Leptin transiently antagonizes ghrelin and long-lastingly orexin in regulation of Ca^(2+) signaling in neuropeptide Y neurons of the arcuate nucleus

AIM： To explore the mechanism for interactions of leptin with ghrelin and orexin in the arcuate nucleus （ARC） activating neuropeptide Y （NPY） neurons during physiological regulation of feeding, METHODS： Single neurons from ARC of adult rats with matured feeding function were isolated. [Ca2＋]i was measured to monitore their activities. The time course of leptin effects on ghrelin-induced versus orexin-induced [Ca2＋]i increases in NPY neurons was studied. RESULTS： Administration of ghrelin or orexin-A at 101~ mol/L increased cytosolic Ca2~ concentration （[Ca2＋]~） in NPY neurons isolated from the ARC of adult rats. Upon administration of leptin at 10^-14-10^-12 mol/L, ghrelin-induced [Ca2＋]i increases were initially （〈 10 min） inhibited but later restored, exhibiting a transient pattern of inhibition. In contrast, orexin-induced [Ca2＋]i increases were inhibited by leptin in a long- lasting manner. Furthermore, a prior administration of leptin inhibited orexin action but not ghrelin action to increase ICa 2＋li, CONCLUSION： Leptin counteracted ghrelin effects transiently and orexin effects long-lastingly in NPY neurons. The transient property with which leptin counteracts ghrelin action in NPY neurons may allow the fasting-associated increase in ghrelin levels to activate NPY neurons in the presence of physiological leptin and to stimulate feeding.

Role of Nociceptive Arcuate Nucleus Neurons in Chloroquine-induced Pruritic Behaviors in Mice

Despite its clinical importance, the underlying central mechanisms of pruritic behaviors are poorly understood. To investigate the role of nociceptive arcuate nucleus neurons in chloro-quine-induced pruritic behaviors in mice, we tested the effect of arcuate nucleus neurons and interscapular brown adipose tissue (IBAT) on itch produced by intradermal injection of chloroquine in the nape of the neck. Our results provide several lines of evidence for an important role of nociceptive arcuate nucleus neurons in chloroquine-induced pruritic behavior: (1) Intradermal microinjection of chloro-quine resulted in a dramatic increase in itch behaviors accompanied by the activation of c-Fos positive neurons in arcuate nucleus; (2) Microinjection of chloroquine significantly increased IBAT temperature in the mice. These findings suggested that chloroquine-induced pruritic behaviors were associated with the activity of nociceptive arcuate nucleus neurons.

Changes in beta-endorphin neuron numbers and serum hormone levels in the arcuate nucleus of ovariectomized rats undergoing treadmill exercise

BACKGROUND： The arcuate nucleus, when damaged in young rats, can lead to pathological changes in adults, such as osteoporosis. Ovariectomized rats suffer from osteoporosis at eight weeks following surgery and the number of β -endorphin immunoreactive neurons in the arcuate nucleus of the hypothalamus is significantly decreased. OBJECTIVE： To establish a rat model of osteoporosis using ovariectomy and to explore changes in the number of β-endorphin neurons and to correlate any such change with serum hormone levels in response to exercise or rest. DESIGN, TIME AND SETTING： The completely randomized block design, neural morphology study was performed at the Key Laboratory of Physiology, Guangdong Medical College, China between March 2004 and January 2005. MATERIALS： Sixteen healthy female rats were selected for ovariectomy. METHODS： Following model establishment, rats were assigned to either rest or exercise groups and each rat was housed individually. Rats in the exercise group underwent an exercise regimen using a treadmill. MAIN OUTCOME MEASURES： Eight weeks following exercise, radioimmunoassay was performed to detect serum growth hormone, estrogen and osteocalcin levels. Immunohistochemistry was used to measure changes in the number of β -endorphin neurons in the arcuate nucleus of the hypothalamus. Changes in bone metabolism were assessed using bone histomorphometry. RESULTS： In the exercise group, the β -endorphin immunoreactive neurons were high in number, darkly stained, and the nucleus was not obvious. In the rest group, the β -endorphin immunoreactive neurons were low in number and lightly stained. The number of β-endorphin immunoreactive neurons in the exercise group was higher compared with the rest group （t = 2.83, P 〈 0.05）. Estrogen levels were similar between the rest and exercise groups （P 〉 0.05）. Serum osteocalcin and growth hormone levels were significantly higher in the exercise group compared with the rest group （t = 2.78, 2.32, P 〈 0.05）. Compared with the rest group, the percentage of trabecular bone area, trabecular thickness, and trabecular number were significantly increased, but trabecular separation was significantly reduced （t=2.48, 2.57, 2.32, 3.06, P 〈 0.05） in the exercise group. In the exercise group, the trabeculae of the tibia were arranged regularly and were high in number. In the rest group, the trabeculae of the tibia were organized in a disorderly manner and were low in number, with many fat particles. CONCLUSION： Exercise promotes bone growth and delays osteoporosis by inducing an increase in the number of β-endorphin immunoreactive neurons in the arcuate nucleus of the hypothalamus and by increasing serum growth hormone and osteocalcin levels.

