Leptin inhibits gastric emptying in rats: role of CCK receptors and vagal afferent fibers

Leptin regulates energy homeostasis and body weight by balancing energy intake and expenditure. It was recently reported that leptin, released into the gut lumen during the cephalic phase of gastric secretion, is capable of initiating intestinal nutrient absorption. Vagal afferent neurons also express receptors for both CCK and leptin, which are believed to interact in controlling food intake. The present study was undertaken to investigate the central and peripheral effects of leptin on gastric emptying rate. Under anesthesia, male Sprague-Dawley rats (250-300 g) were fitted with gastric Gregory cannulas (n=12) and some had additional cerebroventricular cannulas inserted into their right lateral ventricles. Following recovery, the rate of gastric emptying of saline (300 mOsm/kg H(2)O) was determined after instillation into the gastric fistula (3 ml, 37 degrees C, containing phenol red, 60 mg/l as a non-absorbable dilution marker). Gastric emptying rate was determined from the volume and phenol red concentrations recovered after 5 min. Leptin, injected intraperitoneally (i.p.; 10, 30, 60, 100 microg/kg) or intracerebroventricularly (i.c.v.; 5, 15 microg/rat) 15 min before the emptying, delayed gastric emptying rate of saline at the dose of 30 microg/kg or 15 microg/rat (p<0.001). When CCK(1) receptor blocker L-364,718 (1 mg/kg, i.p.), CCK(2) receptor blocker L-365,260 (1 mg/kg, ip) or adrenergic ganglion blocker bretylium tosylate (15 mg/kg, i.p.) was administered 15 min before ip leptin (30 microg/kg) injections, leptin-induced delay in gastric emptying was abolished only by the CCK(1) receptor blocker (p<0.001). However, the inhibitory effect of central leptin on gastric emptying was reversed by adrenergic blockade, but not by either CCK antagonists. Our results demonstrated that leptin delays gastric emptying. The peripheral effect of leptin on gastric motility appears to be mediated by CCK(1) receptors, suggesting the release of CCK and the involvement of vagal afferent fibers. On the other hand, the central effect of leptin on gastric emptying is likely to be mediated by adrenergic neurons. These results indicate the existence of a functional interaction between leptin and CCK receptors leading to inhibition of gastric emptying and short-term suppression of food intake, providing an additional feedback control in producing satiety.     

Preproghrelin-derived peptide, obestatin, fails to influence food intake in lean or obese rodents

Objectives: Obestatin has been initially characterized as a new peptide derived from the ghrelin precursor, which suppresses food intake and inhibits the orexigenic and prokinetic actions of ghrelin when injected peripherally or centrally in lean mice. However, reproducing these data remains controversial. Reasons for the disparity may be the use of different doses, routes, and animal models. We aimed to investigate the effects of peripheral and intracisternal (IC) injection of obestatin on feeding, gastric motility, and blood glucose in rats as well as in diet‐induced obese (DIO) mice. Research Methods and Procedures: Food intake and gastric emptying of a semi‐liquid caloric meal were measured after intraperitoneal (IP) injection of obestatin in rats and DIO mice. Gastric phasic motility and blood glucose were monitored in urethane‐anesthetized rats after IC or intravenous (IV) injection of obestatin. Results: Obestatin injected intraperitoneally at doses ranging from 0.1 to 3 mg/kg influenced neither acute food intake nor gastric emptying in rats. Obestatin injected intravenously at 0.3 or 3 mg/kg and IC at 7.5 or 30 µg/rat modified neither fasted gastric phasic motility nor blood glucose levels, while ghrelin (30 µg/kg, IV) increased and vagotomy suppressed gastric motility, and an oligosomatostatin analog (3 µg/rat, IC) decreased blood glucose. Obestatin, injected intraperitoneally (0.3 mg/kg) in DIO mice, did not alter feeding response to a fast, while urocortin 1 (10 µg/kg, IP) induced a 73.3% inhibition at 2 hours. Discussion: Our data demonstrate that peripheral administration of obestatin did not modify food intake in rats or obese mice or gastric motor function in rats.     

