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Journal of Neuroscience, Vol 9, 1141-1149, Copyright © 1989 by Society for Neuroscience
Excitatory amino acid-stimulated uptake of 22Na+ in primary astrocyte cultures
HK Kimelberg, S Pang and DH Treble
Department of Biochemistry, Albany Medical College, Albany, New York 12208.
In this study we have found that L-glutamic acid, as well as being taken up
by a Na+-dependent mechanism, will stimulate the uptake of 22Na+ by primary
astrocyte cultures from rat brain in the presence of ouabain. By
simultaneously measuring the uptake of 22Na+ and L-3H- glutamate a
stoichiometry of 2-3 Na+ per glutamate was measured, implying electrogenic
uptake. Increasing the medium K+ concentration to depolarize the cells
inhibited L-3H-glutamate uptake, while calculations of the energetics of
the observed L-3H-glutamate accumulation also supported an electrogenic
mechanism of at least 2 Na+:1 glutamate. In contrast, kinetic analysis of
the Na+ dependence of L-3H-glutamate uptake indicated a stoichiometry of
Na+ to glutamate of 1:1, but further analysis showed that the stoichiometry
cannot be resolved by purely kinetic studies. Studies with glutamate
analogs, however, showed that kainic acid was a very effective stimulant of
22Na+ uptake, but 3H-kainic acid showed no Na+ -dependent uptake.
Furthermore, while L-3H-glutamate uptake was very sensitive to lowered
temperatures, glutamate-stimulated 22Na+ uptake was relatively insensitive.
These results indicate that glutamate-stimulated uptake of 22Na+ in primary
astrocytes cultures cannot be explained solely by cotransport of Na+ with
glutamate, and they suggest that direct kainic acid-type receptor induced
stimulation of Na+ uptake also occurs. Since both receptor and uptake
effects involve transport of Na+, accurate measurements of the Na+
:glutamate stoichiometry for uptake can only be done using completely
specific inhibitors of these 2 systems.
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