Topographic mapping of VMH --> arcuate nucleus microcircuits and their reorganization by fasting

In the hypothalamic arcuate nucleus (ARC), pro-opiomelanocortin (POMC) neurons inhibit feeding and neuropeptide-Y (NPY) neurons stimulate feeding. We tested whether neurons in the ventromedial hypothalamic nucleus (VMH), a known satiety center, activate anorexigenic neuronal pathways in the ARC by projecting either excitatory synaptic inputs to POMC neurons and/or inhibitory inputs to NPY neurons. Using laser scanning photostimulation in brain slices from transgenic mice, we found that POMC and NPY neurons, which are interspersed in the ARC, are nevertheless regulated by anatomically distinct synaptic inputs. POMC neurons received strong excitatory input from the medial VMH (mVMH), whereas NPY neurons did not and, instead, received weak inhibitory input only from within the ARC. The strength of the excitatory input from the mVMH to POMC neurons was diminished by fasting. These data identify a new molecularly defined circuit that is dynamically regulated by nutritional state in a manner consistent with the known role of the VMH as a satiety center.     

AGRP neurons are sufficient to orchestrate feeding behavior rapidly and without training

Two intermingled hypothalamic neuron populations specified by expression of agouti-related peptide (AGRP) or pro-opiomelanocortin (POMC) positively and negatively influence feeding behavior, respectively, possibly by reciprocally regulating downstream melanocortin receptors. However, the sufficiency of these neurons to control behavior and the relationship of their activity to the magnitude and dynamics of feeding are unknown. To measure this, we used channelrhodopsin-2 for cell type-specific photostimulation. Activation of only 800 AGRP neurons in mice evoked voracious feeding within minutes. The behavioral response increased with photoexcitable neuron number, photostimulation frequency and stimulus duration. Conversely, POMC neuron stimulation reduced food intake and body weight, which required melanocortin receptor signaling. However, AGRP neuron-mediated feeding was not dependent on suppressing this melanocortin pathway, indicating that AGRP neurons directly engage feeding circuits. Furthermore, feeding was evoked selectively over drinking without training or prior photostimulus exposure, which suggests that AGRP neurons serve a dedicated role coordinating this complex behavior.     

The different time courses of reading different levels of Chinese characters: an ERP study

AMP-activated protein kinase (AMPK) is an energy sensor that is activated by the increase of intracellular AMP:ATP ratio. AMPK in the hypothalamic arcuate nucleus (ARC) is activated during fasting and the activation of AMPK stimulates food intake. To clarify the pathway underlying AMPK-induced feeding, we monitored the activity of single ARC neurons by measuring cytosolic Ca(2+) concentration ([Ca(2+)](i)) with fura-2 fluorescence imaging. An AMPK activator, AICA-riboside (AICAR), at 200 μM increased [Ca(2+)](i) in 24% of ARC neurons. AMPK and acetyl CoA carboxylase were phosphorylated in the neurons with [Ca(2+)](i) responses to AICAR. AICAR-induced [Ca(2+)](i) increases were inhibited by Ca(2+)-free condition but not by thapsigargin, suggesting that AICAR increases [Ca(2+)](i) through Ca(2+) influx from extracellular space. Among AICAR-responding ARC neurons, 38% were neuropeptide Y (NPY)-immunoreactive neurons while no proopiomelanocortin (POMC)-immunoreactive neuron was observed. Intracerebroventricular administration of AICAR increased food intake, and the AICAR-induced food intake was abolished by the co-administration of NPY Y1 receptor antagonist, 1229U91. These results indicate that the activation of AMPK leads to the activation of ARC NPY neurons through Ca(2+) influx, thereby causing NPY-dependent food intake. These mechanisms could be implicated in the stimulation of food intake by physiological orexigenic substances.     

Age-related decrease in proopiomelanocortin gene expression in the arcuate nucleus of the male rat brain.

