Implication of ATP and sodium in arachidonic acid incorporation by placental syncytiotrophoblast brush border and basal plasma membranes in the human.

The human placental syncytiotrophoblast is the main site of exchange of nutrients and minerals between the mother and her fetus. In order to characterize the placental transport of some fatty acids, we studied the incorporation of arachidonic acid, a fetal primordial fatty acid, in purified bipolar syncytiotrophoblast brush border (BBM) and basal plasma membranes (BPM) from human placenta. The basal arachidonic acid incorporation in BBM and BPM was time dependent and reached maximal values of 0.75+/-0.10 and 0.48+/-0.18 pmol/mg protein, respectively, after 2.5 min. The presence of adenosine triphosphate (ATP) (3 m m) increases significantly the maximal incorporation of arachidonic acid by sixfold (4.75+/-0.35 pmol/mg) and ninefold (4.40+/-0.84 pmol/mg) in BBM and BPM, respectively. Moreover, an increase in the arachidonic acid incorporation was also obtained in the presence of sodium where the values achieved 7.68+/-0.98 (10x) and 6.53 pmol/mg (13.6x) for BBM and BPM, respectively. We also showed that the combination of both Na(+)and ATP increases significantly the maximal incorporation of arachidonic acid in BPM to 7.89+/-0.15 pmol/mg protein, while in BBM it did not modify its incorporation (8.18+/-0.25 pmol/mg protein), as compared to the presence of sodium alone. Our results demonstrate that arachidonic acid is incorporated by both placental syncytiotrophoblast membranes, and is ATP and sodium-linked. However, different mechanisms seem to be involved in this fatty acid incorporation through BBM and BPM, since the presence of Na(+)or ATP increased it, while the association of these two elements increased it only in BPM. We also demonstrated by osmolarity experiments that both membranes bind arachidonic acid, potentially involving one or more fatty acids binding proteins.

Human syncytiotrophoblast NPY receptors are located on BBM and activate PLC-to-PKC axis.

Neuropeptide Y (NPY) is abundant in plasma and amniotic fluid of women throughout pregnancy, during which its involvement in placental hormonogenesis has been proposed. In accordance with its putative role, the aim of this study was to characterize the human placental syncytiotrophoblast receptivity to NPY. Thus we performed this study on brush-border membranes (BBM) and basal plasma membranes (BPM). Specific 125I-labeled NPY (125I-NPY) binding to BBM was rapid (20 min), saturable, with a maximum binding capacity of 604 +/- 100 fmol/mg protein, and of high affinity, with a dissociation constant of 11 +/- 3 nM. No saturable binding could be shown in BPM. The rank order of affinity of NPY and related peptides to compete for 125I-NPY binding sites was peptides YY (PYY) > NPY = [Leu31,Pro34]NPY > 13-36NPY >> pancreatic polypeptide (PP). It is noteworthy that PYY displaced only 45% of the binding sites. In BBM, both NPY and PYY were potent phospholipase C (PLC) stimulators, leading to a four- to fivefold increase of control phosphodiesterase activity. The latter effect could be prevented by preincubation of membranes with 5 microM U-73122, a known inhibitor of G protein-linked receptor activation of PLC-beta. Furthermore, 5 microM BIBP-3226, a Y1-receptor antagonist, shifted both dose-response curves to the right in a similar fashion for both peptides. In accordance with the PLC stimulation, both peptides also induced stimulation of protein kinase C (PKC) activity, which could be partially but additively prevented by U-73122 and LY-294002, a selective inhibitor of phosphatidyl-inositol-3 kinase (PI3K). Taken together, these data suggest that placental and blood-derived NPY binds to a mixed population of receptors composed of Y1 and Y3 subtypes on the maternal side of the syncytiotrophoblast, where it can mediate its physiological purposes via PLC-beta and PI3K activation, both of which lead to PKC activation. However, because BIBP-3226 antagonized both effects, the physiological relevance of the apparent Y3 fraction is still unsolved.