Nocturnal Food Intake Research Articles

Central nesfatin-1 reduces the nocturnal food intake in mice by reducing meal size and increasing inter-meal intervals

Nesfatin-1 is well established to reduce food intake upon brain injection in rats, while in mice its anorexigenic action and brain expression are largely unexplored. We characterized the influence of intracerebroventricular (icv) and peripheral (intraperitoneal, ip, subcutaneous, sc) injection of nesfatin-1 on dark phase ingestive behavior using an automated feeding monitoring system and co-localized NUCB2/nesfatin-1 immunoreactivity in the associated brain areas. Nesfatin-1 (0.3, 1 or 3 μg/mouse, icv) caused a dose-related reduction of 4-h dark phase food intake by 13%, 27%, and 46% respectively. Nesfatin-1 (3 μg/mouse, icv) action had a 2-h delayed onset, 82% peak inhibition occurring at 3–4 h post-injection and was long lasting (30% reduction for 12 h period post-injection). Nesfatin-1 (3 μg/mouse, icv)-treated mice had a 46% lower meal frequency associated with 2-times longer inter-meal intervals and a 35% reduction in meal size compared to vehicle during the 1–4 h post-injection ( p < 0.05). NUCB2/nesfatin-1-immunopositive neurons were found in hypothalamic (supraoptic, paraventricular, arcuate, dorsomedial, lateral) and brainstem (dorsal vagal complex) feeding regulatory nuclei. When injected peripherally, neither food intake nor feeding microstructure parameters were altered. These results demonstrate that NUCB2/nesfatin-1 is prominently expressed in mouse hypothalamus and medulla and acts in the brain to curtail the dark phase feeding by inducing satiation and satiety indicated by reduced meal size and prolonged inter-meal intervals respectively. The lack of nesfatin-1 effect when injected peripherally at a 23-times higher dose indicates a primarily central site of the anorexigenic action for nesfatin-1 in mice.

The functional architecture of dehydration-anorexia

The anorexia that accompanies the drinking of hypertonic saline (DE-anorexia) is a critical adaptive behavioral mechanism that helps protect the integrity of fluid compartments during extended periods of cellular dehydration. Feeding is rapidly reinstated once drinking water is made available again. The relative simplicity and reproducibility of these behaviors makes DE-anorexia a very useful model for investigating how the various neural networks that control ingestive behaviors first suppress and then reinstate feeding. We show that DE-anorexia develops primarily because the mechanisms that terminate ongoing meals are upregulated in such a way as to significantly reduce meal size. At the same time however, signals generated by the ensuing negative energy balance appropriately activate neural mechanisms that can increase food intake. But as the output from these two competing processes is integrated, the net result is an increasing reduction of nocturnal food intake, despite the fact that spontaneous meals are initiated with the same frequency as in control animals. Furthermore, hypothalamic NPY injections also stimulate feeding in DE-anorexic animals with the same latency as controls, but again meals are prematurely terminated. Comparing Fos expression patterns across the brain following 2-deoxyglucose administration to control and DE-anorexic animals implicates neurons in the descending part of the parvicellular paraventricular nucleus of the hypothalamus and the lateral hypothalamic areas as key components of the networks that control DE-anorexia. Finally, DE-anorexia generates multiple inhibitory processes to suppress feeding. These are differentially disengaged once drinking water is reinstated. The paper represents an invited review by a symposium, award winner or keynote speaker at the Society for the Study of Ingestive Behavior [SSIB] Annual Meeting in Portland, July 2009.

