QUANTITATIVE IMAGING OF ZINC METABOLISM IN THE BRAIN

大脑中锌代谢的定量成像

• 批准号: 6625455
• 负责人: RICHARD Blair THOMPSON
• 金额: $ 28.8万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-12-17 至 2004-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6625455
• 关键词: bioimaging /biomedical imaging; brain imaging /visualization /scanning; brain metabolism; chemical binding; fluorescence spectrometry; intracellular transport; metal metabolism; microdialysis; sulfonamides; technology /technique development; tissue /cell culture; zinc

Zinc is a metal ion with many functions in the body, including roles in DNA recognition, recognition, enzymatic catalysis, and neural function. Among these, the role(s) of zinc in the brain are the least well understood, yet evidence is mounting that zinc plays a role in nerve cell death following stroke, Alzheimer's disease, epilepsy, and other forms of brain damage. The major goal of this proposal is to develop a group of optical probes for studying zinc in neural tissue by fluorescence microscopy employing our fluorescence-based biosensor technology. The fluorescence-based sensing technology utilizes the high selectivity and affinity of human apo-carbonic anhydrase for zinc; upon binding Zn(II), a covalent or noncovalent label on the protein exhibits changes in fluorescence intensity, wavelength, lifetime, and/or anisotropy. Fluorescence anisotropy microscopy offers significant advantages over wavelength-ratiometric and lifetime-based microscopies. The demonstrated advantages of this approach compared to other fluorescence probes for zinc are thousand-fold better sensitivity (down to picomolar zinc); insensitivity to Mg(II) at 50 mM, Ca at 10 mM, and most other metals; excitation at visible wavelengths, and high efficiency. Specific aims include quantitative imaging of zinc release from mossy fiber boutons in the hippocampus, imaging intracellular free zinc levels and fluxes in cultured cells, observing zinc release in vivo with a microdialysis technique, and developing probes for multiphoton-excitation microscopy.

锌是一种金属离子，在体内具有多种功能，包括在DNA识别、识别、酶催化和神经功能等方面发挥作用。其中，锌在大脑中的作用最不为人所知，但越来越多的证据表明，锌在中风、阿尔茨海默病、癫痫和其他形式的脑损伤后的神经细胞死亡中发挥作用。该提案的主要目标是开发一组光学探针，利用我们基于荧光的生物传感器技术通过荧光显微镜研究神经组织中的锌。基于荧光的传感技术利用人类脱辅基碳酸酐酶对锌的高选择性和亲和力；结合 Zn(II) 后，蛋白质上的共价或非共价标记会表现出荧光强度、波长、寿命和/或各向异性的变化。荧光各向异性显微镜比波长比率显微镜和基于寿命的显微镜具有显着的优势。与其他锌荧光探针相比，这种方法的优点是灵敏度高出千倍（低至皮摩尔锌）；对 50 mM Mg(II)、10 mM Ca 和大多数其他金属不敏感；可见光波长激发，效率高。具体目标包括对海马体中苔藓纤维纽扣释放的锌进行定量成像，对培养细胞中的细胞内游离锌水平和通量进行成像，利用微透析技术观察体内锌的释放，以及开发用于多光子激发显微镜的探针。