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In the rodent brain, astrocytes are known to be the primary source of kynurenate (KYNA), an endogenous antagonist of both the glycine(B) and the alpha7 nicotinic acetylcholine receptor. In the present study, primary human astrocytes were used to examine the characteristics and regulation of de novo KYNA synthesis in vitro. To this end, cells were exposed to KYNA's bioprecursor L-kynurenine, and newly formed KYNA was recovered from the extracellular milieu. The production of KYNA was stereospecific and rose with increasing L-kynurenine concentrations, reaching a plateau in the high microM range. In an analogous experiment, astrocytes also readily produced and liberated the potent, specific glycine(B) receptor antagonist 7-chlorokynurenate from L-4-chlorokynurenine. KYNA synthesis was dose-dependently reduced by L-leucine or L-phenylalanine, two amino acids that compete with L-kynurenine for cellular uptake, and by aminooxyacetate, a non-specific aminotransferase inhibitor. In contrast, KYNA formation was stimulated by 5 mM pyruvate or oxaloacetate, which act as co-substrates of the transamination reaction. Aglycemic or depolarizing (50 mM KCl or 100 microM veratridine) conditions had no effect on KYNA synthesis. Subsequent studies using tissue homogenate showed that both known cerebral kynurenine aminotransferases (KAT I and KAT II) are present in astrocytes, but that KAT II appears to be singularly responsible for KYNA formation under physiological conditions. Taken together with previous results, these data suggest that very similar mechanisms control KYNA synthesis in the rodent and in the human brain. These regulatory events are likely to influence the neuromodulatory effects of astrocyte-derived KYNA in the normal and diseased human brain.

Intrastriatal injection of the endogenous excitotoxin quinolinate in experimental animals causes a lesion which duplicates many features of Huntington's disease (HD). This lesion can be prevented by a related metabolite, kynurenate. Since kynurenate levels are reduced in the HD neostriatum, a deficiency in brain kynurenate may be the cause of neuron loss in HD. In order to investigate the relationship between excitotoxic neurodegeneration and kynurenate formation, effects of a unilateral quinolinate injection on several measures of kynurenate metabolism were studied in the rat striatum and substantia nigra. Within 2 hours, quinolinate caused an approximately 100% increase in striatal kynurenate levels in the absence of changes in its bioprecursor L-kynurenine or its biosynthetic enzymes kynurenine aminotransferases (KATs) I and II. This increase was more dramatic after 2 days (+735%) and was accompanied by an increase in L-kynurenine (+182%). No change or a slight decrease in enzyme activities were detected at this time-point. More chronic excitotoxic lesions produced a substantial increase in kynurenate levels (by approximately 2-, 4- and 4-fold, respectively, after 7 days, 1 and 5 months). Lesion-induced changes in KAT II activity essentially paralleled those seen with kynurenate, whereas KAT I remained slightly decreased at all timepoints. Nigral KAT II activity was increased ipsilaterally 2 days, 1 and 5 months after the striatal quinolinate injection. Kinetic analyses, performed in the striatum 5 months after the quinolinate injection, showed an almost 3-fold decrease in Km values for KAT II in the absence of v(max) changes. These findings indicate that 1) different mechanisms regulate kynurenate production at different stages after an intrastriatal quinolinate injection; 2) an increased substrate affinity to KAT II is responsible for the elevation of kynurenate in the chronically lesioned rat striatum; and 3) qualitative differences in kynurenate metabolism exist between the HD neostriatum and the excitotoxin-lesioned rat striatum, supporting the idea that (a decrease in) kynurenate tone may play a primary role in the pathophysiology of HD.