The overall goal of this K99/R00 career development award proposal is to integrate innovate chemical probes, including in vivo-active small-molecule inhibitors and specialized chemoproteomic assays, with well- established genetic knockout and behavioral mouse models to obtain a global view of the pathophysiological consequence of disrupting 2-arachidonoylglycerol (2-AG) biosynthesis in mouse models of obesity and inflammation. The endogenous cannabinoid (endocannabinoid) 2-AG is a lipid transmitter that activates the G- protein-coupled receptors CB1 and CB2, which are also targets for the psychoactive ingredient in marijuana, 9-tetrahydrocannabinol. The importance of the endocannabinoid system in obesity-associated metabolic disorders is highlighted by the clinical activity of CB1 antagonists as anti-obesity and anti-diabetic drugs. Complementary studies in preclinical models have since shown that many of the beneficial effects are due to blocking CB1 receptors at peripheral sites. These studies highlight the need for a deeper understanding of the central and/or peripheral contributions of the endocannabinoid system in regulating energy homeostasis as well as obesity-related disease states. 2-AG biosynthesis in vivo is differentially regulated by two sequence- related enzymes, diacylglycerol lipase-alpha and beta (DAGL alpha and DAGL beta, respectively), providing an experimental (and, eventually translational) opportunity to uncouple central and peripheral endocannabinoid signaling through selective inactivation of DAGL isoforms. This proposal will test the hypothesis that DAGL alpha and DAGL beta biosynthesize 2-AG involved in energy homeostasis and perform complementary functions in energy balance through regulation of CB1-dependent signaling at anatomically-distinct sites.
The specific aims are to 1) measure energy homeostasis parameters and body mass composition in vivo in DAGL-disrupted mice 2) measure adipose tissue macrophage accumulation in DAGL-disrupted mice and 3) Determine the physiological effects of inactivating DAGLs in the development and progression of metabolic syndrome. Our preliminary studies show that DAGL alpha display a lean phenotype despite increased food intake, providing evidence of altered energy balance upon central and/or peripheral disruption of 2-AG biosynthesis. Our preliminary studies also show the feasibility of using DAGL-selective inhibitors to uncouple central and peripheral 2-AG signaling pathways in vivo. The candidate aims to combine previous training in chemoproteomics, synthetic chemistry, and quantitative proteomics and metabolomics with new mentored training in mouse models of metabolism and inflammation including non-invasive measurements of metabolic parameters governing energy homeostasis and body mass composition as well as in vivo tracking of adipose tissue macrophage accumulation using adoptive cell transfer and flow cytometry.
Obesity and associated metabolic disorders, including type 2 diabetes have been associated with increased signaling activity of the principal endogenous cannabinoid lipid, 2-arachidonoylglycerol (2-AG). The importance of the endogenous cannabinoid system in obesity-associated metabolic disorders is highlighted by the clinical activity of cannabinoid receptor type 1 (CB1) antagonists as anti-obesity and anti-diabetic drugs. The goal of this proposal is to determine the tissue-specific functions of 2-AG biosynthetic pathways in energy balance and test whether blockade of 2-AG production in vivo can produce therapeutic effects akin to systemic CB1 antagonists in mouse models of obesity.
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