Kayoko Ogimoto

Effects of hypothalamic neurodegeneration on energy balance

Normal aging in humans and rodents is accompanied by a progressive increase in adiposity. To investigate the role of hypothalamic neuronal circuits in this process, we used a Cre-lox strategy to create mice with specific and progressive degeneration of hypothalamic neurons that express agouti-related protein (Agrp) or proopiomelanocortin (Pomc), neuropeptides that promote positive or negative energy balance, respectively, through their opposing effects on melanocortin receptor signaling. In previous studies, Pomc mutant mice became obese, but Agrp mutant mice were surprisingly normal, suggesting potential compensation by neuronal circuits or genetic redundancy. Here we find that Pomc-ablation mice develop obesity similar to that described for Pomc knockout mice, but also exhibit defects in compensatory hyperphagia similar to what occurs during normal aging. Agrp-ablation female mice exhibit reduced adiposity with normal compensatory hyperphagia, while animals ablated for both Pomc and Agrp neurons exhibit an additive interaction phenotype. These findings provide new insight into the roles of hypothalamic neurons in energy balance regulation, and provide a model for understanding defects in human energy balance associated with neurodegeneration and aging.

Fibroblast Growth Factor 21 Action in the Brain Increases Energy Expenditure and Insulin Sensitivity in Obese Rats

OBJECTIVE The hormone fibroblast growth factor 21 (FGF21) exerts diverse, beneficial effects on energy balance and insulin sensitivity when administered systemically to rodents with diet-induced obesity (DIO). The current studies investigate whether central FGF21 treatment recapitulates these effects. RESEARCH DESIGN AND METHODS After preliminary dose-finding studies, either saline vehicle or recombinant human FGF21 (0.4 μg/day) was infused continuously for 2 weeks into the lateral cerebral ventricle of male Wistar rats rendered obese by high-fat feeding. Study end points included measures of energy balance (body weight, body composition, food intake, energy expenditure, and circulating and hepatic lipids) and glucose metabolism (insulin tolerance test, euglycemic-hyperinsulinemic clamp, and hepatic expression of genes involved in glucose metabolism). RESULTS Compared with vehicle, continuous intracerebroventricular infusion of FGF21 increased both food intake and energy expenditure in rats with DIO, such that neither body weight nor body composition was altered. Despite unchanged body fat content, rats treated with intracerebroventricular FGF21 displayed a robust increase of insulin sensitivity due to increased insulin-induced suppression of both hepatic glucose production and gluconeogenic gene expression, with no change of glucose utilization. CONCLUSIONS FGF21 action in the brain increases hepatic insulin sensitivity and metabolic rate in rats with DIO. These findings identify the central nervous system as a potentially important target for the beneficial effects of FGF21 in the treatment of diabetes and obesity.

Identification of Body Fat Mass as a Major Determinant of Metabolic Rate in Mice

OBJECTIVE Analysis of energy expenditure (EE) in mice is essential to obesity research. Since EE varies with body mass, comparisons between lean and obese mice are confounded unless EE is normalized to account for body mass differences. We 1) assessed the validity of ratio-based EE normalization involving division of EE by either total body mass (TBM) or lean body mass (LBM), 2) compared the independent contributions of LBM and fat mass (FM) to EE, and 3) investigated whether leptin contributes to the link between FM and EE. RESEARCH DESIGN AND METHODS We used regression modeling of calorimetry and body composition data in 137 mice to estimate the independent contributions of LBM and FM to EE. Subcutaneous administration of leptin or vehicle to 28 obese ob/ob mice and 32 fasting wild-type mice was used to determine if FM affects EE via a leptin-dependent mechanism. RESULTS Division of EE by either TBM or LBM is confounded by body mass variation. The contribution of FM to EE is comparable to that of LBM in normal mice (expressed per gram of tissue) but is absent in leptin-deficient ob/ob mice. When leptin is administered at physiological doses, the plasma leptin concentration supplants FM as an independent determinant of EE in both ob/ob mice and normal mice rendered leptin-deficient by fasting. CONCLUSIONS The contribution of FM to EE is substantially greater than predicted from the metabolic cost of adipose tissue per se, and the mechanism underlying this effect is leptin dependent. Regression-based approaches that account for variation in both FM and LBM are recommended for normalization of EE in mice.

