The Drug of Abuse γ-Hydroxybutyrate Is a Substrate for Sodium-Coupled Monocarboxylate Transporter (SMCT) 1 (SLC5A8): Characterization of SMCT-Mediated Uptake and Inhibition
The Drug of Abuse γ-Hydroxybutyrate Is a Substrate for Sodium-Coupled Monocarboxylate Transporter (SMCT) 1 (SLC5A8): Characterization of SMCT-Mediated Uptake and Inhibition
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DOI：
10.1124/dmd.109.027169
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发表时间：
2009-07-01
影响因子：
3.9
通讯作者：
Morris, Marilyn E.
中科院分区：
文献类型：
--
作者：
Cui, Dapeng;Morris, Marilyn E.
γ-羟基丁酸（GHB）是一种滥用药物，是单羧酸转运蛋白（MCT）的底物。钠偶联单羧酸转运蛋白1（SMCT 1; SLC 5A 8）在肾脏、甲状腺、神经元和肠道中表达，表现出与质子依赖性MCT（SLC 16 A）家族相似的底物特异性。SMCT 1在伽马-羟丁酸处置中的作用尚未确定。在这项研究中，我们的特点是驱动力，运输动力学，GHB摄取抑制剂，以及SMCT和MCT亚型的表达，在大鼠甲状腺滤泡（FRTL-5）细胞。GHB以及丁酸单羧酸盐和D-乳酸盐在pH 7.4时表现出钠依赖性摄取，这可以用简单的Michaelis-Menten方程加上扩散分量来描述[ K-m 0.68 +/- 0.30 mM，V-max 3.50 +/- 1.58 nmol。mg（-1）。min（-1），扩散清除率（P）0.25 ± 0.08 μ l . mg（-1）。min（-1）]。在钠的情况下，GHB的摄取显着增加，在较低的pH值，这表明质子梯度依赖的运输。逆转录-聚合酶链反应和Western分析证明SMCT 1、MCT 1和MCT 2在FRTL-5细胞中表达，支持活性结果。MCT底物（D-乳酸、L-乳酸、丙酮酸和丁酸）、非甾体抗炎药（布洛芬、酮洛芬和萘普生）和丙磺舒抑制FRTL-5细胞中钠依赖性GHB摄取。L-乳酸盐、布洛芬、酮洛芬和丙磺舒的IC 50值分别为101、31.6、64.4和380 μ M。所有四种抑制剂也显着抑制GHB摄取大鼠MCT 1基因转染的MDA/MB 231细胞，这表明他们不是特定的SMCT 1。木犀草素和α-氰基-4-羟基肉桂酸酯代表特异性质子依赖性MCT抑制剂。我们的研究结果表明，GHB是钠和质子依赖性MCT的底物，并确定了MCT的特异性抑制剂。
gamma-Hydroxybutyric acid (GHB), a drug of abuse, is a substrate of monocarboxylate transporters (MCTs). Sodium-coupled monocarboxylate transporter 1 (SMCT1; SLC5A8) is expressed in kidney, thyroid gland, neurons, and intestinal tract and exhibits substrate specificity similar to that of the proton-dependent MCT (SLC16A) family. The role of SMCT1 in GHB disposition has not been determined. In this study we characterized the driving force, transport kinetics, and inhibitors of GHB uptake, as well as expression of SMCT and MCT isoforms, in rat thyroid follicular (FRTL-5) cells. GHB, as well as the monocarboxylates butyrate and D-lactate, exhibited sodium-dependent uptake at pH 7.4, which could be described with a simple Michaelis-Menten equation plus a diffusional component [ K-m 0.68 +/- 0.30 mM, V-max 3.50 +/- 1.58 nmol . mg(-1) . min(-1), and diffusional clearance (P) 0.25 +/- 0.08 mu l . mg(-1) . min(-1)]. In the absence of sodium, GHB uptake was significantly increased at lower pH, suggesting proton-gradient dependent transport. Reverse transcriptase-polymerase chain reaction and Western analyses demonstrated the expression of SMCT1, MCT1, and MCT2 in FRTL-5 cells, supporting the activity results. Sodium-dependent GHB uptake in FRTL-5 cells was inhibited by MCT substrates (D-lactate, L-lactate, pyruvate, and butyrate), nonsteroidal anti-inflammatory drugs (ibuprofen, ketoprofen, and naproxen), and probenecid. IC50 values for L-lactate, ibuprofen, ketoprofen, and probenecid were 101, 31.6, 64.4, and 380 mu M, respectively. All four inhibitors also significantly inhibited GHB uptake in rat MCT1 gene-transfected MDA/MB231 cells, suggesting they are not specific for SMCT1. Luteolin and alpha-cyano-4-hydroxycinnimate represent specific proton-dependent MCT inhibitors. Our findings indicate that GHB is a substrate for both sodium-and proton-dependent MCTs and identified specific inhibitors of MCTs.