Project Summary/Abstract Since teeth are exposed to environmental stimuli, tooth innervation is crucial to their lifelong protection and us- age. The tooth is primarily innervated with sensory nerve fibers from the trigeminal ganglion (TG) that help pro- tect the tooth organ by relaying noxious stimuli. During postnatal development, the dental pulp (DP) secretes neurotrophic factors to guide axonal penetration and sprouting within the tooth in a highly regulated manner. The transforming growth factor β (TGFβ) superfamily regulates many developmental processes in teeth, in- cluding mineralization and innervation. A secreted phosphoprotein downstream of TGFβ signals, osteopontin (OPN), is a major extracellular protein that regulates tooth mineralization and also promotes neurite outgrowth. Following tooth injury, DP cells release neurotrophic factors that modulate neurite growth and dentin repair. Interestingly, OPN-/- mice do not form reparative dentin during pulpal healing, suggesting that OPN plays a cru- cial role in this process. The long-term goal of this project is to understand the cellular mechanisms that protect teeth via sensory innervation. The overall objective is to determine the role of signals from the dental mesen- chyme that regulate tooth innervation. Our central hypothesis is that Tgfbr2 in the dental mesenchyme governs paracrine signaling via OPN to guide tooth sensory innervation. Our laboratory has established a mouse model in which Tgfbr2 is conditionally deleted in odontoblast-producing mesenchyme using an Osterix promoter- driven Cre recombinase (Tgfbr2cko). These mice survive postnatally, but have significant defects in their bones and teeth. We performed a series of experiments to determine whether Tgfbr2 signaling in the DP mesen- chyme regulates their differentiation and ability to guide neurite outgrowth during tooth development. We found that OPN-/- mice do not demonstrate reduced innervation at P7 as originally hypothesized, but do exhibit dis- rupted axonal targeting. The role of neurite outgrowth in regulating reactionary and reparative dentine when the mineralization capacity of the DP cells is compromised remains unknown. Guided by these data, we will test our hypothesis with the following two specific aims: 1) To analyze the OPN-governed signaling cascades in the dental mesenchyme regulating tooth sensory innervation during development; 2) To establish the degree to which the neurite outgrowth associated with dentin repair is regulated by Tgfbr2 and/or OPN signaling in the DP mesenchyme. Under the first aim, we will perform an mRNA Sequence Analysis of control and OPN-/- DP tissue to determine what signals could be disrupting the afferent organization. In the second aim, we will use an in vivo dental injury model and investigate neuronal regeneration in Tgfbr2cko, OPN-/-, and WT mice. The proposed research is significant because it is expected to advance and expand the understanding of how DP cells protect the ...