Conversion of Channelrhodopsin into a Light-Gated Chloride Channel
Conversion of Channelrhodopsin into a Light-Gated Chloride Channel
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DOI：
10.1126/science.1249375
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发表时间：
2014-04
期刊：
影响因子：
56.9
通讯作者：
Jonas Wietek;J. Simon Wiegert;Nona Adeishvili;F. Schneider;Hiroshi C. Watanabe;S. Tsunoda;A. Vogt
中科院分区：
文献类型：
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作者：
Jonas Wietek;J. Simon Wiegert;Nona Adeishvili;F. Schneider;Hiroshi C. Watanabe;S. Tsunoda;A. Vogt
光遗传学洞察力绘制许多行为的功能性神经回路几乎是不可能的，因此Vogelstein等人（第386页，3月27日在线发表;参见O 'Leary和Marder的Perspective）开发了一种广泛适用的神经元行为映射的光遗传学方法，并将其用于果蝇幼虫的表型，从而开发了一个参考图谱。随着光遗传学实验在神经科学研究的某些领域成为常规，创造更专业的工具势在必行（见Hayashi的观点）。Wietek等人（第409页，3月27日在线发表）和Berndt等人（第420页）通过改造通道视紫红质，创造了两种不同的光门控阴离子通道，以阻断突触刺激或去极化电流注射期间的动作电位产生。这些新工具不仅提高了对通道视紫红质的理解，还提供了一种沉默细胞的方法。一类直接光门控阴离子通道可用于以完全可逆的方式阻断神经元输出。[Also参见Hayashi的《透视》和Berndt等人的《报告》。光遗传学领域使用通道视紫红质（ChR）用于光诱导的神经元激活。然而，缺乏在中等光照水平下用于细胞抑制的优化工具。我们发现，在ChR的中央门E90与带正电荷的残基的替代产生氯离子传导ChRs（ChloCs），只有可忽略的阳离子电导。分子动力学模拟揭示了一个高亲和力的Cl-结合位点附近的门已产生。稳定开放状态显著增加了表达细胞（慢ChloC）的操作光敏感性。在CA 1区锥体细胞，ChloCs完全抑制由去极化电流注入或突触刺激触发的动作电位。因此，通过反转选择性过滤器的电荷，我们已经创建了一类直接光门控阴离子通道，其可以用于以完全可逆的方式阻断神经元输出。
Optogenetic Insights Mapping functional neural circuits for many behaviors has been almost impossible, so Vogelstein et al. (p. 386, published online 27 March; see the Perspective by O'Leary and Marder) developed a broadly applicable optogenetic method for neuron-behavior mapping and used it to phenotype larval Drosophila and thus developed a reference atlas. As optogenetic experiments become routine in certain fields of neuroscience research, creating even more specialized tools is imperative (see the Perspective by Hayashi). By engineering channelrhodopsin, Wietek et al. (p. 409, published online 27 March) and Berndt et al. (p. 420) created two different light-gated anion channels to block action potential generation during synaptic stimulation or depolarizing current injections. These new tools not only improve understanding of channelrhodopsins but also provide a way to silence cells. A class of directly light-gated anion channels can be used to block neuronal output in a fully reversible fashion. [Also see Perspective by Hayashi and Report by Berndt et al.] The field of optogenetics uses channelrhodopsins (ChRs) for light-induced neuronal activation. However, optimized tools for cellular inhibition at moderate light levels are lacking. We found that replacement of E90 in the central gate of ChR with positively charged residues produces chloride-conducting ChRs (ChloCs) with only negligible cation conductance. Molecular dynamics modeling unveiled that a high-affinity Cl–-binding site had been generated near the gate. Stabilizing the open state dramatically increased the operational light sensitivity of expressing cells (slow ChloC). In CA1 pyramidal cells, ChloCs completely inhibited action potentials triggered by depolarizing current injections or synaptic stimulation. Thus, by inverting the charge of the selectivity filter, we have created a class of directly light-gated anion channels that can be used to block neuronal output in a fully reversible fashion.