Zhang X Q, Ng Y C, Moore R L, Musch T I, Cheung J Y
Department of Medicine, Milton S. Hershey Medical Center, Pennsylvania State University, Hershey, Pennsylvania 17033, USA.
J Appl Physiol (1985). 1999 Dec;87(6):2143-50. doi: 10.1152/jappl.1999.87.6.2143.
Previous studies have shown lower systolic intracellular Ca(2+) concentrations (Ca(2+)) and reduced sarcoplasmic reticulum (SR)-releasable Ca(2+) contents in myocytes isolated from rat hearts 3 wk after moderate myocardial infarction (MI). Ca(2+) entry via L-type Ca(2+) channels was normal, but that via reverse Na(+)/Ca(2+) exchange was depressed in 3-wk MI myocytes. To elucidate mechanisms of reduced SR Ca(2+) contents in MI myocytes, we measured SR Ca(2+) uptake and SR Ca(2+) leak in situ, i.e., in intact cardiac myocytes. For sham and MI myocytes, we first demonstrated that caffeine application to release SR Ca(2+) and inhibit SR Ca(2+) uptake resulted in a 10-fold prolongation of half-time (t(1/2)) of Ca(2+) transient decline compared with that measured during a normal twitch. These observations indicate that early decline of the Ca(2+) transient during a twitch in rat myocytes was primarily mediated by SR Ca(2+)-ATPase and that the t(1/2) of Ca(2+) decline is a measure of SR Ca(2+) uptake in situ. At 5.0 mM extracellular Ca(2+), systolic Ca(2+) was significantly (P </= 0.05) lower (337 +/- 11 and 416 +/- 18 nM in MI and sham, respectively) and t(1/2) of Ca(2+) decline was significantly longer (0.306 +/- 0.014 and 0.258 +/- 0.014 s in MI and sham, respectively) in MI myocytes. The 19% prolongation of t(1/2) of Ca(2+) decline was associated with a 23% reduction in SR Ca(2+)-ATPase expression (detected by immunoblotting) in MI myocytes. SR Ca(2+) leak was measured by a novel electrophysiological technique that did not require assigning empirical constants for intracellular Ca(2+) buffering. SR Ca(2+) leak rate was not different between sham and MI myocytes: the time constants of SR Ca(2+) loss after thapsigargin were 290 and 268 s, respectively. We conclude that, independent of decreased SR filling by Ca(2+) influx, the lower SR Ca(2+) content in MI myocytes was due to reduced SR Ca(2+) uptake and SR Ca(2+)-ATPase expression, but not to enhanced SR Ca(2+) leak.
先前的研究表明，在中度心肌梗死（MI）3周后从大鼠心脏分离的心肌细胞中，收缩期细胞内Ca(2+)浓度（Ca(2+)）较低，肌浆网（SR）可释放的Ca(2+)含量减少。通过L型Ca(2+)通道的Ca(2+)内流正常，但在MI 3周的心肌细胞中，通过反向Na(+)/Ca(2+)交换的Ca(2+)内流受到抑制。为了阐明MI心肌细胞中SR Ca(2+)含量降低的机制，我们在完整的心肌细胞中原位测量了SR Ca(2+)摄取和SR Ca(2+)泄漏。对于假手术组和MI组的心肌细胞，我们首先证明，应用咖啡因释放SR Ca(2+)并抑制SR Ca(2+)摄取，导致Ca(2+)瞬态下降的半衰期（t(1/2)）比正常收缩期测量值延长了10倍。这些观察结果表明，大鼠心肌细胞收缩期Ca(2+)瞬态的早期下降主要由SR Ca(2+)-ATP酶介导，并且Ca(2+)下降的t(1/2)是原位SR Ca(2+)摄取的一个指标。在细胞外Ca(2+)浓度为5.0 mM时，MI心肌细胞的收缩期Ca(2+)显著（P≤0.05）较低（MI组和假手术组分别为337±11和416±18 nM），并且Ca(2+)下降的t(1/2)显著更长（MI组和假手术组分别为0.306±0.014和0.258±0.014 s）。Ca(2+)下降t(1/2)延长19%与MI心肌细胞中SR Ca(2+)-ATP酶表达降低23%（通过免疫印迹检测）相关。SR Ca(2+)泄漏通过一种新型电生理技术进行测量，该技术不需要为细胞内Ca(2+)缓冲指定经验常数。假手术组和MI组心肌细胞的SR Ca(2+)泄漏率没有差异：毒胡萝卜素处理后SR Ca(2+)损失的时间常数分别为290和268 s。我们得出结论，与Ca(2+)内流导致的SR充盈减少无关，MI心肌细胞中较低的SR Ca(2+)含量是由于SR Ca(2+)摄取和SR Ca(2+)-ATP酶表达降低，而不是由于SR Ca(2+)泄漏增加。
