ABSTRACT Cocaine use disorder (CUD) is a persistent public health concern in the United States, exacerbated by the coronavirus-19 pandemic. Moreover, the only treatment for CUD is behavioral therapy, as there are currently no FDA-approved pharmaceutical therapeutics for CUD. This poses a concern, as individuals experiencing CUD are at substantial risk for relapse across abstinence. A deeper understanding of the cellular and molecular underpinnings of CUD is essential for developing therapeutics and relapse prevention for sustainable abstinence. Although extensive research to date has focused primarily on neuronal mechanisms of cocaine action in the brain, available evidence indicates that neuroglia may significantly contribute to mechanisms of dependence and relapse. The Reissner Lab focuses heavily on understanding the role of astrocytes and microglia in the maintenance of cocaine use disorder and relapse-like behaviors. Published research from the Reissner lab illustrates substantial morphological decreases in the nucleus accumbens core astrocytes following long-access (LgA, 6 h/day, 10 days) cocaine self-administration and a 45-day abstinence period. Preliminary data for this proposal show a significant effect of cocaine and abstinence on the colocalization of astrocyte membranes within microglia, suggesting that microglia may be mediating the effect of cocaine on astrocytes and raising the possibility that microglia phagocytosis is a long-lasting consequence of cocaine use. Thus, this proposal aims to investigate how structural and functional microglial responses in the nucleus accumbens contribute to cocaine dependence and relapse behaviors. In Aim 1, I will explore morphological responses of accumbens microglial, in addition to evidence of engulfment of astrocyte membranes, across abstinence in males and females. In Aim 2, I will test the hypothesis that pharmacological inhibition of microglial phagocytosis will protect against the effects of cocaine on astrocyte structure, microglial engulfment of astrocytes, and cocaine-seeking behavior. A neutrophil inhibitory factor (NIF) peptide which inhibits the protein interaction between complement protein C3 and the microglial C3 receptors will be directly microinjected into the nucleus accumbens, inhibiting phagocytosis. Additionally, this aim will explore the source of phagocytosis signaling by examining “eat me”/“don’t eat me” signals such as C3 colocalization with astrocytes and synapses. The experiments proposed in Aim 3 will characterize the genetic profiles of microglial cells following long-access cocaine self-administration. Through the use of magnetic bead sorting and RT- qPCR, I will be able to identify expression changes of microglia-specific genes. Overall, the findings of this study will provide a foundation for understanding how microglial morphological and functional responses contribute to cocaine dependence in both male and female rats. Further, insights into the role microglia p...