title: Non-neuronal cell-conditioned medium regulates muscarinic receptor expression in cultured sympathetic neurons.
abstract: Regulation of muscarinic receptor expression was examined in cultured sympathetic neurons of the neonatal rat superior cervical ganglion. Receptor concentration was determined by measuring binding of the muscarinic antagonist 1-quinuclidinyl[phenyl-4-3H] benzilate (3H-QNB). 3H-QNB bound to one apparent class of noninteracting sites on sympathetic neuron membranes with a Kd of 28.9 pM and a Bmax of 2.91 pmol/mg protein. Binding increased as a linear function of tissue protein and was saturable. The number of receptors per milligram protein increased approximately 6-fold during 16 d of culture, and receptor numbers were down-regulated by treatment with the agonist carbachol. These observations suggested that measurement of 3H-QNB binding would provide a reliable estimate of muscarinic receptor number on cultured sympathetic neurons. To determine whether nonneuronal cells produce soluble factors that influence muscarinic receptor expression, the effects of treatment with rat fibroblast-conditioned medium (RFCM) were examined. Exposure of sympathetic neurons to 50% RFCM resulted in a 57% decrease in muscarinic receptor numbers without a change in the apparent Kd. The decrease in 3H-QNB binding in response to RFCM was dose-related, with a minimum dose of 15% RFCM required to observe a significant effect. In contrast to the carbachol-induced down-regulation, the reduction in binding after RFCM treatment was not prevented by atropine, indicating that the effect was not mediated by acetylcholine stimulation of muscarinic receptors. Binding of 125I-alpha-bungarotoxin, which labels a nonfunctional membrane site distinct from nicotinic receptors, was not altered by treatment with RFCM, indicating the selectivity of the change in membrane muscarinic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Albert Einstein College of Medicine, Department of Neurology, Bronx, New York 10461.
authors.forename: K E
authors.initials: KE
authors.lastname: Smith
authors.forename: J A
authors.initials: JA
authors.lastname: Kessler
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D003470:Culture Media; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D009116:Muscarine / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D011813:Quinuclidinyl Benzilate / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D011976:Receptors, Muscarinic / Q000378:metabolism*; D013564:Sympathetic Nervous System / Q000166:cytology*
chemical_list: D003470:Culture Media; D011976:Receptors, Muscarinic; D011813:Quinuclidinyl Benzilate; D009116:Muscarine