Stephen C. Woods - US grants

Funds are requested for the purchase of an analytical electron microscope for use on three major research projects by five NIH supported investigators. The three project areas involve the determination of subcellular elemental distributions in studies of: 1) Basic mechanisms of contraction in cardiac muscle. 2) Salivary gland physiology particularly as related to Cystic Fibrosis. 3) The role of intracellular calcium stores in the control of ciliary activity. In these studies the common techniques employed are: 1) Specimen preparation via cryofixation, thin cryosectioning (100-200nm) and freeze drying. 2) High resolution imaging via conventional transmission and scanning transmission electron microscopy. 3) Quantitative elemental microanalysis via energy dispersive x-ray analysis. All three major projects are already underway on an existing JEOL 100C (purchased in 1976). Although this instrument is capable of generating excellent data, it is limited by its nature (oil diffusion pumped). More importantly, however, is the basic limitation of instrument time. The three major projects must share this instrument with several others including, for example, the engineering development of energy loss spectrometry instrumentation (particularly a prototype parallel detection system). Thus, productivity for all is severely limited. If this funding request is approved, a modern, ion pumped, analytical electron microscope will be utilized entirely by the three main projects listed. The existing instrument will then be devoted to: i) developmental projects including instrumental, technique, and new applications, ii) continuing smaller projects, and iii) training and support activities for the major projects. The overall effect of the increased access to this important technology will be to more than double the rate at which progress is made on the major projects while still allowing new instrumentation and new applications to be developed and smaller projects to continue.

health science research support; university;

DESCRIPTION (provided by applicant): The specific aims remain essentially the same for this long-running project; i.e., to elucidate how adiposity signals interact with the brain to influence food intake and body weight. The adiposity signals now include leptin as well as insulin. Specific Aim 1 will determine the interactions of insulin with the hypothalamic propiomelanocoritin (POMC) and neuropeptide Y (NPY) systems. The hypothesis driving the experiments is that insulin will potentiate the POMC system and inhibit the NPY system. Specific Aim 2 will determine the interactions of insulin and leptin within the brain. The hypothesis is that the two peptides will synergistically elicit a net catabolic action. Specific Aim 3 will assess the hypothesis that a major gender difference exists in the central control of energy homeostasis, with insulin controlling the male system and leptin controlling the female system. Finally, Specific Aim 4 will determine the effect of eliminating insulin signaling in the brain by creating mice with brain-specific insulin receptor deletions. All of the experiments are directly relevant to emerging therapeutic approaches to the treatment of obesity.

biomedical equipment resource;

Obesity is a major health problem. For in spite of considerable effort by the scientific and healthcare professions to understand and successfully treat obesity, its incidence continues to rise and the obesity-related costs to society are staggering. The enormity of the problem is reflected in recent surveys and goals of the World Health Organization and the National Institutes of Health. The effort may soon be paying dividends, however, in that two factors have come to light in recent years that shed light on fundamental regulatory system that works to maintain a particular amount of fat in the body. This amount of fat may vary in different environments, and certainly varies considerably among individuals. Nonetheless, in a constant environment, individuals rigorously maintain and defend a particular amount of total stored energy in the form of fat. The second major factor that has come to light concerning human obesity is that when a diet with a high fat content is consumed on a regular basis, the amount of stored fat in the diet increases over time, as has occurred in many nations over the past thirty years, the incidence of obesity also increases. The key negative feedback system that regulates body fat slips. More fat is stored and the individual moves along the scale toward obesity. It is the interaction of these two factors that is the subject of this proposal. More specifically, over three interrelated projects, we ask how consumption of a high-fat diet modifies the negative feedback system that controls body fat. The driving hypothesis of this or any or all of several levels of control. Project 1 explores the processing of ingested fat by the gut and considers the generation and receptor of meal-related signals that control meal size. Project 2 investigates the generation and reception by the brain of adiposity-indicating signals. Finally, Project 3 considers brain neurotransmitter systems that control food intake and metabolism. Identifying the regulatory processes that mediate high-fat diet-induced obesity is the major goal of this project.

