Effect of intraperitoneal CCK-8 on food intake and brain orexin-A after 48 h of fasting in the rat

We investigated the interactions of the peripheral satiety peptide cholecystokinin and the brain orexin-A system in the control of food intake. The effect of an intraperitoneal (i.p.) injection of sulfated cholecystokinin octapeptide (in this article called CCK) (5 microg/kg, 4.4 nmol/kg) or of phosphate-buffered saline (PBS, vehicle control) on 48 h fasting-induced feeding and on orexin-A peptide content was analyzed in diverse brain regions innervated by orexin neurons and involved in the control of food intake. Administration of CCK after a 48 h fast reduced fasting-induced hyperphagia (P<0.05). I.p. CCK increased the orexin-A content in the posterior brainstem of 48 h fasted rats by 35% (P<0.05). Fed animals receiving CCK had 48% higher orexin-A levels in the posterior brainstem than fasted rats (P<0.05). In the lateral hypothalamus, fasting decreased orexin-A levels by 50% as compared to fed rats (P<0.05). In the septal nuclei, the combination of fasting and CCK administration reduced orexin-A contents compared to fed PBS and CCK animals by 13% and 17%, respectively (P<0.05). These results suggest a convergence of pathways activated by peripheral CCK and by fasting on the level of orexin-A released in the posterior brainstem and provide evidence for a novel interaction between peripheral satiety signaling and a brain orexigen in the control of food intake.     

The H3 receptor is involved in cholecystokinin inhibition of food intake in rats.

We investigated the peripheral effects of an H3-receptor agonist and an H3-receptor antagonist (R)alpha-methylhistamine (Ralpha-MeHA) and thioperamide, respectively, on basal feeding and the CCK8-induced inhibition of food intake in rat. Intraperitoneal injection of thioperamide reduced food intake in a dose-dependent manner with maximal inhibition (35%, P<0.01 vs saline) at 3 mg/kg. (R)alpha-MeHA (0.3-3 mg/kg i.p.), an H3-receptor agonist alone had no effect on feeding but reversed the thioperamide-induced inhibition of food intake in a dose-dependent manner. The maximal feeding inhibitory dose of thioperamide (3 mg.kg i.p) increased by 40% and 22 % (P<0.01 vs saline) brain and stomach histamine contents, respectively. Histamine (0.3 - 6 mg/kg i.p.) and CCK-8 (3 - 30 microg/kg i.p) also inhibited food intake in a dose-dependent manner. Inhibition was 20% to 40% for histamine and 40% to 80% (P<0.01 vs saline) for CCK8. CCK-8 inhibition of feeding was increased by thioperamide and prevented by (R)alpha-MeHA in a dose-dependent way. In addition, CCK-8 did not reduce food intake if rats were pretreated with pyrilamine or ranitidine postsynaptic H1- and H2-receptor antagonists respectively. Our data suggest that the H3-receptor is involved in basal feeding. They also suggest that CCK satiety depends upon the release of histamine which acts on the H2- and H1-receptors, the final mediators of this effect.     

Unchanged hypothalamic neuropeptide Y concentrations in hyperphagic, hypoglycemic rats: evidence for specific metabolic regulation of hypothalamic NPY

Hypothalamic concentrations of neuropeptide Y (NPY), a potent central appetite stimulant, increase dramatically in food-restricted and insulin-deficient diabetic rats. This suggest that NPY may drive hyperphagia in these conditions, which are characterized by weight loss and insulin deficiency. To test the hypothesis that insulin deficiency and weight loss are specific stimuli to hypothalamic NPY, we measured NPY concentrations in individual hypothalamic regions in rats with hyperphagia caused by insulin-induced hypoglycemia. Groups of 8 male Wistar rats were injected with ultralente insulin (20-60 U/kg) to induce either acute hypoglycemia (7 h after a single injection) or chronic hypoglycemia (8 days with daily injections). In hypoglycemic rats, plasma insulin concentrations were increased 6- to 7-fold compared with saline-injected controls; food intake was significantly increased with acute and chronic hypoglycemia and weight gain was significantly increased in the chronically hypoglycemic group. NPY concentrations were measured by radioimmunoassay in 8 hypothalamic regions microdissected from fresh brain slices. NPY concentrations were not increased in any region in either acute or chronic hypoglycemia. NPY therefore seems unlikely to mediate hyperphagia in hyperinsulinemia-induced hypoglycemia, supporting the hypothesis that weight loss is a specific stimulus to hypothalamic NPY and that insulin deficiency may be the metabolic signal responsible.     

