Demonstration of kynurenine aminotransferases I and II and characterization of kynurenic acid synthesis in cultured cerebral cortical neurons.
The present study characterizes the synthesis of kynurenic acid (KYNA) from exogenously added kynurenine and its regulation by extrinsic factors, in cultured cerebral cortical neurons and, for comparison, in astrocytes incubated under identical conditions. The neuronal culture showed positive immunostaining for both kynurenic acid aminotransferase (KAT) isoforms I and II. Neurons synthesized KYNA at a rate about 2.3 times higher than astrocytes. Neuronal, but not astrocytic, KYNA synthesis was lowered approximately 30% by ionotropic glutamate receptor agonists [(R,S)-3-hydroxy-5-methoxyloxasole-4-propionic acid (AMPA; 100 microM) and N-methyl-D-aspartic acid (NMDA; 100 microM)] and depolarizing agents [KCl (50 mM) and 4-aminopyridine (4-AP; 10 microM)]. Neuronal and astrocytic synthesis alike were vulnerable to inhibition exerted by the aminotransferase inhibitor aminooxyacetic acid (AOAA), glutamate (IC50: 31 and 85 microM, respectively), substrates of the L-amino transport system [leucine (Leu); IC50: 19 and 42 microM, respectively] and 2-aminobicyclo[2,2,1]heptane-2-carboxylic acid (BCH; IC50: 19 and 28 microM, respectively). Glutamine (Gln), which is a metabolic precursor of glutamate in astrocytes and L-system substrate in both cell types, inhibited KYNA synthesis both in neurons and in astrocytes (IC50: 268 and 318 microM, respectively). alpha-Ketoisocaproic acid (KIC), a Leu transamination product that is produced mainly in astrocytes and shuttled to neurons to modulate intraneuronal concentration of glutamate, stimulated KYNA synthesis in neurons but did not affect the synthesis in astrocytes. In conclusion, this study is the first to demonstrate active, regulation-prone KYNA synthesis in neurons.
Rzeski
Wojciech
W
Department of Virology and Immunology, Institute of Microbiology and Biotechnology, Maria Curie-Skodowska University, Lublin, Poland.
Kocki
Tomasz
T
Dybel
Anna
A
Wejksza
Katarzyna
K
Zdzisińska
Barbara
B
Kandefer-Szerszeń
Martyna
M
Turski
Waldemar A
WA
Okuno
Etsuo
E
Albrecht
Jan
J
D000818:Animals; D000906:Antibodies; D001253:Astrocytes / Q000166:cytology / Q000201:enzymology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology*; D018698:Glutamic Acid / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000276:immunology / Q000378:metabolism*; D007736:Kynurenic Acid / Q000378:metabolism; D007930:Leucine / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000201:enzymology*; D051381:Rats; D017208:Rats, Wistar; D000637:Transaminases / Q000276:immunology / Q000378:metabolism*
D000906:Antibodies; D007527:Isoenzymes; D005973:Glutamine; D018698:Glutamic Acid; D000637:Transaminases; C021481:kynurenine-oxoglutarate transaminase; D007930:Leucine; D007736:Kynurenic Acid