Pretreatment of rat brain slices with ouabain decreases chloride-dependent L-glutamate transport in synaptic membrane.

Possible roles of (Na(+)-K+)-ATPase on regulation of Cl(-)-dependent L-glutamate (L-Glu) transport in rat synaptic membrane were examined. Pretreatment of rat cerebral slices with 20 micrograms/ml veratrine, 56 mM K+ and 20 microM monensin increased Cl(-)-dependent L-[3H]-Glu uptake into membrane vesicles prepared from the slices. Pretreatment with (Na(+)-K+)-ATPase inhibitors, 100 microM ouabain, 100 microM strophanthidin, 100 microM vanadate and K(+)-free medium, decreased the uptake by 25-30%, while 5 microM A23187 and 10 microM ruthenium red had no effect. Ouabain (100 microM) caused the maximal effect in 10-20 min of incubation. The effect of (Na(+)-K+)-ATPase inhibitors was reversible and characterized by a decrease in Vmax of the uptake. Addition of 5 microM ouabain abolished the increases in the uptake induced by either veratrine, 56 mM K+ or monensin. Dose-inhibition curve of ouabain on the increased Cl(-)-dependent L-[3H]-Glu uptake was bi-phasic; ouabain at 0.5-5 microM selectively diminished the stimulatory effect of veratrine and inhibited the basal uptake at higher concentrations. Veratrine had no effect on L-[3H]-Glu uptake in 5 microM strophanthidin containing or K(+)-free medium. These results suggest the involvement of (Na(+)-K+)-ATPase in the veratrine-induced increase in Cl(-)-dependent L-Glu transport in rat brain synaptic membranes.