TRANSCRIPTOME ANALYSIS IN FOCAL EPILEPSY IDENTIFIES CHANGES IN CLOCK EXPRESSION IN EPILEPTOGENIC HUMAN TISSUE LEADING TO NEURONAL SIMPLIFICATION
Abstract number :
2.406
Submission category :
Year :
2014
Submission ID :
1868958
Source :
www.aesnet.org
Presentation date :
12/6/2014 12:00:00 AM
Published date :
Dec 4, 2014, 06:00 AM
Authors :
Judy Liu, Xiaoqin Fu, Peijun Liu, Milagros Tenga, Tammy Tsuchida, Dewi Depositario-Cabacar, Amy Kao, Tesfaye Zelleke, Chima Oluigbo, Amanda Yaun, Suresh Magge, William Gaillard, Gregorio Valdez and Molly Huntsman
Rationale: Medically refractory cases of focal epilepsy may be treated by resection of the region of epileptogenic tissue, which is often curative indicating the importance of the ""seizure focus"" in pathogenesis of epilepsy. Nevertheless, the understanding of the structural and molecular underpinnings of the epileptogenic focus is largely absent. To study how focal seizures arise, we have studied human samples with transcriptome analysis of samples obtained from surgical resection of focal epilepsy cases. Identification of gene expression changes may allow us to identify novel pathways and therapeutic targets for common causes of epilepsy. Methods: Our strategy is to combine transcriptome analysis of human samples with functional studies in mouse models to determine the significance of changes that we observe in focal epilepsy samples. To this end, we have collected human surgical samples from 1) Focal cortical dysplasia (FCD) and 2) tuberous sclerosis complex (TSC). In addition, we have control tissue from non-epileptic regions of brain that was removed in the course of resection of epileptogenic tissue. This is tissue is ordinarily discarded. From these samples we performed transcriptome profiling using microarray (MA). Of the changes identified, we chose to focus on Circadian Locomotor Output Cycles Kaput (Clock). We validated expression changes at the protein level using Western analysis and then performed functional validation in animal models. 1) We show Clock expression in human and mouse brain by immunohistochemistry. 2) Using RNA interference we decreased expression of Clock in mouse neurons and assessed the effect on neuronal morphology. 3) By using a Clock mutant mouse, we studied the functional consequences of Clock deficiency on neuronal physiology. Results: Our preliminary data using microarray demonstrate clear differences between control brain, FCD, and TSC. In addition we show a group of genes regulated in the same way in FCD and TSC. Statistical analysis of gene expression, identified significant downregulation of the transcription factor, Clock expression in both TS and FCD cases compared with non-epileptic brain. In normal brain Clock is is expressed in a laminar specific manner with highest expression in layer 5. Neurons with decreased levels of Clock have simplified dendritic morphology, with a decrease in mature spine morphologies and density. In agreement with the morphology data, electrophysiology of these neurons show decreased sEPSC amplitude. Conclusions: Based on these results we hypothesize that alterations in gene expression of Clock maybe important for pathogenesis of epilepsy. Decreased expression of Clock may affect the normal formation and stability of neuronal circuits, resulting in aberrant neuronal activity and seizures. Thus, epilepsy pathogenesis may involve similar mechanisms to those observed in the oscillatory plasticity governing circadian rhythms, perhaps explaining the propensity of seizures occurring during sleep-wake transitions or during sleep.