Ma J, González A, Chen R
Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106, USA.
J Gen Physiol. 1996 Sep;108(3):221-32. doi: 10.1085/jgp.108.3.221.
Dihydropyridine (DHP) receptors of the transverse tubule membrane play two roles in excitation-contraction coupling in skeletal muscle: (a) they function as the voltage sensor which undergoes fast transition to control release of calcium from sarcoplasmic reticulum, and (b) they provide the conducting unit of a slowly activating L-type calcium channel. To understand this dual function of the DHP receptor, we studied the effect of depolarizing conditioning pulse on the activation kinetics of the skeletal muscle DHP-sensitive calcium channels reconstituted into lipid bilayer membranes. Activation of the incorporated calcium channel was imposed by depolarizing test pulses from a holding potential of -80 mV. The gating kinetics of the channel was studied with ensemble averages of repeated episodes. Based on a first latency analysis, two distinct classes of channel openings occurred after depolarization: most had delayed latencies, distributed with a mode of 70 ms (slow gating); a small number of openings had short first latencies, < 12 ms (fast gating). A depolarizing conditioning pulse to +20 mV placed 200 ms before the test pulse (-10 mV), led to a significant increase in the activation rate of the ensemble averaged-current; the time constant of activation went from tau m = 110 ms (reference) to tau m = 45 ms after conditioning. This enhanced activation by the conditioning pulse was due to the increase in frequency of fast open events, which was a steep function of the intermediate voltage and the interval between the conditioning pulse and the test pulse. Additional analysis demonstrated that fast gating is the property of the same individual channels that normally gate slowly and that the channels adopt this property after a sojourn in the open state. The rapid secondary activation seen after depolarizing prepulses is not compatible with a linear activation model for the calcium channel, but is highly consistent with a cyclical model. A six-state cyclical model is proposed for the DHP-sensitive Ca channel, which pictures the normal pathway of activation of the calcium channel as two voltage-dependent steps in sequence, plus a voltage-independent step which is rate limiting. The model reproduced well the fast and slow gating models of the calcium channel, and the effects of conditioning pulses. It is possible that the voltage-sensitive gating transitions of the DHP receptor, which occur early in the calcium channel activation sequence, could underlie the role of the voltage sensor and yield the rapid excitation-contraction coupling in skeletal muscle, through either electrostatic or allosteric linkage to the ryanodine receptors/calcium release channels.
横管膜上的二氢吡啶（DHP）受体在骨骼肌兴奋 - 收缩偶联中发挥两种作用：（a）它们作为电压传感器，经历快速转变以控制肌浆网中钙的释放；（b）它们构成缓慢激活的L型钙通道的传导单位。为了理解DHP受体的这种双重功能，我们研究了去极化条件脉冲对重构到脂质双分子层膜中的骨骼肌DHP敏感钙通道激活动力学的影响。通过从 - 80 mV的保持电位进行去极化测试脉冲来激活掺入的钙通道。通道的门控动力学通过重复事件的总体平均值进行研究。基于首次潜伏期分析，去极化后出现两种不同类型的通道开放：大多数具有延迟潜伏期，分布模式为70 ms（慢门控）；少数开放具有短的首次潜伏期，<12 ms（快速门控）。在测试脉冲（ - 10 mV）前200 ms施加到 + 20 mV的去极化条件脉冲，导致总体平均电流的激活速率显著增加；激活的时间常数从τm = 110 ms（参考值）变为条件脉冲后的τm = 45 ms。条件脉冲引起的这种激活增强是由于快速开放事件频率的增加，这是中间电压以及条件脉冲与测试脉冲之间间隔的陡峭函数。进一步分析表明，快速门控是通常缓慢门控的相同单个通道的特性，并且通道在处于开放状态一段时间后采用这种特性。去极化预脉冲后出现的快速二次激活与钙通道的线性激活模型不相符，但与循环模型高度一致。提出了一个用于DHP敏感钙通道的六态循环模型，该模型将钙通道激活的正常途径描绘为依次的两个电压依赖性步骤，加上一个限速的电压非依赖性步骤。该模型很好地再现了钙通道的快速和慢门控模型以及条件脉冲的影响。DHP受体的电压敏感门控转变可能发生在钙通道激活序列的早期，它可能通过与兰尼碱受体/钙释放通道的静电或变构连接，成为电压传感器作用的基础，并在骨骼肌中产生快速的兴奋 - 收缩偶联。
