Hypothalamic Leptin Receptor Research Articles

Maternal Choline Intake Programs Hypothalamic Energy Regulatory Pathways and Long-Term Phenotype in Male Wistar Rat Offspring (OR35-04-19)

ObjectivesThe prenatal period is a critical time of brain development. Maternal choline intake is associated with improvement in memory and cognitive function in the offspring. However, the role of choline in the regulation of physiological functions controlled by the hypothalamus has not been reported. The objective of this study is to elucidate the effects of choline intake on the in utero programming of hypothalamic energy regulatory neurons in male Wistar rat offspring. MethodsPregnant Wistar rats received an AIN-93G diet containing recommended choline (RC, 1 g/kg diet), low choline (LC, 0.5-fold), or high choline (HC, 2.5-fold). At birth, brain and blood was collected from male pups. Male pups from each dietary treatment were maintained on the control diet for 17-weeks. Dependent measures include post-weaning food intake, energy expenditure, weight-gain, plasma glucoregulatory hormones, brain choline and 1-carbon metabolite levels, and expression of hypothalamic energy regulatory neurons. ResultsAt birth, pup brain concentrations of choline proportionally reflected the choline content in the maternal diets. HC pups had higher hypothalamic protein expression of the orexigenic neuropeptide-Y neuron than both groups (P < 0.05), but lower activation than LC pups (P < 0.05). Both HC and LC pups had lower plasma leptin concentrations than RC pups (P < 0.01), but LC pups had lower hypothalamic leptin receptor expression compared to both groups at birth (P < 0.05). During adulthood, offspring of HC dams had higher weekly food intake compared to RC (11%, P < 0.01), and higher weight-gain than both RC and LC groups (12%, P < 0.05). LC offspring had lower 24hr energy expenditure and locomotor activity than HC and RC groups (6%, P < 0.05). ConclusionsCholine content of diets consumed by rats during pregnancy impacts in utero development of hypothalamic energy regulatory systems; long-term body weight-gain, food intake and energy expenditure in mature rat offspring. Funding SourcesThis research was supported by the Canadian Institute of Health Research, Institute of Nutrition, Metabolism and Diabetes (CIHR-INMD).

1819-P: Growth Hormone Receptor (GHR) in AgRP Neurons Modulates Energy Balance and Metabolic Adaptation to Fasting via Sirtuin 1 (SIRT1)

We are facing a worldwide public health emergency due to the increasing epidemic of diabetes. In the central nervous system, the hypothalamus is the primary site of integration of the neuroendocrine and autonomic responses which maintain glucose and energy homeostasis. Main physiological effects of growth hormone (GH) in adult brain - is the inhibition of its own secretion, especially critical under fasting states. While the importance of hypothalamic GH receptor (GHR) in GH feedback inhibition is well established, the direct involvement of neuronal GHR signals in the regulation of whole-body homeostasis has not been studied. Previously, we have identified a unique population of hypothalamic leptin receptor (LepRb)-GHR expressing neurons that regulate hepatic glucose production, suggesting a crucial role for these neurons in GHR-neurocircuitry. To characterize hypothalamic GHR-expressing neurons, we have developed GHRcre mice using CRISPR/Cas9 technology. Approximately 60% of GHR+ neurons in the arcuate nucleus (ARC) co-localize with AgRP-expressing neurons and about 25% with GHRH neurons, basically representing most of ARCGHR neurons. By manipulating neuronal activity of neurochemically-defined GHR neurons, we demonstrate that ARCGHR neurons represent distinct neuronal population in glycemic control. Despite similarity in responses in both sexes, only female mice selectively lacking GHR in AgRP neurons showed impaired energy homeostasis and heat production, accompanied by alterations in metabolic adaptation to fasting. Fasting increased the abundance of sirtuin 1 (SIRT1) within ARCGHR+ neurons, while this pattern was diminished in the fasted AgRPEYFP∆GHR female mice. Our data suggests a role of SIRT1 within somatotropic signaling in the AgRPGHR+ neurons in response to alterations in nutritional status and demonstrates the role of AgRP-GHR neurons in assisting the organism in recruiting its resources for survival purposes. Disclosure L.K. Debarba: None. I.T. Ayyar: None. M.C. Koch: None. G. Cady: None. M. Sadagurski: None. Funding American Diabetes Association (1-18-JDF-063 to M.S.)

Leptin actions on the reproductive neuroendocrine axis in sheep

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.

