Agrp mRNA Expression Research Articles

THU190 Deletion Of The Glucocorticoid Receptor From GABAergic Neurons Causes Early Puberty In Male And Female Mice

Abstract Disclosure: S.A. Pereira: None. L. Arvizu-Sanchez: None. A.N. Fontes: None. R.S. Carroll: None. V.M. Navarro: None. U.B. Kaiser: None. Background and Objectives: Glucocorticoid (GC) secretion induced by various types of stress can negatively impact reproduction through effects at several levels of the hypothalamic-pituitary-gonadal (HPG) axis, including inhibition of gonadotrophin-releasing hormone (GnRH) secretion and suppression of luteinizing hormone (LH) release. KNDy neurons, expressing kisspeptin, neurokinin B and dynorphin A, are the GnRH and LH pulse generator and are key regulators of puberty. These neurons are proposed to play an important role in mediating the effects of stress and GC on reproduction. While the precise mechanisms by which stress affects regulation of KNDy neurons remain to be fully characterized, it is known that KNDy neurons are subject to regulation by multiple neural networks, among which GABAergic neurons appear to play a determinant role. Importantly, a large proportion of GABAergic neurons express glucocorticoid receptors (GR). We therefore hypothesized that stress, through the action of GC on GR in GABAergic neurons, affects the activity of KNDy neurons to impact puberty timing. Methods and Results: We generated mice with conditional deletion of GR in GABAergic neurons using VgatCre mice crossed with GRlox/lox mice to generate VgatCre;GRlox/lox mice (GABAGRKO). The pubertal phenotypes of these GABAGRKO mice and their littermate controls (GRlox/lox) were characterized. Male GABAGRKO mice had earlier preputial separation compared to GRlox/lox littermate controls (GABAGRKO 29.8 ± 0.45 days vs. GRlox/lox 33.5 ± 0.57 days; p < 0.0001). Female GABAGRKO mice had a similar age of vaginal opening (GABAGRKO 25.7 ± 0.76 days vs GRlox/lox 24.5 ± 0.87 days; p = 0.2810) but had significantly earlier first estrus compared to their GRlox/lox littermate controls (GABAGRKO 32.4 ± 0.76 days vs GRlox/lox 37.9 ± 0.79 days; p < 0.0001). Male and female mice were euthanized at 30 days of age and their mediobasal hypothalamus (MBH) were collected for mRNA measurement by RT-qPCR of genes known to be related to pubertal development, including Kiss1, Tac2, Pdyn, Agrp and Pomc. Kiss1 and Pdyn expression were significantly higher in the MBH of GABAGRKO males compared to controls; in contrast, Pdyn expression was significantly lower in the MBH of GABAGRKO females and Kiss1 expression was unchanged compared to controls. Agrp expression was lower in the MBH of both GABAGRKO male and female mice compared to controls. No differences were observed in Tac2 and Pomc expression. Data sets are mean ± SEM. Statistical significance was considered at p < 0.05. Conclusions: The deletion of GR in GABAergic neurons results in early puberty onset, sexually dimorphic changes in Pdyn mRNA levels and downregulation of Agrp mRNA expression in male and female GABAGRKO mice. These findings suggest a role of glucocorticoids acting on GR in GABAergic neurons upstream of KNDy neurons to restrain the timing of their activation to initiate puberty. Presentation: Thursday, June 15, 2023

Effects of a Calorie-Restricted Cafeteria Diet and Oleuropein Supplementation on Adiposity and mRNA Expression of Energy Balance Related Genes in Obese Male Rats.

Supplementation with natural bioactive compounds has been proposed to be a complementary tool to the calorie-restricted diets and physical exercise programs used to tackle human overweight, obesity and Metabolic syndrome. Herein, we evaluated the effects of 14 weeks of calorie-restricted cafeteria diet either alone or combined with oral administration of the polyphenol oleuropein in obese adult male rats, compared with a control group fed standard chow and a group fed cafeteria diet. Animals were sacrificed at the age of 26 weeks and several tissues of interest were removed. The results showed that both dietary interventions reduced the adiposity index (p < 0.05 and p < 0.01, respectively), and specifically the abdominal fat depots (mesenteric: p < 0.01 and p < 0.01, respectively; and epididymal: both diets p < 0.001) and restored the decreased soleus skeletal muscle mass. Both interventions decreased leptin mRNA expression in mesenteric white adipose tissue (p < 0.05) and normalized hypothalamic Agrp mRNA expression compared to cafeteria-fed obese rats (p < 0.05). However, only the calorie-restricted cafeteria diet supplemented with oleuropein induced additional lower retroperitoneal adipose accretion (p < 0.05) and increased hypothalamic leptin receptor mRNA levels (p < 0.05). Experiments with female animals, at different doses and longer intervention periods, are needed to better determine the potential benefits of this dietary treatment.

