Pharmacological characterization and distribution of muscarinic receptors in human placental syncytiotrophoblast brush-border and basal plasma membranes

Based on the existence of choline acetyltransferase and acetylcholine in human placenta, we have investigated the presence of muscarinic acetylcholine receptors in brush-border and basal plasma membranes from human term placenta. Radioligand binding assay, using ³H]N-methyl-scopolamine as tracer, showed the existence of acetylcholine muscarinic receptors in brush-border (K_d 0.28±0.04 nM; B_max 9.4±1.6 fmol/mg protein) and basal plasma membranes (K_d 0.24±0.05 nM; B_max 34.3±6.3 fmol/mg protein). In order to perform a pharmacological characterization of these receptors, competition binding experiments were carried out using the muscarinic receptor antagonists pirenzepine, (11(2-diethyl-amino)methyl)-1-piperidinylacetyl-5-11-dihydro-6H-pyrido(14)benzodiazepine (AF-DX 116), himbacine, 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP), dicyclomine and hexahydro-sila-difenidol (HHSD). The results obtained showed that the muscarinic receptors in brush-border and basal plasma membranes belong to different subtypes. In brush-border membranes, the receptor found match in terms of affinity for the antagonists with the muscarinic M₁ receptor subtype (K_i pirenzepine, 13.6±8.2 nM; K_i AF-DX 116, 1680±271 nM; K_i himbacine, 212±6.5 nM; K_i 4-DAMP, 1.5±0.4 nM; K_i dicyclomine, 5.1±0.8 nM; K_i HHSD, 34.3±7.3 nM), whereas the receptor in basal plasma membrane seems to be of the muscarinic M₂ receptor subtype (K_i pirenzepine, 202±48 nM; K_i AF-DX 116, 124±60 nM; K_i himbacine, 20.6±4.8 nM; K_i 4-DAMP, 4.5±1.2 nM; K_i dicyclomine, 54.6±22 nM; K_i HHSD, 89.2±15.8 nM). The results obtained show the existence of muscarinic acetylcholine receptors in brush-border and basal plasma membranes from human term placenta with a different distribution pattern in terms of number of receptors and distribution of different subtypes. The functional significance of these findings is as yet unknown, but these receptors probably mediate different functions as they belong to different subtypes and are coupled to different second messengers.