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Sprague-Dawley rats over-consume calories over a 10 week period and develop diet-induced obesity (c. 100 g body weight differential vs controls) when fed a control pellet diet supplemented with chocolate Ensure liquid. Subsequent withdrawal of Ensure immediately reduces caloric intake by more than 50%, and results in weight loss, despite control pellet being available ad libitum. To assess the molecular underpinnings of this phenomenon, brains were processed for energy balance and food reward-related gene expression analysis at two time points, 24 h and 4 days after the withdrawal of Ensure, when energy intake was suppressed. Gene expression levels in hypothalamic arcuate nucleus and forebrain nucleus accumbens were compared with rats pair-fed to the same energy intake, i.e. imposed negative energy balance, and to controls fed control pellet ad libitum throughout. Cumulative energy intake was approximately 50% lower across the 4 day post-Ensure period, giving rise to a small reduction in body weight although body adiposity and blood leptin remained elevated (c. 100% and 50%, respectively vs controls) in rats that had previously been fed Ensure. In contrast, pair-feeding reduced blood insulin and leptin by 33% and 55%, respectively. Hypothalamic expression of neuropeptide Y and agouti-related peptide was down-regulated at 24 h in rats previously fed Ensure, indicative of the apparent counter-regulatory changes seen in diet-induced obesity, but was normalised between the 24 h and 4 day time points. By contrast, the effect of cumulative negative energy balance in the pair-fed groups increased with time, up-regulating expression of the orexigenic neuropeptides. There was also a reduction of suppressor of cytokine signalling-3 gene expression in pair-fed groups where leptin levels were low. There were no changes in opioid, dopamine receptor or cannabinoid receptor expression in the nucleus accumbens. Feedback from diet-induced obesity appears to drive voluntary hypophagia upon withdrawal of palatable diet, and to override signals from intake restriction that would otherwise set in train an anabolic drive., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Maternal diet ingested during gestation can profoundly alter production and action of hypothalamic neuropeptides involved in feeding and body weight regulation. In this study, we set out to simulate, in a rat model, modifications to feeding habit often observed in pregnant women. Gestating dams were fed a restricted normal diet with the opportunity to complete their energy requirements with either a high-fat (HF) or a high-carbohydrate (HC) diet. Growth and hypothalamic feeding peptides were measured in the offspring at 3 (weaning) and 20 weeks of age. At weaning, body weight was lower in HC pups than in HF pups or control (Ca) pups born to dams fed control diet ad libitum. Expression of neuropeptide Y (NPY) and AgRP mRNA in the arcuate nucleus were increased in HC pups vs Ca and HF pups. By 20 weeks of age, body weight differentials had disappeared, and there was no differences in NPY and AgRP gene expression, although POMC expression was lower in HC rats than in HF rats. NPY and orexin peptide concentrations in the paraventricular nucleus at this age were higher in HC rats than in Ca and HF rats. In HC rats, there was also a greater positive gradient of peptide concentration between the zone of release and the zone of synthesis for NPY and orexin. The early up-regulation of orexigenic peptides in HC rats may be a compensatory adjustment to low body weight. This persisting overactive orexigenic drive might have deleterious metabolic effects in an obesogenic environment at adulthood., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Prenatal nutritional conditions can predispose to development of obesity and metabolic syndrome in adulthood. Gestation with its important modifications in hormonal status is a period of changes in normal feeding habits with pulses of consumption or avoidance of certain categories of food. We tried to mimic in an animal model some changes in food consumption patterns observed in pregnant women. For this purpose, Long-Evans female rats were fed during the dark period, their usual pre-gestational food quantity, and were allowed to complete their daily intake with either a restricted control (Cr), high-fat (HF), or high-carbohydrate (HC) diet available ad libitum during the light period. Dams fed a control diet ad libitum (Ca) served as controls. Body weight and composition, food intake, and metabolic hormones (insulin, leptin) were recorded in male offspring until 20 weeks after birth. Cr and HC females ate less than Ca females (-16%; p < 0.001) and their offspring presented a weight deficit from birth until 6 (HC group) and 10 (Cr group) weeks of age (p < 0.05 or less). Plasma leptin corresponded to low body weight in Cr offspring, but was increased in HC offspring that in addition, had increased plasma insulin, blood glucose, and subcutaneous adipose tissue mass. HF dams ate more than Ca dams (+13%; p < 0.001), but plasma leptin and insulin were similar in their offspring. Hypothalamic Ob-Rb expression was increased in Cr, HC, and HF offspring (+33-100% vs Ca; p < 0.05 or less). HC supplement ingestion during gestation therefore leads to insulin and leptin resistance in adult offspring independently of lower birth weight. These hormonal changes characterize obesity-prone animals. We therefore suggest that attention should be paid to the carbohydrate snacking and overall carbohydrate content in the diet during the last weeks (or months) preceding delivery to limit development of later metabolic disorders in offspring.

