| Project/Area Number |
09670709
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Circulatory organs internal medicine
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| Research Institution | Hamamatsu University School of Medicine |
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Principal Investigator |
HAYASHI Hideharu Hamamatsu University School of Medicine, Photon Medical Research Center, Associate Professor, 光量子医学研究センター, 助教授 (50135258)
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| Co-Investigator(Kenkyū-buntansha) |
SATOH Hiroshi Hamamatsu University School of Medicine, Department of Medicine, Assistant, 医学部附属病院, 助手 (30293632)
TERADA Hajime Hamamatsu University School of Medicine, Department of Medicine, Assistant, 医学部, 助手 (50252177)
金子 雅則 浜松医科大学, 医学部, 助手 (70191989)
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| Project Period (FY) |
1997 – 1998
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| Project Status |
Completed (Fiscal Year 1998)
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| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 1998: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1997: ¥2,700,000 (Direct Cost: ¥2,700,000)
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| Keywords | Ca^<2+> transient / Cell length / indo-1 / Hydrogen peroxide / Ca^<2+> responsiveness / stunned myocardium / laser scanning confocal microscopy / fluo-3 / sarcoplasmic reticulum / Ca^<2+> spark / Na^+ / Ca^<2+>交換 / caffeine transient / 筋小胞体(SR) / pH低下 / ラット / 心室筋細胞 / Ca^<2+> Sensitivity / 虚血 / 再灌流 |
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| Research Abstract |
For the simultaneous measurement of [Ca^<2+>]i and cell length in single myocytes, indo-1 loaded myocytes were excited at 360 nm via an epifluorescence illuminator from a l00-W xenon lamp. The indo-1 emission signal was separated into 405- and 485-nm wavelengths with the appropriate dichroic mirror and band-pass filters. Both fluorescent signals were sampled using two photomultipliers with a photon counting unit. The fluorescent ratios were obtained by dividing the fluorescent intensity at 405-nm by the fluorescent intensity at 480-nm after each background subtraction. The cells were simultaneously illuminated with red light (>600 nm), and the bright-field image was separated from the fluorescence image by 580-nm long-pass dichroic mirror. The cell image was focused on the linear image sensor consisted of 2048 photodiodes. Both edges of a cell were identified by the edge detection system and cell lengths were computed from the numbers of photodiodes between each edges. The simultaneous
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recording of the Ca^<2+> transient and twitch cell shortening was obtained during the control condition and at 5 min after the perfusion of 1 mM hydrogen peroxide. It was shown that the % change in twitch cell shortening was significantly depressed after 2 min perfusion with hydrogen peroxide, while there was no significant change in the % change of the Ca^<2+> transient amplitude. Next, we studied the effect of 1 mM hydrogen peroxide on pH_i in rat ventricular myocytes. The pH_i decreased gradually from 7.234*0.029 to 7.100*0.031 (p<0.0l) after 10 min perfusion of hydrogen peroxide. There was, however, no significant change in pH_i during the first 4 min. In the present study, hydrogen peroxide did not decrease pH_i during the first min of the application of hydrogen peroxide. The reduction in pH_i by hydrogen peroxide could contribute, at least in pail, to the decreased responsiveness of myofilaments after the first min of application. In conclusion, it was shown that hydrogen peroxide decreased Ca^<2+> responsiveness of myofibrils in stunned myocardium.
Next, the line Ca^<2+> transient was recorded in a myocyte loaded with fluo-3/AM using laser scanning confocal microscopy. The decay phase of caffeine transient, which indicates the Ca^<2+> efflux by Na^+/Ca^<2+> exchange was analyzed during low pH and ATP depletion. The contribution of sarcoplasmic reticulum was also analyzed by observing the effects of low pH on the frequency of Ca^<2+> spark. Less
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