Fetal Microglia Become Activated Following Maternal Immune Challenge

Background: Prenatal maternal infection and its accompanying immune response are increasingly recognized as an environmental risk factor for neurodevelopmental disorders such as autism. In mice mid-gestation administration of a viral mimic, polyinosinic:polycytidylic acid (PolyI:C), produces aberrant behavior in the adult offspring. However, though many studies have established an association between prenatal infection and abnormalities in the adult, little work has focused on the mechanisms and events occurring in the fetal brain. We hypothesize that microglia mediate the effects of the maternal immune response in the fetal brain. At mid-gestation in the mouse, microglia are migrating into the brain and are becoming abundant throughout the parenchyma. They are strategically located alongside the cortical-organizing Cajal-Retzius cells in the marginal zone and are notable inhabitants of the ganglionic eminence and thalamus. Microglia in the adult brain can become activated in response to trauma after which they secrete inflammatory cytokines. These cytokines, if elaborated in the fetal brain, could skew brain development altering the differentiation and/or migration of neurons generated there.

Objectives: Though brain cytokines have been measured, a direct investigation of the response of fetal microglia to maternal inflammation is lacking. We will determine whether they are activated and evaluate their cytokine production in control conditions and in response to maternal immune activation.

Methods: In order to study fetal microglia, we have used mice genetically engineered to express EGFP in place of the fractalkine receptor in monocytic derivatives, including microglia (CX3CR1^tim1Litt). We have developed a protocol that employs a process of both mechanical and enzymatic digestion of fetal brain tissue followed by separation using CD11b magnetic beads. Microglia identification is confirmed by examination of fluorescent cells. With this method we have been able to isolate enriched populations of microglia directly from the fetal mouse brain. Expression of cytokines by the fetal microglia from Poly (I:C)-treated dams was compared by real-time PCR to those of saline-injected dams. Immunohistochemistry was used to examine expression of transcription factors in relation to microglial populations.

Results: We found that microglia from dams injected with 10 mg/kg Poly (I:C) on embryonic day E12.5 produced greater amounts of IL-6 mRNA whereas those from saline-injected dams produced higher levels of IL-10 mRNA. A careful time course showed peak production of these cytokines at E16 though levels remained altered through E18, the last time point measured. There were no detectable differences in the production of IFNγ, IL-1β or TNFα between the two groups. Immunohistochemistry showed changes in foxp2 expression in the ganglionic eminence.

Conclusions: Our work demonstrates that fetal microglia can be activated and suggests that their cytokine profile can be altered as a result of maternal immune activation. Therefore, microglia may play an active role in altering normal brain development resulting in the structural and behavioral effects observed in the adult.