The decline in reproductive function with aging is due in part to decreased gonadotropin-releasing hormone (GnRH) secretion. beta-Endorphin (beta E), an endogenous opioid peptide derived from proopiomelanocortin (POMC), is thought to exert a tonic inhibitory effect upon hypothalamic GnRH secretion. We tested the hypothesis that the age-related decrease in GnRH secretion in male rats is due to increased beta E synthesis, by comparing POMC mRNA levels in the arcuate nucleus (ARC) of intact young, middle-aged and old male rats. In an initial study (Study 1), sixteen 20-microns coronal sections each from the ARC of 3- (n = 5) and 23-month-old (n = 4) male Fischer 344 rats were anatomically matched and analyzed. In a second study (Study 2), four anatomically matched sections of caudal arcuate nucleus from 3- (n = 4), 11- (n = 7) and 23-month-old (n = 5) male rats were compared. POMC mRNA levels were quantitated by in situ hybridization histochemistry, using a 35S-labeled oligodeoxynucleotide probe complementary to a portion of rat POMC cDNA and computerized image analysis. The number of grains per cell and cells per section were used as indices of cellular POMC mRNA content and the number of neurons expressing the POMC gene, respectively. Cellular POMC mRNA content was significantly lower in old compared to young animals (Study 1: 54 +/- 3 vs. 74 +/- 2 grains/cell, p less than 0.01; Study 2: 59 +/- 2 vs. 71 +/- 2 grains/cell, p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of oleic acid on distinct populations of neurons in the hypothalamic arcuate nucleus are dependent on extracellular glucose levels

Pharmacological manipulation of fatty acid metabolism in the hypothalamic arcuate nucleus (ARC) alters energy balance and glucose homeostasis. Thus, we tested the hypotheses that distinctive populations of ARC neurons are oleic acid (OA) sensors that exhibit a glucose dependency, independent of whether some of these OA sensors are also glucose-sensing neurons. We used patch-clamp recordings to investigate the effects of OA on ARC neurons in brain slices from 14- to 21-day-old Sprague-Dawley (SD) rats. Additionally, we recorded spontaneous discharge rate in ARC neurons in 8-wk-old fed and fasted SD rats in vivo. Patch-clamp studies showed that in 2.5 mM glucose 12 of 94 (13%) ARC neurons were excited by 2 microM OA (OA-excited or OAE neurons), whereas six of 94 (6%) were inhibited (OA-inhibited2.5 or OAI2.5 neurons). In contrast, in 0.1 mM glucose, OA inhibited six of 20 (30%) ARC neurons (OAI0.1 neurons); none was excited. None of the OAI0.1 neurons responded to OA in 2.5 mM glucose. Thus OAI2.5 and OAI0.1 neurons are distinct. Similarly, in seven of 20 fed rats (35%) the overall response was OAE-like, whereas in three of 20 (15%) it was OAI-like. In contrast, in fasted rats only OAI-like response were observed (three of 15; 20%). There was minimal overlap between OA-sensing neurons and glucose-sensing neurons. In conclusion, OA regulated three distinct subpopulations of ARC neurons in a glucose-dependent fashion. These data suggest that an interaction between glucose and fatty acids regulates OA sensing in ARC neurons.     

Energy balance and hypothalamic effects of a high-protein/low-carbohydrate diet

Diets high in fat or protein and extremely low in carbohydrate are frequently reported to result in weight loss in humans. We previously reported that rats maintained on a low-carbohydrate-high fat diet (LC-HF) consumed similar kcals/day as chow (CH)-fed rats and did not differ in body weight after 7 weeks. LC-HF rats had a 45% decrease in POMC expression in the ARC, decreased plasma insulin, and increased plasma leptin and ghrelin. In the present study we assessed the effects of a low-carbohydrate-high-protein diet (HP: 30% fat, 65% protein, and 5% CHO) on body weight, caloric intake, plasma hormone levels and hypothalamic gene expression. Male rats (n=16) were maintained on CH or HP for 4 weeks. HP rats gained significantly less weight than CH rats (73.4+/-9.4 and 125.0+/-8.2 g) and consumed significantly less kcals/day (94.8+/-1.5 and 123.6+/-1.1). Insulin was significantly reduced in HP rats (HP: 1.8+/-0.6 vs. CH: 4.12+/-0.8 ng/ml), there were no differences between groups in plasma leptin and plasma ghrelin was significantly elevated in HP rats (HP: 127.5+/-45 vs. CH: 76.9+/-8 pg/ml). Maintenance on HP resulted in significantly increased ARC POMC (HP: 121+/-10.0 vs. 100+/-5.9) and DMH NPY (HP: 297+/-82.1 vs. CH: 100+/-37.7) expression compared to CH controls. These data suggest that the macronutrient content of diets differentially influences hypothalamic gene expression in ways that can affect overall intake.     

