3.2 Differential responses of PrL and IL to IDO activation and inflammation induced by ICV-STZ.
In order to investigate whether the ICV-STZ induced behavioral alterations are related to the activation of IDO in the PrL and IL, we examined the levels of kynurenine metabolites, and local injection of 1-MT was used to directly explore the role of IDO in the PrL and IL in ICV-STZ rats. The cannula placements of PrL and IL are shown in Figure 3A and S1A, B. The body weights of all groups maintained the same increasing trend to the end of the experiment (Figure 3B). Besides, ICV-STZ and 1-MT treatment generally had no effect on food and water consumption, as shown in Figure S1C, D. In the PrL, there was main effect of 1-MT treatment (F(1,20)=89.37) for the levels of Trp. Regarding the levels of Kyn, significant effects of ICV-STZ (F(1,20)=47.29), 1-MT treatment (F(1,20)=15.16) and interactions between ICV-STZ and 1-MT treatment (F(1,20)=16.88) were observed. For the ratios of Kyn/Trp, significant effects of ICV-STZ (F(1,20)=29.37), 1-MT treatment (F(1,20)=64.32) and interactions between ICV-STZ and 1-MT treatment (F(1,20)=12.75) were also observed. Post hoc test showed that ICV-STZ did not cause significant alterations in the levels of Trp (Figure 3C) but induced an up-regulation of Kyn levels (Figure 3D) and Kyn/Trp ratios (Figure 3E), which are standardly used as a measure of IDO activity. Intra-PrL injection of 1-MT increased Trp levels and reversed the up-regulation of Kyn levels and Kyn/Trp ratios( Figure 3C-E) in the PrL induced by ICV-STZ. In the IL, there was main effects of ICV-STZ (F(1,20)=23.85) for the levels of Trp. Regarding the levels of Kyn, significant effects of ICV-STZ (F(1,20)=25.35), 1-MT treatment (F(1,20)=18.80) and interactions between ICV-STZ and 1-MT treatment (F(1,20)=19.38) were observed. For the ratios of Kyn/Trp, significant effects of 1-MT treatment (F(1,20)=16.91) and interactions between ICV-STZ and 1-MT treatment (F(1,20)=16.58) were also observed. Different with PrL, the post hoc test revealed that ICV-STZ increased the levels of Trp (Figure 3F), Kyn (Figure 3G) and Kyn/Trp ratios (Figure 3H) in the IL. Intra-IL injection of 1-MT has no effect on Trp levels for vehicle and ICV-STZ rats but normalized the up-regulation of Kyn levels and Kyn/Trp ratios (Figure 3F-H) in the IL induced by ICV-STZ. These results proposed that ICV-STZ induced behavior alterations are likely related to activating IDO in the PrL and IL.
Pro-inflammatory cytokines are the main factors that trigger IDO activation. Hence, we further tested the levels of cytokines to elucidate the involvement of neuroinflammatory response in the PrL and IL. Two-way ANOVA revealed significant effects of 1-MT treatment (F(1,20)=9.711) and interactions between ICV-STZ and 1-MT treatment (F(1,20)=14.92) on the levels of IL-1β in the IL. And there were also significant effects of 1-MT treatment (F(1,20)=15.21) and interactions between ICV-STZ and 1-MT treatment (F(1,20)=8.396) on the levels of IL-6 in the IL. The results showed that the levels of IL-1β (Figure 3I) and IL-6 (Figure 3J) in the PrL were not significantly altered by ICV-STZ and intra-PrL injection of 1-MT. In contrast, the neuroinflammatory response in the IL induced by ICV-STZ was characterized by a significant increase in the levels of IL-1β (Figure 3K) and IL-6 (Figure 3L). Intra-IL injection of 1-MT prevented the increase of IL-1β and IL-6 levels in the IL induced by ICV-STZ.