Amphetamine-induced dopamine efflux -: A voltage-sensitive and intracellular Na+-dependent mechanism
Amphetamine-induced dopamine efflux -: A voltage-sensitive and intracellular Na+-dependent mechanism
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DOI：
10.1074/jbc.m212815200
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发表时间：
2003-04-04
影响因子：
4.8
通讯作者：
Galli, A
中科院分区：
文献类型：
--
作者：
Khoshbouei, H;Wang, HW;Galli, A
安非他明（AMPH）通过作用于多巴胺（DA）转运体（DAT）诱导多巴胺溢出进入突触间隙，从而引发其行为效应。促进交换扩散是描述amph诱导的DA外排的经典模型。该模型假设AMPH诱导的DA外排是由DAT介导的，由AMPH转运进入细胞，然后DA反向运动到细胞外隔室引起。为了进一步表征AMPH的作用，我们在全细胞配置下使用膜片钳技术结合安培法对稳定转染了人DAT （DAT细胞）的人胚胎肾HEK-293细胞进行了检测。在DAT细胞中，amph诱导的DAT介导电流被可卡因阻断。我们证明了由DAT介导的DA外排是电压依赖性的，电致性的，并且依赖于记录电极中的细胞内Na+浓度。使用Na+敏感染料的共聚焦显微镜测量细胞内Na+荧光，通过AMPH应用增强。此外，AMPH诱导DA外流的能力受细胞内Na+浓度的调节，并与dat介导的、AMPH诱导的跨质膜离子通量的大小相关。在缺乏细胞内Na+但存在高细胞内Cl-的情况下，amph诱导的内向电流引起的DA外排与其尺寸和持续时间成正比。因此，我们认为amph诱导的DA外排依赖于两个相关的转运体过程。首先，AMPH与DAT结合并被传输，从而产生向内电流。其次，由于这种amph诱导的内向电流，细胞内的Na+更容易被DAT获取，从而增强了DAT介导的DA的反向转运。
Amphetamine (AMPH) elicits its behavioral effects by acting on the dopamine (DA) transporter (DAT) to induce DA overflow into the synaptic cleft. Facilitated exchange diffusion is the classical model used to describe AMPH-induced DA efflux. This model hypothesizes that AMPH-induced DA efflux is mediated by DAT and results from the transport of AMPH into the cell followed by a counter movement of DA out to the extracellular compartment. To further characterize the action of AMPH, we used the patch clamp technique in the whole-cell configuration combined with amperometry on human embryonic kidney HEK-293 cells stably transfected with the human DAT (DAT cells). In DAT cells, AMPH-induced DAT-mediated currents were blocked by cocaine. We demonstrate that DA efflux mediated by DAT is voltage-dependent, electrogenic, and dependent on intracellular Na+ concentration in the recording electrode. Intracellular Na+ fluorescence, as measured by confocal microscopy using a Na+-sensitive dye, was enhanced by AMPH application. Furthermore, the ability of AMPH to induce DA efflux was regulated by intracellular Na+ concentration and correlated with the size of the DAT-mediated, AMPH-induced ion flux across the plasma membrane. In the absence of intracellular Na+ but the presence of high intracellular Cl-, AMPH-induced inward currents elicited DA efflux proportionally to their dimension and duration. Thus, we propose that AMPH-induced DA efflux depends on two correlated transporter processes. First, AMPH binds to the DAT and is transported, thereby causing an inward current. Second, because of this AMPH-induced inward current, Na+ becomes more available intracellularly to the DAT, thereby enhancing DAT-mediated reverse transport of DA.