Microglia, resident immune sentinels in the brain, are crucial in responding to tissue damage, infection, damage signals like purines (ATP/ ADP), and clearing cellular debris. It is currently unknown how microglial reactivity progresses and contributes to seizure development following Theiler’s Murine Encephalomyelitis Virus (TMEV) infection. Previously, it has been demonstrated that purinergic signaling in microglia is disrupted in the hippocampus of TMEV-infected mice. However, whether reactive cortical microglia also exhibit changes in purinergic signaling, cytokine levels, and purinergic receptors are unknown. Thus, we seek to evaluate region-based differences in microglial reactivity in the TMEV model. We employed a custom triple transgenic mouse line expressing tdTomato and GCaMP6f under a CX3CR1 Cre promoter and exogenously applied ATP/ADP to acute brain slice preparations from TMEV-infected mice and controls of either sex. Interestingly and in contrast to what is observed in hippocampus, we found that despite microglial reactivity in the cortex, microglia can respond to purinergic damage signals and engage calcium signaling pathways, comparable to PBS controls. Using a cytokine panel, we also found that pro-inflammatory cytokine levels (TNF-α, IL-1α and IFN-γ) are brain-region dependent in mice infected with TMEV. Using RNAScope-FISH, we observed increases in expression of purinergic receptors responsible for microglial motility (P2Y 12 R) and inflammation (P2X 7 R) in the cortex. Collectively our results suggest that following TMEV infection, microglial response to novel damage signals, as well as the production of proinflammatory cytokines, varies as a function of brain region. Significance statement Microglia, innate immune brain cells, respond to tissue damage, infection and have elevated calcium transients in epilepsy. Viral infection-induced seizures likely originate in the hippocampus and over time, begin to secondarily generalize to the cortex. Despite recent advances, there is a major gap in understanding the region-specific role of reactive microglia in seizure development. Using a mouse model of viral infection-induced epilepsy, we found that cortical microglia retain their ability to respond to novel purinergic damage cues in acute brain slice preparations, despite being reactive, and have enhanced purinergic receptor and cytokine expression as compared to saline controls. These findings pave the way for future investigation on the impact of regional gene expression changes in reactive microglia in seizure generation.
Balachandar et al. (Mon,) studied this question.