| Budget Amount *help |
¥8,400,000 (Direct Cost: ¥8,400,000)
Fiscal Year 1990: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1989: ¥7,800,000 (Direct Cost: ¥7,800,000)
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| Research Abstract |
A fluorescent microscope system which are capable of (1) fast local application of physiologically active substances such as Ca ions or cyclic nucleotide, with the aid of caged molecules and focused laser beam, simultaneous (2) optical measurement of membrane potential and (3) optical measurement of intracellular concentration of Ca ions, have been constructed. (1) Nitrogen laser was used as a source of ultraviolet laser beam, and the light was led into an inverted microscope using optical fiber. The optical fiber was connected to a lens system with a X-Y positioner mounted on a camera tube of the microscope. The light was further focused through a X40 objective lens. (2) Optical recording of membrane potential was performed with potentiometric dies such as RH155 and with a photodiode. Transmitted light from a halogen lamp which passed through filters and a specimen was detected with a photodiode mounted on a lateral tube of the microscope. (3) Fluoresent signals reflecting intracellular Ca concentration was detected by a photomultiplier mounted on a lateral tube of the microscope. Fluo-3 AM was used as an indicator. Epifluoresent light came from Hg arc lamp was reflected off a dicroic mirror and led to a specimen. Electrical signals from a photodiode and a photomultiplier were current-voltage converted, multiplied, A-D converted, stored in a computer and analysed. The constructed system has two problems. One is that only a small portion of UV laser beam reaches a specimen, and most is lost within the light path. Another is the poor spatial resolution. To overcome these problems, I am planning a few changes such as the introduction of laser beam to different site of the microscope, and the increase of number of optical detectors.
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