Terrar D A, White E
Department of Pharmacology, Oxford.
Q J Exp Physiol. 1989 Mar;74(2):121-39. doi: 10.1113/expphysiol.1989.sp003250.
Possible mechanisms for calcium entry at positive membrane potentials were investigated in single cells isolated from guinea-pig ventricular muscle. The cells were voltage clamped and contraction was measured by an optical technique. When prolonged (200 ms to 2 s) depolarizations at +60 mV were applied, contraction amplitude increased with pulse duration, in contrast to the contraction at 0 mV. When a 'pre-pulse' to 0 mV was applied for 200 ms to inactivate current through 'L-type' calcium channels, contraction nevertheless increased with membrane potential during a subsequent test pulse applied over the range -40 to +60 mV. Contraction during the test pulse at +60 mV was abolished when extracellular calcium was reduced to zero. This effect developed more rapidly than abolition of the contraction in response to the pre-pulse to 0 mV. Reduction of extracellular calcium from 2.5 to 1 mM reduced the contraction at +60 mV to a greater extent than that at 0 mV and caused an inward shift in the current at +60 mV. Nifedipine (5 microM) substantially reduced the contraction during the test pulse to 0 mV but had little effect on the contraction at +60 mV. Conversely, dodecylamine (20 microM) caused little or no decrease in the contraction at 0 mV but substantially reduced the contraction at +60 mV. Following a conditioning pre-pulse to 0 mV the contraction at +60 mV was not consistently reduced by exposure to 3 microM-ryanodine. The interpolation of a single 200 ms pulse to +60 mV in a train of pulses to 0 mV potentiated the following contraction to 0 mV. This potentiation decayed over the first four steps to 0 mV following an interpolated pulse and increased with the voltage of the interpolated pulse over the range -20 to +60 mV. Potentiation was abolished on exposure to 3 microM-ryanodine. These observations are consistent with entry of calcium at positive membrane potentials through voltage-dependent, non-inactivating pathways which are insensitive to nifedipine but inhibited by dodecylamine. The observations support the hypothesis that calcium entry via this mechanism may contribute, at least under some conditions, to the loading of intracellular stores of calcium during the late plateau of the action potential, and thus influence subsequent contraction. Calcium entry through Na+-Ca2+ exchange is a possibility which would allow calcium entry to increase over the range of membrane potentials at which contraction was increased. However, additional calcium entry through other nifedipine-insensitive pathways, such as calcium-activated non-selective channels, cannot be excluded.
在从豚鼠心室肌分离出的单细胞中，研究了在正膜电位下钙内流的可能机制。这些细胞进行电压钳制，并通过光学技术测量收缩情况。当施加持续时间较长（200毫秒至2秒）的+60毫伏去极化时，与0毫伏时的收缩情况相反，收缩幅度随脉冲持续时间增加。当施加一个200毫秒的0毫伏“预脉冲”以使通过“L型”钙通道的电流失活时，在随后施加于-40至+60毫伏范围内的测试脉冲期间，收缩仍随膜电位增加。当细胞外钙降至零时，+60毫伏测试脉冲期间的收缩消失。这种效应比响应0毫伏预脉冲时收缩的消失发展得更快。细胞外钙从2.5毫摩尔降至1毫摩尔时，+60毫伏时的收缩比0毫伏时的收缩减少得更多，并导致+60毫伏时电流向内偏移。硝苯地平（5微摩尔）显著降低了测试脉冲至0毫伏期间的收缩，但对+60毫伏时的收缩影响很小。相反，十二烷基胺（20微摩尔）在0毫伏时对收缩几乎没有或没有降低作用，但显著降低了+60毫伏时的收缩。在施加0毫伏的预处理预脉冲后，暴露于3微摩尔的ryanodine并不能始终如一地降低+60毫伏时的收缩。在一串0毫伏的脉冲中插入一个200毫秒的+60毫伏脉冲会增强随后至0毫伏的收缩。这种增强在插入脉冲后的前四个至0毫伏的步骤中衰减，并在-20至+60毫伏范围内随插入脉冲的电压增加。暴露于3微摩尔的ryanodine会消除这种增强。这些观察结果与钙在正膜电位下通过电压依赖性、非失活途径内流一致，这些途径对硝苯地平不敏感，但被十二烷基胺抑制。这些观察结果支持这样的假设，即至少在某些条件下，通过这种机制的钙内流可能有助于在动作电位晚期平台期期间细胞内钙储存的加载，从而影响随后的收缩。通过钠钙交换的钙内流是一种可能性，这将使钙内流在收缩增加的膜电位范围内增加。然而，并不能排除通过其他对硝苯地平不敏感的途径（如钙激活的非选择性通道）的额外钙内流。
