Short-term agonist-induced loss of cell surface muscarinic receptors and desensitization of receptor-mediated cyclic GMP (cGMP) formation and phosphoinositide hydrolysis were examined in mouse neuroblastoma cells (clone N1E-115) in suspension. This treatment resulted in a time-dependent reduction of approximately 40% of the specific binding of the hydrophilic antagonist [3H]N-methyl-scopolamine ([3H]NMS) with a T 1 2 of down-regulation of 4.83 min. Scatchard analysis revealed that brief exposure to the agonist resulted in a significant reduction in the Bmax with no change in the Kd. Agonist-induced cGMP formation decreased in a similar time-dependent manner with an average T 1 2 of 4.79 min. However, desensitization of muscarinic receptor-stimulated accumulation of inositol phosphates demonstrated a much slower time-course and was accompanied by a reduction in the maximal response with no change in the EC50. In addition, there was rapid partial recovery of cell surface receptors and desensitized cGMP response, with no apparent resensitization of phosphoinositide hydrolysis. Thus, there was a differential rate of short-term desensitization and resensitization of these two muscarinic receptor-mediated responses. Moreover, desensitization of cGMP formation, but not phosphoinositide hydrolysis, closely paralleled loss of cell surface muscarinic receptors.
Bibliographical noteFunding Information:
* Supported in part by Grants NS-24158, NS-25743 and AG-07118 from the National Institutes of Health and by Contracts DAAG-29-85-K-0123 and DAAL03-88-K-0078 from the U.S. Army Research Office. t Recipient of an Emerson Fellowship from the University of Maryland during the course of this work. Present address: Neuroscience Laboratory, University of Michigan, Ann Arbor, MI. $ Recipient of a Research Career Development Award from the National Institutes of Health (AG-~344).