Expression of Kisspeptin/kiss1r System is Down-regulated in the Hypothalamic Arcuate Nucleus of Pubertal Male Rats with High-fat-diet

Objective To explore the potential function of nutrition on kisspeptin/kisspeptin receptor （kisslr） system in the arcuate nucleus （ARC） of male rats. Methods Pregnant rats were obtained and male pups were used to establish obesity model. Body parameters and blood samples were evaluated. Immunohistochemistry was used to determine the localizations and protein levels of kisspeptin, kisslr, and leptin receptor （LepR） immunoreactivity （IR） in ARC. QRT-PCR was used to determine kissl-, kisslr-, LepR-, and GnRH-mRNA levels. Results The establishment of obesity model was successful as body parameters and hormones levels changed noticeably. Kisspeptin-, kisslr-, and LepR-IR were detected and protein levels decreased significantly in high-fat-diets （HFD） rats than controls. The mRNA levels of kissl, LepR and GnRH significantly decreased in the ARC of HFD-fed rats. No difference was observed in kisslr mRNA levels between the two groups. Conclusion These data suggest that failure to increase GnRH levels with HFD may be associated with pubertal down-regulation of LepR and kisspeptin/kisslr system in the ARC of male rats.

Neuropeptide Y and leptin receptor expression in the hypothalamus of rats with chronic immobilization stress

In this study, Sprague-Dawley rats were immobilized to a frame for 3 hours a day for 21 days to establish a model of chronic immobilization stress. The body weight and food intake of rats subjected to chronic immobilization stress were significantly decreased compared with the control group. Dual-labeling immunofluorescence revealed that the expression of leptin receptor and the co-localization coeffient in these leptic receptor neurons in the arcuate nucleus of the hypothalamus were both upregulated, while the number of neuropeptide Y neurons was decreased. Chronic immobilization stress induced high expression of leptin receptor in the arcuate nucleus and suppressed the synthesis and secretion of neuropeptide Y, thereby disrupting the pathways in the arcuate nucleus that regulate feeding behavior, resulting in diminished food intake and reduced body weight.

促生长素抑制素样免疫反应在大白鼠弓状核和正中隆起的超微结构分布

本文用免疫电镜方法证明:促生长素抑制素样免疫反应神经末梢分布于弓状棱并与未标记的树突形成轴树突触。在正中隆起的纤维层和栅状层内均可见上述免疫反应末梢,大多数紧贴门脉毛细血管基底膜周围甚至穿入基底膜内。免疫反应末梢尚可与未标记的末梢形成轴轴突触样结构。

Kisspeptin regulates gonadotropin-releasing hormone secretion in gonadotropin-releasing hormone/enhanced green fluorescent protein transgenic rats

Kisspeptin is essential for activation of the hypothalamo-pituitary-gonadal axis. In this study, we established gonadotropin-releasing hormone/enhanced green fluorescent protein transgenic rats. Rats were injected with 1, 10, or 100 pM kisspeptin-10, a peptide derived from full-length kisspeptin, into the arcuate nucleus and medial preoptic area, and with the kJsspeptJn antagonist peptJde 234 into the lateral cerebral ventricle. The results of immunohistochemical staining revealed that pulsatile luteinizing hormone secretion was suppressed after injection of antagonist peptide 234 into the lateral cerebral ventricle, and a significant increase in luteinizing hormone level was observed after kisspeptin-10 injection into the arcuate nucleus and medial preoptic area. The results of an enzyme-linked immunosorbent assay showed that luteinizing hormone levels during the first hour of kisspeptin-10 infusion into the arcuate nucleus were significantly greater in the 100 pM kisspeptin-10 group than in the 10 pM kisspeptin-10 group. These findings indicate that kisspeptin directly promotes gonadotropin-releasing hormone secretion and luteinizing hormone release in gonadotropin-releasing hormone/enhanced green fluorescent protein transgenic rats. The arcuate nucleus is a key component of the kisspeptin-G protein-coupled receptor 54 signaling pathway underlying regulating luteinizing hormone pulse secretion.

Feeding behavior and gene expression of appetite-related neuropeptides in mice lacking for neuropeptide Y Y5 receptor subclass

Neuropeptide Y (NPY) is a potent neurotransmitter for feeding. Besides NPY, orexigenic neuropeptides such as agouti-related protein (AgRP), and anorexi- genic neuropeptides such as α-melatonin stimulating hormone (MSH) and cocaine-amphetamine-regulated transcript (CART) are also involved in central feeding regulation. During fasting, NPY and AgRP gene expressions are up-regulated and POMC and CART gene ex- pressions are down-regulated in hypothalamus. Based on the network of peptidergic neurons, the former are involved in positive feeding regulation, and the latter are involved in negative feeding, which exert these feeding-regulated peptides especially in paraventricular nucleus (PVN). To clarify the compensatory mecha- nism of knock-out of NPY system on feeding, change in gene expressions of appetite-related neuropeptides and the feeding behavior was studied in NPY Y5-KO mice. Food intake was increased in Y5-KO mice. Fasting increased the amounts of food and water intake in the KO mice more profoundly. These data indicated the compensatory phenomenon of feeding behavior in Y5-KO mice. RT-PCR and ISH suggested that the compensation of feeding is due to change in gene expressions of AgRP, CART and POMC in hypothalamus. Thus, these fi ndings indicated that the compensatory mechanism involves change in POMC/CART gene expression in arcuate nucleus (ARC). The POMC/CART gene expression is important for central compensatory regulation in feeding behavior.

