After 1 h of lithium-pilocarpine status epilepticus (SE), immunocytochemical labeling of NMDA receptor NR1 subunits reveals relocation of subunits from the interior to the cell surface of dentate gyrus granule cells and CA3 pyramidal cells. Simultaneously, an increase in NMDA-miniature excitatory postsynaptic currents (mEPSC) as well as an increase in NMDA receptor-mediated tonic currents is observed in hippocampal slices after SE. Mean-variance analysis of NMDA-mEPSCs estimates that the number of functional postsynaptic NMDA receptors per synapse increases 38% during SE, and antagonism by ifenprodil suggests that an increase in the surface representation of NR2B-containing NMDA receptors is responsible for the augmentation of both the phasic and tonic excitatory currents with SE. These results provide a potential mechanism for an enhancement of glutamatergic excitation that maintains SE and may contribute to excitotoxic injury during SE. Therapies that directly antagonize NMDA receptors may be a useful therapeutic strategy during refractory SE.

SUMMARY Objective Pharmacoresistance remains an unsolved therapeutic challenge in status epilepticus (SE) and in cholinergic SE induced by nerve agent intoxication. SE triggers a rapid internalization of synaptic GABAA receptors and externalization of NMDA receptors that may explain the loss of potency of standard anti-epileptic drugs (AED). We hypothesized that a drug combination aimed at correcting the consequences of receptor trafficking would reduce SE severity and its long-term consequences. Methods A severe model of SE was induced in adult Sprague-Dawley rats with a high dose of lithium and pilocarpine. The GABAA receptor agonist midazolam, the NMDA receptor antagonist ketamine and/or the AED valproate were injected 40 min after SE onset in combination or as monotherapy. Measures of SE severity were the primary outcome. Secondary outcomes were acute neuronal injury, spontaneous recurrent seizures (SRS), and Morris water maze (MWM) deficits. Results Midazolam-ketamine dual therapy was more efficient than double-dose midazolam or ketamine monotherapy or than valproate-midazolam or valproate-ketamine dual therapy in reducing several parameters of SE severity, suggesting a synergistic mechanism. In addition, midazolam-ketamine dual therapy reduced SE-induced acute neuronal injury, epileptogenesis and MWM deficits. Significance This study showed that a treatment aimed at correcting maladaptive GABAA receptor and NMDA receptor trafficking can stop SE and reduce its long-term consequences. Early Midazolam-ketamine dual therapy may be superior to monotherapy in the treatment of benzodiazepine-refractory SE.

Hypoxic necrosis of dentate gyrus neurons in primary culture required the activation of an orderly cell death program independent of protein synthesis. Early mitochondrial swelling and loss of the mitochondrial membrane potential were accompanied by release of cytochrome c and followed by caspase-9-dependent activation of caspase-3. Caspase-3 and -9 inhibitors reduced neuronal necrosis. Calcium directly induced cytochrome c release from isolated mitochondria. Hypoxic neuronal necrosis may be an active process in which the direct effect of hypoxia on mitochondria may lead to the final common pathway of caspase-3-mediated neuronal death.