Lynda M. Brown

Estradiol-dependent decrease in the orexigenic potency of ghrelin in female rats.

Ghrelin, the only known orexigenic gut hormone, is secreted mainly from the stomach, increases with fasting and before meal initiation in humans and rats, and increases food intake after central or peripheral administration. To investigate sex differences in the action of ghrelin, we assessed the effects of exogenous ghrelin in intact male and female rats, the effects of exogenous ghrelin in ovariectomized (OVX) and estradiol (E2)-treated female rats, as well as the effects of OVX on plasma ghrelin and hypothalamic orexigneic neuropeptide expression in rats and on food intake and weight gain in transgenic mice lacking the ghrelin receptor (Ghsr−/− mice). Male and OVX female rats were significantly more sensitive than intact female rats to the orexigenic effects of both centrally (intra–third ventricular, i3vt, 0.01, 0.1, and 1.0 nmol) and systemically (ip, 3, 6, and 9 nmol) administered ghrelin. This difference is likely to be estradiol dependent because E2 attenuated the orexigenic action of ghrelin in OVX female and male rats. Furthermore, OVX increased food intake and body weight in wild-type mice, but not in Ghsr−/− mice, suggesting that OVX increases food intake by releasing ghrelin from a tonic inhibitory effect of estradiol. In addition, following OVX, there was an increase in plasma ghrelin that was temporally associated with increased food intake, body weight, and hypothalamic neuropeptide Y and Agouti-related protein mRNA expression. Collectively, these data suggest that estradiol inhibits the orexigenic action of ghrelin in females, that weight gain associated with OVX is ghrelin mediated, and that this endocrine interaction may account for an important sex differences in food intake and the regulation of body weight.

Consumption of a high-fat diet induces central insulin resistance independent of adiposity

Plasma insulin enters the CNS where it interacts with insulin receptors in areas that are related to energy homeostasis and elicits a decrease of food intake and body weight. Here, we demonstrate that consumption of a high-fat (HF) diet impairs the central actions of insulin. Male Long-Evans rats were given chronic (70-day) or acute (3-day) ad libitum access to HF, low-fat (LF), or chow diets. Insulin administered into the 3rd-cerebral ventricle (i3vt) decreased food intake and body weight of LF and chow rats but had no effect on HF rats in either the chronic or the acute experiment. Rats chronically pair-fed the HF diet to match the caloric intake of LF rats, and with body weights and adiposity levels comparable to those of LF rats, were also unresponsive to i3vt insulin when returned to ad libitum food whereas rats pair-fed the LF diet had reduced food intake and body weight when administered i3vt insulin. Insulins inability to reduce food intake in the presence of the high-fat diet was associated with a reduced ability of insulin to activate its signaling cascade, as measured by pAKT. Finally, i3vt administration of insulin increased hypothalamic expression of POMC mRNA in the LF- but not the HF-fed rats. We conclude that consumption of a HF diet leads to central insulin resistance following short exposure to the diet, and as demonstrated by reductions in insulin signaling and insulin-induced hypothalamic expression of POMC mRNA.

Metabolic impact of sex hormones on obesity.

Obesity and its associated health disorders and costs are increasing. Men and post-menopausal women have greater risk of developing complications of obesity than younger women. Within the brain, the hypothalamus is an important regulator of energy homeostasis. Two of its sub-areas, the ventrolateral portion of the ventral medial nucleus (VL VMN) and the arcuate (ARC) respond to hormones and other signals to control energy intake and expenditure. When large lesions are made in the hypothalamus which includes both the VL VMN and the ARC, animals eat more, have reduced energy expenditure, and become obese. The ARC and the VL VMN, in addition to other regions in the hypothalamus, have been demonstrated to contain estrogen receptors. There are two estrogen receptors, estrogen receptor alpha (ERalpha) and estrogen receptor beta (ERbeta). We and others have previously demonstrated that activation of ERalpha by estrogens reduces food intake and increases body weight. This review focuses on the relative contribution of activation of ERalpha by estrogens in the ARC and the VL VMN in the regulation of food intake and body weight. Additionally, estrogen receptors have been found in many peripheral tissues including adipose tissue. Estrogens are thought to have direct effects on adipose tissue and estrogens may provide anti-inflammatory properties both in the periphery and the in the central nervous system (CNS) which may protect women from diseases associated with inflammation. Understanding the mechanisms by which estrogens regulate body weight and inflammation will assist in determining potential therapeutic agents for menopausal women to decrease the propensity of diseases associated with obesity.