The role of leptin in nutritional status and reproductive function

Infertility associated with suboptimal nutrition is a major concern among livestock producers. Undernourished prepubertal animals will not enter puberty until they are well fed; similarly, adult, normally cyclic females will stop cycling when faced with extreme undernutrition. Work in our laboratory has focused on how body fat (or adiposity) of an animal can communicate to the brain and regulate reproductive competence. In 1994, the discovery in rodents of the obese (ob) gene product leptin, secreted as a hormone from adipocytes, provided a unique opportunity to understand and hence regulate whole body compositional changes. There is now evidence that similar mechanisms are functioning in livestock species in which food intake, body composition, and reproductive performance are of considerable economic importance. Leptin has been reported to be a potent regulator of food intake and reproduction in rodents. There is evidence indicating that at least some of the effects of leptin occur through receptor-mediated regulation of the hypothalamic protein neuropeptide Y (NPY). NPY is a potent stimulator of food intake, is present at high concentrations in feed-restricted cattle and ewes, and is an inhibitor of LH secretion in these livestock species. In our investigations in sheep, we have cloned a partial cDNA corresponding to the ovine long-form leptin receptor, presumably the only fully active form, and have localized the long-form leptin receptor in the ventromedial and arcuate nuclei of the hypothalamus. Leptin receptor mRNA expression was colocalized with NPY mRNA-containing cell bodies in those regions. We have also determined that hypothalamic leptin receptor expression is greater in feed-restricted ewes than in well-fed ewes. These observations provide a foundation for future investigations into the nutritional modulators of reproduction in livestock.

Protein tyrosine phosphatase alpha inhibits hypothalamic leptin receptor signaling and regulates body weight in vivo.

Understanding how body weight is regulated at the molecular level is essential for treating obesity. We show that female mice genetically lacking protein tyrosine phosphatase (PTP) receptor type α (PTPRA) exhibit reduced weight and adiposity and increased energy expenditure, and are more resistant to diet-induced obesity than matched wild-type control mice. These mice also exhibit reduced levels of circulating leptin and are leptin hypersensitive, suggesting that PTPRA inhibits leptin signaling in the hypothalamus. Male and female PTPRA-deficient mice fed a high-fat diet were leaner and displayed increased metabolic rates and lower circulating leptin levels, indicating that the effects of loss of PTPRA persist in the obese state. Molecularly, PTPRA down-regulates leptin receptor signaling by dephosphorylating the receptor-associated kinase JAK2, with which the phosphatase associates constitutively. In contrast to the closely related tyrosine phosphatase ε, leptin induces only weak phosphorylation of PTPRA at its C-terminal regulatory site Y789, and this does not affect the activity of PTPRA toward JAK2. PTPRA is therefore an inhibitor of hypothalamic leptin signaling in vivo and may prevent premature activation of leptin signaling, as well as return signaling to baseline after exposure to leptin.-Cohen-Sharir, Y., Kuperman, Y., Apelblat, D., den Hertog, J., Spiegel, I., Knobler, H., Elson, A. Protein tyrosine phosphatase alpha inhibits hypothalamic leptin receptor signaling and regulates body weight in vivo.

FRUCTOSE ALTERS THE EXPRESSION OF TYPE B LEPTIN RECEPTOR IN HYPHOTALAMUS AND INTESTINUM OF Rattus Novergicus

&lt;p&gt;&lt;strong&gt;Background&lt;/strong&gt;: Several epidemiological data’s reported significant correlation between fructose consumption and diabetes mellitus type 2 by inducing insulin resistance. Leptin resistance induced by fructose was proposed as one novel mechanism that induce insulin resistance but the exact mechanism remains unclear. We hypothesize that fructose diminish the type b leptin receptors in hypothalamus and intestine.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Aim&lt;/strong&gt;: This study was aimed to elucidate fructose effect on the expression of leptin receptor type b in hypothalamus and intestine of Rattus novergicus.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Method&lt;/strong&gt;: twenty eight rats were used and divided into 4 groups: Group 1 was control, group 2 was given fructose 10%, group 3 was given fructose 30% and group 4 was given fructose 55% for 2 months. At the end of treatment, the animal were sacrificed and then the hypothalamus and intestine were collected. The expression of type b leptin receptor were measured by immunohistochemistry technique with primary antibody from &lt;em&gt;Bioss antibodies type&lt;/em&gt; &lt;em&gt;Leptin receptor polyclonal antibody bs-0109R using &lt;/em&gt;Staining kit &lt;em&gt;Skytec Laboratories and DAB chromogen. &lt;/em&gt;A positive expression can be seen as a brown colour in cell cytoplasm and counted in 100 cell. The expression then analysed using SPSS 18 with anova one way tes (p&amp;lt;0,05) followed by post hoc test after the data showed normality and homogeneity using Saphiro wilk and Levene test (p&amp;gt;0,05).&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Result&lt;/strong&gt;: There was significant differences in type b leptin receptors found in hypothalamus between each group (p&amp;lt;0.05). The significant differences also could be seen in the expression between control and group fructose 30 and 50% in intestinal cell (p&amp;lt;0.05).&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Conclusion&lt;/strong&gt;: the consumption of Fructose 55% for 2 months attenuates the expression of type b leptin receptors in hypothalamus and intestine of &lt;em&gt;R&lt;/em&gt;&lt;em&gt;at novergicus&lt;/em&gt;.&lt;/p&gt;&lt;p&gt; &lt;/p&gt;

Phenethyl isothiocyanate activates leptin signaling and decreases food intake.

