Center for Functional Connectomics, Korea Institute of Science and Technology, Seoul, Republic of Korea.
Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore.
J Neurosci Res. 2020 Mar;98(3):410-421. doi: 10.1002/jnr.24152. Epub 2017 Sep 1.
Previous work has demonstrated that fusion of a luciferase to an opsin, to create a luminescent opsin or luminopsin, provides a genetically encoded means of manipulating neuronal activity via both chemogenetic and optogenetic approaches. Here we have expanded and refined the versatility of luminopsin tools by fusing an alternative luciferase variant with high light emission, Gaussia luciferase mutant GLucM23, to depolarizing and hyperpolarizing channelrhodopsins with increased light sensitivity. The combination of GLucM23 with Volvox channelrhodopsin-1 produced LMO4, while combining GLucM23 with the anion channelrhodopsin iChloC yielded iLMO4. We found efficient activation of these channelrhodopsins in the presence of the luciferase substrate, as indicated by responses measured in both single neurons and in neuronal populations of mice and rats, as well as by changes in male rat behavior during amphetamine-induced rotations. We conclude that these new luminopsins will be useful for bimodal opto- and chemogenetic analyses of brain function.
先前的工作表明，通过将荧光素酶与视蛋白融合，创建发光视蛋白或光视蛋白，提供了一种通过化学生物学和光遗传学方法来操纵神经元活动的基因编码手段。在这里，我们通过将具有高光发射的替代荧光素酶变体 Gaussia luciferase mutant GLucM23 与去极化和超极化通道蛋白融合，扩展和改进了光视蛋白工具的多功能性，从而提高了光敏感性。将 GLucM23 与 Volvox 通道蛋白-1 结合产生了 LMO4，而将 GLucM23 与阴离子通道蛋白 iChloC 结合则产生了 iLMO4。我们发现，在存在荧光素酶底物的情况下，这些通道蛋白能够被有效地激活，这一点可以通过在小鼠和大鼠的单个神经元以及神经元群体中测量的反应，以及在雄性大鼠在安非他命诱导的旋转过程中的行为变化来证明。我们得出结论，这些新型光视蛋白将有助于对大脑功能进行双模光遗传学和化学生物学分析。
