Investigation of Intracellular Reactions Using Ultramicroelectrodes

使用超微电极研究细胞内反应

• 批准号: 06805075
• 负责人: MATSUE Tomokazu
• 金额: $ 1.22万
• 依托单位: TOHOKU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06805075/
• 关键词: Ultramicroelectrode; Intracellular measurements; Protoplast; Photosynthesis; Oxygen measurements; Animal cells; Superoxide; Nitric oxide; 脂質二分子膜; ボルタンメトリー; イオンチャンネル

1. Ultramicro Pt disk electrodes with tip radii less than 5mum were fabricated via the processes ; electrochemical etching of Pt wire, sealing the Pt fine wire with a soft glass, and polishing the tip. The electrochemical characteristics of the resultant microelectrodes was investigated and found that constant potential amperometry and differential amperometry for oxygen reduction using the microelectrodes afford the oxygen concentration in the local region.2. Intracellular oxygen concentration in a protoplast from Bryopsis plumosa was determined by constant current amperometry. The intracellular oxygen reduction current increased upon light irradiation, indicating that the real-time tracing of the intracellular oxygen is possible by the present method. From the detailcd analyzes of diferential amperometric measurements using the microelectrode located near the cell, the oxygen influx or efflux was determined at a single cell level.3. The permeability of Ru (NH3) 63+ and Fe (CN) 63- through an ion channel in a planar lipid membrane was evaluated by amperometric measurements using microelectrode located near the membrane. The membrane permselectivity of the viable protoplast was also carried out by amperometry.4. Micro NO sensor and micro O_2 sensor were fabricated by immobilizing respectively Ni porphyrin and superoxide dismutase/catalase onto microelectrodes. We implanted these microsensors into rat brain and measured in vivo concentrations of NO and O_2

1.超微铂盘电极的尖端半径小于5 μ m的制造过程中，电化学腐蚀的铂线，密封的铂细线与软玻璃，和抛光的尖端。研究了微电极的电化学特性，发现微电极用于氧还原的恒电位电流法和微分电流法能够提供局部区域的氧浓度.用恒电流安培法测定了羽状真藓原生质体的胞内氧浓度。细胞内氧还原电流在光照下增加，表明本方法可以实时跟踪细胞内氧。通过对微电极差示电流法的详细分析，确定了单细胞水平上的氧流入或流出.用微电极电流法研究了Ru（NH3）63+和Fe（CN）63-在脂膜离子通道中的渗透性。用安培法测定了活原生质体的膜选择透过性.将镍卟啉和超氧化物歧化酶/过氧化氢酶分别固定在微电极上，制成了微型NO传感器和微型O_2传感器。我们将这些微传感器植入大鼠脑内，并测量了体内NO和O_2的浓度

项目成果

H.Yamada, H.Shiku, T.Matsue, I.Uchida: "Microvoltammetric Characterization of Diaphorase Monolayr at Glass Surfaces" Bioelectrochem. Bioenerg. 33. 91-93 (1994)

H.Yamada、H.Shiku、T.Matsue、I.Uchida：“玻璃表面心肌黄酶单层的微伏安特性”生物电化学。

横山英克: "NOセンサーの中枢神経領域におけるin vivo応用" 生物物理. 35. 35-36 (1995)

Hidekatsu Yokoyama：“NO 传感器在中枢神经系统中的体内应用”生物物理学 35. 35-36 (1995)。

H.Shiku, T.Takeda, H.Yamada, T.Matsue, I.Uchida: "Microfabrication and Characterization of Diaphorase-Patterned Surfaces by Scanning Electrochemical Microscopy" Anal. Chem. 67. 312-317 (1995)

H.Shiku、T.Takeda、H.Yamada、T.Matsue、I.Uchida：“通过扫描电化学显微镜对心肌酶图案表面进行微加工和表征”分析。

H. Yokoyama: "Increase of Central Nitric Oxide During Pentylenetetrazol-Induced Seizures in Rats" Psych, Clinic, Neurosci.49. S277-S279 (1995)

H. Yokoyama：“戊四氮引起的大鼠癫痫发作期间中枢一氧化氮的增加”Psych、Clinic、Neurosci.49。

H.Yokoyama, N.Mori, K.Osonoe, N.Kasai, M.Hiramatsu, T.Yoshimura, T.Matsue, I.Uchida, N.Kobayashi, N.Tsuchihashi, S.Niwa: "Increase of Central Nitric Oxide During Pentylenetetrazol-Induced Seizures in Rats" Psych, Clinic, Neurosci. 49. S277-S279 (1995)

H.Yokoyama、N.Mori、K.Osonoe、N.Kasai、M.Hiramatsu、T.Yoshimura、T.Matsue、I.Uchida、N.Kobayashi、N.Tsuchihashi、S.Niwa：“中心一氧化氮的增加