Characterization of the electrogenic sodium pump in cardiac Purkinje fibres
Abstract
1. The Na pump is examined in sheep cardiac Purkinje fibres using a two micro-electrode voltage clamp technique.
2. After reducing the external K concentration, [K]o, to zero for 2 min or more, subsequent addition of an `activator cation' (known to activate the Na pump in other preparations) produces a transient increase of outward current. This outward current transient is abolished by 10-5 M-strophanthidin (cf. Gadsby & Cranefield, 1979a).
3. It is concluded that this transient increase of outward current is a result of a transient stimulation of the sodium pump by the raised [Na]i following exposure to 0-Ko. Although this current transient may reflect the activity of an electrogenic Na pump, it is difficult to use K as the activator cation to establish this point. This is due to the extracellular K depletion that occurs during Na pump reactivation and the subsequent change that this K depletion produces in the current—voltage relationship of the Purkinje fibre.
4. Rbo or Cso have been used instead of Ko to reactivate the Na pump when examining the transient increase of outward current. On adding either of these cations after exposing a preparation to a solution without such `activator cations', the outward current transient is relatively voltage independent over a wide range of potentials (-90 to +10 mV). It is concluded that, following the addition of Rbo or Cso, the transient increase of outward current is a direct measure of the transient increase of the electrogenic Na pump current.
5. Increasing [Rb]o or [Cs]o over the range of 0-40 mM increases the rate of decay of the electrogenic Na pump current transient. Using a simple model (cf. Rang & Ritchie, 1968), it is shown that the decay rate constant of the electrogenic Na pump current transient is a good measure of the degree of activation of the external site of the Na pump. At a given concentration of activator cation, Rbo produces a greater activation of the Na pump than does Cso. The K0.5 for Rbo is 6.3 mM and for Cso is 14.2 mM. Lio activates the Na pump more weakly than Rbo and Cso.
6. The coupling ratio of the Na pump is shown to be independent of Rbo or Cso over the range 2-40 mM. Furthermore, consistent with the results of Gadsby & Cranefield (1979a), the coupling ratio is independent of Nai over the range considered.
7. The Q10 for the electrogenic Na pump current transient varies between 1.6 and 2.3 over the range of temperature 26-46 °C.
8. A maximum Na pump current of about 0.78 μA cm-2 is obtained. Assuming a coupling ratio of 3Na/2K, the rate of Na ion transport into the cell is estimated to be about 23 p-mole cm-2 sec-1. Assuming a Na pump turnover of 150 sec-1, we estimate that there are about 1000 Na pump sites per μm2 of cell surface.
9. We conclude that the electrogenic Na pump current transient provides a good measure of the activity of the Na pump when Rb or Cs are used as `activator cations'. This measure can be used in the intact preparation to investigate the relationship between Na pump rate and other cellular events such as the regulation of tension (Eisner & Lederer, 1980).
