Feeding after fourth ventricular administration of neuropeptide Y receptor agonists in rats

Neuropeptide Y (NPY) and peptide YY (PYY) stimulate food intake after injection into the fourth cerebral ventricle, suggesting that NPY receptors in the hindbrain are targets for the stimulatory effect of these peptides on food intake. However, the NPY/PYY receptor subtype mediating the feeding response in the hindbrain is not known. To approach to this question we compared dose-effect of several NPY receptor agonists to stimulate food intake in freely-feeding rats 60- and 120-min after injection into the fourth cerebral ventricle. At the 120-min time point, PYY was 2- to 10-times as potent as NPY over the dose-response range and stimulated twice the total intake at the maximally effective dose (2-fold greater efficacy). NPY was 2-times as potent as the Y1, Y5 receptor agonist, [Leu(31)Pro(34)]NPY but acted with comparable efficacy. The Y5-, Y2-differentiating receptor agonist, NPY 2-36, was comparable in potency to PYY at low doses but equal in efficacy NPY and [Leu(31)Pro(34)]NPY. The Y2 receptor agonist, NPY 13-36, produced only a marginal effect on total food intake. The profile of agonist potency after fourth cerebral ventricle administration is similar to the profile obtained when these or related agonists are injected in the region of the hypothalamus. Agonists at both Y1 and Y5 receptors stimulated food intake with a rank order of potency that does not conclusively favor the exclusive involvement of a single known NPY receptor subtype. Thus it is possible that the ingestive effects of NPY and PYY are mediated by multiple or novel receptor subtypes in the hindbrain. And the relatively greater potency and efficacy of PYY raises the possibility that a novel PYY-preferring receptor in the hindbrain is involved in the stimulation of food intake.     

Emerging functions of neuropeptide Y Y(2) receptors in the brain

The Y(2) receptor is the predominant neuropeptide Y (NPY) receptor subtype in the brain. Y(2) receptor mRNA is discretely distributed in the brain, including specific subregions of the hippocampus and the hypothalamus, and is largely consistent with the distribution of Y(2) receptor protein demonstrated by radioligand-binding methods. Y(2) receptor-mediated effects have been reported principally based on the observations using the C-terminal fragments of NPY. Recent studies indicate an involvement of the receptor in food intake, gastrointestinal motility, cardiovascular regulation, and neuronal excitability. Very recently, Y(2) receptor selective antagonist has been developed and Y(2) receptor-deficient animals have been created. These new pharmacological tools will help to clarify the roles of this receptor in brain functions.     

Neuropeptide Y chronically injected into the hypothalamus: A powerful neurochemical inducer of hyperphagia and obesity

Neuropeptide Y (NPY), a putative neurotransmitter abundant in the brain, has recently been shown to act within the hypothalamus, inducing a powerful eating response and a specific appetite for carbohydrates. In the present study, NPY (235 pmol) injected bilaterally in the paraventricular nucleus three times a day for 10 days caused approximately a two-fold increase in daily food intake, a six-fold increase in the rate of body weight gain and a three-fold increase in the body fat of female rats. Subsequently, the food intake and body weight of these subjects decreased precipitously, reaching control levels 20 days postinjection. These findings, demonstrating that exogenous NPY is capable of overriding mechanisms of satiety and body weight control, suggest that disturbances in NPY function may play a role in some disorders of eating behavior and body weight regulation.     

Y1 and Y5 receptors are both required for the regulation of food intake and energy homeostasis in mice

