Neuropeptide Y Gene Expression Research Articles

Neuromodulatory Effects Induced by the Association of Moringa oleifera Lam., Tribulus terrestris L., Rhodiola rosea Lam., and Undaria pinnatidifida Extracts in the Hypothalamus.

The present study investigated the role of a commercial formulation constituted by herbal extracts from Rhodiola rosea, Undaria pinnatifida, Tribulus terrestris, and Moringa oleifera. The formulation was analysed for determining the content in total phenols and flavonoids and scavenging/reducing properties. The formulation was also tested on isolated mouse hypothalamus in order to investigate effects on serotonin, dopamine, neuropeptide Y (NPY), and orexin A. The gene expression of gonadrotopin releasing hormone (GnRH) was also assayed. The formulation was able to reduce dopamine and serotonin turnover, and this could be related, albeit partially, to the capability of different phytochemicals, among which hyperoside and catechin to inhibit monoaminooxidases activity. In parallel, the formulation was effective in reducing the gene expression of NPY and orexin-A and to improve the gene expression of GnRH. In this context, the increased GnRH gene expression induced by the formulation may contribute not only to improve the resistance towards the stress related to ageing, but also to prevent the reduction of libido that could be related with a stimulation of the serotoninergic pathway. According to the in silico analysis, hyperoside could play a pivotal role in modulating the gene expression of GnRH. Regarding NPY and orexin A gene expression, no direct interactions between the formulation phytochemicals and these neuropeptides were anticipated; thus, suggesting that the pattern of gene expression observed following exposure of the hypothalamus to the formulation may be secondary to inhibitory effects of dopamine and serotonin turnover. Concluding, the present study demonstrated the efficacy of the formulation in exerting neuromodulatory effects at the hypothalamic level; thus, suggesting the potential to contrast stress and fatigue.

Effects of Thymoquinone on Hypothalamic NPY and AgRP Gene Expression in Intact and Hypothyroidism- Model Rats

Background & Objective: Thymoquinone is the most important compound of Nigella sativa, which stimulates the activity of thyroid axis in healthy individuals and people with hypothyroidism. The present study investigated the effects of thymoquinone on Neuropeptide Y (NPY) and Agouti-dependent peptide (AgRP) gene expression in the hypothalamus of healthy individuals and hypothyroidism- model rats.
 Material & Methods: Twenty male Wistar rats weighing 190- 220g were used. Hypothyroidism was induced by a daily consumption of Methimazole (20mg/kg) for 42 days via drinking water. Control rats received the intraperitoneal injections of saline. Intact or hypothyroid rats received the intraperitoneal injections of thymoquinone (10mg/kg) for 15 days. One day after last injection, the thyroid gland and hypothalamic samples were dissected. Thyroid gland samples were used for histological study. Relative gene expression of hypothalamic NPY and AgRP was determined by real-time polymerase chain reaction.
 Results: Thymoquinone significantly declined the NPY and AgRP gene expressions in the hypothalamus of intact rats in comparison with control group. Induction of hypothyroidism results in a remarkable increase in the NPY and AgRP gene expressions compared to control rats. In hypothyroid rats receiving thymoquinone, the mean relative NPY and AgRP gene expressions showed an insignificant decrease compared to hypothyroid group.
 Conclusion: Because AgRP/NPY signaling pathway exerts inhibitory effects on thyroid gland function, thymoquinone may stimulate thyroid axis activity partly via inhibiting the hypothalamic AgRP/NPY gene expression in intact rats. In hypothyroidism, used dose of thymoquinone may not able to cause a significant decrease in AgRP/NPY gene expression due to its increased levels.

