A Study on the Excitation-Contraction Coupling of Myocardium ---Analysis on Isolated, perfused Hearts using Multi-Nuclear NMR Methods.

本研究主要探讨缺血再灌注损伤引起细胞内钙稳态紊乱的机制。细胞内离子钙浓度（[Ca^<2+>]_i）用与钙指示剂5 F-BAPTA偶联的氟-19 NMR（F-NMR）测量。结果表明：（1）30 ℃缺血30分钟，心肌[Ca^<2+>]_i在缺血结束时和再灌注早期明显升高; Ca ^<2 +>通道阻断剂预处理可抑制[Ca ^<2+>]_i的升高，提示Ca^<2+>通道开放是缺血时Ca^<2+>升高的机制之一。2)在37 ℃下全脑缺血15分钟后再灌注的心脏显示心肌顿抑。相比之下，心脏重新灌注高[Na]_o。溶液显示出明显更好的回收率。然而，用蔗糖或氯化胆碱替代额外钠的溶液再灌注， ...更多信息 ve功能恢复，表明高[Na] o.再灌注损伤的原因既不是由于高离子强度也不是由于高渗透压。这些结果表明，高[Na]_o.再灌注可防止心肌顿抑，支持Na^+/Ca^+交换在心肌顿抑机制中起重要作用的观点。3)测定心肌细胞内磷酸糖（[SP]）和[Ca^2+] i的含量，以阐明糖酵解对细胞内Ca^2+稳态的贡献。在消耗糖原后，无糖酵解底物的灌注显示舒张末期左室压（EDP）和[SP]无显著变化。即使在糖酵解阻断后，灌注液中葡萄糖的缺失既不增加[SP]也不增加EDP。F-NMR显示EDP与[Ca^<2+>]_i显著相关。这些结果表明，细胞内Ca^2+稳态的破坏是由SP的积累引起的，而不是糖酵解的抑制。少

This research program focused on the mechanism for the disruption of intracellular calcium homeostasis induced by ischemia and reperfusion. Intracellular ionic calcium concentration ([Ca^<2+>]_i) was measured with fluorine-19 NMR (F-NMR) coupled with a calcium indicator, 5F-BAPTA. The results obtained in this program are as follows.1)[Ca^<2+>]_i significantly increased at the end of ischemia and during the early phase of reperfusion when hearts were subjected into 30-min ischemia at30ﾟC. The increase of [Ca^<2+>]_i was attenuated by pretreatment ofCa^<2+> channel blocker, indicating that opening of Ca^<2+> channel is one of the mechanism of Ca^<2+> increase during ischemia. 2)Hearts reperfused after 15-min global ischemia at 37ﾟshowed myocardial stunning. In contrast, hearts reperfused with high-[Na]_o. solution showed significantly better recovery. Nevertheless, reperfusion with solutions in which the additional Na was substituted either by sucrose or by choline chloride did not impro … More ve functional recovery, indicating that the beneficial effects of high-[Na]_o. reperfusion are not due either to high ionic strength or to high osmolarity. These results indicate that high-[Na]_o. reperfusion protects against stunning, supporting the idea that the Na^+/Ca^+ exchange plays an important role in the mechanism of stunned myocardium. 3)Myocardial content of sugar phosphates ([SP]) and [Ca^<2+>]_i were measured to elucidate the contribution of glycolysis to intracellular Ca^<2+> homeostasis. After the depletion of glycogen, the perfusion without substrate of glycolysis showed no significant changes in end-diastolic LV pressure (EDP) and [SP]. Even after the blockage of glycolysis, omission of glucose from the perfusate increased neither [SP] nor EDP. Nevertheless, addition of glucose to perfusate consistently increased EDP with accumulation of SP. F-NMR revealed that EDP correlates significantly with [Ca^<2+>]_i. These results indicate that disruption of intracellular Ca^<2+> homeostasis is caused by the accumulation of SP, rather than the inhibition of glycolysis. Less