Obestatin对血浆ghrelin和nesfatin-1水平及胃排空的影响

目的:探讨侧脑室注射obestatin对大鼠血浆酰基化ghrelin、去酰基化ghrelin、nesfatin-1水平的影响以及对胃排空的调控。方法:侧脑室注射obestatin,采用酶免疫测定(EIA)法检测血浆酰基化ghrelin、去酰基化ghrelin、nesfatin-1水平以及胃排空率的变化。结果:侧脑室分别注射0.1、0.3或1.0 nmol obestatin,大鼠血浆酰基化ghrelin、去酰基化ghrelin以及nesfatin-1水平无显著改变(P0.05),且酰基化ghrelin与去酰基化ghrelin比率无显著改变(P0.05);侧脑室注射obestatin,大鼠摄食量无显著改变,但胃排空率明显增加(P0.05);胃排空率明显延迟(P0.05)。与侧脑室注射1.0 nmol Obestatin组相比,注射1.0 nmol Obestatin+CRF,大鼠摄食量无显著改变,胃排空率明显延迟(P0.05)。各组摄食量及进入十二指肠内食物量无明显差异(P0.05)。结论:中枢obestatin促进大鼠的胃排空,可能与h/r CRF通路有关。     

Lack of obestatin effects on food intake: should obestatin be renamed ghrelin-associated peptide (GAP)?

Obestatin is a newly identified ghrelin-associated peptide (GAP) that is derived from post-translational processing of the prepro-ghrelin gene. Obestatin has been reported initially to be the endogenous ligand for the orphan receptor G protein-coupled receptor 39 (GPR39), and to reduce refeeding- and ghrelin-stimulated food intake and gastric transit in fasted mice, and body weight gain upon chronic peripheral injection. However, recent reports indicate that obestatin is unlikely to be the endogenous ligand for GPR39 based on the lack of specific binding on GRP39 receptor expressing cells and the absence of signal transduction pathway activation. In addition, a number of studies provided convergent evidence that ghrelin injected intracerebroventricularly or peripherally did not influence food intake, body weight gain, gastric transit, gastrointestinal motility, and gastric vagal afferent activity, as well as pituitary hormone secretions, in rats or mice. Similarly, obestatin did not alter ghrelin-induced stimulation of food intake or gastric transit. Therefore, the present state-of-knowledge on obestatin and GPR39 is leaving many unanswered questions that deserve further consideration. Those relate not only to redefining the biological action of obestatin that should be renamed GAP, but also the identification of the native ligand for GPR39.     

Gastric emptying in response to IAPP and CCK in rats with subdiaphragmatic afferent vagotomy

In the subdiaphragmatic vagal deafferentation procedure (SDA), the afferent fibers of the vagus are surgically severed unilaterally where they enter the brain stem. The technique includes a subdiaphragmal truncal vagotomy performed on the contralateral side. This procedure has been used to study the control of food intake, but it has not been used previously to investigate the role of vagal afferent fibers in the control of gastric emptying (GE). The current experiment studied the effect of SDA on the inhibition of GE by islet amyloid polypeptide (IAPP or amylin) and cholecystokinin (CCK) in awake, unrestrained rats with gastric cannulas. The experimental group underwent subdiaphragmatic vagal deafferentation; the control group had sham operations. All rats received 20-min intravenous infusions of IAPP (1, 3, 9, 27, and 81 pmol/kg/min), CCK (3, 30 and 90 pmol/kg/min), and normal saline in random order. Gastric emptying of saline was measured by the phenol red method during the last 5 min of each infusion period. CCK dose-dependently inhibited gastric emptying in both the control and SDA animals. The inhibition of GE by CCK was significantly attenuated by SDA (p<0.01). IAPP also inhibited gastric emptying dose-dependently, but the difference between the SDA and control groups was not significant. The current experiment, which used a different methodology than previous studies, provides support for the hypothesis that the inhibition of gastric emptying by CCK, but not by IAPP, is mediated partly by afferent vagal fibers.     

