by growth hormone secretagogue receptor (GHSR) antagonist [d-Lys-3]-GHRP-6 or BIM28163. RIA and real-time PCR data showed that the levels of ghrelin in the plasma, stomach and ghrelin mRNA in the Arc increased at first but decreased later and the expression of GHSR-1a mRNA in the Arc maintained a low level in DM rats. The present findings indicate that ghrelin could regulate the activity of GD sensitive neurons and gastric motility via ghrelin receptors in the Arc. The reduced effects of promoting gastric motility induced by ghrelin could be connected with the decreased expression of ghrelin receptors in the Arc in diabetes. Our data provide new experimental evidence for the role of ghrelin in gastric motility disorder in diabetes.     

鱼类ghrelin研究进展

1999年日本学者Kojima,et al．^[1]在大鼠胃组织中首次发现了一种多肽,当这种多肽与生长激素促分泌素受体（Growth hormone secretagogue receptor,GHS—R）结合后,能刺激生长激素（Growth hormone,GH）的分泌,因此将其命名为ghrelin（ghre即grow之意）。此后,人们相继发现ghrelin在摄食、能量平衡、体重、心血管等方面显示出多种调节作用。目前有关人类和哺乳动物ghrelin的研究主要集中在药学研究和畜牧饲料开发两大方面,并取得了一定成效。国内外有关鱼类ghrelin的研究资料较少,相关报道仅见于少数的几种鱼类。本文从鱼类ghrelin的结构、组织分布及生理功能三个方面综述了鱼类ghrelin研究的进展情况;     

Sex-associated differences in the leptin and ghrelin systems related with the induction of hyperphagia under high-fat diet exposure in rats

Leptin and ghrelin are known to be main hormones involved in the control of food intake, with opposing effects. Here we have explored whether changes in the leptin and ghrelin system are involved in the long-term effects of high-fat (HF) diet feeding in rats and whether sex-associated differences exist. Male and female Wistar rats were fed until the age of 6 months with a normal-fat (NF) or an HF-diet. Food intake and body weight were followed. Gastric and serum levels of leptin and ghrelin, and mRNA levels of leptin (in stomach and adipose tissue), ghrelin (in stomach), and NPY, POMC, and leptin and ghrelin receptors (OB-Rb and GHS-R) (in the hypothalamus) were measured. In both males and females, total caloric intake and body weight were greater under the HF-diet feeding. In females, circulating ghrelin levels and leptin mRNA expression in the stomach were higher under HF-diet. HF-diet feeding also resulted in higher hypothalamic NPY/POMC mRNA levels, more marked in females, and in lower OB-Rb mRNA levels, more marked in males. In addition, in females, serum ghrelin levels correlated positively with hypothalamic NPY mRNA levels, and these with caloric intake. In males, hypothalamic OB-Rb mRNA levels correlated positively with POMC mRNA levels and these correlated negatively with caloric intake and with body weight. These data reflect differences between sexes in the effects of HF-diet feeding on food intake control systems, suggesting an impairment of the anorexigenic leptin-POMC system in males and an over-stimulation of the orexigenic ghrelin-NPY system in females.     

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.     

Effects of gastric emptying on the postprandial ghrelin response

Distension and chemosensitization of the stomach are insufficient to induce a ghrelin response, suggesting that postgastric feedback is required. This postgastric feedback may be regulated through insulin. We investigated the relation between gastric emptying rate and the postprandial ghrelin response as well as the role of insulin and other hormones possibly mediating this response. Fifteen healthy men [BMI 21.6 kg/m2 (SD 1.9), age 20.5 yr (SD 2.5)] were studied in a single-blind, crossover design. Subjects received two treatments separated by 1 wk: 1) a dairy breakfast in combination with a 3-h intravenous infusion of glucagon-like peptide-1 (GLP-1), which delays gastric emptying, and 2) a dairy breakfast in combination with a 3-h intravenous infusion of saline. Blood samples were drawn before breakfast and during the infusion. Postprandial ghrelin (total) responses were lower following the saline infusion compared with the GLP-1 infusion (P < 0.05). Acetaminophen concentrations, an indirect measurement of gastric emptying rate, were inversely correlated with total ghrelin concentrations (saline r = -0.76; 95% CI = -0.90, -0.49, GLP-1 r = -0.47; 95% CI = -0.76, -0.04). Ghrelin concentrations were only weakly correlated with insulin concentrations (saline r = -0.36; 95% CI = -0.69, 0.09; GLP- 1 r = -0.42; 95% CI = -0.73, 0.03), but strongly inversely correlated with GIP concentrations (saline r = -0.74; 95% CI= -0.89, -0.45; GLP-1 r = -0.63; 95% CI = -0.84, -0.27). In conclusion, our results support the hypothesis that ghrelin requires postgastric feedback, which may not be regulated through insulin. Conversely, our data suggest a role of glucose-dependent insulinotropic polypeptide in ghrelin secretion.     

