Glutamate and GABA: Major Players in Neuronal Metabolism
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Disturbance of neuronal metabolism has implications for a number of neurological and psychiatric conditions, and enhanced knowledge of this is important in developing new methods for treating such disorders. The present research was undertaken to aid understanding of diseases related to disturbance in glutamate and γ-amino butyric acid (GABA) metabolism. Two different types of neuronal cell cultures were used in these studies; one containing GABAergic neurons of cerebral neocortical origin and one containing cerebellar neurons. The latter consists primarily of glutamatergic granule neurons in addition to ~6 % GABAergic neurons and a small number of astrocytes. Metabolism was studied by 13C magnetic resonance spectroscopy (MRS) and mass spectrometry (MS) after adding 13C-labeled precursors ([1-13C]glucose, [U-13C]glutamate or [U-13C]glutamine) to the medium of these cultures. High performance liquid chromatography (HPLC) was used to quantify different amino acids in cell extracts and medium. The amount of protein in the cultures was determined to assess cell damage. In the cerebellar neuronal cultures, GABA was present in surprisingly large amounts compared to neocortical GABAergic cultures. 13C MRS experiments showed that GABA was actively synthesized throughout the culture period by the subpopulation of glutamate decarboxylase (GAD) positive (GABAergic) neurons and subsequently distributed to the other cells in the culture, i.e. to the granule neurons. The function of GABA in these glutamatergic neurons still remains uncertain; however, roles as neurotrophic and neuroprotective agent as well as substrate for energy production have been suggested. As shown previously, both glutamate and glutamine were shown to be excellent precursors for intermediary metabolism in cerebellar neurons. However, it was concluded that glutamate was preferred over glutamine, suggesting that these neurons rely more on reuptake of released glutamate than of supply of glutamine from astrocytes for glutamate homeostasis. This is not surprising when considering the cerebellar structure, with few astrocytes compared to neurons and a relatively large distance between astrocyte and synapse. Exposure of cerebellar cultures to 50 μM kainic acid (KA), a potent glutamate agonist, which is known to eliminate vesicular release of GABA in these cultures, only marginally affected glutamate and GABA metabolism, whereas increasing the KA concentration to 0.5 mM led to a reduction of both GABA and glutamate metabolism compared to unexposed cultures. It was previously believed that treatment with 50 μM KA eliminated the GABAergic neurons in cerebellar cultures, and KA has therefore been added in order to obtain essentially pure glutamatergic granule cell cultures. Although KA treatment abolishes vesicular GABA release, the GABA synthesizing cells are not eliminated by this treatment and still produce GABA in substantial amounts. Results from the present studies can only be understood in terms of inter- and intracellular compartmentation of metabolism. The main focus of metabolic compartmentation studies has been on the two compartments made up by neurons and astrocytes. One pathway previously believed to take place in the astrocytic but not in the neuronal compartment, is the pyruvate recycling pathway for complete tricarboxylic acid (TCA) cycle oxidation of glutamate. Despite this, in one of the present studies, such recycling was clearly present in both astrocytic and neuronal cultures from cerebellum.
Has partsSonnewald, Ursula; Olstad, Elisabeth; Qu, Hong; Babot, Zoila; Cristòfol, Rosa; Suñol, Christina; Schousboe, Arne; Waagepetersen, Helle. First direct demonstration of extensive GABA synthesis in mouse cerebellar neuronal cultures. J Neurochem. 91: 796-803, 2004.
Sonnewald, Ursula; Kortner, Trond M; Qu, Hong; Olstad, Elisabeth; Suñol, Christina; Bak, Lasse K.; Schousboe, Arne; Waagepetersen, Helle. Demonstration of extensive GABA synthesis in the small population of GAD positive neurons in cerebellar cultures by the use of pharmacological tools. Neurochemistry International. 48(6-7): 572-578, 2006.
Olstad, Elisabeth; Qu, Hong; Sonnewald, Ursula. Glutamate is preferred over glutamine for intermediary metabolism in cultured cerebellar neurons. Journal of Cerebral Blood Flow & Metabolism. 27: 811-820, 2007.