Peripheral obestatin has no effect on feeding behavior and brain Fos expression in rodents

Obestatin is produced in the stomach from proghrelin by post-translational cleavage. The initial report claimed anorexigenic effects of obestatin in mice. Contrasting studies indicated no effect of obestatin on food intake (FI). We investigated influences of metabolic state (fed/fasted), environmental factors (dark/light phase) and brain Fos response to intraperitoneal (ip) obestatin in rats, and used the protocol from the original study assessing obestatin effects in mice. FI was determined in male rats injected ip before onset of dark or light phase, with obestatin (1 or 5 μmol/kg), CCK8S (3.5 nmol/kg) or 0.15 M NaCl, after fasting (16 h, n = 8/group) or ad libitum (n = 10–14/group) food intake. Fos expression in hypothalamic and brainstem nuclei was examined in freely fed rats 90 min after obestatin (5 μmol/kg), CCK8S (1.75 nmol/kg) or 0.15 M NaCl (n = 4/group). Additionally, fasted mice were injected ip with obestatin (1 μmol/kg) or urocortin 1 (2 nmol/kg) 15 min before food presentation. No effect on FI was observed after obestatin administration during the light and dark phase under both metabolic conditions while CCK8S reduced FI irrespectively of the conditions. The number of Fos positive neurons was not modified by obestatin while CCK8S increased Fos expression in selective brain nuclei. Obestatin did not influence the refeeding response to a fast in mice, while urocortin was effective. Therefore, peripheral obestatin has no effect on FI under various experimental conditions and did not induce Fos in relevant central neuronal circuitries modulating feeding in rodents.     

Abdominal vagal mediation of the satiety effects of CCK in rats

CCK type 1 (CCK1) receptor antagonists differing in blood-brain barrier permeability were used to test the hypothesis that satiety is mediated in part by CCK action at CCK1 receptors on vagal sensory nerves innervating the small intestine. Devazepide penetrates the blood-brain barrier; A-70104, the dicyclohexylammonium salt of N alpha-3-quinolinoyl-D-Glu-N,N-dipentylamide, does not. At dark onset, non-food-deprived control rats and rats with subdiaphragmatic vagotomies received a bolus injection of devazepide (2.5 micromol/kg i.v.) or a 3-h infusion of A-70104 (3 micromol.kg(-1).h(-1) i.v.) either alone or coadministered with a 2-h intragastric infusion of peptone (0.75 or 1 g/h). Food intake was determined from continuous computer recordings of changes in food bowl weight. In control rats both antagonists stimulated food intake and attenuated the anorexic response to intragastric infusion of peptone. In contrast, only devazepide was effective in stimulating food intake in vagotomized rats. Thus endogenous CCK appears to act both at CCK1 receptors beyond the blood-brain barrier and by a CCK1 receptor-mediated mechanism involving abdominal vagal nerves to inhibit food intake.     

NMDA receptor blockade attenuates CCK-induced reduction of real feeding but not sham feeding

Systemic injection of MK-801, a noncompetitive antagonist of N-methyl-D-aspartate (NMDA) receptor ion channels, increases meal size and delays satiation. We examined whether MK-801 increases food intake by directly interfering with actions of cholecystokinin (CCK). Prior administration of MK-801 (100 microg/kg ip) reversed the inhibitory effects of CCK-8 (2 and 4 microg/kg ip) on real feeding of both liquid and solid foods. MK-801 alone did not alter 30-min sham intake of 15% sucrose compared with intake after saline. Furthermore, while CCK-8 (2 or 4 microg/kg ip) reduced sham intake, this reduction was not attenuated by MK-801 pretreatment. To ascertain whether MK-801 attenuation of CCK-induced reduction of real feeding was associated with attenuated inhibition of gastric emptying, we tested the effect of MK-801 pretreatment on CCK-induced inhibition of gastric emptying of 5-ml saline loads. Ten-minute gastric emptying was accelerated after MK-801 (3.9 +/- 0.2 ml) compared with saline vehicle (2.72 +/- 0.2 ml). CCK-8 (0.5 microg/kg ip) reduced 10-min emptying to 1.36 +/- 0.3 ml. Pretreatment with MK-801 did not significantly attenuate CCK-8-induced reduction of gastric emptying (0.9 +/- 0.4 ml). This series of experiments demonstrates that blockade of NMDA ion channels reverses inhibition of real feeding by CCK. However, neither inhibition of sham feeding nor inhibition of gastric emptying by CCK is attenuated by MK-801. Therefore, increased food intake after NMDA receptor blockade is not caused by a direct interference with CCK-induced satiation. Rather, increased real feeding, either in the presence or absence of CCK, depends on blockade of NMDA receptor participation in other post-oral feedback signals such as gastric sensation or gastric tone.     

