Arcuate nucleus homeostatic systems reflect blood leptin concentration but not feeding behaviour during scheduled feeding on a high-fat diet in mice.

Hypothalamic homeostatic and forebrain reward-related genes were examined in the context of scheduled meal feeding without caloric restriction in C57BL/6 mice. Mice fed ad libitum but allowed access to a palatable high-fat (HF) diet for 2 hours a day rapidly adapted their feeding behaviour and consumed approximately 80% of their daily caloric intake during this 2-hour scheduled feed. Gene expression levels were examined during either the first or second hour of scheduled feeding vs 24 hours ad libitum feeding on the same HF diet. Gene expression of neuropeptide Y, agouti-related peptide, cocaine- and amphetamine-regulated transcript, pro-opiomelanocortin, long-form leptin receptor and suppressor of cytokine signalling-3 in the hypothalamic arcuate nucleus (ARC), as well as enkephalin, dynorphin, dopamine-2-receptor and dopamine-3-receptor in the nucleus accumbens (NAcc) in the forebrain, were measured by in situ hybridisation. Mice fed ad libitum on a HF diet had the highest total caloric intake, body weight gain, fat mass and serum leptin, whereas schedule-fed mice had a mild obese phenotype with intermediate total caloric intake, body weight gain, fat mass and serum leptin. The effects of feeding regime on ARC gene expression were emphasised by significant positive or negative correlations with body weight gain, fat mass and blood leptin, although they did not appear to be related to feeding behaviour in the schedule-fed groups (ie, the large, binge-type meals) and did not reveal any potential candidates for the regulation of these meals. Mechanisms underlying large meal/binge-type eating may be regulated by nonhomeostatic hedonic processes. However, assessment of opioid and dopamine receptor gene expression in the NAcc did not reveal evidence of involvement of these genes in regulating large meals. This complements our previous characterisation of ARC and NAcc genes in schedule-fed mice and rats, although it still leaves open the fundamental question about the underlying mechanisms of meal feeding.

Large, binge-type meals of high fat diet change feeding behaviour and entrain food anticipatory activity in mice.

Male C57BL/6 mice fed ad libitum on control diet but allowed access to a palatable high fat diet (HFD) for 2 h a day during the mid-dark phase rapidly adapt their feeding behaviour and can consume nearly 80% of their daily caloric intake during this 2 h-scheduled feed. We assessed food intake microstructure and meal pattern, and locomotor activity and rearing as markers of food anticipatory activity (FAA). Schedule fed mice reduced their caloric intake from control diet during the first hours of the dark phase but not during the 3-h period immediately preceding the scheduled feed. Large meal/binge-like eating behaviour during the 2-h scheduled feed was characterised by increases in both meal number and meal size. Rearing was increased during the 2-h period running up to scheduled feeding while locomotor activity started to increase 1 h before, indicating that schedule-fed mice display FAA. Meal number and physical activity changes were sustained when HFD was withheld during the anticipated scheduled feeding period, and mice immediately binged when HFD was represented after a week of this "withdrawal" period. These findings provide important context to our previous studies suggesting that energy balance systems in the hypothalamus are not responsible for driving these large, binge-type meals. Evidence of FAA in HFD dark phase schedule-fed mice implicates anticipatory processes in binge eating that do not involve immediately preceding hypophagia or regulatory homeostatic signalling.

Feeding and metabolic consequences of scheduled consumption of large, binge-type meals of high fat diet in the Sprague-Dawley rat.

