Côté K, Proteau S, Teijeira J, Rousseau E
Le Bilarium, Département de Physiologie et Biophysique, Université de Sherbrooke, Sherbrooke, QC, Canada.
J Mol Cell Cardiol. 2000 Nov;32(11):2051-63. doi: 10.1006/jmcc.2000.1236.
Since the role of sarcoplasmic reticulum (SR) in the E-C coupling of mammalian atrial cells has long been a subject of debate, biochemical, electrophysiological and immunological assays were performed in order to define and compare the properties of the Ca(2+)-release channel-ryanodine receptor (RyR)-from atrial and ventricular tissues. Cardiac SR preparations from human, canine and ovine tissues were compared using [(3)H]ryanodine binding, channel reconstitution into planar lipid bilayers and Western blot analysis involving RyR antibodies. [(3)H]ryanodine binding assays revealed a K(d)value of; 2.5 n M for all investigated cardiac tissues. Bound [(3)H]ryanodine was Ca(2+)-dependent with similar EC(50)values of 0.43, 0.49 and 0.79 microM for human atrium, canine ventricle and ovine atrium, respectively. However the density of binding sites was 4.5 times lower in atrial than in ventricular tissues. Beyond the presence of selective K(+)channels (gamma=188 pS) recorded in the SR enriched fraction of human atrium, the activity of a large conducting (gamma=671 pS) cationic channel was also observed. The latter displayed typical characteristics of Ca(2+)-release channels which were activated by 10 microM free [Ca(2+)] and 2 m M ATP. Western blot analysis revealed the presence of the RyR2 isoform in atrial and ventricular samples whereas no immunoreactivity was detected with specific RyR1 and RyR3 antibodies. Our results, obtained at the molecular level, are consistent with the presence of functional SR in human atrial cells. The human atrial Ca(2+)-release channel displays binding and regulating properties typical of the RyR2 isoform.
由于肌浆网（SR）在哺乳动物心房细胞兴奋 - 收缩偶联中的作用长期以来一直存在争议，因此进行了生化、电生理和免疫分析，以定义和比较心房和心室组织中钙释放通道 - 雷诺丁受体（RyR）的特性。使用[³H]雷诺丁结合、将通道重建到平面脂质双分子层以及涉及RyR抗体的蛋白质免疫印迹分析，对来自人类、犬类和绵羊组织的心脏SR制剂进行了比较。[³H]雷诺丁结合分析显示，所有研究的心脏组织的解离常数（Kd）值为2.5 nM。结合的[³H]雷诺丁依赖于Ca²⁺，人类心房、犬类心室和绵羊心房的半数有效浓度（EC50）值分别为0.43、0.49和0.79 μM。然而，心房组织中结合位点的密度比心室组织低4.5倍。除了在人类心房富含SR的部分记录到选择性钾通道（γ = 188 pS）外，还观察到一个大电导（γ = 671 pS）阳离子通道的活性。后者表现出钙释放通道的典型特征，被10 μM游离[Ca²⁺]和2 mM ATP激活。蛋白质免疫印迹分析显示，心房和心室样品中存在RyR2亚型，而用特异性RyR1和RyR3抗体未检测到免疫反应性。我们在分子水平上获得的结果与人类心房细胞中功能性SR的存在一致。人类心房钙释放通道表现出RyR2亚型典型的结合和调节特性。
