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A fluorescent sensor for spatiotemporally resolved imaging of endocannabinoid dynamics in vivo
Nature Biotechnology ( IF 46.9 ) Pub Date : 2021-11-11 , DOI: 10.1038/s41587-021-01074-4
Ao Dong 1, 2, 3 , Kaikai He 1, 2 , Barna Dudok 4 , Jordan S Farrell 4 , Wuqiang Guan 5 , Daniel J Liput 6, 7 , Henry L Puhl 7 , Ruyi Cai 1, 2 , Huan Wang 1, 2 , Jiali Duan 1, 2 , Eddy Albarran 8 , Jun Ding 9 , David M Lovinger 6 , Bo Li 5 , Ivan Soltesz 4 , Yulong Li 1, 2, 3, 10
Affiliation  

Endocannabinoids (eCBs) are retrograde neuromodulators with important functions in a wide range of physiological processes, but their in vivo dynamics remain largely uncharacterized. Here we developed a genetically encoded eCB sensor called GRABeCB2.0. GRABeCB2.0 consists of a circular-permutated EGFP and the human CB1 cannabinoid receptor, providing cell membrane trafficking, second-resolution kinetics with high specificity for eCBs, and shows a robust fluorescence response at physiological eCB concentrations. Using GRABeCB2.0, we monitored evoked and spontaneous changes in eCB dynamics in cultured neurons and acute brain slices. We observed spontaneous compartmentalized eCB transients in cultured neurons and eCB transients from single axonal boutons in acute brain slices, suggesting constrained, localized eCB signaling. When GRABeCB2.0 was expressed in the mouse brain, we observed foot shock-elicited and running-triggered eCB signaling in the basolateral amygdala and hippocampus, respectively. In a mouse model of epilepsy, we observed a spreading wave of eCB release that followed a Ca2+ wave through the hippocampus. GRABeCB2.0 is a robust probe for eCB release in vivo.



中文翻译:

一种用于体内内源性大麻素动力学时空分辨成像的荧光传感器

内源性大麻素 (eCB) 是逆行性神经调节剂,在广泛的生理过程中具有重要功能,但它们的体内动力学在很大程度上仍未得到表征。在这里,我们开发了一种名为 GRAB eCB2.0的基因编码 eCB 传感器。GRAB eCB2.0由循环排列的 EGFP 和人类 CB1 大麻素受体组成,提供细胞膜运输、对 eCB 具有高特异性的第二分辨率动力学,并在生理 eCB 浓度下显示出强大的荧光响应。使用 GRAB eCB2.0, 我们监测了培养的神经元和急性脑切片中 eCB 动力学的诱发和自发变化。我们观察到培养神经元中的自发区室化 eCB 瞬变和急性脑切片中单个轴突 bouton 的 eCB 瞬变,表明受限的局部 eCB 信号。当 GRAB eCB2.0在小鼠大脑中表达时,我们分别在基底外侧杏仁核和海马体中观察到足部电击引起和跑步触发的 eCB 信号。在癫痫小鼠模型中,我们观察到随着 Ca 2+波通过海马体释放的 eCB 扩散波。GRAB eCB2.0是用于体内 eCB 释放的强大探针。

更新日期:2021-11-11
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