Neuropeptide Y (NPY) acting in the hypothalamus is one of the most powerful orexigenic agents known. Of the five known Y receptors, hypothalamic Y1 and Y5 have been most strongly implicated in mediating hyperphagic effects. However, knockout of individual Y1 or Y5 receptors induces late-onset obesity--and Y5 receptor knockout also induces hyperphagia, possibly due to redundancy in functions of these genes. Here we show that food intake in mice requires the combined actions of both Y1 and Y5 receptors. Germline Y1Y5 ablation in Y1Y5(-/-) mice results in hypophagia, an effect that is at least partially mediated by the hypothalamus, since mice with adult-onset Y1Y5 receptor dual ablation targeted to the paraventricular nucleus (PVN) of the hypothalamus (Y1Y5(Hyp/Hyp)) also exhibit reduced spontaneous or fasting-induced food intake when fed a high fat diet. Interestingly, despite hypophagia, mice with germline or hypothalamus-specific Y1Y5 deficiency exhibited increased body weight and/or increased adiposity, possibly due to compensatory responses to gene deletion, such as the decreased energy expenditure observed in male Y1Y5(-/-) animals relative to wildtype values. While Y1 and Y5 receptors expressed in other hypothalamic areas besides the PVN--such as the dorsomedial nucleus and the ventromedial hypothalamus--cannot be excluded from having a role in the regulation of food intake, these studies demonstrate the pivotal, combined role of both Y1 and Y5 receptors in the mediation of food intake.     

Energy Metabolic Profile of Mice after Chronic Activation of Central NPY Y1, Y2, or Y5 Receptors**

Objective: Neuropeptide Y (NPY), a 36‐amino acid peptide with orexigenic properties, is expressed abundantly in the central nervous system and binds to several NPY receptor subtypes. This study examines the roles of the NPY Y1, Y2, and Y5 receptor(s) in energy homeostasis. Research Methods and Procedures: We administered intracerebroventricular NPY (3 μg/d) or selective peptide agonists for the Y1, Y2, and Y5 receptor subtypes to C57Bl/6 mice for 6 days by mini‐osmotic pumps to assess the role of each receptor subtype in NPY‐induced obesity. Energy expenditure (EE) and respiratory quotient (RQ) were studied using indirect calorimetry. Adiposity was measured by DXA scanning and fat pad dissection. Insulin sensitivity was tested by whole‐blood glucose measurement after an insulin challenge. Results: Central administration of the selective Y1 agonist, Y5 agonist, or NPY for 6 days in mice significantly increased body weight, adiposity, and RQ, with significant hyperphagia in the Y5 agonist‐ and NPY‐treated groups but not in the Y1 agonist‐treated group. The NPY, Y1, or Y5 agonist‐treated mice had little change in total EE during ad libitum and pair‐feeding conditions. Conversely, selective activation of the Y2 receptor reduced feeding and resulted in a significant, but transient, weight loss. Discussion: Central activation of both Y1 and Y5 receptors increases RQ and adiposity, whereas only Y5 receptor activation reduces energy expended per energy ingested. Selective activation of Y2 autoreceptors leads to hypophagia and transient weight loss, with little effect on total EE. Our study indicates that all three NPY receptor subtypes may play a role in regulating energy homeostasis in mice.     

Effects of neuropeptide Y on food intake and brain biogenic amines in the rainbow trout (Oncorhynchus mykiss)

Neuropeptide Y (NPY) is one of the most potent stimulants of food intake in mammals, but very little is known about NPY actions in fish. The present study investigated the role of NPY in food intake in the rainbow trout (Oncorhynchus mykiss). Food intake was monitored at different times after intracerebroventricular administration of porcine NPY (4 or 8 microg). Both doses significantly increased food intake at 2 and 3 h, and this effect was dose-dependent. However, 50 h after administration of NPY, food intake was significantly lower than in control fish, and cumulative food intake had returned to levels similar to those seen in the control group. The NPY antagonist (D-Tyr27,36, D-Thr32)-NPY (10 microg) inhibited food intake 2 h after icv administration, but did not block the orexigenic effect of NPY when administered jointly with 4 microg NPY. To identify the NPY receptor subtypes involved in the effects of NPY on food intake, we studied the effects of the Y1 receptor agonist (Leu31, Pro34)-NPY (4 microg), the Y2 receptor agonist NPY(3-36) (4 microg), and the highly specific Y5 receptor agonist (cPP(1-7), NPY19-23, Ala31, Aib32, Gln34)-hPP (4 microg). Short-term (2 h) food intake was moderately stimulated by the Y1 agonist, more strongly stimulated by the Y2 agonist, and unaffected by the Y5 agonist. We found that administration of NPY (8 microg icv) had no effect on aminergic systems in several brain regions 2 and 50 h after NPY administration. These results indicate that NPY stimulates feeding in the rainbow trout, and suggest that this effect is cooperatively mediated by Y2- and Y1-like NPY receptors, not by Y5-like receptors.     