Providing rats and mice with access to palatable high fat diets for a short period each day induces the consumption of substantial binge-like meals. Temporal food intake structure (assessed using the TSE PhenoMaster/LabMaster system) and metabolic outcomes (oral glucose tolerance tests [oGTTs], and dark phase glucose and insulin profiles) were examined in Sprague-Dawley rats given access to 60% high fat diet on one of 3 different feeding regimes: ad libitum access (HF), daily 2 h-scheduled access from 6 to 8 h into the dark phase (2 h-HF), and twice daily 1 h-scheduled access from both 1-2 h and 10-11 h into the dark phase (2×1 h-HF). Control diet remained available during the scheduled access period. HF rats had the highest caloric intake, body weight gain, body fat mass and plasma insulin. Both schedule-fed groups rapidly adapted their feeding behaviour to scheduled access, showing large meal/bingeing behaviour with 44% or 53% of daily calories consumed from high fat diet during the 2 h or 2×1 h scheduled feed(s), respectively. Both schedule-fed groups had an intermediate caloric intake and body fat mass compared to HF and control (CON) groups. Temporal analysis of food intake indicated that schedule-fed rats consumed large binge-type high fat meals without a habitual decrease in preceding intake on control diet, suggesting that a relative hypocaloric state was not responsible or required for driving the binge episode, and substantiating previous indications that binge eating may not be driven by hypothalamic energy balance neuropeptides. In an oGTT, both schedule-fed groups had impaired glucose tolerance with higher glucose and insulin area under the curve, similar to the response in ad libitum HF fed rats, suggesting that palatable feeding schedules represent a potential metabolic threat. Scheduled feeding on high fat diet produces similar metabolic phenotypes to mandatory (no choice) high fat feeding and may be a more realistic platform for mechanistic study of diet-induced obesity.

Photoperiod-dependent regulation of carboxypeptidase E affects the selective processing of neuropeptides in the seasonal Siberian hamster (Phodopus sungorus).

The production of bioactive peptides from biologically inactive precursors involves extensive post-translational processing, including enzymatic cleavage by proteolytic peptidases. Endoproteolytic prohormone-convertases initially cleave the precursors of many neuropeptides at specific amino acid sequences to generate intermediates with basic amino acid extensions on their C-termini. Subsequently, the related exopeptidases, carboxypeptidases D and E (CPD and CPE), are responsible for removing these amino acids before the peptides achieve biological activity. We investigated the effect of photoperiod on the processing of the neuropeptide precursor pro-opiomelanocortin (POMC) and its derived neuropeptides, α-melanocyte-stimulating hormone (MSH) and ß-endorphin (END), within the hypothalamus of the seasonal Siberian hamster (Phodopus sungorus). We thus compared hypothalamic distribution of CPD, CPE, α-MSH and ß-END using immunohistochemistry and measured the enzyme activity of CPE and concentrations of C-terminally cleaved α-MSH in short-day (SD; 8 : 16 h light/dark) and long-day (LD; 16 : 8 h light/dark) acclimatised hamsters. Increased immunoreactivity (-IR) of CPE, as well as higher CPE activity, was observed in SD. This increase was accompanied by more ß-END-IR cells and substantially higher levels of C- terminally cleaved α-MSH, as determined by radioimmunoassay. Our results suggest that exoproteolytic cleavage of POMC-derived neuropeptides is tightly regulated by photoperiod in the Siberian hamster. Higher levels of biological active α-MSH- and ß-END in SD are consistent with the hypothesis that post-translational processing is a key event in the regulation of seasonal energy balance.

Arcuate nucleus homeostatic systems are not altered immediately prior to the scheduled consumption of large, binge-type meals of palatable solid or liquid diet in rats and Mice.

