艾氏腹水癌细胞中钠离子依赖性氨基酸协同转运的能量学

Energetics of Na+-dependent amino acid co-transport in Ehrlich ascites tumor cells.

作者信息

Dawson W D, Smith T C

出版信息

Biochim Biophys Acta. 1987 Feb 12;897(1):5-13. doi: 10.1016/0005-2736(87)90309-9.

Abstract

The energy available from the Na+ electrochemical potential gradient (delta mu Na) has been evaluated in Ehrlich ascites tumor cells during accumulation of 2-aminoisobutyric acid. Cells were incubated in media of varying [Na+] (25-154 mM) in the presence of 0.25 mM 2-aminoisobutyric acid to establish maximum steady-state accumulation of the amino acid. Membrane potential (Vm) and intracellular Na+ activity (aNa) were estimated using standard electrophysiological techniques. In physiological saline ([Na+] = 154 mM) aNa is 4.4 +/- 0.6 mM, giving an apparent Na+ activity coefficient (gamma app) in the cytoplasm of 0.17 +/- 0.02. Vm under these conditions is -20.8 +/- 2.1 mV. From these values, delta mu Na = 9.9 +/- 0.8 kJ/mol. Concomitant determinations of 2-aminoisobutyric acid (AIB) accumulation show an energy requirement (delta mu AIB) of 8.5 +/- 0.5 kJ/mol. Stepwise reductions in extracellular [Na+] give parallel reductions in aNa, Vm and 2-aminoisobutyric acid accumulation. However, under all conditions tested the energy available from the Na+ electrochemical potential gradient exceeds that needed to drive 2-aminoisobutyric acid uptake. The effects of 2-aminoisobutyric acid on Vm have also been determined. Addition of AIB (10 mM) to steady-state cells leads to membrane depolarization (resting Vm = -22.1 +/- 1.3 mV; plus AIB Vm = -16.2 +/- 1.2 mV) within 1 min. Subsequent repolarization of the membrane to resting levels occurs within 10 min. The repolarization phase is blocked in the presence of ouabain (2 mM). The results establish that the energy available from the Na+ gradient is sufficient to serve as a source for 2-aminoisobutyric acid accumulation.

摘要

在2-氨基异丁酸积累过程中，对艾氏腹水瘤细胞中由Na⁺电化学势梯度（δμNa）提供的能量进行了评估。将细胞在含有0.25 mM 2-氨基异丁酸的不同[Na⁺]（25 - 154 mM）培养基中孵育，以建立该氨基酸的最大稳态积累。使用标准电生理技术估算膜电位（Vm）和细胞内Na⁺活性（aNa）。在生理盐水中（[Na⁺] = 154 mM），aNa为4.4±0.6 mM，细胞质中的表观Na⁺活性系数（γapp）为0.17±0.02。在这些条件下Vm为 - 20.8±2.1 mV。根据这些值，δμNa = 9.9±0.8 kJ/mol。同时测定2-氨基异丁酸（AIB）积累显示能量需求（δμAIB）为8.5±0.5 kJ/mol。细胞外[Na⁺]的逐步降低导致aNa、Vm和2-氨基异丁酸积累平行降低。然而，在所有测试条件下，由Na⁺电化学势梯度提供的能量超过驱动2-氨基异丁酸摄取所需的能量。还确定了2-氨基异丁酸对Vm的影响。向稳态细胞中添加AIB（10 mM）会在1分钟内导致膜去极化（静息Vm = - 22.1±1.3 mV；添加AIB后Vm = - 16.2±1.2 mV）。随后膜在10分钟内重新极化至静息水平。在哇巴因（2 mM）存在下，复极化阶段被阻断。结果表明，由Na⁺梯度提供的能量足以作为2-氨基异丁酸积累的来源。