Peripheral leptin administration alters hormone and metabolite levels in the young pig

The present study was conducted to determine if peripheral leptin administration can alter GH secretion or feed intake in young pigs. Six, 6 kg female pigs were fasted overnight and randomly chosen to receive porcine recombinant leptin or saline injections in a crossover design. Three leptin dosages were tested over a 10 day period, 100, 200 or 500 microg/kg body mass (L100, L200 or L500). Leptin was administered in 0.2% bovine serum albumin as a bolus injection into the carotid artery. Blood samples were obtained from the jugular vein over a 24 h period. Leptin delayed feeding in pigs treated with L200 and L500 (P<0.05), while reducing overall intake in pigs treated with L100 (P<0.05). L200 or L500 depressed blood glucose (P<0.05). Plasma insulin levels were elevated by feeding in control animals, while insulin levels were depressed in pigs treated with L200 or L500 (P<0.05). L200 elevated plasma growth hormone (P<0.05) with three peaks apparent at 5, 8, and 13 h post injection. The ability for a single injection of leptin to produce significant changes in hormone and metabolite levels suggests that this peptide has a role in regulation of peripheral metabolism.     

Interactions between gonadotropin-releasing hormone (GnRH) and orexin in the regulation of feeding and reproduction in goldfish (Carassius auratus)

Links between energy homeostasis and reproduction have been demonstrated in vertebrates. As a general rule, abundant food resources favor reproduction whereas low food availability induces an inhibition of reproductive processes. In both mammals and fish, gonadotropin-releasing hormone (GnRH) and orexin (OX) are hypothalamic neuropeptides that play critical roles in the regulation of sexual behavior and appetite, respectively. In order to assess possible interactions between orexin and GnRH in the control of feeding and reproduction in goldfish, we examined the effects of chicken GnRH (cGnRH-II) intracerebroventricular (ICV) injection on feeding behavior and OX brain mRNA expression as well as the effects of orexin ICV injections on spawning behavior and cGnRH-II brain mRNA expression. Treatment with cGnRH-II at doses that stimulate spawning (0.5 ng/g or 1 ng/g) resulted in a decrease in both food intake and hypothalamic orexin mRNA expression. Treatment with orexin A at doses that stimulate feeding (10 ng/g) induced an inhibition of spawning behavior and a decrease in cGnRH-II expression in the hypothalamus and optic tectum-thalamus. Our results suggest that the anorexigenic actions of cGnRH-II in goldfish might be in part mediated by OX and that orexin inhibits reproductive behavior in part via the inhibition of the GnRH system. Our data suggest the existence of a coordinated control of feeding and reproduction by the orexin and GnRH systems in goldfish.     

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.     

