Pharmacological properties of recombinant human N-methyl-D-aspartate receptors comprising NR1a/NR2A and NR1a/NR2B subunit assemblies expressed in permanently transfected mouse fibroblast cells.
Pharmacological properties of recombinant human N-methyl-D-aspartate receptors comprising NR1a/NR2A and NR1a/NR2B subunit assemblies expressed in permanently transfected mouse fibroblast cells.
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发表时间：
1995-11
影响因子：
3.6
通讯作者：
T. Priestley;P. Laughton;J. Myers;B. L. Bourdellès;J. Kerby;P. Whiting
中科院分区：
文献类型：
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作者：
T. Priestley;P. Laughton;J. Myers;B. L. Bourdellès;J. Kerby;P. Whiting
使用全细胞电压钳电生理学比较了两种重组人 N-甲基-D-天冬氨酸 (NMDA) 受体亚型（包含永久转染至小鼠 L(tk-) 细胞中的 NR1a/NR2A 或 NR1a/NR2B 亚基）的药理学特性。谷氨酸是两种受体的完全激动剂，对 NR2B 的亲和力比对含 NR2A 的受体具有适度但统计显着性 (p < 0.002) 的更高亲和力（微观 Kd [mKd] 分别 = 0.76 和 0.43 microM）。与谷氨酸相比，NMDA、喹啉酸和顺-2,3-哌啶二甲酸是两种受体亚型的部分激动剂。当谷氨酸位点激动剂与甘氨酸或 D-丝氨酸结合时，会产生最大幅度电流；因此，这两种氨基酸都被定义为甘氨酸位点的完全激动剂。甘氨酸对 NR2B- 的亲和力 (p < 0.0001) 比对含 NR2A 受体的亲和力高约 10 倍（mKd 分别为 0.057 和 0.53 microM）。 D-环丝氨酸、(+)-(3R)-3-氨基-1-羟基吡咯烷-2-酮、(+)-顺-(4R)-甲基-(3R)-氨基-1-羟基吡咯烷-2-酮和1-氨基环丁烷甲酸对含NR2B的受体也具有较高的亲和力，但与甘氨酸不同，在两种受体亚型上都是部分激动剂。激动剂诱发的全细胞电流被 D-(-)-2-氨基-5-膦酰基戊酸、顺-4-(膦酰基甲基)哌啶-2-羧酸和 3-((R)-2-羧基哌嗪-4-基)-丙基-1-膦酸拮抗，所有这些都具有轻微但统计显着的较高亲和力（2.2-、2.8-和对于含有 NR2A 的受体，分别为 5.5 倍）。甘氨酸位点拮抗剂 7-氯犬尿酸、7-氯-4-羟基-3-(3-苯氧基)苯基喹啉-2-(1H)-酮和 (+/-)-4-(反式)-2-羧基-5,7-二氯-4-苯基氨基羰基氨基-1,2,3,4-四氢喹啉也可拮抗反应。非典型 NMDA 拮抗剂艾芬地尔在功能亲和力方面表现出最大的分离（对于含 NR2B 和 NR2A 的受体，IC50 值分别为 0.6 和 175 microM）。这些实验证明了永久转染的 L(tk-) 细胞在重组 NMDA 受体功能的电生理学研究中的有用性，并首次提供了人类 NMDA 受体亚型的详细功能药理学分析。
The pharmacological properties of two recombinant human N-methyl-D-aspartate (NMDA) receptor subtypes, comprising either NR1a/NR2A or NR1a/NR2B subunits permanently transfected into mouse L(tk-) cells, have been compared using whole-cell voltage-clamp electrophysiology. Glutamate was a full agonist at both receptors, having a modestly but statistically significant (p < 0.002) higher affinity for the NR2B- than the NR2A-containing receptor (microscopic Kd [mKd] = 0.76 and 0.43 microM, respectively). In comparison to glutamate, NMDA, quinolinic acid, and cis-2,3-piperidinedicarboxylic acid were partial agonists at both receptor subtypes. Maximal amplitude currents resulted when glutamate-site agonists were combined with either glycine or D-serine; both of these amino acids were, therefore, defined as full agonists at the glycine site. Glycine had an approximately 10-fold higher affinity (p < 0.0001) for NR2B- than for NR2A-containing receptors (mKd = 0.057 and 0.53 microM, respectively). D-Cycloserine, (+)-(3R)-3-amino-1-hydroxypyrrolidin-2-one, (+)-cis-(4R)-methyl-(3R)-amino-1-hydroxypyrrolidin-2-one, and 1-aminocyclobutanecarboxylic acid also had higher affinities for the NR2B-containing receptor but were partial agonists, at both receptor subtypes, unlike glycine. Agonist-evoked whole-cell currents were antagonized by D-(-)-2-amino-5-phosphonopentanoic acid, cis-4-(phosphonomethyl)piperidine-2-carboxylic acid, and 3-((R)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid, all of which had slightly, but statistically significant, higher affinities (2.2-, 2.8-, and 5.5-fold, respectively) for the NR2A-containing receptor. Responses were also antagonized by the glycine-site antagonists 7-chlorokynurenic acid, 7-chloro-4-hydroxy-3-(3-phenoxy)phenylquinolin-2-(1H)-one, and (+/=)-4-(trans)-2-carboxy-5,7-dichloro-4-phenylaminocarbonylamino- 1,2,3,4- tetrahydroquinoline. The atypical NMDA antagonist ifenprodil showed the largest separation in functional affinity (IC50 values, 0.6 and 175 microM at NR2B- and NR2A-containing receptors, respectively). These experiments demonstrate the usefulness of permanently transfected L(tk-) cells for electrophysiological studies of recombinant NMDA receptor function and provide the first detailed functional pharmacological analysis of human NMDA receptor subtypes.