Protease-activated receptor 2 sensitizes TRPV1 by protein kinase Cε- and A-dependent mechanisms in rats and mice

Proteases that are released during inflammation and injury cleave protease-activated receptor 2 (PAR₂) on primary afferent neurons to cause neurogenic inflammation and hyperalgesia. PAR₂-induced thermal hyperalgesia depends on sensitization of transient receptor potential vanilloid receptor 1 (TRPV1), which is gated by capsaicin, protons and noxious heat. However, the signalling mechanisms by which PAR₂ sensitizes TRPV1 are not fully characterized. Using immunofluorescence and confocal microscopy, we observed that PAR₂ was colocalized with protein kinase (PK) Cε and PKA in a subset of dorsal root ganglia neurons in rats, and that PAR₂ agonists promoted translocation of PKCε and PKA catalytic subunits from the cytosol to the plasma membrane of cultured neurons and HEK 293 cells. Subcellular fractionation and Western blotting confirmed this redistribution of kinases, which is indicative of activation. Although PAR₂ couples to phospholipase Cβ, leading to stimulation of PKC, we also observed that PAR₂ agonists increased cAMP generation in neurons and HEK 293 cells, which would activate PKA. PAR₂ agonists enhanced capsaicin-stimulated increases in [Ca²⁺]_i and whole-cell currents in HEK 293 cells, indicating TRPV1 sensitization. The combined intraplantar injection of non-algesic doses of PAR₂ agonist and capsaicin decreased the latency of paw withdrawal to radiant heat in mice, indicative of thermal hyperalgesia. Antagonists of PKCε and PKA prevented sensitization of TRPV1 Ca²⁺ signals and currents in HEK 293 cells, and suppressed thermal hyperalgesia in mice. Thus, PAR₂ activates PKCε and PKA in sensory neurons, and thereby sensitizes TRPV1 to cause thermal hyperalgesia. These mechanisms may underlie inflammatory pain, where multiple proteases are generated and released. 2006 The Authors. Journal compilation