Sex differences in the cannabinoid regulation of energy homeostasis

This review highlights the progress made thus far in characterizing the behavioral and cellular mechanisms through which cannabinoids regulate energy homeostasis. We performed microstructural analysis of feeding behavior in gonadectomized guinea pigs and gonadally intact, transgenic CB1 receptor knockout mice to determine how cannabinoids affect circadian rhythms in food intake and meal pattern. We also implanted data loggers into the abdominal cavity to correlate the appetite-modulating properties of cannabinoids with changes in core body temperature. We then coupled the effects on feeding behavior and temperature regulation with synaptic changes in the hypothalamic feeding circuitry via whole-cell patch clamp electrophysiological recording from neurons in the arcuate nucleus (ARC), in order to gain a more global perspective on the cannabinoid modulation of energy homeostasis. We observed marked sex differences in cannabinoid effects on food intake and core body temperature--with male guinea pigs exhibiting a comparatively greater sensitivity to the hyperphagia and hypophagia, as well as the hypothermia and hyperthermia, produced by CB1 receptor agonists and antagonists, respectively. In addition, male but not female CB1 receptor knockout mice show a diminished nocturnal food intake and average daily body weight relative to their wildtype littermate controls. The disparity in the CB1 receptor-mediated hyperphagia is paralleled by sex differences in the cellular effects of cannabinoids at anorexigenic, guinea pig proopiomelanocortin (POMC) synapses. Postsynaptically, cannabinoids potentiate an A-type K+ current (I(A)) in POMC neurons from female guinea pigs, whereas in males the activation of an inwardly rectifying K+ current is observed. Presynaptically, while cannabinoids inhibit glutamatergic input onto POMC neurons in males and females to similar degrees, males are more refractory to the cannabinoid-induced inhibition of convergent GABAergic input than females. These data reveal pervasive sex differences in the cannabinoid regulation of energy homeostasis that are consistent with changes in the excitability of POMC neurons.

Identification and Characterization of Nesfatin-1 Immunoreactivity in Endocrine Cell Types of the Rat Gastric Oxyntic Mucosa

Hypothalamic nesfatin-1, derived from the nucleobindin2 (NUCB2) precursor, inhibits nocturnal food intake and body weight gain in rats. Nesfatin-1 is able to cross the blood-brain barrier, suggesting a peripheral source of nesfatin-1. Many centrally acting food intake regulatory neuropeptides are also produced in the periphery, especially in the gastrointestinal tract. Therefore, we investigated the gene expression of NUCB2 and distribution of nesfatin-1-immunoreactive cells in the stomach. Microarray mRNA expression profiles in purified small endocrine cells of the gastric mucosa substantiated by quantitative RT-PCR showed significantly higher NUCB2 mRNA expression compared with brain and heart. Western blot confirmed the expression of NUCB2 protein and its transport into a secretory soluble fraction of gastric mucosal endocrine cell homogenates. Immunohistochemical colabeling for nesfatin-1 and ghrelin, histidine decarboxylase, or somatostatin revealed two subtypes of nesfatin-1-positive endocrine cells. Cells in the midportion of the glands coexpressed nesfatin-1 and ghrelin, whereas few cells in the glandular base coexpressed nesfatin-1 and somatostatin or histidine decarboxylase. High-resolution three-dimensional volume imaging revealed two separate populations of intracytoplasmic vesicles in these cells, one containing nesfatin-1 and the other ghrelin immunoreactivity. Microarray rat genome expression data of NUCB2 in small gastric endocrine cells confirmed by quantitative RT-PCR showed significant down-regulation of NUCB2 after 24 h fasting. In summary, NUCB2 mRNA expression as well as protein content is present in a specific subset of gastric endocrine cells, most of which coexpress ghrelin. NUCB2 gene expression is significantly regulated by nutritional status, suggesting a regulatory role of peripheral nesfatin-1 in energy homeostasis.

Role of the corticotropin‐releasing factor receptor type 2 in the control of food intake in mice: a meal pattern analysis

