Inhibition of the muscarinic receptor-activated K+ current by N-ethylmaleimide in rabbit heart.
The effects of N-ethylmaleimide (NEM), a sulfhydryl alkylating agent, on the ACh-activated K+ current were examined in single cells from rabbit hearts using whole-cell and single channel patch clamp techniques. Bath application of NEM (50 microM) or the muscarinic antagonist, atropine (1 microM) completely inhibited the ACh-activated K+ current in whole-cell recordings. In cell-attached patch conditions, the inhibitory effect of NEM was still observed; in contrast, atropine was ineffective when the agents were bath applied, indicating that the site of action of NEM is different from that of atropine. Inside-out patch recordings confirmed that GTP was required on the internal side of the membrane for activation of this K+ channel when ACh was present in the pipette. NEM abolished this GTP-activated K+ channel activity. GTP gamma S, a non-hydrolysable GTP analogue, was able to activate this K+ channel in the absence of a muscarinic agonist, an effect thought to be due to the direct activation of GTP-binding proteins. Pretreatment with NEM almost completely prevented this effect of GTP gamma S. In contrast, after the activation of the K+ channel by GTP gamma S had reached a steady-state, NEM failed to show a significant inhibitory effect. These results demonstrate that NEM prevents the activation of muscarinic receptor-regulated K+ channel and suggest an involvement of alkylation of the GTP-binding proteins which are coupled to this type of K+ channel.
Nakajima
T
T
Department of Medical Physiology, University of Calgary School of Medicine, Alberta, Canada.
Kaibara
M
M
Irisawa
H
H
Giles
W
W
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D001285:Atropine / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism / Q000502:physiology; D006321:Heart / Q000187:drug effects* / Q000502:physiology; D018727:Muscarinic Antagonists; D009206:Myocardium / Q000166:cytology / Q000648:ultrastructure*; D011188:Potassium / Q000502:physiology*; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology; D011817:Rabbits
D018727:Muscarinic Antagonists; D015221:Potassium Channels; D001285:Atropine; D019204:GTP-Binding Proteins; D000109:Acetylcholine; D005033:Ethylmaleimide; D011188:Potassium