Na+-gradient-dependent stimulation of renal transport of rho-aminohippurate.
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ABSTRACT:
SUBMITTER: Sheikh MI
PROVIDER: S-EPMC1153953 | biostudies-other | 1982 Oct
REPOSITORIES: biostudies-other
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Project description:The transport properties for phenylalanine and glucose in luminal-membrane vesicles from outer cortex (pars convoluta) and outer medulla (pars recta) of rabbit kidney were studied by a spectrophotometric method. Uptake of phenylalanine as well as of glucose by the two types of membrane vesicles was found to be Na+-dependent, electrogenic and stereospecific. Na+-dependent transport of L-phenylalanine by outer-cortical membrane vesicles could be accounted for by one transport system (KA congruent to 1.5 mM). By contrast, in the outer-medullary preparation, L-phenylalanine transport occurred via two transport systems, namely a high-affinity system with K1A congruent to 0.33 mM and a low-affinity system with K2A congruent to 7 mM respectively. Na+-dependent uptake of D-glucose by pars convoluta and pars recta membrane vesicles could be described by single, but different, transport systems, namely a low-affinity system with KA congruent to 3.5 mM and a high-affinity system with KA congruent to 0.30 mM respectively. Attempts to calculate the stoichiometry of the different Na+/D-glucose transport systems by using Hill-type plots revealed that the ratio of the Na+/hexose co-transport probably is 1:1 in the case of pars convoluta and 2:1 in membrane vesicles from pars recta. The Na+/L-phenylalanine stoichiometry of the pars convoluta transporter probably is 1:1. Both the high-affinity and the low-affinity Na+-dependent L-phenylalanine transport system of pars recta membrane vesicles seem to operate with a 1:1 stoichiometry. The physiological importance of the arrangement of low-affinity and high-affinity transport systems along the kidney proximal tubule is discussed.
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Project description:L-Carnitine transport was characterized in Xenopus laevis oocytes before and after injection of mRNA isolated from rat renal cortex. Non-injected oocytes revealed endogenous Na(+)-dependent transport of L-carnitine. After injection of 15 ng of rat kidney mRNA, the Na(+)-dependent L-carnitine transport increased 2-3-fold, reaching maximal activity after 5-6 days. The expressed carnitine transport was maximal at pH 7.5, whereas the endogenous transport showed no clear maximum between pH 6.0 and 8.5. Kinetic analysis revealed apparent Km values for L-carnitine of 66 microM for the endogenous and 149 microM for the expressed transport. Trimethyl-lysine and D-carnitine inhibited both the endogenous and the expressed transport. In contrast, L-acetylcarnitine, L-isovalerylcarnitine, L-palmitoylcarnitine and butyrobetaine inhibited predominantly the expressed transport, whereas glycinebetaine had no inhibitory effect on either transport system. Size-fractionated rat renal-cortex mRNA (median size 2 kb) induced a 3-fold higher L-carnitine transport than did unfractionated mRNA. These studies demonstrate that Xenopus laevis oocytes exhibit Na(+)-dependent L-carnitine transport and provide the basis for expression-cloning of a rat renal Na(+)-dependent L-carnitine transport system.
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Project description:A monoclonal antibody FD19 which removes reconstitutable Na(+)-dependent amino acid transport activity from solubilized bovine renal brush-border membrane vesicles was found to react specifically with the enzyme aminopeptidase N. Cleavage of aminopeptidase N from the membranes with papain inhibited Na(+)-dependent amino acid transport activity without affecting that of alpha-methyl D-glucoside. Removal of aminopeptidase substantially increased the Km values for the Na(+)-dependent transport of alanine, glutamine, leucine and phenylalanine without affecting the Vmax. Both Na(+)-dependent amino acid transport and aminopeptidase activity in intact vesicles were competitively inhibited by amino acids with very similar specificity. These results suggest that the amino acid-binding sites of aminopeptidase N and the transporter interact in some way to increase the Km of the transport process for its substrates. However, independent direct inactivation of the transport system by papain cannot be ruled out.
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