The actions of corticotropin-releasing factor (CRF) and related peptides are mediated by two receptors (CRF(1) and CRF(2)). The respective role of each subtype in the control of food intake remains poorly known. In the present study, we examined the quantity and microstructure of ingestive behavior of knockout (KO) mice lacking CRF(2) receptors and their wild-type (WT) littermates. Under basal conditions, CRF(2) KO mice showed increased nocturnal food intake, evident as an increased zenith in circadian cosinor analysis of food intake. Microstructure analysis revealed that this greater food intake reflected increased meal size, rather than meal frequency, suggesting a decreased satiating value of food. Following acute restraint stress, CRF(2) KO mice showed an intact immediate anorectic response with increased latency to eat and decreased meal size. However, CRF(2) deletion abolished the prolonged phase of restraint-induced anorexia. CRF(2) KO mice did not differ from WT controls in feeding responses to food deprivation or injection of ghrelin receptor agonists. Independent of genotype, food deprivation increased food intake, with dramatic changes in meal size, meal frequency, water : food ratio and eating rate. Acyl-ghrelin or BIM-28131, a potent ghrelin analog, dose-dependently stimulated food intake by increasing meal size (ghrelin, BIM-28131) and meal number (BIM-28131), while slowing the average eating rate (BIM-28131) similarly in WT and KO mice. These results suggest that the CRF(2) receptor is involved in the control of meal size during the active phase of eating and following acute exposure to stress.

Rapid and preferential activation of Fos protein in hypocretin/orexin neurons following the reversal of dehydration‐anorexia

Dehydration (DE)-anorexia is stimulated by chronic consumption of hypertonic saline. Spontaneous nocturnal food intake is markedly reduced with this treatment but is rapidly reversed upon the return of drinking water. Here we examined the neurons in the lateral hypothalamic area (LHA) of chronically dehydrated rats for their peptidergic phenotype, colocalization, and activation profiles following the rapid reversal of anorexia. To do this, we used double-labeling combinations of Fos immunocytochemistry and radioisotopic- and digoxigenin-labeled in situ hybridization. We found that lateral hypothalamic corticotropin-releasing hormone (CRH) neurons show extensive coexpression with neurotensin mRNA, but they are distinct from hypocretin/orexin and melanin-concentrating hormone (MCH) neurons. Chronic dehydration increases Fos-ir in large numbers of neurons in dorsal regions of the LHA. Some of these LHA neurons also show increased CRH, but not hypocretin/orexin or MCH gene expression, as dehydration-anorexia develops. Furthermore, the behavioral sequence of eating and increased activity exhibited by DE animals in the minutes following water drinking is accompanied by a further increase in the number of Fos-ir nuclei in the LHA. Increased Fos activation occurs in a significant number of LHA hypocretin/orexin neurons, but not CRH or MCH neurons, in the LHA. Together these data implicate CRH but not hypocretin/orexin or MCH neurons in the LHA in the motor events associated with dehydration. However, when water is returned, contributions to the network controlling responses evidently come from hypocretin/orexin, but not CRH or MCH, neurons in the LHA.

Circadian Expression of Genes Regulating Food Intake

The Agouti-related protein (AgRP), neuropeptide Y (NPY), proopiomelanocortin (POMC), cocaine and amphetamine-regulated transcript (CART), Orexin, melanin concentrating hormone (MCH), leptin, and its hypothalamic receptor (LR) are key regulators of food intake and energy homeostasis. In the present study, we examined the circadian expression profiles of these genes. We used quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) to measure mRNA levels, spectral analysis to evaluate periodicity, and correlation analysis to examine for associations with diurnal food intake. No gene in particular stood out as a strong candidate, but the overall circadian expression profiles of leptin and its hypothalamic receptor came close to statistically and graphically resembling the diurnal feeding behavior of mice. In mathematical terms, adrenal AgRP exhibited strong circadian expression and had the highest correlation with food intake, followed by leptin. Yet its highest point of expression occurred 8 hours after nocturnal food intake had peaked, suggesting that adrenal AgRP could not play a direct role in the initiation of nocturnal feeding; neither did hypothalamic AgRP, NPY, POMC, CART, Orexin, or MCH. These data show that ad libitum feeding in mice is influenced by complex central and peripheral circuits involving orexigenic and anorectic agents of which leptin and its hypothalamic receptor could play more prevalent roles.