The effect of leptin on feeding-regulating neurons in the rat hypothalamus.

Intense immunoreactivity for the leptin receptor was detected in the hypothalamic arcuate nucleus (ARC), ventromedial nucleus (VMH), and lateral hypothalamus (LH) by immunohistochemistry. Cytosolic Ca2+ concentration ([Ca2+]i) in single neurons isolated from the ARC, VMH and LH was measured with dual wavelength fura-2 fluorescence imaging. A reduction of the superfusate glucose concentration from 10 to 1 mM increased [Ca2+]i in 21% of ARC neurons and 22% of LH neurons. Leptin at 0.1 nM inhibited the [Ca2+]i increase in 66 and 64% of these glucose-sensitive ARC and LH neurons, respectively. Inversely, 10 mM glucose increased [Ca2+]i in 49% of the VMH neurons, and 0.1 nM leptin at 1 mM glucose also increased [Ca2+]i in 84% of these glucose-responsive neurons. These results reveal that leptin inhibits the ARC and LH neurons and stimulates the VMH neurons via the leptin receptor expressed in these cells.     

Leptin regulated calcium channels of neuropeptide Y and proopiomelanocortin neurons by activation of different signal pathways

The fat-derived hormone leptin regulates food intake and body weight in part by modulating the activity of neuropeptide Y (NPY) and proopiomelanocortin (POMC) neurons in the hypothalamic arcuate nucleus (ARC). To investigate the electrophysiological activity of these neurons and their responses to leptin, we recorded whole-cell calcium currents on NPY and POMC neurons in the ARC of rats, which we identified by morphologic features and immunocytochemical identification at the end of recording. Leptin decreased the peak amplitude of high voltage–activated calcium currents (I_HVA) in the isolated neurons from ARC, which were subsequently shown to be immunoreactive for NPY. The inhibition was prevented by pretreatment with inhibitors of Janus kinase 2 (JAK2) and mitogen-activated protein kinases (MAPK). In contrast, leptin increased the amplitude of I_HVA in POMC-containing neurons. The stimulations of I_HVA were inhibited by blockers of JAK2 and phosphatidylino 3-kinase (PI3-k). Both of these effects were counteracted by the L-type calcium channel antagonist nifedipine, suggesting that L-type calcium channels were involved in the regulation induced by leptin. These data indicated that leptin exerted opposite effects on these two classes of neurons. Leptin directly inhibited I_HVA in NPY neurons via leptin receptor (LEPR) –JAK2–MAPK pathways, whereas evoked I_HVA in POMC neurons by LEPR–JAK2–PI3-k pathways. These neural pathways and intracellular signaling mechanisms may play key roles in regulating NPY and POMC neuron activity, anorectic action of leptin and, thereby, feeding.     

Up-regulation of gene expression by hypoxia is mediated predominantly by hypoxia-inducible factor 1 (HIF-1)

