Obesity and type 2 diabetes share several metabolic abnormalities, including an impaired ability of cells to sense increased nutrients flux. Since the brain plays a crucial role in regulating the homeostasis of fuel metabolism and body weight, the ultimate goal of this proposal is to understand the neuronal mechanisms of nutrient sensing. Malonyl CoA, a product of glucose metabolism, has been implicated as a major gauge of fuel metabolism in hypothalamic neurons that monitor energy balance. It is postulated that in the fed state malonyl CoA acts as a sensor of fuel abundance because of its potent allosteric inhibition of the enzyme carnitine palmitoyltransferase 1 (CPT1), which controls the entry of fatty acids into mitochondria where they undergo oxidation. Increased levels of long-chain fatty acyl-Co-enzyme A (LCFA-CoAs) in hypothalamus lead, in turn, to decreased food intake and decreased endogenous glucose production. To test whether the malonyl CoA levels in hypothalamus regulate feeding behavior and metabolism via modulation of fatty acid oxidation, we propose to express in hypothalamus of adult rats a CPT1 gene (CPT1AE3A) that is insensitive to the allosteric inhibition of malonyl CoA, while it retains its full catalytic activity. In order to avoid high constitutive, and therefore supraphysiologic levels of hypothalamic CPT1AE3A expression, we propose in aim 1 to generate gutless adenoviral (gAd) vectors containing a rapalog-inducible expression cassette plus all the transcription factor components, in a single particle. In the second aim we will determine the optimal adenoviral vector and rapalog doses to express CPT1AE3A in hypothalamic neurons in vivo. Finally, in the third aim we will assess whether hypothalamic expression of a malonyl CoA-insensitive CPT1 alters feeding behavior and glucose metabolism. The use of inducible gAd vectors in transduction of hypothalamic neurons will be an invaluable tool for the study of metabolism, since to date no suitable gene expression system has been developed that allows controlled expression of transgenes at moderate levels. Additionally, the application of this technology to the study of the malonyl CoA sensing pathway in the hypothalamus will shed new light in the neuronal mechanisms that couple metabolic nutrient sensing to signal transduction and neurotransmission.
STATEMENT: The prevalence of obesity in affluent societies has reached epidemic proportions, heightening the concern for the health risks associated with obesity, which include type 2 diabetes, hypertension and atherosclerosis. The increased awareness of the health risks associated with obesity has increased the interest in understanding the pathogenesis of obesity. The ultimate goal of this proposal is to delineate novel molecular mechanisms controlling body weight and metabolism and to identify specific molecular targets for the treatment of obesity and diabetes.
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