Coordinated changes in energy intake and expenditure following hypothalamic administration of neuropeptides involved in energy balance.

OBJECTIVE: The hypothalamic control of energy balance is regulated by a complex network of neuropeptide-releasing neurons. Although the effect of these neuropeptides on individual aspects of energy homoeostasis has been studied, the coordinated response of these effects has not been comprehensively investigated. We have simultaneously monitored a number of metabolic parameters following intracerebroventricular (ICV) administration of 1 and 3 nmol of neuropeptides with established roles in the regulation of feeding, activity and metabolism. Ad libitum- fed rats received the orexigenic neuropeptides neuropeptide Y (NPY), agouti-related protein (AgRP), melanin-concentrating hormone (MCH) or orexin-A. Overnight-food-deprived rats received an ICV injection of the anorectic peptides alpha-melanocyte-stimulating hormone (MSH), corticotrophin-releasing factor (CRF) or neuromedin U (NMU). RESULTS: Our results reveal the temporal sequence of the effects of these neuropeptides on both energy intake and expenditure, highlighting key differences in their function as mediators of energy balance. NPY and AgRP increased feeding and decreased oxygen consumption, with the effects of AgRP being more prolonged. In contrast, orexin-A increased both feeding and oxygen consumption, consistent with an observed increase in activity. The potent anorexigenic effects of CRF were accompanied by a prolonged increase in activity, whereas NMU injection resulted in significant but short-lasting inhibition of food intake, ambulatory activity and oxygen consumption. alpha-MSH injection resulted in significant increases in both ambulatory activity and oxygen consumption, and reduced food intake following administration of 3 nmol of the peptide. CONCLUSION: We have for the first time, simultaneously measured several metabolic parameters following hypothalamic administration of a number of neuropeptides within the same experimental system. This work has shown the interrelated effects of these neuropeotides on activity, energy expenditure and food intake, thus facilitating comparison between the different hypothalamic systems.

Co-administration of SR141716 with peptide YY3-36 or oxyntomodulin has additive effects on food intake in mice.

BACKGROUND: SR141716 has been shown to significantly inhibit food intake and reduce body weight by antagonizing CB(1) receptors. The gut hormones peptide YY(3-36) (PYY(3-36)) and oxyntomodulin (OXM) inhibit food intake through Y(2) and Glucagon-Like-Peptide (GLP)-1 receptors respectively. OBJECTIVE: To determine the effects of co-administration of SR141716 with either PYY(3-36) or OXM in mice on food intake. METHODS: Mice (n = 14 per group) were fasted for 16 h prior to study days and given two intraperitoneal injections: study 1, vehicle-saline, SR141716-saline, vehicle-PYY3-36 or SR141716-PYY3-36; study 2: vehicle-saline, SR141716-saline, vehicle-OXM or SR141716-OXM. Food was returned and measured following injections. RESULTS: Co-administration of SR141716-PYY(3-36) or SR141716-OXM showed greater inhibition in food intake when compared with administration of SR141716, PYY(3-36) or OXM alone. CONCLUSION: Our data show that SR141716 in combination with PYY(3-36) or OXM reduces food intake additively in mice.

Low-dose oral tri-iodothyronine does not directly increase food intake in man.

Previously, we have shown that low-dose tri-iodothyronine (T3) increases food intake in rodents. This randomised, double-blind, placebo-controlled study aimed to investigate the effects of low-dose T3 on food intake in normal body weight individuals. However, despite an elevation in fT3 comparable to our earlier studies, administration of low-dose T3 in the fasted state did not stimulate food intake in man.

Effects of acute and chronic relaxin-3 on food intake and energy expenditure in rats.

