On the relationship between the two branches of the kynurenine pathway in the rat brain in vivo

Abstract

In the mammalian brain, kynurenine aminotransferase II (KAT II) and kynurenine 3-monooxygenase (KMO), key enzymes of the kynurenine pathway (KP) of tryptophan degradation, form the neuroactive metabolites kynurenic acid (KYNA) and 3-hydroxykynurenine (3-HK), respectively. Although physically segregated, both enzymes use the pivotal KP metabolite l-kynurenine as a substrate. We studied the functional consequences of this cellular compartmentalization in vivo using two specific tools, the KAT II inhibitor BFF 122 and the KMO inhibitor UPF 648. The acute effects of selective KAT II or KMO inhibition were studied using a radiotracing method in which the de novo synthesis of KYNA, and of 3-HK and its downstream metabolite quinolinic acid (QUIN), is monitored following an intrastriatal injection of (3)H-kynurenine. In naïve rats, intrastriatal BFF 122 decreased newly formed KYNA by 66%, without influencing 3-HK or QUIN production. Conversely, UPF 648 reduced 3-HK synthesis (by 64%) without affecting KYNA formation. Similar, selective effects of KAT II and KMO inhibition were observed when the inhibitors were applied acutely together with the excitotoxin QUIN, which impairs local KP metabolism. Somewhat different effects of KMO (but not KAT II) inhibition were obtained in rats that had received an intrastriatal QUIN injection 7 days earlier. In these neuron-depleted striata, UPF 648 not only decreased both 3-HK and QUIN production (by 77% and 66%, respectively) but also moderately raised KYNA synthesis (by 27%). These results indicate a remarkable functional segregation of the two pathway branches in the brain, boding well for the development of selective KAT II or KMO inhibitors for cognitive enhancement and neuroprotection, respectively.

Figure 1

The kynurenine pathway of tryptophan metabolism in mammalian cells. “X” indicates the selective inhibition of kynurenine aminotransferase II (KAT II; by BFF 122) and kynurenine 3-monooxygenase (KMO; by UPF 648), respectively.

Figure 2

Effect of BFF 122 (1 mM) on the de novo synthesis of tritiated KP metabolites in the naive rat striatum. ³H-Kynurenine was injected in the absence (CTR) or presence (BFF) of the KAT II inhibitor, and the data were analyzed as described in the text. Data are the mean + SEM (n = 6). ** p < 0.01 vs. the contralateral control (paired t-test).

Figure 3

Effect of UPF 648 (0.1 mM) on the de novo synthesis of tritiated KP metabolites in the naïve rat striatum. ³H-Kynurenine was injected in the absence (CTR) or presence (UPF) of the KMO inhibitor, and the data were analyzed as described in the text. Data are the mean + SEM (n = 6). * p < 0.05 vs. the contralateral control (paired t-test).

Figure 4

Effect of BFF 122 (1 mM) on the de novo synthesis of tritiated KP metabolites in the presence of 360 nmoles QUIN. ³H-Kynurenine and QUIN were injected in the absence (CTR) or presence (BFF) of the KAT II inhibitor, as described in the text. Data are the mean + SEM (n = 4). * p < 0.05 vs. the contralateral control (paired t-test).

Figure 5

Effect of UPF 648 (0.1 mM) on the de novo synthesis of tritiated KP metabolites in the presence of 360 nmoles QUIN. ³H-Kynurenine and QUIN were injected in the absence (CTR) or presence (UPF) of the KMO inhibitor, as described in the text. Data are the mean + SEM (n = 7). ** p < 0.01 vs. the contralateral control (paired t-test).

Figure 6

Effect of BFF 122 (1 mM; “B”) on the de novo synthesis of tritiated KP metabolites in unilaterally QUIN-lesioned animals. As detailed in the text, separate groups of animals, unilaterally lesioned with QUIN in the striatum, received bilateral infusions of either ³H-kynurenine alone (n = 11; open bars) or ³H-kynurenine and the KAT II inhibitor (n = 9; solid bars). Bargraphs illustrate between-group comparisons of the effects of BFF 122 (A) in the lesioned (“Q”) and (B) the contralateral, PBS-treated (“S”) striatum, respectively. Data are the mean + SEM. ** p < 0.01 vs. the striatum infused with ³H-kynurenine alone (unpaired t-test).

Figure 7

Effect of UPF 648 (0.1 mM; “U”) on the de novo synthesis of tritiated KP metabolites in unilaterally QUIN-lesioned animals. As detailed in the text, separate groups of animals, unilaterally lesioned with QUIN in the striatum, received bilateral infusions of either ³H-kynurenine alone (n = 11; open bars) or ³H-kynurenine and the KMO inhibitor (n = 7; solid bars). Bargraphs illustrate between-group comparisons of the effects of UPF 648 in (A) the lesioned (“Q”) and (B) the contralateral, PBS-treated (“S”) striatum, respectively. Data are the mean + SEM. ** p < 0.01 and * p < 0.05 vs. the striatum infused with ³H-kynurenine alone (unpaired t-test).

Figure 8

Combined effect of BFF 122 (1 mM) and 0.1 mM UPF 648 (0.1 mM) on the de novo synthesis of tritiated KP metabolites in unilaterally QUIN-lesioned animals. As detailed in the text, separate groups of animals, unilaterally lesioned with QUIN in the striatum, received bilateral infusions of either ³H-kynurenine alone (n = 11; open bars) or ³H-kynurenine and the two enzyme inhibitors (“U+B”; n = 4; solid bars). Bargraphs illustrate between-group comparisons of the effects of BFF 122 + UPF 648 in (A) the lesioned (“Q”) and (B) the contralateral, PBS-treated (“S”) striatum, respectively. Data are the mean + SEM. ** p <0.01 and * p < 0.05 vs. the striatum infused with ³H-kynurenine alone (unpaired t-test).