NPY Neuron-Specific Y2 Receptors Regulate Adipose Tissue and Trabecular Bone but Not Cortical Bone Homeostasis in Mice

Background

Y2 receptor signalling is known to be important in neuropeptide Y (NPY)-mediated effects on energy homeostasis and bone physiology. Y2 receptors are located post-synaptically as well as acting as auto receptors on NPY-expressing neurons, and the different roles of these two populations of Y2 receptors in the regulation of energy homeostasis and body composition are unclear.

Methodology/Principal Findings

We thus generated two conditional knockout mouse models, Y2^lox/lox and NPYCre/+;Y2^lox/lox, in which Y2 receptors can be selectively ablated either in the hypothalamus or specifically in hypothalamic NPY-producing neurons of adult mice. Specific deletion of hypothalamic Y2 receptors increases food intake and body weight compared to controls. Importantly, specific ablation of hypothalamic Y2 receptors on NPY-containing neurons results in a significantly greater adiposity in female but not male mice, accompanied by increased hepatic triglyceride levels, decreased expression of liver cartinine palmitoyltransferase (CPT1) and increased expression of muscle phosphorylated acetyl-CoA carboxylase (ACC). While food intake, body weight, femur length, bone mineral content, density and cortical bone volume and thickness are not significantly altered, trabecular bone volume and number were significantly increased by hypothalamic Y2 deletion on NPY-expressing neurons. Interestingly, in situ hybridisation reveals increased NPY and decreased proopiomelanocortin (POMC) mRNA expression in the arcuate nucleus of mice with hypothalamus-specific deletion of Y2 receptors in NPY neurons, consistent with a negative feedback mechanism between NPY expression and Y2 receptors on NPY-ergic neurons.

Conclusions/Significance

Taken together these data demonstrate the anti-obesogenic role of Y2 receptors in the brain, notably on NPY-ergic neurons, possibly via inhibition of NPY neurons and concomitant stimulation of POMC-expressing neurons in the arcuate nucleus of the hypothalamus, reducing lipogenic pathways in liver and/or skeletal muscle in females. These data also reveal as an anti-osteogenic effect of Y2 receptors on hypothalamic NPY-expressing neurons on trabecular but not on cortical bone.     

Attenuation of hypercholesterolemia and hyperglycemia in ob/ob mice by NPY Y2 receptor ablation

Neuropeptide Y (NPY) is a 36 amino acid peptide well known for its role in regulating food intake and energy homeostasis. It has previously been shown that the NPY Y2 receptor is required for a full biological response to leptin in the central nervous system. We have examined the impact of this receptor on plasma levels of lipid and cholesterol in wild type and obese (ob/ob) mice. The results show that an absence of Y2 in female mice has no effect on cholesterol level in normal lean mice but profoundly decreases serum cholesterol and glucose levels in ob/ob mice. We conclude that NPY, interacting with the Y2 receptor, participates in cholesterol and glucose homeostasis of obese mice.     

Role of dorsomedial hypothalamic neuropeptide Y in energy homeostasis

Hypothalamic neuropeptide Y (NPY) is primarily expressed in the arcuate nucleus (Arc) and the dorsomedial hypothalamus (DMH). Although Arc NPY gene expression is responsive to circulating leptin, the regulation of DMH NPY expression is leptin-independent. DMH NPY expression is increased in response to chronic food restriction, but not acute food deprivation. DMH NPY expression is elevated in pair-fed OLETF rats lacking cholecystokinin (CCK)-1 receptors. A role for CCK in controlling DMH NPY expression is demonstrated by the down-regulation of DMH NPY by parenchymal DMH CCK administration in intact rats. Moreover, access to running wheels normalizes body weight and prevents altered DMH NPY expression of OLETF rats. Together, these data suggest that DMH NPY plays an important role in feeding and body weight control.     

