Characterization of 5,7‐Dichlorokynurenate‐Insensitive d‐[3H]Serine Binding to Synaptosomal Fraction Isolated from Rat Brain Tissues
Characterization of 5,7‐Dichlorokynurenate‐Insensitive d‐[3H]Serine Binding to Synaptosomal Fraction Isolated from Rat Brain Tissues
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DOI：
10.1046/j.1471-4159.1997.69010399.x
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发表时间：
1997-07
影响因子：
4.7
通讯作者：
M. Matoba;U. Tomita;T. Nishikawa
中科院分区：
文献类型：
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作者：
M. Matoba;U. Tomita;T. Nishikawa
摘要：为了探索与N-甲基-d-天冬氨酸（NMDA）型谷氨酸受体的甘氨酸位点不同的内源性d-丝氨酸的靶位点，我们研究了在存在过量浓度（100 µM）甘氨酸位点拮抗剂5，7-二氯犬尿酸（DCK）的情况下，d-[3 H]丝氨酸与从大鼠脑和外周组织制备的突触体P2组分的结合。在1 mM未标记的d-丝氨酸存在下定义非特异性结合。结合、解离和饱和实验表明，在DCK存在下，d-[3 H]丝氨酸与小脑P2组分中的单一识别位点群体快速可逆结合，KD为614 nM，Bmax为2.07 pmol/mg蛋白。d-丝氨酸、l-丝氨酸和甘氨酸对特异性DCK-不敏感的d-[3 H]丝氨酸与小脑的结合产生完全抑制，Ki值相似。士的宁和7-氯犬尿酸盐在10 µM浓度下不能抑制结合。DCK不敏感的d-[3 H]丝氨酸结合被各种氨基酸和谷氨酸和甘氨酸受体相关化合物取代的概况不同于任何其他定义的识别位点。DCK不敏感d-[3 H]丝氨酸结合在大脑皮层和小脑中处于高水平，但在肾脏和肝脏中非常低。目前的研究结果表明，DCK不敏感的d-[3 H]丝氨酸结合位点可能是哺乳动物脑中内源性d-丝氨酸靶点的新候选者。
Abstract: To explore target sites for endogenous d‐serine that are different from the glycine site of the N‐methyl‐d‐aspartate (NMDA) type glutamate receptor, we have studied the binding of d‐[3H]serine to the synaptosomal P2 fraction prepared from the rat brain and peripheral tissues in the presence of an excess concentration (100 µM) of the glycine site antagonist 5,7‐dichlorokynurenate (DCK). Nonspecific binding was defined in the presence of 1 mM unlabeled d‐serine. Association, dissociation, and saturation experiments indicated that d‐[3H]serine bound rapidly and reversibly to a single population of recognition sites in the cerebellar P2 fraction in the presence of DCK, with a KD of 614 nM and a Bmax of 2.07 pmol/mg of protein. d‐Serine, l‐serine, and glycine produced a total inhibition of the specific DCK‐insensitive d‐[3H]serine binding to the cerebellum with similar Ki values. Strychnine and 7‐chlorokynurenate failed to inhibit the binding at 10 µM. The profiles of displacement of the DCK‐insensitive d‐[3H]serine binding by various amino acids and glutamate and glycine receptor‐related compounds differ from those of any other defined recognition sites. DCK‐insensitive d‐[3H]serine binding was at high levels in the cerebral cortex and cerebellum but very low in the kidney and liver. The present findings indicate that the DCK‐insensitive d‐[3H]serine binding site could be a novel candidate for a target for endogenous d‐serine in mammalian brains.