The causes of post-restriction hyperphagia (PRH) represent a target for drug-based therapies to prevent obesity. However, the factors causing PRH are poorly understood. We show that, in mice, the extent of PRH was independent of the time under restriction, but depended on its severity, suggesting that PRH was driven by signals from altered body composition. Signals related to fat mass were important drivers. Circulating levels of leptin and TNFα were significantly depleted following caloric restriction (CR). We experimentally repleted their levels to match those of controls, and found that in both treatment groups the level of PRH was significantly blunted. These data establish a role for TNFα and leptin in the non-pathological regulation of energy homeostasis. Signals from adipose tissue, including but not limited to leptin and TNFα, regulate PRH and might be targets for therapies that support people engaged in CR to reduce obesity.

Central regulation of energy balance in seasonal mammals such as the Siberian hamster is dependent on the precise integration of short-term satiety information arising from the gastrointestinal tract with long-term signals on the status of available energy reserves (e.g. leptin) and prevailing photoperiod. Within the central nervous system, the brainstem nucleus of the solitary tract (NTS) and the parabrachial nucleus (PBN) are major relay nuclei that transmit information from the gastrointestinal tract to higher forebrain centres. We extended studies on the seasonal programming of the hypothalamus to examine the effect of the photoperiod on neuropeptidergic circuitries of this gut-brain axis. In the NTS and PBN we performed gene expression and immunoreactivity (-ir) studies on selected satiety-related neuropeptides and receptors: alpha-melanocyte stimulating hormone, melanocortin-3 receptor, melanocortin-4 receptor (MC4-R), growth hormone secretagogue-receptor, cocaine- and amphetamine-regulated transcript, preproglucagon (PPG), glucagon-like peptide 1 (GLP-1), cholecystokinin (CCK), peptide YY, galanin, neurotensin, and corticotrophin releasing hormone (CRH). Gene expression of PPG and MC4-R, and -ir of CCK and GLP-1, in the NTS were up-regulated after 14 weeks in long-day photoperiod (16 h light:8 h dark) compared to short-days (8 h light:16 h dark), whereas CRH-ir and NT-ir were increased in short-days within the PBN. We suggest that brainstem neuroendocrine mechanisms contribute to the long-term regulation of body mass in the Siberian hamster by a photoperiod-related modulation of satiety signalling.

Epidemiological studies show a link between low birthweight and increased obesity. In contrast, slow growth during the lactation period reduces obesity risk. The present study investigates the potential underlying mechanisms of these observations. Rats were established as follows: (i) control animals [offspring of control dams fed a 20% (w/v) protein diet], (ii) recuperated animals [offspring of dams fed an isocaloric low-protein (8%, w/v) diet during pregnancy and nursed by control dams], and (iii) postnatal low protein animals (offspring of control dams nursed by low-protein-fed dams). Serum and brains were collected from fed and fasted animals at weaning. Expression of hypothalamic energy balance genes was assessed using in situ hybridization. Recuperated pups were smaller at birth, but caught up with controls by day 21 and gained more weight than controls between weaning and 12 weeks of age (P<0.05). At 21 days, they were hypoleptinaemic compared with controls in the fed state, with generally comparable hypothalamic gene expression. Postnatal low protein offspring had significantly lower body weights than controls at weaning and 12 weeks of age (P<0.001). At 21 days, they were hypoglycaemic, hypoinsulinaemic and hypoleptinaemic. Leptin receptor gene expression in the arcuate nucleus was increased in postnatal low protein animals compared with controls. Consistent with hypoleptinaemia, hypothalamic gene expression for the orexigenic neuropeptides NPY (neuropeptide Y) and AgRP (Agouti-related peptide) was increased, and that for the anorexigenic neuropeptides POMC (pro-opiomelanocortin) and CART (cocaine- and amphetamine-regulated transcript) was decreased. These results suggest that the early nutritional environment can affect the development of energy balance circuits and consequently obesity risk.

