ABSTRACT The next goals of INIA-Neuroimmune (INIA-N) emphasize the neuroimmune mechanisms that underlie excessive alcohol drinking behaviors and include the expansion of genomic studies across species, a genomic-based computational approach for drug repurposing, and the development of systems-level approaches. The present proposal falls into the latter goal and also integrates translational and drug- repositioning components. Our past funding period within INIA-West used behavioral analyses of mouse genetic models to show that mu and delta opioid receptors regulate alcohol drinking through reward-based and anxiety-related mechanisms, respectively, and allowed positioning the orphan GPR88 receptor as a novel target to reduce alcohol drinking. We also successfully developed resting-state functional magnetic resonance imaging in living mice that lacked the mu opioid receptor gene. This study revealed the first ever-reported gene-specific functional connectivity signature in the brain in living mice and demonstrated remarkable reshaping of reward/aversion networks. Here, we capitalize on both new knowledge from INIA-N on neuroimmune gene function and our cutting-edge live neuroimaging approach to decipher neural network mechanisms that underlie alcohol drinking behaviors at whole-brain level. Our working hypothesis is that connectivity maps that combine gene alcohol and drug alcohol interactions and the integrated analysis of gene/network/behavior datasets will deliver a deep understanding of biological mechanisms that underlie alcohol drinking and biomarkers with translatable potential for patient stratification and therapy. To test this, we will combine behavioral and neuroimaging analyses of mice undergoing a history of voluntary or forced alcohol exposure. In all three Specific Aims, we will apply a longitudinal design, in which functional/structural images will be acquired before and after a history of voluntary excessive alcohol drinking or upon a forced alcohol diet. A third set of images will be acquired after a 4-week protracted abstinence period, together with behavioral measures of emotional deficits. Specific Aim 1 will set reference datasets in C57/Bl6 mice. Specific Aim 2 will address mu and delta opioid receptors that are responsible for high (mu/reward/nalmefene) and low (delta/anxiety/ARM390) alcohol drinking. Specific Aim 3 will investigate new mechanisms that underlie alcohol-drinking behaviors, including a novel druggable target (GPR88/motivation) and neuroinflammation (PDE4 inhibitors). The latter aim is most novel because neural networks that are involved in the alcohol-induced neuroimmune response are virtually unknown. This proposal follows the natural progression of INIA-N goals toward the clinic. All of the Specific Aims have demonstrated feasibility and fully synergize with several INIA-N investigators, including neuroimaging and drug selection for the clinic. Finally, in addition to alcohol research for INIA-N, the proposal overall will contribute to two growing fields of neuroscience: the development of connectome genetics and translational psychiatry.