Fluorescent ratiometric pH indicator SypHer2: Applications in neuroscience and regenerative biology.
Fluorescent ratiometric pH indicator SypHer2: Applications in neuroscience and regenerative biology.
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DOI：
10.1016/j.bbagen.2015.08.002
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发表时间：
2015-11
期刊：
影响因子：
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通讯作者：
Belousov VV
中科院分区：
文献类型：
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作者：
Matlashov ME;Bogdanova YA;Ermakova GV;Mishina NM;Ermakova YG;Nikitin ES;Balaban PM;Okabe S;Lukyanov S;Enikolopov G;Zaraisky AG;Belousov VV
SypHer is a genetically encoded fluorescent pH-indicator with a ratiometric readout, suitable for measuring fast intracellular pH shifts. However, a relatively low brightness of the indicator limits its use. Here we designed a new version of pH-sensor - SypHer-2, that has up to three times brighter fluorescence signal in cultured mammalian cells compared to the SypHer. Using the new indicator we registered activity-associated pH oscillations in neuronal cell culture. We observed prominent temporal neuronal cytoplasm acidification that occurs in parallel with calcium entry. Furthermore, we monitored pH in presynaptic and postsynaptic termini by targeting SypHer-2 directly to these compartments and revealed marked differences in pH dynamics between synaptic boutons and dendritic spines. Finally, we were able to reveal for the first time the intracellular pH drop which occurs within an extended region of the amputated tail of the Xenopus laevis tadpole before it begins to regenerate. SypHer2 is suitable for quantitative monitoring of pH in biological systems of different scales, from small cellular subcompartments to animal tissues in vivo. The new pH-sensor will help to investigate pH-dependent processes in both in vitro and in vivo studies.
