Role of Intraduodenally Administered Enterostatin in Rats: Inhibition of Food

Central and peripheral administration of enterostatin have been reported to reduce fat or high-fat food intake in rats. Enterostatin is formed in the intestinal lumen by tryptic cleavage of pancreatic procolipase during intraluminal fat digestion. The present experiments were designed to test if enterostatin following intraintestinal infusion would affect food intake in a similar way as intracerebraventricularly or intravenously administered enterostatin. Female Sprague-Dawley rats were fitted with a duodenal catheter and adapted to a feeding schedule for 6 hours each day. After 10 days enterostatin (5.65 and 11.3 nmol/kg/min) or saline were infused into the duodenum and food intake measured. Enterostatin significantly reduced high-fat food intake during the 6 hours of feeding, but had no inhibitory effect on low-fat food intake. Addition of tetracaine to the enterostatin infusates blocked the satiating potency of intestinal enterostatin. These results support the hypothesis of a preabsorptive site of action for enterostatin.     

Effects of Enterostatin on Consumption of Optional Foods in Non-Food-Deprived Rats

Enterostatin, the activation peptide of procolipase, has been reported to reduce high-fat food consumption in rats. This reduction has been reliably demonstrated using procedures in which the test diet was also the maintenance diet of the animals. Other reports, though, have shown that peripherally administered enterostatin had no effect on the consumption of oil provided as an option to the diet, and that centrally administered enterostatin had no effect on the consumption of an optional high-fat mixed food. However, the effects of peripherally administered enterostatin on the consumption of an optional high-fat mixed food have not been examined. This experiment, then, examined the effects of peripherally administered enterostatin on the consumption of optional, mixed foods (no-fat and high-fat cookies) provided in addition to a standard diet under choice and nonchoice conditions. Four experiments were conducted. In experiment I, the effect of enterostatin in a two-choice feeding paradigm was assessed. In experiment II, the effect of enterostatin in a nonchoice feeding paradigm was assessed. In experiment III, the effect of enterostatin administered at five different pretreatment times in a non-choice feeding paradigm was assessed. Enterostatin had no effect on cookie intake in any of these experiments. Finally, experiment IV was undertaken to verify the activity of the peptide. Enterostatin significantly reduced the consumption of a standard diet in overnight food-deprived rats, thus confirming the activity of the peptide used in experiments I to III. Enterostatin may not play a major role in the regulation of food intake that is stimulated by optional foods that are periodically provided in addition to a standard well-balanced diet.     

Different metabolic responses to central and peripheral injection of enterostatin

Enterostatin, a pentapeptide cleaved from procolipase, suppresses fat intake after peripheral and central administration. Chronic treatment of rats with enterostatin decreases body weight and body fat. The effect was greater than could be accounted by the reduction in food intake alone. Hence, we have investigated the effect of enterostatin on energy metabolism. Male Sprague-Dawley rats adapted to a high-fat diet were implanted with lateral cerebral ventricular or amygdala cannulas. The metabolic effects were determined by indirect calorimetry. After habituation to the test cages, fasted rats were injected with either saline vehicle or enterostatin given either intraperitoneally (100 nmol) or intracerebroventricularly (1 nmol) or into specific brain regions [amygdala (0.01 nmol) or paraventricular nucleus (PVN) (0.1 nmol)]. Respiratory quotient (RQ) and energy expenditure were monitored over 2 h. Intraperitoneal enterostatin reduced RQ (saline: 0.81 +/- 0.02 vs. enterostatin: 0.76 +/- 0.01) and increased energy expenditure by 44%. Intracerebroventricular enterostatin increased the energy expenditure without any effects on RQ, whereas PVN enterostatin increased metabolic rate, while preventing the increase in RQ observed in the control animals. In contrast, neither RQ nor energy expenditure was altered after enterostatin was injected into the amygdala. Enterostatin activated AMP-activated protein kinase in primary cultures of human myocytes in a dose- and time-dependent manner and increased the rate of fatty acid beta-oxidation. These findings suggest that enterostatin regulates energy expenditure and substrate partitioning through both peripheral and central effects.     

