犬肾皮质膜囊泡中对氨基马尿酸盐的Na⁺和H⁺梯度依赖性转运

Na+ and H+ gradient-dependent transport of p-aminohippurate in membrane vesicles from dog kidney cortex.

作者信息

Russel F G, van der Linden P E, Vermeulen W G, Heijn M, van Os C H, van Ginneken C A

机构信息

Department of Pharmacology, University of Nijmegen, The Netherlands.

出版信息

Biochem Pharmacol. 1988 Jul 1;37(13):2639-49. doi: 10.1016/0006-2952(88)90258-4.

DOI:10.1016/0006-2952(88)90258-4

PMID:3390224

Abstract

The transport of p-aminohippurate (PAH) was studied in basolateral (BLMV) and brush border membrane vesicles (BBMV) isolated from dog kidney cortex. Imposition of an inwardly directed 100 mN Na+ gradient stimulated the uptake of 50 microM [3H]PAH into BLMV, whereas a pH gradient (pHout = 6.0, pHin = 7.4) only slightly enhanced uptake. The Na+ gradient-dependent uptake of PAH was electroneutral, saturable and sensitive to inhibition by probenecid and several anionic drugs, with (apparent) Km = 0.79 +/- 0.16 mM, Vmax = 0.80 +/- 0.05 nmol/mg protein, 15 sec and Ki for probenecid = 0.08 +/- 0.01 mM. Simultaneous imposition of the pH gradient (outward OH- gradient) and inward Na+ gradient stimulated PAH uptake significantly over that with an Na+ gradient alone. These results are consistent with an Na+ gradient-stimulated PAH/OH- exchange mechanism in the basolateral membrane. In BBMV, PAH uptake could be stimulated by an outwardly directed OH- gradient as well as an inward Na+ gradient. Both gradients could drive PAH transport via a mediated probenecid-sensitive pathway. Na+ gradient-stimulated uptake was electrogenic with a (apparent) Km = 4.93 +/- 0.57 mM, Vmax = 6.71 +/- 0.36 nmol/mg protein, 15 sec and Ki,prob = 0.13 +/- 0.01 mM. The kinetic parameters for PAH/OH- exchange were virtually the same, (apparent) Km = 5.72 +/- 0.49 mM, Vmax = 7.87 +/- 0.33 nmol/mg protein, 15 sec and Ki,prob = 0.16 +/- 0.02 mM. When both the Na+ and pH (outward OH-) gradient were simultaneously imposed an almost twofold stimulation in uptake was observed over that with either an Na+ or pH gradient alone. These results suggested that both gradients stimulate PAH transport in BBMV via the same pathway. However, inhibition experiments with various organic anions showed that the specificities of Na+ and pH gradient-stimulated PAH uptake do not entirely overlap. Thus, our results support a simple transport in BBMV, but it cannot be excluded that two separate pathways are involved.

摘要

对从犬肾皮质分离出的基底外侧膜囊泡（BLMV）和刷状缘膜囊泡（BBMV）中对氨基马尿酸（PAH）的转运进行了研究。施加内向的100 mN Na⁺梯度刺激了50 μM [³H]PAH进入BLMV的摄取，而pH梯度（pH外 = 6.0，pH内 = 7.4）仅略微增强了摄取。PAH的Na⁺梯度依赖性摄取是电中性的、可饱和的，并且对丙磺舒和几种阴离子药物的抑制敏感，（表观）Km = 0.79 ± 0.16 mM，Vmax = 0.80 ± 0.05 nmol/mg蛋白质，15秒，丙磺舒的Ki = 0.08 ± 0.01 mM。同时施加pH梯度（外向OH⁻梯度）和内向Na⁺梯度比单独使用Na⁺梯度时显著刺激了PAH的摄取。这些结果与基底外侧膜中Na⁺梯度刺激的PAH/OH⁻交换机制一致。在BBMV中，外向的OH⁻梯度以及内向的Na⁺梯度都可以刺激PAH的摄取。两种梯度都可以通过介导的对丙磺舒敏感的途径驱动PAH的转运。Na⁺梯度刺激的摄取是生电的，（表观）Km = 4.93 ± 0.57 mM，Vmax = 6.71 ± 0.36 nmol/mg蛋白质，15秒，Ki,prob = 0.13 ± 0.01 mM。PAH/OH⁻交换的动力学参数几乎相同，（表观）Km = 5.72 ± 0.49 mM，Vmax = 7.87 ± 0.33 nmol/mg蛋白质，15秒，Ki,prob = 0.16 ± 0.02 mM。当同时施加Na⁺和pH（外向OH⁻）梯度时，观察到摄取量比单独使用Na⁺或pH梯度时几乎增加了两倍。这些结果表明两种梯度通过相同的途径刺激BBMV中PAH的转运。然而，用各种有机阴离子进行的抑制实验表明，Na⁺和pH梯度刺激的PAH摄取的特异性并不完全重叠。因此，我们的结果支持BBMV中的一种简单转运，但不能排除涉及两条独立途径的可能性。