The Effects of isoflurane on acetylcholine receptor channels: 3. Effects of conservative polar-to-nonpolar mutations within the channel pore.
We performed macroscopic and single-channel current measurements on wild-type (WT) and two mutant muscle-type nicotinic acetylcholine (ACh) receptor channels transiently expressed in HEK-293 cells. The mutants contained polar-to-nonpolar substitutions at the 10' (alpha(2)S10'A beta T10'A gamma delta) and 6' positions (alpha(2)S6'A beta gamma delta S6'A) in the M2 pore region of the channel. We studied the behavior of these channels in the absence and presence of the volatile general anesthetic isoflurane. Both mutations changed the gating behavior of the channel. A comparison of the alpha(2)S10'A beta T10'A gamma delta mutant to WT receptors revealed faster desensitization kinetics, increased sensitivity to ACh, a higher efficacy for activation by the partial nicotinic agonist decamethonium, and a greater number of openings per burst. A comparison of the alpha(2)S6'A beta gamma delta S6'A mutant to WT receptors also revealed increased sensitivity to ACh and an increased burst duration at the single-channel level with ACh as agonist. The alpha(2)S10'A beta T10'A gamma delta mutation increased the sensitivity of the ACh receptor to isoflurane, whereas the alpha(2)S6'A beta gamma delta S6'A mutation did not. These changes were probably not caused by the differential effects of the mutation on channel gating and desensitization. The increased sensitivity of the alpha(2)S10'A beta T10'A gamma delta receptor to isoflurane is state-dependent; the mutation changes the affinity of the closed state but not that of the open state of the channel.
Wenningmann
I
I
Klinik für Anästhesiologie, Universität Bonn, Bonn, Germany.
Barann
M
M
Vidal
A M
AM
Dilger
J P
JP
D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D000818:Animals; D002478:Cells, Cultured; D003650:Decamethonium Compounds / Q000494:pharmacology; D004594:Electrophysiology; D006801:Humans; D007530:Isoflurane / Q000494:pharmacology*; D007700:Kinetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009467:Neuromuscular Depolarizing Agents; D010477:Perfusion; D011950:Receptors, Cholinergic / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D014162:Transfection
D018685:Anesthetics, Inhalation; D003650:Decamethonium Compounds; D009467:Neuromuscular Depolarizing Agents; D011950:Receptors, Cholinergic; C033019:decamethonium; D007530:Isoflurane