Variation of NPY and AGRP MRNA expression in Syrian hamsters according to feeding times

Variation of NPY and AGRP MRNA expression in Syrian hamsters according to feeding times

Gut Microbiota Restores Central Neuropeptide Deficits in Germ-Free Mice.

Recent work has demonstrated the ability of the gut microbiota (GM) to alter the expression and release of gut peptides that control appetite and regulate energy homeostasis. However, little is known about the neuronal response of these hormones in germ-free (GF) animals, especially leptin, which is strikingly low in these animals. Therefore, we aimed to determine the response to exogenous leptin in GF mice as compared to conventionally raised (CONV-R) mice. Specifically, we injected and measured serum leptin in both GF and CONV-R mice and measured expression of orexigenic and anorexigenic peptides NPY, AgRP, POMC, and CART in the hypothalamus and hindbrain to examine whether the GM has an impact on central nervous system regulation of energy homeostasis. We found that GF mice had a significant increase in hypothalamic NPY and AgRP mRNA expression and a decrease in hindbrain NPY and AgRP mRNA, while mRNA expression of POMC and CART remained unchanged. Administration of leptin normalized circulating levels of leptin, GLP-1, PYY, and ghrelin, all of which were significantly decreased in GF mice. Finally, brief conventionalization of GF mice for 10 days restored the deficits in hypothalamic and hindbrain neuropeptides present in GF animals. Taken together, these results show that the GM regulates hypothalamic and hindbrain orexigenic/anorexigenic neuropeptide expression. This is in line with the role of gut microbiota in lipid metabolism and fat deposition that may contribute to excess fat in conventionalized animals under high feeding condition.

Bisphenol S induces Agrp expression through GPER1 activation and alters transcription factor expression in immortalized hypothalamic neurons: A mechanism distinct from BPA-induced upregulation

The increasing prevalence of obesity around the world has brought concern upon ubiquitously present obesogenic environmental compounds, such as bisphenol A (BPA). Increasingly tightened regulations on the industrial use of BPA have prompted a transition to a structurally similar alternative, bisphenol S (BPS). BPS displays endocrine-disrupting behaviours similar to those of BPA and increases body weight, food intake and the hypothalamic expression of Agrp in vivo. However, the mechanisms behind this deleterious effect are unclear. Here, we report an increase in the mRNA level of Agrp at 4 h following BPS treatment in immortalized murine hypothalamic cell lines of embryonic and adult origin (mHypoE-41, mHypoA-59). BPS-induced changes in the expression of transcription factors and estrogen receptors that occurred concurrently with Agrp upregulation demonstrated similarities to BPA-induced changes, however, there were also changes that were unique to BPS. Specifically, while Chop, Atf3, Atf4, Atf6, Klf4, and Creb1 were upregulated and Gper1 was downregulated by both BPA and BPS, Esr1 mRNA levels were upregulated and Foxo1 and Stat3 levels remained unchanged by BPS. Finally, inhibition of GPER1 by G15 prevented BPS-mediated Agrp upregulation, independent of Atf3 and Klf4 upregulation. Overall, our results demonstrate the ability of BPS to increase Agrp mRNA expression through GPER1 signaling and to alter transcription factor expression in hypothalamic neurons, further elucidating the endocrine-disrupting potential of this alternative industrial chemical.

