R D Vaughan-Jones

The dependence of sodium pumping and tension on intracellular sodium activity in voltage-clamped sheep Purkinje fibres.

1. Intracellular Na activity (aiNa) was measured in sheep cardiac Purkinje fibres using a recessed‐tip Na+‐sensitive micro‐electrode. The membrane potentials was controlled with a two‐micro‐electrode voltage clamp. Tension was measured simultaneously. 2. Removing external K produced a rise of aiNa and both twitch and tonic tension. On adding 4‐10 mM‐[Rb]0 to reactivate the Na‐K pump aiNa and tension declined. An electrogenic Na pump current transient accompanied the fall of aiNa. 3. The half‐time of decay of the electrogenic Na pump current transient was similar to that of aiNa, (mean tNa0.5/tI0.5 = 0.97 +/‐ 0.03 (S.E.M.; n = 28)). Following the re‐activation of the Na‐K pump, the electrogenic Na pump current transient was linearly related to aiNa. 4. The duration of exposure to K‐free, Rb‐free solutions was varied to change the level of aiNa. On subsequently re‐activating the Na‐K pump with 10 mM‐[Rb]0, the ratio of the charge extruded to the total change of aiNa was constant. It is concluded that the fraction of Na extruded electrogenically is unaffected by changes of aiNa. About 26% of the total Na extrusion appeared as charge transfer. 5. The relationship between tonic tension and aiNa was usually different during Na‐K pump inhibition in a K‐free, Rb‐free solution compared with the relationship during Na‐K pump re‐activation. In general, a given aiNa was associated with a greater level of tonic tension during Na‐K pump inhibition compared with that during pump re‐activation. A similar hysteresis was often seen between twitch tension and aiNa.

The control of tonic tension by membrane potential and intracellular sodium activity in the sheep cardiac Purkinje fibre.

Intracellular Na activity (aiNa) was measured with recessed‐tip, Na‐selective micro‐electrodes in voltage‐clamped sheep cardiac Purkinje fibres. Tension was measured simultaneously. aiNa was increased reversibly either by exposing the preparation to K‐free, Rb‐free solution of by adding the cardioactive steroid strophanthidin. An increase of aiNa produced an increase of tonic tension which was larger at depolarized membrane potentials. At sufficiently negative membrane potentials, changes of aiNa (over the range 6‐30 mM) had no effect on tonic tension. Therefore, both an increase of aiNa and a depolarization are required to increase tonic tension. It is concluded that either a low level of aiNa or a large negative membrane potential is sufficient to maintain a low intracellular Ca concentration. Tonic tension was measured as a function of aiNa. At a given membrane potential the relationship can be described empirically by an equation of the form: tonic tension = b(aiNa)y, where y is a constant and b depends on membrane potential. In five experiments y was found to be 3.7 +/‐ 0.7 (mean +/‐ S.E.M.) over a range of potentials from ‐60 to ‐10 mV. Tonic tension was measured as a function of membrane potential. At a given aiNa the relationship can be described approximately as: tonic tension = k exp (aV), where a is a constant and k depends on aiNa. In five experiments a was found to be 0.06 +/‐ 0.01 mV‐1 (mean +/‐ S.E.M.). A depolarization of 10 mV increases tonic tension by the same amount as does an increase of aiNa that is equivalent to a 3.7 mV change of the Na equilibrium potential, ENa. Hence ENa is nearly 3 times more effective than membrane potential in controlling tonic tension. During a prolonged depolarization (several minutes) the initial increase of tonic tension decays gradually. This is associated with a fall of aiNa. The relationship between tonic tension and aiNa is similar to that seen when aiNa is increased by inhibiting the Na pump. It is concluded that the fall of aiNa is responsible for the decay of tonic tension. The changes of tonic tension reported in this paper are consistent with the effects of aiNa and membrane potential on a voltage‐dependent Na‐Ca exchange. The possibility that a voltage‐dependent Ca channel contributes to tonic tension is also discussed.

The quantitative relationship between twitch tension and intracellular sodium activity in sheep cardiac Purkinje fibres.

Tension was measured in voltage clamped sheep cardiac Purkinje fibres while simultaneously measuring the intracellular Na activity (aiNa) with a recessed‐tip, Na‐selective micro‐electrode. Inhibiting the Na‐K pump either by exposing the preparation to a K‐free solution or by adding the cardioactive steroid strophanthidin increased both aiNa and twitch tension and resulted in the development of tonic tension, after‐contractions and a transient inward current (ITI). The increase of twitch tension was present at lower aiNa than that required to produce the other phenomena. The relationship between the magnitude of the twitch tension and aiNa was always non‐linear. Twitch tension increased steeply with aiNa at first but the relationship flattened off at higher aiNa and tension eventually decreased. Over the steep range, the relationship between tension and aiNa could be represented as: twitch tension = b (aiNa)y where y had a mean value of 3.2. Changing membrane potential or [Ca2+]o changed b but had little effect on y. Mn (2 mmol/l) greatly decreased twitch tension but, at least initially, had little effect on tonic tension. The steep relationship between twitch tension and aiNa was seen, irrespective of whether the Na‐K pump was inhibited either by exposure to K‐free solution or to strophanthidin and whether the relationship was measured either when aiNa was increasing or after it had reached a steady state. The steep dependence of twitch tension on aiNa observed in the present work means that manoeuvres which produce even small changes of aiNa will have significant effects on contraction.

