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 hormo nogenesis has been proposed. In accordance with its putative role, the aim of this study was to characterize the human placental syncytiotro phoblast receptivity to NPY. Thus we performed this study on brush-bor der membranes (BBM) and basal plasma membranes (BPM). Specific I-125-l abeled NPY (I-125-NPY) binding to BBM was rapid (20 min), saturable, w ith a maximum binding capacity of 604 +/- 100 fmol/mg protein, and of high affinity, with a dissociation constant of 11 +/- 3 nM. No saturab le binding could be shown in BPM. The rank order of affinity of NPY an d related peptides to compete for I-125-NPY binding sites was peptide YY (PYY) > NPY = [Leu(31),Pro(34)]NPY > 13-36NPY >> pancreatic polypep tide (PP). It is noteworthy that PYY displaced only 45% of the binding sites. In BBM, both NPY and PYY were potent phospholipase C (PLC) sti mulators, leading to a four- to fivefold increase of control phosphodi esterase activity. The latter effect could be prevented by preincubati on of membranes with 5 mu M U-73122, a known inhibitor of G protein-li nked receptor activation of PLC-beta. Furthermore, 5 mu M BIBP-3226, a Y-1-receptor antagonist, shifted both dose-response curves to the rig ht 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 phosphatidylinositol- 3 kinase (PI3K). Taken together, these data suggest that placental and blood-derived NPY binds to a mixed population of receptors composed o f Y-1 and Y-3 subtypes on the maternal side of the syncytiotrophoblast , where it can mediate its physiological purposes via PLC-beta and PI3 K activation, both of which lead to PKC activation. However, because B IBP-3226 antagonized both effects, the physiological relevance of the apparent Y-3 fraction is still unsolved.