High fat maintenance diet attenuates hindbrain neuronal response to CCK

Rats maintained on a high fat diet reduce their food intake less in response to exogenous cholecystokinin (CCK) than rats maintained on a low fat diet. In addition, inhibition of gastric emptying by CCK is markedly attenuated in rats maintained on a high fat diet. Both inhibition of food intake and gastric emptying by CCK are mediated by sensory fibers in the vagus nerve. These fibers terminate on dorsal hindbrain neurons of the nucleus of the solitary tract and area postrema. To determine whether diet-induced changes in the control of feeding and gastric emptying are accompanied by altered vagal sensory responsiveness, we examined dorsal hindbrain expression of Fos-like immunoreactivity (Fos-li) following intraperitoneal CCK injection of rats maintained on high fat or low fat diets. Following CCK, there were numerous Fos-li nuclei in the area postrema and in the commissural and medial subnuclei of the nucleus of the solitary tract of rats maintained on a low fat diet. However, Fos-li was absent or rare in the brains of rats maintained on a high fat diet. These data suggest that the vagal sensory response to exogenous CCK is reduced in rats maintained on a high fat diet. Our results also are consistent with our previous findings that CCK-induced reduction of food intake and gastric emptying are both attenuated in rats maintained on a high fat diet. In addition our results support the hypothesis that attenuation of CCK-induced inhibition of food intake and gastric emptying may be due to diet-induced diminution of vagal CCK responsiveness.     

Peripheral urocortin inhibits gastric emptying and food intake in mice: differential role of CRF receptor 2

Intraperitoneal urocortin inhibits gastric emptying and food intake in mice. We investigated corticotropin-releasing factor receptor (CRF-R) subtypes involved in intraperitoneal urocortin actions using selective CRF-R antagonists. Gastric emptying was measured 2 h after a chow meal, and food intake was measured hourly after an 18-h fast in mice. Urocortin (3 microg/kg ip) inhibited gastric emptying by 88%. The CRF-R1/CRF-R2 antagonist astressin B (30 microg/kg ip) and the selective CRF-R2 antagonist antisauvagine-30 (100 microg/kg ip) completely antagonized urocortin action, whereas the selective CRF-R1 antagonist CP-154,526 (10 mg/kg ip) had no effect. Urocortin (1-10 microg/kg ip) dose dependently decreased the 2-h cumulative food intake by 30-62%. Urocortin (3 microg/kg)-induced hypophagia was completely antagonized by astressin B (30 microg/kg ip) and partially (35 and 31%) by antisauvagine-30 (100 or 200 microg/kg ip). The CRF-R1 antagonists CP-154,526 or DMP904 (10 mg/kg ip) had no effect. Capsaicin did not alter urocortin-inhibitory actions while blocking the satiety effect of intraperitoneal CCK. These data indicate that intraperitoneal urocortin-induced decrease in feeding is only partly mediated by CRF-R2, whereas urocortin action to delay gastric emptying of a meal involves primarily CRF-R2.     

Gastroprotection and control of food intake by leptin. Comparison with cholecystokinin and prostaglandins.