Altered ERK‐mediated control of AgRP and metabolic rate during obesity

Adaptation of resting metabolic rate (RMR) contributes to the maintenance of human obesity, yet RMR control and how this system changes during obesity remains poorly defined. Desensitization of the brain to leptin has been proposed to contribute. Leptin normally suppresses Agouti‐related peptide (AgRP) expression in the arcuate nucleus of the hypothalamus (ARC) to disinhibit (increase) melanocortin‐mediated RMR stimulation. Diet‐induced obesity (DIO) increases leptin, but paradoxically exaggerates Agrp expression and causes RMR adaptation. Thus, something connecting leptin to Agrp control becomes dysfunctional during DIO. Our previous data indicate that within the ARC, disruption in the extracellular signal‐regulated kinase (ERK) cascade occurs during DIO. ERK normally phosphorylates Krüppel‐like factor 4 (KLF4), inactivating it. As KLF4 binds to the AgRP promoter to increase transcription, its inactivation results in reduced Agrp expression. We hypothesize that this mechanism becomes dysfunctional within AgRP neurons during DIO, such that KLF4 binding to the AgRP promoter increases despite increased ERK activity.Mouse hypothalamic GT1‐7 cells (model of AgRP neurons) were treated with leptin (1 uM, 15 min or 1 hr) and pERK/ERK ratio was measured by Simple WES and Agrp mRNA expression via qPCR, respectively. Leptin was given to chow‐fed adult mice (2 ug/g ip 30 min) and ARC pERK/ERK ratio was measured, as well as RMR assessed by respirometry. To examine consequences of ERK or KLF4 disruption upon cardiometabolic phenotypes, mice harboring a conditional version of endogenous Mapk1 or Klf4 were crossed with mice expressing Cre via the Agrp locus.In GT1‐7 cells, leptin increased pERK/ERK ratio (1 uM, 15 min leptin n=3, +89% vs vehicle n=2, p&lt;0.05) and suppressed Agrp expression (1 uM, 1 hr leptin n=2, ‐54.1% vs vehicle n=6, p&lt;0.05). In chow‐fed adult mice, leptin injection caused an increase in the ARC pERK/ERK ratio (2 ug/g ip 30 min leptin n=11, +43.8% vs saline n=9, p&lt;0.05). Additionally, leptin stimulated RMR in adult‐chow fed mice (1 ug/g ip leptin n=8, 16.5‐fold of vehicle n=8, p&lt;0.05). DIO caused increases in endogenous plasma leptin and an increase in the ARC pERK/ERK ratio (High Fat Diet (HFD; n=5), +54.4% vs chow n=4, p&lt;0.05), but ARC Agrp expression was not suppressed (HFD n=4, 1.62 fold of chow n=4, p&lt;0.05). There was a possible exaggeration of weight gain in Agrp‐Cre+, Mapk1F/F mice compared to controls following HFD (KOs n=2, +16.8% vs controls n=3). In contrast, weight gain was possibly attenuated in Agrp‐Cre+, Klf4F/F mice compared to controls following HFD (KO n=1, ‐10.7% vs controls n=2). RMR was higher in chow‐fed Agrp‐Cre+, Klf4F/F mice compared to controls (KOs n=5 +8.3% vs controls n=7, p&lt;0.05) and may be exaggerated following HFD (KO n=1 +13.0% vs controls n=3). Together, these findings suggest that ERK and KLF4 within AgRP neurons appear to contribute to the control of AgRP and thus RMR. These findings may identify ERK control of KLF4 and KLF4 as a potential therapeutic target to maintain/restore RMR control after prolonged obesity.

STAT5 ablation in AgRP neurons increases female adiposity and blunts food restriction adaptations

AgRP neurons are important players in the control of energy homeostasis and are responsive to several hormones. In addition, STAT5 signalling in the brain, which is activated by metabolic hormones and growth factors, modulates food intake, body fat and glucose homeostasis. Given that, and the absence of studies that describe STAT5 function in AgRP cells, the present study investigated the metabolic effects of Stat5a/b gene ablation in these neurons. We observed that STAT5 signalling in AgRP neurons regulates body fat in female mice. However, male and female STAT5-knockout mice did not exhibit altered food intake, energy expenditure or glucose homeostasis compared to control mice. The counter-regulatory response or glucoprivic hyperphagia induced by 2-deoxy-d-glucose treatment were also not affected by AgRP-specific STAT5 ablation. However, under 60% food restriction, AgRP STAT5-knockout mice had a blunted upregulation of hypothalamic Agrp mRNA expression and corticosterone serum levels compared to control mice, suggesting a possible role for STAT5 in AgRP neurons for neuroendocrine adaptations to food restriction. Interestingly, ad libitum fed knockout male mice had reduced Pomc and Ucp-1 mRNA expression compared to control group. Taken together, these results suggest that STAT5 signalling in AgRP neurons regulates body adiposity in female mice, as well as some neuroendocrine adaptations to food restriction.