Meal feeding is a critical issue in the over-consumption of calories leading to human obesity. To investigate the mechanisms involved in the regulation of meal feeding in rodents, we studied a scheduled feeding regime that induces substantial food intake over short periods of time. Male Sprague-Dawley rats and C57BL6 mice were fed one of four palatable diets [45% fat pellet, 60% fat pellet or standard pellet supplemented with Ensure (EN; Abbott Laboratories, Maidenhead, UK) or 12.5% sucrose (SUC)] either ad lib. or with daily 2-h scheduled access and standard pellet available for 22 h. Energy balance gene expression in the hypothalamic arcuate nucleus (ARC) and nucleus accumbens (NAcc) reward gene expression were assessed by in situ hybridisation. Rats fed ad lib. on 45% or 60% fat diet were heavier and fatter than controls, and had reduced neuropeptide Y (NPY) gene expression in the ARC. Mice fed ad lib. on any of the palatable diets were heavier, fatter and had higher blood leptin than controls, and had reduced NPY and increased cocaine- and-amphetamine-regulated transcript mRNA in the ARC. Schedule-fed rats and mice quickly adapted their feeding behaviour to 2-h access on palatable food. Three schedule-fed groups binged: the percentage of daily calories consumed in 2 h on 45% fat diet, 60% fat diet or EN, respectively, was 55%, 63% and 49% in rats, and 86%, 86% and 45% in mice. However, changed feeding behaviour was not reflected in an induction of orexigenic neuropeptide or suppression of anorexigenic neuropeptide gene expression in the ARC, in the 2-h period prior to scheduled feeding. The mechanisms underlying large meal/binge-type eating may be regulated by nonhomeostatic processes involving other genes in the hypothalamus or other brain areas. However, assessment of opioid and dopamine receptor gene expression in the NAcc did not reveal evidence of the involvement of these genes in driving large meals, at least at the investigated time point.

Leanness in postnatally nutritionally programmed rats is associated with increased sensitivity to leptin and a melanocortin receptor agonist and decreased sensitivity to neuropeptide Y.

BACKGROUND: Pups of normally nourished dams that are cross-fostered after birth to dams fed a low-protein (8% by weight) diet (postnatal low protein (PLP)) grow slower during the suckling period and remain small and lean throughout adulthood. At weaning, they have increased expression in the arcuate nucleus (ARC) of the hypothalamus of the orexigenic neuropeptide Y (NPY) and decreased expression of pro-opiomelanocortin, the precursor of anorexigenic melanocortins. OBJECTIVES AND METHODS: We investigated, using third ventricle administration, whether 3-month-old male PLP rats display altered sensitivity to leptin with respect to food intake, NPY and the melanocortin 3/4-receptor agonist MTII, and using in situ hybridization or laser capture microdissection of the ARC followed by RT-PCR, whether the differences observed were associated with changes in the hypothalamic expression of NPY or the leptin receptor, NPY receptors and melanocortin receptors. RESULTS: PLP rats were smaller and had reduced percentage body fat content and plasma leptin concentration compared with control rats. Leptin (5 µg) reduced food intake over 0-48 h more in PLP than control rats (P<0.05). Submaximal doses of NPY increased the food intake less in PLP rats than in controls, whereas submaximal doses of MTII reduced the food intake more in PLP rats. Maximal responses did not differ between PLP and control rats. Leptin and melanocortin-3 receptor (MC3R) expression were increased in both ARC and ventromedial hypothalamic nuclei in PLP animals compared with the controls. MC4R, NPY Y1R, Y5R and NPY expression were unchanged. CONCLUSION: Postnatal undernourishment results in food intake in adult rats being more sensitive to reduction by leptin and melanocortins, and less sensitive to stimulation by NPY. We propose that this contributes to increased leptin sensitivity and resistance to obesity. Increased expression of ObRb and MC3R may partly explain these findings but other downstream mechanisms must also be involved.

Short photoperiod-induced decrease of histamine H3 receptors facilitates activation of hypothalamic neurons in the Siberian hamster.

Nonhibernating seasonal mammals have adapted to temporal changes in food availability through behavioral and physiological mechanisms to store food and energy during times of predictable plenty and conserve energy during predicted shortage. Little is known, however, of the hypothalamic neuronal events that lead to a change in behavior or physiology. Here we show for the first time that a shift from long summer-like to short winter-like photoperiod, which induces physiological adaptation to winter in the Siberian hamster, including a body weight decrease of up to 30%, increases neuronal activity in the dorsomedial region of the arcuate nucleus (dmpARC) assessed by electrophysiological patch-clamping recording. Increased neuronal activity in short days is dependent on a photoperiod-driven down-regulation of H3 receptor expression and can be mimicked in long-day dmpARC neurons by the application of the H3 receptor antagonist, clobenproprit. Short-day activation of dmpARC neurons results in increased c-Fos expression. Tract tracing with the trans-synaptic retrograde tracer, pseudorabies virus, delivered into adipose tissue reveals a multisynaptic neuronal sympathetic outflow from dmpARC to white adipose tissue. These data strongly suggest that increased activity of dmpARC neurons, as a consequence of down-regulation of the histamine H3 receptor, contributes to the physiological adaptation of body weight regulation in seasonal photoperiod.

