Regulation of the choline transport system in superfused microcarrier cultures of BeWo cells

BeWo choriocarcinoma cells were cultured onto solid microcarrier beads, packed into syringe barrels and superfused. The unidirectional choline uptake across the microvillous membrane of the cells was measured by a rapid single-circulation paired-tracer dilution procedure using methyl[H-3]choline with D-[C-14]mannitol as the extracellular reference molecule. Choline influx was saturable with a K-t of 214 +/- 15 mu m and a V-max of 45.29 +/- 0.94 nmol/min/mg of cell protein. Uptake of labelled choline was partially inhibited by nicotine, strongly inhibited by hemicolinium-3, and was reduced by about 50 per cent in sodium-free perfusates. A range of agents was added to the stirrer flasks 24 h prior to the experiments to determine if intracellular or extracellular levels of choline or its metabolic product, acetylcholine, regulated choline uptake. Pre-incubation with 2 mM choline reduced the choline maximal uptake by half, while pre-incubation with 100 mu M alpha-NETA [2-(alpha-naphthoyl)ethyltrimethyl-ammonium] reduced the influx by 77 per cent. Choline influx was also reduced to about half in the presence of 100 mu M vesamicol, bethanecol or neostigmine. It is concluded that BeWo cells possess a choline transporter similar to that described in isolated cytotrophoblasts and syncytiotrophoblast microvillous membrane preparations, and that uptake appeared to be regulated by both intracellular and extracellular concentrations of choline and acetylcholine. Therefore, these cells provide a novel model for studying the role of acetylcholine in human placenta. (C) 1998 W. B. Saunders Company Ltd.