Caloric Restriction Prevents Metabolic Dysfunction and the Changes in Hypothalamic Neuropeptides Associated with Obesity Independently of Dietary Fat Content in Rats.

Energy restriction is a first therapy in the treatment of obesity, but the underlying biological mechanisms have not been completely clarified. We analyzed the effects of restriction of high-fat diet (HFD) on weight loss, circulating gut hormone levels and expression of hypothalamic neuropeptides. Ten-week-old male Wistar rats (n = 40) were randomly distributed into four groups: two fed ad libitum a normal diet (ND) (N group) or a HFD (H group) and two subjected to a 25% caloric restriction of ND (NR group) or HFD (HR group) for 9 weeks. A 25% restriction of HFD over 9 weeks leads to a 36% weight loss with regard to the group fed HFD ad libitum accompanied by normal values in adiposity index and food efficiency ratio (FER). This restriction also carried the normalization of NPY, AgRP and POMC hypothalamic mRNA expression, without changes in CART. Caloric restriction did not succeed in improving glucose homeostasis but reduced HFD-induced hyperinsulinemia. In conclusion, 25% restriction of HFD reduced adiposity and improved metabolism in experimental obesity, without changes in glycemia. Restriction of the HFD triggered the normalization of hypothalamic NPY, AgRP and POMC expression, as well as ghrelin and leptin levels.

Vulnerability to psychological stress‐induced anorexia in female mice depends on blockade of ghrelin signal in nucleus tractus solitarius

Background and PurposeWomen have a higher incidence of eating disorders than men. We investigated whether the effects of ghrelin on feeding are affected by sex and stress, and to elucidate the mechanisms that may cause sex differences in stress‐mediated anorexia, focusing on ghrelin.Experimental ApproachAcylated ghrelin was administered to naïve and psychologically stressed male and female C57BL/6J mice, followed by measurements of food intake and plasma hormone levels. Ovariectomy was performed to determine the effects of ovary‐derived oestrogen on stress‐induced eating disorders in female mice. The numbers of Agrp or c‐Fos mRNA‐positive cells and estrogen receptor α/c‐Fos protein‐double‐positive cells were assessed.Key ResultsGhrelin administration to naïve female mice caused a higher increase in food intake, growth hormone secretion, Agrp mRNA expression in the arcuate nucleus and c‐Fos expression in the nucleus tractus solitarius (NTS) than in male mice. In contrast, psychological stress caused a more sustained reduction in food intake in females than males. The high sensitivity of naïve females to exogenous ghrelin was attenuated by stress exposure. The stress‐induced decline in food intake was not abolished by ovariectomy. Estrogen receptor‐α but not ‐β antagonism prevented the decrease in food intake under stress. Estrogen receptor‐α/c‐Fos‐double‐positive cells in the NTS were significantly increased by stress only in females.Conclusion and ImplicationsStress‐mediated eating disorders in females may be due to blockade of ghrelin signalling via estrogen receptor‐α activation in the NTS. Targeting the ghrelin signal in the brain could be a new treatment strategy to prevent these disorders.

