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

Abstract

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 alpha2) 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.

The Nicotinic Acetylcholine Receptor and the Na,K-ATPase α2 Isoform Interact to Regulate Membrane Electrogenesis in Skeletal Muscle

“…However, addition of 100 nM nicotine (closed circles) hyperpolarizes the membrane by Ϫ4.5 mV (p Ͻ 0.01). The magnitude and time course of this effect are similar to that consistently seen in control, Na ϩ -containing solutions (1). This result suggests that Na ϩ entry through the nAChR is not required for its functional interaction with the Na,K-ATPase.…”

“…(20)) to the nAChR was monitored by changes in fluorescence intensity at 557 nm. DC6C was excited using energy transfer from a tryptophan near the ACh binding sites (21,22), as described previously (1). The assays were conducted in Torpedo physiological saline buffer, HTPS (250 mM NaCl, 5 mM KCl, 3 mM CaCl 2 , 2 mM MgCl 2 , and 20 mM Hepes, pH 7.0).…”

“…We previously identified a regulatory mechanism, termed acetylcholine (ACh)-induced hyperpolarization, whereby the nAChR and the Na,K-ATPase functionally interact to modulate the membrane potential of rat skeletal muscle (1)(2)(3)(4). In this interaction, the binding of nanomolar concentrations of ACh to the nAChR stimulates electrogenic transport by the Na,K-ATPase ␣2 isozyme, causing a membrane hyperpolarization of about Ϫ4 mV.…”

Na+,K+-ATPase Functionally Interacts with the Plasma Membrane Na+,Ca2+ Exchanger to Prevent Ca2+ Overload and Neuronal Apoptosis in Excitotoxic Stress

“…The extracellular loops of the catalytic ␣-subunit of Na ϩ ,K ϩ -ATPase form a binding site that is the only known, highly specific receptor for ouabain and other cardiotonic steroids (Ogawa et al, 2009;Lingrel, 2010) and their circulating endogenous analogs (Blaustein, 1993;Schoner and Scheiner-Bobis, 2007;Bagrov and Shapiro, 2008). Upon binding of ouabain to this receptor, Na ϩ ,K ϩ -ATPase interacts with neighboring membrane proteins to affect diverse cell functions such as protein synthesis, proliferation, cell differentiation, gene expression, regulation of intracellular Ca 2ϩ , contractile properties, synaptic efficacy, and neural differentiation (Xie and Askari, 2002;Krivoi et al, 2006;Aperia, 2007;Hazelwood et al, 2008;Desfrere et al, 2009;Li and Xie, 2009;Radzyukevich et al, 2009;Rose et al, 2009;Heiny et al, 2010). In addition, ouabain in nanomolar concentrations stimulates the reversed mode of plasma membrane Na ϩ ,Ca 2ϩ exchange in snail neurons (Saghian et al, 1996).…”

Glial Nax channels control lactate signaling to neurons for brain [Na+] sensing

“…However, monensin treatments alone, which induces extensive Na + influx (Fig. III.6A), had little effect on the Recently, several molecules, including nicotinic ACh receptor, FXYD1, FXYD 2, Src tyrosine kinases, and cofilin, were found to interact with Na + /K + -ATPase (Krivoi, et al, 2006;Feschenko et al, 2003;Zouzoulas and Blostein et al, 2006;Wang and Yu et al, 2005;Jung J, et al, 2002): Some reportedly activate or modify the properties of In Na x -negative C6M16 cells, no such enhancement by the C-terminal of Na x was observed. Fluorescent intensity of Con of the Mock condition was set at 100%.…”