Expression of subunit-omitted mouse nicotinic acetylcholine receptors in Xenopus laevis oocytes

Abstract

1. Nicotinic acetylcholine (ACh) receptors in developing vertebrate skeletal muscle exhibit functional heterogeneity in both conductance and kinetics. To assess the contributions of receptors differing in subunit composition to the heterogeneity, various combinations of mouse alpha beta delta gamma epsilon subunit RNAs were tested for the ability to express functional receptors in Xenopus oocytes. 2. Two combinations of dual-subunit RNAs (alpha delta and alpha gamma) resulted in detectable ACh-activated currents and six different combinations of three or more subunit RNAs produced significant numbers of functional channels. The order of combinations yielding the greatest amount of current was alpha beta gamma > alpha beta delta = alpha delta epsilon > alpha delta gamma > alpha delta > alpha gamma. 3. The extent to which a channel type with three different subunits was expressed was highly dependent upon the ratios of RNAs coding for the different subunits. For alpha beta delta receptors the efficiency of expression was alpha: beta: delta (1/5:1/5:1) >> (1:1:1) >> (1/5:1:1/5) > (1:1/5:1/5). 4. The level of expression of three-subunit combinations was also critically dependent upon the order of RNAs injected. When alpha delta or alpha gamma RNA combinations were co-injected 2 days prior to the injection of beta RNA, the expression was 2-5 times greater than when alpha beta injection was followed by injection of delta or gamma RNA. 5. Single-channel measurements revealed that alpha beta delta channels were not expressed in the presence of alpha beta delta epsilon RNAs, even under conditions when the amount of delta RNA injected was 5-fold higher than the amount of epsilon RNA. 6. These data indicate that the functional expression of subunit-omitted receptors depends critically upon the relative amounts of the five different subunit RNAs. Receptors composed of three different subunits express in the presence of the subunit RNAs characteristic of embryonic muscle (alpha beta delta gamma), but are not observed with the combination of RNAs characteristic of adult muscle (alpha beta delta epsilon).