DESCRIPTION (provided by applicant): We are requesting support to continue a highly successful training program that supports predoctoral and postdoctoral training in neuroendocrinology, with a specific focus on homeostasis and the interactions among neuroendocrine systems controlling energy balance, the hypothalamo-pituitary-adrenal (HPA) axis and reproduction. The University of Cincinnati has enjoyed tremendous growth in neuroscience research over the past decade, with a major concentration of this growth in neuroendocrinology. This is a direct result of the University's explicit commitment to target neuroscience and energy homeostasis as major focus areas for growth and research excellence over the next decade. World-recognized leaders in neuroendocrinology, in the areas of energy balance, HPA axis and reproduction, have been recruited to the faculty ranks in both basic science and clinical Departments in the College of Medicine, and the best of these are mentors on this training grant. These researchers are extremely productive and well funded with a strong history of mentoring successful pre- and postdoctoral trainees. Furthermore, this group of researchers has established a strong set of collaborations and bolstered its ability to provide mentorship and research opportunities to graduate students and fellows working in numerous programs. The strength of the faculty has enabled us to recruit outstanding trainees, and the fruits of this effort are manifest in the productivity and job opportunities that accrued to the trainees supported in the current funding period.

[unreadable] DESCRIPTION (provided by applicant): Numerous systems participate in the integrated processes that cause an animal to seek and ingest food, and to stop eating when more food is available. This project relates to the acute effects of ingesting large numbers of calories at 1 time, for whereas supplying new nutrients to the body is the ultimate goal of eating, it can only be accomplished by perturbing the delicate balance of circulating nutrients, at least on a temporary basis. Because of this, animals have evolved elaborate strategies to minimize the acute impact of meals on blood glucose and other nutrients. Many of these adaptive responses are made in anticipation of eating large meals, and they and their consequences are the focus of this proposal. We have also found that meal-fed animals that consume all of their daily food in a short period of time have greatly improved glucose tolerance at the time they are expecting food. The goal of this project is to understand the mechanisms underlying this adaptation in order to inform future therapies. Based upon our novel observations, we have developed 3 specific aims. Specific Aim 1 will utilize euglycemic hyperinsulinemic or hyperglycemic clamps to identify the metabolic adaptations responsible for improved glucose tolerance, testing specific hypotheses concerning changes in peripheral insulin resistance, glucose disposal rate and/or B-cell function. These experiments will utilize laboratory rats and mice. Specific Aim 2 will test the hypothesis that a comparable battery of food anticipatory responses occurs when ad lib-fed rats anticipate eating large meals. Specific Aim 3 will test the hypothesis that meal-fed rats are enabled to eat large amounts of food in a short period of time because they have become relatively unresponsive to satiety signals. This project will identify behavioral approaches that improve glucose tolerance. Because impaired glucose tolerance is a major symptom of diabetes mellitus and many instances of obesity, and because most current therapies achieve only modest improvements in glucose tolerance, the proposed research has considerable and compelling health implications; i.e., based on the proposed research, novel therapies for individuals with impaired glucose tolerance might include specific eating regimens that would yield powerful health benefits to complement pharmacological approaches. [unreadable] [unreadable]

