Zúñiga Rafael, Mancilla Daniel, Rojas Tamara, Vergara Fernando, González Wendy, Catalán Marcelo A, Zúñiga Leandro
Laboratorio de Fisiología Molecular, Escuela de Medicina, Universidad de Talca, Talca 3460000, Chile.
Centro de Nanomedicina, Diagnóstico y Desarrollo de Fármacos (ND3), Escuela de Medicina, Universidad de Talca, Talca 3460000, Chile.
Biology (Basel). 2022 Jul 23;11(8):1097. doi: 10.3390/biology11081097.
Two pore domain potassium channels (K2P) are strongly expressed in the nervous system (CNS), where they play a central role in excitability. These channels give rise to background K currents, also known as IK (standing-outward potassium current). We detected the expression in primary cultured cerebellar granule neurons (CGNs) of TWIK-1 (K2P1), TASK-1 (K2P3), TASK-3 (K2P9), and TRESK (K2P18) channels by immunocytochemistry and their association with lipid rafts using the specific lipids raft markers flotillin-2 and caveolin-1. At the functional level, methyl-β-cyclodextrin (MβCD, 5 mM) reduced IK currents by ~40% in CGN cells. To dissect out this effect, we heterologously expressed the human TWIK-1, TASK-1, TASK-3, and TRESK channels in HEK-293 cells. MβCD directly blocked TASK-1 and TASK-3 channels and the covalently concatenated heterodimer TASK-1/TASK-3 currents. Conversely, MβCD did not affect TWIK-1- and TRESK-mediated K+ currents. On the other hand, the cholesterol-depleting agent filipin III did not affect TASK-1/TASK-3 channels. Together, the results suggest that neuronal background K channels are associated to lipid raft environments whilst the functional activity is independent of the cholesterol membrane organization.
双孔结构域钾通道（K2P）在神经系统（中枢神经系统，CNS）中大量表达，在该系统中它们对兴奋性起着核心作用。这些通道产生背景钾电流，也称为IK（外向持续性钾电流）。我们通过免疫细胞化学检测了双孔钾通道1（TWIK - 1，K2P1）、串联孔道钾通道1（TASK - 1，K2P3）、串联孔道钾通道3（TASK - 3，K2P9）和TWIK - 相关脊髓小胶质细胞钾通道（TRESK，K2P18）在原代培养的小脑颗粒神经元（CGN）中的表达，并使用特异性脂筏标记物小窝蛋白 - 1和浮舰蛋白 - 2检测了它们与脂筏的关联。在功能水平上，甲基 - β - 环糊精（MβCD，5 mM）使CGN细胞中的IK电流降低了约40%。为了剖析这种效应，我们在人胚肾293（HEK - 293）细胞中异源表达了人TWIK - 1、TASK - 1、TASK - 3和TRESK通道。MβCD直接阻断了TASK - 1和TASK - 3通道以及共价连接的异二聚体TASK - 1/TASK - 3电流。相反，MβCD不影响TWIK - 1和TRESK介导的钾离子电流。另一方面，胆固醇耗竭剂制霉菌素III不影响TASK - 1/TASK - 3通道。总之，结果表明神经元背景钾通道与脂筏环境相关，而其功能活性独立于胆固醇膜结构。
