Asymmetric distribution of muscarinic acetylcholine receptors in Madin-Darby canine kidney cells.
We have characterized the muscarinic ACh receptors (mAChRs) expressed in Madin- Darby canine kidney (MDCK) strain II epithelial cells. Binding studies with the membrane-impermeable antagonist N-[(3)H]methylscopolamine demonstrated that mAChRs are approximately 2.5 times more abundant on the basolateral than on the apical surface. Apical, but not basolateral, mAChRs inhibited forskolin-stimulated adenylyl cyclase activity in response to the agonist carbachol. Neither apical nor basolateral mAChRs exhibited detectable carbachol-stimulated phospholipase C activity. Carbachol application to the apical or the basolateral membrane resulted in a threefold increase in intracellular Ca(2+) concentration, which was completely inhibited by pertussis toxin on the apical side and partially inhibited on the basolateral side. RT-PCR analysis showed that MDCK cells express the M(4) and M(5) receptor mRNAs. These data suggest that M(4) receptors reside on the apical and basolateral membranes of polarized MDCK strain II cells and that the M(5) receptor may reside in the basolateral membrane of a subset of cells.
Nadler
L S
LS
Department of Pharmacology, University of Washington School of Medicine, Seattle, Washington 98195-7750, USA.
Kumar
G
G
Hinds
T R
TR
Migeon
J C
JC
Nathanson
N M
NM
D000255:Adenosine Triphosphate / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D000918:Antibody Specificity; D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D016764:Cell Polarity / Q000502:physiology*; D018679:Cholinergic Agonists / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D017931:DNA Primers; D004285:Dogs; D004789:Enzyme Activation / Q000187:drug effects; D004847:Epithelial Cells / Q000737:chemistry* / Q000166:cytology; D015870:Gene Expression / Q000502:physiology; D007668:Kidney / Q000166:cytology; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000032:analysis; D043588:Receptor, Muscarinic M4; D043642:Receptor, Muscarinic M5; D011976:Receptors, Muscarinic / Q000032:analysis* / Q000235:genetics* / Q000276:immunology
D018679:Cholinergic Agonists; D017931:DNA Primers; D012333:RNA, Messenger; D043588:Receptor, Muscarinic M4; D043642:Receptor, Muscarinic M5; D011976:Receptors, Muscarinic; D005576:Colforsin; D000255:Adenosine Triphosphate; D002217:Carbachol; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D002118:Calcium