Intraventricular insulin and leptin reduce food intake and body weight in C57BL/6J mice

As the incidence of obesity continues to increase, adequate animal models acquire increased importance for the investigation of energy homeostatic mechanisms. Understanding the central mechanism of action of the adiposity hormones, insulin and leptin, has become particularly important as researchers examine ways to treat or prevent obesity. Although the intra-3rd-ventricular (i3vt) administration of insulin reduces food intake in several species, its effects on food intake and body weight have not been previously been assessed in mice. Male C57BL/6J mice were administered insulin i3vt (0.05, 0.1 or 0.4 microU) or leptin i3vt (5 microg/1 microl) as a positive control. As it occurs in other species, i3vt insulin dose-dependently reduced 24-h food intake and body weight, and increased hypothalamic proopiomelanocortin (POMC) mRNA. Hence, genetic manipulations that influence brain insulin sensitivity in mice can now more easily be integrated with the broader literature on energy homeostasis.

Intraventricular (i3vt) ghrelin increases food intake in fatty Zucker rats.

Ghrelin is an orexigenic peptide secreted from the stomach and also made in the brain. Ghrelin receptors are expressed on hypothalamic cells important in appetite and energy balance. We determined that intra-3rd-ventricular (i3vt) ghrelin dose-dependently increases acute (1 and 2 h) food intake in lean and fatty Zucker rats (0, 0.01, 0.1 and 1.0 nmol ghrelin). The percentage increase of food intake in fatty Zucker rats was significantly greater than that in lean rats. Fatty Zucker rats had 4.5 times more ghrelin receptor mRNA in the hypothalamus than lean Zucker rats, suggesting a possible mechanism for the increased sensitivity.

The effects of adrenalectomy and aldosterone replacement in transgenic mice expressing antisense RNA to the type 2 glucocorticoid receptor

Bilateral adrenalectomy (ADX) either prevents or attenuates obesity in several animal models. Mice that express an antisense RNA to the glucocorticoid receptor (GCR) are obese. The present study was conducted to examine the effects of ADX and aldosterone (ALDO) replacement on the rate of weight gain and body composition of mice bearing an antisense GCR gene construct. Twenty-eight male transgenic mice bearing the antisense GCR construct and 16 male B6C/3F1 mice were either bilaterally ADX or given sham operations. At the time of surgery, some of the ADX mice and all of the sham-operated mice were implanted with 100-mg cholesterol (CHOL) pellets inserted subcutaneously in the subscapular region. The remaining ADX mice were implanted with 100-mg 1% w/w ALDO pellets using CHOL as vehicle. All mice were returned to their home cages for 2 weeks. They were then decapitated and the blood was collected for corticosterone, ALDO, insulin, and leptin radioimmunoassay. Carcasses were eviscerated and prepared for gravimetric analyses, including bomb calorimetry. ADX resulted in a significant drop in carcass fat in both transgenic and wildtype groups. ALDO prevented the decrease in carcass fat in both groups. Two weeks after ADX, transgenic mice were as fat as sham-operated wildtype controls, whereas both sham-operated and ALDO-treated transgenic groups were significantly fatter. Despite observing a reliable decrease in carcass fat following ADX, no corresponding decrease in circulating leptin was found.

Comparison of DEXA and QMR for Assessing Fat and Lean Body Mass in Adult Rats

There are several techniques used to measure body composition in experimental models including dual energy X-ray absorptiometry (DEXA) and quantitative magnetic resonance (QMR). DEXA/QMR data have been compared in mice, but have not been compared previously in rats. The goal of this study was to compare DEXA and QMR data in rats. We used rats that varied by sex, diet, and age, in addition we compared dissected samples containing subcutaneous (pelt) or visceral fat (carcass). The data means were compared by focusing on the differences between DEXA/QMR data using a series of scatter plots without assuming that either method is more accurate as suggested by Bland and Altman. DEXA/QMR data did not agree sufficiently in carcass or pelt FM or in pelt LBM. The variation observed within these groups suggests that DEXA and QMR measurements are not comparable. Carcass LBM in young rats did yield comparable data once the data for middle-aged rats was removed. The variation in our data may be a result of different direct and indirect measures that DEXA and QMR technologies use to quantify FM and LBM. DEXA measures FM and estimates fat-free mass. In contrast, QMR uses separate equations of magnetic resonance to measure FM, LBM, total body water and free water. We found that QMR overestimated body mass in our middle-aged rats, and this increased the variation between methods. Our goal was to evaluate the precision of DEXA/QMR data in rats to determine if they agree sufficiently to allow direct comparison of data between methods. However DEXA and QMR did not yield the same estimates of FM or LBM for the majority of our samples.

