Hazama A, Loo D D, Wright E M
Department of Physiology, UCLA School of Medicine, Center for the Health Sciences, Los Angeles, CA 90095-1751, USA.
J Membr Biol. 1997 Jan 15;155(2):175-86. doi: 10.1007/s002329900169.
The rabbit Na+/glucose cotransporter (SGLT1) exhibits a presteady-state current after step changes in membrane voltage in the absence of sugar. These currents reflect voltage-dependent processes involved in cotransport, and provide insight on the partial reactions of the transport cycle. SGLT1 presteady-state currents were studied as a function of external Na+, membrane voltage Vm, phlorizin and temperature. Step changes in membrane voltage-from the holding Vh to test values, elicited transient currents that rose rapidly to a peak (at 3-4 msec), before decaying to the steady state, with time constants tau approximately 4-20 msec, and were blocked by phlorizin (Ki approximately 30 microm). The total charge Q was equal for the application of the voltage pulse and the subsequent removal, and was a function of Vm. The Q-V curves obeyed the Boltzmann relation: the maximal charge Qmax was 4-120 nC; V0.5, the voltage for 50% Qmax was -5 to +30 mV; and z, the apparent valence of the moveable charge, was 1. Qmax and z were independent of Vh (between 0 and -100 mV) and temperature (20-30 degrees C), while increasing temperature shifted V0.5 towards more negative values. Decreasing [Na+]o decreased Qmax, and shifted V0.5 to more negative voltages 9by -100 mV per 10-fold decrease in [Na+]o). The time constant tau was voltage dependent: the tau-V relations were bell-shaped, with maximal taumax 8-20 msec. Decreasing [Na+]o decreased taumax, and shifted the tau-V curves towards more negative voltages. Increasing temperature also shifted the tau-V curves, but did not affect taumax. The maximum temperature coefficient Q10 for tau was 3-4, and corresponds to an activation energy of 25 kcal/mole. Simulations of a 6-state ordered kinetic model for rabbit Na+/glucose cotransport indicate that charge-movements are due to Na+-binding/dissociation and a conformational change of the empty transporter. The model predicts that (i) transient currents rise to a peak before decay to steady-state; (ii) the tau-V relations are bell-shaped, and shift towards more negative voltages as [Na+]o is reduced; (iii) taumax is decreased with decreasing [Na+]o; and (iv) the Q-V relations are shifted towards negative voltages as [Na+]o is reduced. In general, the kinetic properties of the presteady-state currents are qualitatively predicted by the model.
在无糖情况下，兔钠/葡萄糖共转运体（SGLT1）在膜电压阶跃变化后会呈现出一个预稳态电流。这些电流反映了共转运过程中与电压相关的过程，并为转运循环的部分反应提供了见解。研究了SGLT1预稳态电流与细胞外钠离子浓度、膜电压Vm、根皮苷和温度的关系。膜电压从保持电压Vh阶跃到测试值时，会引发瞬态电流，该电流迅速上升至峰值（在3 - 4毫秒时），然后衰减至稳态，时间常数τ约为4 - 20毫秒，且被根皮苷阻断（抑制常数Ki约为30微摩尔）。施加电压脉冲和随后去除电压脉冲时的总电荷量Q相等，且是Vm的函数。Q - V曲线符合玻尔兹曼关系：最大电荷量Qmax为4 - 120纳库仑；V0.5（达到50% Qmax时的电压）为 - 5至 + 30毫伏；可移动电荷的表观价态z为1。Qmax和z与Vh（在0至 - 100毫伏之间）和温度（20 - 30摄氏度）无关，而升高温度会使V0.5向更负的值移动。降低细胞外钠离子浓度[Na + ]o会降低Qmax，并使V0.5向更负的电压移动（每10倍降低[Na + ]o，V0.5移动 - 100毫伏）。时间常数τ与电压有关：τ - V关系呈钟形，最大时间常数τmax为8 - 20毫秒。降低[Na + ]o会降低τmax，并使τ - V曲线向更负的电压移动。升高温度也会使τ - V曲线移动，但不影响τmax。τ的最大温度系数Q10为3 - 4，对应于25千卡/摩尔的活化能。对兔钠/葡萄糖共转运的六态有序动力学模型的模拟表明，电荷移动是由于钠离子结合/解离以及空转运体的构象变化。该模型预测：（i）瞬态电流在衰减至稳态之前上升至峰值；（ii）τ - V关系呈钟形，且随着[Na + ]o降低向更负的电压移动；（iii）随着[Na + ]o降低，τmax减小；（iv）随着[Na + ]o降低，Q - V关系向负电压移动。总体而言，该模型定性地预测了预稳态电流的动力学特性。
