Thursday, November 21, 2024
11/21/2024
Project Summary/Abstract Generalized epilepsy treatments focus on seizure suppression, rather than halting the underlying pathogenesis that is initiated by seizure-associated changes in brain plasticity. Mechanisms contributing to epileptogenesis are difficult to identify in humans due to genetic and environmental diversity, and as such, preclinical models provide advantages for elucidating epileptogenic processes. Our Fos activation pathway and lesion analyses suggest that the ventromedial nucleus of the hypothalamus (VMH) is a critical anatomical node for the propagation of seizure discharge from the forebrain seizure network to the brainstem seizure network. Our preliminary data further support this hypothesis as loss of glutamate transport into presynaptic vesicles by deletion of the vesicular glutamate transporter gene, Vglut2, solely in the VMH can block seizure propagation into the brainstem seizure network, which contains cardio-respiratory centers. Access of seizure discharge into the brainstem results in both brainstem-tonic seizures and increases the probability of Sudden Unexplained Death in Epilepsy (SUDEP). These data suggest the hypothesis that the VMH is a critical anatomical node for seizure-induced plasticity. It also indicates a VMH neuroanatomical pathway, which may be important in SUDEP, since preventing seizure discharge from propagating into brainstem cardio-respiratory centers could prevent SUDEP. Our long-term goal is to identify neuroanatomical substrates and signaling pathways critical for epileptogenesis, so that therapeutic treatments and preventative strategies can be developed for epilepsy and SUDEP in humans. The overall objectives of this proposal build upon our previous work and preliminary data leading to the following aims/objectives: 1) Elucidate the mechanisms by which the VMH is involved in the epileptogenic process for allowing ictal discharge propagation from the forebrain seizure network into the brainstem seizure network and 2) Determine the genomic landscape in the VMH using spatial transcriptomics before and during epileptogenesis. In Aim 1, we will assess VMH reorganizational process using 1) mice with VMH-targeted deletion of genes (i.e., Vglut2, trkB, Bdnf) in our repeated flurothyl model to assess seizure outcomes and 2) assess changes in afferent and efferent connectivity to the VMH via Timm’s staining (fiber sprouting) and tract tracing. For Aim 2, we will utilize the novel approach of spatial transcriptomics to perform cell-type and regional whole transcriptome analysis using a NanoString GeoMx Digital Spatial Profiler and Illumina sequencing to determine gene expression changes in different populations of VMH neurons. Expression differences will be confirmed by qRT-PCR and/or immunolabeling. Overall, our proposal will elucidate seizure response mechanisms that initiate and contribute to epileptogenesis and provide a framework of transcriptional networks contributing to epileptogenesis that could be targeted for therapeutic intervention for epilepsy and/or SUDEP.
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