Capsaicin Up-Regulates Protease-Activated Receptor-4 mRNA and Protein in Primary Cultured Dorsal Root Ganglion Neurons
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Previous study has shown that there is a functional link between the transient receptor potential vanilloid type 1 (TRPV1) receptor and protease-activated receptor-4 (PAR4) in modulation of inflammation and pain. Capsaicin activation of TRPV1 is involved in enhancement of the expression of TRPV1 in mRNA and protein in dorsal root ganglion (DRG) in vivo. Whether capsaicin could influence expression of PAR4 in primary sensory neurons remains unknown. In the present study, expression of PAR4 in cultured rat DRG neurons was observed using immunofluorescence, real-time PCR and Western blots to examine whether increases in PAR4 mRNA and protein levels are induced by capsaicin treatment with or without pre-treatment of forskolin, a cyclic AMP/protein kinase A (cAMP/PKA) activator or PKA inhibitor fragment 14-22 (PKI14-22), a PKA inhibitor. Capsaicin treatment of cultured DRG neurons significantly increased the expression of PAR4 in mRNA and protein levels. The percentage of PAR4-, TRPV1-immunoreactive neurons and their co-localization in cultured DRG neurons increased significantly in the presence of capsaicin as compared with that in the absence of capsaicin. Compared with capsaicin-only group, pre-incubation with forskolin strongly enhanced the capsaicin-induced increase of PAR4 in mRNA and protein levels. Consistent with the involvement of PKA in the modulation of PAR4 expression, this evoked expression both at mRNA and protein levels was significantly inhibited after PKA was inhibited by pre-incubation with PKI14-22. Taken together, these results provide evidence that TRPV1 activation significantly increases the expression of PAR4 mRNA and protein levels in primary cultures of DRG neurons after capsaicin incubation. Effects of capsaicin on PAR4 expression appear to be mediated by cAMP/PKA signal pathways in DRG neurons.
KeywordsProtease-activated receptor-4 Transient receptor potential vanilloid type 1 Cultured primary afferent neurons Protein kinase A Real-time PCR Rat
This study was supported by the National Nature Science Foundation of China (81070898) and Nature Science Foundation of Shandong Province (2009ZRB01209).
- Amadesi S, Cottrell GS, Divino L, Chapman K, Grady EF, Bautista F, Karanjia R, Barajas-Lopez C, Vanner S, Vergnolle N, Bunnett NW (2006) Protease-activated receptor 2 sensitizes TRPV1 by protein kinase C epsilon- and A-dependent mechanisms in rats and mice. J Physiol 575:555–571PubMedCrossRefGoogle Scholar
- Augé C, Balz-Hara D, Steinhoff M, Vergnolle N, Cenac N (2009) Protease-activated receptor-4 (PAR 4): a role as inhibitor of visceral pain and hypersensitivity. Neurogastroenterol Motil 21:1189–e107Google Scholar
- Nakanishi M, Hata K, Nagayama T, Sakurai T, Nishisho T, Wakabayashi H, Hiraga T, Ebisu S, Yoneda T (2010) Acid activation of Trpv1 leads to an up-regulation of calcitonin gene-related peptide expression in dorsal root ganglion neurons via the CaMK-CREB cascade: a potential mechanism of inflammatory pain. Mol Biol Cell 21:2568–2577PubMedCrossRefGoogle Scholar
- Nishimura S, Ishikura H, Matsunami M, Shinozaki Y, Sekiguchi F, Naruse M, Kitamura T, Akashi R, Matsumura K, Kawabata A (2010) The proteinase/proteinase-activated receptor-2/transient receptor potential vanilloid-1 cascade impacts pancreatic pain in mice. Life Sci 87:643–650PubMedCrossRefGoogle Scholar
- Ritchie E, Saka M, Mackenzie C, Drummond R, Wheeler-Jones C, Kanke T, Plevin R (2007) Cytokine upregulation of proteinase-activated receptors 2 and 4 expression mediated by p38 MAP kinase and inhibitory kappa B kinase b in human endothelial cells. Br J Pharmacol 150:1044–1054PubMedCrossRefGoogle Scholar
- Schnizler K, Shutov LP, Van Kanegan MJ, Merrill MA, Nichols B, McKnight GS, Strack S, Hell JW, Usachev YM (2008) Protein kinase A anchoring via AKAP150 is essential for TRPV1 modulation by forskolin and prostaglandin E2 in mouse sensory neurons. J Neurosci 28(19):4904–4917PubMedCrossRefGoogle Scholar
- Vellani V, Petrosino S, De Petrocellis L, Valenti M, Prandini M, Magherini PC, McNaughton PA (2008) Functional lipidomics. Calcium-independent activation of endocannabinoid/endovanilloid lipid signalling in sensory neurons by protein kinases C and A and thrombin. Neuropharmacology 55:1274–1279PubMedCrossRefGoogle Scholar