The hypoxia-inducible factor 1 (HIF-1) plays a critical role in cellular responses to hypoxia. The aim of the present study was to evaluate which genes are induced by hypoxia, and whether this induction is mediated by HIF-1, by expression microarray analysis of wt and HIF-1alpha null mouse fibroblasts. Forty-five genes were up-regulated by hypoxia and 40 (89%) of these were regulated by HIF-1. Of the 114 genes down-regulated by hypoxia, 19 (17%) were HIF-1-dependent. All glycolytic enzymes were strongly up-regulated by hypoxia in a HIF-1-dependent manner. Genes already known to be related to hypoxia, such as glucose transporter 1, BNIP3, and hypoxia-induced gene 1, were induced. In addition, multiple new HIF-1-regulated genes were identified, including genes involved in metabolism (adenylate kinase 4, galactokinase), apoptosis (galectin-3 and gelsolin), and invasion (RhoA). Genes down-regulated by hypoxia were involved in cytoskeleton maintenance (Rho kinase), mRNA processing (heterogeneous nuclear ribonucleoprotein H1 and splicing factor), and DNA repair (REV3). Furthermore, seven cDNAs from genes with unknown function or expressed sequence tags (ESTs) were up-regulated and 27 such cDNAs were down-regulated. In conclusion, hypoxia causes down- rather than up-regulation of gene expression and HIF-1 seems to play a major role in the regulation of hypoxia-induced genes.     

Somatostatin modulation of excitatory synaptic transmission between periventricular and arcuate hypothalamic nuclei in vitro

Hypophysiotropic somatostatin (SRIF) and growth hormone-releasing hormone (GHRH) neurons are primarily involved in the neurohormonal control of growth hormone (GH) secretion. They are located in periventricular (PEV) and arcuate (ARC) hypothalamic nuclei, respectively, but their connectivity is not well defined. To better understand the neuronal network involved in the control of GH secretion, connections from PEV to ARC neurons were reconstructed in vitro and neuronal phenotypes assessed by single-cell multiplex RT-PCR. Of 814 stimulated PEV neurons, monosynaptic responses were detected in only 45 ARC neurons. Monosynaptic excitatory currents were detected in 29 ARC neurons and inhibitory currents in 16, indicating a 2/1 ratio for excitatory versus inhibitory connections. Galanin (GAL), NPY, pro-opiomelanocortin (POMC), and SRIF mRNAs were detected in neurons from both nuclei but GHRH mRNA almost exclusively in ARC. Among the five SRIF receptors, only sst1 and sst2 were expressed, in 94% of ARC and 59% of PEV neurons, respectively. Of 128 theoritical combinations between neuropeptides and sst receptors, only 22 were represented in PEV and 25 in ARC. For PEV neurons, neuropeptide phenotypes did not influence excitatory connections. However, the occurrence of presynaptic sst receptors on GAL and SRIF PEV neurons significantly increased their probability of connection to ARC neurons. GHRH ARC neurons expressing sst2, but not sst1, receptors were always connected with PEV neurons. Physiological responses to sst1 (CH-275) or sst2 (Octreotide) agonists were always correlated with the detection of respective sst mRNAs. In conclusion, 1) SRIF-modulated excitatory transmission develops in vitro from PEV to ARC neurons, 2) ARC GHRH neurons bearing sst2 receptors appears directly controlled by fast glutamatergic transmission from PEV neurons simultaneously expressing one to four neuropeptides, 3) GHRH neurons bearing sst1 receptors lack this control, and 4) these results suggest that fast excitatory neurotransmission and neuropeptide modulation can derive from a small subset of PEV hypothalamic neurons targeted at ARC neuronal subpopulations.     

Characterization of apolipoprotein A-IV in brain areas involved in energy homeostasis

Apolipoprotein A-IV (apo A-IV) is a satiation protein synthesized in the small intestine and hypothalamus. To further understand its anorectic mechanisms, we used immunohistochemical techniques to characterize the distribution of apo A-IV in brain areas involved in energy homeostasis. Dense apo A-IV staining was detected in the arcuate (ARC) and ventromedial hypothalamic nuclei with less staining in cells in the paraventricular and dorsomedial nuclei. In the brainstem, apo A-IV staining was found in the nucleus of the solitary tract. Double-staining immunohistochemistry revealed co-existence of apo A-IV with neuronal nuclei (a neuronal marker), but less with glial fibrillary acidic protein (a glial marker), in ARC, suggesting that apo A-IV is largely present in neurons. In the ARC, apo A-IV was co-localized with pro-opiomelanocortin (POMC), and apo A-IV administration stimulated hypothalamic POMC gene expression, suggesting that the brain apo A-IV system suppresses food intake by stimulating the ARC POMC system. To ascertain whether the apo A-IV detected in the brain is derived from the circulation, (125)I-labeled recombinant rat apo A-IV was intravenously injected into mice. No increase of radioactive apo A-IV was found in the brain, consistent with a lack of uptake of co-injected (99m)Tc-labeled albumin, indicating that circulating apo A-IV is unable to cross the blood brain barrier. These data collectively support the hypothesis that apo A-IV, produced by neuronal cells, may exert its anorectic action by interacting with catabolic regulatory neuropeptides.     

