Identifying the brain network changes produced by alcohol dependence and neuropharmacological interventions to reduce alcohol drinking and relapse in animal models is critical in understanding the consequences of drinking alcohol and identifying the mechanisms responsible for the therapeutic efficacy of current and novel medications. However, the functional connectivity of whole-brain networks during alcohol abstinence is poorly known. Because of technical limitations, visualizing changes throughout the whole brain at single-cell resolution has not been possible until recently. A key evolution of The Scripps Research Institute Alcohol Research Center (TSRI-ARC) is a novel Functional Connectomics Component that evolved from the previous Neurocircuitry Component to identify these functional whole-brain networks using single-cell whole-brain imaging (iDisco+) of immediate early gene (c-fos) expression in mice and rats. During the previous funding cycle, we found that alcohol-dependent mice exhibit hyperactivity of the extended amygdala network and a functional disconnection between the infralimbic cortex (IL) and both the central nucleus of the amygdala (CeA) and the lateral hypothalamus (LH). These changes were associated with widespread increases in coordinated brain activity during alcohol abstinence and decreased whole-brain network modularity. We also identified how two neuropharmacological treatments (corticotropin-releasing factor 1 [CRF1] receptor antagonist and opioid receptor antagonist) reverse specific brain network changes produced by alcohol withdrawal. In this renewal, we will test the hypothesis that FDA-approved medications and TSRI-ARC-related compound candidates for the treatment of alcohol use disorder normalize whole-brain network modularity, promote cortico-subcortical functional connectivity, and reduce the influence of the extended amygdala on the whole-brain network. We will also test the hypothesis that inactivation of the LH®IL circuit and activation of the IL®CeA circuit, two key interventions expected to reduce alcohol seeking, will restore whole-brain modularity and identify the specific subnetwork mechanisms associated with these effects. The use of single-cell whole-brain imaging in animals characterized for addiction-like behaviors is a significant advance for the field as it provides an innovative, unbiased, and comprehensive mapping of the whole-brain network mechanisms associated with behavioral states and interventions relevant to alcohol use disorder.
Specific Aim 1: To test if FDA-approved medications for AUD and ARC-related compounds restore whole-brain network modularity and functional connectivity in dependent animals. We will use iDisco+/Fos whole-brain imaging to test the effect of naltrexone, acamprosate, and the TSRI-ARC compounds suvorexant (a dual orexin antagonist), R121919/CP376395 (CRF1 antagonists), nor-binaltorphimine (a k-opioid receptor antagonist), AT403 (a nociceptin receptor agonist), and 2-PMPA (a GCPII inhibitor) on whole-brain functional connectivity in naive, nondependent, and alcohol-dependent mice (two-bottle choice/chronic intermittent ethanol model). The naive mice will be used to identify each drug's pharmacological “brainprint” for future drug development studies. The nondependent and dependent mice will be used to test the hypothesis that the treatments will restore network modularity and identify the subnetwork mechanisms responsible for their behavioral effects.
Specific Aim 2: To identify the functional neuronal networks recruited by inactivation of the LH®IL and activation of the IL®CeA pathways in dependent animals. We will 1) identify whole-brain networks recruited after chemogenetic inactivation of the LH®IL and activation of the IL®CeA pathways in dependent animals and 2) test if inactivation of the LH®IL and activation of the IL®CeA pathways decreases whole-brain modularity and restores cortico-subcortical functional connectivity. This information will be critical to understanding the role of the LH®IL and IL®CeA pathways in addiction-like behavior and neuroadaptations of the whole-brain networks in alcohol-dependent animals. It will also be critical for the TSRI-ARC components to understand better the consequences of their circuit-specific manipulations on whole-brain network function.
Outcomes: This component will produce comprehensive single-cell whole-brain functional connectomics atlases of the effect of alcohol abstinence in males and females, identify the network mechanisms responsible for the behavioral effects of FDA-approved medications and experimental treatments for alcohol use disorder, and identify the network mechanisms associated with modulation of key cortico-subcortical circuits identified by other TSRI-ARC Components.
Integration with The Other Components: This Component stands on its own as its success does not depend on the other components; however, it is fully integrated with the other TSRI-ARC components as it will test key hypotheses generated by the Martin-Fardon, Contet, Zorrilla, and Roberto Components. Moreover, our results will provide considerable insights into the consequences of the pharmacological and biological interventions used by the other components to enrich the interpretation of their results.