Choline transport in human placental brush-border membrane vesicles

Abstract

Pathways for transport of choline by human placental epithelia were investigated using brush border membrane vesicles isolated by divalent cation precipitation. The presence of choline transport mechanisms mediating Na(+)-choline cotransport, choline/H+ exchange and facilitated diffusion were assessed from [3H]choline tracer flux measurements. The rate and magnitude of intravesicular choline accumulation was unaffected by the imposition of an inwardly directed Na+ gradient suggesting an absence of a mechanism mediating brush border membrane Na(+)-choline cotransport. The imposition of inside-acid or inside-alkaline pH gradients was observed to have no significant effect on choline uptake suggesting choline is not a substrate for placental epithelial organic cation/H+ exchange. Conditions favoring the development of an inside-negative K+ diffusion potential was observed to induce a concentrative accumulation of choline to levels exceeding equilibrium suggesting the presence of a conductive uptake pathway for choline in placental brush border membrane. Evidence to suggest conductive choline uptake resulted from a mediated transport process includes a demonstration of the counterflow phenomena, the concentration-dependent inhibition by hemicholinium-3 (IC50 approximately equal to 100 microM) and the saturable rate of conductive choline uptake (Km approximately equal to 300 microM, Vmax approximately equal to 30 nmol/mg per min). Substrate specificity studies of the mechanism mediating conductive choline uptake suggest the interaction of choline with the transport protein occurs at a minimum of two sites: a site of negativity with the positively charged nitrogen group and a site of hydrogen bonding to the primary alcohol. Several commonly prescribed pharmaceuticals known to cross the placental barrier including imipramine, verapamil, propranolol, quinine, flurazepam, amiloride and ritodrin were observed to inhibit conductive choline uptake suggesting an interaction with the mechanism mediating conductive choline transport. Conductive choline uptake was unaffected by the presence of the basic amino acids lysine, arginine and histidine; the neurotransmitters serotonin, dopamine and histamine and the vitamins thiamine and carnitine which suggests the mechanism mediating conductive choline transport is not a pathway for placental uptake of these compounds.

The effect of a series of organic cations upon the plasmalemmal serotonin transporter, SERT

“…The diuretic amiloride is known to decrease the placental uptake of the organic cations choline (Grassl, 1994) and guanidine (Zevin et al, 1997). However, in the present study, both guanidine and choline did not modify 3 H-5HT transport by JAR cells.…”

Nicotine Transport in a Human Choriocarcinoma Cell Line (JAR)

“…10,17,18 Trans-stimulation of nicotine uptake was demonstrated after preloading JAR cells with unlabeled nicotine, indicating that nicotine uptake in JAR cells is carrier-mediated. Nicotine uptake was exquisitely sensitive to temperature and was almost abolished at 4°C, consistent with a carrier-mediated process.…”

Molecular and functional characterization of choline transporter in the human trophoblastic cell line JEG-3 cells

“…Furthermore, Muller et al found that JEG-3 cells express a low-affinity (K m value ¼ 206 mM) and Na þ -independent choline transport system that has not yet been identified at a molecular level [12]. In human placental brush-border membrane vesicles, the presence of choline transport mechanisms that mediate Na þ -choline co-transport, choline/H þ exchange and facilitated diffusion were assessed based on [ 3 H]choline tracer flux measurements [13]. Thus, there is no consensus regarding the functional expression of choline transporters in the BPB.…”