Enterostatin inhibition of dietary fat intake is dependent on CCK-A receptors

Enterostatin, a pentapeptide released from the exocrine pancreas and gastrointestinal tract, selectively inhibits fat intake through activation of an afferent vagal signaling pathway. This study investigated if the effects of enterostatin were mediated through a CCK-dependent pathway. The series of in vivo and in vitro experiments included studies of 1) the feeding effect of peripheral enterostatin on Otsuka Long Evans Tokushima Fatty (OLETF) rats lacking CCK-A receptors, 2) the effect of CCK-8S on the intake of a two-choice high-fat (HF)/low-fat (LF) diet, 3) the effects of peripheral or central injection of the CCK-A receptor antagonist lorglumide on the feeding inhibition induced by either central or peripheral enterostatin, and 4) the ability of enterostatin to displace CCK binding in a 3T3 cell line expressing CCK-A receptor gene and in rat brain sections. The results showed that OLTEF rats did not respond to enterostatin (300 microg/kg ip) in contrast to the 23% reduction in intake of HF diet in Long Evans Tokushima Otsuka (LETO) control rats. CCK (1 microg/kg ip) decreased the intake of the HF diet in a two-choice diet regime with a compensatory increase in intake of the LF diet. Peripheral injection of lorglumide (300 microg/kg) blocked the feeding inhibition induced by either near-celiac arterial or intracerebroventricular enterostatin, whereas intracerebroventricular lorglumide (5 nmol icv) only blocked the response to intracerebroventricular enterostatin but not to arterial enterostatin. Enterostatin did not bind on CCK-A receptors because neither enterostatin nor its analogs VPDPR and beta-casomorphin displaced [3H]L-364,718 from CCK-A receptors expressed in 3T3 cells or the binding of 125I-CCK-8S from rat brain sections. The data suggest that both the peripheral and central responses to enterostatin are mediated through or dependent on peripheral and central CCK-A receptors.     

Neuropeptide Y receptor(s) mediating feeding in the rat: characterization with antagonists

The present study evaluated the effect of the neuropeptide Y (NPY) Y1 receptor antagonists BIBO 3304 and SR 120562A and of the Y5 receptor antagonists JCF 104, JCF 109, and CGP 71683A on feeding induced either by NPY or food deprivation. In a preliminary experiment, NPY was injected into the third cerebroventricle (3V) at doses of 0.07, 0.15, 0.3, or 0.6 nmol/rat. The dose of 0.3 nmol/rat, which produced a cumulative 2-h food intake of 11.2 +/- 1.9 g/kg body weight, was chosen for the following experiments. The antagonists were injected in the 3V 1 min before NPY. The Y1 receptor antagonist BIBO 3304 significantly inhibited NPY-induced feeding at doses of 1 or 10 nmol/rat. The Y1 receptor antagonist SR 120562A, at the dose of 10 but not of 1 nmol/rat, significantly reduced the hyperphagic effect of NPY, 0.3 nmol/rat. The Y5 receptor antagonists JCF 104 and JCF 109 (1 or 10 nmol/rat) and CGP 71683A (10 or 100 nmol/rat) did not significantly modify the effect of NPY, 0.3 nmol/rat. However, JCF 104 (10 nmol/rat) and CGP 71683A (100 nmol/rat), but not JCF 109 (10 nmol/rat), significantly reduced food intake during the interval from 2 to 4 h after injection of a higher dose, 0.6 nmol/rat, of NPY. Feeding induced by 16 h of food deprivation was significantly reduced by the Y1 receptor antagonist BIBO 3304 (10 nmol/rat), but it was not significantly modified by the same dose of SR 120562A or JCF 104. These findings support the idea that the hyperphagic effect of NPY is mainly mediated by Y1 receptors. The results obtained with JCF 104 and CGP 71683A suggest that Y5 receptors may have a modulatory role in the maintenance of feeding induced by rather high doses of NPY after the main initial feeding response.     

Enterostatin suppresses food intake in rats after near-celiac and intracarotid arterial injection

Enterostatin (Ent) selectively suppresses the intake of dietary fat after peripheral and central administration. To further investigate the site of action of Ent, we compared the feeding responses to Ent injected intra-arterially near the celiac artery, into the carotid artery, or intravenously in rats adapted to a high-fat diet. After near-celiac arterial injection there was an immediate dose-dependent (0.05-13.5 nmol) inhibition of food intake occurring within 5 min in overnight-fasted rats that lasted up to 20 min. Carotid arterial Ent had a similar, immediate dose-related response, and the inhibitory effect was long lasting. The response to intravenous Ent was only evident at the highest dose (13.5 nmol) and was delayed for at least 120 min. Pretreatment with capsaicin, which causes degeneration of vagal sensory neurons, abolished the inhibitory responses to near-celiac Ent but not to intravenous or intracarotid Ent. These results provide further evidence for both a gastrointestinal site of action for peripheral Ent and a central site of action for intracarotid Ent and suggest that the delayed response to intravenous Ent may reflect either binding or slow uptake of this peptide into the central nervous system.     

