Friedman S , Tauber M , Ben-Chaim Y .

	Figure 1. The effect of extracellular Na+ on the potency of the M2R. (a,b) Recordings of ACh-induced GIRK currents in 72 mM Na+ solution (a) and in Na+ free solution (b) from the same oocyte. 3 concentrations of ACh were applied sequentially (1, 2 and 3 stand for 10, 100 and 10,000 nM, respectively). (c) dose response curves assembled from various experiments conducted at 72 mM Na+ solution (black circles) and in Na+ free solution (red circles). Each point represents mean (± SEM) from 7–21 oocytes from 6 batches of oocytes. The solid black and red lines were generated by fitting a 3 parameters equation to the data (see “Methods”). (d) The maximal amplitude of IACh, evoked by 100 µM ACh in 72 mM Na+ solution (black) and in Na+ free solution (red). The two bars are not significantly different (unpaired t test, p = 0.71).
	Figure 2. Extracellular Na+ does not affect the basal GIRK current (IK). (a) Recording of IK from an oocyte that expresses the GIRK channel but not the M2R. Removing Na+ from the extracellular solution did not affect IK. (b) Collected results from 10 oocytes. Each two circles connected with a dashed line represent the amplitude of IK from one oocyte at 72 mM Na+ (black) and at Na+ free solution (red). The order of recordings was selected randomly. Both here and in (c), IK was measured by subtracting the current at 24 mM K+ solution in the presence of 1 mM Ba2+ from the current in its absence. (c) Current–voltage relationship of IK at 72 mM Na+ solution (black circles) and in Na+ free solution (red circles). Results are given as mean (± SEM) from 11 oocytes.
	Figure 3. The effect of extracellular Na+ on the dissociation of ACh from the M2R. (a) The basic experimental protocol. The oocyte was clamped at − 80 mV and IACh was evoked by application of 1 µM ACh in 24 mM K+ solution. Following washout of the ACh with ACh-free solution the current declined and the time constant of the decay was fitted by a standard exponential equation (red dashed line). (b) Representative recordings from one oocyte in 72 mM Na+ solution (black) and in Na+ free solution (red). The currents shown are normalized to enable comparison of the kinetics of the current decline. (c) Collected results from 11 oocytes. Each two circles connected with a dashed line represent the time constant of the decay of IACh following ACh washout from one oocyte at 72 mM Na+ (black) and at Na+ free solution (red). The order of recordings was selected randomly.
	Figure 4. Mutation of Asp69 abolished the Na+ dependence of the M2R. (a) Dose response curves assembled from oocytes expressing the Asp69ASn mutant at 72 mM Na+ solution (black circles) and in Na+ free solution (red circles). The number of repetitions for each ACh concentration varied between 7 and 21 from 4 batches of oocytes. The solid black and red lines were generated by fitting a 3 parameters equation to the data (see “Methods”). (b) Representative recordings from one oocyte in 72 mM Na+ solution (black) and in Na+ free solution (red). The currents shown are normalized to enable comparison of the kinetics of the current decline. (c) Collected results from 10 oocytes. Each two circles connected with a dashed line represent the time constant of the decay of IACh following ACh washout from one oocyte at 72 mM Na+ (black) and at Na+ free solution (red). The order of recordings was selected randomly. The two conditions are not significantly different (p = 0.45).
	Figure 5. The effect of extracellular Na+ on the voltage-dependence of the potency of ACh toward the M2R. (a) Representative recordings from one oocyte at − 80 mV (left) and at + 40 mV (right) at Na+ free solution. (b,c) Dose response curves assembled from 5 experiments at 72 mM Na+ solution (b) and in Na+ free solution (c). For each oocyte, IACh was measured at − 80 mV (black circles) and at + 40 mV (red circles) at either Na+ concentration, and the response of each concentration was normalized to the response evoked by 10 µM ACh at the same holding potential. Each point represents mean (± SEM) from 8–30 oocytes. The solid black and red lines were generated by fitting a 3 parameters equation to the data (see “Methods”).

Ågren, Point mutation of a conserved aspartate, D69, in the muscarinic M2 receptor does not modify voltage-sensitive agonist potency. 2018, Pubmed, Xenbase

