Functional expression of the norepinephrine transporter in cultured rat astrocytes
Functional expression of the norepinephrine transporter in cultured rat astrocytes
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DOI：
10.1046/j.1471-4159.2003.01514.x
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发表时间：
2003-01-01
影响因子：
4.7
通讯作者：
Matsumiya, T
中科院分区：
文献类型：
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作者：
Inazu, M;Takeda, H;Matsumiya, T
我们评估了去甲肾上腺素（NE）转运蛋白（NET）在培养的大鼠皮质星形胶质细胞的功能表达。特异性[H-3]NE摄取以时间依赖性方式增加，并且这种摄取涉及温度和Na+敏感机制。Na+依赖的[H-3]NE摄取是饱和的，该过程的Km为539.3 +/- 55.4 nM，V-max为141 +/- 0.03 pmol/mg protein/min。哇巴因，一种Na+-K+ ATP酶抑制剂，显著抑制Na+依赖的[H-3]NE摄取。选择性NE摄取抑制剂尼索西汀、三环类抗抑郁药地昔帕明和丙咪嗪以及5-羟色胺和NE再摄取抑制剂（SNRI）米那普仑非常有效地抑制Na+依赖性[H-3]NE摄取。另一方面，GBR-12935（一种选择性多巴胺摄取抑制剂）、氟伏沙明（一种选择性5-羟色胺再摄取抑制剂）、文拉法辛（一种SNRI）和可卡因具有较弱的抑制活性。RT-PCR结果显示，星形胶质细胞表达NET蛋白，具有神经元NET的特征。Western blot结果显示，抗NET多克隆抗体在星形胶质细胞中识别一条80 kDa的主带。这些数据表明神经元NET在培养的大鼠星形胶质细胞中功能性表达。神经胶质细胞可以通过失活NE来显著控制去甲肾上腺素能活性，NE在远离末端的位点逃避神经元再摄取，因此是抑制NE摄取的抗抑郁药物的细胞靶点。
We assessed the functional expression of the norepinephrine (NE) transporter (NET) in cultured rat cortical astrocytes. Specific [H-3]NE uptake increased in a time-dependent manner, and this uptake involves temperature- and Na+ -sensitive mechanisms. The Na+ -dependent [H-3]NE uptake was saturable, and the K-m for the process was 539.3 +/- 55.4 nM and the V-max was 141 +/- 0.03 pmol/mg protein/min. Ouabain, a Na+-K+ ATPase inhibitor, significantly inhibited Na+ -dependent [H-3]NE uptake. The selective NE uptake inhibitor nisoxetine, the tricyclic antidepressants desipramine and imipramine, and the serotonin and NE reuptake inhibitor (SNRI) milnacipran very potently inhibited Na+-dependent [H-3]NE uptake. On the other hand, GBR-12935 (a selective dopamine uptake inhibitor), fluvoxamine (a selective serotonin reuptake inhibitor), venlafaxine (a SNRI) and cocaine had weaker inhibitory activities. RT-PCR demonstrated that astrocytes expressed mRNA for the cloned NET protein, which was characterized as neuronal NET. Western blots indicated that anti-NET polyclonal antibody recognized a major band of 80 kDa in astrocytes. These data indicate that the neuronal NET is functionally expressed in cultured rat astrocytes. Glial cells may exert significant control of noradrenergic activity by inactivating NE that escapes neuronal re-uptake in sites distant from terminals, and are thus cellular targets for antidepressant drugs that inhibit NE uptake.