カルシウムイメージング法の開発　その生物科学への貢献

<p>Now, the importance of Ca²⁺ in the cell has been well recognized. However, year of 1983, when we wanted to measure of intracellular Ca²⁺ ion（[Ca²⁺]i）in a single cell such as neuronal cells, we could not find any reliable method. Only one possibility was to use a fluorescence Ca²⁺ indicator（Ca²⁺ chelator）quin-2, which was invented by RY Tsein, who loaded the reagent into cells by modifying the reagent cell permeable form using esterification. They showed successful measurement of [Ca²⁺]i in cell suspension of lymphocytes by a fluorescence spectrophotometer. In 1985 RY Tsein developed a new indicator called fura-2, which had lower affinity to Ca²⁺ than quin-2. The reagent encouraged us to utilize it for measuring [Ca²⁺]i of the neuronal cell. Since neuronal cells might have diversity in their excitability, we should deal with them separately as a single cell. Thus we intended to develop a new method to measure [Ca²⁺]i in single cells by using a fluorescence microscope. Although we encountered so many obstacles before establishing our method, we conquered them and succeeded in developing a primitive, but efficient [Ca²⁺]i measurement system. The method detected the microscopic fluorescence image obtained by a high sensitive video camera. We measured the brightness of the video image using optical fibers and integrating them and made them measurable data. We succeeded in detecting the [Ca²⁺]i. increase in cultured hippocampal neurons during exposure to glutamate in 1986. This was the first report on the real time measurement of Ca²⁺ increase in a single neuronal cell. We improved our method little by little and five years later we introduced an image-processing system. This method became a standard Ca²⁺ imaging system in Japan. We applied this method and found many important Ca²⁺ related biological functions. Recently, we can use a high-performance computer for analyzing the image data and high-capacity memory system for image analysis. As a fluorescence image detector, we can use a multiphoton microscope. Moreover, the molecular biological technique established new Ca²⁺ indicators called GCaMP family by modifying green fluorescence protein. Now these methods are working at the front of biosciences.</p>