M(2) and M(3) muscarinic receptor-mediated contractions in longitudinal smooth muscle of the ileum studied with receptor knockout mice.
Isometric contractile responses to carbachol were studied in ileal longitudinal smooth muscle strips from wild-type mice and mice genetically lacking M(2) or M(3) muscarinic receptors, in order to characterize the mechanisms involved in M(2) and M(3) receptor-mediated contractile responses. Single applications of carbachol (0.1-100 microM) produced concentration-dependent contractions in preparations from M(2)-knockout (KO) and M(3)-KO mice, mediated via M(3) and M(2) receptors, respectively, as judged by the sensitivity of contractile responses to blockade by the M(2)-preferring antagonist methoctramine (300 nM) or the M(3)-preferring antagonist 4-DAMP (30 nM). The M(2)-mediated contractions were mimicked in shape by submaximal stimulation with high K(+) concentrations (up to 35 mM), almost abolished by voltage-dependent Ca(2+) channel (VDCC) antagonists or depolarization with 140 mM K(+) medium, and greatly reduced by pertussis toxin (PTX) treatment. The M(3)-mediated contractions were only partially inhibited by VDCC antagonists or 140 mM K(+)-depolarization medium, and remained unaffected by PTX treatment. The contractions observed during high K(+) depolarization consisted of different components, either sensitive or insensitive to extracellular Ca(2+). The carbachol contractions observed with wild-type preparations consisted of PTX-sensitive and -insensitive components. The PTX-sensitive component was functionally significant only at low carbachol concentrations. The results suggest that the M(2) receptor, through PTX-sensitive mechanisms, induces ileal contractions that depend on voltage-dependent Ca(2+) entry, especially associated with action potential discharge, and that the M(3) receptor, through PTX-insensitive mechanisms, induces contractions that depend on voltage-dependent and -independent Ca(2+) entry and intracellular Ca(2+) release. In intact tissues coexpressing M(2) and M(3) receptors, M(2) receptor activity appears functionally relevant only when fractional receptor occupation is relatively small.
Unno
Toshihiro
T
Laboratory of Pharmacology, Department of Veterinary Medicine, Faculty of Applied Biological Science, Gifu University, Japan.
Matsuyama
Hayato
H
Sakamoto
Takashi
T
Uchiyama
Mai
M
Izumi
Yusuke
Y
Okamoto
Hiroyuki
H
Yamada
Masahisa
M
Wess
Jürgen
J
Komori
Seiichi
S
D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology*; D018679:Cholinergic Agonists / Q000494:pharmacology*; D003959:Diamines / Q000494:pharmacology; D005260:Female; D007082:Ileum / Q000187:drug effects / Q000502:physiology; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D018727:Muscarinic Antagonists / Q000494:pharmacology; D009119:Muscle Contraction / Q000187:drug effects*; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology; D009529:Nicardipine / Q000494:pharmacology; D037342:Pertussis Toxin / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D043585:Receptor, Muscarinic M2 / Q000819:agonists* / Q000172:deficiency / Q000502:physiology; D043587:Receptor, Muscarinic M3 / Q000819:agonists* / Q000172:deficiency / Q000502:physiology
D002121:Calcium Channel Blockers; D018679:Cholinergic Agonists; D003959:Diamines; D018727:Muscarinic Antagonists; D010880:Piperidines; D043585:Receptor, Muscarinic M2; D043587:Receptor, Muscarinic M3; C042375:4-diphenylacetoxy-1,1-dimethylpiperidinium; D002217:Carbachol; D009529:Nicardipine; D037342:Pertussis Toxin; C054938:methoctramine