The effects of acute and repeated intraparaventricular (iPVN) administration of human relaxin-3 (H3) were examined on food intake, energy expenditure, and the hypothalamo-pituitary thyroid axis in male Wistar rats. An acute high dose iPVN injection of H3 significantly increased food intake 1 h post-administration [0.4+/-0.1 g (vehicle) vs 1.6+/-0.5 g (180 pmol H3), 2.4+/-0.5 g (540 pmol H3) and 2.2+/-0.5 g (1,620 pmol H3), p<0.05 for all doses vs vehicle]. Repeated iPVN H3 injection (180 pmol/twice a day for 7 days) significantly increased cumulative food intake in ad libitum fed animals compared with vehicle [211.8+/-7.1 g (vehicle) vs 261.6+/-6.7 g (ad libitum fed H3), p<0.05]. Plasma leptin was increased in the H3 ad libitum fed group. Plasma thyroid stimulating hormone was significantly decreased after acute and repeated administration of H3. These data suggest H3 may play a role in long-term control of food intake.

Oxyntomodulin increases energy expenditure in addition to decreasing energy intake in overweight and obese humans: a randomised controlled trial.

BACKGROUND: Oxyntomodulin has recently been found to decrease body-weight in obese humans and may be a potential anti-obesity therapy. OBJECTIVE: To determine whether oxyntomodulin alters energy expenditure, in addition to reducing energy intake, in 'free-living' overweight and obese volunteers. DESIGN: Randomized double-blind controlled cross-over trial. SETTING: Community and hospital-based. PARTICIPANTS: Fifteen healthy overweight and obese men and women (age: 23-49 years, BMI: 25.1-39.0 kg/m(2)). All volunteers completed the study protocol. INTERVENTIONS: Four-day subcutaneous self-administration of pre-prandial oxyntomodulin, three times daily. Participants were advised to maintain their normal dietary and exercise regimen. MEASUREMENTS: (1) Energy expenditure, measured by indirect calorimetry and combined heart rate and movement monitoring; (2) energy intake, measured during a study meal. RESULTS: Oxyntomodulin administration reduced energy intake at the study meal by 128+/-29 kcal (P=0.0006) or 17.3+/-5.5% (P=0.0071), with no change in meal palatability. Oxyntomodulin did not alter resting energy expenditure; but increased activity-related energy expenditure by 143+/-109 kcal/day or 26.2+/-9.9% (P=0.0221); total energy expenditure by 9.4+/-4.8% (P=0.0454) and physical activity level by 9.5+/-4.6% (P=0.0495). A reduction in body weight of 0.5+/-0.2% was observed during the oxyntomodulin administration period (P=0.0232). CONCLUSION: Oxyntomodulin increases energy expenditure while reducing energy intake resulting in negative energy balance. This data supports the role of oxyntomodulin as a potential anti-obesity therapy.

Evidence for a stimulatory action of melanin-concentrating hormone on luteinising hormone release involving MCH1 and melanocortin-5 receptors.

The present series of studies aimed to further our understanding of the role of melanin-concentrating hormone (MCH) neurones in the central regulation of luteinising hormone (LH) release in the female rat. LH release was stimulated when MCH was injected bilaterally into the rostral preoptic area (rPOA) or medial preoptic area (mPOA), but not when injected into the zona incerta (ZI), of oestrogen-primed ovariectomised rats. In rats that were steroid-primed to generate a surge-like release of LH, MCH administration into the ZI blocked this rise in LH release: no such effect occurred when MCH was injected into the rPOA or mPOA. In vitro, MCH stimulated gonadotrophin-releasing hormone (GnRH) release from hypothalamic explants. Double-label immunohistochemistry showed GnRH-immunoreactive neurones in the vicinity of and intermingled with immunoreactive MCH processes. MCH is the endogenous ligand of the MCH type 1 receptor (MCH1-R). Previously, we have shown a role for melanocortin-5 receptors (MC5-R) in the stimulatory action of MCH, so we next investigated the involvement of both MCH1-R and/or MC5-R in mediating the actions of MCH on GnRH and hence LH release. The stimulatory action of MCH in the rPOA was inhibited by administration of antagonists for either MCH1-R or MC5-R. However, in the mPOA, the action of MCH was blocked only by the MC5-R antagonist. LH release was stimulated by an agonist for MC5-R injected into the rPOA or mPOA; this was blocked by the MC5-R antagonist but not the MCH1-R antagonist. These results indicate that both MCH1-R and MC5-R are involved in the central control of LH release by MCH.

The importance of acclimatisation and habituation to experimental conditions when investigating the anorectic effects of gastrointestinal hormones in the rat.

