IMIDAZOLINE BINDING-SITES IN HUMAN PLACENTA - EVIDENCE FOR HETEROGENEITY AND A SEARCH FOR PHYSIOLOGICAL-FUNCTION

1 An alpha-2-adrenoceptor antagonist, idazoxan, that binds to both alpha-2-adrenoceptors and to imidazoline sites (IR), has been used to characterize human placental IR. Human placenta is shown to be the richest source of IR (1800 +/- 100 fmol mg-1 protein; K(d) 38.9 +/- 3.4 nm). 2 Primary cells derived from human placenta and grown in monolayers, also displayed a high density of receptors (3209 +/- 136 fmol mg-1 in cytotrophoblasts and 3642 +/- 144 fmol mg-1 protein in syncytiotrophoblast enriched cell culture). 3 [H-3]-idazoxan did not show binding characteristics of alpha-2-adrenoceptors in human placental membranes or human trophoblastic cells, thus making it a ligand of choice to study the imidazoline site. The tissue appeared to be lacking alpha-2-adrenoceptors in that other alpha-2-adrenoceptor ligands, [H-3]-rauwolscine and [H-3]-clonidine, do not bind to alpha-2-adrenoceptors in human placenta. 4 IRs are localized on the cell surface, as determined by the release of bound [H-3]-idazoxan from cells, when washed with high ionic/acidic medium. 5 Imidazoline receptors of human placenta display high affinity for amiloride (72 +/- 27 nm). The high affinity was used as a criterion to classify IR to IR(a) subtype (placenta, rabbit kidney, rabbit liver and rabbit adipose cells) as opposed to the IR(b) subtype which display low affinity for amiloride (> 2-mu-M, in all the other tissues). 6 Several novel ligands comprising a guanido functional group attached to an aromatic residue (e.g. benziliden-amino-guanidine (BAG), guanido pyrrole) display pronounced selectivity for IR over the alpha-2-adrenoceptors as the affinity of BAG is about 40 fold higher (K(d) = 18.9 +/- 13.8 nM in human placenta), than the affinity for alpha-2-adrenoceptors (K(d) = 768 +/- 299 nM in human platelets). Imidazoline sites bind selectively BAG and other guanido ligands thus indicating a distinct structural requirement at its site of binding. 7 K+ channel blockers and monovalent ions (e.g. Cs+ and NH4+) interfere with idazoxan binding to IR, indicating a possible involvement of IR in K+ transport.