PHYSICS AND CHEMISTRY OF PHOTORECEPTION

光接收的物理和化学

• 批准号: 6161930
• 负责人: W A HAGINS
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6161930
• 关键词: Anura; biomedical equipment development; biophysics; calcium; calcium indicator; calorimetry; chemical synthesis; chromophore; divalent cations; fluorescence microscopy; fluorescent dye /probe; image processing; photochemistry; retina; rod cell; visual photoreceptor; visual phototransduction

1. Members of a family of fluorescent calcium-sensitive indicator dyes including Calcium Green, Calcium Orange, Calcium Crimson, Flo-3 and Rhod-2 have been transferred to retinal rods of frogs by the silylated-dye method described in previous reports in 1993-4. Dye levels exceeding 1 micromolar in the outer segments of rods have been achieved with one our incubations. The dyes partially emerge from the labeled cells when the cell membranes are ruptured osmotically. 2. Studies by image-intensified fluorescence microscopy of the responses of the dyes to calcium ions show that the indicators are almost completely bound to the rod disk membranes and are almost totally insensitive to changes in calcium ion activity in the bathing solutions. Analysis of the polarization of the fluorescence emitted by the dyes Calcium Green, Calcium Crimson and Calcium Orange show that the E-vectors of the emitting oscillators are parallel to the surfaces of the disk membranes, while that of the membrane responsive fluorophore N,N' Bis(dansyl cystine) is parallel to the fatty acid side chains in the membranes. (Yoshikami, Hagins, Sahu) 3. A family of precursors for chelation of divalent ions, including Ca++, have been synthesis with functional groups ready for coupling to fluorescent chromophores. The aim is to produce a group of ned Calcium-sensitive fluorophores that are not inactivated by binding cell organelles as are the above studied dyes available commercially. A total of eight chelators have been made and studies of the coupling reactions are under way. (Hagins, Mani) 4. A pyroelectric photocalorimeter that uses evanescent-waves from stripped optical fibers to introduce photo excitation into small fluid samples is under construction. (Hagins) 5. A pyroelectric calorimeter with improved signal-to-noise properties is under construction for studying phototransduction in frog retinal rods at very low light intensities. Improvements will make it possible to measure heat bursts from nucleotide hydrolysis within 1-2 minutes after the tissue is introduced into the calorimeter. (Yoshikami, Hagins)

1. 荧光钙敏感指示染料家族成员 包括钙绿色、钙橙子、钙深红、Flo-3和 Rhod-2已被转移到青蛙的视网膜杆， 甲硅烷基化染料法在1993-4年的先前报告中描述。 染料 水平超过1微摩尔的外节杆已经 实现了我们的孵化。 染料部分地从 当细胞膜破裂时标记的细胞。 2. 通过图像增强荧光显微镜研究 的染料钙离子显示，指示剂几乎 完全结合在杆盘膜上，几乎完全 对浴液中钙离子活性的变化不敏感。 染料荧光偏振特性的分析 钙绿色、钙深红和钙橙子表明， 发射振荡器的E矢量平行于 圆盘膜，而膜响应荧光团的 N，N'双（丹酰胱氨酸）平行于脂肪酸侧链， 膜。（Yoshikami，Hagins，Sahu） 3. 前体家族 用于螯合二价离子，包括Ca++，已经合成， 准备好与荧光发色团偶联的官能团。 的 目的是产生一组ned钙敏感荧光团， 不像上述研究的染料那样通过结合细胞器而灭活 可商购获得。 共制备了8种螯合剂， 偶联反应的研究正在进行中。（Hagins，Mani）4. 一 热释电光热量计，它使用来自剥离的 将光激发引入小流体样品的光纤 网站正在建设中（哈金斯） 5. 热释电量能器， 改进的信号噪声特性正在建设中，用于研究 青蛙视网膜杆在非常低的光强度下的光转导。 改进将使测量热爆发成为可能， 在引入组织后1-2分钟内核苷酸水解 放进量热计（Yoshikami，Hagins）