Intracellular [Ca2+] and Vo2 after manipulation of the Free-energy of the Na+/Ca2+-exchanger in isolated rat ventricular myocytes

We have investigated whether the Na+/Ca2+-exchanger has a functional regulatory role in the control of oxidative metabolism in suspensions of isolated rat ventricular myocytes. Therefore we simultaneously measured intracellular [Ca2+] ([Ca2+]staggeredi) with Indo-1 and respiratory rate (Vo2) after abrupt manipulation of the free-energy of the Na+/Ca2+-exchanger (ΔGexch). The average fraction of viable myocytes was about 90% (82% rod-shaped plus 8% viable round cells). ΔGexch was manipulated either by an abrupt decrease of [Na+]o (in combination with an increase of [K+]o or [Ca2+]o) or by changing membrane potential and/or intracellular cation activities with the use of gramicidin or veratridine. A change of extracellular cation composition caused a transient increase of [Ca2+]staggeredi and Vo2, with peak values after 30 to 40 s and a new steady state near control values after 180 to 240 s. Peak values of the transients were associated with the magnitude of the thermodynamic disturbance. Inhibition of sodium-pump activity with ouabain greatly enhanced peak values and reduced the rate of return to a new steady state. Reversal of the initial disturbance of ΔGexch by restoring [Na+]o or reduction of [Ca2+]o during the time course of the transients greatly accelerated return to a new steady-state. An increase of sarcolemmal sodium permeability with the Na-channel ligand veratridine or manipulation of [Na+]staggeredi and [K+]staggeredi with the Na+/K+-exchanger gramicidin caused monophasic increase of both [Ca2+]staggeredi and Vo2. The relationship between Vo2 and [Ca2+]staggeredi was the same, irrespective of the nature of the intervention (either extracellular or intracellular manipulation of ΔGexch). We conclude that cytoplasmic [Ca2+] (thermodynamically controlled by the Na+/Ca2+-exchanger) is a major regulator of the respiratory rate in (quiescent) myocytes.