Obesity, a principal risk factor for the development of diabetes mellitus, heart disease, and hypertension, is a growing and serious health problem all over the world. Leptin is a weight-reducing hormone produced by adipose tissue, which decreases food intake via hypothalamic leptin receptors (Ob-Rb) and the Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway. Protein tyrosine phosphatase 1B (PTP1B) negatively regulates leptin signaling by dephosphorylating JAK2, and the increased activity of PTP1B is implicated in the pathogenesis of obesity. Hence, inhibition of PTP1B may help prevent and reduce obesity. In this study, we revealed that phenethyl isothiocyanate (PEITC), a naturally occurring isothiocyanate in certain cruciferous vegetables, potently inhibits recombinant PTP1B by binding to the reactive cysteinyl thiol. Moreover, we found that PEITC causes the ligand-independent phosphorylation of Ob-Rb, JAK2, and STAT3 by inhibiting cellular PTP1B in differentiated human SH-SY5Y neuronal cells. PEITC treatment also induced nuclear accumulation of phosphorylated STAT3, resulting in enhanced anorexigenic POMC expression and suppressed orexigenic NPY/AGRP expression. We demonstrated that oral administration of PEITC to mice significantly reduces food intake, and stimulates hypothalamic leptin signaling. Our results suggest that PEITC might help prevent and improve obesity.

The intracellular domain of the leptin receptor prevents mitochondrial depolarization and mitophagy

Hypothalamic leptin receptor (LR) signaling regulates body weight by balancing food intake and energy expenditure. It is well established that the human LR undergoes ectodomain shedding, but little is known about the fate of the remaining cytosolic domain. This study demonstrates that regulated intramembrane proteolysis (RIP) releases the LR intracellular domain (LR ICD), which translocates to the mitochondria where it binds to SOCS6. This LR ICD-SOCS6 interaction stabilizes both proteins on the mitochondrial outer membrane and requires a functional BC box in SOCS6 for mitochondrial association and a central motif in the LR ICD for SOCS6 binding. The LR ICD prevents CCCP-induced mitochondrial depolarization and mitophagy as shown by lowered Parkin translocation and p62 accumulation. Strict regulation of mitochondrial dynamics in the hypothalamus is known to be essential for body weight homeostasis. This is the first study showing that the LR can directly modulate mitochondrial biology.

A high nutrient dense diet alters hypothalamic gene expressions to influence energy intake in pigs born with low birth weight

The low birth weight (LBW) individual had greater risk of developing metabolic dysfunction in adulthood. The aim of this study was to test whether the LBW individual is more prone to glucose intolerance on a high nutrient dense (HND) diet, and to investigate the associated hypothalamic gene expressions using pigs as model. The intake of digestible energy intake, if calculated on a body weight basis, was greater in LBW pigs than that of normal birth weight (NBW) pigs. The LBW pigs fed the HND diet had greater digestible energy intake than those fed the NND diet at adulthood, which did not occur for NBW pigs. Notably, up-regulated hypothalamic toll-like receptor 4, interleukin 6 and phospho-NFκB p65 expressions, and the altered expressions of hypothalamic leptin receptor, suppressor of cytokine signaling 3, agouti-related protein and proopiomelanocortin predicted the overconsumption of energy intake and development of glucose intolerance in LBW pigs fed the HND diet. Collectively, pigs born with LBW had a distinct hypothalamic leptin signaling to a high nutrient dense diet, which contributed to greater energy intake and glucose intolerance.

