| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1994: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1993: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1992: ¥900,000 (Direct Cost: ¥900,000)
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| Research Abstract |
Complex changes in ionic movement through the sarcolemma occur after ischemia or hypoxia, and may be restored or aggravated after reperfusion or reoxygenation. Earliest changes are increases in [Ca^<2+>]_i, H_i^+ and [Na^+]_i, However, the interactions between these ions are not fully understood. In this study, the effect of ischemia or hypoxia on intracellular and extracellular ionic changes particularly Na^+/Ca^<2+> exchange was investigated in both isolated cardiac myocytes and isolated perfused rat hearts.[Ca^<2+>]_i, pH_i and [Na^+]_i were measured using fluorescence indicators, Fura-2, BCECF and SBFI,respectively. We have measured increases in the [Ca^<2+>]_i of isolated cardiac myocytes during Na^+-free (Na^+ substituted by TMA^+) exposures using Fura-2 in normoxic and hypoxic solutions and when varing pH_i and pH_0. Hypoxia was produced either by (1) bubbling solution with N_2 or (2) bubbling with N_2 and adding 2-deoxy-D-glucose. After 5 min of hypoxia, the rate of increase in
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[Ca^<2+>]_i during exposure to Na^+-free solutions decreased significantly when compared with that observed in aerobic condition. When pH_i is acidified by adding and removing NH_4Cl, similar inhibitory effects to those observed during hypoxia are produced.
When the isolated hearts were perfused with aerobic and ischemic conditions, [Ca^<2+>]_i increases in a biphasic pattern while pH_i decreases rapidly from 7.2 to 6.8 at 3 min add 6.6 at 10 min after hypoxia starts. [Na^+]_i increases gradually during hypoxia. Both the forward and reverse modes of the Na^+/Ca^<2+> exchanger were markedly inhibited during hypoxia and recovered slowly on reoxygenation. The forward mode of Na/Ca exchange was estimated by transient increase in [Ca^<2+>]_i followed by the decay in [Ca^<2+>]_i induced by addition of caffeine. Na^+ channel blocker partially prevented an increase in [Ca^<2+>]_i during hypoxia. These data collectively indicate the complex spectrum of ionic changes, particularly Na^+/Ca^<2+> exchanger that occur during both ischemia/hypoxia and reperfusion/reoxgenation. These results suggested that the Na^+/Ca^<2+> exchange systems, both forward and reverse mode, were inhibited during hypoxia/ischemia. However, an increase in [Ca^<2+>]_i is likely due to the inhibition of forward mode of Na^+/Ca^<2+> exchange since this exchanger normally represents the major efflux pathway for Ca^<2+> in the heart.pH_i changes during hypoxia have also strong modulatory influence on Na^+/Ca^<2+> exchange. Less
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