On the functional interaction between nicotinic acetylcholine receptor and Na+,K+-ATPase

Previous studies have shown that nanomolar acetylcholine (ACh) produces a 2 to 4-mV hyperpolarization of skeletal muscle fibers putatively due to Na+,K+-ATPase activation. The present study elucidates the involvement of the nicotinic ACh receptor (nAChR) and of Na+,K+-ATPase isoform(s) in ACh-induced hyperpolarization of rat diaphragm muscle fibers. A variety of ligands of specific binding sites of nAChR and Na+,K+-ATPase were used. Dose–response curves for ouabain, a specific Na+,K+-ATPase inhibitor, were obtained to ascertain which Na+,K+-ATPase isoform(s) is involved. The ACh dose–response relationship for the hyperpolarization was also determined. The functional relationship between these two proteins was also studied in a less complex system, a membrane preparation from Torpedo electric organ. The possibility of a direct ACh effect on Na+,K+-ATPase was studied in purified lamb kidney Na+,K+-ATPase and in rat red blood cells, systems where no nAChR is present. The results indicate that binding of nAChR agonists to their specific sites results in modulation of ouabain-sensitive (most probably α2) isoform of Na+,K+-ATPase, leading to muscle membrane hyperpolarization. In the Torpedo preparation, ouabain modulates dansyl-C6-choline binding to nAChR, and vice versa. These results provide the first evidence of a functional interaction between nAChR and Na+,K+-ATPase. Possible interaction mechanisms are discussed.