Nachshen D A
J Gen Physiol. 1984 Jun;83(6):941-67. doi: 10.1085/jgp.83.6.941.
K-stimulated (voltage-dependent) influx of 45Ca was measured in synaptosomes (isolated presynaptic nerve terminals) from rat brain. Influx was terminated at 1 s with a rapid-filtration technique, so that most of the Ca uptake was mediated by inactivating ("fast") Ca channels (Nachshen, D. A., and Blaustein, M. P., 1980, J. Gen. Physiol., 76:709-728). This influx was blocked by multivalent cations with half-inhibition constants (K1) that clustered in three distinct groups: (a) K1 greater than 1 mM (Mg2+, Sr2+, and Ba2+); (b) K1 = 30-100 microM (Mn2+, Co2+, Ni2+, Cu2+, Zn2+, and Hg2+); (c) K1 less than 1 micro M (Cd2+, Y3+, La3+ and the trivalent lanthanides, and Pb2+). Most of these ions had very little effect on synaptosome steady state membrane potential, which was monitored with a voltage-sensitive fluorescent dye, or on the voltage dependence of Ca influx, which was assessed by measuring voltage-dependent Ca uptake at two levels of depolarization. The blockers inhibited Ca influx by competing with Ca for the channel site that is involved in the transport of divalent cations. Onset of fast channel inhibition by Mg, Co, Ni, Cu, Zn, Cd, La, Hg, and Pb was rapid, occurring within 1 s; inhibition was similar after 1 s or 30 min of exposure to these ions. The inhibition produced by Co, Cu, Zn, Cd, La, and Pb could be substantially reversed within 1 s by removing the inhibitory cation. The relative efficacies of the lanthanides as fast channel blockers were compared; there was a decrease in inhibitory potency with decreasing ionic radius. A model of the Ca channel binding site is considered, in which inhibitory polyvalent cation selectivity is determined primarily by coulombic interactions between the binding site and the different cations. The site is envisaged as consisting of two anions (radius 1 A) with a separation of 2 A between them. Small cations are unable to bind effectively to both anions. The selectivity sequences predicted for the alkaline earth cations, lanthanides, and transition metals are in substantial agreement with the selectivity sequences observed for inhibition of the fast Ca channel.
在大鼠脑突触体（分离出的突触前神经末梢）中测量了钾离子刺激（电压依赖性）的45Ca内流。采用快速过滤技术在1秒时终止内流，这样大部分的钙摄取是由失活的（“快速”）钙通道介导的（Nachshen, D. A., 和Blaustein, M. P., 1980, J. Gen. Physiol., 76:709 - 728）。这种内流被多价阳离子阻断，其半抑制常数（K1）聚集成三个不同的组：（a）K1大于1 mM（Mg2 +、Sr2 +和Ba2 +）；（b）K1 = 30 - 100 microM（Mn2 +、Co2 +、Ni2 +、Cu2 +、Zn2 +和Hg2 +）；（c）K1小于1 microM（Cd2 +、Y3 +、La3 +和三价镧系元素以及Pb2 +）。这些离子中的大多数对突触体稳态膜电位影响很小，膜电位用电压敏感荧光染料监测，对钙内流的电压依赖性影响也很小，钙内流的电压依赖性通过在两个去极化水平测量电压依赖性钙摄取来评估。这些阻滞剂通过与钙竞争参与二价阳离子转运的通道位点来抑制钙内流。Mg、Co、Ni、Cu、Zn、Cd、La、Hg和Pb对快速通道的抑制作用起效迅速，在1秒内发生；在暴露于这些离子1秒或30分钟后抑制作用相似。通过去除抑制性阳离子，Co、Cu、Zn、Cd、La和Pb产生的抑制作用可在1秒内基本逆转。比较了镧系元素作为快速通道阻滞剂的相对效力；随着离子半径减小，抑制效力降低。考虑了一个钙通道结合位点模型，其中抑制性多价阳离子选择性主要由结合位点与不同阳离子之间的库仑相互作用决定。该位点设想由两个阴离子（半径1 Å）组成，它们之间的间距为2 Å。小阳离子不能有效地与两个阴离子结合。预测的碱土金属阳离子、镧系元素和过渡金属的选择性序列与观察到的快速钙通道抑制的选择性序列基本一致。
