Morphology of P2X3‐immunoreactive nerve endings in the rat tracheal mucosa
Airway sensory receptors are in involved in respiratory modulation and defense against noxious stimuli in various animals including rats (Sant'Ambrogio & Widdicombe, 2001; Widdicombe, 2003). Sensory nerve endings in the tracheobronchial tree have been classified into several types based on morphological characteristics; that is, smooth muscle‐associated nerve endings, intraepithelial free nerve endings, and nerve endings in neuroepithelial bodies (review see; Lee & Yu, 2014; Widdicombe, 2001). Smooth muscle‐associated nerve endings are plant‐like in shape, located in the smooth muscle layers of the trachea and bronchi of various animal species, and identified by silver impregnation (Larsell, 1922; Larsell & Dow, 1933; Fisher, 1964), the reconstruction of electron micrographs (Düring, Andres, & Iravani, 1974; Krauhs, 1984), and immunohistochemistry (Lembrechts et al., 2011; Yamamoto, Atoji, Kuramoto, & Suzuki, 1998; Yamamoto, Atoji, & Suzuki, 1995; Yamamoto, Hayashi, Atoji, & Suzuki, 1994; Yu, Wang, & Zhang, 2003). Yu et al. (2003) identified these structures using a combination of electrophysiology with a field mechanical stimulation and immunohistochemistry for Na+‐K+‐ATPase, the α3‐subunit, and concluded that these endings are slowly adapting receptors (SARs) activated by lung inflation. On the other hand, intraepithelial free nerve endings have been observed using immunohistochemistry for substance P and/or the calcitonin gene‐related peptide (CGRP; Baluk, Nadel, & McDonald, 1992; Cadieux et al., 1986; Springall et al., 1987). Intraepithelial nerve endings also express capsaicin‐sensitive transient receptor potential vanilloid 1 (TRPV1) channels (Watanabe et al., 2006; Watanabe et al., 2005; Yamamoto, Sato, & Taniguchi, 2007), which suggests that they are unmyelinated C‐fiber nociceptors sensitive to noxious stimuli. Furthermore, other types of sensory receptors have been classified by electrophysiological properties; rapidly adapting receptors (RARs) evoked by mechanical stretch by lung deflation and irritant stimuli, for example, touch, cigarette smoke, and hypertonic solution (Bergren & Peterson, 1993; Davies, Pirie, & Eyre‐Todd, 1996; Ho, Gu, Lin, & Lee, 2001; Tsubone, 1986). Although RAR is thought to exist in the deeper region of submucosal layer (Mortola, Sant'ambrogio, & Clement, 1975; Undem, Carr, & Kollarik, 2002), the morphological characteristics of the RARs are unknown.
Extracellular adenosine 3′‐triphosphate (ATP) is one of the important excitatory neurotransmitters/neuromodulators in the peripheral nervous system (Burnstock, 2000; Nakatsuka & Gu, 2006). In the sensory system, ATP is bind to ionotrophic P2X purinoreceptors to activate the sensory nerves (Burnstock, 2000). Functionally, ATP plays a role in mechanosensory transduction by P2X3 homomeric and/or P2X2/P2X3 heteromeric receptors, in various organs including urinary bladder, ureter, esophagus, and stomach (review see, Burnstock, 2009). On the other hand, in the taste buds, P2X2 and/or P2X3 receptors on taste fibers are activated by ATP released by type II taste cells when taste stimuli are received (review see, Kinnamon & Finger, 2013). Immunohistochemical studies revealed that P2X2 and/or P2X3 subtypes are expressed in the sensory neurons of the dorsal root ganglion, trigeminal ganglion, and nodose ganglion of the rat using immunohistochemistry (Hubscher, Petruska, Rau, & Johnson, 2001; Petruska, Cooper, Gu, Rau, & Johnson, 2000; Staikopoulos, Sessle, Furness, & Jennings, 2007; Wang & Neuhuber, 2003; Xiang, Bo, & Burnstock, 1998). In the airway, ATP was released from epithelial cells by various stimuli, for example, mechanical and osmotic stimuli, and seems to activate sensory nerve endings (Burnstock, Brouns, Timmermans, & Adriaensen, 2012). By immunohistochemical analyses, P2X3 immunoreactivity has been reported in various sensory endings in the airway; ramified intraepithelial nerve endings and the nerve endings associated with solitary and clustered GNAT3‐immunoreactivitive chemosensory cells and SNAP25‐immunoreactive neuroendocrine cells in the rat larynx (Takahashi, Nakamuta, & Yamamoto, 2016), nerve endings associated with the bronchial smooth muscle layer and nerve endings within neuroepithelial bodies in the rat bronchi (Brouns, Adriaensen, Burnstock, & Timmermans, 2000).