Intranasal delivery of N-terminal modified leptin-pluronic conjugate for treatment of obesity

Leptin is an adipocyte-secreted hormone that is delivered via a specific transport system across the blood-brain barrier (BBB) to the brain where it acts on the hypothalamus receptors to control appetite and thermogenesis. Peripheral resistance to leptin due to its impaired brain delivery prevents therapeutic use of leptin in overweight and moderately obese patients. To address this problem, we modified the N-terminal amine of leptin with Pluronic P85 (LepNP85) and administered this conjugate intranasally using the nose-to-brain (INB) route to bypass the BBB. We compared this conjugate with the native leptin, the N-terminal leptin conjugate with poly(ethylene glycol) (LepNPEG5K), and two conjugates of leptin with Pluronic P85 attached randomly to the lysine amino groups of the hormone. Compared to the random conjugates of leptin with P85, LepNP85 has shown higher affinity upon binding with the leptin receptor, and similarly to native hormone activated hypothalamus receptors after direct injection into brain. After INB delivery, LepNP85 conjugate was transported to the brain and accumulated in the hypothalamus and hippocampus to a greater extent than the native leptin and LepNPEG5K and activated leptin receptors in hypothalamus at lower dose than native leptin. Our work suggests that LepNP85 can access the brain directly after INB delivery and confirms our hypothesis that the improvement in brain accumulation of this conjugate is due to its enhanced brain absorption. In conclusion, the LepNP85 with optimized conjugation chemistry is a promising candidate for treatment of obesity.

Corticotropin-releasing factor overexpression in mice abrogates sex differences in body weight, visceral fat, and food intake response to a fast and alters levels of feeding regulatory hormones

BackgroundCorticotropin-releasing factor overexpressing (CRF-OE) male mice showed an inhibited feeding response to a fast, and lower plasma acyl ghrelin and Fos expression in the arcuate nucleus compared to wild-type (WT) mice. We investigated whether hormones and hypothalamic feeding signals are impaired in CRF-OE mice and the influence of sex.MethodsMale and female CRF-OE mice and WT littermates (4–6 months old) fed ad libitum or overnight fasted were assessed for body, adrenal glands and perigonadal fat weights, food intake, plasma hormones, blood glucose, and mRNA hypothalamic signals.ResultsUnder fed conditions, compared to WT, CRF-OE mice have increased adrenal glands and perigonadal fat weight, plasma corticosterone, leptin and insulin, and hypothalamic leptin receptor and decreased plasma acyl ghrelin. Compared to male, female WT mice have lower body and perigonadal fat and plasma leptin but higher adrenal glands weights. CRF-OE mice lost these sex differences except for the adrenals. Male CRF-OE and WT mice did not differ in hypothalamic expression of neuropeptide Y (NPY) and proopiomelanocortin (POMC), while female CRF-OE compared to female WT and male CRF-OE had higher NPY mRNA levels. After fasting, female WT mice lost more body weight and ate more food than male WT, while CRF-OE mice had reduced body weight loss and inhibited food intake without sex difference. In male WT mice, fasting reduced plasma insulin and leptin and increased acyl ghrelin and corticosterone while female WT showed only a rise in corticosterone. In CRF-OE mice, fasting reduced insulin while leptin, acyl ghrelin and corticosterone were unchanged with no sex difference. Fasting blood glucose was higher in CRF-OE with female > male. In WT mice, fasting increased hypothalamic NPY expression in both sexes and decreased POMC only in males, while in CRF-OE mice, NPY did not change, and POMC decreased in males and increased in females.ConclusionsThese data indicate that CRF-OE mice have abnormal basal and fasting circulating hormones and hypothalamic feeding-related signals. CRF-OE also abolishes the sex difference in body weight, abdominal fat, and fasting-induced feeding and changes in plasma levels of leptin and acyl ghrelin.

Blockage of neonatal leptin signaling induces changes in the hypothalamus associated with delayed pubertal onset and modifications in neuropeptide expression during adulthood in male rats

The neonatal leptin surge, occurring from postnatal day (PND) 5 to 13 and peaking at PND9 in rodents, is important for the development of neuroendocrine circuits involved in metabolic control and reproductive function. We previously demonstrated that treatment with a leptin antagonist from PND 5 to 9, coincident with peak leptin levels in the neonatal surge, modified trophic factors and markers of cell turnover and neuronal maturation in the hypothalamus of peri-pubertal rats. The kisspeptin system and metabolic neuropeptide and hormone levels were also modified. Here our aim was to investigate if the timing of pubertal onset is altered by neonatal leptin antagonism and if the previously observed peripubertal modifications in hormones and neuropeptides persist into adulthood and affect male sexual behavior. To this end, male Wistar rats were treated with a pegylated super leptin antagonist (5mg/kg, s.c.) from PND 5 to 9 and killed at PND102–103. The appearance of external signs of pubertal onset was delayed. Hypothalamic kiss1 mRNA levels were decreased in adult animals, but sexual behavior was not significantly modified. Although there was no effect on body weight or food intake, circulating leptin, insulin and triglyceride levels were increased, while hypothalamic leptin receptor, POMC and AgRP mRNA levels were decreased. In conclusion, alteration of the neonatal leptin surge can modify the timing of pubertal onset and have long-term effects on hypothalamic expression of reproductive and metabolic neuropeptides.