Circulating peptide leptin which is the product of the ob gene is known to provide feedback information on the size of fat stores to central OB-receptors that control food intake. Recently, leptin messenger RNA and leptin protein have been detected in gastric epithelium and leptin was found to be released by CCK into circulation but the physiological role of this gastric leptin remains unknown. As CCK has been reported to protect gastric mucosa against various noxious agents, we designed the study to determine the influence of leptin and CCK on the gastroprotection and the control of food intake and to compare them with classic gastroprotective substance, prostaglandin E2, in rats with acute gastric mucosal lesions induced by topical application of 75% ethanol. Four series of Wistar rats (A, B, C and D) were used to determine; A) the effects of various doses of leptin (0.1-10 microg/kg) given intraperitoneally (i.p.) on ethanol-induced gastric lesions, gastric blood flow (GBF) and plasma levels of immunoreactive leptin; B) the effects of various doses of CCK-8 (0.1-10 microg/kg i.p.) on ethanol-induced gastric lesions, GBF and plasma levels of leptin; C) the effects of various doses of PGE2 (12.5--100 microg/kg) given intragastrically (i.g.) on ethanol-induced gastric lesions and GBF and D) the influence of leptin, CCK and PGE2 on the intake of liquid meal in rats. Rats were anesthetized with ether 1 h after i.g. administration of 75% ethanol to measure the GBF using H2-gas clearance technique and blood samples were withdrawn for the measurement of plasma leptin levels by radioimmunoassay (RIA). Food intake was assessed in separate group of rats fasted 18 h and then fed with liquid caloric meal. Leptin, CCK and PGE2 reduced dose-dependently gastric lesions induced by 75% ethanol, the dose reducing these lesions by 50% (ED50) being, respectively, 1 microg/kg, 5 microg/kg and 20 microg/kg. The protective effects of leptin, CCK-8 and PGE2 were accompanied by significant attenuation of the fall of the GBF caused by ethanol. Leptin and CCK reduced also dose-dependently the food intake while PGE2 was not effective. Leptin and CCK resulted a dose-dependent increment in the plasma leptin levels. We conclude that: 1) exogenous leptin and CCK, causing similar increments in plasma immunoreactive leptin levels, protect dose-dependently gastric mucosa against the damage provoked by 75% ethanol; 2) Leptin and CCK afford similar gastroprotective activity to that attained with PGE2 but unlike PGE2 were highly effective in the reduction in food intake and 3) the protective effects of leptin, CCK and PGE2 were accompanied by significant increase of GBF suggesting that the protection afforded by these substances are mediated, at least in part, by gastric hyperemia.     

GABA(B) receptors on vagal afferent pathways: peripheral and central inhibition

To investigate GABA(B) receptors along vagal afferent pathways, we recorded from vagal afferents, medullary neurons, and vagal efferents in ferrets. Baclofen (7-14 micromol/kg i.v.) reduced gastric tension receptor and nucleus tractus solitarii neuronal responses to gastric distension but not gastroduodenal mucosal receptor responses to cholecystokinin (CCK). GABA(B) antagonists CGP-35348 or CGP-62349 reversed effects of baclofen. Vagal efferents showed excitatory and inhibitory responses to distension and CCK. Baclofen (3 nmol i.c.v. or 7-14 micromol/kg i.v.) reduced both distension response types but reduced only inhibitory responses to CCK. CGP-35348 (100 nmol i.c.v. or 100 micromol/kg i.v.) reversed baclofen's effect on distension responses, but inhibitory responses to CCK remained attenuated. They were, however, reversed by CGP-62349 (0.4 nmol i.c.v.). In conclusion, GABA(B) receptors inhibit mechanosensitivity, not chemosensitivity, of vagal afferents peripherally. Mechanosensory input to brain stem neurons is also reduced centrally by GABA(B) receptors, but excitatory chemosensory input is unaffected. Inhibitory mechano- and chemosensory inputs to brain stem neurons (via inhibitory interneurons) are both reduced, but the pathway taken by chemosensory input involves GABA(B) receptors that are insensitive to CGP-35348.     

