System A consists of three subtypes, sodium-coupled neutral amino acid transporter 1 (SNAT1), SNAT2, and SNAT4, which are all expressed in the placenta. The aim of this study was to evaluate the contributions of each of the three subtypes to total system A-mediated uptake in placental MVM of human and rat, using betaine and l-arginine as subtype-selective inhibitors of SNAT2 and SNAT4, respectively. Appropriate concentrations of betaine and l-arginine for subtype-selective inhibition in SNAT-overexpressing cells were identified. It was found that 10 mM betaine specifically and almost completely inhibited human and rat SNAT2-mediated [14C]α-methylaminoisobutyric acid ([14C]MeAIB) uptake, while 5 mM l-arginine specifically and completely inhibited [3H]glycine uptake via human SNAT4, as well as [14C]MeAIB uptake via rat SNAT4. In both human and rat placental MVM vesicles, sodium-dependent uptake of [14C]MeAIB was almost completely inhibited by 20 mM unlabeled MeAIB. l-Arginine (5 mM) partly inhibited the uptake in humans, but hardly affected that in rats. Betaine (10 mM) partly inhibited the uptake in rats, but hardly affected it in humans. These results suggest that SNAT1 is most likely the major contributor to system A-mediated MeAIB uptake by human and rat MVM vesicles and that the remaining uptake is mainly mediated by SNAT4 in humans and SNAT2 in rats. Thus, inhibition studies using betaine and l-arginine are useful to characterize the molecular mechanisms of system A-mediated transport.
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