Qhobosheane Monde, Wu Donghai, Gu Yunrong, Tan Weihong
Department of Chemistry, The McKnight Brain Institute, University of Florida, Gainesville, FL 32611, USA.
J Neurosci Methods. 2004 May 30;135(1-2):71-8. doi: 10.1016/j.jneumeth.2003.12.005.
In this study, we have imaged and monitored real-time release of neurotransmitter glutamate from mouse brain slices stimulated by nicotine and physiological salts using a newly developed two-dimensional (2D) imaging biosensor. Nicotine, the addictive substance contained in cigarettes and other tobacco baring products, affects human body through interactions with both central and peripheral nervous system receptors and exists in various concentrations in body organs including the brain. The 2D imaging biosensor, designed for sensitive glutamate monitoring, is prepared through the use of a flat silica plate that is covalently attached with an enzyme, glutamate dehydrogenase (GDH). The 2D imaging biosensor can spatially resolve glutamate release and has a detection limit of 40 nM in detecting glutamate release from glutaminergic neurons in the brain slices. Using this biosensor, we have shown that the administration of nicotine (50 nM) on the brain slice stimulates repetitive glutamate release. To the best of our knowledge, this is the first time that a biosensor was used to define the vital role of nicotine in glutamate release. Physiological salts such as KCl and CaCl(2) have also been used to characterize the biosensor in measuring glutamate release. It is found in this study that nicotine stimulation is much more potent than physiological salts. The real-time detection of newly released glutamate from mouse brain slices clearly demonstrates the feasibility of the two-dimensional glutamate biosensor for real-time monitoring of dynamic glutamate release from living specimen.
在本研究中，我们使用新开发的二维（2D）成像生物传感器，对尼古丁和生理盐溶液刺激的小鼠脑片释放神经递质谷氨酸的过程进行了成像和实时监测。尼古丁是香烟及其他烟草制品中的成瘾物质，通过与中枢和外周神经系统受体相互作用影响人体，并以不同浓度存在于包括大脑在内的身体器官中。用于灵敏监测谷氨酸的二维成像生物传感器，是通过使用与谷氨酸脱氢酶（GDH）共价连接的平面硅胶板制备而成。该二维成像生物传感器能够在空间上分辨谷氨酸的释放，在检测脑片中谷氨酰胺能神经元释放谷氨酸时的检测限为40 nM。使用这种生物传感器，我们发现向脑片施用尼古丁（50 nM）会刺激谷氨酸的重复性释放。据我们所知，这是首次使用生物传感器来确定尼古丁在谷氨酸释放中的重要作用。氯化钾和氯化钙等生理盐溶液也已用于表征该生物传感器在测量谷氨酸释放方面的性能。本研究发现，尼古丁刺激比生理盐溶液更有效。从小鼠脑片中实时检测新释放的谷氨酸，清楚地证明了二维谷氨酸生物传感器用于实时监测活体标本中动态谷氨酸释放的可行性。
