The mechanisms of protease-activated receptor 2 in mediating pain behaviors through nonselective cation channel signaling

Abstract Background: Activation of purinergic receptor P2X ligand-gated ion channel 3 (P2X3), transient receptor potential vanilloid type 1 (TRPV1), and transient receptor potential ankyrin 1 (TRPA1) by their specific ligands is a major mechanism contributing to magnified pain responses. The relationship between these nonselective cation channels and proteinase-activated receptor 2 (PAR2) activation mediated pain is still to be clarified. Methods: In this study, both in vitro model of dorsal root ganglion (DRG) neurons with PAR2 agonist SL-NH2 challenge and SL-NH2-induced pain rat model were used to approach these questions. The expression of P2X3, TRPV1, and TRPA1 in DRG neurons was investigated by quantitative real-time RT-PCR, Western blot, and immunofluorescence. The involvement of the PLCβ3/PKCε signaling pathway was also determined. The behavior test for mechanical allodynia and thermal hyperalgesia was performed. Results: SL-NH2 induced upregulation of P2X3, TRPV1, and TRPA1 through phosphorylation of phospholipase Cβ3 (PLCβ3) and protein kinase Cε (PKCε) signaling pathway in DRG neurons in vitro and in vivo. SL-NH2 also elevated the proportion of P2X3-, TRPV1-, and TRPA1-expressing neurons. The upregulation of P2X3, TRPV1, and TRPA1 and phosphorylation of PLCβ3 and PKCε in DRG neurons was paralleled with mechanical allodynia and thermal hyperalgesia behaviors in rats. Conclusions: The data of the present study imply that SL-NH2 as a noxious stimulus activates PAR2 which induces TRPV1, TRPA1, and P2X3 upregulation through PLCβ3/PKCε signaling pathway, thereby decreasing activation thresholds and increasing excitability, resulting in sustained nociceptive activity in DRG neurons, and then causing mechanical allodynia and thermal hyperalgesia behaviors. These data expanded our knowledge about PAR2-mediated pain sensitivity and its relationship with TRPV1, TRPA1, and P2X3 and provided new opportunities on management of pain behaviors.