Hypothalamic Pomc‐deficiency Impairs the Function of Leptin to Decrease Food Intake and Bodyweight

Proopiomelanocortin (Pomc) in the arcuate nucleus (Arc) of the hypothalamus is necessary for normal energy homeostasis; therefore, its deficiency leads to obesity. We recently reported that ArcPomc‐rescue by genetic intervention prevents obesity in otherwise obesity‐destined ArcPomc‐knockout (KO) mice. However, the efficiency of ArcPomc‐rescue to normalize body weight (BW) was contingent on the magnitude of obesity at the time of genetic intervention. To dissect the molecular mechanism(s) responsible for that observation, we hypothesized that ArcPomc‐rescue mediated reversal of obesity is linked to the function of leptin to reduce BW and food intake (FI). To test our hypothesis, we performed a longitudinal study during which we assessed BW, FI, plasma leptin levels and leptin sensitivity in the same set of ArcPomc‐KO mice when they were obese, weight‐matched to WT mice by calorie restriction, and after ArcPomc rescue. ArcPomc was rescued in the weight‐matched mice by tamoxifen‐inducible Cre recombinase technology. We found that obese KO mice exhibited significantly higher plasma leptin levels (68 ±4.1 vs. 5.6 ±0.4 ng/ml), which were reduced by 78% in the weight‐matched group (14.5 ±1.2 ng/ml) but still remained higher, relative to WT group (P&lt;0.01). Moreover, leptin treatment did not reduce BW relative to saline‐group (Δ BW, 0.52 ±0.06 vs. 0.56 ±0.07 g/24 h when mice were obese; 0.8 ± 0.08 vs 0.6 ±0.07 g/24 h when mice were weight‐matched) and FI (Δ FI, 5.8 ±0.2 vs. 6.1 ±0.1 g/24 h when mice were obese; 6.0 ±0.1 vs. 6.2 ±0.1 g/24 h when mice were weight‐matched) in the KO mice at different stages of the longitudinal study. Remarkably, we found that ArcPomc‐rescue in the weight‐matched KO mice restored normal BW (32.6 ±1.0 g), FI (4.4 ±0.6 g), leptin levels (4.6 ±1.3 ng/ml) and sensitivity [Δ BW, −1.6 ±0.1 (leptin) vs. 0.01 ±0.06 (saline) g/24 h] in the mice that had a history of obesity. In contrast, mice that remained ArcPomc deficient regained massive obesity (BW: 68.4 ±1.5 g, P&lt;0.001) and hyperleptinemia (70.2 ±5.4 ng/ml, P&lt;0.001). To further confirm the association of hypothalamic Pomc with leptin action, we treated weight‐matched ArcPomc KO mice without a history of obesity with leptin. Intraperitoneal and intracerebroventricular leptin administration failed to reduce BW and FI in the KO mice. Interestingly, there was no change in hypothalamic leptin receptor gene expression; however, relative hypothalamic pSTAT3 levels after leptin administration were 20% lower in the KO mice compared to wildtype mice. Altogether, our study suggests that restoration of normal body weight and food intake in ArcPomc KO mice after ArcPomc‐rescue is linked with leptin sensitivity. Moreover, hypothalamic Pomc appears to mediate the function of leptin to control BW and FI. Therefore, stimulating hypothalamic Pomc signaling provides an alternative approach to treat obesity in face of leptin resistance.Support or Funding InformationNIH grants R01 DK066604 (to M.J.L.), R01 DK068400 (to M.J.L. and M.R.). NIH grants P30 DK020572 and P30 DK089503 to University of Michigan Animal Phenotyping and Chemistry Cores supported by the Michigan Diabetes Research Center and the Michigan Nutrition and Obesity Research Center

The expression of orexigenic and anorexigenic factors in middle-aged female rats that had been subjected to prenatal undernutrition

