Degree Type

Dissertation

Date of Award

2019

Degree Name

Doctor of Philosophy

Department

Veterinary Microbiology and Preventive Medicine

Major

Biomedical Sciences

First Advisor

Thimmasettappa Thippeswamy

Abstract

Epilepsy is a neurological disorder characterized by spontaneous recurrent seizures (SRS) that affects over 65 million people worldwide, making it one of the most common neurological disorder globally. Etiology for epilepsy is unknown, however, gene mutation and exposure to neurotoxins or head trauma can cause seizures predisposing brain to epilepsy. In an acquired experimental models of temporal lobe epilepsy (TLE), epileptogenesis occurs soon after the induction of status epilepticus. We show herein, through the characterization of a mouse kainate model, the classical features of epileptogenesis such as development of SRS and epileptiform spiking patterns to understand neuronal hyper-excitability, and immunohistochemistry (IHC) on brain sections to investigate the mechanisms of neuroinflammation caused by reactive gliosis, synaptic reorganization, and neurodegeneration. These parameters served as a readout for interventional studies in the rat KA model of TLE.

An inducible nitric oxide synthase (iNOS) and Src family kinase (SFK) mutually regulate each other’s expression and promote neuroinflammation and neurodegeneration. Therefore, we tested the disease modifying properties of 1400W, an iNOS inhibitor, and Saracatinib (SAR), SFK inhibitor, in our rat model. The results conclusively demonstrated that both 1400W and SAR significantly reduced spike frequency and SRS in 4-6 month of continuous EEG studies when compared to the vehicle-treated group. IHC on brain sections and Western blot results revealed reduced gliosis, serum albumin and neurodegeneration and enhanced Kir4.1 channel levels after 1400W treatment. In addition, SAR decreased translocation of Fyn and PKCδ into the nucleus with diminished expression of TNFα, IL-1β and iNOS mRNA; serum IL-6, IL-12, TNFα, IL-1β levels; and nitro-oxidative stress markers 4-HNE, gp91phox, 3-NT and iNOS in the hippocampus suggesting the role of Fyn as an upstream mediator of neuroinflammation in epileptogenesis. IHC results further revealed a significant reduction in Fyn and PKCδ immunopositive reactive glia and neurons and neurodegeneration in the brain after SAR treatment. In the vehicle treated group, Fyn and PKCδ levels were persistently upregulated during post-SE suggesting that Fyn-PKCδ drives neuroinflammation during epileptogenesis. Collectively, these findings reveal that iNOS and SFK/Fyn are principal mediators of neuroinflammation during epileptogenesis, and are potential therapeutic targets to prevent/treat epilepsy.

Copyright Owner

Shaunik Sharma

Language

en

File Format

application/pdf

File Size

323 pages

Available for download on Wednesday, September 08, 2021

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