Developmental changes in hypothalamic leptin receptor: relationship with the postnatal leptin surge and energy balance neuropeptides in the postnatal rat.

In the adult brain, leptin regulates energy homeostasis primarily via hypothalamic circuitry that affects food intake and energy expenditure. Evidence from rodent models has demonstrated that during early postnatal life, leptin is relatively ineffective in modulating these pathways, despite the high circulating levels and the presence of leptin receptors within the central nervous system. Furthermore, in recent years, a neurotrophic role for leptin in the establishment of energy balance circuits has emerged. The precise way in which leptin exerts these effects, and the site of leptin action, is unclear. To provide a detailed description of the development of energy balance systems in the postnatal rat in relation to leptin concentrations during this time, endogenous leptin levels were measured, along with gene expression of leptin receptors and energy balance neuropeptides in the medial basal hypothalamus, using in situ hybridization. Expression of leptin receptors and both orexigenic and anorexigenic neuropeptides increased in the arcuate nucleus during the early postnatal period. At postnatal day 4 (P4), we detected dense leptin receptor expression in ependymal cells of the third ventricle (3V), which showed a dramatic reduction over the first postnatal weeks, coinciding with marked morphological changes in this region. An acute leptin challenge robustly induced suppressor of cytokine signaling-3 expression in the 3V of P4 but not P14 animals, revealing a clear change in the location of leptin action over this period. These findings suggest that the neurotrophic actions of leptin may involve signaling at the 3V during a restricted period of postnatal development.

Photoperiod regulates genes encoding melanocortin 3 and serotonin receptors and secretogranins in the dorsomedial posterior arcuate of the Siberian hamster.

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.

The thyrotropin-releasing hormone secretory system in the hypothalamus of the Siberian hamster in long and short photoperiods.

Thyrotropin-releasing hormone (TRH) is not only essential for the regulation of the pituitary-thyroid axis, but also exerts complementary effects on energy metabolism within the brain. We hypothesised that increased activity of the TRH secretory system may contribute to seasonal adaptations in the Siberian hamster whereby food intake is decreased in winter, and catabolism of fat stores is increased to support thermogenesis. We determined the distribution of TRH producing neurones and TRH-R1 receptor expressing cells in the hypothalamus, and investigated whether photoperiod regulated this system. TRH-immunoreactive (ir) cell somata and preproTRH mRNA expression were found to be widely distributed throughout the medial hypothalamus, with particular clusters in the paraventricular nucleus, the medial preoptic area and periventricular nucleus, and in the dorsomedial hypothalamus extending into the lateral hypothalamic area. A partial sequence encoding TRH-R1 was cloned from hamster hypothalamic cDNA and used to generate a riboprobe for in situ hybridisation studies. TRH-R1 mRNA expressing cells were abundant throughout the hypothalamus, corresponding to the widespread presence of TRH-ir fibres. Photoperiod did not affect the expression of preproTRH mRNA in any region, and the only significant change in TRH-R1 expression was in the dorsomedial posterior arcuate region. This wide distribution of TRH-producing and receptive cells in the hypothalamus is consistent with its hypothesised neuromodulatory roles in the short-term homeostatic control of appetite, thermoregulation and energy expenditure, but the lack of photoperiodic change in TRH mRNA expression does not support the hypothesis that changes in this system underlie long-term seasonal changes in body weight.

Early life programming of energy balance.

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.