Inverse age-related changes between hypothalamic NPY and KISS1 gene expression during pubertal initiation in male rhesus monkey

The neuroendocrine mechanism underlying the sinusoidal wave nature of gonadotropin–releasing hormone pulse generator activity from infantile to adult age still needs to be meticulously defined. Direct inhibition of kisspeptin neurons by neuropeptide Y (NPY) and close intimacy between the two rekindle the importance of these two neuropeptides controlling reproductive axis activity. Thus, the present study was undertaken to decipher simultaneous fluctuations and to profile correlative changes in the relative expression of KISS1, NPY, and their receptor genes from the mediobasal hypothalamus of infant (n = 3), juvenile, pre-pubertal, and adult (n = 4 in each stage) male rhesus monkey (Macaca mulatta) by RT-qPCR. Significant elevation (p < 0.05−0.01) in KISS1 and KISS1R and low (p < 0.05) expression in NPY and NPY1R mRNA in the adult group as compared to the pre-pubertal group was observed. Moreover, significantly high (p < 0.05) expression of NPY and NPY1R mRNA with non-significant (p> 0.05) decline in KISS1 and KISS1R in pre-pubertal animals in comparison to infants describe inverse correlative age-associated changes during pubertal development. Current findings imply that NPY may contribute as a neurobiological brake for the dormancy of kisspeptin neurons before pubertal onset, while dwindling of this brake is likely to occasion kisspeptin dependent hypothalamic–pituitary–gonadal axis activation at puberty. These findings may help in the development of clinical and therapeutic strategies to regulate fertility in humans.

Influence of peripheral lipopolysaccharide (LPS) on feed intake, body temperature and hypothalamic expression of neuropeptides involved in appetite regulation in broilers and layer chicks

ABSTRACT 1. This study examined the expression of genes related to appetite-regulating neuropeptides in the hypothalamus of broiler and layer chicks (Gallus gallus) after intraperitoneal (IP) injection of lipopolysaccharide (LPS). 2. Both broiler and layer chicks received (n = 10 per group) LPS at doses of 0 and 200 µg and feed intake was measured up to 6 h after injection. In a further experiment, (n = 8 per group) mRNA abundance of some hypothalamic neuropeptides was measured 2 h after injection. The rectal temperature of each chick was measured before and 2 h post-injection. 3. Feed intake was significantly decreased by LPS from 2 h after injection and thereafter, while the rectal temperature did not change. 4. LPS decreased the expression of appetite-enhancing neuropeptides: neuropeptide Y (NPY) and agouti-related peptide (AgRP) in broilers and, NPY in layer chicks. The expression of appetite-suppressing neuropeptides (corticotrophin-releasing factor (CRF), proopiomelanocortin (POMC) and, cocaine and amphetamine regulated-transcript (CART) was not changed in broilers, while CRF tended to decrease and POMC was significantly decreased in layers. The abundance of the cytokine tumour necrosis factor-alpha (TNF-α) did not change in broilers but was decreased in layers. 5. The findings indicated that the reduction in gene expression of hypothalamic appetite-enhancing neuropeptides NPY and AgRP is responsible for anorexia caused by LPS at a dose that did not influence body temperature.

Relationship between perirenal adipose tissue neuropeptide Y and insulin resistance in nutritional transition model

To observe the changes of neuropeptide Y(NPY) expression in perirenal adipose tissue and its relationship with insulin resistance in the nutritional transition models of refeeding after calorie restriction. SPF Male SD rats, aged 8 weeks, were randomly divided into normal chow group and refeeding with normal chow after calorie restriction for 4 weeks group. NPY gene expression in perirenal adipose tissue were detected by real-time quantitative PCR at the end of 4 and 12 weeks, along with fasting plasma glucose, fasting serum lisulin, free fatty acids and average glucose infusion rate(GIR_(60-120)) of hyperinsulinemic-euglycemic clamp test for 60-120 minutes. NPY gene mRNA expression in perirenal adipose tissue was detected by real-time quantitative PCR. And the relationship between NPY gene expression and insulin resistance was detected by Spearman correlation analysis. The expression level of NPY gene in perirenal adipose tissue in caloric restriction for 4 weeks group was significantly increased by calorie restriction(P&lt;0. 01). After refeeding, the expression level of NPY gene in refeeding with normal group was still slightly increased, which was significantly higher than that in normal group at the end of the experiment(P&lt;0. 01). The levels of fasting plasma glucose and fasting insulin in caloric restriction for 4 weeks group decreased slightly, GIR_(60-120) increased slightly, but there were no statistical differences compared with normal group(P&gt;0. 05), but free fatty acid levels increased significantly(P&lt;0. 01). After refeeding, the levels of fasting insulin, free fatty acid in refeeding with normal group increased significantly, GIR_(60-120) decreased evidently(P&lt;0. 01), but the changes of fasting blood glucose were not obvious. The result of stepwise regression showed that the expression level of NPY gene in perirenal adipose tissue was closely related to GIR_(60-120) and fasting insulin, with R values of-0. 816 and 0. 789 respectively(R~2=0. 892, P&lt;0. 01). The result of correlation analysis showed that in the 4-week group, the mRNA expression level of NPY gene in perirenal adipose tissue was closely related to GIR_( 60-120)、fasting insulin and free fatty acid, with R values of-0. 765, 0. 716 and 0. 657 respectively(P&lt;0. 01). In the 12 week group, the mRNA expression level of NPY gene in perirenal adipose tissue was closely related to GIR_(60-120), fasting insulin and free fatty acid, with R values of-0. 853, 0. 622 and 0. 608 respectively(P&lt;0. 01). The mRNA expression of NPY gene in perirenal adipose tissue was closely related to indicators of insulin resistance. It is an important factor affecting insulin sensitivity.