Enterostatin decreases postprandial pancreatic UCP2 mRNA levels and increases plasma insulin and amylin

This study investigated the chronic effect of enterostatin on body weight and some of the associated changes in postprandial metabolism. Rats were adapted to 6 h of food access/day and a choice of low-fat and high-fat (HF) food and then given enterostatin or vehicle by an intraperitoneally implanted minipump delivering 160 nmol enterostatin/h continuously over a 5-day infusion period. Enterostatin resulted in a slight but significant reduction of HF intake and body weight. After the last 6-h food access period, enterostatin-treated animals had lower plasma triglyceride and free fatty acid but higher plasma glucose and lactate levels than control animals. Enterostatin infusion resulted in increased uncoupling protein-2 (UCP2) expression in various tissues, including epididymal fat and liver. UCP2 was reduced in the pancreas of enterostatin-treated animals, and this was associated with increased plasma levels of insulin and amylin. Whether these two hormones are involved in the observed decreased food intake due to enterostatin remains to be determined. As lipid metabolism appeared to be altered by enterostatin, we measured peroxisome proliferator-activated receptor (PPAR) expression in tissues and observed that PPARalpha, -beta, -gamma1, and -gamma2 expression were modified by enterostatin in epididymal fat, pancreas, and liver. This further links altered lipid metabolism with body weight loss. Our data suggest that alterations in UCP2 and PPARgamma2 play a role in the control of insulin and amylin release from the pancreas. This implies that enterostatin changes lipid and carbohydrate metabolic pathways in addition to its effects on food intake and energy expenditure.     

Effect of Enterostatin on the Feeding Responses to Galanin and NPY

We have investigated the possibility that enterostatin may inhibit the intake of dietary fat by inhibiting either galanin or NPY-induced feeding pathways. Rats, adapted to either high fat (HF) or low fat-high carbohydrate (HC) diets and fitted with third ventricular cannulas were used to study the effects of intracerebroventricular (icv) enterostatin on icv NPY and galanin induced feeding responses in satiated rats. An equimolar dose of enterostatin (0.1nmoles) inhibited, while a tenfold excess of entersotatin abolished the feeding response to galanin in rats adapted to a HF diet. The galanin stimulation of food intake was reduced in rats adapted to the HC diet and this response was less sensitive to inhibition by enterostatin. Enterostatin had no inhibitory effects on NPY-induced feeding in rats adapted to the HC diet and only a small inhibitory effect, at tenfold molar excess, in rats adapted to the HF diet. The ability of enterostatin to bind to galanin or NPY Y-1 receptors was investigated in lig and binding studies. Enterstatin fialed to dispace ¹²⁵I-galanin or ¹²⁵I-PYY from specific binding sites in rat forebrain homogenates or SK-N-MC cells respectively. The data provide support for the hypothesis that enterostatin specifically inhibits a galanin-responsive fat intake system, but indicate that this effect is not modulated by direct interaction with either galanin or NPY-Y1 receptors.     

Injection of neuropeptide W into paraventricular nucleus of hypothalamus increases food intake

Neuropeptide W (NPW) is an endogenous ligand for G protein-coupled receptor 7 (GPR7). There are two forms of the peptide, designated as neuropeptide W-23 (NPW23) and neuropeptide W-30 (NPW30). In the current study we found that intracerebroventricular administration of NPW23 increased c-Fos immunoreactivity (IR) in a variety of brain sites, many of which are involved in the regulation of feeding. In particular, we noted that c-Fos IR levels were increased in hypocretin-expressing neurons in the perifornical region of the lateral hypothalamus (LH). We then studied whether injection of NPW23 into the paraventricular nucleus of the hypothalamus (PVN) and the LH increased food intake over a 24-h time period. Intra-PVN injection of NPW23 at doses ranging from 0.1 to 3 nmol increased feeding for up to 4 h, and doses ranging from 0.3 to 3 nmol increased feeding for up to 24 h. In contrast, only the 3-nmol dose of NPW23 increased feeding after administration into the LH. Together, these data suggest a modulatory role for NPW in the control of food intake.     