OBJECTIVE: Peptide YY3-36 (PYY(3-36)), glucagon-like peptide-1 (GLP-1), oxyntomodulin and cholecystokinin (CCK) are gastrointestinal-derived hormones that are released postprandially in proportion to the amount of calories ingested. All significantly reduce food intake following peripheral administration to rodents. We have investigated the effect of handling, exposure to a novel environment or to environmental enrichment on the anorectic effect of these gut hormones. RESULTS: Results suggest that the transfer of a rat into a novel environment (cage change) inhibits the anorectic response to peripherally administered PYY(3-36) and oxyntomodulin (1 h food intake reduction (% saline control): PYY/home cage 82.3 +/- 5.9%, P < 0.05; PYY/clean cage 103.4 +/- 9.7%; oxyntomodulin/home cage 71.6 +/- 12.1%, P < 0.05; oxyntomodulin/clean cage 103.0 +/- 8.5%) and attenuates the anorectic response to GLP-1 and CCK (1 h food intake reduction (% saline control): GLP-1/home cage 68.8 +/- 6.4%, P < 0.01; GLP-1/clean cage 80.0 +/- 9.3%; CCK/home cage 49.8 +/- 6.2%, P < 0.001; CCK/clean cage 69.4 +/- 10.6%, P < 0.05). We have also observed that exposure to a novel environment does not alter anorectic effect of peripherally administered melanocortin 3/4 receptor agonist, melanotan II (MTII) (1 h food intake reduction (% saline control): MTII/home cage 32.0 +/- 6.3%, P < 0.001; MTII/clean cage 24.8 +/- 4.2%, P < 0.001). The attenuation in food intake observed following exposure to a novel environment can be attributed, in part, to a significant reduction in the food intake of the saline treated animals. In a further study, the anorectic effect of peripherally administered PYY(3-36) is attenuated in unhandled rats (88 +/- 4.2% saline control, P = ns) or rats exposed to environmental enrichment (103.3 +/- 9.7% saline control, P = ns), but not in animals that were handled extensively prior to the study (80.1 +/- 7.3% saline control, P < 0.05). CONCLUSION: These studies highlight the importance of handling, acclimatisation and habituation of rodents to experimental conditions prior to investigating the ability of gut hormones to alter food intake.

Subcutaneous administration of ghrelin stimulates energy intake in healthy lean human volunteers.

BACKGROUND: The gastric hormone ghrelin appears a useful agent to stimulate food intake in people with anorexia of illness. The loss of ghrelin's acyl group renders it inactive, thus it has been thought that subcutaneous administration may be problematic. OBJECTIVE: To investigate whether human subjects are sensitive to the effects of ghrelin administered by single subcutaneous injection. STUDY DESIGN: Randomized, double-blind, placebo-controlled trial. SUBJECTS: Sixteen healthy lean volunteers (eight men and eight women). PROTOCOL: Fasted subjects received subcutaneous injections of ghrelin (3.6 nmol/kg) or saline. After 30 min, a buffet breakfast was served. RESULTS: Ghrelin injection increased energy intake by 27% (ghrelin 5076 +/- 691 kJ versus saline 4230+/-607 kJ, P = 0.04). Ghrelin appeared to enhance the perceived palatability of the food offered (palatability score: ghrelin 81.1 +/- 3.6 versus saline 70.0 +/- 4.4; P = 0.03). CONCLUSIONS: These results suggest that subcutaneous ghrelin is effective at stimulating energy intake and improving palatability and may be of direct use in the treatment of appetite loss.

Abnormalities of the somatotrophic axis in the obese agouti mouse.