SAT-603 Growth Hormone-Releasing Hormone (GHRH) Antagonists Stimulate Feeding in Mice

Growth hormone-releasing hormone (GHRH) is a hypothalamic neuropeptide which stimulates the synthesis and secretion of growth hormone (GH) in pituitary gland. GHRH was also found to modulate food intake in mammals. MIA-690 is a synthetic GHRH antagonist of the Miami (MIA) series with potent antitumor effects. To date, its role in hypothalamic feeding modulation has not been evaluated. In the present study, we aimed to investigate the effects of chronic MIA-690 administration on feeding behavior, locomotor activity and hypothalamic dopamine (DA), norepinephrine (NE), serotonin (5-hydroxytriptamine, 5-HT), orexigenic peptides [agouti-related peptide (AgRP) and neuropeptide Y (NPY)] and anorexigenic peptides [cocaine and amphetamine-regulated transcript (CART) and proopiomelanocortin (POMC)] activity.Adult C57/BL6 mice were treated daily for 4 weeks by subcutaneous administration of (5 µg) MIA-690 or vehicle solution. Food intake and body weight were recorded every 4 days throughout the study. Immediately after the last injection, locomotor activity in the home cage was recorded, and thereafter animals were sacrificed. Visceral, subcutaneous and brown fat depots were quickly excised and weighed. Hypothalamus was also dissected for evaluating gene expression of AgRP, NPY, CART and POMC by real-time reverse transcription polymerase chain reaction. In addition, hypothalamic DA, NE and 5-HT levels were measured by high performance liquid chromatography (HPLC) coupled to electrochemical detection.Our findings show that administration of MIA-690 increased food intake and body weight, without affecting locomotor activity. No difference was observed in visceral, subcutaneous and brown fat mass in animals treated with MIA-690 or vehicle. As for neuromodulatory effects, a significant increase of AgRP gene expression and NE levels, along with a reduction of 5-HT levels were found after MIA-690 treatment. On the other hand, we did not observe any alteration in NPY, POMC and CART gene expression, as well as DA levels, following MIA-690 administration.In conclusion, chronic peripheral administration of MIA-690 could play an orexigenic role paralleled by increased body weight. The stimulation of feeding could be mediated, at least in part, by increased AgRP gene expression and NE levels and decreased 5-HT levels, in the hypothalamus.

Neuropeptide Y neurons in the nucleus accumbens modulate anxiety-like behavior

Neuropeptide Y (NPY) is a 36-amino acid neuropeptide that is widely expressed in the central nervous system, including the cerebral cortex, nucleus accumbens (NAc) and hypothalamus. We previously analyzed the behavior of transgenic mice exclusively expressing an unedited RNA isoform of the 5-HT2C receptor. These mice showed decreased NPY gene expression in the NAc and exhibited behavioral despair, suggesting that NAc NPY neurons may be involved in mood disorder; however, their role in this behavior remained unknown. Therefore, in the present study, we investigated the functional role of NAc NPY neurons in anxiety-like behavior by examining the impact of specific ablation or activation of NAc NPY neurons using NPY-Cre mice and Cre-dependent adeno-associated virus. Diphtheria toxin-mediated ablation of NAc NPY neurons significantly increased anxiety-like behavior in the open field and elevated plus maze tests, compared with before toxin treatment. Moreover, chemogenetic activation of NAc NPY neurons reduced anxiety-like behavior in both behavioral tests compared with control mice. These results suggest that NPY neurons in the NAc are involved in the modulation of anxiety in mice.

