Activation Of Fyn Kinase By CAMP-dependent Protein Kinase In Spinal Dorsal Horn Was Involved In Inflammatory Pain

Peripheral tissue injury activates cAMP-dependent protein kinase (PKA) in spinal dorsal horn, which plays a critical and unique role in the initiation and development of chronic inflammatory pain. However, the intracellular signaling cascades that PKA triggers to enhance the nociceptive neuronal excitability and its responsiveness to peripheral stimuli are still largely unknown as yet. Previous studies have demonstrated that src-family protein tyrosine kinases (SFKs) are also required for the establishment of central sensitization. The present study systematically investigated the relationship between PKA and SFKs, finding that:(1) Direct activation of PKA in spinal dorsal horn evoked pronounced mechanical allodynia in naive mice; (2) PKA was able to specifically activate SFKs member Fyn in spinal dorsal horn, which, however, produced minimal effect on the activity of another important SFKs member---Src; (3) The specific activation of Fyn kinase was responsible for PKA-induced mechanical allodynia, because intrathecal application of SFKs inhibitor PP2 or overexpression of a kinase-defective Fyn mutant---Fyn(K296M), greatly attenuated the reduction of paw withdrawal threshold evoked by PKA agonist forskolin; (4) Peripheral tissue injury also stimulated spinal PKA to enhance the enzymatic activity of Fyn kinase; (5) PKA was found to disrupt the association of Fyn with its inhibitory regulatory protein--- Striatal-enriched protein phosphatase 61 (STEP61), which might serve as the key mechanism for PKA to activate spinal Fyn; (6) PKA-activated Fyn operated to regulate the function of NMD A (N-methyl-D-aspartate) subtype glutamate receptors. Our results showed that PKA activity significantly promoted the phosphorylation of NMDA receptor subunit NR2B at tyrosine 1472, which could be totally eliminated by SFKs inhibitor PP2; (7) Although peripheral tissue injury also increased Src activity in spinal dorsal horn, PKA inhibition could not abolish the effect of tissue injury on Src, suggesting that tissue injury might activate Fyn and Src via distinct signaling pathways, which synergistically contributed to the process of central sensitization.Taken together, these data implicated the presence of such a signaling pathway in spinal dorsal horn, that is, PKA→Fyn→NR2B. This signaling pathway might operate to enhance the function of NMDA receptor to initiate and maintain the chronic inflammatory pain following peripheral tissue injury.