OBJECTIVE: Abnormalities of the melanocortin system produce obesity and increased linear growth. While the obesity phenotype is well characterised, the mechanism responsible for increased linear growth is unclear. The somatotrophic axis was studied in the obese agouti (A(y)/a) mouse as a model of a perturbed melanocortin system. DESIGN: Adult obese A(y)/a mice were compared to age- and sex-matched wild-type (WT) controls. Weight and body length (nose-anus) were recorded. Plasma growth hormone (GH), insulin-like growth factor-I (IGFI), insulin and leptin were measured using radioimmunoassay. Since ghrelin is a potent GH secretagogue, plasma ghrelin, stomach ghrelin peptide and stomach ghrelin mRNA expression were studied. Hypothalamic periventricular (PeVN) somatostatin neurones and arcuate (Arc) neuropeptide Y (NPY) neurones inhibit the growth axis, whereas Arc growth hormone-releasing hormone (GHRH) neurones are stimulatory. Therefore, specific hypothalamic expression of somatostatin, NPY and GHRH was measured using quantitative in situ hybridisation. RESULTS: Obese A(y)/a mice were significantly heavier and longer than WT controls. Plasma IGFI concentrations were 30% greater in obese A(y)/a mice. Obese A(y) /a mice were hyperinsulinaemic and hyperleptinaemic, yet plasma ghrelin, and stomach ghrelin peptide and mRNA were significantly reduced. In obese A(y)/a mice, PeVN somatostatin and Arc NPY mRNA expression were reduced by 50% compared to WT controls, whereas Arc GHRH mRNA expression was unchanged. CONCLUSION: Increased body length in adult obese A(y)/a mice may result from reduced Arc NPY and PeVN somatostatin mRNA expression, which in turn, may increase plasma IGFI concentrations and upregulate the somatotrophic axis.

Ghrelin does not stimulate food intake in patients with surgical procedures involving vagotomy.

CONTEXT: Patients with gastric or esophageal surgery and transection of the vagus nerve may suffer from appetite and weight loss but without dysphagia or mechanical obstruction to eating. The gastric hormone ghrelin stimulates food intake and GH release in rodents and man. However, rodents with vagotomy are not sensitive to the feeding effects of ghrelin. OBJECTIVE: The objective of the study was to determine whether humans with vagotomy are sensitive to ghrelin. STUDY DESIGN: The design was a double-blind, randomized, placebo-controlled trial. SETTING: This was a hospital-based study. PATIENTS: Six men and one woman who all had a previous complete truncal vagotomy with lower esophageal or gastric surgery entered and completed the study. INTERVENTION: Each patient received 120-min infusions of saline, 1 pmol/kg.min ghrelin, and 5 pmol/kg.min ghrelin on 3 separate days. After 90 min, a buffet meal was served. MAIN OUTCOME MEASURE: Energy intake at the buffet meal was measured. RESULTS: Ghrelin-stimulated GH release in a dose-dependent manner was measured, confirming bioactivity. However, no change in energy intake was observed with either dose of ghrelin [energy intake (kilojoules): saline 2805 +/- 812; ghrelin 1 pmol/kg.min, 2486 +/- 767; ghrelin 5 pmol/kg.min, 2382 +/- 543; P = not significant]. CONCLUSIONS: Ghrelin is unlikely to be an effective appetite-stimulatory treatment for patients with vagotomy and esophageal or gastric surgery. Our results suggest that an intact vagus nerve may be required for exogenous ghrelin to increase appetite and food intake in man.

Regulation of rat pituitary cocaine- and amphetamine-regulated transcript (CART) by CRH and glucocorticoids.

Cocaine- and amphetamine-regulated transcript (CART) was originally isolated from rat brain, but CART is also synthesized and stored in the anterior pituitary. The localization of pituitary CART and factors regulating its synthesis are largely unknown. The regulation of pituitary CART synthesis and release in response to CRH and glucocorticoids was examined in vitro and in vivo. CART immunoreactivity (CART-IR) was released from anterior pituitary segments. This release was increased 15-fold in response to corticotropin-releasing hormone (CRH). Intraperitoneal administration of CRH to rats significantly increased plasma CART-IR. Furthermore, CART-IR content and plasma CART-IR were significantly increased in adrenalectomized rats, and anterior pituitary CART mRNA expression, CART-IR content, and plasma CART-IR were significantly decreased in corticosterone-treated rats. Plasma CART-IR showed a pattern of diurnal variation similar to that of ACTH and corticosterone, and plasma CART-IR was positively correlated with corticosterone. CART-IR was detectable in the medium of the corticotroph cell line AtT-20. Dual in situ hybridization for prepro-CART (ppCART) mRNA expression and immunocytochemistry for ACTH showed localization of ppCART mRNA to a subpopulation of ACTH-immunoreactive cells. These findings demonstrate that pituitary CART expression and release are regulated by CRH and the glucocorticoid environment and that pituitary CART is partly localized to corticotrophs.

