Kameyama A, Yazawa K, Kaibara M, Ozono K, Kameyama M
Department of Physiology, Faculty of Medicine, Kagoshima University, Sakura-ga-oka, Kagoshima 890, Japan.
Pflugers Arch. 1997 Mar;433(5):547-56. doi: 10.1007/s004240050313.
Possible mechanisms for run-down in the Ca2+ channel, such as proteolysis or dephosphorylation of the channel, were examined in guinea-pig ventricular myocytes. The Ca2+ channel current, recorded in inside-out patches using a pipette solution containing 50 mM Ba2+ and 3 microM Bay K 8644, ran down with a mean survival time of 2.35 min. The survival time was not significantly affected by adenosine triphosphate (ATP) (3 mM), 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid (BAPTA) (2 mM), isoprenaline (l-5 microM), phosphate (l20 mM) and leupeptin (l0 microM). Stimulation of guanosine triphosphate (GTP)-binding proteins was also ineffective. The catalytic subunit of adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase (PKA, 0.5-2 microM) slightly and transiently increased channel activity, but had minimal effects on the channel when applied after complete run-down. On the other hand, cytoplasm from the heart, skeletal muscle, brain and liver, but not kidney, induced channel activity. There was a positive correlation between NPo (the product of the number of channels N and the open probability Po) value before run-down and that after the application of cytoplasm, suggesting that the activity of once-active channels was restored ba the exogenous cytoplasm. The potency of cytoplasm in tissues in inducing channel activity was not related to PKA activity nor to the number of dihydropyridine binding sites. These results suggest that the run-down of the cardiac Ca2+ channel is not mediated by dephosphorylation or proteolysis of the channel, but involves other factor(s), possibly interaction of the channel protein with a cytoplasmic regulatory protein.
在豚鼠心室肌细胞中研究了钙通道功能衰退的可能机制，如通道的蛋白水解或去磷酸化。使用含有50 mM Ba2+和3 microM Bay K 8644的移液管溶液在膜内面向外的膜片中记录钙通道电流，该电流以平均存活时间2.35分钟逐渐衰退。存活时间不受三磷酸腺苷（ATP）（3 mM）、1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸（BAPTA）（2 mM）、异丙肾上腺素（1-5 microM）、磷酸盐（120 mM）和亮抑酶肽（10 microM）的显著影响。对鸟苷三磷酸（GTP）结合蛋白的刺激也无效。3',5'-环磷酸腺苷（cAMP）依赖性蛋白激酶（PKA，0.5-2 microM）的催化亚基轻微且短暂地增加了通道活性，但在完全衰退后应用时对通道的影响最小。另一方面，心脏、骨骼肌、脑和肝脏而非肾脏的细胞质可诱导通道活性。衰退前的NPo（通道数量N与开放概率Po的乘积）值与应用细胞质后的NPo值呈正相关，表明曾经活跃的通道活性通过外源性细胞质得以恢复。组织中细胞质诱导通道活性的效力与PKA活性和二氢吡啶结合位点的数量均无关。这些结果表明，心脏钙通道的衰退不是由通道的去磷酸化或蛋白水解介导的，而是涉及其他因素，可能是通道蛋白与细胞质调节蛋白的相互作用。