The mechanism(s) involved in the regulation of the seasonal-appropriate body weight of the Siberian hamster are currently unknown. We have identified photoperiodically regulated genes including VGF in a sub-region of the arcuate nucleus termed the dorsomedial posterior arcuate (dmpARC). Gene expression changes in this nucleus so far account for a significant number of those reported as photoperiodically regulated and are therefore likely to contribute to seasonal physiological responses of the hamsters. The present study aimed to identify additional genes expressed in the dmpARC regulated by photoperiod that could be involved in regulating the activity of this nucleus with respect to seasonal physiology of the Siberian hamster. Using laser capture microdissection coupled with a microarray analysis and a candidate gene approach, we have identified several photoperiodically regulated genes in the dmpARC that are known to have roles in secretory and intracellular signalling pathways. These include secretogranin (sg) III and SgVI (secretory pathway), melanocortin 3 receptor (MC3-R) and serotonin (5-HT) receptors 2A and 7 (signalling pathway), all of which increase in expression under a short photoperiod. The spatial relationship between receptor signalling and potential secretory pathways was investigated by dual in situ hybridisation, which revealed that 5-HT2A and 5-HT7 receptors are expressed in neurones expressing VGF mRNA and that a sub-population (approximately 40%) of these neurones express MC3-R. These gene expression changes in dmpARC neurones may reflect the functional requirement of these neurones for seasonal physiological responses of the hamster.

A wealth of detailed mechanistic information relating to obesity and body weight regulation has emerged from study of single gene mutation models, and continues to be generated by engineered rodent models targeting specific genes. However, as an early step in translational research, many researchers are turning to models of diet-induced obesity. Interpretation of data generated from such models is not aided by the variety of diets and rodent strains employed in these studies and a strong case could be made for rationalisation. Differences in experimental protocol, which may deploy a single obligatory solid diet, a choice of solid diets, or liquid/solid combinations, and which may or may not allow a preferred macronutrient composition to be selected, mean that different models of diet-induced obesity achieve that obesity by different routes. The priority should be to mimic the palatability- and choice-driven over-consumption that probably underlies the majority of human obesity. Some of the hypothalamic energy balance genes apparently 'recognise' developing diet-induced obesity as indicated by counter-regulatory changes in expression levels. However, substantial changes in gene expression on long-term exposure to obesogenic diets are not able to prevent weight gain. Forebrain reward systems are widely assumed to be overriding hypothalamic homeostatic energy balance systems under these circumstances. More mechanism-based research at the homeostatic/reward/diet interface may enable diets to be manipulated with therapeutic benefit, or define the contribution of these interactions to susceptibility to diet-induced obesity.

Obesity prevalence is increasing worldwide, leading to increased morbidity, mortality and health care costs due to its role as a key risk factor in many diseases. Early life growth and nutrition has been implicated in determining susceptibility to obesity in both childhood and adulthood; however, the mechanisms underlying this link are poorly understood. A variety of animal models have been established to try and uncover the developmental programming effects of maternal early life nutrition on energy balance regulation and the mechanisms behind them.

Previously, 40% food restriction of male Siberian hamsters over 21 days in short-day (SD) photoperiod induced characteristic changes in expression of hypothalamic arcuate nucleus energy balance genes; mRNAs for neuropeptide Y, agouti-related peptide, and leptin receptor were upregulated, and those of proopiomelanocortin and cocaine- and amphetamine-regulated transcript were depressed. The present study examined the effect of refeeding hamsters for 6 days (approximately 50% recovery of weight differential) or 19 days (resumption of appropriate weight trajectory). Hyperphagia continued throughout refeeding, but differences in fat pad weights and leptin levels had disappeared after 19 days. Cocaine- and amphetamine-regulated transcript gene expression was depressed by prior restriction in both refed groups. The depressive effect of prior restriction on proopiomelanocortin gene expression had disappeared after 19 days of refeeding. There was no effect of prior food restriction on neuropeptide Y or agouti-related peptide gene expression. Expression of the anorexigenic brain-derived neurotrophic factor was downregulated in the ventromedial nucleus after SD exposure for 12 wk. In the SD food restriction study, there were effects of photoperiod on brain-derived neurotrophic factor gene expression but not of prior food restriction. Hypothalamic energy balance genes in the hamster respond asynchronously to return to a seasonally appropriate body weight. The achievement of this weight rather than the weight at which caloric restriction was imposed is the critical factor. The differential responses of hypothalamic energy balance genes to food restriction and refeeding are poorly characterized in any species, a critical issue given their potential relevance to human weight loss strategies that involve caloric restriction.