Fetal growth retardation, which affects short- and long-term fetal brain development, is associated with metabolic, hematological, and thermal disturbances, which can increase the risk of metabolic syndrome later in life. Orexigenic and anorexigenic factors regulate food intake and energy expenditure. We studied how the expression of these factors was affected by food deprivation (FD) in middle-aged female rats that had been subjected to prenatal undernutrition. Eight pregnant rats were divided into two groups, the normal nutrition (NN) (n=4) group and the undernutrition (UN) (n=4) group, which received 50% (approximately 11g) of the daily food intake of the normal nutrition rats from day 13 of pregnancy to delivery. The pups from these dams were defined as the maternal NN (mNN) and maternal UN (mUN) groups, respectively. After weaning, all of the pups were housed and allowed ad libitum access to food and water. At the age of 6 months, both groups of pups were sub-divided into three groups. One group was allowed to consume normal amounts of food (Fed), and the other two groups were subjected to 24h or 48h FD (n=7–8 per group). The rats’ serum leptin levels and hypothalamic mRNA expression levels of various orexigenic or anorexigenic factors were measured. In both the mNN and mUN rats, the serum leptin levels of the 24h and 48h FD groups tended to be lower than those of the Fed group, and the serum leptin levels of the 24h FD mUN rats and the Fed mUN rats differed significantly. The hypothalamic neuropeptide Y (NPY) mRNA expression levels of the 24h and 48h FD groups were significantly higher in the mUN rats than in the mNN rats. In addition, among the mUN rats the hypothalamic NPY mRNA expression levels of the 48h FD group were significantly higher than those of the Fed group. In both the mNN and mUN rats, prepro-orexin mRNA expression was lower in the 48h FD group than in the corresponding Fed group. Among the mUN rats, the 48h FD group exhibited significantly lower hypothalamic proopiomelanocortin (POMC) mRNA expression than the Fed group, and a similar tendency was seen among the mNN rats. Among the mNN rats, the 24h FD group displayed significantly higher hypothalamic leptin receptor (OBRb) mRNA levels than the Fed group. However, no such differences were seen among the mUN rats. As a result, the hypothalamic OBRb mRNA expression levels of the mUN rats in the 24h and 48h FD groups were lower than those of the corresponding mNN rat groups. These findings indicate that rats that are subjected to prenatal undernutrition exhibit upregulated expression of orexigenic factors and are more sensitive to FD in middle age, which might increase their risk of developing metabolic disorders in later life.

Hypothalamic Amylin Acts in Concert with Leptin to Regulate Food Intake

In this report we evaluated the functions of hypothalamic amylin invivo and invitro. Profiling of hypothalamic neurons revealed that islet amyloid polypeptide (Iapp, precursor to amylin) is expressed in neurons in the lateral hypothalamus, arcuate nucleus, medial preoptic area, and elsewhere. Hypothalamic expression of lapp is markedly decreased in ob/ob mice and normalized by exogenous leptin. In slices, amylin and leptin had similar electrophysiologic effects on lateral hypothalamic leptin receptor ObRb-expressing neurons, while the amylin antagonist AC187 inhibited their activity and blunted the effect of leptin. Finally, i.c.v. infusion of AC187 acutely reduced the anorectic effects of leptin. These data show that hypothalamic amylin is transcriptionally regulated by leptin, that it can act directly on ObRb neurons in concert with leptin, and that it regulates feeding. These findings provide a potential mechanism for the increased efficacy of a metreleptin/pramlintide combination therapy for obesity.

The expression of leptin, hypothalamic neuropeptides and UCP1 before, during and after fattening in the Daurian ground squirrel (Spermophilus dauricus)

The Daurian ground squirrel (Spermophilus dauricus) accumulates large amounts of body fat during pre-hibernation fattening. Leptin, an adipose-derived hormone, plays important roles in energy balance and thermogenesis. We predicted that body fat accumulation would lead to the elevation of leptin concentration while its effect on satiety would be suppressed in hypothalamus during fattening. In addition, the uncoupling protein 1 (UCP1) in brown adipose tissue (BAT) would increase and correlated positively with leptin concentration before hibernation. Here, we measured serum leptin concentration and leptin mRNA in white adipose tissue (WAT), hypothalamic neuropeptides involved in energy regulation and UCP1 in BAT before, during and after fattening in squirrels. The fat mass gradually increased during fattening but serum leptin increased mainly in the late phase of fattening, which was consistent with leptin mRNA expression in WAT. During fattening, the mRNA of hypothalamic leptin receptor was up-regulated and correlated positively with serum leptin. Orexigenic neuropeptide Y mRNA increased by 67%; however agouti-related peptide remained unchanged before hibernation. There was no significant change in anorexigenic neuropeptide mRNA. No change in suppressor of cytokine signaling-3 and protein tyrosine phosphatase-1B was detected. UCP1 mRNA expression and protein content in BAT increased significantly after fattening. These changes were independent of environmental conditions and serum leptin concentration. Our results suggest that the dissociation of leptin production and adiposity during fattening may facilitate fat accumulation. No evidence of suppressed leptin signal was found in fattening squirrels. The UCP1 recruitment in post-fattening squirrels could occur without winter-like acclimation and increased leptin.

Prolonged REM sleep restriction induces metabolic syndrome-related changes: Mediation by pro-inflammatory cytokines.

