PIP₂ Mediated Inhibition of TREK Potassium Currents by Bradykinin in Mouse Sympathetic Neurons.

Bradykinin (BK), a hormone inducing pain and inflammation, is known to inhibit potassium M-currents (I_M) and to increase the excitability of the superior cervical ganglion (SCG) neurons by activating the Ca²⁺-calmodulin pathway. M-current is also reduced by muscarinic agonists through the depletion of membrane phosphatidylinositol 4,5-biphosphate (PIP₂). Similarly, the activation of muscarinic receptors inhibits the current through two-pore domain potassium channels (K2P) of the "Tandem of pore-domains in a Weakly Inward rectifying K⁺ channel (TWIK)-related channels" (TREK) subfamily by reducing PIP₂ in mouse SCG neurons (mSCG). The aim of this work was to test and characterize the modulation of TREK channels by bradykinin. We used the perforated-patch technique to investigate riluzole (RIL) activated currents in voltage- and current-clamp experiments. RIL is a drug used in the palliative treatment of amyotrophic lateral sclerosis and, in addition to blocking voltage-dependent sodium channels, it also selectively activates the K2P channels of the TREK subfamily. A cell-attached patch-clamp was also used to investigate TREK-2 single channel currents. We report here that BK reduces spike frequency adaptation (SFA), inhibits the riluzole-activated current (I_RIL), which flows mainly through TREK-2 channels, by about 45%, and reduces the open probability of identified single TREK-2 channels in cultured mSCG cells. The effect of BK on I_RIL was precluded by the bradykinin receptor (B₂R) antagonist HOE-140 (d-Arg-[Hyp³, Thi⁵, d-Tic⁷, Oic⁸]BK) but also by diC₈PIP₂ which prevents PIP₂ depletion when phospholipase C (PLC) is activated. On the contrary, antagonizing inositol triphosphate receptors (IP₃R) using 2-aminoethoxydiphenylborane (2-APB) or inhibiting protein kinase C with bisindolylmaleimide did not affect the inhibition of I_RIL by BK. In conclusion, bradykinin inhibits TREK-2 channels through the activation of B₂Rs resulting in PIP₂ depletion, much like we have demonstrated for muscarinic agonists. This mechanism implies that TREK channels must be relevant for the capture of information about pain and visceral inflammation.

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