Na(+)-dependent and Na(+)-independent systems of choline transport by plasma membrane vesicles of A549 cell line.
Membrane vesicles of A549 lung cells accumulate choline by two pathways: the Na(+)-independent uphill uptake of choline [Michaelis-Menten constant (Km) approximately 44 microM; steady-state gradient approximately 45 at 5 microM external choline] is dependent on a transmembrane H+ gradient, is relatively insensitive to hemicholinium-3, is amiloride sensitive, and is abolished by valinomycin plus carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP). The Na(+)-dependent active choline uptake (Km approximately 4 microM, inhibitor constant for hemicholinium-3 approximately 0.1 microM), is specific for Na+, is amiloride and FCCP sensitive, and is electrogenic: the overshoot using K(+)-loaded vesicles and NaCl gradient was increased by valinomycin. The time of the overshoot peak, T was approximately 90 s in a NaSCN medium (or in presence of other lipid-soluble anions), a value close to that for alpha-aminoisobutyrate as substrate (T = approximately 1.5 min). T was lengthened in NaCl medium to approximately 10 min, and the overshoot was abolished by impermeant anions. External Cl- is not required for the choline uptake: valinomycin produced an overshoot in the presence of only impermeant anions, with T approximately 90 s. Most of the above properties are shared by the high-affinity Na(+)-dependent choline transport in synaptosomes. The characteristics of the Na(+)-dependent choline uptake by membrane vesicles of A549 cells are consistent with an electrogenic choline(+)-Na+ cotransport, with the rate-limiting anion (e.g., Cl-) influx balancing the positive charges transferred into the vesicles. The data are also consistent with an involvement of an amiloride-sensitive choline+/H+ antiport (or choline(+)-OH- symport) in the low- and high-affinity choline uptake pathways.
Institute for Environmental Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104.
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Kleinzeller
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Dodia
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Chander
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Fisher
D000838:Anions / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects; D002412:Cations / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002794:Choline / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008168:Lung / Q000166:cytology / Q000378:metabolism*; D012964:Sodium / Q000494:pharmacology*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism
D000838:Anions; D002412:Cations; D017923:Sodium-Hydrogen Exchangers; D012964:Sodium; D002794:Choline