Stimulation of the Na+/K+ pump by external [K+] is regulated by voltage-dependent gating.

Vasilets, Larisa A.; Omay, Halit; Ohta, Toshiko; Noguchi, Shunsuke; Kawamura, Masaru; Schwarz, Wolfgang

doi:10.1016/s0021-9258(18)55292-3

Cited by 57 publications

(12 citation statements)

References 15 publications

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“…In recent years, various protein preparations and experimental techniques have been used to analyze the electrogenicity of the Na,K-ATPase Gadsby, 1986, 1989;Lauger and Apell, 1988;Borlinghaus et al, 1987;Rakowski et al, 1991;Sturmer et al, 1991;Vasilets et al, 1991;Sturmer and Apell, 1992;Gadsby et al, 1993;Sagar and Rakowski, 1994;Hilgemann, 1994;Heyse et al, 1994 :rbCP j rf FIGURE 9 Kinetic reaction cycles of the Na+ transport in the Na,K-ATPase. Cycle A represents the cycle proposed by Heyse et al (1994) in the absence of K+ ions.…”

Section: Discussionmentioning

confidence: 99%

“…The pump cycle of the Na,K-ATPase can be described by the generally accepted Post-Albers cycle (Fig. 1), in which four partial reactions are electrogenic: the cytoplasmic Na+ binding, E1 + 3 Nacyt -> Na3El, (Goldshleger et al, 1987;Stiirmer et al, 1991;Heyse et al, 1994); the conformational transition, (Na3)E,-P --P-E2(Na3) as well as the subsequent release of Na+ ions, P-E2(Na3) -> P-E2 + 3 Nae, Gadsby, 1986, 1989;Lauger and Apell, 1988; Borlinghaus et al, 1987;Sturmer et al, 1991;Stiirmer and Apell, 1992;Gadsby et al, 1993;Sagar and Rakowski, 1994;Heyse et al, 1994); and finally the extracellular binding of K+ ions, P-E2 + 2 Kext P-E2(K2), (Rakowski et al, 1991;Stiirmer et al, 1991;Vasilets et al, 1991;Sagar and Rakowski, 1994).…”

Section: Introductionmentioning

confidence: 99%

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Electrogenicity of the sodium transport pathway in the Na,K-ATPase probed by charge-pulse experiments

Wuddel¹,

Apell²

1995

Biophysical Journal

101

191

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A charge-pulse technique was designed to measure charge movements in the Na-transport mode of the Na,K-ATPase in membrane fragments adsorbed to a planar lipid bilayer with high time resolution. 1) Na+ transport was measured as a function of membrane potential, and 2) voltage-dependent extracellular ion binding and release were analyzed as a function of Na+ concentration and membrane potential. The results could be fitted and explained on the basis of a Post-Albers cycle by simulations with a mathematical model. The minimal reaction sequence explaining the electrogenicity of the pump consists of the following steps: (Na3)E1-P <--> P-E2(Na3) <--> P-E2(Na2) <--> P-E2(Na) <--> P-E2. The conformational change, E1 to E2, is electrogenic (beta 0 < or = 0.1) and the rate-limiting step of forward Na+ transport with a rate constant of 25 s-1 (T = 20 degrees C). The first ion release step, P-E2(Na3) <--> P-E2(Na2), is the major charge translocating process (delta 0 = 0.65). It is probably accompanied by a protein relaxation in which the access structure between aqueous phase and binding site reduces the dielectric distance. The release of the subsequent Na+ ions has a significantly lower dielectric coefficient (delta1 = delta 2 = 0.2). Compared with other partial reactions, the ion release rates are fast (1400 s-1, 700 s-1, and 4000 s-1). On the basis of these findings, a refined electrostatic model of the transport cycle is proposed.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Electrogenicity of the sodium transport pathway in the Na,K-ATPase probed by charge-pulse experiments

Wuddel¹,

Apell²

1995

Biophysical Journal

101

191

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“…The coefficient of determination (R 2 ) for goodness of fit is also provided. (Vasilets et al, 1991). The Na i -loading solution was washed out and replaced with KCM211 external solution plus 10 µM ouabain immediately before recording currents.…”