Assessing the Effects of Leptin and Angiotensin II Upon Expression of Agouti‐Related Peptide in the GT1‐7 Cell Model

Leptin acts at its receptor (Lepr) within the arcuate nucleus of the hypothalamus (ARC) to reduce feeding behavior and increase resting metabolic rate (RMR) in part through the inhibition of Agouti‐related peptide (Agrp) signaling. Previously we demonstrated that a unique subset of Agrp neurons express the angiotensin II (ANG) type 1a receptor (Agtr1a). Genetic deletion of Agtr1a from Agrp neurons of mice was sufficient to disinhibit expression of Agrp within the ARC, and to attenuate adipose sympathetic nerve activity responses to leptin. In contrast, RMR responses to brain renin‐angiotensin system (RAS) stimulation are intact in the dysfunctional Lepr db/db mouse model, and acute leptin administration does not additively exaggerate RMR activation in a transgenic mouse model of increased brain RAS activity. New unpublished data also indicate that ANG signaling within the ARC stimulates RMR through suppression of Agrp expression and Agrp neuron electrical activity, and thereby disinhibition of melanocortin Mc4r receptor signaling. To further dissect the interaction of Lepr and Agtr1a within Agrp neurons, here we examined the effects of leptin and ANG stimulation of the “GT1‐7” immortalized cell culture model of mouse Agrp‐expressing cells. We confirmed that cultured GT1‐7 cells express critical markers of the Agtr1a‐positive Agrp neuron subtype, including Agrp, Agtr1a, and Neuropeptide‐Y (Npy), but not pro‐opiomelanocortin (Pomc). Dose‐ and time‐course experiments were performed to identify mid‐range doses and time courses for Agrp gene expression responses to acute application of leptin and ANG. Subsequently, Agrp expression responses to low doses of leptin (100 nM) and ANG (100 nM), which would allow detection of any additive or synergistic effect, were assessed after 1 and 2–2.5 hours to evaluate the hypothesis that leptin and ANG signaling are non‐additive for the suppression of Agrp expression. After 1 hour, ANG (0.66 (0.49–0.91), fold of vehicle (± 1 sem), n=0.5) and leptin (0.61 (0.44–0.84), n=6) had similar effects on Agrp mRNA expression compared to vehicle (1.00 (0.86–1.17), n=7), but co‐administration of both peptides had no synergistic effect (0.96 (0.78–1.18), n=5). Likewise after 2–2.5 hours, compared to vehicle (1.00 (0.78–1.28), n=6), ANG (0.56 (0.34–0.94), n=6) and leptin (0.69 (0.57–0.85), n=6) had similar effects that were not additive (0.81 (0.67–0.98), n=6). These findings support the preliminary conclusion that the effects of leptin and ANG upon Agrp expression are not additive in the GT1‐7 cell culture model of Agrp neurons. These findings support our working model in which leptin action at the ARC results in increased Agtr1a signaling to suppress Agrp, ultimately to disinhibit Mc4r signaling and thus increase resting energy expenditure.Support or Funding InformationHL134850, HL084207

STAT5 ablation in AgRP neurons increases female adiposity and blunts food restriction adaptations.

AgRP neurons are important players in the control of energy homeostasis and are responsive to several hormones. In addition, STAT5 signalling in the brain, which is activated by metabolic hormones and growth factors, modulates food intake, body fat and glucose homeostasis. Given that, and the absence of studies that describe STAT5 function in AgRP cells, the present study investigated the metabolic effects of Stat5a/b gene ablation in these neurons. We observed that STAT5 signalling in AgRP neurons regulates body fat in female mice. However, male and female STAT5-knockout mice did not exhibit altered food intake, energy expenditure or glucose homeostasis compared to control mice. The counter-regulatory response or glucoprivic hyperphagia induced by 2-deoxy-d-glucose treatment were also not affected by AgRP-specific STAT5 ablation. However, under 60% food restriction, AgRP STAT5-knockout mice had a blunted upregulation of hypothalamic Agrp mRNA expression and corticosterone serum levels compared to control mice, suggesting a possible role for STAT5 in AgRP neurons for neuroendocrine adaptations to food restriction. Interestingly, ad libitum fed knockout male mice had reduced Pomc and Ucp-1 mRNA expression compared to control group. Taken together, these results suggest that STAT5 signalling in AgRP neurons regulates body adiposity in female mice, as well as some neuroendocrine adaptations to food restriction.