Acupuncture Targeting SIRT1 in the Hypothalamic Arcuate Nucleus Can Improve Obesity in High-Fat-Diet-Induced Rats with Insulin Resistance via an Anorectic Effect

Objective: To investigate the anorexigenic and anti-obesity effectiveness of electroacupuncture (EA) on high-fat-diet-induced (HFDI) obese rats with insulin resistance (IR) and to reveal the possible mechanisms of EA affecting SIRT1 (silent mating type information regulation 2 homolog 1) in the central nervous system (CNS). Methods: We divided 60 rats into 6 groups. All interventions, including EA and intracerebroventricular administration, were performed after 8 weeks of model establishment. We tested obesity phenotypes like body weight (BW) gain; food intake; and IR levels including glucose infusion rate, intraperitoneal insulin tolerance test (IPITT), and intraperitoneal glucose tolerance test (IPGTT) during treatment. We detected protein expression and microscopic locations in hypothalamic SIRT1, the transcription factor FOXO1 (forkhead box protein O1), acetylated FOXO1 (Ac-FOXO1), pro-opiomelanocortin (POMC), and neuropeptide Y (NPY) via Western blotting and immunofluorescence, and monitored gene expression by real-time polymerase chain reaction. Results: Like the SIRT1 agonist, EA suppressed BW gain and IR levels in obese rats, but this was only partially blocked by the SIRT1 antagonist. EA could upregulate protein expression of hypothalamic SIRT1 and downregulate the acetylation level of FOXO1 in the hypothalamic arcuate nucleus (ARC), which decreased gene expression of NPY and increased that of POMC. The agonist targeted the hypothalamic SIRT1 gene, unlike EA, which targeted posttranscriptional regulation. Conclusion: EA could improve obesity in HFDI rats with IR via its anorectic effect. This effect targeted posttranscriptional regulation of the SIRT1 gene, which induced upregulation of ARC FOXO1 deacetylation and mediated the gene expression of POMC and NPY.

Relation of neuropeptide Y gene expression and genotyping with hypertension in chronic kidney disease

ObjectivesThe prognosis of high-risk patients might be greatly ameliorated using genetic predisposition risk factors. Sympathetic activity and innate immunity related to neuropeptide Y function may be related to dyslipidemia and atherosclerosis. The aim of this study is to detect the correlation between Neuropeptide Y (NPY) SNP rs16147 and its gene expression in chronic kidney disease with and without hypertension. MethodsThis study carried out on 150 subjects who were divided into 3 main groups group (I) 50 CKD patients with hypertension, group (II) 50 CKD patients without hypertension and group (III) 50 healthy individuals. Carotid intima media thickness (CIMT) was measured by Ultrasound. Kidney function test and lipid profile were performed. Genotyping and gene expression of neuropeptide Y (NPY) were performed using real time PCR. ResultsThere was a significant increase in number and percentage of CC genotype and C allele of NPY SNP distribution in CKD patients with and without hypertension when compared to controls. A significant association was found between CC genotype and C allele and the risk of CKD with hypertension with odd ratio 3.26 and 1.77, respectively. There is a significant positive correlation between NPY gene expression level and CIMT among chronic kidney disease patients with highest level of TC, LDLc and CIMT among CC genotype of NPY gene. ConclusionA significant association was found between CC genotype and C allele of NPY at rs16147 with increase NPY gene expression and risk of developing hypertension in CKD.

Effects of Kisspeptin-10 on Hypothalamic Neuropeptides and Neurotransmitters Involved in Appetite Control.

