Gow I F, Latham T, Ellis D, Flatman P W
Department of Physiology, University Medical School, Edinburgh, UK.
Magnes Res. 1995 Sep;8(3):223-32.
The preparation by collagenase dispersion is described of isolated, calcium-tolerant myocytes from the septomarginal (moderator) band dissected from the sheep heart. Cells obtained were small rods (85 x 9 microns), with pronounced striations characteristic of cardiac myocytes. Isolated cells were loaded with the fluorescent ion-sensitive probes Mag-fura-2 or fura-2, for use in microspectrofluorimetry experiments to measure cytosolic free magnesium ([Mg2+]i) and calcium ([Ca2+]i) respectively; cells remained usable for up to 8 h after isolation. Glycolysis and electron transport were inhibited by a short exposure (4 min) to deoxyglucose (15 mM) and cyanide (2 mM), added simultaneously. This appeared to produce a small, but not statistically significant (P = 0.056) rise in [Mg2+]i, presumably resulting from Mg2+ liberated following consumption of MgATP. Inhibition of the Na pump by strophanthidin (20 microM), followed by removal of external Na in the presence of strophanthidin, caused an increase in both [Mg2+]i and [Ca2+]i. The time course of changes in the two ions were dissimilar, so it seems unlikely that the observed rise in the [Mg2+]i was due solely to a direct effect of [Ca2+]i on Mag-fura-2 or due to the displacement of [Mg2+]i by Ca from binding sites. Evidence is presented that suggests sodium-magnesium exchange plays a role in the regulation of myocyte [Mg2+]i.
本文描述了从绵羊心脏的隔缘（节制）束中分离出耐钙心肌细胞的胶原酶分散制备方法。获得的细胞为小棒状（85×9微米），具有心肌细胞特有的明显条纹。分离的细胞加载了荧光离子敏感探针Mag - fura - 2或fura - 2，分别用于显微荧光光谱实验以测量胞质游离镁（[Mg2 + ]i）和钙（[Ca2 + ]i）；分离后细胞在长达8小时内仍可使用。通过同时加入脱氧葡萄糖（15 mM）和氰化物（2 mM）短时间暴露（4分钟）来抑制糖酵解和电子传递。这似乎导致[Mg2 + ]i出现小幅升高，但无统计学意义（P = 0.056），推测这是由于MgATP消耗后释放出Mg2 + 所致。毒毛花苷（20 microM）抑制钠泵，然后在毒毛花苷存在下去除细胞外钠，导致[Mg2 + ]i和[Ca2 + ]i均升高。两种离子变化的时间进程不同，所以观察到的[Mg2 + ]i升高似乎不太可能仅仅是由于[Ca2 + ]i对Mag - fura - 2的直接作用或Ca从结合位点置换[Mg2 + ]i所致。有证据表明钠 - 镁交换在心肌细胞[Mg2 + ]i的调节中起作用。
