Central Regulation Of Energy Homeostasis Research Articles

Neuropeptide W in the rat pancreas: Potentiation of glucose-induced insulin release and Ca 2+ influx through L-type Ca 2+ channels in β-cells and localization in islets

Neuropeptide W (NPW) is a regulatory peptide that acts via two subtypes of G protein-coupled receptors, GPR7 and GPR8. Evidence has been provided that NPW is involved in the central regulation of energy homeostasis and feeding behavior. In this study, we examined the effects of NPW on insulin release and localization of NPW in the rat pancreas. NPW (10–100 nM) significantly increased insulin release in the presence of 8.3 mM, but not 2.8 mM, glucose in the isolated rat islets. By fura-2 microfluorometry, NPW (1–100 nM) concentration-dependently increased cytosolic Ca 2+ concentration ([Ca 2+] i) at 8.3 mM glucose in rat single β-cells. The NPW-induced [Ca 2+] i increase was abolished under external Ca 2+-free conditions and by an L-type Ca 2+ channel blocker nifedipine (10 μM). RT-PCR analysis revealed that mRNA for NPW was expressed in the rat pancreas and hypothalamus. Double immunohistochemical analysis showed that NPW-immunoreactivity was found in islets and co-localized with insulin-containing β-cells, but not glucagon-containing α-cells and somatostatin-containing δ-cells. These results suggest that NPW could serve as a local modulator of glucose-induced insulin release in rat islets. NPW directly activates β-cells to enhance Ca 2+ influx through voltage-dependent L-type Ca 2+ channels and potentiates glucose-induced insulin release.

Structure of the Bateman2 Domain of Yeast Snf4: Dimeric Association and Relevance for AMP Binding

AMP-activated protein kinase (AMPK) is a central regulator of energy homeostasis in mammals. AMP is believed to control the activity of AMPK by binding to the gamma subunit of this heterotrimeric enzyme. This subunit contains two Bateman domains, each of which is composed of a tandem pair of cystathionine beta-synthase (CBS) motifs. No structural information is currently available on this subunit, and the molecular basis for its interactions with AMP is not well understood. We report here the crystal structure at 1.9 Angstrom resolution of the Bateman2 domain of Snf4, the gamma subunit of the yeast ortholog of AMPK. The structure revealed a dimer of the Bateman2 domain, and this dimerization is supported by our light-scattering, mutagenesis, and biochemical studies. There is a prominent pocket at the center of this dimer, and most of the disease-causing mutations are located in or near this pocket.

Central Regulation of Energy Homeostasis

Insulin has two important functions that relate to overall metabolic homeostasis. The phylogenetically oldest is the maintenance of sufficient energy stores to allow for development, growth, and reproduction. The newer is as a feedback regulator of plasma glucose. The key role of the central nervous system in both functions is reviewed from a personal perspective, and the development of the concept that both body weight (adiposity) and plasma glucose are critically regulated by the same hormone is described. The recent suggestion that diabetes and obesity are linked by their common reliance on this central nervous system insulin signaling system is reviewed. Recent efforts to understand the hypothalamic mechanisms involved are described, and the common use of insulin receptor substrate 2 and the phosphatidylinositol 3-kinase signaling mechanism is emphasized. Potential consequences of defects in the secretion of insulin or the action of insulin in the central nervous system are given, and linkage between obesity and diabetes is illustrated with a potential clinical representative.

Somatostatin, a negative‐regulator of central leptin action in the rat hypothalamus

Leptin-responsive neurons of the hypothalamus constitute a heterogeneous population expressing a vast array of different neuropeptides and neurotransmitters, some of which participate in the regulation of hunger and satiety. Here we report that somatostatin modulates the efficacy of leptin-signalling in the rat hypothalamus. Using a two-pulse paradigm at 30-min intervals, we delivered somatostatin or somatostatin receptor subtype-selective agonists in combination with leptin into the lateral cerebral ventricle of stereotaxically cannulated rats. To monitor the effect of somatostatin on the leptin-signalling pathway, we quantified changes in the leptin-mediated activation of STAT3, the signal transducer and activator of transcription 3. Successive administration of somatostatin and leptin diminished the level of STAT3-phosphorylation and nuclear STAT3 translocation in the ventromedial and dorsomedial hypothalamic nuclei, the lateral hypothalamic area, and the arcuate nucleus by about 40% compared to leptin administration alone. Furthermore, application of subtype-selective somatostatin receptor agonists suggests that the observed reduction in leptin-responsiveness is predominantly mediated by the sst3 receptor-subtype, followed by sst1 and sst2. In addition, the intensity of the negative-regulatory effect of somatostatin on leptin-signalling displayed regional differences for the three receptor-subtypes involved. Addressing the functional consequences of the diminished leptin-signalling, behavioural analyses showed that centrally applied somatostatin counteracts the leptin-mediated suppression of food intake. These results suggest that the pleiotropic effector somatostatin also plays a role in the central regulation of energy homeostasis.