The effects of rubidium ions and membrane potentials on the intracellular sodium activity of sheep Purkinje fibres.

1. Intracellular Na activity, aiNa, was measured in voltage‐clamped sheep cardiac Purkinke fibres. 2. Increasing [Rb]0 from 0 to 4 mM in K‐free solutions (at a fixed membrane potential) decreased aiNa. Further increases of [Rb]0 (up to 20 mM) had little or no effect. 3. Following exposure to Rb‐free, K‐free solution, the addition of a test concentration of Rb produced an exponential decrease of aiNa. The rate constant of decay of aiNa increased with increasing [Rb]0 over the measured range (0‐20 mM). 4. The accompanying electrogenic Na pump current transient decayed with the same rate constant as aiNa over the range of [Rb]0 examined. During this decay the electrogenic Na pump current was a linear function of aiNa. Increasing [Rb]0 increased the steepness of the dependence of the electrogenic current on aiNa. 5. A constant fraction of the net Na efflux was electrogenic. This fraction was not affected by varying [Rb]0 over the range 0‐20 mM. 6. Using a simple model, it is shown that the dependence of steady‐state aiNa on [Rb]0 is half‐saturated by less than 1 mM‐[Rb]0. The rate constant of decay of aiNa and the slope of the relationship between electrogenic Na pump current and aiNa are, however, better fitted with a lower affinity for Rb (K0.5 = 4 mM‐[Rb]0). 7. Depolarizing the membrane potential with the voltage clamp decreased aiNa; hyperpolarization increased it. These effects persisted in the presence of 10(‐5) M‐strophanthidin. An effect of membrane potential on the net passive Na influx can account for the observations. 8. The effects of membrane potential on the net passive Na influx were examined by measuring the maximum rate of rise of aiNa at different holding potentials after inhibiting the Na‐K pump in a K‐free, Rb‐free solution. Depolarization decreased the Na influx. 9. Using the constant field equation, the net passive Na influx was used to estimate the apparent Na permeability coefficient, PNa. This was between 0.8 x 10(‐8) and 1.5 x 10(‐8) cm sec‐1.

Do calcium-activated potassium channels exist in the heart?

Changes of intracellular Ca concentration in cardiac muscle have significant effects on transmembrane currents and many of these effects can be accounted for by postulating the existence of Ca-activated K channels in the heart. However, the evidence that such channels exist is equivocal. This is partly because of technical problems, for example the difficulty of identifying an individual ionic current amongst the many currents that exist in the heart. An additional problem, however, is posed by the fact that other currents may also be modulated by Ca ions. It is important therefore to distinguish between these currents and those caused by Ca-activated, K-specific channels. In this review we consider the evidence for Ca activated currents in the heart and, in particular, we discuss whether or not these currents are carried exclusively by K ions.

The Effects of Intracellular Na on Contraction and Intracellular pH in Mammalian Cardiac Muscle

Intracellular Na and pH were measured with recessed-tip ion-selective microelectrodes in voltage-clamped sheep cardiac Purkinje fibers. Intracellular Na activity (aiNa) was elevated by inhibiting the Na/K pump. This produced an increase of twitch tension that had a steep dependence on the increase of aiNa. These effects of aiNa on twitch tension are probably mediated by an Na-Ca exchange. An increase of aiNa also produced a component of tonic tension that appears to be produced directly by the Na-Ca exchange. The dependence of tonic tension and aiNa on membrane potential suggests that this exchange process may be voltage-sensitive. The increase of aiNa is associated with an intracellular acidification that appears to be secondary to an increase of [Ca2+]i produced by Na-Ca exchange. Therefore, as well as affecting [Ca2+]i, Na-Ca exchange can under some circumstances influence pHi indirectly, and this complicates the interpretation of changes in tension, since protons and Ca ions have opposite effects on contractile force.

Effects of Sodium Pump Inhibition on Contraction in Sheep Cardiac Purkinje Fibers

Publisher Summary It is well established that inhibiting the sodium pump increases the force of contraction of cardiac muscle. This effect appears to be mediated via an increase of intracellular sodium activity. One theory for the mechanism of this effect involves a Na/Ca exchange. Increasing would produce an increase of Ca influx and decrease Ca efflux through such an exchange, leading to increased contraction. To investigate the mechanism of the positive inotropic effects of a raised sodium activity one needs to measure sodium activity and tension simultaneously. The chapter investigates the steady-state relationship between sodium activity and tension; it also characterizes the effects of sudden changes of sodium activity. The only feasible way to measure sodium activity with adequate time resolution is with an ion-selective microelectrode. Recessed-tip Na-sensitive microelectrodes are used in this chapter. The experiments are performed on voltage-clamped sheep cardiac Purkinje fibers. The use of the voltage clamp makes it possible to investigate the effects of changes of sodium activity without secondary effects because of the changes of membrane potential. Sodium pump activity is controlled by changing the external rubidium concentration.