Sex differences in basal hypothalamic anorectic and orexigenic gene expression and the effect of quantitative and qualitative food restriction

BackgroundResearch into energy balance and growth has infrequently considered genetic sex, yet there is sexual dimorphism for growth across the animal kingdom. We test the hypothesis that in the chicken, there is a sex difference in arcuate nucleus neuropeptide gene expression, since previous research indicates hypothalamic AGRP expression is correlated with growth potential and that males grow faster than females. Because growth has been heavily selected in some chicken lines, food restriction is necessary to improve reproductive performance and welfare, but this increases hunger. Dietary dilution has been proposed to ameliorate this undesirable effect. We aimed to distinguish the effects of gut fullness from nutritional feedback on hypothalamic gene expression and its interaction with sex.MethodsTwelve-week-old male and female fast-growing chickens were either released from restriction and fed ad libitum or a restricted diet plus 15% w/w ispaghula husk, a non-nutritive bulking agent, for 2 days. A control group remained on quantitative restriction. Hypothalamic arcuate nucleus neuropeptides were measured using real-time PCR. To confirm observed sex differences, the experiment was repeated using only ad libitum and restricted fed fast-growing chickens and in a genetically distinct breed of ad libitum fed male and female chickens. Linear mixed models (Genstat 18) were used for statistical analysis with transformation where appropriate.ResultsThere were pronounced sex differences: expression of the orexigenic genes AGRP (P < 0.001) and NPY (P < 0.002) was higher in males of the fast-growing strain. In genetically distinct chickens, males had higher AGRP mRNA (P = 0.002) expression than females, suggesting sex difference was not restricted to a fast-growing strain. AGRP (P < 0.001) expression was significantly decreased in ad libitum fed birds but was high and indistinguishable between birds on a quantitative versus qualitative restricted diet. Inversely, gene expression of the anorectic genes POMC and CART was significantly higher in ad libitum fed birds but no consistent sex differences were observed.ConclusionExpression of orexigenic peptides in the avian hypothalamus are significantly different between sexes. This could be useful starting point of investigating further if AGRP is an indicator of growth potential. Results also demonstrate that gut fill alone does not reduce orexigenic gene expression.

Does obestatin modulate the hypothalamic appetite‐regulating network in peripubertal sheep?

The participation of peripheral peptides in the processes regulating the food intake (energy homeostasis) at the central nervous system level remains unclear. This study focuses on the role of obestatin in neuronal activity within the hypothalamic appetite-regulating network in ruminants. The animals (n=28) were randomly divided into two groups. The sheep in the control group received intracerebroventricular infusions of the Ringer-Locke solution, and the sheep in obestatin group were infused with obestatin (diluted in the Ringer-Locke solution) at 25μg per 120μl/hr. The series of four 1-hr infusions on 3consecutive days were performed, and immediately after the experiment, the sheep were decapitated. Selected brain regions were fixed in situ for further immunohistochemical analysis, while the remaining ones were frozen for real-time RT-qPCR analysis. Obestatin infusion elicited changes in the neuropeptide Y (NPY) neuronal network in the hypothalamus. The results obtained show that exogenous obestatin evoked an increase in npy and agrpmRNA expression in the mediobasal hypothalamus (MBH), while the immunoreactivity for NPY was decreased in the arcuate and periventricular nuclei. The increase in cart and pomcmRNA expression in the MBH was also observed. Moreover, increased levels of gpr39 receptor and npy receptor 1 mRNA expression were evident in obestatin-infused sheep. Based on these results, it can be concluded that obestatin plays a role in the modulation of appetite-regulating network at the central level in sheep. The results obtained suggest that the underlying mechanism may involve the modification of the activity of NPY/AgRP and CART/α-MSH neurons in the arcuate nucleus.

Involvement of CD36 in Modulating the Decrease of NPY and AgRP Induced by Acute Palmitic Acid Stimulation in N1E-115 Cells.

Central nervous system (CNS) fatty acid sensing plays an important role in the regulation of food intake, and palmitic acid (PA) is the most important long chain fatty acid (LCFA) in the mammalian diet. To explore the effect of PA on central neuropeptide expression and the role of the cluster of the differentiation of 36 (CD36) in the process, N1E-115 cells were cultured with PA in the presence or absence of sulfosuccinimidyl-oleate (SSO), a CD36 inhibitor. Results showed that 10 μmol/L PA significantly reduced NPY and AgRP mRNA expression after 20 min of exposure, while the expression of CD36 was upregulated. The presence of SSO significantly attenuated the decrease of NPY and AgRP expression that was induced by PA alone, although no notable effect on PA- induced CD36 gene expression was observed. In conclusion, our study suggests the involvement of CD36 in the PA-induced decrease of NPY and AgRP in N1E-115 cells.