Besides its role as key regulator in gonadotropin releasing hormone secretion, reproductive function, and puberty onset, kisspeptin has been proposed to act as a bridge between energy homeostasis and reproduction. In the present study, to characterize the role of hypothalamic kisspeptin as metabolic regulator, we evaluated the effects of kisspeptin-10 on neuropeptide Y (NPY) and brain-derived neurotrophic factor (BDNF) gene expression and the extracellular dopamine (DA), norepinephrine (NE), serotonin (5-hydroxytriptamine, 5-HT), dihydroxyphenylacetic acid (DOPAC), and 5-hydroxyindoleacetic acid (5-HIIA) concentrations in rat hypothalamic (Hypo-E22) cells. Our study showed that kisspeptin-10 in the concentration range 1 nM–10 μM was well tolerated by the Hypo-E22 cell line. Moreover, kisspeptin-10 (100 nM–10 μM) concentration independently increased the gene expression of NPY while BDNF was inhibited only at the concentration of 10 μM. Finally, kisspeptin-10 decreased 5-HT and DA, leaving unaffected NE levels. The inhibitory effect on DA and 5-HT is consistent with the increased peptide-induced DOPAC/DA and 5-HIIA/5-HT ratios. In conclusion, our current findings suggesting the increased NPY together with decreased BDNF and 5-HT activity following kisspeptin-10 would be consistent with a possible orexigenic effect induced by the peptide.

Short communication: Gender and heat stress effects on hypothalamic gene expression and feed intake in broilers

Our study aims to evaluate gender and heat stress effects on animal performance and on the expression of five hypothalamic genes related to feed consumption: neuropeptide Y (NPY), ghrelin (GHRL), pro-opiomelanocortin (POMC), AMP-activated protein kinase (AMPKα-1), and liver kinase B1 (LKB1). To assay these effects, 42-day-old male and female broilers were maintained in thermal comfort or were subjected to heat stress (HS, 38°C for 24 hours). All animals were fed with diets formulated to meet their nutritional requirements. Broilers subjected to HS showed lower weight gain (p=0.0065) and tended to have lower feed intake (p=0.0687) than broilers kept in comfortable conditions. We observed gender and heat stress interaction effects on NPY (p=0.0225), AMPKα-1 (p=0.0398), and POMC expression (p=0.0072). The highest NPY gene expression was observed in male broilers from the thermal comfort group. Male broilers exposed to HS showed the highest AMPKα-1 gene expression levels. Comparing POMC expression between males and females at the comfortable temperature, we observed that females showed higher POMC expression levels than male broilers. A gender effect was also observed on LKB1 and AMPKα-1 gene expression (p=0.0256 and p=0.0001, respectively); increased expression was observed in male broilers. Our results indicate that the expression of some hypothalamic genes related to food consumption may contribute to the observed differences in voluntary feed intake between animals of different gender exposed to different environmental conditions.

Strategies of behavior, energetic and thermogenesis of striped hamsters in response to food deprivation.

The life history of many animals includes periods of food shortage. Two behavioral strategies are involved in small mammals in response to food shortage: an increase in activity behavior, representing the increased foraging or migratory behavior, and a decrease in activity level, serving as a mechanism for conserving energy. However, it is uncertain whether animals adopt both strategies in response to food shortage, and whether hormone and neuroendocrine mechanisms are involved in both strategies. In the present study, changes in behavior and metabolic rate were examined in food-deprived striped hamsters (Cricetulus barabensis). The effects of leptin supplement on activity behavior, metabolic rate and hypothalamic neuropeptide gene expression were also examined. The behavior of food-deprived hamsters significantly changed with photoperiod phases: with increasing activity during the dark phase compared to those fed ad libitum, whereas decreasing activity and simultaneously increasing resting behavior during the light phase. Resting metabolic rate, body mass, and masses of fat depots and digestive tracts significantly decreased in food-deprived hamsters compared with ad libitum controls. Leptin supplement tended to attenuate the increased activity in the dark phase. Gene expression of hypothalamic neuropeptide Y (NPY) was significantly upregulated in food-deprived hamsters, while was significantly attenuated by exogenous leptin. These findings suggest that both behavior strategies are important behavioral adjustments in free-living animals to cope with food shortage. Leptin and hypothalamic NPY gene expression may be involved in the adjustments of physiology and behavior in animals demonstrating a hyperactivity strategy in response to food shortage.