Altered expression of SOCS3 in the hypothalamic arcuate nucleus during seasonal body mass changes in the field vole, Microtus agrestis.

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.

G protein-coupled receptor 101 mRNA expression in the mouse brain: altered expression in the posterior hypothalamus and amygdala by energetic challenges.

GPCR101 is a recently identified orphan G protein-coupled receptor (GPCR) expressed abundantly in the human and mouse hypothalamus. In the absence of a ligand, a direct approach to determine the function(s) of this receptor is not possible. However, clues to the possible functions of GPCR101 may yield from information on the distribution of the receptor and the effect of in vivo manipulation upon the expression level of the receptor. In situ hybridisation on mouse brain sections revealed GPCR101 expression in a number of nuclei, including the amygdala, lateral parabrachial nucleus and nucleus of the solitary tract, as well as in the arcuate nucleus, posterior hypothalamus and paraventricular nucleus of the hypothalamus. Food-deprivation was found to increase GPCR101 mRNA level in the posterior hypothalamus and amygdala. In obese mice bearing the ob gene mutation, GPCR101 mRNA level decreased in the posterior hypothalamus and remained unaltered in the amygdala. By contrast, in both nuclei, GPCR101 mRNA level did not change significantly in obese ob/ob mice after intraperitoneal injection of leptin or in mice fed with a high fat diet. These data suggest that GPCR101 mRNA expression in the posterior hypothalamus and amygdala is regulated by a factor(s) other than leptin. Dual in situ hybridisation was used to establish the relationship between GPCR101 and neuropeptides expressed in the hypothalamus. In the arcuate nucleus, GPCR101 mRNA was expressed in approximately half of the population of neurones expressing the mRNA for the anorexigenic neuropeptide, pro-opiomelanocortin, which suggests a potential functional relationship.

PC1/3 and PC2 gene expression and post-translational endoproteolytic pro-opiomelanocortin processing is regulated by photoperiod in the seasonal Siberian hamster (Phodopus sungorus).

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.

The suppressor of cytokine signalling 3, SOCS3, may be one critical modulator of seasonal body weight changes in the Siberian hamster, Phodopus sungorus.

The Siberian hamster, Phodopus sungorus, exhibits a remarkable cycle of body weight, reproduction and leptin sensitivity in response to a seasonal change in photoperiod. In the present study, we investigated the hypothesis that the suppressor of cytokine signalling 3 (SOCS3) plays a critical role in the regulation of the seasonal body weight cycle. We analysed arcuate nucleus SOCS3 gene expression in short day length (SD; 8 : 16 h light/dark) acclimated Siberian hamsters that were transferred back to long day length (LD; 16 : 8 h light/dark) and in hamsters that spontaneously became photorefractory to SD induced by prolonged exposure. SD acclimated hamsters that were transferred back to LD for 1, 2, 3, 4 or 6 weeks, increased arcuate nucleus SOCS3 gene expression to the LD level within 2 weeks, and maintained this higher level thereafter. The early increase of SOCS3 gene expression preceded the LD-induced rise in body weight by approximately 3 weeks. Hamsters kept in SD for an extended period (25 weeks), began to become refractory to SD and to increase body weight. By this time, there was no difference in level of SOCS3 gene expression between LD and SD photoperiods, although body weight was still suppressed in SD hamsters. Finally, we addressed whether SOCS3 gene expression is related to SD-induced gonadal regression or to body weight decrease by comparing Siberian hamsters with Syrian hamsters. The latter exhibited substantial SD-induced gonadal regression but only limited seasonal changes in body weight. Acclimation to either LD or SD for 14 weeks had no effect on SOCS3 gene expression. This implies that arcuate nucleus SOCS3 gene expression is unlikely to be related to seasonal cycles in reproductive activity. Taken together, the findings further strengthen our hypothesis that SOCS3 may be one molecular trigger of seasonal cycles in body weight.

Photoperiod regulates leptin sensitivity in field voles, Microtus agrestis.

