Notes: The production of the neuroinhibitory and neuroprotective metabolite kynurenic acid (KYNA) was investigated in rat brain by examining its biosynthetic enzyme, kynurenine aminotransferase (KAT). By using physiological (low micromolar) concentrations of the substrate L-kynurenine (KYN) and by determining the irreversible conversion of [3H] KYN to [3H] KYNA as a measure of KAT activity, a novel, simple, and sensitive assay was developed which permitted the detailed characterization of the enzyme. Only a single protein, which under routine assay conditions showed approximately equal activity with 2-oxoglutarate and pyruvate as the aminoacceptor, was found in rat brain. The enzyme was distributed heterogeneously between the nine brain regions studied, with the KAT-rich olfactory bulb displaying approximately five times higher activity than the cerebellum, the area with lowest KAT activity. In subcellular fractionation studies, the majority of KAT was recovered in mitochondria. In contrast to many known aminotransferases, partially purified KAT was shown to be highly substrate-specific. Thus, of the amino acids tested, only α-aminoadipate and tryptophan displayed moderate competition with KYN. Notably, 3-hydroxykynurenine, reportedly a very good substrate of KAT, competed rather poorly with KYN as well. Aminooxyacetic acid, a nonspecific transaminase inhibitor, blocked KAT activity with an apparent Ki of 5 μM. Kinetic analyses with partially purified rat brain KAT revealed a Km of 17 μMfor KYN with 1 mM 2-oxoglutarate, but a much higher Km(910 μM) with 1 mM pyruvate. Km values for 2-oxoglutarate and pyruvate were 150 and 160 μM, respectively. The cellular localization of KAT was examined in striatal homogenates obtained from rats 7 days after an intrastriatal injection of quinolinate. At that time of almost complete neuronal destruction and pronounced astrocytic proliferation, enzyme activity in the lesioned striatum was almost twice as high as in controls, suggesting a preferential astroglial localization of brain KAT. The characteristics of KAT described here are compatible with a central role of the enzyme in brain KYNA function in vivo. Abnormal KAT activity, therefore, should be considered as an etiological factor in pathological phenomena related to dysfunction of excitatory amino acid receptors in the brain.

Notes: Recent studies have revealed the effectiveness of 2-methyl-6-(phenylethynyl)pyridine (MPEP), a highly selective antagonist of metabotropic glutamate receptors subtype 5 (mGluR5), in conditioned drug reward. In a previous study we showed that MPEP blocks expression of context-conditioned morphine- but not cocaine reward in the rat. The present study now examines the effectiveness of MPEP in the expression of context-conditioned food, MDMA (‘ecstasy?) or amphetamine reward. Therefore, three groups of rats were conditioned either to food, MDMA or amphetamine in the conditioned place preference (CPP) paradigm. After conditioning, CPP expression and locomotion were determined simultaneously in the presence and absence of the respective reward (i.e. food or drug), or after application of 50?mg/kg MPEP (the dose that was most effective in reducing morphine CPP expression in our previous study). As a result, MPEP reduced locomotion in all groups. Furthermore, only expression of amphetamine CPP was inhibited by MPEP, while expression of food and MDMA CPP was not affected, suggesting that the MPEP-induced inhibition of amphetamine CPP expression was not causally linked to the reduction of locomotion. Overall, we conclude that MPEP reduces expression of context-conditioned amphetamine but not MDMA or food reward.