The 2-Pore Domain Potassium Channel TREK-1 Regulates Cytokine Secretion from Human Alveolar Epithelial Cells Independently of Potassium Currents

Abstract

We previously identified the 2-pore domain potassium (K2P) channel TREK-1 as an important regulator of tumor necrosis factor-α (TNF-α)-induced interleukin-6 (IL-6) and monocyte chemotactic protein-1 (MCP-1) secretion from alveolar epithelial cells (AECs). Furthermore, in an in vivo model of Acute Lung Injury (ALI) we found that TREK-1 deficiency led to increased lung damage but decreased broncho-alveolar lavage cytokine levels. Unfortunately, the mechanisms underlying TREK-1 regulation of inflammatory cytokine secretion remain poorly understood.This study was designed to determine whether the changes in cytokine secretion from TREK-1 deficient cells are due to the absence of TREK-1 protein or the lack of TREK-1-mediated potassium currents. For this purpose, we stimulated control and TREK-1 deficient human AECs (A549) with TNF-α and measured IL-6 and MCP-1 release by ELISA in the presence and absence of three TREK-1 blockers, spadin (0.5mM), amlodipine (2μM) and charybdotoxin (100nM). While spadin is recognized as a specific TREK-1 inhibitor, amlodipine and charybdotoxin at these concentrations also inhibit TREK-1.We found that TREK-1 deficient AECs released less IL-6 and more MCP-1 upon TNF-α stimulation but this cytokine release was unaffected by any of the three TREK-1 blockers. To determine whether cytokine secretion occurred independently of potassium currents in general, we raised the extracellular potassium concentration (100mM) to minimize the electrochemical driving force for potassium. Interestingly, we found that TNF-α-induced IL-6 and MCP-1 secretion were significantly impaired in the absence of a potassium gradient.In summary, TREK-1 protein, but not TREK-1 mediated potassium current, is required for IL-6 and MCP-1 secretion from AECs, although other potassium conductances appear important. These data suggest that TREK-1 may alter cytokine secretion from AECs by functioning as a regulatory molecule rather than a potassium channel.