We have previously shown that cold-acclimated (8 degrees C) male field voles (Microtus agrestis) transferred from short (SD, 8:16 h L:D) to long photoperiod (LD, 16:8 h L:D) exhibit increases in body mass, adiposity and food intake. To assess whether these increases were associated with decreased leptin sensitivity, we infused LD and SD voles with physiological doses of murine leptin (or saline) delivered peripherally for 7 days via mini-osmotic pumps. Measurements were made of body mass (weight-reducing effect of leptin), food intake (anorectic effect of leptin) and gene expression of uncoupling protein 1 (UCP1) in brown adipose tissue (BAT) (thermogenic effect of leptin). The SD animals were sensitive to the weight-reducing effects of leptin (mean body mass decrease of 1.2 g over 7 days) and appetite-reducing effect of leptin (mean food intake decrease of 2.5 g over 7 days), whereas LD voles were resistant to the hormone treatment. The switch from a leptin-sensitive to leptin-resistant state appears to act as a desensitisation mechanism that allows voles transferred from SD to LD to ignore elevated leptin levels generated by increased body fat and accumulate adipose tissue without stimulating compensatory changes opposing the weight gain. Neither SD nor LD voles responded to infusion of leptin by changes in BAT UCP1 gene expression, suggesting dissociation of anorectic and thermogenic effects of leptin, possibly related to chronic cold exposure. Our results indicate that cold-acclimated voles show photoperiod-regulated changes in leptin sensitivity and may provide an attractive model for elucidating molecular mechanisms of leptin resistance.

Diet-induced obesity in the Sprague-Dawley rat: dietary manipulations and their effect on hypothalamic neuropeptide energy balance systems.

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.

Hypothalamic energy balance gene responses in the Sprague-Dawley rat to supplementation of high-energy diet with liquid ensure and subsequent transfer to chow.

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.

Introduction of a high-energy diet acutely up-regulates hypothalamic cocaine and amphetamine-regulated transcript, Mc4R and brown adipose tissue uncoupling protein-1 gene expression in male Sprague-Dawley rats.

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.

Effect of photoperiod on body mass, food intake and body composition in the field vole, Microtus agrestis.

Many small mammals respond to seasonal changes in photoperiod by altering body mass and adiposity. These animals may provide valuable models for understanding the regulation of energy balance. Here, we present data on the field vole (Microtus agrestis) - a previously uncharacterised example of photoperiod-induced changes in body mass. We examined the effect of increased day length on body mass, food intake, apparent digestive efficiency, body composition, de novo lipogenesis and fatty acid composition of adipose tissue in cold-acclimated (8 degrees C) male field voles by transferring them from a short (SD, 8 h:16 h L:D) to long day photoperiod (LD, 16 h:8 h L:D). During the first 4 weeks of exposure to LD, voles underwent a substantial increase in body mass, after which the average difference between body masses of LD and SD voles stabilized at 7.5 g. This 24.8% increase in body mass reflected significant increases in absolute amounts of all body components, including dry fat mass, dry lean mass and body water mass. After correcting body composition and organ morphology data for the differences in body mass, only gonads (testes and seminal vesicles) were enlarged due to photoperiod treatment. To meet energetic demands of deposition and maintenance of extra tissue, voles adjusted their food intake to an increasing body mass and improved their apparent digestive efficiency. Consequently, although mass-corrected food intake did not differ between the photoperiod groups, the LD voles undergoing body mass increase assimilated on average 8.4 kJ day(-1) more than animals maintained in SD. The majority (73-77%) of the fat accumulated as adipose tissue had dietary origin. The rate of de novo lipogenesis and fatty acid composition of adipose tissue were not affected by photoperiod. The most important characteristics of the photoperiodic regulation of energy balance in the field vole are the clear delineation between phases where animals regulate body mass at two different levels and the rate at which animals are able to switch between different levels of energy homeostasis. Our data indicate that the field vole may provide an attractive novel animal model for investigation of the regulation of body mass and energy homeostasis at both organism and molecular levels.