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

Abstract

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). 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). [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. 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. 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). 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. 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.

Autoradiographic Comparison of [3H]-Clonidine Binding to Non-Adrenergic Sites and α2-Adrenergic Receptors in Human Brain

“…For instance, the correlation observed in Figure 2 may (Table 1). The possibility that the I-sites labeled by [ 3 H]-clonidine are actually I 2 sites also is unlikely because higher than expected affinities were observed for several agents (i.e., BDF6143 and rilmenidine affinities were 4-fold and 54-fold higher, respectively, than expected for human I 2 sites) ( Table 2) (Diamant et al 1992). Furthermore, [ 3 H]-clonidine binding was examined in human medullary slices with the goal of identifying a typical I 1 site (i.e., moxonidine-displaceable).…”

[3H]‐idazoxan binding to rabbit cerebral cortex recognises multiple imidazoline I2‐type receptors: pharmacological characterization and relationship to monoamine oxidase

“…Rabbit kidney, adipocytes and liver and human placenta all show a high affinity for amiloride whilst human kidney, guinea-pig ileum and rat liver have lower affinities for amiloride (Michel & Insel, 1989;Diamant et al, 1992). Many of the pharmacological differences in amiloride affinity can be attributed to species differences, for example, rabbit tissues generally show a higher affinity for amiloride than do tissues from rats, guinea-pigs and man (Michel & Insel, 1989;Diamant et al, 1992). However, recent evidence is emerging for two types of 12 sites in the same species, for example human liver, kidney and adipocytes show an approximately 50 fold lower affinity for amiloride than does the human placenta (Daimant et al, 1992).…”

“…[3H]-idazoxan binding to 12 sites can be divided into two subtypes, based on their affinity for amiloride (Michel & Insel 1989;Diamant et al, 1992). Rabbit kidney, adipocytes and liver and human placenta all show a high affinity for amiloride whilst human kidney, guinea-pig ileum and rat liver have lower affinities for amiloride (Michel & Insel, 1989;Diamant et al, 1992).…”

“…There is now emerging pharmacological evidence that I2 sites in certain species, in-cluding rabbits, may constitute a heterogeneous population of sites (Michel & Insel, 1989;Wikberg et al, 1991;Diamant et al, 1992;Stewart et al, 1992).…”

Rilmenidine Elevates Cytosolic Free Calcium Concentration in Suspended Cerebral Astrocytes

“…The likelihood of another imidazoline receptor subtype (13) or a rilmenidine-binding site distinct from imidazo!ine and a 2-adrenergic receptors is supported by unidentified nlmenidine binding sites in the cerebral cortex (King et al, 1995). Other tissues expressing mitochondnial imidazo!ine receptors also express imidazoline receptors on the plasma membrane and on another unidentified cellular compartment (LachaudPettiti eta!., 1991; Diamant et a!., 1992).…”