Diet-induced obesity and prenatal undernutrition lead to differential neuroendocrine gene expression in the hypothalamic arcuate nuclei.

Previously we reported that prenatal undernutrition (UN) leads to a dysregulation of appetite suppression through alterations in hypothalamic neuropeptide gene expression. In the current study, we expand our observations and investigate neuroendocrine transcriptional responses and central leptin sensitivity within the arcuate nucleus of rats exposed to prenatal UN or a postnatal high-fat diet (HF). Pregnant Wistar rats were fed a standard chow diet either ad libitum (AD) or at 30% of AD intake throughout gestation (UN) resulting in either control or intrauterine growth-restricted female offspring. At weaning, AD offspring were fed either a chow (C) or a HF (30% fat wt/wt) diet ad libitum for the remainder of the study, whereas UN offspring were fed a chow diet only. At ~142days, AD and UN offspring received either recombinant rat leptin (L) or saline (S) subcutaneously for 14days. Prenatal UN had a significant effect on hypothalamic NPY (P<0.0001), AgRP (P<0.01) and ObRb (P<0.02) mRNA expression compared to AD chow-fed offspring. A postnatal HF diet had a significant effect on AgRP mRNA expression (P<0.001), compared to AD chow-fed offspring, but no effect on NPY and ObRb expression. Leptin treatment, in both UN and HF offspring, was ineffective in reducing NPY and AgRP mRNA expression, and had no effect on ObRb expression. These findings suggest that prenatal UN and a postnatal HF diet lead to differential neuroendocrine gene expression in the hypothalamic arcuate nuclei and reduced sensitivity to leptin's anorexigenic effects.

GPR17 gene disruption does not alter food intake or glucose homeostasis in mice

G protein-coupled receptor 17 (GPR17) was recently reported to be a Foxo1 target in agouti-related peptide (AGRP) neurons. Intracerebroventricular injection of GPR17 agonists induced food intake, whereas administration of an antagonist to the receptor reduced feeding. These data lead to the conclusion that pharmacological modulation of GPR17 has therapeutic potential to treat obesity. Here we report that mice deficient in Gpr17 (Gpr17(-/-)) have similar food intake and body weight compared with their wild-type littermates. Gpr17(-/-) mice have normal hypothalamic Agrp mRNA expression, AGRP plasma levels, and metabolic rate. GPR17 deficiency in mice did not affect glucose homeostasis or prevent fat-induced insulin resistance. These data do not support a role for GPR17 in the control of food intake, body weight, or glycemic control.

Effect of tributyltin on the food intake and brain neuropeptide expression in rats.

Tributyltin (TBT) is a largely diffused environmental pollutant. Several studies have demonstrated that TBT is involved in the development of obesity. However, few studies addressing the effects of TBT on the brain neuropeptides involved in appetite and body weight homeostasis have been published. Experiments were carried out on female and male Sprague-Dawley rats. Animals were exposed to TBT (0.5 μg/kg body weight) for 54 days. The hepatic triglyceride and total cholesterol were determined using commercial enzyme kits. The NPY, AgRP, POMC and CART mRNA expression in brains were quantified by real-time PCR. TBT exposure resulted in significant increases in the hepatic total cholesterol and triglyceride concentration of both male and female rats. Interestingly, increases in body weight and fat mass were only found in the TBT-treated male rats. TBT exposure also led to a significant increase in food intake by the female rats, while no change was observed in the male rats. Moreover, the neuropeptides expression was different between males and females after TBT exposure. TBT induced brain NPY expression in the female rats, and depressed brain POMC, AgRP and CART expression in the males. TBT can increase food intake in female rats, which is associated with the disturbance of NPY in brains. TBT had sex-different effects on brain NPY, AgRP, POMC and CART mRNA expression, which indicates a complex neuroendocrine mechanism of TBT.

Changes in FGF21 serum concentrations and liver mRNA expression in an experimental model of complete lipodystrophy and insulin-resistant diabetes.

