ABSTRACT Chronic tic disorders (CTD), including Tourette’s disorder and persistent tic disorders, are a significant source of disability for children and adolescents, yet pharmacological therapies remain highly unsatisfactory due to serious adverse events. An ideal direction to develop safer treatments for CTD would be to target the same biological processes whereby tics spontaneously wax and wane over time; however, the mechanisms underlying these fluctuations remain poorly understood. A novel opportunity to solve this problem may come from recent evidence documenting that tic severity is influenced by the gut microbiota. Understanding whether changes in the composition of the gut microbiota may account for tic exacerbations - and through which molecular mechanisms - may lead to therapeutic breakthroughs in CTD; this issue, however, remains unexplored. The goal of the studies proposed in this R21 application is to explore whether and how variations in the gut microbiota contribute to tic exacerbations. The gut microbiota may influence tic severity through multiple mechanisms, including the synthesis of inflammatory cytokines. Ample correlational evidence points to tumor necrosis factor-α (TNF) as the inflammatory cytokine best associated with tic exacerbations. Thus, to test whether this cytokine increases tic severity, we evaluated its effects in a mouse model of CTD. These preliminary studies showed that low TNF doses that do not elicit sickness behavior significantly increase tic-like stereotypies in these mice. Based on these findings, we hypothesize that, in CTD patients, gut microbiota alterations lead to increased production of TNF or other inflammatory cytokines, which exacerbate tics. To test this hypothesis, the exploratory studies proposed in this R21 application will use a translational platform, combining clinical analyses in CTD patients and mechanistic experiments in mouse models. Specifically: - In Aim 1, we will assess the alterations of gut microbiota associated with CTD and test whether tic exacerbations are associated with alterations of the gut microbiota and inflammatory responses by testing fecal and blood samples from forty CTD patients and forty non-affected controls. - In Aim 2, we will test the causal link between gut microbiota alterations and tic exacerbations by transplanting fecal specimens from CTD-affected individuals into our mouse models of Tourette’s disorder; we will also test whether TNF or other inflammatory cytokines contribute to potential changes in tic severity. The translational studies proposed in this R21 application will be the first systematic analyses of the role of gut microbiota in tic fluctuations. If our hypothesis is verified, future R01-funded studies will test whether fecal material transplant from non-affected individuals and/or probiotic treatments can reduce tic severity. We will also study the downstream mechanisms whereby inflammation increases tic severity. Thus, our results may lead to the development of new, safer, mechanistically based treatments for CTD.

PROJECT NARRATIVE Recent evidence suggests that the course of tic disorders is robustly influenced by the gut microbiota, i.e. the community of bacteria, archaea, and fungi inhabiting the gastrointestinal system. Building on this evidence, we propose clinical studies and experiments in animal models to explore how gut microbiota imbalances increase tic severity.