Project Summary Opioid use disorder (OUD) is a debilitating and life-long addictive disease with significant public health costs. OUD is characterized by various symptoms, including a compulsive need to use, loss of control over use, and a negative emotional state induced by withdrawal. Mu-opioid receptors (MORs) in the ventral tegmental area (VTA) are known to contribute to both the rewarding effects of opioid use and the aversive signs and symptoms of opioid withdrawal. However, there is growing evidence that additional populations of MOR, which is broadly expressed throughout emotional and motivational brain circuits, critically contribute to opioid-related behaviors. Striatal cholinergic interneurons (CINs) regulate dopamine release and responding for rewards. Given these important functions of CINs, and because this neuronal subpopulation expresses MOR, we hypothesize that MOR stimulation alters CIN activity and striatal dopamine release to promote opioid reward, consumption, and withdrawal-induced aversion. Our experiments will test this hypothesis through completion of two primary specific aims. We will determine 1) whether MORs on cholinergic interneurons participate in opioid reward and withdrawal and 2) whether CIN MORs regulate dopamine release during opioid reward and withdrawal. Opioid reward will be tested using a conditioned place preference (CPP) task and withdrawal using conditioned place aversion (CPA). Opioid consumption will be assessed with a limited access, two-bottle choice fentanyl drinking task developed in our lab. Collectively, completion of these aims will demonstrate that MORs on NAc CINs contribute to opioid reward, consumption, and withdrawal through modulation of DA release. Furthermore, we will train undergraduate students in a broad array of modern behavioral neuroscience techniques, such as chemogenetics, fluorescent in situ hybridization, and fluorophotometry. These activities will expand access to meaningful research opportunities to a diverse set of talented undergraduate students interested in neuroscience and other biomedical research fields at Miami University.