The hypothalamus and the control of energy homeostasis: different circuits, different purposes.

The hypothalamus regulates many aspects of energy homeostasis, adjusting both the drive to eat and the expenditure of energy in response to a wide range of nutritional and other signals. It is becoming clear that various neural circuits operate to different degrees and probably serve specific functions under particular conditions of altered feeding behaviour. This review will discuss this functional diversity by illustrating hypothalamic neurones that express neuropeptide Y (NPY), the melanocortin-4 receptor (MC4-R) and the orexins. NPY neurones in the arcuate nucleus (ARC) release NPY, a powerful inducer of feeding and obesity, in the paraventricular nucleus (PVN) and the lateral hypothalamic area (LHA). ARC-NPY neurones are inhibited by leptin and insulin and become overactive when levels of these hormones fall during undernutrition. They may function physiologically to protect against starvation. With disruption of the inhibitory leptin signals due to gene mutations, the NPY neurones are overactive, which contributes to hyperphagia and obesity in the ob/ob and db/db mice and fa/fa Zucker rat. The MC4-R is activated by alpha-melanocyte-stimulating hormone [alpha-MSH; a cleavage product of pro-opiomelanocortin (POMC), which is expressed in the other ARC neurones] and inhibits feeding. This effect is antagonised by agouti gene-related peptide (AGRP), which is coexpressed by the ARC-NPY neurones only. Activation of MC4-R, possibly mediated by blockade of AGRP release, appears to restrain overeating of a palatable diet. This response may be programmed by a transient rise in leptin soon after presentation of palatable food, and rats that fail to do this will overeat and become obese. Orexin-A and -B (corresponding to hypocretins 1 and 2) are expressed in specific LHA neurones. These have extensive reciprocal connections with many areas involved in appetite control, including the nucleus of the solitary tracts (NTS), which relays vagal afferent satiety signals from the viscera. Orexin neurones also have close anatomical connections with LHA glucose-sensitive neurones. Orexin-A induces acute feeding but does not cause obesity. Orexin neurones are stimulated by hypoglycaemia partly via the NTS and inhibited by food ingestion. These neurones may therefore be involved in the severe hyperphagia of hypoglycaemia and short-term control of feeding.     

Ins2Akita mice exhibit hyperphagia and anxiety behavior via the melanocortin system

Elevated anxiety symptoms have been reported to be present in many patients with diabetes mellitus. The underlying mechanisms by which diabetes mellitus influences behavior remain to be determined. We assessed feeding and anxiety behaviors in spontaneously diabetic Ins2Akita mice. We measured blood glucose, body weight, and food and water intakes in C57BL/6 heterozygote Ins2Akita mice. The behavioral properties of Ins2Akita mice were assessed in an open-field test and an elevated plus-maze. The gene expression of hypothalamic neuropeptides was examined in non-food-deprived Ins2Akita mice. Body weights of the Ins2Akita mice were less than those of the age-matched C57BL/6 mice, as controls. Food and water intakes were increased in the Ins2Akita mice. In the open-field test, the Ins2Akita mice had decreased locomotor activity and increased immobilization time. The Ins2Akita mice exhibited anxiety behavior in the elevated plus-maze. RT-PCR analysis showed decreased proopiomelanocortin (POMC) mRNA expression and increased agouti-related protein (AGRP) mRNA expression in Ins2Akita mice. There were no significant differences in hypothalamic ghrelin mRNA expression. These observations indicate that Ins2Akita mice, which are characterized by hypoinsulinemia and hyperglycemia, exhibited hyperphagia and anxiety behavior; the mechanism of action involved the activation of hypothalamic AGRP and the inactivation of hypothalamic POMC. In addition, Ins2Akita mice are a useful model for understanding the mechanisms involved in the psychological complications of diabetes mellitus. Further, melanocortin systems may be therapeutic targets not only for diabetes but also for its associated complications.     

