Klöckner U, Storck T, Conradt M, Stoffel W
Institute of Biochemistry, Medical Faculty of the University of Cologne, Germany.
J Biol Chem. 1993 Jul 15;268(20):14594-6.
The transport of L-glutamate into Xenopus laevis oocytes expressing the cloned L-glutamate/L-aspartate transporter (GLAST-1) from rat brain was studied using the voltage clamp technique. At a holding potential of -90 mV, a bath application of 100 microM L-glutamate induced an inward current (IGLAST) with an amplitude ranging from -5 to -30 nA. IGLAST did not require extracellular Ca2+, Mg2+, or Cl-, was larger at negative potentials, and did not reverse up to +80 mV. The current was dependent on external L-glutamate and Na+ with half-maximal amplitudes at 11 microM L-glutamate and 41 mM Na+. IGLAST saturated at 100 microM L-glutamate and 80 mM Na+. The Hill coefficient for Na+ and L-glutamate was 3.3 and 1.3, respectively, suggesting that 3 Na+ accompany the transport of 1 L-glutamate molecule. At low [Na+]o, IGLAST was enhanced by reducing [K+]o, an indication for the countertransport of K+. Reducing external pH from 7.4 to 6.0 did not change the amplitude of IGLAST. This argues against a glutamate/proton cotransport. The results provide evidence for GLAST-1 carrying out a high affinity, sodium-dependent L-glutamate transport with a proposed stoichiometry of 3 Na+, 1 L-glutamate-/1 K+.
利用电压钳技术研究了L-谷氨酸转运至表达来自大鼠脑克隆的L-谷氨酸/L-天冬氨酸转运体（GLAST-1）的非洲爪蟾卵母细胞中的过程。在-90 mV的钳制电位下，向浴槽中施加100 μM的L-谷氨酸可诱导出内向电流（IGLAST），其幅度在-5至-30 nA之间。IGLAST不需要细胞外的Ca2+、Mg2+或Cl-，在负电位时更大，并且在高达+80 mV时不会反转。该电流依赖于细胞外的L-谷氨酸和Na+，在11 μM的L-谷氨酸和41 mM的Na+时具有半数最大幅度。IGLAST在100 μM的L-谷氨酸和80 mM的Na+时达到饱和。Na+和L-谷氨酸的希尔系数分别为3.3和1.3，表明1个L-谷氨酸分子的转运伴随着3个Na+。在低[Na+]o时，降低[K+]o可增强IGLAST，这表明存在K+的反向转运。将细胞外pH从7.4降至6.0不会改变IGLAST的幅度。这排除了谷氨酸/质子共转运的可能性。这些结果为GLAST-1进行高亲和力、钠依赖性的L-谷氨酸转运提供了证据，推测其化学计量为3个Na+、1个L-谷氨酸-/1个K+。