Implantation of fibre encapsulated RIN 1056a cells transfected with NPY cDNA into the lateral ventricle of rats alters body weight

Neuropeptide Y (NPY) is a hypothalamic neuropeptide thought to play an important role in the regulation of food intake and energy expenditure. Our aim was to over-express bioactive NPY in the lateral ventricle by implanting cells transfected with NPY cDNA. Cells from the RIN 1056a clonal rat islet cell line were transfected with NPY cDNA. Radioimmunoassay, chromatography and receptor binding assays were used to ensure the secreted NPY was bioactive, before and after implantation. NPY cDNA transfected and untransfected control cells were encapsulated in PVDF hollow fibres to prevent tumour formation and implanted into the lateral ventricle of male Wistar rats. The effects on body weight and food intake were measured for 15 days. Animals implanted with NPY cDNA transfected RIN 1056a cells showed a greater rise in body weight than controls. This difference was statistically significant five days after implantation, and remained so until the end of the experiment. Cumulative food intake was also increased in rats implanted with NPY cDNA transfected RIN 1056a cells, but this difference failed to reach statistical significance. We have demonstrated that implantation of NPY over-expressing cells into the lateral hypothalamus of rats increases body weight gain.     

Central prostaglandin D(2) stimulates food intake via the neuropeptide Y system in mice

We found that prostaglandin (PG) D(2), the most abundant PG in the central nervous system, stimulates food intake after intracerebroventricular administration in mice. The orexigenic effect of PGD(2) was mimicked by a selective agonist for the DP(1) receptor among two receptor subtypes for PGD(2), and abolished by its antagonist. Central administration of an antagonist or antisense oligodeoxynucleotide for the DP(1) receptor remarkably decreased food intake, body weight and fat mass. Hypothalamic mRNA levels of lipocalin-type PGD synthase were up-regulated after fasting. The orexigenic activity of PGD(2) was also abolished by an antagonist for neuropeptide Y (NPY) Y(1) receptor. Taken together, PGD(2) may stimulate food intake through central DP(1) receptor coupled to the NPY system.     

Orexigenic effect of the melanocortin MC4 receptor antagonist HS014 is inhibited only partially by neuropeptide Y Y1 receptor selective antagonists

Neuropeptide Y (NPY) and melanocortin (MC) peptides have opposite effects on food intake: NPY-like peptides and MC receptor antagonists stimulate feeding and increase body weight, whereas melanocortins and NPY antagonists inhibit food intake. In this study we tested whether the orexigenic effect of the selective MC4 receptor antagonist HS014 (1 nmol) could be inhibited by three different NPY antagonists, (R)-N2-(diphenylacetyl)-N-[(4-hydroxy-phenyl)methyl]D-argininam ide (BIBP3226), (R)-N-[[4-(aminocarbonylaminomethyl)-phenyl]methyl]-N2(diphenyl acetyl)-argininamidetrifluoroacetate (BIBO3304), and decapeptide [D-Tyr(27,36)D-Thr32]NPY(27-36), after icv administration in freely feeding male rats. All three NPY receptor antagonists inhibited the orexigenic effects of HS014 partially and with markedly different potency. [D-Tyr(27,36)D-Thr32]NPY(27-36) was active only in subconvulsive dose. The NPY Y1 selective antagonist BIBP3226 was more effective in inhibiting the effect of HS014 than BIBO3304 despite in vitro data indicating that BIBP3226 is about 10 times less potent than BIBO3304 at NPY Y1 receptor. An enantiomer of BIBO3304, BIBO3457, failed to inhibit HS014-induced feeding, indicating that the effects of BIBO3304 were stereoselective. These results suggest that stimulation of food intake caused by weakening of melanocortinergic tone at the MC4 receptor is partially but not exclusively related to NPY Y1 receptor activation.     

