Diminished Functional Activity of Newly Synthesized Muscarinic Acetylcholine Receptors in Stably Transfected Y1 Adrenal Cells

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Abstract
We have examined the functional responsiveness of newly synthesized m2 muscarinic acetylcholine receptors in stably transfected Y1 adrenal cells. After inactivation of preexisting receptors with the covalent alkylating antagonist propylbenzilylcholine mustard, the number of cell surface receptors returned to control values over a 3-h period. After a 3-h recovery, the cells exhibited diminished sensitivity for muscarinic receptor-mediated inhibition of adenylyl cyclase activity, with much higher concentrations of agonist being required to elicit a response. The functional sensitivity returned to control values over a 12-18-h period. The decreased functional activity was not due to a decreased affinity of the newly synthesized receptors for agonist or to a decrease in the levels of inhibitory G proteins in the cells. The results suggest that muscarinic receptors may be synthesized in a form with diminished functional activity. The ability to study the maturation of receptor function in a transfected cell system should allow a combination of biochemical and molecular genetic approaches to analyze the synthesis and functional responsiveness of muscarinic receptors.