Identity of the organic cation transporter OCT3 as the extraneuronal monoamine transporter (uptake2) and evidence for the expression of the transporter in the brain
Identity of the organic cation transporter OCT3 as the extraneuronal monoamine transporter (uptake2) and evidence for the expression of the transporter in the brain
复制标题
DOI：
10.1074/jbc.273.49.32776
复制
发表时间：
1998-12-04
影响因子：
4.8
通讯作者：
Ganapathy, V
中科院分区：
文献类型：
--
作者：
Wu, X;Kekuda, R;Ganapathy, V
我们研究了转运阳离子神经毒素和神经递质的潜在敏感的有机转运蛋白OCT 3和类固醇敏感性使用异源表达系统，并分析了OCT 3在大脑中的表达。当在哺乳动物细胞中表达时，OCT 3介导神经毒素1-甲基-4-苯基吡啶（MPP+）和神经递质多巴胺的摄取。竞争实验表明，几种阳离子神经活性剂，包括安非他明与OCT 3相互作用，当在非洲爪蟾卵母细胞中表达时，OCT 3介导的MPP+摄取与电压钳条件下的内向电流相关。MPP+诱导的电流相对于MPP+浓度是饱和的，并且半最大饱和（K-0.5）发生在约25 μ M MPP+，膜电位钳位在-50 mV。膜电位对MPP ~+的K ~（-0.5）有显著影响。OCT 3被几种类固醇抑制，β-雌二醇是最有效的抑制剂（Ki类似于1 μ M）。OCT 3的类固醇敏感性模式与OCT 1和OCT 2不同，但与神经元单胺转运蛋白（摄取）显著相关（2）。转运特性和类固醇敏感性为OCT 3作为摄取的分子身份提供了强有力的证据。通过北方印迹分析、逆转录-聚合酶链反应和使用OCT 3特异性探针的原位杂交证实OCT 3在脑中表达。通过对逆转录-聚合酶链反应产物进行测序以及从脑cDNA文库中分离OCT 3 cDNA，已确定与探针杂交的转录物的分子身份。原位杂交的区域分布研究表明，OCT 3在脑内不同区域广泛表达，尤其是在海马、小脑和大脑皮层，OCT 3可能在脑内阳离子神经毒素和神经递质的处置中发挥重要作用。
We investigated the transport of cationic neurotoxins and neurotransmitters by the potential-sensitive organic transporter OCT3 and its steroid sensitivity using heterologous expression systems and also analyzed the expression of OCT3 in the brain. When expressed in mammalian cells, OCT3 mediates the uptake of the neurotoxin 1-methyl-4-phenylpyridinium (MPP+) and the neurotransmitter dopamine. Competition experiments show that several cationic neuroactive agents including amphetamines interact with OCT3, When expressed in Xenopus laevis oocytes, OCT3-mediated MPP+ uptake is associated with inward currents under voltage-clamp conditions. The MPP+-induced currents are saturable with respect to MPP+ concentration, and half-maximal saturation (K-0.5) occurs at about 25 mu M MPP+ with membrane potential clamped at -50 mV. The K-0.5 for MPP+ is markedly influenced by membrane potential. OCT3 is inhibited by several steroids, and beta-estradiol is the most potent inhibitor (K-i similar to 1 mu M). The pattern of steroid sensitivity of OCT3 is different from that of OCT1 and OCT2 but correlates significantly with that of the extraneuronal monoamine transporter (uptake(2)). The transport characteristics and steroid sensitivity provide strong evidence for the molecular identity of OCT3 as uptake,. OCT3 is expressed in the brain as evidenced from Northern blot analysis, reverse transcription-polymerase chain reaction, and in situ hybridization using OCT3-specific probes. The molecular identity of the transcript hybridizing to the probe has been established by sequencing the reverse transcription-polymerase chain reaction product and also by the isolation of the OCT3 cDNA from a brain cDNA library. Regional distribution studies with in situ hybridization show that OCT3 is expressed widely in different brain regions, especially in the hippocampus, cerebellum, and cerebral cortex, OCT3 is likely to play a significant role in the disposition of cationic neurotoxins and neurotransmitters in the brain.