Glucose-sensitive neurons in the rat arcuate nucleus contain neuropeptide Y.

Glucose is known to regulate the activity of the hypothalamic feeding centers. Neuropeptide Y (NPY)-containing neurons in the hypothalamic arcuate nucleus (ARC) have been implicated in the stimulation of feeding. We examined the presence of glucose-sensitive neurons in the ARC and their coincidence with NPY-containing neurons. Cytosolic Ca2+ concentration ([Ca2+]i) in single ARC neurons isolated from rat hypothalamus was measured with fura-2 fluorescence imaging; the cells were then stained immunocytochemically with an anti-NPY antiserum. Lowering the glucose concentration from 10 to 1 mM increased [Ca2+]i in 36 out of 180 neurons (20%), the majority of which (34 neurons, 94%) were immunoreactive for NPY. In conclusion, the ARC contains glucose-sensitive NPY-containing neurons. The suggested role of these neurons is to transduce a reduction in the glucose concentration in the brain to the release of NPY and, subsequently, stimulation of feeding.     

Nicotinamide induces male-specific body weight loss in the postnatal period through molecular regulation of the hypothalamus and liver

Molecular mechanisms of body weight control have been discovered recently and much research focuses on the hypothalamic regulation of food intake and the hepatic regulation of glucose utility. We previously reported that postnatal nicotinamide treatment reduced brain dopamine and body weight. To further investigate the differential effects of nicotinamide-mediated body weight loss, nicotinamide (i.p. 100 mg/kg) was injected into postnatal and adult mice twice a week for 4 weeks. Interestingly, following nicotinamide treatment, male postnatal mice displayed reduced body weight and spontaneous motor activity. No significant changes were observed in adult and postnatal female mice or adult male mice following nicotinamide treatment. In male postnatal mice, hypothalamic agouti-related peptide (AGRP) and proopiomelanocortin (POMC) levels were increased in the arcuate nucleus following nicotinamide treatment. Neuropeptide Y (NPY) levels were unchanged in both male and female mice. Additionally, nicotinamide-injected male postnatal mice had increased glucose 6-phosphatase (G6Pase) and decreased phosphoenolpyruvate carboxykinase (PEPCK) expression in liver. These results indicate that hypothalamic POMC and hepatic PEPCK are important molecules that mediate nicotinamide-induced weight loss in postnatal male mice.     

Hormone and glucose signalling in POMC and AgRP neurons

In the wake of the obesity pandemic, increased research efforts are under way to define how peripheral hormones and metabolites regulate energy homeostasis. The melanocortin system, comprising anorexigenic proopiomelanocortin (POMC) expressing neurons and orexigenic agouti-related protein (AgRP)/neuropeptide Y (NPY) coexpressing neurons in the arcuate nucleus of the hypothalamus are crucial for normal energy homeostasis both in rodents and humans. They are regulated by peripheral hormones such as leptin and insulin, as well as nutrients such as glucose, amino acids and fatty acids. Although much progress has been made, recent reports continue to underline how restricted our understanding of POMC and AgRP/NPY neuron regulation by these signals is. Importantly, ATP-dependent potassium (K_ATP) channels are regulated both by ATP (from glucose metabolism) and by leptin and insulin, and directly control electrical excitability of both POMC and AgRP neurons. Thus, this review attempts to offer an integrative overview about how peripheral signals, particularly leptin, insulin and glucose, converge on a molecular level in POMC and AgRP neurons of the arcuate nucleus of the hypothalamus to control energy homeostasis.     

