Effect of the lipid environment on the differential affinity of purified cerebral and atrial muscarinic acetylcholine receptors for pirenzepine.

Muscarinic acetylcholine receptors (mAChRs) of porcine cerebral membrane (predominantly M1 subtype) and porcine atrial membrane (M2 subtype) showed the same affinity for the muscarinic antagonist [3H]quinuclidinylbenzylate [( 3H]QNB). In contrast, the affinity for pirenzepine (another muscarinic antagonist) of 86% of binding sites in the cerebral membrane (H sites) was 34-fold higher than that in the atrial membrane. After purification of mAChRs by affinity chromatography, this difference was less than 3-fold. This phenomenon was fully reversed by insertion of purified mAChRs into either cerebral or atrial membranes whose native muscarinic binding sites had been alkylated with propylbenzilycholine mustard, indicating that the purified receptors recovered their original affinities for pirenzepine upon interaction with membrane components. To examine the effect of the interaction between receptors and lipid components on the affinities for [3H]QNB and pirenzepine, binding experiments were carried out with mAChRs inserted into various lipid preparations. When purified cerebral and atrial mAChRs were inserted into cholesteryl hemisuccinate, their affinities for [3H]QNB and pirenzepine became close to the membrane values and were 7- and 50- to 60-fold higher than those of receptors inserted into phosphatidylcholine, respectively. When insertion was carried out into either cholesteryl hemisuccinate, phosphatidylcholine, or cholesteryl hemisuccinate/phosphatidylcholine mixtures, (80:20 and 50:50, w/w), the affinity of cerebral H sites for pirenzepine was only 3- to 5-fold higher than that of atrial receptors, but it became 20- and 60-fold higher when the receptors were inserted in a cholesteryl hemisuccinate/phosphatidylcholine mixture (20:80, w/w) and in a cholesteryl hemisuccinate/phosphatidylcholine/phosphatidylinositol mixture (4:48:48, w/w), respectively. These results suggest that the affinities of mAChRs for antagonists, in particular the differential affinities of cerebral and atrial mAChRs for pirenzepine, are modulated by the lipid environment.