Gut hormones in relation to body mass and torpor pattern changes during food restriction and re-feeding in the gray mouse lemur

Potential implications of gut hormones in body mass and torpor and behavioral pattern changes induced by an incremental (40 and 80%) calorie restriction (CR) in long-days (LD, summer) and short-days (SD, winter) were investigated in gray mouse lemurs. Only 80% food-deprived LD and SD animals showed a continuous mass loss resulting in a 10 and 15% mass reduction, respectively. Ghrelin levels of all food-deprived groups increased by 2.6-fold on average and remained high after re-feeding while peptide YY (PYY) levels increased by 3.8-fold only in LD animals under 80% CR. In the re-fed SD group, body mass was positively associated with ghrelin and negatively associated with PYY, while no correlations were noted in the re-fed LD animals. Plasma glucagon-like peptide-1 (GLP-1) increased by 2.9-fold only in LD food-restricted mouse lemurs and was negatively associated with the minimal body temperature. No significant correlations were reported in food-deprived SD animals. These results suggest that ghrelin, PYY and GLP-1 may be related to pre-wintering fattening mechanisms and to the modulation of torpor expression, respectively. Such observation clearly warrants further investigations, but it opens an interesting area of research in torpor regulation.     

Desacyl ghrelin inhibits the orexigenic effect of peripherally injected ghrelin in rats

Studies showed that the metabolic unlike the neuroendocrine effects of ghrelin could be abrogated by co-administered unacylated ghrelin. The aim was to investigate the interaction between ghrelin and desacyl ghrelin administered intraperitoneally on food intake and neuronal activity (c-Fos) in the arcuate nucleus in non-fasted rats. Ghrelin (13 μg/kg) significantly increased food intake within the first 30 min post-injection. Desacyl ghrelin at 64 and 127 μg/kg injected simultaneously with ghrelin abolished the stimulatory effect of ghrelin on food intake. Desacyl ghrelin alone at both doses did not alter food intake. Both doses of desacyl ghrelin injected separately in the light phase had no effects on food intake when rats were fasted for 12 h. Ghrelin and desacyl ghrelin (64 μg/kg) injected alone increased the number of Fos positive neurons in the arcuate nucleus compared to vehicle. The effect on neuronal activity induced by ghrelin was significantly reduced when injected simultaneously with desacyl ghrelin. Double labeling revealed that nesfatin-1 immunoreactive neurons in the arcuate nucleus are activated by simultaneous injection of ghrelin and desacyl ghrelin. These results suggest that desacyl ghrelin suppresses ghrelin-induced food intake by curbing ghrelin-induced increased neuronal activity in the arcuate nucleus and recruiting nesfatin-1 immunopositive neurons.     

Effect of ghrelin on chronic liver injury and fibrogenesis in male rats: Possible role of nitric oxide

Recent studies have revealed that ghrelin may be an antioxidant and anti-inflammatory agent in many organs, however its role in chronic liver injury (CLI) remains unclear. The role of nitric oxide (NO) in CLI is controversial as evidence suggests that NO is either a primary mediator of liver cell injury or exhibits a protective effect against injurious stimuli. Recent evidence demonstrated that the therapeutic potential for ghrelin was through eNOS activation and increase in NO production. However, its role on NO production in the liver has not been previously investigated. The aim of this study was to investigate the role of ghrelin in treatment of CLI, and whether this action is mediated through NO. Forty male rats were divided into four groups: Group I: Control; Group II: chronic liver injury (CLI); Group III: CLI + Ghrelin; and Group IV: CLI + Ghrelin + l-NAME. Liver enzymes and tumor necrosis factor alpha (TNF-α), were measured to assess hepatocellular injury. Liver tissue collagen content, malondialdehyde (MDA), gene expression of Bax, Bcl-2, and eNOS were assessed to determine the mechanism of ghrelin action. Results showed that ghrelin decreased serum liver enzymes and TNF-α levels. Ghrelin also reduced liver tissue collagen, MDA, and Bax gene expression, and increased Bcl-2 and eNOS gene expression. The effects on TNF-α, collagen, MDA, Bax, and eNOS were partially reversed in Group IV, suggesting that ghrelin's action could be through modulation of NO levels. Therefore, ghrelin's hepatoprotective effect is partially mediated by NO release.     

