Abstracts

Rationale:
NG2 cells are a glial cell type abundantly tiled throughout the central nervous system. Due to their capacity to generate and regenerate oligodendrocytes, they are often referred to as oligodendrocyte precursor cells (OPCs). However, a growing body of evidence indicates much broader and more complex roles for NG2 cells in nervous system function and disease and has led to their classification as the fourth major glial cell type. NG2 cells receive synaptic inputs and also express a distinguishing array of ion channels, receptors, and an immuneproteosome, uniquely priming them to monitor and respond to changes in the neural environment. While many studies have shown a role for glial cells in the process of epileptogenesis, the role of NG2 cells remains unclear. Theiler’s murine encephalomyelitis virus (TMEV) is a neurotropic picoronavirus which initially triggers acute hippocampal seizures, and although the virus is cleared within 14 days, leads to Temporal Lobe Epilepsy (TLE) in > 50% of animals. While previous studies have identified a reactive state of microglia and astrocytes as major contributors to initiating and maintaining inflammation after TMEV infection, the role of NG2 cells has not been determined. Therefore, the goal of this study is to investigate NG2 cells in response to TMEV infection and acute seizures.
Method:
Male and female C57BL6/J mice were injected intracortically with 2x105 plaque forming units of TMEV or PBS (n=8 per group). Mice were graded twice daily for seizures on a modified Racine scale between 3 – 7 days post-injection (dpi). At 4 and 14 dpi, brains were fixed (n=4 per group at each timepoint) and stained for NG2 and Iba1, or NG2 and Ki67. Confocal z-stacks were acquired in both the ipsilateral CA1 region of the hippocampus and the overlying cortex. Total field areas stained by each cell marker and total field area of colocalized pixels between NG2 and Ki67 were measured through each section then averaged across triplicate sections.
Results:
Both NG2 cells and microglia displayed reactive morphologies in the CA1 region of TMEV-injected mice at 4 and 14 dpi as measured by significantly greater immunoreactivity of NG2 and Iba1. While increased immunoreactivity for Iba1 was also present in the cortex, there was no significant change in NG2 immunoreactivity in the cortex. Glial activation in response to viral infection may also be manifested by increased proliferation. Colocalization analysis for NG2 and Ki67 revealed a significant increase in overlap between NG2 and Ki67 in CA1 of TMEV-injected mice at both 4 and 14 dpi, but no significant difference in cortex.
Conclusion:
NG2 cells acquire a reactive phenotype and increased proliferation in response to TMEV-infection. The reactive NG2 response is tightly localized to CA1, the site of active viral infection and seizure focus. These results suggest that NG2 cells alter their function in response to viral encephalopathy. Future studies are aimed at understanding whether these changes are promoting or preventative of neuronal hyperexcitability, seizures, and inflammation in the development of epilepsy.
Funding:
:NSF Graduate Research Fellowship and NIH/NINDS R37NS065434