Movements of labelled sodium ions in isolated rat superior cervical ganglia

Abstract

1. Isolated rat superior cervical ganglia were incubated in Krebs solution containing (24)Na and carbachol for 4 min at 25 degrees C. They were then washed at 3 degrees C for 15 min to remove extracellular (24)Na and the efflux of residual intracellular (24)Na stimulated by warming to 25 degrees C.2. During the 15 min wash at 3 degrees C desaturation curves became exponential with a rate constant of 0.012 +/- 0.001 min(-1) (n = 24). This was assumed to represent loss of intracellular (24)Na, and initial uptake of (24)Na was calculated therefrom by back-extrapolation to zero wash-time. After 4 min in (24)Na + 180 muM carbachol intracellular [(24)Na] so calculated was 61.6 +/- 3.1 mM (n = 18), representing 83% labelling of intracellular Na. In the absence of carbachol intracellular [(24)Na] was 10.0 +/- 0.5 mM, representing 49% labelling. Extracellular Na was labelled by > 90% after 4 min in (24)Na. The apparent rate constant for washout of extracellular (24)Na was 0.6 min(-1) at 3 degrees C and 0.95 min(-1) at 25 degrees C.3. The loss of the residual intracellular (24)Na during temperature stimulation was interpreted quantitatively in terms of an exponential decline of the bulk of intracellular (24)Na with an extrusion rate constant of 0.39 +/- 0.1 min(-1) (n = 18), efflux being delayed by passage through the extracellular space with an effective rate constant of 0.8-1.2 min(-1).4. The peak rate constant (k(C)) for the desaturation curve at 25 degrees C was 0.35 +/- 0.01 min(-1). An Arrhenius plot of log k(C)/T degrees K(-1) yielded a two-stage linear regression with a transition at 20 degrees C. Activation energies of 8 and 31 kcal. mole(-1) were calculated above and below this transition respectively.5. Omission of K from the 25 degrees C temperature-stimulating solution reduced k(C) by 62%. The K-sensitive component of extrusion rate constant was a hyperbolic function of [K](e) with half-saturation at 5.6 mM-[K](e) and maximum k(C) of 0.58 min(-1).6. Cyanide (2 mM), 2,4-dinitrophenol (1 mM) and ouabain (1.4 mM) reduced k(C) by 50-90%. The half-maximally inhibiting concentration of ouabain was about 60 muM.7. Substitution of sucrose, Li or choline for external Na did not reduce the extrusion rate of (24)Na in either 6 mM-[K](e) or 0 mM-[K](e). Li stimulated (24)Na extrusion in Na-free, K-free solution.8. The properties of the ganglionic Na pump deduced from rates of temperature-stimulated (24)Na extrusion accord with the view that the ganglion hyperpolarization observed after Na loading by exposure to nicotinic depolarizing agents results from electrogenic Na extrusion. A comparable hyperpolarization is observed after temperature stimulation following Na loading.