Release of alpha-ketoglutarate, malate and succinate from cultured astrocytes: possible role in amino acid neurotransmitter homeostasis

Abstract

The rates of release of the tricarboxylic acid (TCA) cycle constituents alpha-ketoglutarate (alpha-KG), malate and succinate were determined in cultured mouse cerebellar astrocytes, cerebellar granule neurons and cerebral cortical neurons. In addition, its dependence on the external HCO3- concentration was investigated together with effects of K+, glutamate and glutamine. The rate of release of these TCA cycle constituents was linear with time for at least 48 h regardless of the cell type. The release was for all 3 compounds much higher in the astrocytes (13.1, 3.8 and 1.5 nmol.h-1.mg-1 for alpha-KG, malate and succinate, respectively) than in cerebellar (6.5 and 1.5 for alpha-KG and malate) and cortical (3.5 and 1.2 for alpha-KG and malate) neurons. Release of succinate in the neurons could not be determined accurately due to the sensitivity of the assay. In the astrocytes the release of alpha-KG and malate was dependent on HCO3- in a saturable manner with Km values around 6 and 1 mM for alpha-KG and malate, respectively. The release of alpha-KG and malate from astrocytes was stimulated by glutamate (0.5 mM) whereas K+ (15 and 55 mM) and glutamine (0.5 mM) had no effect. The results clearly demonstrate that astrocytes but not neurons release appreciable amounts of TCA cycle intermediates reflecting the presence of pyruvate carboxylase in these cells. The exact functional importance of this release remains to be established but it could play some albeit a minor quantitative role for neuronal homeostasis of the neurotransmitter amino acids glutamate and GABA.