Long-term changes in the muscarinic M1 receptor induced by instantaneous formation of wash-resistant xanomeline-receptor complex.
Unlike other M1 muscarinic acetylcholine receptor agonists, xanomeline demonstrates a unique mode of binding to the receptor. It not only binds reversibly to the receptor's conventional orthosteric site but also binds persistently at a secondary binding domain(s) on the M1 receptor. This results in persistent activation of the receptor even after extensive washout, and allosteric modulation of the orthosteric site. In the current study, we investigated how the effects of very brief exposure (1 min) of intact Chinese hamster ovary cells expressing M1 receptors to xanomeline followed by washout change with time. Pretreatment with xanomeline for 1 min resulted in a concentration-dependent wash-resistant inhibition of [3H]N-methylscopolamine (NMS) binding, with a lower potency than that observed in the continuous presence of xanomeline in the binding assay medium. This effect was associated with wash-resistant receptor activation. Incubation of pretreated and washed cells in control medium for 24 h transformed the monophasic xanomeline wash-resistant binding curve to one that exhibits two distinct potencies. This was the result of the appearance of a new very high-potency binding component without a change in the low-potency state. The delayed effects of persistently bound xanomeline are mainly due to reduction of the maximal binding of [3H]NMS without a change in its affinity. These treatment conditions also reversed persistent receptor activation by xanomeline. Our results imply that brief exposure to xanomeline followed by washing and prolonged waiting may result in delayed receptor desensitization accompanied by internalization or down-regulation.
De Lorme
Kayla C
KC
Division of Neuroscience Research in Psychiatry, University of Minnesota Medical School, Minneapolis, MN 55455, USA.
Grant
Marianne K O
MK
Noetzel
Meredith J
MJ
Polson
Sonya B
SB
El-Fakahany
Esam E
EE
D000495:Allosteric Site; D000818:Animals; D001667:Binding, Competitive; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D008024:Ligands; D018721:Muscarinic Agonists / Q000494:pharmacology*; D019832:N-Methylscopolamine / Q000494:pharmacology; D010053:Ovary / Q000166:cytology; D011485:Protein Binding; D011725:Pyridines / Q000494:pharmacology*; D011869:Radioligand Assay; D043643:Receptor, Muscarinic M1 / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D013830:Thiadiazoles / Q000494:pharmacology*; D013997:Time Factors; D014162:Transfection
D007295:Inositol Phosphates; D008024:Ligands; D018721:Muscarinic Agonists; D011725:Pyridines; D043643:Receptor, Muscarinic M1; D013830:Thiadiazoles; D002217:Carbachol; C075257:xanomeline; D019832:N-Methylscopolamine