Vaginal microbiome-nociceptor interactions in endometriosis pain

Project Abstract Pain is one of the two cardinal symptoms of endometriosis, a gynecological disease that is diagnostically defined by the presence of endometrium-like tissue outside of the uterus. Despite symptom prevalence, the cellular and molecular basis of endometriosis pain is poorly understood. Removal of ectopic endometrial lesions and/or hysterectomy provide pain relief in some, but not all patients suggesting that there must be other factors contributing to endometriosis pain. Our preliminary data clearly demonstrate that the vaginal microbiome contributes to endometriosis pain. Using a mouse model of endometriosis, intravaginal antibiotic treatment, and vaginal microbiota transplant (VMT) paradigms, we determined that the endometriosis vaginal microbiota is both sufficient and required for pain and nociceptor sensitization in this model. Intra-vaginal administration of probiotic bacteria also decreased endometriosis pain and sensitization of uterine-innervating neurons. Thus, in this project we will now explore the bi-directional relationship between the microbiome and uterine-innervating nociceptors in more detail, specifically determining the contributions of calcitonin gene related peptide (CGRP) signaling. Given that nearly all vaginal- and uterine-innervating afferents are CGRP+, we hypothesize that introduction of the endometriosis-associated vaginal microbiome increases the activity of and subsequent release of CGRP from uterine-innervating afferents; this CGRP release then puts additional selective pressures on the vaginal microbiome, thus perpetuating endometriosis pain. To test this hypothesis, we will pursue three aims. In Aim 1, we will use chemogenetics and behavior testing to first determine if activity in CGRP+ uterine-innervating afferents is both sufficient to model endometriosis pain-like behaviors and necessary for endometriosis and endometriosis VMT pain. In Aim 2, we will determine if the endometriosis vaginal microbiome, in the absence of other disease pathology, is sufficient to induce the nociceptor sprouting and sensitization that is observed in this disease. Note that these experiments will be performed in parallel using mouse and human dorsal root ganglia tissue and host-specific vaginal bacterial communities. Finally, in Aim 3 we will determine if neuronal CGRP signaling is the primary factor that drives endometriosis-associated shifts in the vaginal microbiome. These experiments will again be performed in parallel using mouse and human vaginal microbiome libraries. In summary, the experiments outlined in this proposal will provide foundational insight into host-microbiome signaling in the female reproductive tract, and furthermore, lay the groundwork for personalized microbial therapies for endometriosis pain.