Release of noradrenaline from slices of cat spleen by pre-treatment with calcium, strontium and barium

Abstract

1. Spleen slices pre-incubated for different periods at 4 degrees C in Krebs solution containing varying concentrations of calcium, up to 96 mM, lost their endogenous noradrenaline stores when reincubated in normal Krebs solution at 37 degrees C for 2 hr. Rate of loss of noradrenaline was roughly related to the calcium concentration of the pre-incubation medium and the pre-exposure time.2. Pre-treatment with isotonic barium or strontium (96 mM) Krebs solution also induced release of noradrenaline from spleen slices when re-exposed to normal Krebs solution. Barium was more effective than either calcium or strontium.3. The enhanced release induced by calcium pre-treatment occurred in the absence of calcium, with or without EGTA.4. Tissue calcium concentration of spleen slices was 0.68 m-mole/kg. Pre-treatment of slices with normal or 96 mM calcium-Krebs solution for 4 hr at 4 degrees C increased the calcium concentration to 2.57 and 9.9 m-mole/kg, respectively.5. Ouabain, which caused a dose-dependent release of noradrenaline, did not modify the release induced by calcium pre-treatment.6. Spleen slices prepared from cats anaesthetized with sodium pentobarbitone instead of ether were resistant to noradrenaline depletion by calcium pre-treatment.7. Evoked release of [(3)H]noradrenaline by high potassium from calcium-pre-treated slices did not occur in the absence of external calcium, even though the calcium pre-treatment enhanced the tissue concentration of this ion by nearly tenfold.8. Net uptake of noradrenaline in normal and in treated slices whose noradrenaline content was severely reduced by barium pre-treatment or sodium withdrawal was comparable.9. Specific activity of released and endogenous [(3)H]noradrenaline increased as the tissue stores of noradrenaline were reduced.10. It is suggested that the spontaneous loss of tissue noradrenaline after pre-treatment with high-calcium solution was due to inhibition of sodium-potassium-activated ATPase by intracellular accumulation of calcium ions. Evidence is presented to suggest that vesicles depleted of their endogenous transmitter by pre-treatment with calcium, strontium or barium, or by sodium withdrawal, are re-used for the storage and release of exogenous noradrenaline.