Genetic Dissection of Neuronal Pathways Controlling Energy Homeostasis

Recent research has identified a number of genes playing critical roles in the central regulation of energy homeostasis. Subsequently, models of the neurocircuitry regulating energy balance have been suggested, although their physiological relevance remains mostly untested. Using the Cre/loxP system, we can now genetically dissect these neurocircuits and establish the specific roles of these genes in small neuronal subpopulations. Here we focus on two receptors shown to be critical in the central regulation of energy homeostasis: leptin (LepR) and melanocortin-4 receptors (MC4R). Mice and humans deficient in either leptin or melanocortin signaling are severely obese. A prominent model of leptin action places the arcuate nucleus of the hypothalamus, and in particular arcuate proopiomelanocortin (POMC) neurons, at the center stage of energy balance regulation. By deleting LepR specifically from POMC neurons in mice, we showed that LepR on POMC neurons are required but not solely responsible for leptin's regulation of body weight homeostasis. Thus, LepR on other neurons must also be critically important in leptin-mediated regulation of body weight homeostasis. Data from MC4R-deficient mice have shown that MC4Rs regulate both sides of the energy intake/energy expenditure balance. Our recent experiments used MC4R-deficient mice with restored MC4R expression only in the paraventricular hypothalamus and a subpopulation of amygdala neurons. We showed that MC4Rs in the paraventricular hypothalamus and/or amygdala are sufficient to control food intake but that MC4Rs elsewhere control energy expenditure, thereby discovering the novel concept of functional and anatomical divergence of MC4Rs.

Sumoylation of CCAAT/Enhancer-binding Protein α and Its Functional Roles in Hepatocyte Differentiation

The sumoylation of CCAAT/enhancer-binding proteins (C/EBPs) by small ubiquitin-related modifier-1 (SUMO-1) has been reported recently. In this study, we investigated the functional role of the sumoylation of C/EBPalpha in the differentiation of hepatocytes. The amount of sumoylated C/EBPalpha gradually decreased during the differentiation, which suggests that the sumoylation is important for the control of growth/differentiation especially in the fetal liver. To analyze the function of the sumoylation of C/EBPalpha in liver-specific gene expression, we studied its effects on the expression of the albumin gene. The C/EBPalpha-mediated transactivation of the albumin gene was reduced by sumoylation of C/EBPalpha in primary fetal hepatocytes. The enhancement of C/EBPalpha-mediated transactivation by BRG1, a core subunit of the SWI/SNF chromatin remodeling complex, was hampered by sumoylation in a luciferase reporter assay. In addition, we discovered that sumoylation of C/EBPalpha blocked its inhibitory effect on cell proliferation by leading to the disruption of a proliferation-inhibitory complex because of a failure of the sumoylated C/EBPalpha to interact with BRG1. BRG1 was recruited to the dihydrofolate reductase promoter in nonproliferating C33a cells but was not detected in proliferating cells where C/EBPalpha, BRG1, and SUMO-1 were overexpressed. This result suggests that BRG1 down-regulates the expression of the dihydrofolate reductase gene. These findings provide the insight that SUMO acts as a space regulator, which affects protein-protein interactions.