Pre-obese and obese agouti mice are sensitive to the anorectic effects of peptide YY(3-36) but resistant to ghrelin.

OBJECTIVE: The role of the melanocortin system in the feeding effects of peripheral peptide YY(3-36) (PYY(3-36)) and ghrelin was investigated using the agouti (A(y)/a) mouse as a model of abnormal melanocortin signalling. Furthermore, we examined whether the ectopic expression of agouti protein in A(y)/a mice results in complete MC4-R inhibition, by studying the effects of peripheral alpha-melanocyte-stimulating hormone (alpha-MSH) and leptin on food intake. DESIGN: Adult A(y)/a mice were studied in the pre-obese state (7-8 weeks) and obese state (14-15 weeks). Animals received PYY(3-36) (0.02 micromol/kg), NDP-alpha-MSH (0.2 micromol/kg), leptin (2 micromol/kg) (all 24 h fasted state) and ghrelin (0.2 micromol/kg) (fed state) by intraperitoneal (i.p.) injection. Age-matched A(y)/a controls received i.p. saline. A separate cohort of wild-type (WT), age-matched controls received the same peptide dose or saline. Food intake was measured at 1, 2, 4, 8 and 24 h post-injection and compared in all four groups. Plasma leptin-, ghrelin- and PYY-like immunoreactivity (IR) were measured using radioimmunoassay (RIA). RESULTS: At 2 h post-injection, PYY(3-36) reduced food intake in pre-obese and obese A(y)/a mice, whereas ghrelin had no effect. Plasma ghrelin levels were significantly reduced in pre-obese and obese A(y)/a mice compared to WT controls. Peripheral administration of NDP-alpha-MSH and leptin acutely suppressed feeding (0-2 h) in pre-obese and obese A(y)/a mice. CONCLUSIONS: Responsiveness of pre-obese and obese A(y)/a mice to PYY(3-36) suggests that the melanocortin system may not be essential for the anorectic effects of this peptide. Melanocortinergic antagonism by agouti protein in A(y)/a mice may be sufficient to block the effects of endogenous, but not exogenous PYY(3-36), alpha-MSH and leptin. The mechanism underlying ghrelin resistance in A(y)/a mice may result from antagonism of hypothalamic melanocortin receptors-4 by agouti protein, supporting a role for the melanocortin system in mediating ghrelin's actions.

Central orexin A has site-specific effects on luteinizing hormone release in female rats.

Orexin A stimulates GnRH release from hypothalamic explants in vitro. The sites of action of orexin A in the regulation of LH release have been investigated in vivo in ovariectomized rats that were given vehicle or estradiol benzoate (EB), with or without an injection of progesterone 48 h later. Orexin A was administered intrahypothalamically under Saffan anesthesia, 50 h after the EB or vehicle; its effects on plasma LH levels were monitored in sequential blood samples. Orexin A (1.0 microg/side) injected into the rostral preoptic area (rPOA) at the level of the organum vasculosum of the lamina terminalis had a stimulatory effect on LH release in EB-treated ovariectomized rats. When orexin A was injected into the medial POA (mPOA) or the arcuate/median eminence, it had an inhibitory effect on the LH surge that occurs in ovariectomized rats primed with EB plus progesterone. Orexin A injected into the mPOA also reduced LH levels in ovariectomized rats untreated with ovarian steroids. Both the stimulatory and inhibitory effects of orexin A were antagonized by SB334867A, a selective orexin 1 receptor antagonist. Furthermore, when given alone into the rPOA, this antagonist attenuated the LH surge induced by EB plus progesterone. Thus, orexin appears to have a dual effect on LH release, being stimulatory in the rPOA and inhibitory in the mPOA or arcuate/median eminence. Both effects may be mediated, at least in part, by the orexin 1 receptor. Double label immunohistochemistry revealed close appositions between orexin A immunoreactive varicosities and a small proportion of GnRH cell bodies in the rPOA. It is suggested that the stimulatory effect of orexin A on LH release may involve direct actions on GnRH neurons.