Intragastric beta-casomorphin(1-7) attenuates the suppression of fat intake by enterostatin

The current experiments were designed to compare the feeding response to enterostatin and beta-casomorphin(1-7) injected intragastrically. Sprague-Dawley rats with a gastric cannula were allowed to chose from high-fat diet (HF) or low-fat diet (LF) in separate jars. Enterostatin injected intragastrically into overnight fasted rats caused a U-shaped dose-dependent reduction in the intake of the HF diet for the first two hours after infusion but had no effect on the LF intake. beta-Casomorphin(1-7) stimulated the intake of the HF diet but had no effect on the LF diet. Finally, beta-casomorphin(1-7) blocked the inhibitory effect of enterostatin on HF intake in fasted rats.     

Plasma Insulin in Response to Enterostatin and Effect of Adrenalectomy in Rats

Enterostatin has previously been reported to alter serum insulin and corticosterone levels after central administration of the peptide. The purpose of the present study was to investigate the effect of peripheral administration of enterostatin on insulin and corticosterone levels as well as the response of plasma insulin to enterostatin administration in adrena-lectomized rats. Female Sprague-Dawley rats were given a bolus injection intravenously with enterostatin alone or together with glucose. Enterostatin increased basal plasma levels of insulin, but significantly inhibited the increase in plasma insulin stimulated by glucose. Plasma corticosterone levels were not altered after a single intravenous injection of enterostatin. In rats infused chronically with enterostatin, plasma insulin levels were significantly reduced and plasma corticosterone levels were increased. The daily food intake was lower in these rats, but there was no effect on body weight. After adrenalectomy, the responsiveness of plasma insulin to enterostatin infusion was completely abolished. Furthermore, adrenalectomy itself reduced basal plasma levels of insulin and increased plasma levels of endogenous enterostatin. These results suggest that peripheral enterostatin administration produces a similar effect as central infusion of the peptide, and that the glucocorticoid hormones are involved in the regulation of plasma insulin by enterostatin.     

Plasma Enterostatin: Identification and Release in Rats in Response to a Meal

Objective: To discover a possible absorption and/or secretion of enterostatin into the circulating blood, as well as to compare the levels of circulating enterostatin after high‐fat feeding and low‐fat feeding. Research Methods and Procedures: Using a specific enzyme‐linked immunosorbent assay, plasma enterostatin levels were determined after feeding a high‐fat, a high‐fat/‐sucrose, or a low‐fat meal to Sprague‐Dawley rats deprived of food overnight. Results: The enterostatin levels were increased by all diets; the response to the high‐fat and the high‐fat/‐sucrose meals was greater in magnitude and duration than that to the low‐fat meal. In addition, enterostatin levels correlated with the intake of dietary fat. Plasma enterostatin levels after high‐fat feeding were found to be similar to those after intravenous administration of exogenous enterostatin known to inhibit high‐fat food intake. Gel chromatography of pooled postprandial plasma extracts followed by high‐performance liquid chromatography analysis showed that plasma enterostatin was identical to synthetic enterostatin. Affinity cross‐linking of plasma proteins with ¹²⁵I‐enterostatin on sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, followed by autoradiography, revealed a single band with a molecular weight of about 66 kDa, indicating the presence of a potential enterostatin‐binding protein in plasma. Discussion: The measurements of plasma enterostatin may be a sensitive indicator for the measurement of fat intake.     

Intravenous infusion of glucagon-like peptide-1 potently inhibits food intake, sham feeding, and gastric emptying in rats

