Imidazoline binding sites in human placenta: evidence for heterogeneity and a search for physiological function

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Abstract
1 An α2-adrenoceptor antagonist, idazoxan, that binds to both α2-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; Kd 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 [3H]-idazoxan did not show binding characteristics of α2-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 α2-adrenoceptors in that other α2-adrenoceptor ligands, [3H]-rauwolscine and [3H]-clonidine, do not bind to α2-adrenoceptors in human placenta. 4 IRs are localized on the cell surface, as determined by the release of bound [3H]-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 IRa subtype (placenta, rabbit kidney, rabbit liver and rabbit adipose cells) as opposed to the IRb subtype which display low affinity for amiloride (> 2 μm, 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 pyrole) display pronounced selectivity for IR over the α2-adrenoceptors as the affinity of BAG is about 40 fold higher (Kd = 18.9 ± 13.8 nm in human placenta), than the affinity for α2-adrenoceptors (Kd = 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.