[unreadable] DESCRIPTION (provided by applicant): [unreadable] In spite of considerable effort by the scientific and healthcare professions to understand and successfully treat obesity, its incidence continues to rise and the obesity-related costs to society are staggering. The enormity of the problem is reflected in recent surveys and goals of the World Health Organization and the National Institutes of Health. Two factors have come to light in recent years that shed light on fundamental aspects of the problem and may hold a key for future therapies. The first factor is that the body possesses a remarkably efficient regulatory system that works to maintain a particular amount of fat in the body. This amount of fat may vary in different environments, and it certainly varies considerably among individuals. Nonetheless, in a constant environment, individuals rigorously defend a particular amount of total stored energy in the form of fat. The second major factor concerning human obesity is that when a diet with a high saturated fat content is consumed on a regular basis, the amount of stored fat that the body defends increases. Hence, epidemiological studies have identified a significant correlation between average dietary fat intake and the incidence of obesity and its related complications mad risk factors among nations. And when the average amount of fat in the diet increases over time, as has occurred in many nations over the past thirty years, the incidence of obesity also increases. The key point is that when individuals are exposed, on a chronic basis, to a higher average level of dietary saturated fat, the otherwise incredibly robust negative feedback system that regulates body fat slips. More fat is stored and the individual moves toward obesity. It is the interaction of these two factors that is the subject of this proposal. More specifically, over three interrelated projects, we ask how consumption of a high saturated fat diet modifies a number of critical factors that determine body fat and the predisposition to become diabetic. Project 1 focuses on how the gut and CNS protein Apo A-IV regulate food intake and can be altered by diets with different amounts and types of fat. Project 2 tests several hypotheses about how saturated versus monounsaturated fats influence the adaptation of the pancreatic beta-cell to increasing insulin resistance. Project 3 seeks to elucidate the mechanisms in the CNS that contribute to the defense of increased body fat that occurs on high saturated fat diet. [unreadable] [unreadable] [unreadable]