Differential body weight and feeding responses to high-fat diets in rats and mice lacking cholecystokinin 1 receptors

Prior data demonstrated differential roles for cholecystokinin (CCK)1 receptors in maintaining energy balance in rats and mice. CCK1 receptor deficiency results in hyperphagia and obesity of Otsuka Long-Evans Tokushima Fatty (OLETF) rats but not in mice. To ascertain the role of CCK1 receptors in high-fat-diet (HFD)-induced obesity, we compared alterations in food intake, body weight, fat mass, plasma glucose, and leptin levels, and patterns of hypothalamic gene expression in OLETF rats and mice lacking CCK1 receptors in response to a 10-wk exposure to HFD. Compared with Long-Evans Tokushima Otsuka (LETO) control rats, OLETF rats on HFD had sustained overconsumption over the 10-wk period. High fat feeding resulted in greater increases in body weight and plasma leptin levels in OLETF than in LETO rats. In situ hybridization determinations revealed that, while HFD reduced neuropeptide Y (NPY) mRNA expression in both the arcuate nucleus (Arc) and the dorsomedial hypothalamus (DMH) of LETO rats, HFD resulted in decreased NPY expression in the Arc but not in the DMH of OLETF rats. In contrast to these results in OLETF rats, HFD increased food intake and induced obesity to an equal degree in both wild-type and CCK1 receptor(-/-) mice. NPY gene expression was decreased in the Arc in response to HFD, but was not detectable in the DMH in both wild-type and CCK1 receptor(-/-) mice. Together, these data provide further evidence for differential roles of CCK1 receptors in the controls of food intake and body weight in rats and mice.     

Selective activation of central NPY Y1 vs. Y5 receptor elicits hyperinsulinemia via distinct mechanisms

Central administration of neuropeptide Y (NPY) stimulates hyperphagia and hyperinsulinemia. Recent evidence has suggested that the Y1 and Y5 receptor subtypes may both mediate NPY-stimulated feeding. The present study attempts to further characterize the role of central NPY receptor subtypes involved in hyperinsulinemia. NPY and peptide analogs of NPY that selectively activated the NPY Y1 or Y5 receptor subtype induced feeding and hyperinsulinemia in satiated Long Evans rats, whereas NPY analogs that selectively activated the NPY Y2 or Y4 receptor subtype did not. To determine whether NPY-induced hyperinsulinemia is secondary to its hyperphagic effect, we compared the plasma insulin levels in the presence and absence of food after a 1-min central infusion of NPY and its analogs at 15, 60, and 120 min postinfusion. Our data suggest that selective activation of central NPY Y1 receptor subtype induced hyperinsulinemia independent of food ingestion, whereas the NPY Y5 receptor-induced hyperinsulinemia was dependent on food ingestion. Central administration of the selective Y1 receptor agonist D-Arg25 NPY eventually decreased plasma glucose levels 2 h postinfusion in Long Evans rats.     

ICV NPY Y1 receptor agonist but not Y5 agonist induces torpor-like hypothermia in cold-acclimated Siberian hamsters

