Abstracts

Rationale: Dendritic spines represent critical structural and functional components of synapses that could be involved in the pathophysiology of epilepsy and its comorbidities. The mTOR pathway has been implicated in regulating dendritic spine morphology under both normal and pathological conditions. Our lab has found that kainate-induced status epilepticus in mice causes acute severe dendritic beading and spine loss within minutes. Kainate seizures also cause an acute activation of the mTOR pathway, and rapamycin can block this seizure-induced mTOR activation. In this study, we tested the hypothesis that mTOR inhibitors attenuate acute seizure-induced dendritic injury. Methods: Adult male transgenic mice expressing green fluorescent protein (GFP) in neocortical neurons were used. Four experimental groups were utilized: control, kainate, rapamycin pretreatment (30 mg/kg, i.p, 24 and 48 hr before kainate) and rapamycin posttreatment (30 mg/kg, i.p, immediately after status epilepticus). In vivo multiphoton imaging was used to monitor dendritic changes in mice after kainate induced status epilepticus. A cranial window was created over neocortex and screw electrodes were placed adjacent to the cranial window to record EEG. GFP-expressing dendrites from neocortical neurons were imaged with a multiphoton microscope. After pre-seizure images were obtained, seizures were induced in mice by i.p. injection of 20 mg/kg kainate. After 30 min cumulative duration of electrographic seizures, seizures were terminated by isoflurane anesthesia induction for subsequent post-seizure imaging at 0 hr and 4 hr. Post-hoc image analysis was performed to evaluate changes in the number of dendritic spines and the beading of dendrites. The beading was categorized as no beading, mild beading, or severe beading, based on previous criteria. Results: There was no significant spine loss and no signs of dendritic beading in control mice over a 4 hr period. Kainate induced status epilepticus caused ~60% spine loss and ~90% dendritic beading, with severe beading in most dendrites (~50%) immediately following seizure termination. Rapamycin pretreated mice only had ~15% spine loss and mild beading in ~13% of dendrites after kainate induced seizures. There was a ~70% spine loss and beading in ~90% of dendrites in rapamycin posttreated mice. Pretreatment with rapamycin had significant protective effects at 0 hr and 4 hr, but posttreatment had no significant effect. Conclusions: Pretreatment with the mTOR inhibitor, rapamycin, attenuates acute seizure induced dendritic injury, at least for the first few hours following the seizures. However, treatment with rapamycin initiated after status epilepticus has no significant neuroprotective effects against acute seizure induced dendritic injury. Treatment with mTOR inhibitors may represent a rational neuroprotective strategy for preventing seizure-related dendritic injury in high risk patients or with known epilepsy. Additional studies are needed to determine whether rapamycin pretreatment or postreatment has protective effects over a longer time course.