Molecular Pathways Leading to Development of Epilepsy After Brain Injury and Advanced Optical Methods for Epilepsy Study

Abstract Epilepsy is regarded as a kind of brain disease caused by the abnormal activities of neurons after brain injuries. Long term severe neuron activities, or excitotoxicity, may lead to the death of neurons, causing irreversible damage to the brain. Epilepsy is not curable and hence its treatment is widely investigated. In this dissertation, in order to understand epilepsy, three main projects are undertaken. These projects are: 1) Insulin-Like Growth Factor-1 (IGF-1) regulation in epileptogenesis; 2) effect of siRNA silencing on axon growth; and 3) optical coherence microscopy (OCM) in the study of epilepsy.The first project is to investigate IGF-1 Signaling in Posttraumatic Epileptogenesis. It is reported that concentration of Insulin-like Growth Factor -1 (IGF-1) increases in the brain tissue after head injury, and the phosphorylation in the receptor of IGF-1 causes activation of mammalian target of rapamycin (mTOR) cascade, which involves epileptogenesis, suggesting that IGF-1 is involved in the downstream regulation of neuronal behavior after traumatic brain injury (TBI). In our experiment, the role of IGF-1 in epileptogenesis in an organotypic hippocampal culture (OHC) model of posttraumatic epilepsy is investigated. Lactate production, lactate dehydrogenase (LDH), electrical activity, and phosphorylation of Akt, MAPK, and S6 proteins were measured and their corresponding inhibitors are applied to investigate epileptogenesis. Results show that short-term application of IGF-1 result in a decreased risk of epileptogenesis while chronic application leads to an increase in ictal activities. In addition, chronic IGF-1 application promotes the activation of the Akt-mTOR signaling cascade, but not the MAPK signaling cascade. The second project uses small interfering RNA (siRNA) silencing to inhibit the epileptic pathway. In neurons, actin binding proteins (ABP) and tubulin binding proteins contribute to axon projection. Expression of these proteins is directly associated with the growth of neural circuits. Since epilepsy is deemed to be caused by the formation of abnormal neural circuits, logically speaking, if these proteins are appropriately silenced, epileptogenesis should be inhibited. In this research, cofilin, as an important ABP, is proposed as a targeted protein to control the growth of axons in order to understand the relationship between axon growth and epilepsy at a neuron level. The third project is to evaluate seizure-induced neural injury using optical coherence microscopy. Since this technology can image living tissue based on an intrinsic scattering index, it can be used to investigate neural activities before and after epilepsy and monitor the development of seizure activities at a tissue level. In this project, we firstly validate neuron imaging by OCM with confocal imaging, demonstrating that OCM can be applied to the study of neuron death in the OHC model. Then, after integrating the OCM system with a perfusion system, we use this system to study neural activities and show that there are different optical signals from neurons and surrounding neuropil. The remaining parts of the thesis include other projects done in collaboration with Dr. Liu's and Dr. Haas's laboratories.