Estrogens, inflammation and obesity: an overview

Emerging research has suggested that inflammatory stress may play a role in the development of obesity. Both the leptin and insulin receptor are sensitive to intracellular inflammatory signaling that can be stimulated through toll-like receptor 4 activation by saturated fat. Pharmacological intervention within this cascade often protects animals from becoming obese, thus highlighting inflammatory pathways as a possible site of study in the prevention of pathologic weight gain. It has been well established in animal models that females display a marked reduction in the susceptibility to weight gain on high-fat diets compared to males. In addition, it has been widely accepted that females are partially protected from inflammatory-related diseases. At the molecular level, this reduction in disease susceptibility has been suggested to be due to the anti-inflammatory properties of 17 β-estradiol. Through direct free radical scavenging, transcriptional regulation, and protein interactions, chronic exposure to estradiol can reduce systemic inflammatory stress. As the knowledge base continues to grow on the etiology of obesity, further research is needed on the precise molecular pathways that can be inhibited by estradiol. Understanding of such pathways may provide a basis for the future use of estrogen and its related compounds (daidzein, genistein, resveratrol) to prevent weight gain in peri- and post-menopausal females.

The effect of TNFα on food intake and central insulin sensitivity in rats

Circulating and tissue levels of the proinflammatory cytokine tumor necrosis factor α (TNFα) are elevated in obesity. TNFα interferes with insulin signaling in many tissues and also plays a causal role in the anorexia that accompanies severe challenges to the immune system. The interactions between TNFα and insulin in the control of eating are less well known. The present study evaluated the role of TNFα in the central nervous system control of food intake by insulin in adult male Long Evans rats. We first determined the ability of several doses of TNFα injected into the 3rd cerebral ventricle (i3vt) to reduce food intake in male rats. Subsequently, we assessed the ability of a subthreshold dose of TNFα to modulate the effect of i3vt insulin on food intake in male rats fed a low-fat chow or a high-fat (HF) diet. TNFα administered i3vt dose-dependently reduced food intake in rats fed a standard low-fat chow diet. Moreover, a low, sub-threshold dose of TNFα diminished the reduction in food intake by insulin in rats maintained on a chow diet, but enhanced insulin action in rats maintained on a HF diet. These data suggest that the interaction of TNFα with central insulin varies with nutritional and/or dietary conditions.

Acute exposure to high-fat diets increases hepatic expression of genes related to cell repair and remodeling in female rats

High-fat diets (HFD) promote the development of both obesity and fatty liver disease through the up-regulation of hepatic lipogenesis. Insulin resistance, a hallmark of both conditions, causes dysfunctional fuel partitioning and increases in lipogenesis. Recent work has demonstrated that systemic insulin resistance occurs in as little as the first 72 hours of an HFD, suggesting the potential for hepatic disruption with HFD at this time point. The current study sought to determine differences in expression of lipogenic genes between sexes in 3-month-old male and female Long-Evans rats after 72 hours of a 40% HFD or a 17% fat (chow) diet. Owing to the response of estrogen on hepatic signaling, we hypothesized that a sexual dimorphic response would occur in the expression of lipogenic enzymes, inflammatory cytokines, apoptotic, and cell repair and remodeling genes. Both sexes consumed more energy when fed an HFD compared with their low fat-fed controls. However, only the males fed the HFD had a significant increase in body fat. Regardless of sex, HFD caused down-regulation of lipogenic and inflammatory genes. Interestingly, females fed an HFD had up-regulated expression of apoptotic and cell repair-related genes compared with the males. This may suggest that females are more responsive to the acute hepatic injury effects caused by HFDs. In summary, neither male nor female rats displayed disrupted hepatic metabolic pathways after 72 hours of the HFD treatment. In addition, female rats appear to have protection from increases in fat deposition, possibly due to increased caloric expenditure; male rats fed an HFD were less active, as demonstrated by distance traveled in their home cage.