Neuroscience Paris Seine - Institut de Biologie Paris Seine (NPS - IBPS), CNRS, INSERM, Sorbonne Université, Paris, France.
CNRS, Université de Paris, UPR 9080, Laboratoire de Biochimie Théorique, Paris, France.
Elife. 2020 Oct 28;9:e59026. doi: 10.7554/eLife.59026.
Glutamate delta (GluD) receptors belong to the ionotropic glutamate receptor family, yet they don't bind glutamate and are considered orphan. Progress in defining the ion channel function of GluDs in neurons has been hindered by a lack of pharmacological tools. Here, we used a chemo-genetic approach to engineer specific and photo-reversible pharmacology in GluD2 receptor. We incorporated a cysteine mutation in the cavity located above the putative ion channel pore, for site-specific conjugation with a photoswitchable pore blocker. In the constitutively open GluD2 Lurcher mutant, current could be rapidly and reversibly decreased with light. We then transposed the cysteine mutation to the native receptor, to demonstrate with high pharmacological specificity that metabotropic glutamate receptor signaling triggers opening of GluD2. Our results assess the functional relevance of GluD2 ion channel and introduce an optogenetic tool that will provide a novel and powerful means for probing GluD2 ionotropic contribution to neuronal physiology.
谷氨酸 δ（GluD）受体属于离子型谷氨酸受体家族，但它们不结合谷氨酸，被认为是孤儿受体。由于缺乏药理学工具，定义神经元中 GluD 的离子通道功能的进展一直受到阻碍。在这里，我们使用化学遗传方法在 GluD2 受体中设计特定的和光可逆药理学。我们在位于假定的离子通道孔上方的腔中引入半胱氨酸突变，用于与光可切换的孔阻断剂进行位点特异性缀合。在组成型开放的 GluD2 Lurcher 突变体中，电流可以快速且可逆地用光减少。然后，我们将半胱氨酸突变转移到天然受体上，以高药理学特异性证明代谢型谷氨酸受体信号触发 GluD2 的打开。我们的结果评估了 GluD2 离子通道的功能相关性，并引入了一种光遗传学工具，这将为研究 GluD2 离子型对神经元生理学的贡献提供一种新颖而强大的手段。