A dahlia flower extract has antidiabetic properties by improving insulin function in the brain

Butein,a rare chalcone found in the toxic plant Toxicodendron vernicifluum,has been shown to regulate glucose homeostasis via inhibition of the nuclear factor kappa-B kinase subunit beta(IKKβ)/nuclear factor kappa B(NF-κB)pathway in the brain.Here,we investigated whether the nonpoisonous plant Dahlia pinnata could be a source of butein as a potential treatment for type 2 diabetes(T2D).In mice fed a high-fat diet(HFD)to induce glucose intolerance,an oral D.pinnata petal extract improved glucose tolerance at doses of 3.3 mg/kg body weight and 10 mg/kg body weight.Surprisingly,this effect was not mediated by butein alone but by butein combined with the closely related flavonoids,sulfuretin and/or isoliquiritigenin.Mechanistically,the extract improved systemic insulin tolerance.Inhibition of phosphatidylinositol 3-kinase to block insulin signaling in the brain abrogated the glucoregulatory effect of the orally administered extract.The extract reinstated central insulin signaling and normalized astrogliosis in the hypothalamus of HFD-fed mice.Using NF-κB reporter zebrafish to determine IKKβ/NF-κB activity,a potent anti-inflammatory action of the extract was found.A randomized controlled crossover clinical trial on participants with predia-betes or T2D confirmed the safety and efficacy of the extract in humans.In conclusion,we identified an extract from the flower petals of D.pinnata as a novel treatment option for T2D,potentially targeting the central regulation of glucose homeostasis as a root cause of the disease.

Loss of the centrosomal protein Cenpj leads to dysfunction of the hypothalamus and obesity in mice

Cenpj is a centrosomal protein located at the centrosomes and the base of cilia,it plays essential roles in regulating neurogenesis and cerebral cortex development.Although centrosomal and cilium dysfunction are one of the causes of obesity,insulin resistance,and type 2 diabetes,the role that Cenpj plays in the regulation of body weight remains unclear.Here,we deleted Cenpj by crossing Cenpjflox/flox mice with Nkx2.1-Cre mice.Loss of the centrosomal protein Cenpj in Nkx2.1-expressing cells causes morbid obesity in mice at approximately 4 months of age with expended brain ventricles but no change of brain size.We found that hypothalamic cells exhibited reduced proliferation and increased apoptosis upon Cenpj depletion at the embryonic stages,resulting in a dramatic decrease in the number of Proopiomelanocortin(POMC)neurons and electrophysiological dysfunction of NPY neurons in the arcuate nucleus(ARC)in adults.Furthermore,depletion of Cenpj also reduced the neuronal projection from the ARC to the paraventricular nucleus(PVN),with decreased melanocortin-4 receptors(MC4R)expression in PVN neurons.The study defines the roles that Cenpj plays in regulating hypothalamus development and body weight,providing a foundation for further understanding of the pathological mechanisms of related diseases.

nr0b1(DAX1) loss of function in zebrafish causes hypothalamic defects via abnormal progenitor proliferation and differentiation

The nuclear receptor DAX-1, encoded by the NR0B1 gene, is presented in the hypothalamic tissues in humans and other vertebrates. Human patients with NR0B1 mutations often have hypothalamic-pituitary defects, but the involvement of NR0B1 in hypothalamic development and function is not well understood. Here, we report the disruption of the nr0b1 gene in zebrafish causes abnormal expression of gonadotropins, a reduction in fertilization rate, and an increase in postfasting food intake, which are indicators of abnormal hypothalamic functions. We find that loss of nr0b1 increases the number of prodynorphin(pdyn)-expressing neurons but decreases the number of pro-opiomelanocortin(pomcb)-expressing neurons in the zebrafish hypothalamic arcuate region(ARC). Further examination reveals that the proliferation of progenitor cells is reduced in the hypothalamus of nr0b1 mutant embryos accompanying the decreased expression of genes in the Notch signaling pathway. Additionally, the inhibition of Notch signaling in wildtype embryos increases the number of pdyn neurons, mimicking the nr0b1 mutant phenotype. In contrast,ectopic activation of Notch signaling in nr0b1 mutant embryos decreases the number of pdyn neurons.Taken together, our results suggest that nr0b1 regulates neural progenitor proliferation and maintenance to ensure normal hypothalamic neuron development.