The prevalence of obesity and its associated deleterious health consequences is growing in the United States making it a critical area of research. This research will focus on the endogenous stomach-released 'hunger peptide', ghrelin, that is secreted in direct proportion to the last time food was eaten. Circulating ghrelin is highest in humans with the Prader-Willi Syndrome (PWS), the most common genetic form of life-threatening obesity, who are prodigious food hoarders. Our animal model, the Siberian hamster, is obese (~50% body fat) and hoards food. Exogenous ghrelin, within the physiological range, increases ingestive behaviors in Siberian hamsters to nearly identical levels as food deprivation [36-56 hrs (IACUC approved), as shorter ones do not impact these behaviors due to their ample lipid stores]. The necessity of ghrelin release during food deprivation to stimulate these increases in ingestive behaviors will be tested using a 'mirror-image'compound (a Spiegelmer) designed to bind and render ghrelin inactive. In addition, I will test the role of central-acting ghrelin in promoting obesity by inhibiting brown adipose tissue (BAT) thermogenesis, while simultaneously inhibiting white adipose tissue (WAT) lipid mobilization (lipolysis). Because increases in the sympathetic nervous system (SNS) activity are necessary to increase BAT thermogenesis and WAT lipolysis, I will test how centrally-acting ghrelin promotes obesity by decreasing the SNS drive to BAT/WAT. First, the co-localization of the ghrelin receptor [growth hormone secretagogue receptor-1a (GHS-R1a)] mRNA with the SNS outflow to BAT and to WAT will be tested using the retrograde, transynaptic tract tracer, pseudorabies virus (PRV) and visualized using in situ hybridization and immunohistochemistry, respectively. To test the function of centrally-acting ghrelin, site-specific microinjections of ghrelin will be made into brain loci with high SNS outflow to BAT/WAT + GHS-R1a colocalization and measure BAT temperature, uncoupling protein-1 (the critical factor for BAT thermogenesis) protein, and norepinephrine turnover rate (NETO, a neurochemical measure of SNS drive) and from WAT, and two SNS-induced intracellular markers of lipolysis (phosphorylation of hormone sensitive lipase and perilipin A) by Western blot. Several key new techniques will be learned herein: 1) Western blot protein analysis, 2) HPLC, 3) viral transneuronal tract tracing in combination with 4) in situ hybridization and 5) in vivo BAT thermogenesis methodologies. This fellowship will allow for the participation in a two week extensive hands-on course in molecular biology methods, attendance to international scientific meetings to present data and professional development and scientific seminars, and learning the art of successful grant writing and importance of ethics. Testing the behavioral and neuro/biochemical mechanisms by which ghrelin increases obesity will deepen our understanding of obesity genesis in normal and PWS humans, allowing for more effective designs of intervention at both the food intake and energy expenditure levels.
Prevention and reversal of obesity is difficult and is poorly understood. Obesity arises when food intake is greater than energy expenditure, and understanding the systemic and neural mechanisms leading to food intake and energy expenditure are critical. Here we test the dual role of the stomach 'hunger hormone'ghrelin in causing increases in food seeking, storage, and hoarding and decreases in energy expenditure through brown fat using an animal model that naturally hoards food and exhibits reversal of obesity.
