Project Summary/Abstract The overall goal of this proposal is to understand how diacylglycerol lipase beta (DAGLβ) attenuates neuroinflammatory signaling and consequential pain. In response to tissue damage, proinflammatory signals (lipids or cytokines) are released by injured cells and initiate cell-based adaptative and innate immunity mechanisms to protect the host. The activation of these innate immune cells results in a series of immunomodulatory cascades that triggers additional inflammation and the stimulation of nearby nerves, eliciting pain. One of these cascades involves the production of proinflammatory eicosanoid (i.e. prostaglandin) lipids while another cascade centers around the transcription of proinflammatory genes key to cytokine expression. Current pharmacological interventions either work at the site of trauma (i.e. non-steroidal anti-inflammatory drugs, NSAIDs) or in the transmission of resulting pain responses (i.e. opioids). However, none of these therapies provide efficacious long-term treatment as the eventual drug tolerance that builds elicits adverse side effects ranging from chronic disease to addiction. What is desperately needed is a therapy that displays efficacy in not only thwarting inflammatory signaling, but also relieving pain without consequence over long periods of exposure. DAGLβ is an enzyme that directly attenuates eicosanoid production through the hydrolysis of diacylglycerol lipids upstream of the prostaglandin synthetic pathway. Inhibition of DAGLβ has been found to be efficacious in promoting antinociception sourced through either inflammatory (acute) or neuropathic (chronic) pain models. Moreover, long-term DAGLβ inhibitor exposure does not produce characteristic metabolic, behavioral or addictive side effects. A critical gap in our understanding is how DAGLβ attenuates cytokine production as evidence strongly suggests that its role in eicosanoid production does not explain its pain-relieving effects in chronic pain as NSAIDs, which also target eicosanoid production, are often ineffective in these same chronic models. Here we describe a novel connection between DAGLβ activity and kinase-based signaling that is known to regulate proinflammatory cytokine gene transcription. We seek to identify the mechanisms by which DAGLβ regulates neuroinflammatory signaling for the eventual translation of DAGLβ small molecule inhibitors into efficacious therapies for chronic pain alleviation. In this proposal, our first aim is the determine the mechanisms through which DAGLβ activity modulates a kinase-based signaling network, separate from the canonical prostaglandin lipid production pathway. Our second aim is to test the applicability of DAGLβ inhibitors in suppressing nociceptive behaviors displayed by chronically inflamed mice through repeated exposure. Successful completion of this project will broaden our understanding of lipid signaling involved in neuroinflammation, identify novel cross-talk between distinct me...