Neuroinflammation and gut diversity effects due to opioid self-administration

Purpose: Opioids are a great resource to treat pain in humans, but prolonged use can lead users to physical dependence and opioid use disorders (OUD). The use of opioids increases the risk of developing OUDs in humans via activation ofμ-opioid receptors. OUD is extensively studied for its effects in the central nervous system including the brain; however, Gut-Brain Axis (GBA) research will potentially expand our understanding of addiction and provide a new paradigm for developing new substance use disorder (SUD) therapeutics. GBA in animal models of OUDs can elucidate the complex interactions between the brain and gut that lead to pathological drug seeking and consumption and their relation to GBA components (i.e., bacterial populations, gut peptides, and gut signaling). In this study, we will share the determining temporal hallmarks of gut alterations in rats self-administering morphine for 15 days and relate it to neuroinflammatory features in the brain. One of our hypotheses is that abuse of drugs, such as morphine, starts by inducing inflammation of the brain and gut taxonomic changes similar to those observed in human opioid users. Methods: In this project, we used Sprague Dawley (SD) rats, and both the experimental and control groups were surgically implanted with intravenous (IV) catheters. The control group was exposed to the self-administration box, but only received sucrose pellets instead of morphine to avoid differences in behavior due to instrument learning or exposure to self-administration chambers. The experimental group was trained to self-administer morphine dosed to their body weight of 0.4 mg/kg for 14 days. The experimental timeline was (1) baseline: 7 days after catheter implantation, (2) acute: 24 hours after the second day of self-administration, and (3) chronic: 24 hours after completion of the 12 days of self-administration. Fecal samples were acquired at the three-time points and analyzed with 16s DNA sequencing to determine the relative abundance of microbial species at the time points. After the last day, we collected the brains from all animals and prepared tissue FFPE for immunohistochemistry and spatial transcriptomics analysis. Concurrently we acquired MRI diffusion-weighted scans in a 7.0 T preclinical scanner. Rats were restrained under sedation (isoflurane 5% induction, 2% maintenance) reducing the stressor of noise and restriction while scanning. This project will help us identify whether neuroinflammation markers occur due to large doses of morphine repetitively in rodents. Results: Preliminary analysis points out that chronic and voluntary administration of morphine leads to a neuroinflammatory response in the habenula possibly detected with diffusion MRI. Our preliminary16s DNA sequencing analysis has shown some key microbiome differences in our experimental drug group versus our control pellet group. Conclusion: As we continue analyzing our data, we hope to provide more insight on early effects of chronic drug use on gut-brain axis.