Mutated-leptin gene transfer induces increases in body weight by electroporation and hydrodynamics-based gene delivery in mice

To investigate whether in vivo gene transfer causes leptin-antagonistic effects on food intake, animal body weight and fat tissue weight, the R128Q mutated-leptin gene, an R to Q substitution at position 128 of mouse leptin, was transferred into mouse liver and leg muscle by electroporation and hydrodynamics-based gene delivery. Mutated-leptin gene transfer by electroporation caused significant increases in body weight at 5 days and after (5.4% increase relative to control; p<0.05). Hydrodynamics-based gene delivery of the mutated-leptin gene also caused an increase in body weight (3.0% increase relative to control; p<0.05). Mutated-leptin gene transfer by electroporation significantly increased the tissue weight of epididymal white fat and neuropeptide Y mRNA expression in the hypothalamus compared with those of the control group 3 weeks after gene transfer (p<0.05). These results suggest that mutated-leptin gene transfer successfully produced leptin-antagonistic effects by modulating the central regulator of energy homeostasis. Also, the extent of leptin-antagonistic effects by electroporation was much higher than hydrodynamics-based gene delivery, with at least single gene transfer.

Polymorphisms in the leptin and leptin receptor genes in relation to resting metabolic rate and respiratory quotient in the Québec Family Study

Leptin (LEP) is an endocrine hormone that participates in many metabolic pathways, including those associated with the central regulation of energy homeostasis. We examined the associations between polymorphisms in the LEP and leptin receptor (LEPR) genes and resting metabolic rate (RMR) and respiratory quotient (RQ) in the Quebec Family Study. Three polymorphisms in LEPR (K109R, Q223R and K656N) and one in LEP (19A > G) were genotyped in 678 subjects. RMR, RQ at rest and RQ while sitting, standing and walking at 4.5 km/h (RQ45) were adjusted for age, sex, fat mass and fat-free mass. RQ45 was associated with the LEPR-K109R (P = 0.004) and Q223R (P = 0.03) polymorphisms, and RMR showed association with the LEPR-K656N polymorphism (P = 0.006). For the LEP-19A > G polymorphism, no significant associations were observed. However, LEP-A19A homozygotes who were carriers of the LEPR N656 allele had a significantly lower RQ45 compared to other genotype combinations (P for interaction=0.003). These findings suggest that DNA sequence variation in the LEPR gene contributes to human variation in RMR and in the relative rates of substrate oxidation during low-intensity exercise in steady state but not in a resting state.

Developmental changes in the long form leptin receptor and related neuropeptide gene expression in the pig brain.

The hypothalamus is the key site of central regulation of energy homeostasis, appetite, and reproduction. The long form leptin receptor (Ob-Rl) is localized within the hypothalamus along with several neuropeptides that are involved in regulation of the neuroendocrine axis. In the present study, developmental changes in gene expression of the Ob-Rl, preproorexin, proopiomelanocortin (POMC), corticotropin releasing factor (CRF), somatostatin, and GnRH in the hypothalamus was studied. Expression of Ob-Rl and neuropeptide mRNA was examined by semiquantitative reverse transcription-polymerase chain reaction in hypothalami collected from 106-day-old fetus (n = 3) and 7-day-old (n = 3), 3.5-mo-old (n = 3), and 6-mo-old (n = 2) gilts. In addition, leptin mRNA expression in the first three ages was examined in back fat. Leptin mRNA expression increased (P < 0.05) by 7 days postnatal, but Ob-Rl mRNA expression increased (P < 0.01) by 3.5 mo. Expression of preproorexin (P < 0.05), somatostatin, and GnRH (P < 0.01) mRNA peaked by 3.5 mo of age while POMC mRNA expression increased markedly (P < 0.01) by 6 mo of age. The CRF mRNA expression did not change across ages. These findings suggest a possible relationship among Ob-Rl and a number of hypothalamic and peripheral peptides in the development of the neuroendocrine axis. These peptides may serve as messengers that link mechanisms that regulate reproduction and energy balance.

Central melanocortins and the regulation of weight during acute and chronic disease.

Recent advances in our understanding of the regulation of body weight, appetite, and metabolic rate have highlighted the role of the adipose-derived hormone leptin and its receptor as fundamental modulators of these processes. Investigations of the neural targets for leptin action--as well as characterization of the agouti obesity syndrome--have, in turn, led to the discovery of fundamental neural pathways involved in the central regulation of energy homeostasis. In particular, the central melanocortin system has been shown to regulate appetite and metabolic rate in rodents; mutations in this system have been demonstrated to result in obesity in humans. Overall, the melanocortin system appears to function as a bidirectional rheostat in the regulation of energy intake and expenditure in rodents and potentially in humans. The first section of this chapter will focus on the development of our understanding of melanocortin physiology in the context of obesity. In particular, recent data regarding the interplay between melanocortin and neuropeptide Y (NPY) signaling at a cellular level will be discussed. The following section will discuss the hypothesis that melanocortin signaling plays a role in pathological weight loss and hypermetabolism observed in murine cachexia models. The potential role of this system in integrating a variety of anorexic and cachexic signals, as well as the potential for its pharmacological manipulation in the treatment of human cachexia, will be discussed.