Seasonal adaptations in physiology exhibited by many animals involve an interface between biological timing and specific neuroendocrine systems, but the molecular basis of this interface is unknown. In this study of Siberian hamsters, we show that the availability of thyroid hormone within the hypothalamus is a key determinant of seasonal transitions. The expression of the gene encoding type III deiodinase (Dio3) and Dio3 activity in vivo (catabolism of T(4) and T(3)) is dynamically and temporally regulated by photoperiod, consistent with the loss of hypothalamic T(3) concentrations under short photoperiods. Chronic replacement of T(3) in the hypothalamus of male hamsters exposed to short photoperiods, thus bypassing synthetic or catabolic deiodinase enzymes located in cells of the ependyma of the third ventricle, prevented the onset of short-day physiology: hamsters maintained a long-day body weight phenotype and failed to undergo testicular and epididymal regression. However, pelage moult to a winter coat was not affected. Type II deiodinase gene expression was not regulated by photoperiod in these hamsters. Collectively, these data point to a pivotal role for hypothalamic DIO3 and T(3) catabolism in seasonal cycles of body weight and reproduction in mammals.

Contemporary foods and beverages that constitute the diets of adults and children almost certainly contribute to the obesity problem. To develop a model of childhood obesity, we examined the effects of feeding juvenile rats 2 solid diets, either alone or in combination [nonpurified control diet (C), high-energy (HE), or C+HE] with or without the liquid supplement Ensure (EN). Rats were fed C until 4 wk of age and then were assigned to 1 of 6 weight-matched groups that were fed C, HE, C+HE, C+EN, HE+EN, or C+HE+EN for 5 wk. EN accelerated weight gain and increased energy intake and adiposity irrespective of the solid diet consumed. Serum leptin concentrations were increased after the consumption of all diets when compared with C rats, but there was dissociation between leptin levels and adiposity. The type of solid diet had no effect on the expression of a panel of hypothalamic genes except for glutamate-decarboxylase-67. EN decreased mRNA for agouti-related peptide and neuropeptide Y in the arcuate nucleus and DYN in the paraventricular nucleus. Dynorphin and CART mRNA were decreased in the supraoptic retrochiasmatic nucleus. The reduction in orexigenic signaling in the hypothalamus suggests that overconsumption of EN is sensed by the hypothalamus but that any initiated physiological responses fail to compensate effectively and may be negated or overwhelmed by other systems. Providing diets in solid and liquid form, with choice, mimics more closely the human environment. Understanding the interactions between these diets and peripheral and central energy balance systems could be crucial in unraveling the events underlying human obesity and its early development.

We have previously shown that cold-acclimated (8 degrees C) male field voles (Microtus agrestis) transferred from short day (SD, 8 h light) to long day (LD, 16 h light) photoperiod exhibit an increase in body mass lasting 4 weeks, after which they stabilise at a new plateau approximately 7.5 g (24.8%) higher than animals maintained in SD. By infusing voles with exogenous leptin, we have also demonstrated that SD voles respond to the hormone by reducing body mass and food intake, whereas LD animals increasing body mass are resistant to leptin treatment. In the present study, we investigated whether seasonal changes in body mass could be linked to modulation of the leptin signal by suppressor of cytokine signalling-3 (SOCS3). We used in situ hybridisation to examine hypothalamic arcuate nucleus (ARC) expression of SOCS3, neuropeptide Y (NPY), agouti-related peptide (AgRP), pro-opiomelanocortin (POMC) and cocaine- and amphetamine-regulated transcript (CART) genes in 90 voles exposed to either SD or LD for up to 11 weeks. LD voles increasing body mass had significantly higher levels of SOCS3 mRNA than SD or LD voles with a stable body mass. There were no associated changes in expression of NPY, AgRP, POMC and CART genes. These results suggest that voles that regulate body mass at either the lower (SD) or upper (LD) plateau remain sensitive to leptin action, whereas SOCS3-mediated leptin resistance is a short-term mechanism that enables animals to move between the stable body mass plateaus. Our data provide evidence that expression of SOCS3 in the ARC is involved in the modulation of the strength of the leptin signal to facilitate seasonal cycles in body mass and adiposity.