Hyperleptinemia in Neonatally Overfed Female Rats Does Not Dysregulate Feeding Circuitry.

Neonatal overfeeding during the first weeks of life in male rats is associated with a disruption in the peripheral and central leptin systems. Neonatally overfed male rats have increased circulating leptin in the first 2 weeks of life, which corresponds to an increase in body weight compared to normally fed counterparts. These effects are associated with a short-term disruption in the connectivity of neuropeptide Y (NPY), agouti-related peptide (AgRP), and pro-opiomelanocortin (POMC) neurons within the regions of the hypothalamus responsible for control of energy balance and food intake. Female rats that are overfed during the first weeks of their life experience similar changes in circulating leptin levels as well as in their body weight. However, it has not yet been studied whether these metabolic changes are associated with the same central effects as observed in males. Here, we hypothesized that hyperleptinemia associated with neonatal overfeeding would lead to changes in central feeding circuitry in females as it does in males. We assessed hypothalamic NPY, AgRP, and POMC gene expression and immunoreactivity at 7, 12, or 14 days of age, as well as neuronal activation in response to exogenous leptin in neonatally overfed and control female rats. Neonatally overfed female rats were hyperleptinemic and were heavier than controls. However, these metabolic changes were not mirrored centrally by changes in hypothalamic NPY, AGRP, and POMC fiber density. These findings are suggestive of sex differences in the effects of neonatal overfeeding and of differences in the ability of the female and male central systems to respond to changes in the early life nutritional environment.

Abstract 477: Nuclear Receptor Nur77 Modulates Cardiac Function via Neuropeptide Y and Neuropeptide Y Receptor 1 Signalling

Cardiac function is modulated by the sympathetic nervous system via catecholamines released from cardiac sympathetic nerve endings and adrenal chromaffin cells. The co-transmitter Neuropeptide Y (NPY) is co-released with catecholamines and also modulates cardiac function. NPY has previously been reported as an independent predictor of mortality in heart failure patients. In experimental models, NPY induces cardiac hypertrophy and elevates cardiomyocyte intracellular Ca 2+ concentrations and Ca 2+ -dependent signalling. However, not much is known yet about the upstream regulation of NPY in the context of cardiac function modulation. Interestingly, a robust upregulation of NPY gene expression has been reported in macrophages lacking the nuclear receptor Nur77. We have shown that Nur77 deficiency in mice leads to alterations in cardiomyocyte Ca 2+ homeostasis and adverse cardiac remodelling in response to isoproterenol. As these effects may be explained by enhanced NPY action in Nur77-deficient (Nur77-KO) mice, we hypothesized that Nur77 is a regulator of NPY. We now show that Nur77 down-regulates NPY expression in and secretion from sympathetic cells in vitro . Furthermore, NPY levels are significantly higher in blood plasma and hearts from Nur77-KO compared to wildtype mice. Antagonism of NPY receptor 1 (NPY1R) attenuates cardiac hypertrophy and elevated cardiomyocyte Ca 2+ responses caused by Nur77 deficiency. Interestingly, cardiomyocyte-specific Nur77-deficient mice also exhibit elevated cardiomyocyte Ca 2+ levels, which is partially reduced by NPY1R antagonism, even though plasma and cardiac NPY levels are similar to control mice. In conclusion, the transcription factor Nur77 regulates cardiac function both via systemic mechanisms and in cardiomyocytes intrinsically, via NPY/NPY1R modulation. These results imply a novel role for Nur77 as a potential modifier gene in heart failure.

Traumatic stress-induced persistent changes in DNA methylation regulate neuropeptide Y expression in rat jejunum.