Reduced CCK signaling in obese-prone rats fed a high fat diet

Deficits in satiation signaling during obesogenic feeding have been proposed to play a role in hyperphagia and weight gain in animals prone to become obese. However, whether this impaired signaling is due to high fat (HF) feeding or to their obese phenotype is still unknown. Therefore, in the current study, we examined the effects of CCK-8 (0.5, 1.0, 2.0, and 4.0 μg/kg) on suppression of food intake of HF-fed obese prone (OP) and resistant (OR) rats. Additionally, we determined the role of endogenous CCK in lipid-induced satiation by measuring plasma CCK levels following a lipid gavage, and tested the effect of pretreatment with devazepide, a CCK-1R antagonist on intragastric lipid-induced satiation. Finally, we examined CCK-1R mRNA levels in the nodose ganglia. We show that OP rats have reduced feeding responses to the low doses of exogenous CCK-8 compared to OR rats. Furthermore, OP rats exhibit deficits in endogenous CCK signaling, as pretreatment with devazepide failed to abolish the reduction in food intake following lipid gavage. These effects were associated with reduced plasma CCK after intragastric lipid in OP but not OR rats. Furthermore, HF feeding resulted in downregulation of CCK-1Rs in the nodose ganglia of OP rats. Collectively, these results demonstrate that HF feeding leads to impairments in lipid-induced CCK satiation signaling in obese-prone rats, potentially contributing to hyperphagia and weight gain.     

MK-329 blocks the inhibition of alcohol intake by CCK-8

Peripheral administration of sulfated cholecystokinin octapeptide (CCK-8) potently reduces alcohol intake, preference, and blood levels in rats. MK-329 (L-364,718 or Devazepide) acts at peripheral cholecystokinin (CCK_A) receptors to antagonize CCK-8's physiological and behavioral effects, such as pancreatic stimulation and inhibition of feeding. We determined whether CCK_A receptor blockade would also prevent CCK-8's alcohol satiety effect. Water-deprived female and male rats (n = 7 for each) received randomized combinations of intraperitoneal injections of MK-329 (0, 100, 200, or 400 μg/kg) followed by CCK-8 (0 or 4 μg/kg). Rats were then given access to 5% w/v ethanol for 30 min, followed by 30-min access to water, with food ad lib. MK-329 at all doses significantly (p < 0.05) reduced the suppression of alcohol intake and food intake by CCK-8. MK-329 alone increased alcohol intake at 400 μg/kg, and increased food intake, in females and males at 100 and 200 μg/kg, respectively. We concluded that CCK-8's alcohol and food satiation effects depend on specific, peripheral CCK_A receptors, and satiation of alcohol consumption and drinking-associated feeding reflect an endogenous functional interaction of CCK-8 with CCK_A receptors.     

Food intake in rhesus monkeys following central administration of orexins

Orexin A has been reported to stimulate food intake in rats while orexin B does not. The purpose of this study was to determine the role of orexin A or orexin B administration on food intake in adult, male rhesus monkeys. Food intake was measured at 2 and 8 h after the morning feeding following central injections of vehicle, orexin A (10, or 20 microg) or orexin B (10, 30, or 100 microg). When compared to vehicle injections, the 10 and 20 microg doses of orexin A decreased food intake at 2 h post-dose by 45% and 64%, respectively. Eight-hour food intake was decreased at only the 20 microg orexin A dose. Orexin B at all doses and time points did not alter food intake when compared to vehicle. These results indicate that orexin A exhibits anorectic activity while orexin B does not affect food intake in the rhesus monkey.     

Endogenous amylin contributes to the anorectic effects of cholecystokinin and bombesin

Previous studies indicated that amylin contributes to the anorectic effects of cholecystokinin (CCK) and bombesin (BBS), possibly by enhancing the release of pancreatic amylin or by modulating their anorectic actions within the central nervous system (CNS). To elucidate the interaction between amylin and CCK or BBS, respectively, we investigated the influence of an IP injection of CCK or BBS on feeding in amylin-deficient mice (IAPP(-/-)). The anorectic effects of CCK and BBS were nearly abolished in IAPP(-/-) mice compared to wildtype (WT) mice (e.g. 20 microg/kg CCK, 1-h food intake: WT/NaCl 0.53 +/- 0.03 g; WT/CCK 0.16 +/- 0.03 g (P < 0.001); IAPP(-/-)/NaCl 0.49 +/- 0.05 g; IAPP(-/-)/CCK 0.39 +/- 0.04 g). Acute amylin replacement restored the anorectic effect of CCK in IAPP(-/-) mice.To find out whether CCK or BBS enhance the feeding-induced release of pancreatic amylin, we injected rats with CCK-8 (0.5-50 microg/kg) or BBS (5 microg/kg) and measured plasma amylin levels after injections. Neither CCK nor BBS increased the plasma amylin level in rats. We suggest that the mediation of the anorectic effects of CCK and BBS by amylin is not dependent on a CCK- or BBS-induced release of pancreatic amylin, but may rather be due to a modulation of their effects by amylin within the CNS.     