Extended Access to Nicotine Self-Administration Leads to Dependence: Circadian Measures, Withdrawal Measures, and Extinction Behavior in Rats

The present study characterized nicotine intake, circadian patterns of food and water intake, precipitated somatic signs of withdrawal, and extinction of nicotine-seeking behavior in rats with 23-h access to intravenous self-administration (IVSA). Separate groups of animals were allowed access to nicotine IVSA (0.015, n = 9; 0.03, n = 14; 0.06, n = 16; mg/kg/0.1 ml infusion/s; fixed ratio 1) and trained to nosepoke for food and water 23 h/day for 40 consecutive days. Somatic signs of nicotine withdrawal were examined following saline or mecamylamine administration (1.5 mg/kg i.p.), and extinction of nicotine-seeking behavior was assessed. A dose-dependent decrease in lever responding and an increase in nicotine intake were observed, with the highest nicotine dose producing the lowest amount of lever responding and the highest amount of nicotine intake. Nicotine acutely reduced diurnal and nocturnal food intake, producing smaller and fewer meals, and an increased rate of eating. Differences in rate of nicotine intake between the light and dark phase decreased significantly, especially in rats receiving higher unit nicotine doses (0.03 and 0.06 mg/kg), along with long-term decreases in the circadian profile and amplitude of feeding. Mecamylamine precipitated robust withdrawal signs, the magnitude of which was positively correlated with the total amount of self-administered nicotine. Extinction of nicotine-seeking behavior was observed and was facilitated by removal of nicotine-associated cues. The results demonstrate that rats will self-administer nicotine to the point of producing dependence, as measured by somatic signs, resistance to extinction, and measures of food intake.

Delayed Satiety-Like Actions and Altered Feeding Microstructure by a Selective Type 2 Corticotropin-Releasing Factor Agonist in Rats: Intra-Hypothalamic Urocortin 3 Administration Reduces Food Intake by Prolonging the Post-Meal Interval

Brain corticotropin-releasing factor/urocortin (CRF/Ucn) systems are hypothesized to control feeding, with central administration of 'type 2' urocortins producing delayed anorexia. The present study sought to identify the receptor subtype, brain site, and behavioral mode of action through which Ucn 3 reduces nocturnal food intake in rats. Non-food-deprived male Wistar rats (n=176) were administered Ucn 3 into the lateral (LV) or fourth ventricle, or into the ventromedial or paraventricular nuclei of the hypothalamus (VMN, PVN) or the medial amygdala (MeA), regions in which Ucn 3 is expressed in proximity to CRF(2) receptors. LV Ucn 3 suppressed ingestion during the third-fourth post-injection hours. LV Ucn 3 anorexia was reversed by cotreatment with astressin(2)-B, a selective CRF(2) antagonist and not observed following equimole subcutaneous or fourth ventricle administration. Bilateral intra-VMN and intra-PVN infusion, more potently than LV infusion, reduced the quantity (57-73%) and duration of ingestion (32-68%) during the third-fourth post-infusion hours. LV, intra-PVN and intra-VMN infusion of Ucn 3 slowed the eating rate and reduced intake by prolonging the post-meal interval. Intra-VMN Ucn 3 reduced feeding bout size, and intra-PVN Ucn 3 reduced the regularity of eating from pellet to pellet. Ucn 3 effects were behaviorally specific, because minimal effective anorectic Ucn 3 doses did not alter drinking rate or promote a conditioned taste aversion, and site-specific, because intra-MeA Ucn 3 produced a nibbling pattern of more, but smaller meals without altering total intake. The results implicate the VMN and PVN of the hypothalamus as sites for Ucn 3-CRF(2) control of food intake.

