Light-regulated voltage-gated potassium channels for acute interrogation of channel function in neurons and behavior.
Light-regulated voltage-gated potassium channels for acute interrogation of channel function in neurons and behavior.
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DOI：
10.1371/journal.pone.0248688
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发表时间：
2021
期刊：
影响因子：
3.7
通讯作者：
Pfaffinger PJ
中科院分区：
文献类型：
--
作者：
Jerng HH;Patel JM;Khan TA;Arenkiel BR;Pfaffinger PJ
电压门控钾通道调节可兴奋细胞的膜电位和电导，控制动作电位的放电速率和波形。尽管对Kv通道进行了70多年的深入研究，但令人惊讶的是，人们对各种神经元中特定通道的表达以及它们的功能特性如何影响体内神经元网络的活动和行为知之甚少。尽管已经尝试了许多离子通道的体内遗传操作，但对这些结果的解释由于强大的稳态可塑性机制而变得复杂，这些机制在兴奋性扰动后起着维持功能的作用。为了更好地理解Kv通道是如何塑造网络功能和行为的，我们开发了一种新的光遗传学技术，通过将冷冻沃切菌金黄色色素1的光敏LOV结构域融合到Kv1亚基蛋白的n端，从而形成一个optokv1通道，从而用光敏锐地调节Kv通道的表达。对非洲爪蟾卵母细胞、哺乳动物细胞和神经元中Opto-Kv1通道表达的记录表明，蓝光在所有测试系统中都强烈诱导Opto-Kv1通道的当前表达。我们还发现含有显性负孔突变的Opto-Kv1结构（Opto-Kv1(V400D)）可用于以蓝光依赖的方式下调Kv1电流。最后，为了确定Opto-Kv1通道是否能在体内引发光依赖性行为效应，我们将Opto-Kv1 （V400D）的表达靶向于小鼠嗅球二尖瓣细胞中表达kv1.3的细胞。将灯泡暴露在蓝光下2-3小时，在对类似气味初步习惯后，灯泡对新气味的敏感性显著增加，这与Kv1.3基因敲除动物的行为变化相当。总之，我们已经开发了新的光激活Kv通道，提供了新的方法来询问体内的神经回路，并检查正常和致病突变Kv通道在脑功能和行为中的作用。
Voltage-gated potassium (Kv) channels regulate the membrane potential and conductance of excitable cells to control the firing rate and waveform of action potentials. Even though Kv channels have been intensely studied for over 70 year, surprisingly little is known about how specific channels expressed in various neurons and their functional properties impact neuronal network activity and behavior in vivo. Although many in vivo genetic manipulations of ion channels have been tried, interpretation of these results is complicated by powerful homeostatic plasticity mechanisms that act to maintain function following perturbations in excitability. To better understand how Kv channels shape network function and behavior, we have developed a novel optogenetic technology to acutely regulate Kv channel expression with light by fusing the light-sensitive LOV domain of Vaucheria frigida Aureochrome 1 to the N-terminus of the Kv1 subunit protein to make an Opto-Kv1 channel. Recording of Opto-Kv1 channels expressed in Xenopus oocytes, mammalian cells, and neurons show that blue light strongly induces the current expression of Opto-Kv1 channels in all systems tested. We also find that an Opto-Kv1 construct containing a dominant-negative pore mutation (Opto-Kv1(V400D)) can be used to down-regulate Kv1 currents in a blue light-dependent manner. Finally, to determine whether Opto-Kv1 channels can elicit light-dependent behavioral effect in vivo, we targeted Opto-Kv1 (V400D) expression to Kv1.3-expressing mitral cells of the olfactory bulb in mice. Exposure of the bulb to blue light for 2–3 hours produced a significant increase in sensitivity to novel odors after initial habituation to a similar odor, comparable to behavioral changes seen in Kv1.3 knockout animals. In summary, we have developed novel photoactivatable Kv channels that provide new ways to interrogate neural circuits in vivo and to examine the roles of normal and disease-causing mutant Kv channels in brain function and behavior.
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DOI：
10.1083/jcb.201210119
发表时间：
2013-05-13
期刊：
The Journal of cell biology
影响因子：
--
作者：
Chen D;Gibson ES;Kennedy MJ
通讯作者：
Kennedy MJ
影响因子：
48
作者：
Kennedy, Matthew J.;Hughes, Robert M.;Peteya, Leslie A.;Schwartz, Joel W.;Ehlers, Michael D.;Tucker, Chandra L.
通讯作者：
Tucker, Chandra L.
影响因子：
25
作者：
Banghart, M;Borges, K;Kramer, RH
通讯作者：
Kramer, RH
影响因子：
2.5
作者：
Fortin, Doris L.;Dunn, Timothy W.;Kramer, Richard H.
通讯作者：
Kramer, Richard H.
影响因子：
1.5
作者：
Nagai, Y;Sano, H;Yokoi, M
通讯作者：
Yokoi, M