[unreadable] DESCRIPTION (provided by applicant): Multiple inputs and controls participate in the maintenance of energy homeostasis, and numerous neural and endocrine systems are involved in the integrated responses that maintain energy stores in the body at a level. Since most stored energy is in the form of adipose tissue (fat), and since the brain is the major controller of energy homeostasis, a fundamental requirement is that the brain receive accurate and timely information about the amount and distribution of body fat. This information is then integrated with other factors to determine whether an individual seeks and eats food (i.e., energy intake), as well as the efficiency with which the ingested food is utilized (energy expenditure) or stored. Information regarding the amount of fat stored in the body reaches the brain via the circulating signals insulin and leptin. Each is secreted into the blood in direct proportion to total body fat, and each is transported from the blood into the brain where it interacts with neurons situated to influence energy intake and expenditure. Hence, insulin and leptin are considered to be adiposity signals, and the long-term goal of this project is to elucidate how adiposity signals interact with other factors to influence energy homeostasis. We anticipate that the coming decade will see considerable progress in this arena, as well as the translation of the new-found information into potential novel therapeutic strategies to tackle clinical problems of dysregulation of energy homeostasis (i.e., obesity, eating disorders). With this as an ultimate end point, the present proposal has three specific aims that address critical and as yet unanswered questions regarding the actions of adiposity signals in the brain. The first assesses the hypothesis that insulin and leptin act in the hypothalamus by enhancing the signal provided by local levels of nutrients, including glucose and fatty acids. The second assesses several hypotheses following from our observation that there are important gender differences in the actions of insulin and leptin in the central control of energy homeostasis, and that males and females utilize different strategies to regulate energy homeostasis and defend their body fat stores. The final aim assesses hypotheses regarding the effect of reducing insulin signaling in specific brain regions on food intake and body weight, and upon related levels of metabolic hormones and neuropeptides, utilizing lentiviral technology. PUBLIC HEALTH RELEVANCE: Hormones such as insulin and leptin that are secreted in direct proportion to body fat interact in the brain with nutrients such as glucose, fatty acids and amino acids to determine food intake and energy expenditure. Proposed research will determine how these signals interact within neurons in the hypothalamus and intervene by manipulating the ability of the brain of laboratory rats to detect and respond to insulin or leptin. Experiments will assess behavior as well as molecular signals within cells. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): This project is based upon the relatively recent realization that obesity, and especially visceral obesity, is a chronic inflammatory condition characterized by migration of high numbers of macrophages into the adipose tissue, with consequent increased local and plasma levels of cytokines. The consequences of these changes are thought to be causally linked to insulin resistance and the metabolic syndrome. In particular, high levels of inflammatory cytokines in the plasma, and especially tumor necrosis factor-1 (TNF1), have been recognized as an important risk factor for several metabolic and cardiovascular disorders, and in animal models reductions in TNF1 ameliorate many of the deleterious effects of obesity. There is compelling evidence that an important initial step along the road to obesity is an increase of TNF1 synthesis by adipose tissue, and that this in turn stimulates preadipocytes and local endothelial cells to recruit macrophages into the adipose tissue. This continues as a spiraling situation of increased inflammatory signaling molecules and mutual cross-stimulation of adipocytes and macrophages. TNF1 and other inflammatory signaling molecules eventually reach high enough levels to be detected in the plasma. Because TNF1 from adipose tissue circulates throughout the body and reaches diverse receptor populations, the role of TNF1 activity at specific sites, and at selective receptors, in the etiology of obesity and insulin resistance is not known. We have preliminary evidence that one important site of TNF1 action is in the brain, where it interacts with other signals that influence energy intake. Proposed experiments will (1) test the hypothesis that TNF1 acts in the brain to control energy homeostasis, and specifically that TNF1 action within the brain increases central insulin sensitivity while simultaneously decreasing systemic insulin sensitivity. A related hypothesis is that TNF1 is more effective in the brains of animals that have systemic insulin resistance, such as high-fat diet (HFD) induced obesity;(2) assess the relative contributions of free (unbound;soluble) TNF1 and membrane-bound TNF1 in the effects of TNF1 on energy balance;and (3) determine whether subcutaneous adipose tissue, mesenteric adipose tissue, or adipose tissue-resident macrophages are individually (or in combination) sufficient sources of TNF1 to elicit symptoms of the metabolic syndrome. Experiments will use normal (wild-type) mice and mice lacking TNF1 or its receptors or else mice lacking TACE, the enzyme that cleaves membrane-bound TNF1 to free TNF1. PUBLIC HEALTH RELEVANCE: Visceral obesity is recognized to be a chronic inflammatory condition characterized by high numbers of macrophages in adipose tissue and elevated plasma levels of cytokines such as tumor necrosis factor-1 (TNF1) that is thought to lead to insulin resistance. Proposed experiments will test the hypothesis that TNF1 acts in the brain to control food intake and systemic insulin sensitivity, and determine which form of TNF1 is critical. Experiments will use normal (wild-type) mice and mice lacking TNF1 or its receptors.

DESCRIPTION (provided by applicant): There is a fundamental gap in understanding the mechanism(s) by which estrogens potently inhibit food intake and reduce body weight in many species, including humans. This gap represents an important problem because unless it is closed, it is unlikely that estrogenic intermediaries and pathways can be targeted as a means of obesity prevention and treatment in women with impaired estrogen signaling. Compelling evidence suggests that E2 exerts its anorexigenic action through an indirect mechanism; i.e., that E2 increases the strength of other physiological signals that reduce meal size. Apolipoprotein A-IV (apoA-IV), an important satiation factor, is a compelling candidate in this regard. The objective of this proposal is to identify the mechanisms through which E2 stimulates apoA-IV gene expression in the NTS and to determine how this effect becomes impaired in female rats by chronic high-fat diet (HFD) consumption. The central hypothesis is that E2 normally stimulates apoA-IV gene expression in the NTS and that this effect is impaired by chronic consumption of a HFD, leading to increased food intake and the development of dietary obesity. This hypothesis is based on our preliminary data in HFD-induced obese ovariectomized (OVX) rats. The rationale for the proposed research is that once the particular mechanisms as to how E2 stimulates apoA-IV gene expression and how such effect of E2 is altered by chronic consumption of a HFD are understood, the key component(s) of estrogen signaling could be manipulated pharmacologically, leading to innovative targets to the prevention and treatment of dietary obesity in women. Guided by strong preliminary data, we propose testing this hypothesis by pursuing three specific aims. First, to identify estrogen receptor and DNA response elements that mediate E2's effect on apoA-IV gene expression. Second, to determine whether E2's reduced effect on apoA-IV gene expression contributes to the development of dietary obesity, and whether this can be circumvented by administration of apoA-IV. Third, to determine the mechanism of impaired estrogen signaling in HFD-induced obese female rats. The proposed research is innovative because it translates basic research discovery to a pre-clinical model designed to identify novel treatment targets. This approach represents a substantial departure from the status quo, and is expected to result in an efficacious therapy of obesity in women. This application is significant because it is expected to advance the understanding of how estrogens regulate energy homeostasis in normal female rats, and how such effect is impaired when the animals are fed a HF diet. Ultimately, such knowledge will facilitate the development of preventive and therapeutic interventions to address the epidemic of obesity and, consequently, to improve quality of life for many afflicted individuals and to decrease health care costs in the United States.

