Lacerda A E, Brown A M
Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, Texas 77030.
J Gen Physiol. 1989 Jun;93(6):1243-73. doi: 10.1085/jgp.93.6.1243.
The hypothesis that dihydropyridine (DHP)-sensitive calcium channels have three distinct modes of gating has been examined. The major prediction is that the relative frequencies among modes depend on DHP concentration while the kinetics within a mode do not. We tested this by studying whole-cell and single-channel calcium currents in neonatal rat and adult guinea pig cardiac myocytes in different concentrations of several DHPs. In the absence of DHPs calcium currents declined with time but the kinetics, which are the focus of this study, were unchanged. Open-time frequency distributions had insignificant numbers of prolonged openings and were well fit by single tau's. Agonist DHP stereoisomers produced concentration-dependent changes in whole-cell tail current tau's. The frequency distribution of single calcium channel current open times became biexponential and the tau's were concentration dependent. The average number of openings per trace of channels with customary open times increased with increases in DHP concentration. Latencies to first opening for the customary openings and for prolonged openings were shorter in the presence of DHPs. A second larger conductance is another important feature of DHP-bound single calcium channels. Thus DHPs not only caused prolonged openings; they produced numerous changes in the kinetics of customary openings and increased channel conductance. It follows that these effects of DHPs do not support the hypothesis of modal gating of calcium channels. The mode model is not the only model excluded by the results; models in which DHPs are allowed to act only or mainly on open states are excluded, as are models in which the effects are restricted to inactivated states. We suggest a different type of model in which cooperative binding of DHPs at two sites produces the essential changes in kinetics and conductance.
二氢吡啶（DHP）敏感性钙通道具有三种不同门控模式的假说已得到检验。主要预测是，模式之间的相对频率取决于DHP浓度，而模式内的动力学则不受影响。我们通过研究新生大鼠和成年豚鼠心肌细胞在不同浓度几种DHP下的全细胞和单通道钙电流来对此进行测试。在没有DHP的情况下，钙电流随时间下降，但本研究重点关注的动力学保持不变。开放时间频率分布中延长开放的数量很少，并且能很好地用单时间常数拟合。激动剂DHP立体异构体使全细胞尾电流时间常数产生浓度依赖性变化。单个钙通道电流开放时间的频率分布变为双指数分布，且时间常数呈浓度依赖性。具有常规开放时间的通道每次记录的平均开放次数随DHP浓度增加而增加。在存在DHP的情况下，常规开放和延长开放的首次开放潜伏期较短。另一个较大的电导是与DHP结合的单个钙通道的另一个重要特征。因此，DHP不仅导致延长开放；它们还使常规开放的动力学产生了许多变化，并增加了通道电导。由此可见，DHP的这些作用不支持钙通道模式门控的假说。模式模型并不是结果排除的唯一模型；那些允许DHP仅作用于开放状态或主要作用于开放状态的模型也被排除，那些作用仅限于失活状态的模型同样被排除。我们提出了一种不同类型的模型，其中DHP在两个位点的协同结合产生了动力学和电导的本质变化。