Gut and mind.

Obesity is a growing epidemic, causally associated with a number of serious medical conditions, including diabetes mellitus, coronary heart disease, and several cancers. The gut hormones ghrelin and peptide YY are secreted from the gut in response to changes to nutritional status. While food intake is stimulated by ghrelin, it is inhibited by peptide YY. The discovery, anatomy, and physiology of ghrelin and peptide YY are discussed, focusing on the adaptive changes in diseases such as obesity and anorexia nervosa. Ghrelin and PYY are important therapeutic targets in the quest to find an effective antiobesity treatment.

gamma-MSH increases intracellular cAMP accumulation and GnRH release in vitro and LH release in vivo.

The roles of the melanocortin 3 receptor (MC3-R) and its agonist, gamma(2)-melanocyte-stimulating hormone (gamma(2)-MSH) in the regulation of the hypothalamo-pituitary-gonadal (HPG) axis are poorly understood. Here we show gamma(2)-MSH stimulated intracellular cAMP accumulation and gonadotrophin-releasing hormone (GnRH) secretion in the immortalised GnRH cell line GT(1)-7. The MC3/4-R antagonist Agrp blocked these actions. Reverse transcriptase polymerase chain reaction demonstrated GT(1)-7 cells express MC3-R mRNA. gamma(2)-MSH also stimulated GnRH release from hypothalamic explants. In vivo, gamma(2)-MSH administration into the medial preoptic area significantly increased plasma luteinising hormone. MC3-R and gamma(2)-MSH may modulate the HPG axis.

Chronic CNS administration of Agouti-related protein (Agrp) reduces energy expenditure.

OBJECTIVE: To investigate whether the Agouti-related protein (Agrp), the melanocortin receptor antagonist, alters oxygen consumption, as a measure of energy expenditure. DESIGN: A 7-day intracerebroventricular administration of Agrp (1 nmol/day) in rats. MEASUREMENTS: Oxygen consumption was determined in closed-circuit respirometers on days 1 and 8. BRL-35135, a beta3-adrenoreceptor agonist known to activate the brown adipose tissue (BAT) thermogenesis directly and increase core temperature, was administered i.p. (40 microg/kg) on day 9 to challenge functionally the BAT. RESULTS: Agrp treatment caused a 54% increase in daily food intake and a 12% increase in body weight. An 8% decrease in VO(2) measurements was observed following ICV Agrp treatment on day 1. A similar decrease (7%) was observed on day 8. BRL-35135 stimulated colonic temperature in control rats. However, in the rats that had previously been treated with Agrp this effect was significantly blunted. CONCLUSION: Chronic CNS administration of Agrp decreases oxygen consumption and decreases the capacity of BAT to expend energy. The obesity observed following CNS administration of Agrp is the result of decreased energy expenditure and increased food intake.

Agouti-related protein has an inhibitory paracrine role in the rat adrenal gland.

alpha-Melanocyte-stimulating-hormone (alpha-MSH) is an agonist at the melanocortin 3 receptor (MC3-R) and melanocortin 4 receptor (MC4-R). alpha-MSH stimulates corticosterone release from rat adrenal glomerulosa cells in vitro. Agouti-related protein (AgRP) an endogenous antagonist at the MC3-R and MC4-R, is expressed in the adrenal gland. We investigated the expression of the MC3-R and MC4-R and the role of AgRP in the adrenal gland. MC3-R and MC4-R expression was detected in rat adrenal gland using RT-PCR. The effect of AgRP on alpha-MSH-induced corticosterone release was investigated using dispersed rat adrenal glomerulosa cells. AgRP administered alone did not affect corticosterone release, but co-administration of AgRP and alpha-MSH attenuated alpha-MSH-induced corticosterone release. To investigate glucocorticoid feedback, adrenal AgRP expression was compared in rats treated with dexamethasone to controls. AgRP mRNA was increased in rats treated with dexamethasone treatment compared to controls. Our findings demonstrate that adrenal AgRP mRNA is regulated by glucocorticoids. AgRP acting via the MC3-R or MC4-R may have an inhibitory paracrine role, blocking alpha-MSH-induced corticosterone secretion.

