Real-time, spatially resolved analysis of serotonin transporter activity and regulation using the fluorescent substrate, ASP+.
Real-time, spatially resolved analysis of serotonin transporter activity and regulation using the fluorescent substrate, ASP+.
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DOI：
10.1111/j.1471-4159.2010.06828.x
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发表时间：
2010-08
影响因子：
4.7
通讯作者：
Shippenberg TS
中科院分区：
文献类型：
--
作者：
Oz M;Libby T;Kivell B;Jaligam V;Ramamoorthy S;Shippenberg TS
The serotonin transporter (SERT) mediates clearance of serotonin from the synapse, thereby, regulating extracellular serotonin concentrations. Radioligand uptake techniques are typically used to assess SERT function in tissue and heterologous expression systems. The need for sufficient protein in samples, however, requires use of homogenate preparations, potentially masking effects limited to specific cell populations. 4-(4-(dimethylamino)-styryl)-N-methylpyridinium (ASP+) is a fluorescent monoamine transporter substrate that has been used for real-time monitoring of dopamine and norepinephrine transporter function in single cells. The present live cell imaging studies examine the utility of ASP+ for quantifying hSERT function in HEK-293 and neuroblastoma cells. We show rapid membrane binding and intracellular ASP+ accumulation in hSERT expressing cells. Accumulation is saturable; dependent on temperature and the presence of sodium and chloride in the media, and attenuated by serotonin. Acute or prolonged exposure of cells to serotonin re-uptake inhibitors produces a concentration-dependent decrease in accumulation. Similar effects are produced by PKC activation whereas p38MAPK activation increases ASP+ accumulation. These data demonstrate the validity of ASP+ as a probe for monitoring SERT function in living cells. Alterations in SERT binding and uptake can be quantified in the same cell and use of a within cell design permits analysis of time-related alterations in SERT function.
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影响因子：
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作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
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