Ghrelin对大鼠摄食的影响及orexins信号通路的调控

目的:探究Ghrelin对大鼠摄食的影响及orexins信号通路的调控作用。方法:采用免疫组织化学染色的方法观察Ghrelin免疫阳性神经元轴突末梢与orexin神经元的突触联系以及下丘脑外侧区(LHA)内c-fos的表达。侧脑室注射抗-orexin-A IgG和抗-orexin-B IgG混合液、抗-黑色素浓集激素(MCH)IgG、NPY-1受体拮抗剂后测量大鼠摄食量,观察其对ghrelin诱导摄食的影响。结果:Ghrelin免疫阳性神经元轴突末梢与orexin神经元的突触相接触。侧脑室注射ghrelin可诱导orexin神经元内c-fos表达,但是没有引起MCH神经元内c-fos的表达。预先注射抗-NPY IgG抗体,ghrelin仍然可诱导orexin神经元内c-fos表达。侧脑室预先注射抗-orexin-A IgG和抗-orexin-B IgG抗体可减弱ghrelin促摄食作用,但是预先注射抗-MCH IgG抗体对ghrelin诱导的摄食作用没有明显影响。注射NPY受体拮抗剂可进一步加强抗-orexin-A IgG抗体和抗-orexin-B IgG抗体对ghrelin诱导摄食的抑制效应。结论:ghrelin可能与orexin系统相互作用共同参与摄食和能量平衡的调控。     

Alpha-melanocyte-stimulating hormone mediates melanin-concentrating hormone-induced anorexigenic action in goldfish

In goldfish, intracerebroventricular (ICV) administration of melanin-concentrating hormone (MCH) inhibits feeding behavior, and fasting decreases hypothalamic MCH-like immunoreactivity. However, while MCH acts as an anorexigenic factor in goldfish, in rodents MCH has an orexigenic effect. Therefore, we examined the involvement of two anorexigenic neuropeptides, alpha-melanocyte-stimulating hormone (alpha-MSH) and corticotropin-releasing hormone (CRH), in the anorexigenic action of MCH in goldfish, using an alpha-MSH receptor antagonist, HS024, and a CRH receptor antagonist, alpha-helical CRH((9-41)). ICV injection of HS024, but not alpha-helical CRH((9-41)), suppressed MCH-induced anorexigenic action for a 60-min observation period. We then examined, using a real-time PCR method, whether MCH affects the levels of mRNAs encoding various orexigenic neuropeptides, including neuropeptide Y (NPY), orexin, ghrelin and Agouti-related peptide (AgRP), in the goldfish diencephalon. ICV administration of MCH at a dose sufficient to inhibit food consumption decreased the expression of mRNAs for NPY and ghrelin, but not for orexin and AgRP. These results indicate that the anorexigenic action of MCH in the goldfish brain is mediated by the alpha-MSH signaling pathway and is accompanied by inhibition of NPY and ghrelin synthesis.     