Outbred male Sprague-Dawley rats were provided with one of the four flavours of the liquid diet, Ensure, in addition to chow pellets, to examine whether differences in flavour lead to differences in energy intake i.e. degree of over-consumption. For half the rats, the Ensure supplement was provided for 14 days and then withdrawn for the final 8 days of the study, whereas the remaining animals were allowed to consume Ensure for 22 days. All four flavours of Ensure, chocolate, vanilla, coffee and asparagus, induced a sustained increase in daily energy intake of approximately 15%. There was an effect of flavour on initial consumption of the Ensure diet, with coffee and asparagus flavours being consumed less avidly than vanilla or chocolate. However, this effect was short-lived. Overall, there was no effect of flavour on body weight gain, energy intake from Ensure, total energy intake, body composition, or measured blood hormones and metabolites. Withdrawal of Ensure resulted in reductions in body weight gain, total energy intake, fat but not lean tissue mass, and concentrations of blood leptin, non-esterified fatty acids and triglycerides, but there was no effect of the flavour of Ensure previously supplied on any of these parameters. The ability of the liquid diet, Ensure, to stimulate long-term caloric over-consumption is not due to its flavouring. Rather, other attributes of Ensure must be more important, such as its intrinsic flavour, liquid formulation, macronutrient composition, and ease of ingestion, digestion and absorption.

A remarkable feature of the seasonal adaptation displayed by the Siberian hamster (Phodopus sungorus) is the ability to decrease food intake and body weight (by up to 40%) in response to shortening photoperiod. The regulating neuroendocrine systems involved in this adaptation and their neuroanatomical and molecular bases are poorly understood. We investigated the effect of photoperiod on the expression of prohormone convertases 1 (PC1/3) and 2 (PC2) and the endoproteolytic processing of the neuropeptide precursor pro-opiomelanocortin (POMC) within key energy balance regulating centres of the hypothalamus. We compared mRNA levels and protein distribution of PC1/3, PC2, POMC, adrenocorticotrophic hormone (ACTH), alpha-melanocyte-stimulating hormone (MSH), beta-endorphin and orexin-A in selected hypothalamic areas of long day (LD, 16:8 h light:dark), short day (SD, 8:16 h light:dark) and natural-day (ND, photoperiod depending on time of the year) acclimated Siberian hamsters. The gene expression of PC2 was significantly higher within the arcuate nucleus (ARC, P < 0.01) in SD and in ND (versus LD), and is reflected in the day length profile between October and April in the latter. PC1/3 gene expression in the ARC and lateral hypothalamus was higher in ND but not in SD compared to the respective LD controls. The immunoreactivity of PC1/3 cleaved neuropeptide ACTH in the ARC and PC1/3-colocalised orexin-A in the lateral hypothalamus were not affected by photoperiod changes. However, increased levels of PC2 mRNA and protein were associated with higher abundance of the mature neuropeptides alpha-MSH and beta-endorphin (P < 0.01) in SD. This study provides a possible explanation for previous paradoxical findings showing lower food intake in SD associated with decreased POMC mRNA levels. Our results suggest that a major part of neuroendocrine body weight control in seasonal adaptation may be effected by post-translational processing mediated by the prohormone convertases PC1/3 and PC2, in addition to regulation of gene expression of neuropeptide precursors.

Energy dense, high fat, high sugar, foods and beverages in our diet are a major contributor to the escalating global obesity problem. Here, we examine the physiological and neuroendocrine effects of feeding rats a solid high-energy (HE) diet with or without a liquid supplement (Ensure) and the consequence of subsequently transferring animals back to chow (C). Outbred Sprague-Dawley rats were fed C until 49-56 days of age, and then transferred a HE diet for 3 weeks before allocation to one of two weight-matched groups. Over the next 10 weeks, one group remained on HE diet, whereas the other had access to the liquid diet, chocolate Ensure (EN), in addition to HE diet (HE + EN). Half the rats from each group were then killed, and the remainder were returned to C for 3 weeks. Supplementation of the HE diet with EN accelerated weight gain and increased daily energy intake, adipose tissue mass, and circulating leptin levels. Transferring animals back to C caused a decrease in bodyweight in the HE + EN group, whereas HE animals were weight stable. Both groups also exhibited voluntary hypophagia, although the magnitude and duration of this response was greater in HE + EN animals. The only effect of Ensure on the hypothalamic genes studied was on tyrosine kinase B expression in the ventromedial hypothalamic nucleus (VMH), which was increased in rats given the supplement. Withdrawal of the obesogenic diets decreased gene expression for cocaine-and-amphetamine regulated transcript (CART) and dynorphin (DYN) in the arcuate nucleus (ARC), and DYN and brain-derived neurotrophic factor (BDNF) in the VMH, whereas neuropeptide Y (NPY) gene expression in the ARC was increased. These changes were independent of previous dietary history. EN supplementation generates distinct physiological responses, yet has a minimal effect on hypothalamic neuropeptide or receptor gene expression, possibly due to the development of leptin resistance. Withdrawal of obesogenic diets induces changes in the gene expression consistent with NPY, CART and BDNF attempting to oppose weight gain on either HE or HE + EN.