Neuroendocrine Profiles Associated with Energy Intake, Sleep, and Stress in the Night Eating Syndrome

Night eating syndrome (NES) is characterized by evening hyperphagia and frequent awakenings with ingestion of food. It is associated with obesity and depressed mood. Greater understanding of hormonal influences on NES is desirable. Our objective was to evaluate 25-h profiles of hormones involved in energy balance, sleep, and stress in NES. Blood assays for glucose, insulin, ghrelin, leptin, melatonin, cortisol, TSH, and prolactin were sampled repeatedly among NES and control subjects. Food intake and depressive symptoms were assessed. Fifteen NES and 14 matched control participants stayed three nights in a General Clinical Research Center. We assessed differences between NES and control participants in the 25-h profiles of eight hormones. Nocturnal food intake was higher among NES participants, although their daily calorie intake was similar to that of controls. Reflecting their increased nocturnal intake, insulin (P < 0.001) and glucose levels (P = 0.07) among NES participants were higher than those of controls. Ghrelin levels were significantly lower in NES participants than in controls from 0100-0900 h (P = 0.003). Levels of plasma cortisol, melatonin, leptin, and prolactin did not differ between groups, but there was a trend for TSH levels (P = 0.07) to be higher during the 25 h in NES. NES participants had greater depressive symptoms than controls (P < 0.001). The differences in the levels of glucose, insulin, and ghrelin between NES and controls are closely associated with nocturnal food intake.

A new way of looking at eating

the analysis of meals, especially of spontaneous meals, has long had a problematic place in the behavioral neuroscience of eating. Only a small minority of eating researchers now, or ever, have concerned themselves with meal patterns. On first glance this seems paradoxical. After all, eating occurs

Physiological characteristics in SAMP1 and SAMP2 mice

We examined the physiological and behavioral characteristics of senescence-accelerated mice, SAMP1 and SAMP2 strains. In SAMP1 (P1) mice, we examined the diurnal rhythms of food and water intake, spontaneous motor activity (SMA), and spontaneous wheel running (SWR). Without a SWR exercise, food and water intake and SMA in the P1 mice were significantly higher during the light (L)-phase of the light/dark (LD) cycle than for the control SAMR1 (R1) mice. Additionally, the SWR and SMA activity split into two peaks. SWR exercise significantly increased the percentage of nocturnal food intake and SMA in P1 mice. Thus disordered diurnal rhythms in P1 mice can be normalized partially by the SWR exercise. In SAMP2 (P2) mice, we examined the physiological characteristics throughout the life span. The food and water intake, oxygen uptake ( V O 2), and SWR were higher than those of the control R1 mice, whereas the body weight, colonic temperature (Tco) and pelt weight were significantly lower. These results suggest that: (1) lower Tco in P2 mice is caused by morphological or insulated characteristics, and (2) and the short life span in P2 mice is caused by excessive food intake and higher V O 2 and SWR activity.

Repeated administration of the anorectic factor prolactin-releasing peptide leads to tolerance to its effects on energy homeostasis.

Central administration of a single dose of prolactin-releasing peptide (PrRP) causes a reduction in both fast-induced and nocturnal food intake and body weight gain. The aim of this study was to examine the effect of repeated administration of PrRP on energy homeostasis, including a measure of the expression of the mitochondrial uncoupling protein-1 (UCP-1) in brown adipose tissue. Conscious, free-feeding animals received central injections of PrRP (4 nmol icv) or vehicle. A single injection at 1000 caused a sustained hyperthermia over the 4-h test period and an increase in the expression of UCP-1 mRNA. Repeated, twice daily injection caused a reduction in body weight gain greater than that seen in pair-fed animals for the first 48-72 h. After 72 h, the animals became refractory to the actions of PrRP. The pair-fed group showed a reduction in UCP-1 mRNA expression at 48 h, which was reversed by PrRP treatment. This study indicates that PrRP exerts its effects on energy homeostasis in the short-medium term by reducing food intake and increasing energy expenditure.

Diurnal rhythm disorder of behavioral activity in SAMP1 mice is partially normalized by spontaneous wheel running

We examined the diurnal rhythms of food and water intake, spontaneous wheel running (SWR), and spontaneous motor activity (SMA) in the SAMP1 strain, a mouse model of accelerated senescence. Without SWR exercise, food, and water intake in the SAMP1 mice was significantly higher during the light (L)-phase of the light–darkness (LD) cycle than in the control SAMR1 strain. Additionally, SWR and SMA activity rhythms were split in SAMP1 mice, as demonstrated by the appearance of a secondary peak starting from the end of the dark (D)-phase. SWR exercise significantly increased the percentages of nocturnal food and water intake and SMA in the SAMP1 mice, although food and water intake did not reach the level of control SAMR1 mice. Thus, the disordered diurnal rhythms in SAMP1 mice can be normalized, even if only partially, by SWR exercise.

