Hori M, Sato H, Kitakaze M, Iwai K, Takeda H, Inoue M, Kamada T
First Department of Medicine, Osaka University School of Medicine, Suita, Japan.
Circ Res. 1994 Aug;75(2):324-34. doi: 10.1161/01.res.75.2.324.
Catecholamine cardiotoxicity is attributable in part to Ca2+ overload. To test whether the cytoskeletal structures of microtubules in cardiomyocytes are reversibly injured by catecholamine through excessive Ca2+ influx, morphological changes in the microtubules of neonatal rat myocytes were studied by immunohistochemical technique during exposure to norepinephrine (NE). In intact myocytes, microtubules appeared as a filamentous network throughout the cytoplasm and around the nucleus. NE exposure (10 mumol/L) for > 30 minutes elicited microtubular disassembly in a duration-dependent fashion without any irreversible change in sarcomere structure, and this abnormality recovered within 24 hours after cessation of stimulation. Microtubular disruption scores obtained by semiquantitative assessment were significantly increased in a dose-dependent manner (10.8 +/- 4.0 in the control condition, 23.4 +/- 4.7 at 60 minutes with 10 mumol/L NE), whereas they were significantly attenuated by pretreatment with propranolol (100 mumol/L; score, 11.8 +/- 3.3) but not with phentolamine (100 mumol/L; score, 26.4 +/- 4.8). Isoproterenol (1 mumol/L) and denopamine (10 mumol/L) mimicked the effects of NE, but phenylephrine did not, indicating that NE-induced microtubular disassembly is mediated by beta 1-adrenergic receptor stimulation. This beta-adrenergic receptor-mediated insult was significantly attenuated by a decrease in Ca2+ concentration in the medium from 2 to 0.5 mmol/L and by pretreatment with diltiazem (1 mumol/L). In contrast, microtubular disassembly was induced by an increase in Ca2+ concentration in the medium and an administration of the Ca2+ ionophore A23187, even without beta-adrenergic receptor stimulation. Involvement of intracellular hypoxia and activation of Ca(2+)-calmodulin-dependent kinase or Ca(2+)-dependent neutral protease were excluded from possible mechanisms; however, inhibition of tubulin polymerization by excessive Ca2+ influx during beta-adrenergic receptor stimulation may be primarily involved. We conclude that microtubular structures that support cellular integrity are reversibly injured by beta-adrenergic receptor stimulation through excessive Ca2+ influx.
儿茶酚胺心脏毒性部分归因于Ca2+超载。为了测试心肌细胞中微管的细胞骨架结构是否会因儿茶酚胺通过过量Ca2+内流而受到可逆性损伤，在暴露于去甲肾上腺素（NE）期间，采用免疫组织化学技术研究了新生大鼠心肌细胞微管的形态变化。在完整的心肌细胞中，微管呈现为贯穿整个细胞质和细胞核周围的丝状网络。暴露于NE（10 μmol/L）超过30分钟会以时间依赖性方式引发微管解聚，而肌节结构没有任何不可逆变化，并且这种异常在刺激停止后24小时内恢复。通过半定量评估获得的微管破坏评分以剂量依赖性方式显著增加（对照条件下为10.8±4.0，10 μmol/L NE处理60分钟时为23.4±4.7），而普萘洛尔（100 μmol/L）预处理可显著减轻该评分（评分为11.8±3.3），但酚妥拉明（100 μmol/L）预处理则不能（评分为26.4±4.8）。异丙肾上腺素（1 μmol/L）和多巴胺（10 μmol/L）模拟了NE的作用，但去氧肾上腺素则没有，这表明NE诱导的微管解聚是由β1 - 肾上腺素能受体刺激介导的。这种β - 肾上腺素能受体介导的损伤通过将培养基中Ca2+浓度从2 mmol/L降低到0.5 mmol/L以及用硫氮䓬酮（1 μmol/L）预处理而显著减轻。相反，即使没有β - 肾上腺素能受体刺激，培养基中Ca2+浓度的增加和Ca2+离子载体A23187的给药也会诱导微管解聚。细胞内缺氧以及Ca(2+)-钙调蛋白依赖性激酶或Ca(2+)-依赖性中性蛋白酶的激活被排除在可能的机制之外；然而，β - 肾上腺素能受体刺激期间过量Ca2+内流对微管蛋白聚合的抑制可能是主要原因。我们得出结论，支持细胞完整性的微管结构会因β - 肾上腺素能受体刺激通过过量Ca2+内流而受到可逆性损伤。