糖尿病大鼠弓状核-海马肥胖抑素通路构成及对胃运动和胃排空的影响

目的: 探究糖尿病大鼠弓状核(ARC)-海马肥胖抑素(obestatin)神经通路构成,以及该通路对大鼠胃运动、胃排空的影响。方法: 健康雄性Wistar大鼠采用果糖溶液诱导胰岛素抵抗加腹腔注射链脲佐菌素的方法制备糖尿病模型,造模之后,随机分为5组:对照组(NS组)、0.1、1和10 pmol obestatin组、obestatin+NBI27914组,每组7只;各组通过置管分别向海马内注射0.5 μl 生理盐水(NS)、obestatin(0.1 pmol、1 pmol、10 pmol)和混合液(10 pmol obestatin + 60 pmol NBI27914),给药后立即记录大鼠胃运动,15 min后进行胃排空研究;通过荧光金(FG)逆行追踪及免疫组化方法比较正常及糖尿病大鼠ARC-海马obestatin神经通路构及ARC obestatin mRNA表达的异同。结果: 与正常大鼠相比,糖尿病大鼠ARC FG/obestatin双标神经元数目显著减少(P＜0.05),ARC obestatin mRNA表达量显著下降(P＜0.05);obestatin各组可剂量依赖性的抑制大鼠胃运动及胃排空(P＜0.05~0.01),obestatin的这些效应可被促肾上腺皮质激素受体1(CRFR1)阻断剂NBI27914部分阻断(P＜0.05);obestatin对糖尿病大鼠胃运动和胃排空的抑制效应显著减弱(P＜0.05)。结论: ARC-海马之间存在obestatin神经和功能通路,参与糖尿病大鼠胃运动及胃排空调控,且CRFR1信号通路参与该过程。该通路功能的减弱可能参与了糖尿病早期胃动力紊乱的发病。     

Metabolic effects of chronic obestatin infusion in rats

Obestatin is purported to be a peptide hormone encoded in preproghrelin. We studied the metabolic effects of continuous infusion of obestatin via subcutaneously implanted osmotic mini-pumps. Administration of up to 500nmol/kg body weight/day obestatin did not change 24h cumulative food intake or body weight in rats. Similarly, no effects were observed when obestatin was infused at 1000nmol/kg body weight/day for seven days. This dose of obestatin infused during a 24h fast did not alter weight loss, suggesting that obestatin has no effect on energy expenditure, and this dose did not alter glucose or insulin responses during an IPGTT. Obestatin was originally proposed to interact with GPR39 and subsequently the receptor for GLP-1. While both receptors are expressed in pancreatic islets, incubation with obestatin did not alter insulin release from islets in vitro. Moreover, obestatin did not bind to INS-1 beta-cells or HEK cells overexpressing GLP-1 receptors or displace GLP-1 binding to these cells. Our findings do not support the concept that obestatin is a hormone with metabolic actions.     

Targeted disruption of the murine CCK1 receptor gene reduces intestinal lipid-induced feedback inhibition of gastric function

Cholecystokinin (CCK), acting at CCK1 receptors (CCK1Rs) on intestinal vagal afferent terminals, has been implicated in the control of gastrointestinal function and food intake. Using CCK1R(-/-) mice, we tested the hypothesis that lipid-induced activation of the vagal afferent pathway and intestinal feedback of gastric function is CCK1R dependent. In anesthetized CCK1R(+/+) ("wild type") mice, meal-stimulated gastric acid secretion was inhibited by intestinal lipid infusion; this was abolished in CCK1R(-/-) mice. Gastric emptying of whole egg, measured by nuclear scintigraphy in awake mice, was significantly faster in CCK1R(-/-) than CCK1R(+/+) mice. Gastric emptying of chow was significantly slowed in response to administration of CCK-8 (22 pmol) in CCK1R(+/+) but not CCK1R(-/-) mice. Activation of the vagal afferent pathway was measured by immunohistochemical localization of Fos protein in the nucleus of the solitary tract (NTS; a region where vagal afferents terminate). CCK-8 (22 pmol ip) increased neuronal Fos expression in the NTS of fasted CCK1R(+/+) mice; CCK-induced Fos expression was reduced by 97% in CCK1R(-/-) compared with CCK1R(+/+) mice. Intralipid (0.2 ml of 20% Intralipid and 0.04 g lipid), but not saline, gavage increased Fos expression in the NTS of fasted CCK1R(+/+) mice; lipid-induced Fos expression was decreased by 47% in CCK1R(-/-) compared with CCK1R(+/+)mice. We conclude that intestinal lipid activates the vagal afferent pathway, decreases gastric acid secretion, and delays gastric emptying via a CCK1R-dependent mechanism. Thus, despite a relatively normal phenotype, intestinal feedback in response to lipid is severely impaired in these mice.     

