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. 1995 Sep;116(2):1723–1728. doi: 10.1111/j.1476-5381.1995.tb16654.x

Relationship between agonist binding, phosphorylation and immunoprecipitation of the m3-muscarinic receptor, and second messenger responses.

A B Tobin 1, G B Willars 1, N T Burford 1, S R Nahorski 1
PMCID: PMC1909083  PMID: 8528551

Abstract

1. Phosphoinositidase C-linked m3-muscarinic receptors expressed in Chinese hamster ovary cells (CHO-m3 cells) are phosphorylated on serine following agonist stimulation. 2. m3-Muscarinic receptor phosphorylation is concentration-dependent requiring a carbachol concentration of 13.2 microM for half maximal stimulation. 3. The phosphorylation concentration-response curve lies to the left of the curve for carbachol binding to muscarinic receptors (KD = 100 microM) in membranes from CHO-m3 cells. In contrast, receptor phosphorylation closely correlates with receptor-mediated phosphoinositidase C activation (EC50 for inositol 1,4,5 trisphosphate accumulation during the peak and plateau phases were 7.14 microM and 5.92 microM respectively) but not with rapid agonist-mediated calcium elevation (EC50 = 0.32 microM) measured in fura-2-AM loaded cells. 4. These data suggest a dissociation of receptor phosphorylation from agonist occupation. Such an apparent 'receptor reserve' for m3-muscarinic receptor phosphorylation may be indicative of a mechanism that is dependent on a small amplification of the receptor signal, though probably dissociated from the calcium signal.

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