Pernot Pierre, Mothet Jean-Pierre, Schuvailo Oleg, Soldatkin Alexey, Pollegioni Loredano, Pilone Mirella, Adeline Marie-Thérèse, Cespuglio Raymond, Marinesco Stéphane
CNRS, Institut de Neurobiologie Alfred Fessard-FRC2118, Laboratoire de Neurobiologie Cellulaire et Moléculaire-UPR9040, 91198 Gif sur Yvette, France.
Anal Chem. 2008 Mar 1;80(5):1589-97. doi: 10.1021/ac702230w. Epub 2008 Jan 30.
d-Serine is an endogenous ligand for N-methyl-d-aspartate (NMDA) receptors, and alterations in its concentration have been related to several brain disorders, especially schizophrenia. It is therefore an important target neuromodulator for the pharmaceutical industry. To monitor d-serine levels in vivo, we have developed a microbiosensor based on cylindrical platinum microelectrodes, covered with a membrane of poly-m-phenylenediamine (PPD) and a layer of immobilized d-amino acid oxidase from the yeast Rhodotorula gracilis (RgDAAO). By detecting the hydrogen peroxide produced by enzymatic degradation of d-serine, this microbiosensor shows a detection limit of 16 nM and a mean response time of 2 s. Interferences by ascorbic acid, uric acid, l-cysteine, and by biogenic amines and their metabolites are rejected at more than 97% by the PPD layer. Although several d-amino acids are potential substrates for RgDAAO, d-serine was the only endogenous substrate present in sufficient concentration to be detected by our microbiosensor in the central nervous system. When implanted in the cortex of anesthetized rats, this microbiosensor detected the increase in concentration of d-serine resulting from its diffusion across the blood-brain barrier after an intraperitoneal injection. This new device will make it possible to investigate in vivo the variations in d-serine concentrations occurring under normal and pathological conditions and to assess the pharmacological potency of new drugs designed to impact d-serine metabolism.
D-丝氨酸是N-甲基-D-天冬氨酸（NMDA）受体的内源性配体，其浓度变化与多种脑部疾病相关，尤其是精神分裂症。因此，它是制药行业重要的目标神经调节剂。为了监测体内D-丝氨酸水平，我们开发了一种基于圆柱形铂微电极的微生物传感器，该电极覆盖有聚间苯二胺（PPD）膜和一层固定化的来自纤细红酵母（RgDAAO）的D-氨基酸氧化酶。通过检测D-丝氨酸酶促降解产生的过氧化氢，这种微生物传感器的检测限为16 nM，平均响应时间为2 s。PPD层对维生素C、尿酸、L-半胱氨酸以及生物胺及其代谢物的干扰排除率超过97%。尽管几种D-氨基酸是RgDAAO的潜在底物，但D-丝氨酸是中枢神经系统中唯一浓度足够高、能被我们的微生物传感器检测到的内源性底物。当植入麻醉大鼠的皮层时，这种微生物传感器检测到腹腔注射后D-丝氨酸因扩散穿过血脑屏障而导致的浓度升高。这种新装置将使在体内研究正常和病理条件下D-丝氨酸浓度的变化以及评估旨在影响D-丝氨酸代谢的新药的药理效力成为可能。