The acquisition of obesity: insights from cellular and genetic research.

The acquisition of increased adipose tissue mass in man occurs during prolonged periods of positive energy balance. Normally, energy homeostasis in children and adults is regulated strictly and the energy stores are kept within the defined age-dependent physiological range. Susceptibility to definitive increases in the level of energy balance during times of reduced energy consumption or increased energy intake, leading to changes in body composition and/or changes in relative body weight, seems to be genetically determined. Although at present much information on the regulation of energy homeostasis and related unfavourable factors exists from animal studies, knowledge of the regulation of energy balance in human subjects is still insufficient. Some evidence on relevant factors involved in the regulation of energy balance in man has been obtained from epidemiological data, as well as from studies of patients with rare monogenetic forms of obesity. In the present article a special focus will be put on the regulation of body energy stores at the level of the adipose tissue, with emphasis on the regulation of human adipocyte differentiation. In addition to the currently intensive scientific interest in the central regulation of energy homeostasis in man, there is sufficient evidence to support the idea that the acquisition of an increased adipose tissue mass is also dependent on the susceptibility of pre-adipocytes to proliferate, to differentiate or to enter into apoptosis.

Effects of leptin on insulin secretion from isolated rat pancreatic islets.

Leptin is a hormone secreted by adipocytes as a peripheral metabolic signal for the central regulation of energy homeostasis or the reproductive system. Recent studies demonstrated that leptin receptor mRNA is expressed in pancreatic islets of rodents and that leptin at relatively high doses inhibits glucose-induced insulin secretion from rat islets. However, the physiological mechanism of leptin on insulin secretion has not been identified. In this study, we report that leptin inhibits glucose-induced insulin secretion at lower concentrations ranging from 25 to 50 ng/ml using a static incubation method. A perifusion study revealed that leptin (50 ng/ml) affected the second phase of insulin secretion but not the first phase. Leptin did not affect insulin secretion stimulated by glibenclamide (1 and 5 micromol/l) or forskolin (1 micromol/l). Leptin (50 ng/ml) significantly inhibited insulin secretion induced by the phorbol ester phorbol 12-myristate 13-acetate (TPA) in the presence of Ca2+ but not in the absence of Ca2+. Because TPA is known to activate protein kinase C (PKC), these present results suggest that leptin, at a physiological concentration, suppresses the second phase of insulin secretion by reducing activity of the Ca2+-dependent PKC isoform.

NPY mRNA and peptide immunoreactivity in the arcuate nucleus are increased by osmotic stimuli: correlation with dehydration anorexia.

The role of neuropeptide Y (NPY) in the central control of appetite and energy balance is now established, but its involvement in the control of drinking and fluid homeostasis is less well characterized. Central administration of NPY stimulates drinking in rats, an effect believed to be independent of its orexigenic effects. Recent studies have demonstrated increased preproneuropeptide Y (preproNPY) mRNA in the arcuate nucleus (ARC) of the rat following food deprivation (FD) or water deprivation (WD). Because WD also suppresses food intake, it was not clear whether the osmotic or the anorectic effects of this stimulus were responsible for increased ARC preproNPY mRNA. In an attempt to distinguish between these possibilities, the present study further examined the effects of hyperosmotic stimuli on preproNPY mRNA in the ARC. Salt loading (4 or 7 days) and WD (4 days) both increased the abundance of preproNPY mRNA in the ARC. These increases were proportional to the severity and duration of treatment and were related to the degree of anorexia and weight loss. In a separate study WD, FD, or combined food and water deprivation (4 days) all produced similar decreases in body weight, but WD produced a smaller increase in ARC preproNPY mRNA. All of these treatments resulted in the appearance of NPY-like immunoreactivity in ARC neuronal perikarya. Together these findings suggest that NPY neuron activity in the ARC may be regulated by decreases in food intake rather than changes in body weight per se or increased osmolarity and support other data implicating NPY in the central regulation of energy homeostasis.