Stress-induced chronic neuropsychiatric conditions such as anxiety are often co-morbid with gastrointestinal malfunctions. While we find enduring anxiety-like symptoms following minimal traumatic brain injury (MTBI) in rats, gastrointestinal consequences of MTBI remain elusive. In this study, we examined the effects of MTBI on a major gut peptide, neuropeptide Y (NPY) and gut motility. DNA methylation was studied as a possible epigenetic mechanism operative in the regulation of NPY expression in the gut. Minimal traumatic brain injury reduced the gut motility 48hours and 30days after trauma. The expression of DNA methyltransferase isoforms (DNMT1, DNMT3a, and DNMT3b) was altered in the jejunum 48hours and 30days after MTBI. However, the mRNA levels of growth arrest and DNA damage 45 (GADD45) isoforms, GADD45a, and GADD45b, which are believed to be involved in active DNA demethylation, initially decreased at 48hours but subsequently increased after 30days of trauma. Similarly, DNA hypomethylation at the NPY promoter region in the jejunum was correlated with the increase in NPY mRNA and protein levels 30days post-trauma. On the other hand, DNA hypomethylation at 48hours was associated with a decline in NPY expression. Treatment with 5-azacytidine (5-AzaC), a DNMT inhibitor, retarded DNA methylation and restored the NPY mRNA levels in the jejunum of MTBI-induced rats. These results suggest that DNA demethylation could be operative as an epigenetic mechanism in the long-term regulation of NPY gene expression to alter the gut motility during traumatic stress.

A potential role for the secretogranin II-derived peptide EM66 in the hypothalamic regulation of feeding behaviour.

EM66 is a conserved 66-amino acid peptide derived from secretogranin II (SgII), a member of the granin protein family. EM66 is widely distributed in secretory granules of endocrine and neuroendocrine cells, as well as in hypothalamic neurones. Although EM66 is abundant in the hypothalamus, its physiological function remains to be determined. The present study aimed to investigate a possible involvement of EM66 in the hypothalamic regulation of feeding behaviour. We show that i.c.v. administration of EM66 induces a drastic dose-dependent inhibition of food intake in mice deprived of food for 18hours, which is associated with an increase of hypothalamic pro-opiomelanocortin (POMC) and melanocortin-3 receptor mRNA levels and c-Fos immunoreactivity in the POMC neurones of the arcuate nucleus. By contrast, i.c.v. injection of EM66 does not alter the hypothalamic expression of neuropeptide Y (NPY), or that of its Y1 and Y5 receptors. A 3-month high-fat diet (HFD) leads to an important decrease of POMC and SgII mRNA levels in the hypothalamus, whereas NPY gene expression is not affected. Finally, we show that a 48hours of fasting in HFD mice decreases the expression of POMC and SgII mRNA, which is not observed in mice fed a standard chow. Taken together, the present findings support the view that EM66 is a novel anorexigenic neuropeptide regulating hypothalamic feeding behaviour, at least in part, by activating the POMC neurones of the arcuate nucleus.

A Role of Central NELL2 in the Regulation of Feeding Behavior in Rats.

A brain-enriched secreting signal peptide, NELL2, has been suggested to play multiple roles in the development, survival, and activity of neurons in mammal. We investigated here a possible involvement of central NELL2 in regulating feeding behavior and metabolism. In situ hybridization and an im-munohistochemical approach were used to determine expression of NELL2 as well as its colocalization with proopiomelanocortin (POMC) and neuropeptide Y (NPY) in the rat hypothalamus. To investigate the effect of NELL2 on feeding behavior, 2 nmole of antisense NELL2 oligodeoxynucleotide was administered into the lateral ventricle of adult male rat brains for 6 consecutive days, and changes in daily body weight, food, and water intake were monitored. Metabolic state-dependent NELL2 expression in the hypothalamus was tested in vivo using a fasting model. NELL2 was noticeably expressed in the hypothalamic nuclei controlling feeding behavior. Furthermore, all arcuatic POMC and NPY positive neurons produced NELL2. The NELL2 gene expression in the hypothalamus was up-regulated by fasting. However, NELL2 did not affect POMC and NPY gene expression in the hypothalamus. A blockade of NELL2 production in the hypothalamus led to a reduction in daily food intake, followed by a loss in body weight without a change in daily water intake in normal diet condition. NELL2 did not affect short-term hunger dependent appetite behavior. Our data suggests that hypothalamic NELL2 is associated with appetite behavior, and thus central NELL2 could be a new therapeutic target for obesity.

Dorsomedial hypothalamic NPY affects cholecystokinin-induced satiety via modulation of brain stem catecholamine neuronal signaling.