Section: Single-channel Recordingmentioning

confidence: 99%

Activation of Slo2.1 channels by niflumic acid

Dai

Garg

Sanguinetti

2010

Journal of General Physiology

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Slo2.1 channels conduct an outwardly rectifying K+ current when activated by high [Na+]i. Here, we show that gating of these channels can also be activated by fenamates such as niflumic acid (NFA), even in the absence of intracellular Na+. In Xenopus oocytes injected with <10 ng cRNA, heterologously expressed human Slo2.1 current was negligible, but rapidly activated by extracellular application of NFA (EC50 = 2.1 mM) or flufenamic acid (EC50 = 1.4 mM). Slo2.1 channels activated by 1 mM NFA exhibited weak voltage dependence. In high [K+]e, the conductance–voltage (G-V) relationship had a V1/2 of +95 mV and an effective valence, z, of 0.48 e. Higher concentrations of NFA shifted V1/2 to more negative potentials (EC50 = 2.1 mM) and increased the minimum value of G/Gmax (EC50 = 2.4 mM); at 6 mM NFA, Slo2.1 channel activation was voltage independent. In contrast, V1/2 of the G-V relationship was shifted to more positive potentials when [K+]e was elevated from 1 to 300 mM (EC50 = 21.2 mM). The slope conductance measured at the reversal potential exhibited the same [K+]e dependency (EC50 = 23.5 mM). Conductance was also [Na+]e dependent. Outward currents were reduced when Na+ was replaced with choline or mannitol, but unaffected by substitution with Rb+ or Li+. Neutralization of charged residues in the S1–S4 domains did not appreciably alter the voltage dependence of Slo2.1 activation. Thus, the weak voltage dependence of Slo2.1 channel activation is independent of charged residues in the S1–S4 segments. In contrast, mutation of R190 located in the adjacent S4–S5 linker to a neutral (Ala or Gln) or acidic (Glu) residue induced constitutive channel activity that was reduced by high [K+]e. Collectively, these findings indicate that Slo2.1 channel gating is modulated by [K+]e and [Na+]e, and that NFA uncouples channel activation from its modulation by transmembrane voltage and intracellular Na+.

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“…The grouped parameters FNpa represent the theoretical maximum pump current (/max) that will result under conditions in which the voltage-dependent rate coefficients are no longer rate determining, but instead the pump cycle is rate limited by voltage-independent steps. Because there is considerable variability in the density of pump sites in different oocytes (Schweigert, Lafaire, and Schwarz, 1988;Vasilets, Omay, Ohta, Noguchi, Kawamura, and Schwarz, 1991) FNoa was allowed to vary in the fit to each experimental case. No consistent trend in the value of FNpa is seen in Table III as [Na]o is increased.…”

Section: Calculation Of Individual Rate Coefficients Given the Density Of Pump Sites (N)mentioning

confidence: 99%

Access channel model for the voltage dependence of the forward-running Na+/K+ pump.

Sagar

Rakowski

1994

The Journal of General Physiology

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The voltage dependence of steady state current produced by the forward mode of operation of the endogenous electrogenic Na+/K + pump in Na+-loaded Xenopus oocytes has been examined using a two-microelectrode voltage clamp technique. Four experimental cases (in a total of 18 different experimental conditions) were explored: variation of external [Na § ([Na], and activation of pump current by various [K]o at 0, 15, and 120 mM [Na]o (tetramethylammonium replacement). Ionic current through K + channels was blocked by Ba 2+ (5 raM) and tetraethylammonium (20 raM), thereby allowing pump-mediated current to be measured by addition or removal of external K § Control measurements and corrections were made for pump current run-down and holding current drift. Additional controls were done to estimate the magnitude of the inwardly directed pump-mediated current that was present in K+-free solution and the residual K+-channel current. A pseudo two-state access channel model is described in the Appendix in which only the pseudo first-order rate coefficients for binding of external Na § and K § are assumed to be voltage dependent and all transitions between states in the Na+/K + pump cycle are assumed to be voltage independent. Any three-state or higher order model with only two oppositely directed voltage-dependent rate coefficients can be reduced to an equivalent pseudo two-state model. The steady state current-voltage (I-V) equations derived from the model for each case were simultaneously fit to the I-V data for all four experimental cases and yielded least-squares estimates of the model parameters. The apparent fractional depth of the external access channel for Na § is 0.486 _+ 0.010; for K + it is 0.256 -+ 0.009. The Hill coefficient for Na + is 2.18 -0.06, and the Hill coefficient for K + (which is dependent on [Na]o) ranges from 0.581 --0.019 to 1.35 -0.034 for 0 and 120 mM [Na]o, respectively. The model provides a reasonable fit to the data and supports the hypothesis that under conditions of saturating internal [Na+], the principal voltage dependence of the Na+/K + pump cycle is a consequence of the existence of an external high-field access channel in the pump molecule through which Na + and K § ions must pass in order to reach their binding sites.

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Stimulation of the Na+/K+ pump by external [K+] is regulated by voltage-dependent gating.

Cited by 57 publications

References 15 publications

Electrogenicity of the sodium transport pathway in the Na,K-ATPase probed by charge-pulse experiments

Electrogenicity of the sodium transport pathway in the Na,K-ATPase probed by charge-pulse experiments

Activation of Slo2.1 channels by niflumic acid

Access channel model for the voltage dependence of the forward-running Na+/K+ pump.

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