Abstracts

This is a Late Breaking abstract

Rationale: The cellular and molecular mechanisms that drive the generation of pro-epileptogenic aberrant adult-born granule cells (abGCs) remains elusive. Our lab reported that 2-week-old (2w) abGCs in the mouse dentate gyrus display elevated Ca2+ activity after pilocarpine (pilo) induced seizures compared to sham. Moreover, this amplified Ca2+ activity is reduced by chemogenetic DREADDs (hM4Di) mediated silencing in pilo treated mice (Lybrand et al 2021). Though Ca2+ transients have been shown to promote the migration of cerebellar GCs via voltage gated Ca2+ channels (Komuro et al 1996), we do not fully understand the mechanisms by which aberrant Ca2+ activity controls abGC maturation. We hypothesize that Ca2+ activity offers a potential mechanism for abnormal regulation of gene expression programs in abGCs during seizures. Hence, the identification and functional dissection of Ca2+ activity-mediated genes in abGCs may reveal the molecular mechanism of aberrant neurogenesis, including changes in neural progenitor proliferation, ectopic cell migration and dendritic development._x000D_
Methods: RNA-seq analysis of 2w old abGCs after pilo treatment and hM4Di silencing revealed genes downregulated with pilo treatment and upregulated with hM4Di silencing – among which the Timp3 gene was one of the most significantly differentially expressed genes. Timp3 functions as an extracellular matrix (ECM) modifying gene by inhibiting matrix metalloproteinases (MMPs), thereby regulating ECM integrity. Downregulation of Timp3 is reported to regulate migration, invasion, and metastasis of tumor cells by modifying MMPs (Garofalo et al, 2009). We hypothesized that seizure induced changes in the expression of Timp3 may play a role in controlling aberrant abGC development. To address this, we firstly confirmed in RNAscope analysis that Timp3 is expressed in abGCs which is downregulated in pilo treated mice. To determine the role of Timp3 in aberrant abGC development, we used the LXR agonist T0901317 (T09) reported to knockdown its expression._x000D_
Results: T09 treatment invivo for 2w lead to an increased neural progenitor proliferation, as well as the number of DCX+ and NeuroD1+ abGCs. We also found T09 treatment increased hilar ectopic abGCs and granule cell dispersion. These results suggest that alterations in Ca2+ activity within 2w old abGCs after pilo leads to changes in gene expression, including Timp3, which could play a role in aberrant development of abGCs. Further experiments are being performed to address whether Timp3-mediated aberrant migration is sufficient to promote spontaneous recurrent seizures._x000D_
Conclusions: Our work provides new insight in the role of ECM modifying genes in epileptogenesis. We show a new way to manipulate potential aberrant gene regulatory pathways and define the functional role of our top candidate gene Timp3 in aberrant abGC development. Moreover, our study suggests that ECM modifying small molecules could be a potential therapeutic target for pro-epileptogenic aberrant neurogenesis._x000D_
Funding: The work was supported by grants from the NIH (R01NS093992; R01NS113516; R21AG066496) and funding support from the Robert J. Kleberg, Jr. and Helen C. Kleberg Foundation, and the Semmes Foundation.