The three groups of specific aims of this project are intended to determine how metabolic signals, particularly insulin and adrenal glucocorticoid hormones, contribute to the normal regulation of body energy balance and adiposity. The first of these aims proposes to quantify insulin uptake into the CNS and its regulation in the dog. The effects of hormones, food deprivation, and overfeeding on this uptake will be evaluated and compared with the changes in food intake during refeeding. We hypothesize that regulated insulin uptake is a critical contributor to energy balance and is predictive of the response to dietary intervention. The second group of aims will test the hypothesis that insulin and glucocorticoids interact in the brain to regulate energy balance by controlling the biosynthesis and release of neuropeptide Y, a critical neuroregulator of food intake synthesized in the hypothalamic arcuate nucleus and released into the paraventricular nucleus. We also propose to determine how abnormal regulation of this system participates in experimental obesity in the Zucker fa/fa rat. The role of insulin and glucocorticoids in the regulation of NPY mRNA expression will be studied in the rat brain during caloric restriction and refeeding hyperphagia using in situ hybridization. The importance of NPY to food intake will also be assessed by infusion of an NPY antagonist into the cerebral ventricle of normal and genetically obese rats. The third group of aims proposes to assess metabolic control of NPY gene transcription in vivo in phenotypically normal mice heterozygous for a targeted mutation (""""""""knockout"""""""") of the NPY gene and in cell culture systems. The goal is to determine whether increased hypothalamic NPY gene expression induced by food deprivation in vivo reflects increased transcription of the NPY gene. This will be accomplished using mice expressing an NPY promoter-reporter gene construct. Additional studies in transgenic mice expressing a full-length mouse NPY gene are proposed to determine the biologic effect of excessive arcuate nucleus NPY gene expression on food intake and body weight. Cell culture systems will then be used to evaluate the site of hormone action on the NPY promoter and/or the effect of hormones on NPY mRNA stability. Ultimately, we wish to understand the insulin/glucocorticoid/NPY body weight regulatory system and determine how this system might be modulated to influence the development of obesity.
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