Improved expression of halorhodopsin for light-induced silencing of neuronal activity.
Improved expression of halorhodopsin for light-induced silencing of neuronal activity.
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DOI：
10.1007/s11068-008-9034-7
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发表时间：
2008-08
期刊：
影响因子：
--
通讯作者：
Feng G
中科院分区：
文献类型：
--
作者：
Zhao S;Cunha C;Zhang F;Liu Q;Gloss B;Deisseroth K;Augustine GJ;Feng G
在完整的哺乳动物大脑中控制和操纵神经元活动的能力对于映射功能连接和解剖行为背后的神经回路至关重要。我们以前已经产生了转基因小鼠，表达通道视紫红质-2光诱导激活的神经元和映射的神经回路。在这里，我们描述了转基因小鼠，表达盐视紫红质（NpHR），光驱动的氯泵，可用于沉默神经元活动通过光。使用Thy-1启动子将NpHR表达靶向神经元，我们发现这些小鼠中的神经元表达高水平的NpHR-YFP，并且表达NpHR-YFP的皮质锥体神经元的光照导致这些细胞中动作电位放电的快速、可逆的光抑制。然而，NpHR-YFP表达导致许多细胞内泡的形成，这可能破坏神经元功能。各种亚细胞标记物的标记表明，泡产生于内质网中NpHR-YFP的保留。通过改进信号肽序列并向NpHR-YFP添加ER输出信号，我们消除了水泡的形成并显著增加了NpHR-YFP的膜表达。因此，NpHR的改进版本应该作为一个很好的工具，在体外和体内的神经元沉默。
The ability to control and manipulate neuronal activity within an intact mammalian brain is of key importance for mapping functional connectivity and for dissecting the neural circuitry underlying behaviors. We have previously generated transgenic mice that express channelrhodopsin-2 for light-induced activation of neurons and mapping of neural circuits. Here we describe transgenic mice that express halorhodopsin (NpHR), a light-driven chloride pump, that can be used to silence neuronal activity via light. Using the Thy-1 promoter to target NpHR expression to neurons, we found that neurons in these mice expressed high levels of NpHR-YFP and that illumination of cortical pyramidal neurons expressing NpHR-YFP led to rapid, reversible photoinhibition of action potential firing in these cells. However, NpHR-YFP expression led to the formation of numerous intracellular blebs, which may disrupt neuronal function. Labeling of various subcellular markers indicated that the blebs arise from retention of NpHR-YFP in the endoplasmic reticulum. By improving the signal peptide sequence and adding an ER export signal to NpHR-YFP, we eliminated the formation of blebs and dramatically increased the membrane expression of NpHR-YFP. Thus, the improved version of NpHR should serve as an excellent tool for neuronal silencing in vitro and in vivo.
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DOI：
10.1073/pnas.0700293104
发表时间：
2007-03-20
影响因子：
11.1
作者：
Armbruster, Blaine N.;Li, Xiang;Roth, Bryan L.
通讯作者：
Roth, Bryan L.
DOI：
10.1083/jcb.131.6.1715
发表时间：
1995-12
期刊：
The Journal of cell biology
影响因子：
--
作者：
Nakamura N;Rabouille C;Watson R;Nilsson T;Hui N;Slusarewicz P;Kreis TE;Warren G
通讯作者：
Warren G
影响因子：
16.2
作者：
Arenkiel, Benjamin R.;Peca, Joao;Feng, Guoping
通讯作者：
Feng, Guoping
影响因子：
9.2
作者：
Nagel, G;Brauner, M;Gottschalk, A
通讯作者：
Gottschalk, A
影响因子：
25
作者：
Boyden, ES;Zhang, F;Deisseroth, K
通讯作者：
Deisseroth, K