Cultured microglial cells also expressed both mRNAs (Fig. 2F). Given the high level of expression of GM-CSFRα-mRNA in the cultured microglia, it is likely that the main source of GM-CSFRα-mRNA in the ventral
midbrain is microglia. On the other hand, DArgic neurons may be the main source of IL-3Rα-mRNA. Figure 2 Expression of receptors Inhibitors,research,lifescience,medical for GM-CSF and IL-3 in the SNpc and primary cultured microglia. (A) BLU9931 mw immunoreactivity of GM-CSFRα was localized to microglial cells (yellow arrowheads) and DArgic neurons (blue arrowheads). Microglial cells were identified … Increased expression of Bcl-xL in DArgic neurons in the SNpc of the cytokine-injected rats Both GM-CSF and IL-3 have been reported to increase the expression of antiapoptotic factors belonging to the Bcl-2 family in isolated neurons (Wen et al. 1998; Huang et al. 2007; Schabitz et al. 2008). Immunohistochemical staining with antibodies to Iba1, TH, and Bcl-xL Inhibitors,research,lifescience,medical revealed that Bcl-xL immunoreactivity was localized to capillary-like Inhibitors,research,lifescience,medical structures (yellow arrowheads) in and around the SNpc in a sham rat
(Fig. 3A). Bcl-xL-immunoreactivity was similarly localized in a saline-injected Parkinsonian rat, although the immunoreactivity was markedly suppressed (Fig. 3B). By contrast, strong Bcl-xL-immunoreactivity was localized to DArgic neurons of a cytokine-injected rat (Fig. 3C, blue arrowheads). Note that the activated morphology of microglia was found in the SNpc, only in the ipsilateral side of the 6-OHDA-treated rats. Furthermore, immunoreactivity at a similar level was observed in DArgic neurons in the contralateral SNpc of the cytokine-injected rat, Inhibitors,research,lifescience,medical where microglia display resting ramified Inhibitors,research,lifescience,medical morphology (Fig. 3D). qRT-PCR showed a significant increase of Bcl-xL-mRNA in the cytokine group (Fig. 3E), and the proapoptotic factor Bax-mRNA did not significantly change among the three groups (Fig. 3F).
In comparison with the mRNA data, Bcl-xL protein was not increased in the cytokine group compared with the sham group. However, the Bcl-xL protein was markedly decreased until in the saline group (Fig. 3G). These data suggest that 6-OHDA administration accelerates degradation of Bcl-xL protein and that the cytokine injection increased the transcription of Bcl-xL mRNA in DArgic neurons. Figure 3 Antiapoptotic factor Bcl-xL expression in the SNpc. (A–D) Representative immunohistochemical data showing expression of Bcl-xL protein in the SNpc of sham, saline, and cytokine group at 1 week after 6-OHDA administrations. Localization of Bcl-xL … Phenotypic changes of microglia in response to the cytokines It has been shown previously that primary cultured rat microglial cells change their morphology in response to GM-CSF and IL-3 (Fujita et al. 1996).