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Abstract：
G-protein-gated inward rectifier K+ (GIRK) channels play a critical role in the regulation of the excitability of cardiomyocytes and neurons and include GIRK1, GIRK2, GIRK3 and GIRK4 subfamily members. BD1047 dihydrobromide (BD1047) is one of the representative antagonists of the multifunctional Sigma-1 receptor (S1R). In the analysis of the effect of BD1047 on the regulation of Gi-coupled receptors by S1R using GIRK channel as an effector, we observed that BD1047, as well as BD1063, directly inhibited GIRK currents even in the absence of S1R and in a voltage-independent manner. Thus, we aimed to clarify the effect of BD1047 on GIRK channels and identify the structural determinants. By electrophysiological recordings in Xenopus oocytes, we observed that BD1047 directly inhibited GIRK channel currents, producing a much stronger inhibition of GIRK4 compared to GIRK2. It also inhibited ACh-induced native GIRK current in isolated rat atrial myocytes. Chimeric and mutagenesis studies of GIRK2 and GIRK4 combined with molecular docking analysis demonstrated the importance of Leu77 and Leu84 within the cytoplasmic, proximal N-terminal region and Glu147 within the pore-forming region of GIRK4 for inhibition by BD1047. The activator of GIRK channels, ivermectin, competed with BD1047 at Leu77 on GIRK4. This study provides us with a novel inhibitor of GIRK channels and information for developing pharmacological treatments for GIRK4-associated diseases.
摘要：
G蛋白门控内向整流K（GIRK）通道在调节心肌细胞和神经元的兴奋性中起关键作用，包括GIRK1，GIRK2，GIRK3和GIRK4亚家族成员。BD1047二氢溴酸盐(BD1047)是多功能Sigma-1受体(S1R)的代表性拮抗剂之一。在使用GIRK通道作为效应子分析BD1047对S1R调节Gi偶联受体的作用中，我们观察到BD1047和BD1063即使在没有S1R的情况下也直接抑制GIRK电流，并且以电压无关的方式。因此，我们旨在阐明BD1047对GIRK通道的影响，并确定结构决定因素。通过在非洲爪的卵母细胞中的电生理记录，我们观察到BD1047直接抑制GIRK通道电流，与GIRK2相比，对GIRK4的抑制作用更强。它还抑制分离的大鼠心房肌细胞中ACh诱导的天然GIRK电流。GIRK2和GIRK4的嵌合和诱变研究结合分子对接分析，证明了Leu77和Leu84在细胞质中的重要性，近端N端区域和GIRK4孔形成区域内的Glu147被BD1047抑制。GIRK通道的活化剂，伊维菌素,在GIRK4的Leu77上与BD1047竞争。这项研究为我们提供了一种新型的GIRK通道抑制剂，并为开发GIRK4相关疾病的药物治疗提供了信息。
