INFLUENCE OF N‐ETHYLMALEIMIDE ON CHOLINOCEPTORS AND RESPONSES IN LONGITUDINAL MUSCLES FROM GUINEA‐PIG ILEUM

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Abstract
The binding of carbamylcholine to membranes prepared from the longitudinal muscle of guinea‐pig ileum was determined from its inhibition of the binding of [3H]‐3‐quinuclidinyl benzilate. Carbamylcholine binding was resolved into high and low affinity components with apparent dissociation constants of 0.11 ± 0.02 and 11 ± 1 μm 42% of the receptors displayed high affinity carbamylcholine binding. Alkylation of longitudinal muscle membranes with N‐ethylmaleimide increased muscarinic receptor affinity for carbamylcholine in a manner consistent with a conversion of low affinity to high affinity receptors. After exposure the muscle membrane fragments to 1 mm N‐ethylmaleimide for 20 min at 35°C, carbamylcholine binding was resolved into two components with apparent dissociation constants of 0.11 ± 0.01 and 9 ± 2 μm, with 74% of the receptors displaying the higher affinity. Exposure of longitudinal membranes mounted in an organ chamber to 1 mm N‐ethylmaleimide for 30 s depressed isometric contractions in response to acetylcholine by 80%, while contractions induced by K+ and Ba2+ were reduced by less than 20% and 10%, respectively. Acetylcholine dose‐response curves were shifted to the right while Ba2+ curves were unaffected. It is suggested that N‐ethylmaleimide has a selective effect on muscarinic responses in the longitudinal muscle by disrupting processes occurring after receptor occupancy but before the induction of phospholipid turnover or calcium influx in the postsynaptic membrane.