Increased neuropeptide Y (NPY) gene expression in the dorsomedial hypothalamus (DMH) has been shown to cause hyperphagia, but the pathway underlying this effect remains less clear. Hypothalamic neural systems play a key role in the control of food intake, in part, by modulating the effects of meal-related signals, such as cholecystokinin (CCK). An increase in DMH NPY gene expression decreases CCK-induced satiety. Since activation of catecholaminergic neurons within the nucleus of solitary tract (NTS) contributes to the feeding effects of CCK, we hypothesized that DMH NPY modulates NTS neural catecholaminergic signaling to affect food intake. We used an adeno-associated virus system to manipulate DMH NPY gene expression in rats to examine this pathway. Viral-mediated hrGFP anterograde tracing revealed that DMH NPY neurons project to the NTS; the projections were in close proximity to catecholaminergic neurons, and some contained NPY. Viral-mediated DMH NPY overexpression resulted in an increase in NPY content in the NTS, a decrease in NTS tyrosine hydroxylase (TH) expression, and reduced exogenous CCK-induced satiety. Knockdown of DMH NPY produced the opposite effects. Direct NPY administration into the fourth ventricle of intact rats limited CCK-induced satiety and overall TH phosphorylation. Taken together, these results demonstrate that DMH NPY descending signals affect CCK-induced satiety, at least in part, via modulation of NTS catecholaminergic neuronal signaling.

Neuropeptide Y input to the rat basolateral amygdala complex and modulation by conditioned fear.

Within the basolateral amygdaloid complex (BLA), neuropeptide Y (NPY) buffers against protracted anxiety and fear. Although the importance of NPY's actions in the BLA is well documented, little is known about the source(s) of NPY fibers to this region. The current studies identified sources of NPY projections to the BLA by using a combination of anatomical and neurochemical approaches. NPY innervation of the BLA was assessed in rats by examining the degree of NPY coexpression within interneurons or catecholaminergic fibers with somatostatin and tyrosine hydroxylase (TH) or dopamine β-hydroxylase (DβH), respectively. Numerous NPY(+) /somatostatin(+) and NPY(+) /somatostatin(-) fibers were observed, suggesting at least two populations of NPY fibers within the BLA. No colocalization was noted between NPY and TH or DβH immunoreactivities. Additionally, Fluorogold (FG) retrograde tracing with immunohistochemistry was used to identify the precise origin of NPY projections to the BLA. FG(+) /NPY(+) cells were identified within the amygdalostriatal transition area (AStr) and stria terminalis and scattered throughout the bed nucleus of the stria terminalis. The subpopulation of NPY neurons in the AStr also coexpressed somatostatin. Subjecting animals to a conditioned fear paradigm increased NPY gene expression within the AStr, whereas no changes were observed within the BLA or stria terminalis. Overall, these studies identified limbic regions associated with stress circuits providing NPY input to the BLA and demonstrated that a unique NPY projection from the AStr may participate in the regulation of conditioned fear. J. Comp. Neurol. 524:2418-2439, 2016. © 2016 Wiley Periodicals, Inc.

Effect of electroconvulsive stimulation on messenger RNA expression in the prefrontal cortex in a rat pain model.

Previous reports have shown that electroconvulsive therapy (ECT) is efficacious in the treatment of neuropathic pain; however, its mechanism of action remains unclear. The present study aimed to understand these mechanisms by investigating the alterations in the expression of neuropeptide Y (NPY) and interleukin-1β (IL-1β) in the prefrontal cortex. A rat model of neuropathic pain produced by chronic constrictive injury of the sciatic nerve was used, and mechanical and thermal hyperalgesia were evaluated starting 2 days after the injury. Using a pulse generator, ECT was administered to the rodents for 6 days from days 7-12 after the injury. Thermal and mechanical stimulation were administered to assess pain thresholds. Quantitative polymerase chain reaction, used to measure gene expression levels in the prefrontal cortex, showed that NPY and IL-1β gene expression levels in the prefrontal cortex increased following the injury. The present results indicate that these gene expression level variations may be associated with the mechanisms underlying the effect of ECT in treating neuropathic pain.