Peripheral oxytocin suppresses food intake and causes weight loss in diet-induced obese rats

Growing evidence suggests that oxytocin plays an important role in the regulation of energy balance and that central oxytocin administration induces weight loss in diet-induced obese (DIO) animals. To gain a better understanding of how oxytocin mediates these effects, we examined feeding and neuronal responses to oxytocin in animals rendered obese following exposure to either a high-fat (HFD) or low-fat diet (LFD). Our findings demonstrate that peripheral administration of oxytocin dose-dependently reduces food intake and body weight to a similar extent in rats maintained on either diet. Moreover, the effect of oxytocin to induce weight loss remained intact in leptin receptor-deficient Koletsky (fa(k)/fa(k)) rats relative to their lean littermates. To determine whether systemically administered oxytocin activates hindbrain areas that regulate meal size, we measured neuronal c-Fos induction in the nucleus of the solitary tract (NTS) and area postrema (AP). We observed a robust neuronal response to oxytocin in these hindbrain areas that was unexpectedly increased in rats rendered obese on a HFD relative to lean, LFD-fed controls. Finally, we report that repeated daily peripheral administration of oxytocin in DIO animals elicited a sustained reduction of food intake and body weight while preventing the reduction of energy expenditure characteristic of weight-reduced animals. These findings extend recent evidence suggesting that oxytocin circumvents leptin resistance and induces weight-loss in DIO animals through a mechanism involving activation of neurons in the NTS and AP, key hindbrain areas for processing satiety-related inputs.

Leptin Deficiency Causes Insulin Resistance Induced by Uncontrolled Diabetes

OBJECTIVE Depletion of body fat stores during uncontrolled, insulin-deficient diabetes (uDM) results in markedly reduced plasma leptin levels. This study investigated the role of leptin deficiency in the genesis of severe insulin resistance and related metabolic and neuroendocrine derangements induced by uDM. RESEARCH DESIGN AND METHODS Adult male Wistar rats remained nondiabetic or were injected with the β-cell toxin, streptozotocin (STZ) to induce uDM and subsequently underwent subcutaneous implantation of an osmotic minipump containing either vehicle or leptin at a dose (150 μg/kg/day) designed to replace leptin at nondiabetic plasma levels. To control for leptin effects on food intake, another group of STZ-injected animals were pair fed to the intake of those receiving leptin. Food intake, body weight, and blood glucose levels were measured daily, with body composition and indirect calorimetry performed on day 11, and an insulin tolerance test to measure insulin sensitivity performed on day 16. Plasma hormone and substrate levels, hepatic gluconeogenic gene expression, and measures of tissue insulin signal transduction were also measured. RESULTS Physiologic leptin replacement prevented insulin resistance in uDM via a mechanism unrelated to changes in food intake or body weight. This effect was associated with reduced total body fat and hepatic triglyceride content, preservation of lean mass, and improved insulin signal transduction via the insulin receptor substrate–phosphatidylinositol-3-hydroxy kinase pathway in the liver, but not in skeletal muscle or adipose tissue. Although physiologic leptin replacement lowered blood glucose levels only slightly, it fully normalized elevated plasma glucagon and corticosterone levels and reversed the increased hepatic expression of gluconeogenic enzymes characteristic of rats with uDM. CONCLUSIONS We conclude that leptin deficiency plays a key role in the pathogenesis of severe insulin resistance and related endocrine disorders in uDM. Treatment of diabetes in humans may benefit from correction of leptin deficiency as well as insulin deficiency.

Identification of a physiological role for leptin in the regulation of ambulatory activity and wheel running in mice

Mechanisms regulating spontaneous physical activity remain poorly characterized despite evidence of influential genetic and acquired factors. We evaluated ambulatory activity and wheel running in leptin-deficient ob/ob mice and in wild-type mice rendered hypoleptinemic by fasting in both the presence and absence of subcutaneous leptin administration. In ob/ob mice, leptin treatment to plasma levels characteristic of wild-type mice acutely increased both ambulatory activity (by 4,000 ± 200 beam breaks/dark cycle, P < 0.05) and total energy expenditure (TEE; by 0.11 ± 0.01 kcal/h during the dark cycle, P < 0.05) in a dose-dependent manner and acutely increased wheel running (+350%, P < 0.05). Fasting potently increased ambulatory activity and wheel running in wild-type mice (AA: +25%, P < 0.05; wheel running: +80%, P < 0.05), and the effect of fasting was more pronounced in ob/ob mice (AA: +400%, P < 0.05; wheel running: +1,600%, P < 0.05). However, unlike what occurred in ad libitum-fed ob/ob mice, physiological leptin replacement attenuated or prevented fasting-induced increases of ambulatory activity and wheel running in both wild-type and ob/ob mice. Thus, plasma leptin is a physiological regulator of spontaneous physical activity, but the nature of leptins effect on activity is dependent on food availability.

Deficiency of TNFα converting enzyme (TACE/ADAM17) causes a lean, hypermetabolic phenotype in mice

Energy homeostasis involves central nervous system integration of afferent inputs that coordinately regulate food intake and energy expenditure. Here, we report that adult homozygous TNFalpha converting enzyme (TACE)-deficient mice exhibit one of the most dramatic examples of hypermetabolism yet reported in a rodent system. Because this effect is not matched by increased food intake, mice lacking TACE exhibit a lean phenotype. In the hypothalamus of these mice, neurons in the arcuate nucleus exhibit intact responses to reduced fat mass and low circulating leptin levels, suggesting that defects in other components of the energy homeostasis system explain the phenotype of Tace(DeltaZn/DeltaZn) mice. Elevated levels of uncoupling protein-1 in brown adipose tissue from Tace(DeltaZn/DeltaZn) mice when compared with weight-matched controls suggest that deficient TACE activity is linked to increased sympathetic outflow. These findings collectively identify a novel and potentially important role for TACE in energy homeostasis.