Glucagon-like peptide-1(7-36)-amide (GLP-1) is postulated to act as a hormonal signal from gut to brain to inhibit food intake and gastric emptying. A mixed-nutrient meal produces a 2 to 3-h increase in plasma GLP-1. We determined the effects of intravenous infusions of GLP-1 on food intake, sham feeding, and gastric emptying in rats to assess whether GLP-1 inhibits food intake, in part, by slowing gastric emptying. A 3-h intravenous infusion of GLP-1 (0.5-170 pmol.kg(-1).min(-1)) at dark onset dose-dependently inhibited food intake in rats that were normally fed with a potency (mean effective dose) and efficacy (maximal % inhibition) of 23 pmol.kg(-1).min(-1) and 82%, respectively. Similar total doses of GLP-1 administered over a 15-min period were less potent and effective. In gastric emptying experiments, GLP-1 (1.7-50 pmol.kg(-1).min(-1)) dose-dependently inhibited gastric emptying of saline and ingested chow with potencies of 18 and 6 pmol.kg(-1).min(-1) and maximal inhibitions of 74 and 83%, respectively. In sham-feeding experiments, GLP-1 (5-50 pmol.kg(-1).min(-1)) dose-dependently reduced 15% aqueous sucrose intake in a similar manner when gastric cannulas were closed (real feeding) and open (sham feeding). These results demonstrate that intravenous infusions of GLP-1 dose-dependently inhibit food intake, sham feeding, and gastric emptying with a similar potency and efficacy. Thus GLP-1 may inhibit food intake in part by reducing gastric emptying, yet can also inhibit food intake independently of its action to reduce gastric emptying. It remains to be determined whether intravenous doses of GLP-1 that reproduce postprandial increases in plasma GLP-1 are sufficient to inhibit food intake and gastric emptying.     

The CCKB antagonist CI988 reduces food intake in fasted rats via a dopamine mediated pathway

Studies have shown a reduction of food intake following peripheral and brain injection of CCK. However, it remains to be established whether endogenous central CCK is involved in the regulation of food intake. We investigated the role of central CCK in the regulation of food intake by pharmacological manipulation of the CCK_B (CCK₂) receptor system. Intracerebroventricularly (ICV) cannulated male Sprague Dawley rats were fasted for 24 h and received an ICV injection of the CCK_B receptor antagonist CI988 at a dose of 10 nmol or 49 nmol or vehicle. Another group received two consecutive ICV injections consisting of the corticotropin-releasing factor (CRF) receptor-1 (CRF₁) antagonist, CP376395 (3 nmol) or the CRF₂ receptor antagonist, K41498 (2 nmol) alone, or followed by CI988 (49 nmol). Lastly, another group of rats received an intraperitoneal (IP) injection of the dopamine antagonist, flupentixol (∼197 and ∼493 nmol/kg) alone, or followed by CI988 (49 nmol, ICV). Cumulative food intake was assessed for 11 h. Vehicle injected rats showed a robust feeding response. CI988 at 49 nmol reduced food intake by 30% starting at 2 h post injection. CP376395 and K41498 had no effect on food intake. Flupentixol injected IP at a dose of 197 and 493 nmol/kg alone did not modulate food intake whereas the higher dose blocked the CI988-induced reduction of feeding. During the dark phase, CI988 had no effect on food intake in unfasted rats. In summary, CCK_B signaling is involved in the regulation of food intake after a fast likely by downstream dopamine signaling.     

Intracerebroventricular injections of cholecystokinin octapeptide suppress feeding in rats--pharmacological characterization of this action

Cholecystokinin octapeptide (CCK-8), administered intracerebroventricularly (i.c.v.), will suppress feeding. The aim of the present study was to determine the pharmacological characteristics of this satiety inducing effect in rats. For this purpose, we employed a feeding bioassay model in 24 h fasted rats and examined the effects of CCK-8 and a variety of structurally related analogs on latency to feed after i.c.v. injection and on the amount of food and water consumed as measured after the initiation of feeding in sequential 20-min epochs for 1 h. CCK-8, given in doses of 0.1, 1 and 10 nmol, produced a dose-dependent increase in feeding latency and a reduction of food intake during the first 20 min after initiation of feeding. Food intake during the next 40 min and water consumption were not altered. Plasma levels of CCK-like immunoreactivity after an i.c.v. injection of a dose of CCK-8 which blocked feeding (10 nmol) rose insignificantly from 117 to 125 pg/ml. In contrast, at the minimally effective dose of CCK-8 after i.v. administration (10 nmol), which also produced an inhibition of feeding, the plasma level was 1430 pg/ml. This difference indicates that plasma levels of CCK after i.c.v. CCK-8 are not adequate to produce the observed feeding suppression and suggests that the effects of i.c.v. CCK-8 are not mediated by a peripheral redistribution. Systematic dose response studies revealed the following rank order of potencies: CCK-8 greater than or equal to G-17 II much greater than CCK-8 NS = G-17 I greater than or equal to CCK-4 = CCK 26-29 = 0. Only gastrin-17 II (sulfated) produced an effect comparably significant to CCK-8. I.c.v. proglumide at 2500 nmol failed to modify the effects of CCK-8 at 10 nmol after i.c.v. injection. These data demonstrate that the structural requirements for feeding suppressive activity in rat brain are the carboxyterminus with a sulfated tyrosine residue, located 6 to 7 residues from the carboxyterminus, as present in CCK-8 and gastrin-17 II.     