The SD (Sprague-Dawley) rat model of DIO (diet-induced obesity) is reported to exhibit a clear segregation into susceptible and resistant subpopulations shortly after transfer to a HE (high energy) diet. This does not appear to be the case for rats sourced in the U.K., where body weight gain on obesogenic HE diet is normally distributed, as might be anticipated for a polygenic trait in an outbred population. Many of the energy balance effects of dietary manipulation in this model (e.g. supplementation of HE diet with the liquid diet, Ensure; energy intake and defence of body weight following withdrawal of obesogenic diet) appear to be characteristics of the diets being manipulated rather than subject traits. The activities of energy balance-related hypothalamic signals are affected by diet and the development of DIO, but may not be able to differentiate between different diets and the relative levels of obesity that develop.

Nutritional feedback provided by systemic hormones, such as insulin and leptin, influences reproductive neuroendocrine output within the hypothalamus, yet the mechanisms and their interaction with photoperiodic cues remain unresolved in seasonal species. Here, peripheral glucose (G) infusion was used to increase endogenous concentrations of insulin and leptin in food-restricted sheep kept in either long-day (LD) or short-day (SD) photoperiod, and responses were examined in terms of pulsatile luteinising hormone (LH) (gonadotrophin-releasing hormone by inference) output and hypothalamic gene expression for nutritionally sensitive neuropeptides and receptors. We addressed the hypothesis that these hypothalamic responses were correlated and influenced by photoperiod. Oestradiol-implanted, castrated male sheep were kept 16 weeks in SD (8 h light/day) or LD (16 h light/day) and then transferred to the opposite photoperiods for 8 weeks, during which food was restricted to 90% requirement to maintain body weight (maintenance). For the final 6 days, food was reduced to 75% maintenance, and sheep in both photoperiods were infused intravenously with G (60 mM/h) or saline (S) (n = 8/group). G-infused sheep had higher mean plasma concentrations of G, insulin and leptin than S-infused sheep, with no effect of photoperiod. In LD, but not in SD, G infusion increased LH pulse frequency and pulse amplitude. In LD, but not in SD, gene expression in the hypothalamic arcuate nucleus was lower in G- than S-infused sheep for neuropeptide Y (NPY) and agouti-related peptide (AGRP) and was higher in G- than S-infused sheep for pro-opiomelanocortin (POMC). Gene expression for leptin and insulin receptors was not affected by photoperiod or infusion. These results are consistent with the involvement of NPY, AGRP and POMC in mediating the reproductive neuroendocrine response to increased systemic nutritional feedback, and they support the hypothesis that hypothalamic responses to nutritional feedback are influenced by photoperiod in sheep.

Obesity is an escalating problem in Western societies. Susceptibility to weight gain within an obesogenic environment is variable. It remains unclear how the range of weight gain responses are generated. It is possible that an individual's immediate and/or sustained appetite for apparently palatable foods, or metabolic adaptations to a new diet could be important. The present study therefore examined the short- to medium-term effects of a high-energy (HE) diet on bodyweight, food intake, and energy balance-related signalling systems. Sprague-Dawley rats were fed either chow or an HE diet for 12 h, 24 h, 48 h or 14 days. Blood hormones and metabolites were assayed, and expression of uncoupling protein-1 (UCP-1) and hypothalamic energy-balance related genes were determined by Northern blotting or in situ hybridisation, respectively. Short-term exposure (12 h, 24 h, 48 h) to the HE diet had no effect on grams of food consumed, but caloric intake was increased. Exposure to HE diet for 14 days (medium term) established a bodyweight differential of 7.7 g, and animals exhibited a transient increase in caloric intake of 5 days duration. Terminal levels of leptin, insulin, glucose and non-esterified fatty acids (NEFAs) were all increased in HE-fed animals. UCP-1 mRNA was elevated in interscapular brown adipose tissue from HE-fed rats only at 12 h. Cocaine and amphetamine-regulated transcript (CART) and Mc4R gene expression in the hypothalamus were increased after 12 h and 24 h on an HE diet, respectively. The rats appear to passively over-consume calories as a result of consuming a similar weight of a more energy dense food. This evokes physiological responses, which adjust caloric intake over several days. Circulating NEFA and insulin concentrations, UCP-1, Mc4R and CART gene expression are increased as an immediate consequence of consuming HE diet, and may be involved in countering hypercaloric intake. Circulating leptin is increased in the HE-fed animals after 48 h, reflecting their increasing adiposity.

