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Am J Physiol Cell Physiol 264: C1550-C1560, 1993;
0363-6143/93 $5.00
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AJP - Cell Physiology, Vol 264, Issue 6 C1550-C1560, Copyright © 1993 by American Physiological Society

ARTICLES

M3 muscarinic acetylcholine receptor coupling to PLC in rat exorbital lacrimal acinar cells

P. Mauduit, H. Jammes and B. Rossignol
Laboratoire de Biochimie des Transports Cellulaires, Centre National de la Recherche Scientifique, Unite de Recherche Associee 1116, Universite Paris-Sud, Orsay, France.

This study was designed to characterize the muscarinic acetylcholine receptor (mAChR) subtype present in rat exorbital lacrimal gland as well as its biochemical coupling. The nonselective muscarinic antagonist [N-methyl-3H]scopolamine ([3H]NMS) binds with high affinity to a homogeneous population of binding sites in both membranes [dissociation constant (Kd) = 82.3 +/- 3.2 pM] and acinar cell (Kd = 170.3 +/- 20 pM) preparations. Muscarinic antagonist inhibition of [3H]NMS binding is homogeneous with the following order of potency: atropine > or = 4-diphenylacetoxy-N-methylpiperidine (4-DAMP) > pirenzepine > 11-([2-(diethylamino)-ethyl]-1-piperidinyl)-acetyl- 5,11-dihydro-6H-pirido[2,3-b]1,4,benzo diazepine-6-one (AFDX 116). Both the affinity of the selective antagonists 4-DAMP, pirenzepine, and AFDX 116 and Northern blot analysis of lacrimal gland mRNAs show a single mAChR population of the M3 subtype. Muscarinic agonist inhibition of [3H]NMS binding displays both high (approximately 20%)- and low-affinity sites (approximately 80%). Both the receptor occupancy and the stimulation by agonists or the inhibition by antagonists of the accumulation of [3H]inositol phosphate were examined under identical conditions with respect to tissue preparations (acinar cells) and buffer (Krebs-Ringer). Results demonstrate 1) the efficient coupling of the M3 mAChR subtype with the phosphatidylinositol (4,5))bisphosphate-specific phospholipase C activity and 2) that the efficacy of a muscarinic agonist is dependent on its structure. Lastly, comparison of the agonists affinity and potency to trigger the [3H]inositol phosphate accumulation suggests that the occupation of the high-affinity agonist binding state of the M3 mAChR was involved in the cellular response.


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