Hove-Madsen L, Tort L
Departamento de Biologia Cellular i Fisiologia, Facultat de Ciencies, Universitat Autònoma de Barcelona, Spain.
Pflugers Arch. 2001 Feb;441(5):701-8. doi: 10.1007/s004240000470.
The whole-cell patch-clamp technique combined with rapid caffeine (CAF) applications was used to measure Na+-Ca2+ exchange (NCX) currents (I(NCX)). The rate of Ca2+ extrusion and the amount of Ca2+ extruded from the cell upon a rapid CAF exposure were obtained from I(NCX) and its time integral, respectively. This gave a maximal NCX rate (V(NCX)) of 151 amol pF(-1) s(-1) or 2.3 mM s(-1) and a half-maximal V(NCX) (K0.5) at a total cellular [Ca2+] ([Ca2+]tot) of 15.4 amol pF(-1). Using the same approach for the tail current induced by repolarization to -80 mV gave a K0.5 of 7.0 amol pF(-1) corresponding to 108 microM total or 2-4 microM free Ca2+. The relationship between [Ca2+]tot and V(NCX) was linear in the physiological range. Inhibition of the SR function with cyclopiazonic acid plus ryanodine reduced the slope significantly from 23.2+/-1.4 to 17.6+/-1.6 s(-1), while ryanodine alone had no effect. The relationship between [Ca2+]tot and V(NCX) was steeper at more negative membrane potentials, and with identical SR Ca2+ loads the maximal VNCX at -10 mV was reduced to 39.7+/-2.7% of the value at -90 mV. Long depolarizations caused SR Ca2+ loading through reverse-mode NCX. Between -30 and +10 mV reverse mode V(NCX)=Vm.0.047 amol pF(-1) s(-1) mV(-1)+2.51 amol pF(-1) s(-1), giving a reversal potential of -54 mV. In conclusion, the relationship between V(NCX) and [Ca2+]tot shows that the NCX is capable of removing a total Ca2+ transient of 60 microM at physiological heart rates, while reverse-mode NCX reloads the sarcoplasmic reticulum (SR) during depolarization. Furthermore, small alterations in the action potential configuration are predicted to change significantly the relative importance of the NCX in the regulation of cytosolic [Ca2+] and SR Ca2+ loading.
采用全细胞膜片钳技术结合快速施加咖啡因（CAF）来测量钠钙交换（NCX）电流（I(NCX)）。Ca2+外排速率以及快速暴露于CAF后从细胞中挤出的Ca2+量分别从I(NCX)及其时间积分中获得。由此得到最大NCX速率（V(NCX)）为151 amol pF(-1) s(-1)或2.3 mM s(-1)，在总细胞[Ca2+]（[Ca2+]tot）为15.4 amol pF(-1)时的半最大V(NCX)（K0.5）。对复极化至 -80 mV所诱导的尾电流采用相同方法，得到的K0.5为7.0 amol pF(-1)，对应于108 microM总Ca2+或2 - 4 microM游离Ca2+。在生理范围内，[Ca2+]tot与V(NCX)之间的关系呈线性。用环匹阿尼酸加ryanodine抑制肌浆网（SR）功能可使斜率从23.2±1.4显著降低至17.6±1.6 s(-1)，而单独使用ryanodine则无影响。在更负的膜电位下，[Ca2+]tot与V(NCX)之间的关系更陡峭，并且在相同的SR Ca2+负载下，-10 mV时的最大VNCX降至 -90 mV时值的39.7±2.7%。长时间去极化通过反向模式NCX导致SR Ca2+负载。在 -30至 +10 mV之间，反向模式V(NCX)=Vm.0.047 amol pF(-1) s(-1) mV(-1)+2.51 amol pF(-1) s(-1)，给出的反转电位为 -54 mV。总之，V(NCX)与[Ca2+]tot之间的关系表明，在生理心率下，NCX能够去除60 microM的总Ca2+瞬变，而反向模式NCX在去极化过程中使肌浆网（SR）重新加载Ca2+。此外，预计动作电位构型的微小变化会显著改变NCX在调节胞质[Ca2+]和SR Ca2+负载中的相对重要性。