Ghrelin is an endogenous ligand for the growth hormone secretagogue receptor (GHSR). However, the functional interaction of ligand and receptor is not very well understood. We demonstrate that GHSR mRNA is up-regulated after food deprivation (48 h) in the hypothalamic arcuate nucleus and ventromedial nucleus of the seasonal Siberian hamster, Phodopus sungorus. This increase is accompanied by a two-fold elevation of circulating ghrelin concentration. Chronic changes in feeding state imposed by food restriction over a period of 12 weeks during long day-length induced increased GHSR gene expression, whereas food restriction for 6 weeks had no effect. Phodopus sungorus reveals remarkable seasonal changes in body weight, fat mass and circulating leptin levels. Ghrelin is generally regarded as having opposing effects on appetite and body weight with respect to those exhibited by leptin. However, our study revealed that seasonal adaptations were not accompanied by changes in either GHSR gene expression or circulating ghrelin concentration. Therefore, we suggest that ghrelin only plays a minor role in modulating long-term seasonal body weight cycles. Our findings imply that ghrelin predominantly acts as a short-term regulator of feeding.

The anorexigenic and orexigenic hormones leptin and ghrelin act in opposition to one another. When leptin signaling is reduced, as in the Zucker fatty rat, or when circulating ghrelin is increased during fasting, the effect of ghrelin becomes more dominant, indicating an influence of both hormones on ghrelin action. This effect could be mediated via the level of expression of ghrelin receptor (growth hormone secretagogue receptor [GHS-R]). For testing this, GHS-R expression was measured using in situ hybridization in Zucker fatty versus lean rats; in fed versus fasted (48 h) rats, treated with either ghrelin or leptin; and in GH-deficient, dwarf versus control rats. In the arcuate nuclei of the Zucker fatty rat and in fasted rats, GHS-R expression is significantly increased. A single leptin intracerebroventricular injection attenuated the fasting-induced increase in GHS-R but had no effect in fed rats 2 h after injection, whereas leptin infusion for 24 h or longer significantly decreased GHS-R expression in fed rats. Ghrelin significantly increased GHS-R expression but not in dwarf rats. These results show that the level of GHS-R expression in the ARC is reduced by leptin and increased by ghrelin and that the effect of ghrelin may be GH dependent.

The incidence of obesity, with its associated health risks, is on the increase throughout the western world affecting all age groups, including children. The typical western diet is high in fat and sugar and low in complex carbohydrates. This study looks at the effects of feeding an equivalent high-energy (HE) diet to growing rats. Juvenile male Sprague-Dawley rats that were fed an HE (18.9 kJ/g) diet starting approximately 10 d after weaning gained less weight than littermates fed a nonpurified (14 kJ/g) diet. Despite an initial hyperphagia following the change in diet, HE rats also consumed less energy. Although they exhibited reduced weight gain, HE rats were relatively obese; fat pad weights were elevated for all 4 dissected depots. HE-fed rats exhibited symptoms of developing metabolic syndrome with elevated plasma concentrations of glucose, triglycerides, nonesterified fatty acids, insulin, and leptin. In addition, leptin receptor gene expression in the hypothalamic arcuate nucleus (ARC) and ventromedial nucleus of HE rats was reduced. Consistent with the elevated serum leptin and other peripheral signals in HE rats, hypothalamic gene expression for the orexigenic neuropeptides, neuropeptide Y (ARC and dorsomedial nucleus), and agouti-related peptide (AgRP), was reduced. This reduction in orexigenic signaling and decline in energy intake is consistent with an apparent attempt to counter the further development of an obese state in rats consuming an energy-dense diet. The juvenile Sprague-Dawley rat has potential in the development of a model of childhood diet-induced obesity.

Objective: To investigate the effect of a high-energy (HE) diet on caloric intake, body weight, and related parameters in outbred male Sprague-Dawley (SD) rats., Research Methods and Procedures: Twenty-eight SD rats were fed either chow (C) for 19 weeks or HE diet for 14 weeks and then C for 5 weeks. Blood hormones and metabolites were assayed, and expression of uncoupling protein-1 and hypothalamic energy-balance-related genes were determined by Northern blotting and in situ hybridization, respectively., Results: HE rats gained body weight more rapidly than C animals with a range of weight gains, but there was no evidence that weight gain was bimodally distributed. Caloric intake was transiently elevated after introduction of the HE diet. Transfer of HE rats back to C resulted in a drop in caloric intake, but a stable body weight. In terminal analysis, two of four dissected adipose tissue depots were heavier in rats that had previously been fed HE diet. Blood leptin, insulin, glucose, and nonesterified fatty acids were not different between the groups. Uncoupling protein-1 mRNA was elevated in interscapular brown adipose tissue from HE rats. There was a trend for agouti-related peptide mRNA in the hypothalamic arcuate nucleus to be higher in HE rats., Discussion: Contrary to other studies of the SD rat on HE diet, body weight and other measured parameters were normally distributed. There was no segregation into two distinct populations on the basis of susceptibility to diet-induced obesity. This characteristic may be dependent on the breeding colony from which animals were sourced.

