Investigating the Role of Microglia in Viral Infection-Induced Epilepsy
Abstract number :
1.427
Submission category :
1. Basic Mechanisms / 1A. Epileptogenesis of acquired epilepsies
Year :
2022
Submission ID :
2233025
Source :
www.aesnet.org
Presentation date :
12/3/2022 12:00:00 PM
Published date :
Nov 22, 2022, 05:29 AM
Authors :
Lakshmini Balachandar, PhD – University of Utah; Matthew Stefanic, BS – Undergraduate Researcher, University of Utah; Glenna Wallis, PhD – University of Utah; Lauren Buxton, BS – Senior Lab Specialist, University of Utah; Karen Wilcox, PhD – Chair, Pharmacology and Toxicology, University of Utah
This is a Late Breaking abstract
Rationale: Microglia—resident immune cells in the central nervous system (CNS), play a crucial role in responding to tissue damage and infection. Microglial activation constitutes their innate immune response to pathogens, damage signals and cellular debris but chronic persistence of this activation can lead to exacerbation of the neuroinflammatory milieu. It is currently unknown how this phenomenon persists or contributes to seizure development following Theiler’s Murine Encephalomyelitis Virus (TMEV) infection. Previous work in our laboratory has demonstrated that purinergic signaling in microglia is disrupted in the hippocampus of TMEV infected mice. However, to determine if activated microglia in the cortex also exhibit changes in purinergic signaling, we exogenously applied ATP and ADP to newly activated microglia in cortical brain slices obtained from TMEV-treated and control mice.
Methods: CX3CR1CreER (020940), td-Tomato (B6.Cg-Gt(ROSA)26Sortm14(CAG-tdTomato)/J) and Lck-GCaMP6f (Rosa26-CAG-LSL-Lck-GCaMP6f; 029626) knock-in mouse lines were obtained from the Jackson Laboratory and a custom triple transgenic mouse line expressing tdTom and GCamp6f in microglia was generated. Mice were injected with 20uL of either 2.5 x 105 plaque forming units of Daniels strain of TMEV or phosphate buffered saline (PBS) intracortically, 2 mm deep in the right hemisphere of the posterior parietal cortex. We employed two-photon microscopy in acute cortical brain slices to study the functional ability of microglia to detect purinergic damage signals (ATP and ADP) and respond to them during the peak period of viral encephalitis (~5 days post-infection) in the TMEV mouse model. The GCaMP6f calcium signals in microglia, including both processes and soma, were analyzed using custom scripts using FIJI, CellProfiler, and MATLAB.
Results: While the hippocampus is the primary site of viral infection in this model, microglial activation in the cortex of these infected mice is also evident. Microglia have a diminished calcium response to exogenous application of ATP during infection in the hippocampus. Interestingly and in contrast to what is observed in hippocampus, we have observed that despite this additional activation in the cortex, the ability of microglia to respond to purinergic damage signals and engage in Ca2+ signaling pathways is comparable to our PBS controls.
Conclusions: Our results suggest that following TMEV infection, microglial response to novel damage signals varies as a function of brain region. It is particularly interesting to notice this, despite microglial activation in the cortex, as it suggests microglial roles in other potential mechanisms in the brain including homeostasis, apoptosis etc.
Funding: This work was funded by R37 NS065434 (KSW) and the Skaggs Scholars Fellowship (GJW).
Basic Mechanisms