Although the viscoelastic properties of the bladder wall depend on it, and large numbers of patients suffer from diseases that give rise to stiff, fibrotic bladders (e.g., neurogenic bladder, outlet obstruction, underactive bladder, radiation cystitis), we know little about the bladder's connective tissue including its constituent fibroblasts. We recently reported that the PDGFRA (platelet derived growth factor alpha)-expressing “interstitial cells” of the mouse bladder are fibroblasts, and that multiple, regionally distinct populations of these cells are found in the bladder wall. One of these populations, the suburothelial fibroblasts (SUFs), reside in the lamina propria just underneath the urothelium. They have a classical fibroblast-like ultrastructure, and they can be differentiated from other bladder fibroblasts by the selective expression of ACTA2 (smooth muscle actin), MYH9 (non-muscle myosin IIA), and MYH10 (non-muscle myosin IIB), along with several other canonical and universal fibroblast markers. Strikingly, we observe that SUFs are contractile, and that diphtheria toxin (DT)-mediated ablation of them leads to an underactive bladder phenotype. Our studies will test the hypothesis that SUF-mediated contraction of the lamina-propria associated collagen matrix promotes rugae re-formation after voiding, and that afferent nerve processes distributed within and attached to the lamina propria collagen become activated as the collagen fibers in the rugae unfurl, relaying information about the state of bladder filling to the CNS. In Subaim 1.1 we will use conditional SUF Piezo1 knockout (KO) mice to determine whether SUF-mediated rugae formation is dependent on the mechanosensitive ion channel PIEZO1. In Subaim 1.2 we will use conditional SUF Itgb1 KO mice to determine if SUF- collagen interactions are necessary for the post voiding formation of rugae. In Subaim 2.1 we will assess whether SUFs are integral to mucosal contraction, collagen organization, and bladder function in SUF-DT ablated mice or conditional SUF Piezo1 KO and Itgb1 KO mice. In Subaim 2.2, we will determine the contribution that mucosal unfolding makes to bladder afferent outflow using SUF ablation and conditional deletion of Itgb1 in SUFs and sensory neurons. Impact: Upon completion of our studies, we will have defined the molecular machinery that allows SUFs to refold the mucosa into rugae when the bladder empties and established a novel role for rugae in regulating bladder afferent signaling and function as the bladder fills.

The inner lining of the urinary bladder is covered with mucosal rugae that unfurl during bladder filling, but how they re-form after voiding and their purpose is not well understood. Our studies will define how contraction of a specialized population of suburothelial fibroblasts regulates rugae formation, and then determine how unfolding of these rugae contributes to afferent nerve signaling and bladder function.