There is a growing literature on the role of leptin in appetite and neuroendocrine regulation in domestic ruminants. Circulating leptin concentration is higher in fat than in thin sheep, is reduced by chronic underfeeding and is higher in sheep subjected to long-day rather than short-day photoperiods. Leptin is reduced acutely by fasting and increases after meals so that there are long- and short-term components to the systemic leptin signal. Nutritional stimulation of reproductive neuroendocrine output is associated with increased circulating concentrations of leptin; peripheral leptin administration restores LH secretion in fasted sheep, and leptin is permissive (although not a trigger) for puberty. Intracerebroventricular (i.c.v.) pharmacological leptin infusion stimulates LH in underfed but not in well-fed sheep, and reduces food intake in well-fed sheep. A single i.c.v. pharmacological injection or physiological infusion of leptin stimulates LH in well-fed sheep, with or without a concomitant decrease in appetite. Furthermore, these appetite and LH responses are differentially affected by photoperiod, indicating that different neuronal pathways may mediate the two responses. Hypothalamic leptin receptors co-localize with orexigenic and anorexigenic neurones, some of which contact GnRH cells, but the confluence of leptin signalling with photoperiod (melatonin) signalling remains unresolved. Photoperiod-entrained sheep provide potential models of altered central leptin sensitivity, in which downstream mechanisms regulating appetite and GnRH may be dissociated.

Hypothalamic pathways involved in the regulation of energy balance have not been widely studied in ruminants to date. Here, we used in situ hybridisation to study the gene expression of a number of leptin-sensitive receptors and neuropeptides in the ovine hypothalamus. Gene expression was first localised for cocaine- and amphetamine-regulated transcript (CART) and agouti-related peptide (AGRP). We then examined in adult male castrated sheep the effects of acute negative energy balance induced by a 4-day fast on the amounts of these mRNAs and those for leptin receptor (OB-Rb), neuropeptide Y (NPY) and pro-opiomelanocortin (POMC). CART mRNA was localised in the arcuate nucleus (ARC), paraventricular nucleus, median eminence and ventromedial hypothalamic nucleus, and extensive co-localisation with POMC mRNA was demonstrated in the ARC. AGRP mRNA was localised in the ARC. Fasting up-regulated gene expression for OB-Rb and for the orexigenic neuropeptides NPY and AGRP in the ARC. There was a trend towards down-regulation of gene expression for the anorexigenic neuropeptide CART and no effect on POMC in the ARC, although these results are inconclusive. The presence or absence of oestradiol-containing subcutaneous implants did not influence gene expression or the effects of fasting. The hypothalamic changes were consistent with responses to the observed reduction in circulation leptin and suggest that the peripheral feedback and central mechanisms for restoring the energy balance may be largely conserved across monogastric and ruminant species., (Copyright 2002 S. Karger AG, Basel)

Hypothalamic orexin gene expression has not been reported in the ruminant. Here, we describe the localization of preproorexin mRNA in the ovine lateral hypothalamic area (LHA) and zona incerta (ZI) using in situ hybridization. The hypothalamic localization of the orexin gene expression was similar in sheep to rodent models. Since appetite in sheep is seasonally (photoperiodically) regulated, we compared the amounts of preproorexin mRNA between long- (LD) and short-day (SD) photoperiods in both freely feeding (food intake is 20% higher in LD than SD) and food-restricted sheep (50% liveweight maintenance for 11 weeks). Gene expression was higher in SDs than in LDs but was not affected by chronic food restriction. In a second study, hypothalamic orexin gene expression in castrate sheep was not affected by a 4-day fast, irrespective of gonadal steroid (estradiol) replacement, and was not affected by the gonadal steroid per se. The results demonstrate the sensitivity of orexin gene expression to photoperiod, but up-regulation occurs in SDs when the appetite is characteristically low and no sensitivity to imposed changes in food intake. This supports the concept that orexins may not have a primary role in appetite regulation and correction of negative energy balance but since the sheep breed only in SDs, their role in seasonal reproductive activation deserves further study.