Inflammatory monocytes and microglia play independent roles in inflammatory ictogenesis

Background: The pathogenic contribution of neuroinflammation to ictogenesis and epilepsy may provide a therapeutic target for reduction of seizure burden in patients that are currently underserved by traditional anti-seizure medications. The Theiler's murine encephalomyelitis virus (TMEV) model has provided important insights into the role of inflammation in ictogenesis, but questions remain regarding the relative contribution of microglia and inflammatory monocytes in this model. Methods: Female C57BL/6 mice were inoculated by intracranial injection of 2 × 10⁵, 5 × 10⁴, 1.25 × 10⁴, or 3.125 × 10³ plaque-forming units (PFU) of the Daniel’s strain of TMEV at 4–6 weeks of age. Infiltration of inflammatory monocytes, microglial activation, and cytokine production were measured at 24 h post-infection (hpi). Viral load, hippocampal injury, cognitive performance, and seizure burden were assessed at several timepoints. Results: The intensity of inflammatory infiltration and the extent of hippocampal injury induced during TMEV encephalitis scaled with the amount of infectious virus in the initial inoculum. Cognitive performance was preserved in mice inoculated with 1.25 × 10⁴ PFU TMEV relative to 2 × 10⁵ PFU TMEV, but peak viral load at 72 hpi was equivalent between the inocula. CCL2 production in the brain was attenuated by 90% and TNFα and IL6 production was absent in mice inoculated with 1.25 × 10⁴ PFU TMEV. Acute infiltration of inflammatory monocytes was attenuated by more than 80% in mice inoculated with 1.25 × 10⁴ PFU TMEV relative to 2 × 10⁵ PFU TMEV but microglial activation was equivalent between groups. Seizure burden was attenuated and the threshold to kainic acid-induced seizures was higher in mice inoculated with 1.25 × 10⁴ PFU TMEV but low-level behavioral seizures persisted and the EEG exhibited reduced but detectable abnormalities. Conclusions: The size of the inflammatory monocyte response induced by TMEV scales with the amount of infectious virus in the initial inoculum, despite the development of equivalent peak infectious viral load. In contrast, the microglial response does not scale with the inoculum, as microglial hyper-ramification and increased Iba-1 expression were evident in mice inoculated with either 1.25 × 10⁴ or 2 × 10⁵ PFU TMEV. Inoculation conditions that drive inflammatory monocyte infiltration resulted in robust behavioral seizures and EEG abnormalities, but the low inoculum condition, associated with only microglial activation, drove a more subtle seizure and EEG phenotype.