Inhibitory Effect of Apelin-12 on Nocturnal Food Intake in the Rat

Apelin, the endogenous peptide ligand of the APJ receptor, is expressed in brain regions implicated in food and water intake. This study reports for the first time, the effect of apelin-12, one of the most potent apelin peptides, on spontaneous (nocturnal) feeding. Randomised intracerebroventricular injection of 1, 3 and 10 nmol apelin-12 or saline vehicle, 10 min prior to lights out, led to dose-dependent reductions in food intake 2-4 h after injection (n=7; p<0.05). This suggests that apelin-12 exerts a delayed inhibitory effect on nocturnal feeding. Relative to saline, no effect was observed on total 24-h food intake post injection. In contrast, day-time administration of 10 nmol apelin-12 to satiated rats stimulated feeding (n=5-11; p<0.05); lower doses had no effect. No changes in water intake were observed after apelin-12. These results suggest that apelin is involved in the central control of feeding.

Evidence for involvement of neuropeptide Y and melanocortin systems in the hyperphagia of lactation in rats

Hypothalamic neuropeptide Y (NPY) systems are upregulated during lactation in rats. Because NPY is central to the hypothalamic control of energy balance, the present studies tested the hypothesis that NPY contributes to the marked hyperphagia during lactation. A 4-day infusion of [ d-tyr (27,36), d-thr (32)] NPY (27–36) ( d-NPY 27–36), a peptide analogue of NPY that antagonizes NPY-induced feeding, into the third ventricle at 1 μg/h transiently inhibited nocturnal feeding in nonlactating female rats. However, this antagonist had no effect on nocturnal feeding, but did transiently reduce food intake during the light hours, when infused into the third ventricle at the same dose in lactating females. An essentially similar pattern of results was obtained with chronic infusion into the third ventricle of the anorexigenic peptide α-melanocyte-stimulating hormone (α-MSH, 1 μg/h), in nonlactating and lactating rats. Both d-NPY 27–36 and α-MSH transiently reduced nocturnal food intake in lactating rats by approximately 10% when infused at the higher dose of 5 μg/h, and a marked inhibition of approximately 40% of both nocturnal and diurnal feeding was produced by a combined infusion of both at 5 μg/h. These results provide the first pharmacological evidence implicating specific neuromessengers in mediating the hyperphagia of lactation, and suggest that, while an action of NPY may contribute to the increased food intake seen in lactating animals, other systems are also involved. In particular, a reduction in melanocortin signaling during lactation may allow for an increased orexigenic influence of the agouti-related protein (AgRP), which is co-expressed with NPY.

PRL-releasing peptide interacts with leptin to reduce food intake and body weight.

PRL-releasing peptide (PrRP) is a novel anorexigen that reduces food intake and body weight gain in rats. In common with other anorexigens, PrRP mRNA expression is reduced during states of negative energy balance, i.e. lactation and fasting in female rats. In this study, we examined the interaction between PrRP and the adiposity signal, leptin, which interacts with a number of peptidergic systems in the brain to regulate energy homeostasis. Intracerebroventricular coadministration of 4 nmol PrRP and 1 microg leptin in rats resulted in additive reductions in nocturnal food intake and body weight gain and an increase in core body temperature compared with each peptide alone. We show also, by quantitative in situ hybridization, that PrRP mRNA is reduced in fasted male rats and obese Zucker rats, indicating that PrRP mRNA expression, like that of other anorexigens, may be regulated by leptin. Finally we show, using immunohistochemistry, that greater than 90% of PrRP neurons in all regions where PrRP is expressed contain leptin receptors. Thus, we provide evidence for PrRP neurons forming part of the leptin-sensitive brain circuitry involved in the regulation of food intake and energy homeostasis.