Dose-dependent effects of peptide YY(3-36) on conditioned taste aversion in rats

We used a conditioned taste aversion test to assess whether PYY(3-36) reduces food intake by producing malaise. Two-hour IV infusion of PYY(3-36) (8, 15, and 30 pmol/kg/min) at dark onset in non-food-deprived rats produced a dose-dependent inhibition of feeding and a conditioned aversion to the flavored chow paired with PYY(3-36) infusion. In food-deprived rats, PYY(3-36) at 2 and 4 pmol/kg/min inhibited intake of a flavored saccharin solution without producing conditioned taste aversion, whereas higher doses (8 and 15 pmol/kg/min) inhibited saccharin intake and produced taste aversion. These results suggest that anorexic doses of PYY(3-36) may produce a dose-dependent malaise in rats, which is similar to that reported for PYY(3-36) infusion in humans. Previous studies have shown that PYY(3-36) potently inhibits gastric emptying, and that gut distention can produce a conditioned taste aversion. Thus, PYY(3-36) may produce conditioned taste aversion in part by slowing gastric emptying.     

Mouse pancreatic polypeptide modulates food intake, while not influencing anxiety in mice

This study was designed to investigate the effects of synthetic mouse pancreatic polypeptide (mPP) on feeding and anxiety in mice. The intracerebroventricular (ICV) injection of mPP (0.003–3 nmol) dose-dependently increased food intake. A significant increase was observed 20 min after ICV injection and continued for 4 h. The intraperitoneal (IP) injection of mPP (0.03–30 nmol) dose-dependently decreased food intake. A significant decrease was observed 20 min after IP injection and continued for 4 h. In the elevated plus maze test, the ICV injection of mPP (0.003–3 nmol) did not affect anxiety behavior. These results suggest that mPP modulates food intake and the Y4 receptor in the brain may contribute to the regulation of feeding, whereas appearing not to influence anxiety in mice.     

Differential inhibition of galanin- and ghrelin-induced food intake by i.c.v. GLP-1(7-36)-amide

Feeding regulation involves both anorectic and orexigenic neuropepetides mainly located in the hypothalamus. To gain further insight into the interaction between these two groups of regulators inhibition of feeding induced by glucagon-like peptide-1 (GLP-1) was examined during stimulation of food intake by equimolar doses of ghrelin and galanin. The experiments were carried out in freely feeding rats. Intracerebroventricular (i.c.v.) injections were accomplished through stereotaxically implanted cannulae aimed at the lateral cerebral ventricle. Food intake of standard rat chow pellets was subsequently recorded for 2 h. Ghrelin and galanin stimulated food intake significantly with no difference between these two peptides. During ghrelin stimulation GLP-1 inhibited feeding in doses between 0.015 and 1.5 nmol. During galanin stimulation of food intake a ten fold higher dose (0.15 nmol) was required to inhibit food intake. In conclusion equimolar doses of i.c.v. ghrelin and galanin are similarly effective stimuli of food intake when given alone. However in combination with an anorectic neuropeptide such as GLP-1 they have substantially different potencies of feeding stimulation. Such interaction could also be of interest for therapeutic strategies involving both regulating groups of neuropeptides.     

Mouse pancreatic polypeptide modulates food intake, while not influencing anxiety in mice

This study was designed to investigate the effects of synthetic mouse pancreatic polypeptide (mPP) on feeding and anxiety in mice. The intracerebroventricular (i.c.v.) injection of mPP (0.003-3 nmol) dose-dependently increased food intake. A significant increase was observed 20 min after i.c.v. injection and continued for 4 h. The intraperitoneal (i.p.) injection of mPP (0.03-30 nmol) dose-dependently decreased food intake. A significant decrease was observed 20 min after i.p. injection and continued for 4 h. In the elevated plus maze test, the i.c.v. injection of mPP (0.003-3 nmol) did not affect anxiety behavior. These results suggest that mPP modulates food intake and the Y4 receptor in the brain may contribute to the regulation of feeding, whereas appearing not to influence anxiety in mice.