DESCRIPTION (provided by applicant): Obesity and type-2 diabetes are national and worldwide epidemics. Since currently available anti-obesity and anti-diabetes drugs have limited efficacy as well as safety concerns, identifying new target compounds, particularly with dual properties in controlling both body weight and blood glucose, is a high priority. Recently, we have identified novel functions of ginsenoside Rb1 (Rb1), the most abundant and biologically active compound in ginseng. Ginseng has been used as a traditional crude medicine in Asian countries to restore and enhance well-being without notable toxic side effects for thousands of years, and is one of the best-selling herbal supplements in the United States. Peripheral administration of Rb1 to rats potently suppresses food intake without eliciting signs of toxicity. Chronic treatment with Rb1 significantly reduces food intake and body weight gain in high-fat diet (HFD)-induced obese rats and also significantly decreases fasting blood glucose and improves impaired glucose tolerance to a greater extent than what occurs in pair-fed controls. These results demonstrate potential novel roles for Rb1 as an anti-obesity and anti-hyperglycemic agent. Our central hypothesis is that Rb1 increases leptin sensitivity to maintain body energy and glucose homeostasis, and it is strongly supported by our preliminary data in lean and HFD-induced obese rats. The rationale for the proposed research is that once the particular mechanisms of Rb1 in the regulation of body weight and blood glucose are understood, the newly acquired information will allow rational design of future pre-clinical studies and clinical trials to develop Rb1 as a novel agent for the prevention and treatment of obesity and diabetes. Guided by strong preliminary data, we propose testing this hypothesis by pursuing three specific aims. First, to determine the interaction of Rb1 and leptin in reducing food intake in rats. Second, to identify the mechanisms through which Rb1 improves energy homeostasis in HFD-induced obese rats. Third, to test the hypothesis that Rb1, like leptin, regulates glucose metabolism by increasing insulin sensitivity at peripheral tissues and/or by increasing insulin secretion. The proposed work is innovative because it assesses novel pharmacological functions of Rb1 in the regulation of energy and glucose homeostasis. Successful completion of the proposed research will enhance our understanding of the mechanisms by which Rb1 prevents and treats obesity and associated symptoms of the metabolic syndrome. Given the continuing epidemics of obesity and diabetes, identifying and understanding a novel pharmacological agent, such as Rb1, with the dual properties of controlling body weight and reducing blood glucose, is expected to have a significant public health impact. PUBLIC HEALTH RELEVANCE: The objective of this project is to elucidate the mechanisms by which the compound Rb1 prevents and treats obesity, diabetes and other symptoms of the metabolic syndrome. Given the growing incidence of obesity and diabetes, identifying and understanding a new pharmacological agent, like Rb1, with dual properties of controlling body weight and reducing blood glucose, is expected to have a significant public health impact.