CCK inhibits the orexigenic effect of peripheral ghrelin

CCK and ghrelin exert antagonistic effects on ingestive behavior. The aim of the present study was to investigate the interaction between ghrelin and CCK administered peripherally on food intake and neuronal activity in specific hypothalamic and brain stem nuclei, as assessed by c-Fos-like immunoreactivity (c-FLI) in nonfasted rats. Ghrelin (13 microg/kg body wt) injected intraperitoneally significantly increased the cumulative food intake when measured at 30 min and 1 h after injection, compared with the vehicle group (2.9 +/- 1.0 g/kg body wt vs. 1.2 +/- 0.5 g/kg body wt, P < 0.028). Sulfated CCK octapeptide (CCK-8S) (2 or 25 microg/kg body wt) injected simultaneously blocked the orexigenic effect of ghrelin (0.22 +/- 0.13 g/kg body wt, P < 0.001 and 0.33 +/- 0.23 g/kg body wt, P < 0.0008), while injected alone, both doses of CCK-8S exerted a nonsignificant trend to reduce food intake. Ghrelin (13 microg/kg body wt ip) markedly increased the number of c-FLI-positive neurons per section in the arcuate nucleus (ARC) compared with vehicle (median: 31.35 vs. 9.86, P < 0.0001). CCK-8S (2 or 25 microg/kg body wt ip) had no effect on neuronal activity in the ARC, as assessed by c-FLI (median: 5.33 and 11.21 cells per section), but blocked the ghrelin-induced increase of c-fos expression in this area when both peptides were administered simultaneously (median: 13.33 and 12.86 cells per section, respectively). Ghrelin at this dose had no effect on CCK-induced stimulation of c-fos expression in the paraventricular nucleus of the hypothalamus and the nucleus of the solitary tract. These results suggest that CCK abolishes ghrelin-induced food intake through dampening increased ARC neuronal activity.     

Triiodothyronine (T3) stimulates food intake via enhanced hypothalamic AMP-activated kinase activity

Thyroid hormone regulates food intake. We previously reported that rats with triiodothyronine (T3)-induced thyrotoxicosis display hyperphagia associated with suppressed circulating leptin levels, increased hypothalamic neuropeptide Y (NPY) mRNA and decreased hypothalamic pro-opiomelanocortin (POMC) mRNA. AMP-activated kinase (AMPK) is a serine/threonine protein kinase that is activated when cellular energy is depleted. We hypothesized that T3 causes an increase in hypothalamic AMPK activity, which in turn contributes to the development of T3-induced hyperphagia. Rats that were given s.c. injections of T3 (4.5 nmol/kg) had increased food intake 2 h later without alterations in NPY and POMC mRNA levels, but with increased hypothalamic phosphorylated AMPK (169%) and phosphorylated acetyl-CoA carboxylase (194%). To determine the more chronic effects of T3, rats were given 6 daily s.c. injection of T3 or the vehicle. Food intake was significantly increased. Multiple T3 injections increased hypothalamic phosphorylated AMPK (278%) and phosphorylated acetyl-CoA carboxylase (335%) compared to the controls. Intracerebroventricular administration of compound C, an AMPK inhibitor, blocked the food intake induced by a single or multiple injections of T3. Taken together, these results suggest that enhanced hypothalamic AMPK phosphorylation contributes to T3-induced hyperphagia. Hypothalamic AMPK plays an important role in the regulation of food intake and body weight.     

The role of cholecystokinin octapeptide in the central control of food intake in the dog

Cholecystokinin octapeptide (CCK-8, 1, 190 pmol/5 min) decreased food intake and water consumption in two models of ingestive behavior, i.e., food deprivation-induced feeding and insulin-induced feeding, when administered into the third (3V) and lateral (LV) cerebral ventricles. In fasted dogs, the suppression of food intake was more prominent after 3V CCK-8, whereas intravenously administered CCK-8 was without effect. Neuropeptide Y (NPY, 1, 190 pmol) had no significant stimulatory effect on food intake and water consumption in fasted as well as satiated dogs, and actually reduced both food and water intake in insulin-treated dogs. There was a slight but significant decrease in food and water intake after 275 nmol naloxone administration in both feeding models, and some of the dogs vomited. In insulin-treated animals, CCK-8 reversed, but NPY potentiated the hypothermic phase of temperature response observed after saline administration, whereas naloxone failed to alter rectal temperature. These results suggest that the effect of CCK-8 on feeding seems to involve central mechanisms in the dog, and that the mechanisms by which CCK-8, NPY and naloxone affect feeding behavior are different.     