A new simple mouse model for the in vivo evaluation of cholecystokinin (CCK) antagonists: comparative potencies and durations of action of nonpeptide antagonists

A new simple mouse assay for the in vivo evaluation of CCK antagonists which is based upon visual determination of the gastric emptying of a charcoal meal is described. CCK-8 (24 micrograms/kg s.c.) but not various other peptide and nonpeptide agents effectively inhibited gastric emptying in this test system. The effect of CCK-8 was antagonized by established peripheral CCK antagonists but not representative agents of various other pharmacological classes. The rank order of potency of the CCK antagonists were: L-364,718 (ED50 = 0.01 mg/kg, i.v.; 0.04 mg/kg, p.o.) greater than Compound 16 (ED50 = 1.5 mg/kg, i.v.; 2.0 mg/kg p.o.) greater than asperlicin (ED50 = 14.8 mg/kg i.v.) greater than proglumide (ED50 = 184 mg/kg i.v.; 890 mg/kg, p.o.). Duration of action studies based upon ED50 values determined at various time intervals after oral administration showed that L-364,718 and proglumide are considerably longer acting than Compound 16. Asperlicin (ED50 greater than 300 mg/kg, p.o.) was ineffective as a CCK antagonist when administered orally. These data provide the first direct comparisons of the in vivo potencies of current CCK antagonists and demonstrate the utility of a new simple mouse assay for the in vivo characterization of peripheral CCK antagonists.     

Inhibitory effects of beta-alanine on the vagal efferent and afferent excitatory responses of the stomach to stimulation of vagal trunk in cats.

The effects of beta-alanine on the electrically evoked vagal efferent (hexamethonium-sensitive initial excitatory response) and afferent (hexamethonium-resistant delayed excitatory response) responses of the cat stomach were studied. beta-alanine (30 to 300 micrograms/kg, i.v.) dose-dependently inhibited both the efferent and afferent response. The IC50 values of beta-alanine on the efferent and afferent response were 296 +/- 65 micrograms/kg and 128 +/- 35 microgram/kg, respectively. Maximal inhibitory effects of beta-alanine (300 micrograms/kg, i.v.) appeared about 1 hr after the injection. Glycine and taurine (100 to 10,000 micrograms/kg) did not affect these responses. Treatment with hexamethonium (10 mg/kg, i.v.) prevented the efferent response, but augmented the afferent response. The treatment with hexamethonium abolished the inhibitory effect of beta-alanine on the afferent response. Both picrotoxin (100 and 500 micrograms/kg, i.v.) and bicuculline (2000 micrograms/kg, i.v.) antagonized the inhibitory effects of beta-alanine on the vagal efferent and afferent responses of the stomach. The present experiments clearly demonstrated that beta-alanine inhibited both the vagal efferent and afferent excitatory responses of stomach to electrical stimulation of vagal trunk in cats.     

c-Kit mutant mouse behavioral phenotype: altered meal patterns and CCK sensitivity but normal daily food intake and body weight