Chronic sleep restriction in human beings results in metabolic abnormalities, including changes in the control of glucose homeostasis, increased body mass and risk of cardiovascular disease. In rats, 96h of REM sleep deprivation increases caloric intake, but retards body weight gain. Moreover, this procedure increases the expression of pro-inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), which may be involved with the molecular mechanism proposed to mediate insulin resistance. The goal of the present study was to assess the effects of a chronic protocol of sleep restriction on parameters of energy balance (food intake and body weight), leptin plasma levels and its hypothalamic receptors and mediators of the immune system in the retroperitoneal adipose tissue (RPAT). Thirty-four Wistar rats were distributed in control (CTL) and sleep restriction groups; the latter was kept onto individual narrow platforms immersed in water for 18h/day (from 16:00h to 10:00h), for 21days (SR21). Food intake was assessed daily, after each sleep restriction period and body weight was measured daily, after the animals were taken from the sleep deprivation chambers. At the end of the 21day of sleep restriction, rats were decapitated and RPAT was obtained for morphological and immune functional assays and expression of insulin receptor substrate 1 (IRS-1) was assessed in skeletal muscle. Another subset of animals was used to evaluate blood glucose clearance. The results replicated previous findings on energy balance, e.g., increased food intake and reduced body weight gain. There was a significant reduction of RPAT mass (p<0.001), of leptin plasma levels and hypothalamic leptin receptors. Conversely, increased levels of TNF-α and IL-6 and expression of phosphorylated NFκ-β in the RPAT of SR21 compared to CTL rats (p<0.01, for all parameters). SR21 rats also displayed reduced glucose clearance and IRS-1 expression than CTL rats (p<0.01). The present results indicated that 21days of sleep restriction by the platform method induced metabolic syndrome-related alterations that may be mediated by inflammation of the RPAT.

Relationship status update for PTP1B and LepR: It׳s complicated.

The adipocyte-derived hormone leptin is an important regulator of systemic energy metabolism. Transported via the circulation to the brain in proportion to fat mass, leptin promotes its biological action through activation of the long form of the leptin receptor (LepRb). Accordingly, binding of leptin to the extracellular domain of LepRb promotes transphosphorylation of the LepRb-associated Janus kinase 2 (JAK2), which in turn phosphorylates other tyrosine residues (Tyr985, Tyr1077, Tyr1138) along the intracellular tail of LepRb in order to facilitate downstream signaling [1]. The protein tyrosine phosphatase 1B (PTP1B) is an important negative regulator of leptin action due to its ability to directly inhibit LepRb-Jak2 signaling (Figure 1). The relevance of PTP1B to regulate energy metabolism was first shown in mice with global PTP1B deletion [2] and was later confirmed in a series of other studies. These whole body PTP1B deficient mice show decreased body weight, food intake and adiposity, leading to resistance to diet-induced obesity (DIO) and improved leptin sensitivity and glucose metabolism when exposed to a high-fat diet (HFD) [2]. Of appreciable note, mice with specific deletion of PTP1B in muscle, liver or adipose tissue show no overt changes in body weight, adiposity or glucose tolerance [3–5]. However, deletion of PTP1B specifically in the central nervous system (CNS) [6], in LepRb-expressing cells [7] and POMC neurons [8,9] mimics the results observed in the global PTP1B deficient mice. These studies strongly support the relevance of central PTP1B signaling in regulating energy metabolism. It remains unclear, however, whether and to what extent the metabolic benefits arising from central PTP1B deficiency depend on PTP1B׳s action within the hypothalamus and whether the observed effects on metabolism are mediated via hypothalamic leptin receptor signaling. An elegant step in solving these questions has now been taken by the group of Kendra Bence from the University of Pennsylvania [10]. In the current issue of Molecular Metabolism, Tsou and colleagues used mice with hypothalamus-specific expression of Cre recombinase (Nkx2.1 cre) to specifically delete PTP1B in the hypothalamus. Compared to wildtype control mice, these hypothalamus-specific PTP1B deficient mice (Nkx2.1 PTP1B−/−) show decreased body weight, adiposity and food intake under chronic HFD exposure. These data corroborate previous reports about the relevance of central PTP1B signaling in the regulation of systems metabolism. Moreover, they show for the first time that the hypothalamus plays a key role in mediating PTP1B׳s action on metabolism and that hypothalamic selective PTP1B deletion mimics the phenotype of the whole body, whole brain, LepRb-expressing cell and POMC neuronal PTP1B knockout mice [6–9]. To further assess whether the metabolic benefits observed in the hypothalamus-specific PTP1B−/− mice depend on hypothalamic leptin receptor signaling, the authors generated mice with concomitant deletion of PTP1B and the leptin receptor in the hypothalamus (Nkx2.1 PTP1B−/−: LepRb−/−). Interestingly, when comparing these double mutant mice with hypothalamus-specific LepRb deficient mice, the metabolic benefits of PTP1B deletion vanished. No difference in body weight, food intake, adiposity or glucose tolerance was observed when double mutant mice were compared to mice that lack only the leptin receptor, but not PTP1B, in the hypothalamus. Together, the data indicate that the improved body weight and adiposity that is observed upon hypothalamic PTP1B deletion is mediated via interaction of PTP1B and the leptin receptor in the hypothalamus and that the metabolic benefits arising from PTP1B deficiency thus depend on a functional leptin receptor in the hypothalamus. These results underscore the importance of hypothalamic leptin receptor signaling for the regulation of energy and glucose metabolism and highlight the role of PTP1B in systems metabolism. In summary, the recent data by Tsou et al., combined with a series of previous studies, make PTP1B an interesting target for studies aiming to improve leptin sensitivity in states of pathologically increased body weight. Figure 1 Schematic of how PTP1B affects intracellular leptin receptor signaling. Binding of leptin to the extracellular domain of LepRb leads to autophosphorylation of the Janus kinase 2 (JAK2). JAK2 in turn phosphorylates three LepRb tyrosine residues (Tyr985 ...

