Nicotinic acetylcholine receptors (nAChRs) mediate fast synaptic transmission in muscles and autonomic ganglia and regulate cytokine and neurotransmitter release in the brain and non-excitable cells. The α7 nAChRs localized in the outer membrane of mitochondria regulate cytochrome c release stimulated by apoptosis-inducing agents. However, the mechanisms through which nAChRs influence mitochondrial permeability remain obscure. Here we put an aim to explore the interaction of nAChRs with voltage-dependent anion channels (VDAC1) and pro-apoptotic protein Bax in the course of apoptosis induction. By using molecular modeling in silico, it was shown that both Bax and VDAC1 can bind within the 4th transmembrane portion (M4) of nAChR subunits. Experimentally, α7 nAChR-Bax and α7 nAChR-VDAC1 complexes were identified by sandwich ELISA in mitochondria isolated from astrocytoma U373 cells. Stimulating apoptosis of U373 cells by H₂O₂ disrupted α7-VDAC complexes and favored formation of α7-Bax complexes accompanied by cytochrome c release from mitochondria. α7-selective agonist PNU282987 or type 2 positive allosteric modulator PNU120596 disrupted α7-Bax and returned α7 nAChR to complex with VDAC1 resulting in attenuation of cytochrome c release. It is concluded that mitochondrial nAChRs regulate apoptosis-induced mitochondrial channel formation by modulating the interplay of apoptosis-related proteins in mitochondria outer membrane.

烟碱型乙酰胆碱受体 (nAChR) 介导肌肉和自主神经节中的快速突触传递，并调节大脑和非兴奋性细胞中细胞因子和神经递质的释放。位于线粒体外膜中的 α7 nAChRs 调节细胞色素 c 的释放，由细胞凋亡诱导剂刺激。然而，nAChR 影响线粒体通透性的机制仍不清楚。在这里，我们旨在探讨 nAChRs 与电压依赖性阴离子通道 (VDAC1) 和促凋亡蛋白 Bax 在凋亡诱导过程中的相互作用。通过在计算机中使用分子建模，表明 Bax 和 VDAC1 都可以在 nAChR 亚基的第 4 跨膜部分 (M4) 内结合。实验上，α7 nAChR-Bax 和 α7 nAChR-VDAC1 复合物通过夹心 ELISA 在从星形细胞瘤 U373 细胞中分离的线粒体中进行鉴定。H 刺激 U373 细胞凋亡₂ O ₂破坏了 α7-VDAC 复合物，并有利于 α7-Bax 复合物的形成，同时细胞色素 c 从线粒体中释放。α7 选择性激动剂 PNU282987 或 2 型正变构调节剂 PNU120596 破坏 α7-Bax 并使 α7 nAChR 与 VDAC1 复合，导致细胞色素 c 释放减弱。结论是线粒体nAChRs通过调节线粒体外膜中凋亡相关蛋白的相互作用来调节凋亡诱导的线粒体通道形成。