Tachykinin-induced changes in β-endorphin gene expression in the rat arcuate nucleus

Previous neuroanatomical data have indicated the presence of synaptic connections between tachykinergic terminals and proopiomelanocortin (POMC) neurons in the arcuate nucleus. Consequently, tachykinins may regulate the activity of POMC neurons. To evaluate the functional signification of this regulation, the effect of intracerebroventricular injections of neurokinin A (NKA) on POMC mRNA levels was studied by using in situ hybridization. Repeated injection of NKA (40 μg/animal per day during 3 days) induced a 48% increase in POMC mRNA expression as compared to NaCl injected control animals. In conclusion the results of this study show an excitatory effect of tachykinin on POMC neurons and suggest a direct and/or indirect excitatory control of POMC neuronal activity by endogenous tachykinins.     

Hypothalamic mTOR pathway mediates thyroid hormone-induced hyperphagia in hyperthyroidism

Hyperthyroidism is characterized in rats by increased energy expenditure and marked hyperphagia. Alterations of thermogenesis linked to hyperthyroidism are associated with dysregulation of hypothalamic AMPK and fatty acid metabolism; however, the central mechanisms mediating hyperthyroidism-induced hyperphagia remain largely unclear. Here, we demonstrate that hyperthyroid rats exhibit marked up-regulation of the hypothalamic mammalian target of rapamycin (mTOR) signalling pathway associated with increased mRNA levels of agouti-related protein (AgRP) and neuropeptide Y (NPY), and decreased mRNA levels of pro-opiomelanocortin (POMC) in the arcuate nucleus of the hypothalamus (ARC), an area where mTOR co-localizes with thyroid hormone receptor-α (TRα). Central administration of thyroid hormone (T3) or genetic activation of thyroid hormone signalling in the ARC recapitulated hyperthyroidism effects on feeding and the mTOR pathway. In turn, central inhibition of mTOR signalling with rapamycin in hyperthyroid rats reversed hyperphagia and normalized the expression of ARC-derived neuropeptides, resulting in substantial body weight loss. The data indicate that in the hyperthyroid state, increased feeding is associated with thyroid hormone-induced up-regulation of mTOR signalling. Furthermore, our findings that different neuronal modulations influence food intake and energy expenditure in hyperthyroidism pave the way for a more rational design of specific and selective therapeutic compounds aimed at reversing the metabolic consequences of this disease.     

The effect of intracerebroventricular infusions of leptin on the immunoreactivity of neuropeptide Y and gonadotrophin releasing hormone neurons in the hypothalamus of prepubertal sheep in conditions of short fasting

In the study we evaluated the effects of infusion of exogenous leptin to the third ventricle of the brain on the expression of immunoreactive (ir) neuropeptide Y (NPY) neurons in the hypothalamus and ir gonadotrophin releasing hormone (GnRH) nerve terminals in the median eminence of prepubertal lambs in the conditions of short fasting. Merino female sheep (n=16) were randomly divided into four groups, two fed with standard feeds and two fasted for 72 h. One standard and one fasted groups were infused with Ringer saline (controls), remaining standard and fasted groups with leptin (25 microg/120 microl/h), for 4 h during three consecutive days, and then slaughtered. Ir NPY and ir GnRH were localized by immunohistochemistry using specific polyclonal antibodies. Detection of both hormones was followed by the image analysis and expressed as the percent area stained and integral density of immunostaining. In the hypothalami from all groups the ir NPY perikarya and varicose nerve fibers were localized in three distinct sub-areas, in the arcuate (ARC), paraventricular and periventricular nuclei. In fasted sheep the percent area and integral density for immunoreactivity of NPY increased significantly (P<0.001) in three sub-areas compared to the standard-fed animals. Leptin infusion lowered the both parameters (P<0.001) but solely in the ARC NPY population of fasted sheep. The percent area and integral density of immunostaining for ir GnRH in fasted sheep revealed the augmentation (P<0.001) compared to standard-fed sheep. Leptin infusions diminished (P<0.001) both parameters in fasted, without effects in standard-fed lambs. In conclusion, the enhanced by fasting immunoreactivity of the ARC NPY perikarya and varicose nerve fibers and restrained immunoreaction of GnRH terminals in the median eminence were reversed by exogenous leptin. It is suggested that leptin can affect GnRH release via ARC NPY neurons in conditions of deficit of nutrients in prepubertal, female lambs.