Hypothalamic expression of anorexigenic and orexigenic hormone receptors in obese females Neotomodon alstoni: Effect of fasting

Obesity is a world problem that requires a better understanding of its physiological and genetic basis, as well as the mechanisms by which the hypothalamus controls feeding behavior. The volcano mouse Neotomodon alstoni develops obesity in captivity when fed with regular chow diet, providing a novel model for the study of obesity. Females develop obesity more often than males; therefore, in this study, we analysed in females, in proestrous lean and obese, the differences in hypothalamus expression of receptors for leptin, ghrelin (growth hormone secretagogue receptor GHS-R), and VPAC, and correlates for plasma levels of total ghrelin. The main comparisons are between mice fed ad libitum and mice after 24 hours of fasting. Mice above 65 g body weight were considered obese, based on behavioral and physiological parameters such as food intake, plasma free fatty acids, and glucose tolerance. Hypothalamic tissue from obese and lean mice was analysed by western blot. Our results indicate that after ad libitum food access, obese mice show no significant differences in hypothalamic leptin receptors, but a significant increase of 60% in the GHS-R, and a nearly 62% decrease in VPAC2 was noted. After a 24-hour fast, plasma ghrelin increased nearly two fold in both lean and obese mice; increases of hypothalamic leptin receptors and GHS-R were also noted, while VPAC2 did not change significantly; levels of plasma free fatty acids were 50% less after fasting in obese than in lean animals. Our results indicate that in obese N. alstoni mice, the levels of orexigenic receptors in the hypothalamus correlate with overfeeding, and the fact that lean and obese females respond in different ways to a metabolic demand such as a 24-hour fast.

Molecular cloning, characterization and expression profiles of multiple leptin genes and a leptin receptor gene in orange-spotted grouper (Epinephelus coioides)

Leptin plays key roles in body weight regulation, energy metabolism, food intake, reproduction and immunity in mammals. However, its function in teleosts is still unclear. In the present study, two leptin genes (gLepA and gLepB) and one leptin receptor gene (gLepR) were cloned and characterized in orange-spotted grouper (Epinephelus coioides). The cDNAs of gLepA and gLepB were 671bp and 684bp in length, encoding for proteins of 161 amino acid (aa) and 158 aa, respectively. The three-dimensional (3D) structures modeling of gLepA and gLepB showed strong conservation of tertiary structure with that of other vertebrates. The total length of gLepR cDNA was 4242bp, encoding a protein of 1169 aa which contained all functionally important domains conserved among vertebrate LEPR. Tissue distribution analysis showed that gLepA was highly expressed in cerebellum, liver and ovary, while gLepB mRNA abundantly in the brain regions, as well as in the ovary with some extend. The gLepR was mainly expressed in kidney, head kidney and most of brain regions. Analysis of expression profiles of gLep and gLepR genes during the embryonic stages showed that high expression of gLepR was observed in the brain vesicle stage, while neither gLepA nor gLepB mRNA was detected during different embryonic stages. Finally, fasting and refeeding experiments were carried out to investigate the possible function of leptin genes in food intake and energy metabolism, and the results showed that a significant increase of gLepA expression in the liver was induced by food deprivation in both short-term (7days) and long-term (3weeks) fasting and gLepA mRNA upregulation was eliminated after refeeding, while gLepB wasn’t detected in the liver of grouper during fasting. No significant differences in hypothalamic leptin and leptin receptor expression were found during short-term fasting and refeeding. Hepatic expression of gLepA mRNA increased significantly 9h after a single meal. These results suggested gLepA, other than gLepB, functioned in the regulation of energy metabolism and food intake in this Perciform fish.