CCK and 5-HT act synergistically to suppress food intake through simultaneous activation of CCK-1 and 5-HT3 receptors

Cholecystokinin (CCK) and serotonin (5-HT) systems have been shown to cooperate interdependently in control of food intake. To assess mechanisms by which CCK and 5-HT systems interact in control of food intake we examined: (1) participation of CCK-1 and 5-HT3 receptors in 5-HT-induced suppression of sucrose intake; (2) the interaction between CCK and 5-HT in suppression of food intake; (3) the role of CCK-1 and 5-HT3 receptors in mediating this interaction. Intraperitoneal administration of 5-HT (0.25, 0.5 and 1.0 mg/kg) significantly reduced intake compared to control in a dose responsive fashion (r2=0.989). Suppression of food intake by 5-HT was significantly attenuated by prior treatment with the 5-HT3 receptor antagonist ondansetron at each 5-HT dose tested (P<0.05), while blockade of CCK-1 receptors by lorglumide had no effect on 5-HT-induced suppression of intake. Administration of CCK-8 (0.5 microg/kg) or 5-HT (0.5 mg/kg) alone significantly reduced sucrose intake by 22.9 and 22.2% respectively, compared to control (P<0.0001). Co-administration of CCK and 5-HT resulted in a synergistic suppression of intake leading to an overall 48.4% reduction in sucrose intake compared to saline (P<0.0001). Concomitant CCK-1 and 5-HT3 receptor blockade by lorglumide and ondansetron respectively, resulted in a complete reversal of the combined CCK and 5-HT-induced suppression of intake. Independent administration of lorglumide or ondansetron did not alter intake compared to control. These studies provide evidence that 5-HT causes suppression in food intake by acting at 5-HT3, not CCK-1 receptors. Furthermore, CCK and 5-HT interact to produce an enhanced suppression of food intake, an effect mediated through concomitant activation of CCK-1 and 5-HT3 receptors.     

Exogeneous and endogenous CCK inhibit ethanol ingestion in Sardinian alcohol-preferring rats

Ethanol ingestion, like food ingestion, stimulates release of the signaling molecule cholecystokinin (CCK) from the small intestine. Here, we investigated the possibility that ethanol-induced CCK release might be a negative-feedback control of ethanol ingestion, similar to its function as part of the mechanism by which ingested food produces meal-ending satiation. We used Sardinian alcohol-preferring (sP) and Marchesian Sardinian (msP) alcohol-preferring rats, two apparently identical substrains that spontaneously ingest pharmacologically relevant amounts of ethanol, as well as their background strain, Wistar (W) rats. We demonstrated that: (1) intraperitoneal (IP), but not intracerebroventricular, injections of 0.5-4 microg/kg CCK-8 produced transient, dose-related reductions in 10% ethanol ingestion; (2) this inhibitory effect of CCK-8 on ethanol intake appeared behaviorally similar to its inhibitory action on ingestion of sucrose solutions; (3) the inhibitory effect of IP CCK-8 on ethanol ingestion occurred without evidence of tolerance when tests were repeated on consecutive days; (4) IP CCK-8 reduced ethanol intake despite simultaneously reducing blood ethanol levels (BALs); and (5) antagonism of CCK1 receptors with devazepide increased ethanol intake, indicating that endogenous CCK normally limits the size of bouts of ethanol ingestion. These results implicate peripheral CCK in the control of ethanol ingestion in sP and msP alcohol-preferring rats.