Central administration of interleukin-4 exacerbates hypothalamic inflammation and weight gain during high-fat feeding

In peripheral tissues, the link between obesity and insulin resistance involves low-grade inflammation induced by macrophage activation and proinflammatory cytokine signaling. Since proinflammatory cytokines are also induced in the hypothalamus of animals placed on a high-fat (HF) diet and can inhibit neuronal signal transduction pathways required for normal energy homeostasis, hypothalamic inflammation is hypothesized to contribute to the pathogenesis of diet-induced obesity (DIO). We addressed this hypothesis by perturbing the inflammatory milieu of the hypothalamus in adult male Wistar rats using intracerebroventricular (icv) administration of interleukin-4 (IL-4), a Th2 cytokine that promotes alternative activation (M2) of macrophages and microglia. During HF feeding, icv IL-4 administration increased hypothalamic proinflammatory cytokine gene expression and caused excess weight gain. Intracerebroventricular pretreatment with PS1145, an inhibitor of IKKbeta (a key intracellular mediator of inflammatory signaling), blocked both IL-4 effects, suggesting a causal relationship between IL-4-induced weight gain and hypothalamic inflammation. These observations add to growing evidence linking hypothalamic inflammation to obesity pathogenesis.

Increased energy expenditure and leptin sensitivity account for low fat mass in myostatin-deficient mice.

Myostatin deficiency causes dramatically increased skeletal muscle mass and reduced fat mass. Previously, myostatin-deficient mice were reported to have unexpectedly low total energy expenditure (EE) after normalizing to body mass, and thus, a metabolic cause for low fat mass was discounted. To clarify how myostatin deficiency affects the control of body fat mass and energy balance, we compared rates of oxygen consumption, body composition, and food intake in young myostatin-deficient mice relative to wild-type (WT) and heterozygous (HET) controls. We report that after adjusting for total body mass using regression analysis, young myostatin-deficient mice display significantly increased EE relative to both WT (+0.81 ± 0.28 kcal/day, P = 0.004) and HET controls (+0.92 ± 0.31 kcal/day, P = 0.005). Since food intake was not different between groups, increased EE likely accounts for the reduced body fat mass (KO: 8.8 ± 1.1% vs. WT: 14.5 ± 1.3%, P = 0.003) and circulating leptin levels (KO: 0.7 ± 0.2 ng/ml vs. WT: 1.9 ± 0.3 ng/ml, P = 0.008). Interestingly, the observed increase in adjusted EE in myostatin-deficient mice occurred despite dramatically reduced ambulatory activity levels (-50% vs. WT, P < 0.05). The absence of hyperphagia together with increased EE in myostatin-deficient mice suggests that increased leptin sensitivity may contribute to their lean phenotype. Indeed, leptin-induced anorexia (KO: -17 ± 1.2% vs. WT: -5 ± 0.3%) and weight loss (KO: -2.2 ± 0.2 g vs. WT: -1.6 ± 0.1, P < 0.05) were increased in myostatin-deficient mice compared with WT controls. We conclude that increased EE, together with increased leptin sensitivity, contributes to low fat mass in mice lacking myostatin.

Mice Lacking Hepatic Lipase Are Lean and Protected against Diet-Induced Obesity and Hepatic Steatosis

Hepatic lipase (HL)-mediated lipoprotein hydrolysis provides free fatty acids for energy, storage, and nutrient signaling and may play a role in energy homeostasis. Because HL-activity increases with increased visceral fat, we hypothesized that increased HL-activity favors weight gain and obesity and consequently, that HL deficiency would reduce body fat stores and protect against diet-induced obesity. To test this hypothesis, we compared wild-type mice (with endogenous HL) and mice genetically deficient in HL with respect to daily body weight and food intake, body composition, and adipocyte size on both chow and high-fat (HF) diets. Key determinants of energy expenditure, including rate of oxygen consumption, heat production, and locomotor activity, were measured by indirect calorimetry. HL-deficient mice exhibited reduced weight gain on both diets (by 32%, chow; by 50%, HF; both P < 0.0001, n = 6-7 per genotype), effects that were associated with reduced average daily food intake (by 22-30% on both diets, P < 0.0001) and a modest increase in the rate of oxygen consumption (by 25%, P < 0.003) during the light cycle. Moreover, in mice fed the HF diet, HL deficiency reduced both body fat (by 30%, P < 0.0001) and adipocyte size (by 53%, P < 0.01) and fully prevented the development of hepatic steatosis. Also, HL deficiency reduced adipose tissue macrophage content, consistent with reduced inflammation and a lean phenotype. Our results demonstrate that in mice, HL deficiency protects against diet-induced obesity and its hepatic sequelae. Inhibition of HL-activity may therefore have value in the prevention and/or treatment of obesity.