Patients with obesity and type 2 diabetes often display high levels of the anti-diabetic factor fibroblast growth factor-21 (FGF21), suggesting that the overproduction of FGF21 may result from increased adiposity in an attempt by white adipose tissue (WAT) to counteract insulin resistance. However, the production of FGF21 diabetes in the absence of WAT has not been examined. In this study, we investigated the effects of lipodystrophy in A-ZIP F-1 mice on FGF21 production in relation to diabetes. A-ZIP F-1 mice displayed high FGF21 plasma levels resulting from enhanced FGF21 mRNA expression in the liver. Concomitant enhancement of FGF21 receptor (FGFR1) and glucose transporter 1 (GLUT-1) mRNA expression was observed in the muscles of A-ZIP F-1 mice. Furthermore, the activation of hypothalamic NPY and AgRP mRNA expression positively correlated with plasma levels of FGF21 but not active ghrelin. Our study demonstrates that an increased FGF21 plasma level in lipodystrophic A-ZIP F-1 mice results mainly from up-regulated liver production but does not suffice to overcome the lipodystrophy-induced severe type 2-diabetes and insulin resistance in the liver linked to the augmented liver fat deposition.

The increase in body weight induced by lack of methyl CpG binding protein‐2 is associated with altered leptin signalling in the hypothalamus

Methyl CpG binding protein-2 (MECP2) is a chromatin-remodelling factor with a dual role in gene expression. Evidence from patients carrying MECP2 mutations and from transgenic mouse models demonstrates that this protein is involved in the control of body weight. However, the mechanism for this has not been fully elucidated. To address this, we used a previously characterized Mecp2-null mouse model and found that the increase in body weight is associated with an increased amount of adipose tissue and high leptin levels. Appropriate body weight control requires the proper expression of pro-opiomelanocortin (Pomc) and agouti-related peptide (Agrp), two neuropeptides essential for satiety and appetite signals, respectively. Our results show that in the absence of Mecp2, Pomc and Agrp mRNA expression are altered, and the mice are leptin resistant. To determine the mechanism underlying the defective leptin sensing, we evaluated the expression of genes and the post-translational modifications associated with leptin signalling, which are fundamental to Pomc and Agrp transcriptional control and proper leptin response. We found a decrease in the phosphorylation level of Akt and its target protein Foxo1, which indicate an alteration in leptin-induced signal transduction. Our results demonstrate that the absence of Mecp2 disrupted body weight balance by altering post-translational modifications in leptin-signalling components that regulate Pomc and Agrp expression.

Hormonal, hypothalamic and striatal responses to reduced body weight gain are attenuated in anorectic rats bearing small tumors

Lack of compensatory or even reduced food intake is frequently observed in weight-losing cancer patients and contributes to increased morbidity and mortality. Our previous work has shown increased transcription factor expression in the hypothalamus and ventral striatum of anorectic rats bearing small tumors. mRNA expression of molecules known to be involved in pathways regulating appetite in these structures was therefore assessed in this study. Given that pain, pro-inflammatory cytokines and metabolic hormones can modify food intake, spinal cord cellular activation patterns and plasma concentrations of cytokines and hormones were also studied. Morris hepatoma 7777 cells injected subcutaneously in Buffalo rats provoked a 10% lower body weight and 15% reduction in food intake compared to free-feeding tumor-free animals 4weeks later when the tumor represented 1–2% of body mass. No differences in spinal cord activation patterns or plasma concentration of pro-inflammatory cytokines were observed between groups. However, the changes in plasma ghrelin and leptin concentrations found in food-restricted weight-matched rats in comparison to ad libitum-fed animals did not occur in anorectic tumor-bearing animals. Real-time PCR showed that tumor-bearing rats did not display the increase in hypothalamic agouti-related peptide mRNA observed in food-restricted weight-matched animals. In addition, microarray analysis and real-time PCR revealed increased ventral striatal prostaglandin D synthase expression in food-restricted animals compared to anorectic tumor-bearing rats. These findings indicate that blunted hypothalamic AgRP mRNA expression, probably as a consequence of relatively high leptin and low ghrelin concentrations, and reduced ventral striatal prostaglandin D synthesis play a role in maintaining cancer-associated anorexia.