Ågren, Point mutation of a conserved aspartate, D69, in the muscarinic M2 receptor does not modify voltage-sensitive agonist potency. 2018, Pubmed , Xenbase
Barbhaiya, Site-directed mutagenesis of the human A1 adenosine receptor: influences of acidic and hydroxy residues in the first four transmembrane domains on ligand binding. 1996, Pubmed
Barchad-Avitzur, A Novel Voltage Sensor in the Orthosteric Binding Site of the M2 Muscarinic Receptor. 2016, Pubmed , Xenbase
Ben Chaim, Voltage affects the dissociation rate constant of the m2 muscarinic receptor. 2013, Pubmed , Xenbase
Ben-Chaim, The M2 muscarinic G-protein-coupled receptor is voltage-sensitive. 2003, Pubmed , Xenbase
Ben-Chaim, Movement of 'gating charge' is coupled to ligand binding in a G-protein-coupled receptor. 2006, Pubmed , Xenbase
Birk, Membrane Potential Controls the Efficacy of Catecholamine-induced β1-Adrenoceptor Activity. 2015, Pubmed
Dascal, Signalling via the G protein-activated K+ channels. 1997, Pubmed
Dascal, Expression of an atrial G-protein-activated potassium channel in Xenopus oocytes. 1993, Pubmed , Xenbase
Dekel, Depolarization induces a conformational change in the binding site region of the M2 muscarinic receptor. 2012, Pubmed , Xenbase
Evans, Differential ontogeny of putative M1 and M2 muscarinic receptor binding sites in the murine cerebral cortex and heart. 1985, Pubmed
Fenalti, Molecular control of δ-opioid receptor signalling. 2014, Pubmed
Gao, Allosteric modulation of A(2A) adenosine receptors by amiloride analogues and sodium ions. 2000, Pubmed
Haga, Structure of the human M2 muscarinic acetylcholine receptor bound to an antagonist. 2012, Pubmed
Ho, Molecular determinants for sodium-dependent activation of G protein-gated K+ channels. 1999, Pubmed , Xenbase
Horstman, An aspartate conserved among G-protein receptors confers allosteric regulation of alpha 2-adrenergic receptors by sodium. 1990, Pubmed
Katritch, Allosteric sodium in class A GPCR signaling. 2014, Pubmed
Kruse, Muscarinic acetylcholine receptors: novel opportunities for drug development. 2014, Pubmed
Kruse, Activation and allosteric modulation of a muscarinic acetylcholine receptor. 2013, Pubmed
Liu, Structural basis for allosteric regulation of GPCRs by sodium ions. 2012, Pubmed
Mahaut-Smith, A role for membrane potential in regulating GPCRs? 2008, Pubmed
Martinez-Pinna, Direct voltage control of signaling via P2Y1 and other Galphaq-coupled receptors. 2005, Pubmed
Miao, Allosteric effects of sodium ion binding on activation of the m3 muscarinic g-protein-coupled receptor. 2015, Pubmed
Miller-Gallacher, The 2.1 Å resolution structure of cyanopindolol-bound β1-adrenoceptor identifies an intramembrane Na+ ion that stabilises the ligand-free receptor. 2014, Pubmed
Mosser, Kinetic analysis of M2 muscarinic receptor activation of Gi in Sf9 insect cell membranes. 2002, Pubmed
Motulsky, Influence of sodium on the alpha 2-adrenergic receptor system of human platelets. Role for intraplatelet sodium in receptor binding. 1983, Pubmed
Navarro-Polanco, Conformational changes in the M2 muscarinic receptor induced by membrane voltage and agonist binding. 2011, Pubmed
Neve, Regulation of dopamine D2 receptors by sodium and pH. 1991, Pubmed
Neve, Pivotal role for aspartate-80 in the regulation of dopamine D2 receptor affinity for drugs and inhibition of adenylyl cyclase. 1991, Pubmed
Ohana, The metabotropic glutamate G-protein-coupled receptors mGluR3 and mGluR1a are voltage-sensitive. 2006, Pubmed , Xenbase
Peleg, G(alpha)(i) controls the gating of the G protein-activated K(+) channel, GIRK. 2002, Pubmed , Xenbase
Rinne, Voltage regulates adrenergic receptor function. 2013, Pubmed
Rosenberger, Cardiac muscarinic cholinergic receptor binding is regulated by Na+ and guanyl nucleotides. 1980, Pubmed
Sahlholm, Voltage-dependence of the human dopamine D2 receptor. 2008, Pubmed , Xenbase
Sharma, Molecular and functional identification of m1 muscarinic acetylcholine receptors in rat ventricular myocytes. 1996, Pubmed
Sui, Activation of the atrial KACh channel by the betagamma subunits of G proteins or intracellular Na+ ions depends on the presence of phosphatidylinositol phosphates. 1998, Pubmed , Xenbase
Sui, Na+ activation of the muscarinic K+ channel by a G-protein-independent mechanism. 1996, Pubmed , Xenbase
Suno, Structural insights into the subtype-selective antagonist binding to the M2 muscarinic receptor. 2018, Pubmed
Tsai, Agonist-specific effects of monovalent and divalent cations on adenylate cyclase-coupled alpha adrenergic receptors in rabbit platelets. 1978, Pubmed
Vickery, Membrane potentials regulating GPCRs: insights from experiments and molecular dynamics simulations. 2016, Pubmed
Vickery, Structural Mechanisms of Voltage Sensing in G Protein-Coupled Receptors. 2016, Pubmed
Vogel, Double mutant cycle analysis of aspartate 69, 97, and 103 to asparagine mutants in the m2 muscarinic acetylcholine receptor. 1999, Pubmed
Vorobiov, Coupling of the muscarinic m2 receptor to G protein-activated K(+) channels via Galpha(z) and a receptor-Galpha(z) fusion protein. Fusion between the receptor and Galpha(z) eliminates catalytic (collision) coupling. 2000, Pubmed , Xenbase
Wang, Functional M3 muscarinic acetylcholine receptors in mammalian hearts. 2004, Pubmed
Wilson, The role of a conserved inter-transmembrane domain interface in regulating alpha(2a)-adrenergic receptor conformational stability and cell-surface turnover. 2001, Pubmed
Zarzycka, Harnessing Ion-Binding Sites for GPCR Pharmacology. 2019, Pubmed