cAMP/PKA Pathways and S56 Phosphorylation Are Involved in AA/PGE2-Induced Increases in rNaV1.4 Current.
Arachidonic acid (AA) and its metabolites are important second messengers for ion channel modulation. The effects of extracellular application of AA and its non-metabolized analogue on muscle rNaV1.4 Na+ current has been studied, but little is known about the effects of intracellular application of AA on this channel isoform. Here, we report that intracellular application of AA significantly augmented the rNaV1.4 current peak without modulating the steady-state activation and inactivation properties of the rNaV1.4 channel. These results differed from the effects of extracellular application of AA on rNaV1.4 current. The effects of intracellular AA were mimicked by prostaglandin E2 but not eicosatetraynoic acid (ETYA), the non-metabolized analogue of AA, and were eliminated by treatment with cyclooxygenase inhibitors, flufenamic acid, or indomethacin. AA/PGE2-induced activation of rNaV1.4 channels was mimicked by a cAMP analogue (db-cAMP) and eliminated by a PKA inhibitor, PKAi. Furthermore, inhibition of EP2 and EP4 (PGE2 receptors) with AH6809 and AH23848 reduced the intracellular AA/PGE2-induced increase of rNaV1.4 current. Two mutated channels, rNaV1.4S56A and rNaV1.4T21A, were designed to investigate the role of predicted phosphorylation sites in the AA/PGE2-mediated regulation of rNaV1.4 currents. In rNaV1.4S56A, the effects of intracellular db-cAMP, AA, and PGE2 were significantly reduced. The results of the present study suggest that intracellular AA augments rNaV1.4 current by PGE2/EP receptor-mediated activation of the cAMP/PKA pathway, and that the S56 residue on the channel protein is important for this process.
School of Life Science and Technology, Tongji University, Shanghai 200092, PR China.
Hua
H
Gu
School of Life Sciences, Institute of Brain Science and State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai 200433, PR China.
Yan-Jia
YJ
Fang
School of Life Sciences, Institute of Brain Science and State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai 200433, PR China.
Dong-Dong
DD
Liu
School of Life Science and Technology, Tongji University, Shanghai 200092, PR China.
Ping
P
Chen
School of Life Sciences, Institute of Brain Science and State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai 200433, PR China.
Yan-Ai
YA
Mei
D016718:Arachidonic Acid / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D015232:Dinoprostone / Q000494:pharmacology*; D005439:Flufenamic Acid / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D062553:NAV1.4 Voltage-Gated Sodium Channel / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects* / Q000502:physiology
D016861:Cyclooxygenase Inhibitors; D062553:NAV1.4 Voltage-Gated Sodium Channel; D016718:Arachidonic Acid; D005439:Flufenamic Acid; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D015232:Dinoprostone; D007213:Indomethacin