Hypothalamic actions of neuromedin U.

The central nervous system and gut peptide neuromedin U (NMU) inhibits feeding after intracerebroventricular injection. This study explored the hypothalamic actions of NMU on feeding and the hypothalamo-pituitary-adrenal axis. Intraparaventricular nucleus (intra-PVN) NMU dose-dependently inhibited food intake, with a minimum effective dose of 0.1 nmol and a robust effect at 0.3 nmol. Feeding inhibition was mapped by NMU injection into eight hypothalamic areas. NMU (0.3 nmol) inhibited food intake in the PVN (0-1 h, 59 +/- 6.9% of the control value; P < 0.001) and arcuate nucleus (0-1 h, 76 +/- 10.4% of the control value; P < 0.05). Intra-PVN NMU markedly increased grooming and locomotor behavior and dose-dependently increased plasma ACTH (0.3 nmol NMU, 24.8 +/- 1.9 pg/ml; saline, 11.4 +/- 1.0; P < 0.001) and corticosterone (0.3 nmol NMU, 275.4 +/- 40.5 ng/ml; saline, 129.4 +/- 25.0; P < 0.01). Using hypothalamic explants in vitro, NMU stimulated CRH (100 nM NMU, 5.9 +/- 0.95 pmol/explant; basal, 3.8 +/- 0.39; P < 0.01) and arginine vasopressin release (100 nM NMU, 124.5 +/- 21.8 fmol/explant; basal, 74.5 +/- 7.6; P < 0.01). Leptin stimulated NMU release (141.9 +/- 20.4 fmol/explant; basal, 92.9 +/- 9.4; P < 0.01). Thus, we describe a novel role for NMU in the PVN to stimulate the hypothalamo-pituitary-adrenal axis and locomotor and grooming behavior and to inhibit feeding.

Hypothalamic interactions between neuropeptide Y, agouti-related protein, cocaine- and amphetamine-regulated transcript and alpha-melanocyte-stimulating hormone in vitro in male rats.

A number of neuropeptides implicated in the hypothalamic regulation of appetite are synthesized in the arcuate nucleus (Arc). Neuropeptide Y (NPY) and agouti-related protein (Agrp) are orexigenic. The pro-opiomelanocortin (POMC) product alpha-melanocyte-stimulating hormone (alpha-MSH) is anorectic. Intracerebroventricular administration of cocaine- and amphetamine-regulated transcript (CART) decreases food intake. However, recent results show that CART is orexigenic when injected into discrete hypothalamic nuclei. There is almost complete coexpression of NPY and Agrp mRNA in Arc neurones, and the majority of CART-containing neurones in the Arc also contain POMC mRNA. We investigated possible interactions between these neuropeptides in vitro using a rat hypothalamic explant system. Administration of 1, 10 and 100 nm of NPY to hypothalamic explants significantly increased release of Agrp(83-132)-immunoreactivity (IR). NPY (10 and 100 nm) significantly increased the release of CART(55-102)-IR and alpha-MSH-IR from hypothalamic explants. Agrp(83-132) (10 nm) administered to hypothalamic explants significantly increased the release of NPY-IR. Agrp(83-132) (10 and 100 nm) significantly decreased the release of CART(55-102)-IR from hypothalamic explants. Administration of 1, 10 and 100 nm CART(55-102) to hypothalamic explants resulted in a significant increase in NPY-IR release. Administration of 10 nm CART(55-102) to hypothalamic explants significantly increased the release of Agrp(83-132)-IR. NDP-MSH (10 nm) administered to hypothalamic explants significantly increased the release of NPY-IR. NDP-MSH (10 and 100 nm) significantly increased the release of Agrp(83-132)-IR from hypothalamic explants. These data suggest that orexigenic neuropeptides in the arcuate nucleus stimulate the release of each other, perhaps reinforcing orexigenic behaviour via a positive-feedback loop. Our results are also in keeping with the possibility that the melanocortin-3 receptor in the arcuate nucleus may influence the release of arcuate neuropeptides.