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Optical Measurements of Intracellular Free Magnesium Concentration in Skeletal and Cardiac Muscle Cardiac

骨骼肌和心肌细胞内游离镁浓度的光学测量心脏

基本信息

批准号：
03670043
负责人：
KONISHI Masato
金额：
$ 1.09万
依托单位：
The Jikei University School of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1991
资助国家：
日本
起止时间：
1991 至 1992
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-03670043/
关键词：
Skeletal muscle Cardiac muscle Intracellular magnesium Fluorescent Mg indicator Acidosis Intracellular ATP 細胞内Mg濃度 Mg指示薬 furaptra 骨格筋線維 NaーMg交換系

项目摘要

The fluorescent Mg^<2+> indicator, furaptra, was injected into single frog skeletal muscle fibers, and the indicator's fluorescence signals were measured at rest and analyzed in terms of the intracellular free Mg^<2+> concentration ([Mg^<2+>]_i)(17ﾟC). Based on the fluorescence excitation spectrum of furaptra, the calibrated [Mg^<2+>]_i level averaged 0.54 mM, if the value of dissociation constant (K_D) obtained in vitro (5.5 mM) was used. However, if the indicator reacts with Mg^<2+> with a two-fold larger K_D in myoplasm, as previously suggested for the furaptra-Ca^<2+> reaction (Konishi et al, 1991, J. Gen. Physiol. 97 : 271-301), the calculated [Mg^<2+>]_i would average 1.1 mM. Thus, the value 1.1 mM Probably represents the best estimate from furaptra of [Mg^<2+>]_i in resting muscle fibers. Extracellular perfusion of muscle fibers with high Mg^<2+> concentration solution or low Na^+ concentration did not cause any detectable changes in [Mg^<2+>]_i within 4 min.Ventricular myocytes enzymatically isolated from rat heart were loaded with furaptra. The fluorescence signals were measured from single myocytes (32ﾟC), and analyzed as in the skeletal muscle fibers The estimated [Mg^<2+>]_i for the resting single myocytes was between 0.8-1.3 mM. Superfusion with a high extracellular Mg^<2+> concentration caused a very slow and small elevation in [Mg^<2+>]_i over a period of hours. Other experimental interventions, including a low extracellular Na^+ concentration, isoproterenol and CO_2 acidosis, did not cause detectable change in [Mg^<2+>]_i, whereas the application of a mitochondrial uncoupler caused a rapid and large increase in [Mg^<2+>]_i.The results suggest that the [Mg^<2+>]_i level of around 1 mM is similar in different muscle types (skeletal and cardiac muscles). It is also suggested that [Mg^<2+>]_i is highly maintained at around 1 mM in both skeletal and cardiac muscles, unless intracellular ATP is depleted.

将荧光Mg^<2+>指示剂furaptra注射到单个青蛙骨骼肌纤维中，在静息状态下测量指示剂的荧光信号，并根据细胞内游离Mg^<2+>浓度（[Mg^<2+>] i）（17 ℃）进行分析。以呋喃甲素的荧光激发光谱为基础，用离体测定的解离常数（KD）值（5.5mM）校正[Mg^<2+>] i的平均值为0.54mM。然而，如果指示剂与Mg^<2+>反应，则在肌浆中的K_D大两倍，如先前对furaptra-Ca^<2+>反应所建议的，（Konishi等人，1991，J.Gen.Physiol.97：271-301），计算的[Mg^<2+>]_i将平均为1.1 mM。因此，1.1mM的值可能代表静息肌纤维中[Mg^2+]_i的最佳估计值。用高浓度Mg^<2+>溶液或低浓度Na^+溶液对肌纤维进行细胞外灌流，在4分钟内均未引起[Mg^<2+>]_i的任何可检测的变化。从单个肌细胞（32 ℃）中测量荧光信号，并像在骨骼肌纤维中一样进行分析。静息单个肌细胞的[Mg^2+] i估计值在0.8-1.3 mM之间。在数小时内，高浓度的细胞外Mg^2+灌流导致[Mg^2+] i非常缓慢和微小的升高。其他实验干预，包括低细胞外Na^+浓度、异丙肾上腺素和CO_2酸中毒，均未引起[Mg^<2+>]_i的可检测变化，而线粒体解偶联剂的应用引起[Mg^<2+>]_i的快速和大量增加。结果表明，在不同的肌肉类型中，约1 mM的[Mg^<2+>]_i水平是相似的（骨骼肌和心肌）。还表明，[Mg^<2+>]_i在骨骼肌和心肌中高度维持在约1 mM，除非细胞内ATP耗尽。