Gambassi G
Laboratory of Cardiovascular Science, National Institute on Aging, National Institutes of Health, Baltimore, MD.
Cardiologia. 1993 Jan;38(1):25-36.
Myocardial ischemia is associated with an intracellular acidosis recovering after reperfusion. Alpha 1-adrenergic stimulation (alpha 1) exacerbates ischemic injury and triggers ventricular arrhythmias during the reperfusion phase. We have tested the hypothesis that the arrhythmogenic effect of alpha 1 is due to a cytosolic alkalinization secondary to proteinkinase C (PKC)-dependent activation of the sarcolemmal Na+/H+ exchanger. In addition, the effect of the 2 distinct receptor subtypes, alpha 1A and alpha 1B, has also been evaluated. We used single, enzymatically dissociated, adult rat ventricular myocytes. Cells were loaded with the ester derivative of the Ca2+ probe, indo-1 or with the intracellular pH probe SNARF-1. Fluorescence was monitored simultaneously with contractility and was taken as an index of intracellular [Ca2+] or as pHi value. Cells on a stage of an inverted microscope were superfused with a bicarbonate buffer continuously gassed with 95% O2 and 5% CO2 (pH = 7.37; Ca2+ 1.5 mM) and electrically stimulated at 0.5 Hz, at 25 degrees C. Alpha 1 (phenylephrine 50 microM + nadolol 1 microM) increased twitch amplitude and pHi (delta pHi = 0.05 +/- 0.01; pHi in control = 7.24 +/- 0.06, n = 10; p < 0.05). This effect was abolished by the PKC inhibitor staurosporine (5 nM), by overnight (12-24 hours) exposure to 0.2 microM phorbol esters and by the perfusion with 10 microM ethylisopropylamiloride (EIPA), a Na+/H+ exchange inhibitor. During 15 min of hypercarbic acidosis, achieved by switching to a buffer equilibrated with 85% O2 and 15% CO2 (pH = 7.01), alpha 1 had a more pronounced effect on pHi (delta pHi = 0.11 +/- 0.02; pHi in control = 7.02 +/- 0.04, n = 10; p < 0.05). During the 10 min following the removal of acidosis, alpha 1 induced aftercontractions in 75% (n = 20) versus only 25% (n = 8) of the cells in the absence of phenylephrine (p < 0.05). Superfusion with 10 microM EIPA (n = 6) abolished the occurrence of aftercontractions. Selective alpha 1A-adrenergic receptors stimulation (alpha 1 + 2 microM CEC, which inactivates alpha 1B receptors) further increased them (92%, n = 12) whereas selective alpha 1B-adrenergic receptors stimulation (alpha 1 + 2 microM WB-4101, a alpha 1A receptors antagonist) greatly reduced the occurrence of aftercontractions (11%, n = 18). These results show that the PKC-mediated activation of the Na+/H+ exchanger is the mechanism for the arrhythmias induced by alpha 1 during simulated reperfusion after a period of acidosis.(ABSTRACT TRUNCATED AT 400 WORDS)
心肌缺血与再灌注后细胞内酸中毒的恢复有关。α1肾上腺素能刺激（α1）会加重缺血性损伤，并在再灌注阶段引发室性心律失常。我们检验了这样一个假说：α1的致心律失常作用是由于蛋白激酶C（PKC）依赖性激活肌膜钠氢交换体后导致的胞质碱化。此外，还评估了两种不同受体亚型α1A和α1B的作用。我们使用了成年大鼠单个酶解心室肌细胞。细胞用Ca2+探针indo-1的酯衍生物或细胞内pH探针SNARF-1进行负载。荧光与收缩性同时监测，并作为细胞内[Ca2+]的指标或pHi值。倒置显微镜载物台上的细胞用持续通以95% O2和5% CO2的碳酸氢盐缓冲液灌流（pH = 7.37；Ca2+ 1.5 mM），并在25℃以0.5 Hz进行电刺激。α1（去氧肾上腺素50 μM + 纳多洛尔1 μM）增加了收缩幅度和pHi（ΔpHi = 0.05 ± 0.01；对照pHi = 7.24 ± 0.06，n = 10；p < 0.05）。PKC抑制剂星形孢菌素（5 nM）、过夜（12 - 24小时）暴露于0.2 μM佛波酯以及用10 μM乙基异丙基氨氯吡咪（EIPA，一种钠氢交换抑制剂）灌流均可消除此效应。在通过切换到用85% O2和15% CO2平衡的缓冲液实现的15分钟高碳酸性酸中毒期间（pH = 7.01），α1对pHi有更明显的作用（ΔpHi = 0.11 ± 0.02；对照pHi = 7.02 ± 0.04，n = 10；p < 0.05）。在去除酸中毒后的10分钟内，α1在75%（n = 20）的细胞中诱导了后收缩，而在无去氧肾上腺素的情况下仅25%（n = 8）的细胞出现后收缩（p < 0.05）。用10 μM EIPA灌流（n = 6）消除了后收缩的发生。选择性刺激α1A肾上腺素能受体（α1 + 2 μM CEC，可使α1B受体失活）进一步增加了后收缩（92%，n = 12），而选择性刺激α1B肾上腺素能受体（α1 + 2 μM WB - 4101，一种α1A受体拮抗剂）则大大减少了后收缩的发生（11%，n = 18）。这些结果表明，PKC介导的钠氢交换体激活是酸中毒一段时间后模拟再灌注期间α1诱导心律失常的机制。（摘要截断于400字）