The reduced metabolism derived from daily torpor enables numerous small mammals, including Siberian hamsters, to survive periods of energetic challenge. Little is known of the neural mechanisms underlying the initiation and expression of torpor. Hypothalamic neuropeptide Y (NPY) contributes to surviving energetic challenges by both increasing food ingestion and reducing metabolic expenditure. Intracerebroventricular injections of NPY in cold-acclimated Siberian hamsters induce torpor-like hypothermia comparable to natural torpor. Multiple NPY receptor subtypes have been identified, and the Y1 receptor and Y5 receptor both contribute to the orexigenic effect of NPY. The purpose of this research was to compare and contrast the effects of Y1 receptor activation by a specific Y1 agonist ([D-Arg25]-NPY) or Y5 receptor activation by a specific Y5 agonist ([D-Trp34]-NPY) on body temperature and subsequent food intake in cold-acclimated Siberian hamsters. Intracerebroventricular injections of Y1 agonist produced torporlike hypothermia closely resembling that induced by intracerebroventricular NPY. The intracerebroventricular Y5 agonist infrequently produced hypothermia reaching criterion for torpor and that failed to resemble either NPY-induced or natural torpor. Combined injections of Y1 and Y5 agonists resulted in hypothermia comparable to Y5 agonist treatments alone, negating the mimicry of NPY treatment seen with Y1 agonist alone. Prior treatment with Y1 agonist or Y5 agonist surprisingly had lingering effects on NPY-induced torpor expression, Y1 agonist enhanced and Y5 agonist inhibited the effect of NPY. The ability of NPY to induce torporlike hypothermia, especially its initiation, most likely involves activation of the NPY Y1 receptor subtype.     

Leptin and neuropeptide Y (NPY) modulate nitric oxide synthase: further evidence for a role of nitric oxide in feeding.

Recent studies have suggested a role for nitric oxide in the regulation of food intake. Neuropeptide Y (NPY) is one of the most potent orexigenic agents. Chronic administration of leptin decreases food intake. This study examined the effects of NPY and leptin on nitric oxide synthase (NOS) in the hypothalamus. Previously it has been demonstrated that obese (ob/ob) mice have elevated NOS levels in the hypothalamus. In this study we demonstrated that the administration of leptin (6 microg/day) subcutaneously (SC) for 3 days decreased body weight (P < 0.001) and food intake P < 0.001) in obese (ob/ob) mice as expected. In addition, leptin decreased NOS in the hypothalamus nu 37% (P < 0.01) and in brown adipose tissue by 69% (P < 0.01) but not in white adipose tissue. NPY was administered intracerebroventricularly to CD-1 mice at doses of 0.25 and 0.50 microg. Mice were sacrificed 15 min after injection and NOS was measured in their hypothalami. NPY at the lower dose increased NOS in the hypothalamus by 147%. These results, taken together, with previously published studies support the concept that nitric oxide may play a role as a mediator of the effects of NPY and leptin on food intake. The alterations of NOS in brown adipose tissue following leptin administration could result in changes in blood flow or metabolism in the brown fat.     

Neither agouti-related protein nor neuropeptide Y is critically required for the regulation of energy homeostasis in mice

Agouti-related protein (AgRP), a neuropeptide abundantly expressed in the arcuate nucleus of the hypothalamus, potently stimulates feeding and body weight gain in rodents. AgRP is believed to exert its effects through the blockade of signaling by alpha-melanocyte-stimulating hormone at central nervous system (CNS) melanocortin-3 receptor (Mc3r) and Mc4r. We generated AgRP-deficient (Agrp(-/-)) mice to examine the physiological role of AgRP. Agrp(-/-) mice are viable and exhibit normal locomotor activity, growth rates, body composition, and food intake. Additionally, Agrp(-/-) mice display normal responses to starvation, diet-induced obesity, and the administration of exogenous leptin or neuropeptide Y (NPY). In situ hybridization failed to detect altered CNS expression levels for proopiomelanocortin, Mc3r, Mc4r, or NPY mRNAs in Agrp(-/-) mice. As AgRP and the orexigenic peptide NPY are coexpressed in neurons of the arcuate nucleus, we generated AgRP and NPY double-knockout (Agrp(-/-);Npy(-/-)) mice to determine whether NPY or AgRP plays a compensatory role in Agrp(-/-) or NPY-deficient (Npy(-/-)) mice, respectively. Similarly to mice deficient in either AgRP or NPY, Agrp(-/-);Npy(-/-) mice suffer no obvious feeding or body weight deficits and maintain a normal response to starvation. Our results demonstrate that neither AgRP nor NPY is a critically required orexigenic factor, suggesting that other pathways capable of regulating energy homeostasis can compensate for the loss of both AgRP and NPY.     

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.