Project summary/abstract Cocaine addiction is a multidimensional psychiatric disorder with pathophysiology that seems to involve abnormally strong learned associations. These types of learned associations are thought to be encoding within patterns of sparsely distributed neurons, called neuronal ensembles. Relatively little is known about how neuronal ensembles controlling cocaine-seeking differ from those underlying natural reward seeking. We have recently found that a small molecule, Rac1, is differentially expressed in cocaine-paired neuronal ensembles compared to food-paired ensembles. This molecule may represent a unique molecular adaptation that defines the cocaine-seeking ensemble. There is, therefore, a critical need to determine the role or Rac1 in cocaine- seeking ensembles. The long-term goal is to determine the neural mechanisms underlying drug memories to enable development of clinically useful therapies to alleviate craving and relapse of cocaine use disorder. The overall objective in this application is to the role of Rac1 in the behavioral, structural, and activity of cocaine- ensemble neurons. Our central hypothesis is that Rac1 signaling is integral to cocaine-induced neuroadaptations, driving neuroadaptations that increase the cue-reactivity of cocaine ensemble neurons and drive cocaine-seeking behavior. The rationale for the proposed research is that understanding how Rac1 expression effects the neuronal ensembles governing cocaine-seeking behavior will provide new opportunities for developing experimental therapeutics to treating cocaine use disorder. To attain the overall objectives, the following specific aims will be pursued: 1) Determine the impact of IL ensemble Rac1 activity on cocaine-seeking behavior; 2) Identify changes in cellular structure within cocaine-associated neuronal ensembles; and 3) Determine the role of Rac1 activity on cue reactivity of cocaine ensemble neurons. The research proposed in this application is innovative because it dissects the role of Rac1 within neuronal ensembles in cocaine self- administration compared to food-seeking ensembles using several cutting-edge methods. These contributions will have significant impact because they are expected to have determined how Rac1 drives adaptations within the neuronal ensembles mediating cocaine-seeking.