In contrast to plasma membrane-expressed nicotinic acetylcholine receptors (nAChRs), mitochondrial nAChRs function in an ion-independent manner by triggering intra-mitochondrial kinases that regulate the release of cytochrome c (Cyt c), an important step in cellular apoptosis. The aim of this study is to determine the structural requirements for mitochondrial α3β4* nAChR activation by measuring the modulatory effects of two noncompetitive antagonists of these receptors, (+)-catharanthine and (±)-18-methoxycoronaridine [(±)-18-MC], on Cyt c release from wild-type and α7−/− mice mitochondria. The sandwich ELISA results indicated that α3β4* nAChRs are present in liver mitochondria in higher amounts compared to that in brain mitochondria and that these receptors are up-regulated in α7−/− mice. Correspondingly, (±)-18-MC decreased Cyt c release from liver mitochondria of wild-type mice and from brain and liver mitochondria of α7−/− mice. The effect in wild-type mice mitochondria was mediated mainly by the Src-dependent pathway, regulating the apoptogenic activity of reactive oxygen species, while in α7−/− mice mitochondria, (±)-18-MC strongly affected the calcium-calmodulin kinase II-dependent pathway. In contrast, (+)-catharanthine was much less potent than (±)-18-MC and triggered several signaling pathways, suggesting the involvement of multiple nAChR subtypes. These results show for the first time that noncompetitive antagonists can induce mitochondrial α3β4* nAChR signaling, giving a more comprehensive understanding on the function of intracellular nAChR subtypes.

与质膜表达的烟碱型乙酰胆碱受体（nAChRs）相反，线粒体nAChRs通过触发调节细胞色素c（Cyt c）释放的线粒体内激酶（一种细胞凋亡的重要步骤），以离子独立的方式发挥作用。这项研究的目的是通过测量这些受体的两种非竞争性拮抗剂（+）-catharanthine和（±）-18-甲氧基Coronaridine [（±）-18- MC]，从野生型和α7-/-小鼠线粒体中释放出Cyt c。夹心ELISA结果表明，与脑线粒体相比，肝线粒体中α3β4* nAChRs的含量更高，并且这些受体在α7-/-小鼠中被上调。相应地，（±）-18-MC降低了野生型小鼠肝线粒体以及α7-/-小鼠脑和肝线粒体中Cyt c的释放。在野生型小鼠线粒体中的作用主要由Src依赖性途径介导，调节活性氧的凋亡活性，而在α7-/-小鼠线粒体中，（±）-18-MC强烈影响钙钙调蛋白激酶II依赖性途径。相反，（+）-金盏花碱的效力比（±）-18-MC低得多，并触发了几种信号传导途径，表明涉及多种nAChR亚型。这些结果首次表明非竞争性拮抗剂可以诱导线粒体α3β4* nAChR信号传导，从而对细胞内nAChR亚型的功能有了更全面的了解。在野生型小鼠线粒体中的作用主要由Src依赖性途径介导，调节活性氧的凋亡活性，而在α7-/-小鼠线粒体中，（±）-18-MC强烈影响钙钙调蛋白激酶II依赖性途径。相反，（+）-金盏花碱的效力比（±）-18-MC低得多，并触发了几种信号传导途径，表明涉及多种nAChR亚型。这些结果首次表明非竞争性拮抗剂可以诱导线粒体α3β4* nAChR信号传导，从而对细胞内nAChR亚型的功能有了更全面的了解。在野生型小鼠线粒体中的作用主要由Src依赖性途径介导，调节活性氧的凋亡活性，而在α7-/-小鼠线粒体中，（±）-18-MC强烈影响钙钙调蛋白激酶II依赖性途径。相反，（+）-金盏花碱的效力比（±）-18-MC低得多，并触发了几种信号传导途径，表明涉及多种nAChR亚型。这些结果首次表明非竞争性拮抗剂可以诱导线粒体α3β4* nAChR信号传导，从而对细胞内nAChR亚型的功能有了更全面的了解。（±）-18-MC强烈影响钙钙调蛋白激酶II依赖性途径。相反，（+）-金盏花碱的效力比（±）-18-MC低得多，并触发了几种信号传导途径，表明涉及多种nAChR亚型。这些结果首次表明非竞争性拮抗剂可以诱导线粒体α3β4* nAChR信号传导，从而对细胞内nAChR亚型的功能有了更全面的了解。（±）-18-MC强烈影响钙钙调蛋白激酶II依赖性途径。相反，（+）-金盏花碱的效力比（±）-18-MC低得多，并触发了几种信号传导途径，表明涉及多种nAChR亚型。这些结果首次表明非竞争性拮抗剂可以诱导线粒体α3β4* nAChR信号传导，从而对细胞内nAChR亚型的功能有了更全面的了解。