Teeth are composed of many tissues, covered by an inflexible and obdurate enamel. Unlike most other tissues, teeth become extremely cold sensitive when inflamed. The mechanisms of this cold sensation are not understood. Here, we clarify the molecular and cellular components of the dental cold sensing system and show that sensory transduction of cold stimuli in teeth requires odontoblasts. TRPC5 is a cold sensor in healthy teeth and, with TRPA1, is sufficient for cold sensing. The odontoblast appears as the direct site of TRPC5 cold transduction and provides a mechanism for prolonged cold sensing via TRPC5’s relative sensitivity to intracellular calcium and lack of desensitization. Our data provide concrete functional evidence that equipping odontoblasts with the cold-sensor TRPC5 expands traditional odontoblast functions and renders it a previously unknown integral cellular component of the dental cold sensing system.

The spatial and temporal distribution of receptors constitutes an important mechanism for controlling the magnitude of cellular responses. Several members of the transient receptor potential (TRP) ion channel family can regulate their function by modulating their expression at the plasma membrane (PM) through rapid vesicular translocation and fusion. The mechanisms underlying this regulation are not completely understood, and the contribution of vesicular trafficking to physiological function is unknown. TRPM8 receptors are expressed in mammalian peripheral sensory neurons and are essential for the detection of cold temperatures. Previously, we showed that TRPM8-containing vesicles are segregated into three main pools, immobile at the PM, simple diffusive and corralled-hopping. Here, we show that channel expression at the PM is modulated by TRPM8 agonists in F11 and HEK293T cells. Our results support a model in which the activation of TRPM8 channels, located at the PM, induces a short-lived recruitment of a TRPM8-containing vesicular pool to the cell surface causing a transitory increase in the number of functional channels, affecting intrinsic properties of cold receptor responses. We further demonstrate the requirement of intact vesicular trafficking to support sustained cold responses in the skin of mice.

BackgroundThe purpose of this study is to determine the effectiveness of warming anesthesia on the control of the pain produced during the administration of dental anesthesia injection and to analyze the role of Transient Receptor Potential Vanilloid-1 nociceptor channels in this effect.Patients and methodsA double-blind, split-mouth randomized clinical trial was designed. Seventy-two volunteer students (22.1±2.45 years old; 51 men) from the School of Dentistry at the Universidad Austral de Chile (Valdivia, Chile) participated. They were each administered 0.9 mL of lidocaine HCl 2% with epinephrine 1:100,000 (Alphacaine®) using two injections in the buccal vestibule at the level of the upper lateral incisor teeth. Anesthesia was administered in a hemiarch at 42°C (107.6°F) and after 1 week, anesthesia was administered by randomized sequence on the contralateral side at room temperature (21°C–69.8°F) at a standardized speed. The intensity of pain perceived during the injection was compared using a 100 mm visual analog scale (VAS; Wilcoxon test p<0.05).ResultsThe use of anesthesia at room temperature produced an average VAS for pain of 35.3±16.71 mm and anesthesia at 42°C produced VAS for pain of 15±14.67 mm (p<0.001).ConclusionThe use of anesthesia at 42°C significantly reduced the pain during the injection of anesthesia compared to its use at room temperature during maxillary injections. The physiological mechanism of the temperature on pain reduction could be due to a synergic action on the permeabilization of the Transient Receptor Potential Vanilloid-1 channels, allowing the passage of anesthetic inside the nociceptors.

Retrograde fluorescent labeling of dental primary afferent neurons (DPANs) has been described in rats through crystalline fluorescent DiI, while in the mouse, this technique was achieved with only Fluoro-Gold, a neurotoxic fluorescent dye with membrane penetration characteristics superior to the carbocyanine dyes. We reevaluated this technique in the rat with the aim to transfer it to the mouse because comprehensive physiologic studies require access to the mouse as a model organism. Using conventional immunohistochemistry, we assessed in rats and mice the speed of axonal dye transport from the application site to the trigeminal ganglion, the numbers of stained DPANs, and the fluorescence intensity via 1) conventional crystalline DiI and 2) a novel DiI formulation with improved penetration properties and staining efficiency. A 3-dimensional reconstruction of an entire trigeminal ganglion with 2-photon laser scanning fluorescence microscopy permitted visualization of DPANs in all 3 divisions of the trigeminal nerve. We quantified DPANs in mice expressing the farnesylated enhanced green fluorescent protein (EGFPf) from the transient receptor potential cation channel subfamily M member 8 (TRPM8) locus in the 3 branches. We also evaluated the viability of the labeled DPANs in dissociated trigeminal ganglion cultures using calcium microfluorometry, and we assessed the sensitivity to capsaicin, an agonist of the TRPV1 receptor. Reproducible DiI labeling of DPANs in the mouse is an important tool 1) to investigate the molecular and functional specialization of DPANs within the trigeminal nociceptive system and 2) to recognize exclusive molecular characteristics that differentiate nociception in the trigeminal system from that in the somatic system. A versatile tool to enhance our understanding of the molecular composition and characteristics of DPANs will be essential for the development of mechanism-based therapeutic approaches for dentine hypersensitivity and inflammatory tooth pain.