类阿片
阿片受体
HEK 293细胞
报告基因
化学
体内
细胞生物学
受体
荧光
计算生物学
药理学
生物
基因
生物化学
基因表达
物理
遗传学
量子力学
作者
Kayla E. Kroning,Wenjing Wang
标识
DOI:10.1002/anie.202101262
摘要
Abstract Mu‐opioid receptor (MOR) signaling regulates multiple neuronal pathways, including those involved in pain, reward, and respiration. To advance the understanding of MOR's roles in pain modulation, there is a need for high‐throughput screening methods of opioids in vitro and high‐resolution mapping of opioids in the brain. To fill this need, we designed and characterized a genetically encoded fluorescent reporter, called S ingle‐chain P rotein‐based O pioid T ransmission I ndicator T ool for M OR (M‐SPOTIT). M‐SPOTIT represents a new and unique mechanism for fluorescent reporter design and can detect MOR activation, leaving a persistent green fluorescence mark for image analysis. M‐SPOTIT showed an opioid‐dependent signal to noise ratio (S/N) up to 12.5 and was able to detect as fast as a 30‐second opioid exposure in HEK293T cell culture. Additionally, it showed an opioid‐dependent S/N up to 4.6 in neuronal culture and detected fentanyl with an EC 50 of 15 nM. M‐SPOTIT will potentially be useful for high‐throughput detection of opioids in cell cultures and cellular‐resolution detection of opioids in vivo. M‐SPOTIT's novel mechanism can be used as a platform to design other G‐protein‐coupled receptor‐based sensors.
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