Chronic intraparaventricular nuclear administration of orexin A in male rats does not alter thyroid axis or uncoupling protein-1 in brown adipose tissue

Orexin A, synthesised in the posterolateral hypothalamus, has widespread distribution including the paraventricular nucleus (PVN), which is rich in thyrotropin-releasing hormone (TRH) neurones. Nerve fibres in the PVN synapse on neurones that send polysynaptic projections to brown adipose tissue (BAT), which is important in thermogenesis. A number of observations suggests orexin A may be involved in regulation of metabolism and thermogenesis. We investigated the effect of orexin A injected intracerebroventricularly (ICV) on thyroid-stimulating hormone (TSH) and thyroid hormones in male rats. We then examined the effect of chronic iPVN injections of orexin A on plasma TSH and uncoupling protein-1 (UCP-1) protein in BAT. Orexin A (3 nmol) administered ICV significantly suppressed plasma TSH at 10 and 90 min. Orexin A (0.3 nmol) administered into the PVN twice daily for 3 days significantly increased day-time 2-h food intake, but did not significantly alter nocturnal food intake. Though chronic iPVN orexin A altered diurnal food intake, there was no effect on 24-h food intake or body weight. Furthermore, orexin A administered chronically into the PVN did not alter UCP-1 level in BAT, or plasma hormones relative to saline injected animals. Chronic iPVN orexin A does not appear to influence thermogenesis through activation of UCP-1 or the thyroid axis.

Circadian variation in the effects of nitric oxide synthase inhibitors on body temperature, feeding and activity in rats.

We have investigated whether there is circadian variation in the effects of nitric oxide synthase inhibitors on body temperature, physical activity and feeding. We used nocturnally active Sprague-Dawley rats, housed at approximately 24 degrees C with a 12:12 h light:dark cycle (lights on 07:00 hours) and provided with food and water ad libitum. Nitric oxide synthesis was inhibited by intraperitoneal injection of the unspecific nitric oxide synthase inhibitor N-nitro- L-arginine methyl ester ( L-NAME, 100, 50, 25, 10 mg/kg), or the relatively selective inducible nitric oxide synthase inhibitor aminoguanidine (100, 50 mg/kg), during the day ( approximately 09:00 hours) or night ( approximately 21:00 hours). Body temperature and physical activity were measured using radiotelemetry, while food intake was calculated by weighing each animal's food before as well as 12 and 24 h after each injection. We found that daytime injection of L-NAME and aminoguanidine had no effect on daytime body temperature. However, daytime injection of both drugs did decrease nocturnal food intake ( P<0.05) and activity ( P<0.05). When injected at night, L-NAME reduced night-time body temperature ( P<0.01), activity ( P<0.05) and food intake ( P<0.05) in a dose-dependent manner, but night-time injection of aminoguanidine inhibited only night-time activity ( P<0.05). The effects of nitric oxide synthase inhibition on body temperature, feeding and activity therefore are primarily a consequence of inhibiting constitutively expressed nitric oxide synthase, and are subject to circadian variation.

Anorectic effect of leptin is mediated by hypothalamic corticotropin-releasing hormone, but not by urocortin, in rats

Leptin is a key afferent signal that decreases food intake and increases energy expenditure by acting on the specific receptors in the hypothalamus. Corticotropin-releasing hormone (CRH) and its homologous peptide, urocortin, are also known to have a potent anorectic effect when given intracranially. To determine possible involvement of these two peptides in the leptin-induced anorexia, in the present study, food intake was measured in rats pretreated with antibodies against CRH and urocortin. In the control rats without antibody pretreatment, intracerebroventricular (icv) injection of leptin (0.1–1 μg/rat) suppressed nocturnal food intake. The anorectic effect of leptin was substantially attenuated in rats pretreated with icv injection of an anti-CRH antibody, but not with an anti-urocortin antibody. These results suggest that the anorectic effect of leptin is mediated by CRH, but not by urocortin, in the hypothalamus.