The mouse W/Wv mutation of the c-Kit receptor causes extensive loss of gastrointestinal interstitial cells of Cajal and vagal intramuscular arrays (IMAs; one of the two putative mechanoreceptors in gastrointestinal smooth muscle). To characterize the behavioral phenotype of the c-Kit mouse and to evaluate the roles of these mechanoreceptors in controlling food intake, meal patterns and daily intakes of W/Wv mice and controls were examined using solid (20-mg pellets) and liquid (Isocal) maintenance diets. After the meal pattern experiments, CCK (0.5, 1, 2, 4, 8, and 16 microg/kg ip) was administered to examine the role of the interstitial cells and vagal IMA mechanoreceptors in relaying peripheral signals of satiety activated by CCK-A receptors, whereas the specificity of the response was assessed with the antagonist devazepide (300 microg/kg ip). On both diets, the W/Wv mice ate smaller meals for shorter durations, with a compensatory increase in meal number, resulting in daily intakes and body weights similar to the controls. After CCK injections, the mutant mice consistently suppressed intake more ( approximately 2x) in 30-min tests, regardless of the test diet (12.5% glucose, chow, pellets, and Isocal). The increased sensitivity of W/Wv mice to CCK reflected an increased potency of the hormone (c-Kit mouse ED50 = 2.4 microg/kg; control ED50 = 6.4 microg/kg) and a shift of the dose-response curve to the left. Devazepide blocked the CCK suppression of ingestion. These results indicate that the selective loss of the interstitial cells and IMAs disrupts short-term feeding of the W/Wv mice by inducing an earlier satiety, possibly by altering gastric accommodation and/or emptying, without affecting the long-term mechanisms controlling overall intake or body weight. The results also suggest that the reduction of interstitial cells and IMAs augments the sensitivity to or increases the efficiency of exogenous CCK.     

Effects of juice from Morinda citrifolia (Noni) on gastric emptying in male rats

The effects of juice from Morinda citrifolia (noni) on gastric emptying, gastrointestinal transit, and plasma level of cholecystokinin (CCK) in rats were studied. Male rats were given noni by gavage at levels of 0.25, 1, or 4 ml/kg once per day for one or 7 days. The rats in the control group were given water, while the rats in the experimental group were fasted overnight before measurement of gastrointestinal motility. Gastrointestinal motility was assessed in rats 15 min after intragastric instillation of a test meal containing charcoal (10%) and Na251CrO4 (0.5 microCi/ml). Gastric emptying was determined by measuring the amount of radiolabeled chromium contained in the small intestine as a percentage of the initial amount received. Then, gastrointestinal transit was evaluated by calculating the geometric center of distribution of the radiolabeled marker. Finally, blood samples were collected for measurement of CCK by radioimmunoassay. The administration of noni at 0.25 ml/kg, but not at 1 ml/kg and 4 ml/kg, for 1 day significantly inhibited gastric emptying. In contrast, gastric emptying was significantly inhibited by oral noni (0.25, 1, or 4 ml/kg) for 7 days. Intraperitoneal injection of lorglumide (5 or 10 mg/kg), a selective CCK1 receptor antagonist, effectively attenuated the noni-induced inhibition of gastric emptying. The intestinal transit and body weight, food intake, water intake, urine volume as well as feces weight were not altered by the administration of noni either acutely or chronically, but the administration of oral noni (1 ml/kg) for 7 days increased the level of plasma CCK in male rats. These results suggest that oral noni inhibits gastric emptying in male rats via a mechanism involving stimulation of CCK secretion and CCK1 receptor activation.     

Exogenous and endogenous ghrelin in gastroprotection against stress-induced gastric damage

Ghrelin, identified in the gastric mucosa has been involved in control of food intake and growth hormone (GH) release but little is known about its influence on gastric secretion and mucosal integrity. The effects of ghrelin on gastric secretion, plasma gastrin and gastric lesions induced in rats by 75% ethanol or 3.5 h of water immersion and restraint stress (WRS) were determined. Exogenous ghrelin (5, 10, 20, 40 and 80 microg/kg i.p.) increased gastric acid secretion and attenuated gastric lesions induced by ethanol and WRS and this was accompanied by the significant rise in plasma ghrelin level, gastric mucosal blood flow (GBF) and luminal NO concentrations. Ghrelin-induced protection was abolished by vagotomy and attenuated by suppression of COX, deactivation of afferent nerves with neurotoxic dose of capsaicin or CGRP(8-37) and by inhibition of NOS with L-NNA but not influenced by medullectomy and administration of 6-hydroxydopamine. We conclude that ghrelin exerts a potent protective action on the stomach of rats exposed to ethanol and WRS, and these effects depend upon vagal activity, sensory nerves and hyperemia mediated by NOS-NO and COX-PG systems.