Luminopsins integrate opto- and chemogenetics by using physical and biological light sources for opsin activation
Luminopsins integrate opto- and chemogenetics by using physical and biological light sources for opsin activation
复制标题
DOI：
10.1073/pnas.1510899113
复制
发表时间：
2016-01-19
影响因子：
11.1
通讯作者：
Hochgeschwender, Ute
中科院分区：
文献类型：
--
作者：
Berglund, Ken;Clissold, Kara;Hochgeschwender, Ute
发光蛋白是荧光素酶和视蛋白的融合蛋白，通过选择外在物理光或内在生物光进行激活，可以以不同的时间和空间分辨率询问神经元回路。在之前开发的野生型高斯荧光素酶与视紫红质通道融合的基础上，我们通过将明亮的高斯荧光素酶变体与视紫红质通道融合来激发神经元（发光视蛋白，LMO）或与抑制神经元的质子泵（抑制性LMO，iLMO）融合，从而扩展了发光蛋白的用途。这些改进的改性活生物体可以在体外和体内可靠地激活或沉默神经元。改进的 LMO 在海马回路中的表达不仅能够以精细的时空分辨率绘制 CA1 神经元的突触激活图，而且还可以在大时空尺度上驱动节律性回路兴奋。此外，体内黑质中病毒介导的 LMO 或 iLMO 表达不仅产生了预期的对单个单元活性的双向控制，而且还对响应荧光素酶底物全身注射的循环行为产生相反的影响。因此，虽然保留了被外部光源激活的能力，但改性活生物体通过使相同的视蛋白可以进行非侵入性的化学遗传学控制来扩展光遗传学的用途，从而允许相同的探针在一定范围的空间和时间尺度上操纵神经元活动。
Luminopsins are fusion proteins of luciferase and opsin that allow interrogation of neuronal circuits at different temporal and spatial resolutions by choosing either extrinsic physical or intrinsic biological light for its activation. Building on previous development of fusions of wild-type Gaussia luciferase with channelrhodopsin, here we expanded the utility of luminopsins by fusing bright Gaussia luciferase variants with either channelrhodopsin to excite neurons (luminescent opsin, LMO) or a proton pump to inhibit neurons (inhibitory LMO, iLMO). These improved LMOs could reliably activate or silence neurons in vitro and in vivo. Expression of the improved LMO in hippocampal circuits not only enabled mapping of synaptic activation of CA1 neurons with fine spatiotemporal resolution but also could drive rhythmic circuit excitation over a large spatiotemporal scale. Furthermore, virus-mediated expression of either LMO or iLMO in the substantia nigra in vivo produced not only the expected bidirectional control of single unit activity but also opposing effects on circling behavior in response to systemic injection of a luciferase substrate. Thus, although preserving the ability to be activated by external light sources, LMOs expand the use of optogenetics by making the same opsins accessible to noninvasive, chemogenetic control, thereby allowing the same probe to manipulate neuronal activity over a range of spatial and temporal scales.