Establishment of Innervation and Sensory Heterogeneity of Trigeminal Dental Neurons

Objectives: While sensory innervation of the mammalian tooth serves a vital role in protecting our dentition from both injury and infection, our knowledge of the signaling mechanisms regulating the onset and patterning of tooth innervation is lacking. Similarly, our understanding of the molecular and functional properties of the neuronal populations within the trigeminal ganglion (TG) that afferently serve the tooth pulp remains rudimentary. In this study, we analyzed the expression and function of GDNF and its receptor, Ret, in the spatiotemporal regulation of tooth innervation. Furthermore, we explored the molecular and functional properties of a novel Ret+ subpopulation of trigeminal dental neurons in the adult. Methods: We explored the expression of Ret in mouse mandibular molars and TG neurons during tooth innervation using tamoxifen-inducible reporters. By crossing a ubiquitously-expressed tamoxifen-inducible Cre (UBC-Cre/ERT2) to Ret conditional knockout mice (Retfx/fx), as well as a highly specific pharmacologic inhibitor of Ret (1NM-PP1), Ret was deleted immediately prior to tooth innervation and the neural projections into P7 molars were analyzed. The same animal models were examined to define the physiological function of Ret+ neurons in the adult. Results: Ret is expressed by a subpopulation of neurons projecting to the tooth pulp at both P7 and in the adult. GDNF is highly expressed by many cell types throughout the dental pulp. Genetic deletion, and pharmacogenetic inhibition of Ret, led to a significant reduction in the neurites present within the pulp, with a corresponding accumulation of aberrant fibers outside the tooth germ. Ret+ TG dental neurons represented a molecularly-unique population of neurons in the adult. Conclusions: Ret has at least two distinct roles in tooth organogenesis and TG biology: a perinatal axon guidance function in regulating target innervation of dental pulpal afferents, and a subsequent maintenance function in specifying a unique population of TG dental neurons.