Authors :
Presenting Author: Thach-Vu Nguyen, BS – University of Maryland School of Medicine
Sandesh Kamdi, PhD – Department of Neurosurgery – University of Maryland School of Medicine; John Page, BS – Department of Neurology – University of Maryland School of Medicine; Janice Babus, BS – Department of Neurology – University of Maryland School of Medicine; Marianna Baybis, MS – Department of Neurosurgery, Department of Neurology – University of Maryland School of Medicine; Peter Crino, MD, PhD – Department of Neurology – University of Maryland School of Medicine; Philip Iffland, PhD – Department of Neurology – University of Maryland School of Medicine; Whitney Parker, MD, PhD – Department of Neurosurgery – University of Maryland School of Medicine
Rationale:
Pretzel syndrome, or polyhydramnios, megalencephaly, and symptomatic epilepsy (PMSE) is a rare neurodevelopmental disorder resulting from a homozygous loss-of-function mutation of the homonymous gene for STRADA pseudokinase. Our previous work demonstrated that STRADA plays a critical role in neuronal migration via modulation of mechanistic target of rapamycin (mTOR) signaling, and that mTOR complex 1 (mTORC1) inhibition with rapamycin (Rap) can prevent disrupted pathfinding of neural progenitor cells in vitro and reduce seizure frequency in PMSE patients. To examine the link between disrupted cortical development and epileptic potential in PMSE, we generated a germline Strada knockout (KO) mouse line, which was shown to have ectopic neurons in the subcortical white matter at postnatal day ten (P10), recapitulating this feature of the human disease. In the present study, we investigated the seizure threshold potential and associated neuropathological changes in adult Strada KO mice.
Methods:
To model PMSE and STRADA loss as an epileptogenic mTORopathy, we tested our Strada KO mouse strain, which was found in prior work to have a high rate of perinatal lethality. We implanted surviving P50-60 Strada KO mice with electrodes and established a baseline EEG profile. Video and EEG recordings were then compared between age-matched Strada+/+, Strada+/-, and Strada-/- animals following pentylenetetrazol (PTZ, 40 mg/kg, intraperitoneal) administration. Blinded observers used a modified Racine scale to behaviorally measure the severity of induced seizures. Manual event-based EEG examination and spectral analysis were performed for seizure onset detection and total power quantification. Hippocampal sections were immunostained for the nuclear oncogene c-Fos and co-labeled with excitatory and inhibitory neuronal markers VGLUT and GAD1/67, respectively. Cortical lamination was visualized with layer markers Ctip2 and SATB2. Neurons were labeled with NeuN, and mTORC1 activity was gauged via S6 phosphorylation.Results:
While there was no statistically significant behavioral difference between the different genotypes, the power spectral density compared between the different groups suggests increased PTZ-induced epileptic activity in KO mice. Immunostained hippocampal sections of these mice show a higher proportion of active excitatory to inhibitory neurons after seizure induction, compared to control animals. Phosphorylation of S6 was enhanced in KO cortex and hippocampus, consistent with an upregulation in mTORC1 signaling and further supporting PMSE as an mTORopathy.
Conclusions:
EEG analysis suggests a reduced seizure threshold in Strada KO mice, associated with cortical and hippocampal enhanced mTORC1 activity and increased excitatory tone. Our future studies will investigate the potential of a compensatory overexpression of the isoform Stradb to rescue neuronal heterotopia in Strada-deficient mice, as well as the rescue potential of mTORC1 inhibition with Rap.
Funding:
This study is funded by the Department of Neurosurgery of the University of Maryland, Baltimore.