Impact of Raptor and Rictor Deletion on Hippocampal Pathology Following Status Epilepticus
Abstract number :
1.454
Submission category :
1. Basic Mechanisms / 1E. Models
Year :
2023
Submission ID :
1253
Source :
www.aesnet.org
Presentation date :
12/2/2023 12:00:00 AM
Published date :
Authors :
Presenting Author: Sarah Yaser, PhD Student – University of Cincinnati (UC), Cincinnati Children’s Hospital Medical Center (CCHMC)
Christin Godale, PhD – UC Neuroscience Graduate Program, CCHMC Department of Anesthesia – University of Cincinnati (UC), Cincinnati Children’s Hospital Medical Center (CCHMC); Austin Drake, BS – UC Medical Scientist Training Program, UC Neuroscience Graduate Program, CCHMC Department of Anesthesia – University of Cincinnati (UC), Cincinnati Children’s Hospital Medical Center (CCHMC); Danielle Tapp, PhD – CCHMC Department of Anesthesia – Cincinnati Children’s Hospital Medical Center; Kimberly Kraus, PhD – UC Medical Scientist Training Program, UC Neuroscience Graduate Program, CCHMC Department of Anesthesia – University of Cincinnati (UC), Cincinnati Children’s Hospital Medical Center (CCHMC); Natasha Mayer, BS – Summer Undergraduate Research Program – University of Cincinnati; Mary Dusing, BS – CCHMC Department of Anesthesia – Cincinnati Children’s Hospital Medical Center; Candi LaSarge, PhD – UC Neuroscience Graduate Program, CCHMC Department of Anesthesia – University of Cincinnati (UC), Cincinnati Children’s Hospital Medical Center (CCHMC); Steve Danzer, PhD – UC Medical Scientist Training Program, UC Neuroscience Graduate Program, CCHMC Department of Anesthesia, UC Department of Anesthesia – University of Cincinnati, Cincinnati Children’s Hospital Medical Center
Rationale: Mechanistic target of rapamycin (mTOR) signaling is mediated through mTORC1 and mTORC2. mTORC1 signaling requires Raptor (rapamycin-sensitive companion of mTOR), while mTORC2 signaling requires Rictor (rapamycin-insensitive companion of mTOR). Both pathways are implicated in epileptogenesis. To discriminate the roles of mTORC1 and mTORC2, we selectively deleted the genes encoding Raptor or Rictor from a subset of granule cells after an epileptogenic brain injury.
Methods: Male and female Raptorfl/fl, tdTomato reporter (tdT+/-) and Rictorfl/fl, tdT+/- mice were given pilocarpine, to induce status epilepticus (SE), or saline. A week later, all mice received hippocampal injections containing a mix of AAV9.CamKII.eGFP and AAV9.CamKII.Cre to produce GFP+ wildtype (WT) granule cells and adjacent tdT+ knockout cells (KO). Brains were collected two months post-injection for analyses of granule cell soma area, spine density, mossy fiber axon sprouting and mossy fiber giant bouton (presynaptic terminal) volume.
Results: Granule cell soma area was significantly reduced for tdT+ Raptor and Rictor KO cells relative to GFP+ WT cells (two-way RM ANOVA with cell genotype and treatment [SE vs no SE] as factors; main effect of genotype, Raptor, p< 0.001; Rictor, p=0.044). Reduced granule cell mossy fiber bouton volume was observed among Raptor KO cells relative to adjacent WT cells (main effect of genotype, p=0.006). A cell intrinsic effect of Raptor and Rictor deletion was also evident in reduced spine density of KOs cells relative to WT cells (two-way RM ANOVA; main effect of genotype, Raptor, p = 0.010; Rictor, p = 0.007). Mossy fiber sprouting significantly increased in pilocarpine treated mice relative to controls as assessed by Znt3 immunostaining (two-way ANOVA; main effect of treatment, p< 0.001). To assess KO granule cell participation in mossy fiber sprouting, the relative proportions of GFP+ WT and tdT+ KO puncta were determined and normalized to the proportion of GFP+ WT and tdT+ granule cells labeled in each mouse. Raptor KO puncta were significantly underrepresented relative to WT (one-way ANOVA, p=0.018) and Rictor KO puncta (p = 0.011), revealing a cell-intrinsic effect of Raptor in mossy fiber sprouting.
Basic Mechanisms