Regulation of ghrelin gene expression in stomach and feeding response to a ghrelin analogue in two strains of rats

Ghrelin is a peptide produced by the stomach and released into the circulation. As a natural ligand of the growth hormone secretagogue (GHS) receptor, it stimulates growth hormone secretion but it also stimulates feeding in humans and rodents. The orexigenic effect of ghrelin has been related to AgRP/NPY and orexin pathways. We proposed that ghrelin might be involved in the susceptibility to diet induced obesity and in the regulation of macronutrient selection. We have investigated these hypotheses in two strains of rat, the Osborne–Mendel (OM) rat that prefers diets high in fat and is sensitive to dietary obesity and the S5B/P1 (S5B) rat that prefers a low fat diet and is resistant to high fat diet induced obesity.

OM and S5B rats were adapted to a choice of high fat (HF) and low fat (LF) diet for 2 weeks. GHRP-2, an analogue of ghrelin, was injected intraperitoneally into satiated and 24 h fasted rats at doses of 10, 30 and 90 nmol. Food intake was measured over the next 4 h period. In satiated S5B rats, GHRP-2 stimulated intake of the LF diet in a dose dependent manner but did not affect the intake of the HF diet. In satiated OM rats, 90 nmol of GHRP-2 stimulated HF intake. In contrast, neither fasted OM nor S5B rats increased the intake of either HF or LF diet in response to GHRP-2. Fasting for 18 h induced a large rise in ghrelin mRNA in stomach of OM rats but not in S5B rats. There were no significant differences in plasma total ghrelin. An increase in ghrelin mRNA in stomach immediately before the onset of the dark cycle was observed in OM but not in S5B rats. Active ghrelin level was significantly affected by different feeding conditions in both OM and S5B rats adapted on HF diet with a trend to increase after 48 h of fasting and to decline to basal levels following 10 h of refeeding. These data suggest that ghrelin stimulates the intake of the preferred macronutrient. In addition, a differential regulation of ghrelin gene expression between OM and S5B rats may be important in their differential sensitivity to HF diet-induced obesity.     

Ghrelin与生殖系统研究进展

Ghrelin是1999年发现的生长激素促分泌素受体(growth hormone secretagogue receptor,GHS-R)的天然配体,由28个氨基酸残基组成.除具有促进生长激素的释放、增加摄食、刺激胃蠕动和胃酸分泌,尚有其它许多功能.近年来发现Ghrelin及其受体在生殖系统也广泛分布,提示Ghrelin对生殖系统也具有重要的调节作用,进一步的研究发现Ghrelin具有调节生殖激素黄体生成素、催乳素、雌二醇和孕酮的分泌,促进颗粒细胞的增殖等作用.本文就Ghrelin在生殖系统的研究进展做如下综述.     

Ghrelin, des-acyl ghrelin and obestatin: three pieces of the same puzzle

The major active product of ghrelin gene is a 28-amino acid peptide acylated at the serine 3 position with an octanoyl group, called simply ghrelin. Ghrelin has a multiplicity of physiological functions, affecting GH release, food intake, energy and glucose homeostasis, gastrointestinal, cardiovascular, pulmonary and immune function, cell proliferation and differentiation and bone physiology. Nevertheless, recent developments have shown that ghrelin gene can generate various bioactive molecules besides ghrelin, mainly des-acyl ghrelin and obestatin, obtained from alternative splicing or from extensive post-translational modification. Although their receptors have not yet been identified, they have already proven to be active, having intriguingly subtle but opposite physiological actions to ghrelin. This suggests the existence of a novel endocrine system with multiple effector elements which not only may have opposite actions but may regulate the action of each other. In this review, we summarize the steps which lead to the production of the different ghrelin gene products and examine the most significant differences between them in terms of structure and actions.