*3.2. PIEZO Channels*

Since PIEZO family ion channels were cloned in mammals, the PIEZO gene family have been considered as putative mechanosensitive ion channel proteins [108–114]. PIEZO1 and PIEZO2 were identified by efforts to elucidate mammalian mechanosensing mechanisms which could not be clearly understood by TRP channels. While PIEZO channels are broadly expressed in a wide range of mammalian mechanosensitive cell types, PIEZO2 channels are identified as low-threshold mechanoreceptors in sensory DRG neurons and Merkel cells [115–117]. Moreover, the depletion of PIEZO2 in sensory DRG neurons and Merkel cells resulted in the dramatic reduction of rapidly adapting mechanically induced currents, suggesting a critical role of PIEZO2 as low threshold mechanoreceptor [118]. These findings have great implications for tooth pain research because low threshold mechanoreceptors are regarded as major players in tooth pain sensory systems, considering that mild mechanical stimuli could cause severe tooth pain. Moreover, several studies revealed that major populations of dental primary afferent neurons consist of A-fibers regarded as low-threshold mechanoreceptors [2,53]. Recently, many groups have examined the functional expression of PIEZO2 in the dental sensory system. Won et al. demonstrated PIEZO2 expression in murine dental primary afferent neurons by single-cell RT-PCR and in situ hybridization and function by recording rapidly adapting inward current induced by direct pocking [119]. Interestingly, PIEZO2 positive dental primary afferent neurons were medium-to-large sized and co-expressed with TRPV1, Nav1.8, and CGRP, which are regarded as nociceptive neuronal marker genes. These results indicate that PIEZO2 positive low-threshold mechanoreceptor neurons innervating teeth are 'algoneurons' that also, paradoxically, act as nociceptors.

To verify the role of these low-threshold mechanoreceptors in the odontoblast cells, other studies have been performed to verify PIEZO expression in odontoblast cells. An electrophysiological study with odontoblast cells co-cultured with IB4-negative medium-sized TG neurons elucidated the role of odontoblasts as mechanosensitive transducer cells [120]. Inward currents were detected from TG neurons when mechanical stimulation was applied to neighboring odontoblast cells. Interestingly, this odontoblast-induced inward current from TG neurons was antagonized with a PIEZO1 selective blocker. Three-dimensional imaging with focused ion beam-scanning electron microscopy revealed that PIEZO2 is expressed in nearly all rodent matured odontoblast cells and is absent in immature cells [121]. PIEZO2 proteins were detected selectively in odontoblastic processes that protrude into dentinal tubules. In another study, however, odontoblastic response to mechanical stimulation was inhibited by a specific antagonist of PIEZO1 [120]. These controversial results indicate the essential

role of PIEZO ion channels in dental sensory systems as putative mechanosensors but also suggest that more research is needed to comprehensively understand complex dental mechanosensing systems.
