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

Abstract

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.

Altered Na + transport after an intracellular α-subunit deletion reveals strict external sequential release of Na + from the Na/K pump

“…Thus, the position of the I P -V curve on the voltage axis depends on the [Na o ϩ ] and the apparent Na o ϩ affinity. Mutations that affect Na o ϩ binding have been shown to induce parallel shifts in this curve across the voltage axis (6), similar to those produced by changing [Na o ϩ ] in normal pumps (9,13). In the absence of K o ϩ , but presence of Na o ϩ , the pump shuttles between Na ϩ -bound and unbound states in a voltage-dependent manner.…”

“…The more voltage-sensitive partial reactions are those reactions involving ion binding and release through a channel-like structure accessible from the extracellular space ( Fig. 1) (7)(8)(9)(10). Because the Na ϩ -transport branch is more voltage-sensitive (8, 11) than the K ϩ -transport branch (7,12), under physiological conditions with high Na o ϩ , negative transmembrane voltages inhibit the pump by forcing Na ϩ ions back into the access channel producing a steep positive slope in the steady-state Na/K pump current (I P ) versus V curve.…”

Energy landscape of the reactions governing the Na + deeply occluded state of the Na + /K + -ATPase in the giant axon of the Humboldt squid

“…By establishing the Na þ and K þ gradients across cell membranes, the Na þ ∕K þ pump enables action potentials, synaptic signaling, and most solute transport in and out of cells. Two consequences of the unequal transport stoichiometry of Na þ and K þ are that steady pumping produces an outwardly directed current (1), proportional in magnitude to the turnover rate (2), which can be monitored electrically (3)(4)(5)(6)(7)(8), and that at least one step in the transport cycle must move charge through the membrane field (9). The latter implies that, under favorable conditions, charge relaxations following voltage jumps can be used to learn details about specific steps during the transport cycle (10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28).…”

A simple recipe for setting up the flux equations of cyclic and linear reaction schemes of ion transport with a high number of states: The arrow scheme

“…For example, for m D 1 this is (7) In the definition of D Cn m (Eqs. 7,8), all indices i greater than n are equivalent to i-n, due to the cyclic nature of the model, as illustrated by the equations for…”

“…11 Of special interest are cyclic reaction schemes. It is immediately obvious that they apply to electrogenic pumps 3,8,12,13,16,19,22 and cotransporters. 6,14,15,23,24 However, it turned out that cyclic reaction schemes also provided the best fits of IV curves from ion channels, 7,[25][26][27] from bacteriorhodopsin, 9,17 the viral M2 channel 11 or from Polytheonamide B, a marine cytotoxic b-helical peptide.…”