Effects of thiopental on transport and metabolism of glutamate in cultured cerebellar granule neurons.
This study was performed to analyze the effects of the barbiturate thiopental on neuronal glutamate uptake, release and metabolism. Since barbiturates are known to bind to the GABA(A) receptor, some experiments were carried out in the presence of GABA. Cerebellar granule neurons were incubated for 2 h in medium containing 0.25 mM [U-(13)C]glutamate, 3 mM glucose, 50 microM GABA and 0.1 or 1 mM thiopental when indicated. When analyzing cell extracts, it was surprisingly found that in addition to glutamate, aspartate and glutathione, GABA was also labeled. In the medium, label was observed in glutamate, aspartate and lactate. Glutamate exhibited different labeling patterns, indicating metabolism in the tricarboxylic acid cycle, and subsequent release. A net uptake of [U-(13)C]glutamate and unlabeled glucose was seen under all conditions. The amounts of most metabolites synthesized from [U-(13)C]glutamate were unchanged in the presence of GABA with or without 0.1 mM thiopental. In the presence of 1 mM thiopental, regardless of the presence of GABA, decreased amounts of [1,2, 3-(13)C]glutamate and [U-(13)C]aspartate were found in the medium. In the cell extracts increased [U-(13)C]glutamate, [1,2, 3-(13)C]glutamate, labeled glutathione and [U-(13)C]aspartate were observed in the 1 mM thiopental groups. Glutamate efflux and uptake were studied using [(3)H]D-aspartate. While efflux was substantially reduced in the presence of 1 mM thiopental, this barbiturate only marginally inhibited uptake even at 3 mM. These results may suggest that the previously demonstrated neuroprotective action of thiopental could be related to its ability to reduce excessive glutamate outflow. Additionally, thiopental decreased the oxidative metabolism of [U-(13)C]glutamate but at the same time increased the detectable metabolites derived from the TCA cycle. These latter effects were also exerted by GABA.
Qu
H
H
Department of Pharmacology and Toxicology, Norwegian University of Science and Technology, Trondheim.
Waagepetersen
H S
HS
van Hengel
M
M
Wolt
S
S
Dale
O
O
Unsgård
G
G
Sletvold
O
O
Schousboe
A
A
Sonnewald
U
U
D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism; D001693:Biological Transport, Active / Q000187:drug effects; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D003594:Cytoplasmic Granules / Q000187:drug effects / Q000378:metabolism; D004734:Energy Metabolism / Q000187:drug effects; D018757:GABA Modulators / Q000494:pharmacology*; D008401:Gas Chromatography-Mass Spectrometry; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D013874:Thiopental / Q000494:pharmacology*; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
D018757:GABA Modulators; D001224:Aspartic Acid; D005680:gamma-Aminobutyric Acid; D013874:Thiopental