Protease-activated receptor 2 sensitizes TRPV1 by protein kinase Cε- and A-dependent mechanisms in rats and mice
Protease-activated receptor 2 sensitizes TRPV1 by protein kinase Cε- and A-dependent mechanisms in rats and mice
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DOI：
10.1113/jphysiol.2006.111534
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发表时间：
2006-09-01
影响因子：
5.5
通讯作者：
Bunnett, Nigel W.
中科院分区：
文献类型：
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作者：
Amadesi, Silvia;Cottrell, Graeme S.;Bunnett, Nigel W.
在炎症和损伤过程中释放的蛋白酶切割初级传入神经元上的蛋白酶激活受体2（PAR（2）），引起神经源性炎症和痛觉过敏。PAR（2）诱导的热痛敏依赖于瞬时受体电位香草酸受体1（TRPV 1）的增敏，该增敏由辣椒素、质子和有害热门控。然而，PAR（2）致敏TRPV1的信号机制尚未完全确定。利用免疫荧光和共聚焦显微镜观察到PAR（2）与蛋白激酶（PK）C β和PKA共定位于大鼠背根神经节神经元的一个亚群中，并且PAR（2）激动剂促进PKC β和PKA催化亚单位从培养的神经元和HEK 293细胞的胞浆移位到质膜。亚细胞分级分离和蛋白质印迹法证实了这种激酶的重新分布，这是激活的指示。尽管PAR（2）与磷脂酶C β偶联，导致PKC刺激，但我们还观察到PAR（2）激动剂增加了神经元和HEK 293细胞中cAMP的产生，从而激活PKA。PAR（2）激动剂增强辣椒素刺激的HEK 293细胞[Ca 2 +]（i）和全细胞电流的增加，表明TRPV 1敏化。足底联合注射非痛觉剂量的PAR（2）激动剂和辣椒素可减少小鼠对辐射热的缩爪潜伏期，提示热痛觉过敏。PKC β和PKA的拮抗剂阻止HEK 293细胞中TRPV 1 Ca 2+信号和电流的敏化，并抑制小鼠的热痛觉过敏。因此，PAR（2）激活感觉神经元中的PKC β和PKA，从而使TRPV 1增敏而引起热痛觉过敏。这些机制可能是炎症性疼痛的基础，其中产生和释放多种蛋白酶。
Proteases that are released during inflammation and injury cleave protease-activated receptor 2 (PAR(2)) on primary afferent neurons to cause neurogenic inflammation and hyperalgesia. PAR(2)-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 PAR(2) sensitizes TRPV1 are not fully characterized. Using immunofluorescence and confocal microscopy, we observed that PAR(2) was colocalized with protein kinase (PK) C epsilon and PKA in a subset of dorsal root ganglia neurons in rats, and that PAR(2) agonists promoted translocation of PKC epsilon 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 PAR(2) couples to phospholipase C beta, leading to stimulation of PKC, we also observed that PAR(2) agonists increased cAMP generation in neurons and HEK 293 cells, which would activate PKA. PAR(2) 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 PAR(2) agonist and capsaicin decreased the latency of paw withdrawal to radiant heat in mice, indicative of thermal hyperalgesia. Antagonists of PKC epsilon and PKA prevented sensitization of TRPV1 Ca2+ signals and currents in HEK 293 cells, and suppressed thermal hyperalgesia in mice. Thus, PAR(2) activates PKC epsilon 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.