Project summary Substance use disorder is a public health concern with no effective, long-lasting pharmacotherapies. A large amount of effort has been put into defining the neural basis of addictive behaviors as well as the factors that lead to the development of drug addiction. Past research has focused on the role of entire brain regions in mediating addiction and recently, with advances in genetic approaches, genetically defined cellular populations. However, recent work has shown that even within these defined populations, only a small percentage of cells are activated to a given stimulus. Thus, moving forward, it will be important to identify the role of functionally defined neural populations – termed ensembles – in addictive behaviors. Here, we aim to understand how ensembles in the nucleus accumbens (NAc) that are activated by cocaine encode information and drive volitional cocaine consumption. In this proposal, we will combine translational addiction models (mouse self-administration) with optical tools for recording (fiber photometry) and manipulating (optogenetics) functionally defined ensembles to elucidate what information is encoded within them and how they control complex behaviors. Together, this proposal will give me the technical and theoretical training to answer complex questions about the neural control of behavior and its dysregulation in substance use disorder. In Aim 1, I will define what aspects of cocaine- associated behaviors these cells encode by using fiber photometry to record from cocaine-activated ensembles in vivo while animals self-administer cocaine. In Aim 2, I will determine if these ensembles are sufficient to drive self-administration by allowing mice to perform optical self-stimulation tasks in ChR2-tagged cocaine-activated ensembles. Finally, in Aim 3, we will define how temporally specific activation or inhibition of these cells during specific stimuli alters self-administration. By photo-stimulating or -inhibiting these cocaine ensembles during self- administration, we can define the sufficiency and necessity of these neural populations in various aspects of drug-taking (acquisition, consumption, and drug seeking/relapse). These experiments will provide new information about the cell-specific populations that drive both initial drug taking as well as the behaviors that define substance use disorder. By focusing on functionally defined neuronal populations, we will discover subpopulations that would be unobservable when applying a priori broad genetic markers. Through these novel findings, we will learn more about the mechanisms that mediate progression to drug addiction in specific subpopulations so that we will be able, in the future, to decrease cocaine-taking behavior without producing addictive or aversive effects.