Epilepsy is a complex disorder affecting the central nervous system and is characterised by spontaneously recurring seizures (SRSs). Epileptic patients undergo symptomatic pharmacological treatments, however, in 30% of cases, they are ineffective, mostly in patients with temporal lobe epilepsy. Therefore, there is a need for developing novel treatment strategies. Transplantation of cells releasing γ-aminobutyric acid (GABA) could be used to counteract the imbalance between excitation and inhibition within epileptic neuronal networks. We generated GABAergic interneuron precursors from human embryonic stem cells (hESCs) and grafted them in the hippocampi of rats developing chronic SRSs after kainic acid-induced status epilepticus. Using whole-cell patch-clamp recordings, we characterised the maturation of the grafted cells into functional GABAergic interneurons in the host brain, and we confirmed the presence of functional inhibitory synaptic connections from grafted cells onto the host neurons. Moreover, optogenetic stimulation of grafted hESC-derived interneurons reduced the rate of epileptiform discharges in vitro. We also observed decreased SRS frequency and total time spent in SRSs in these animals in vivo as compared to non-grafted controls. These data represent a proof-of-concept that hESC-derived GABAergic neurons can exert a therapeutic effect on epileptic animals presumably through establishing inhibitory synapses with host neurons.

Glial cell line-derived neurotrophic factor (GDNF) has been shown to counteract seizures when overexpressed or delivered into the brain in various animal models of epileptogenesis or chronic epilepsy. The mechanisms underlying this effect have not been investigated. We here demonstrate for the first time that GDNF enhances GABAergic inhibitory drive onto mouse pyramidal neurons by modulating postsynaptic GABA_A receptors, particularly in perisomatic inhibitory synapses, by GFRα1 mediated activation of the Ret receptor pathway. Other GDNF receptors, such as NCAM or Syndecan3, are not contributing to this effect. We observed similar alterations by GDNF in human hippocampal slices resected from epilepsy patients. These data indicate that GDNF may exert its seizure-suppressant action by enhancing GABAergic inhibitory transmission in the hippocampal network, thus counteracting the increased excitability of the epileptic brain. This new knowledge can contribute to the development of novel, more precise treatment strategies based on a GDNF gene therapy approach.

Convulsive status epilepticus (SE) in immature life is often associated with lasting neurobiological changes. We provoked SE by pentylenetetrazole in postnatal day 20 rat pups and examined communication modalities between the temporal hippocampus and medial entorhinal cortex (mEC) in vitro . After a minimum of 40 days post-SE, we prepared combined temporal hippocampal - medial entorhinal cortex (mEC) slices from conditioned (SE) and naïve (N) adult rats and recorded 4-aminopyridine-induced spontaneous epileptiform interictal-like discharges (IED) simultaneously from CA3 and mEC layer V-VI. We analyzed IED frequency and high frequency oscillations (HFOs) in intact slices and after surgical separation of hippocampus from mEC, by two successive incisions (Schaffer collateral cut, Parasubiculum cut). In all slices, IED frequency was higher in CA3 vs mEC and Raster plots indicated no temporal coincidence between them either in intact or in CA1-cut slices. IED frequency was significantly higher in SE mEC, but similar in SE and N CA3, independently of connectivity state. Ripples (R) and Fast Ripples (FR) coincided with IEDs and their power differed between SE and N intact slices, both in CA3 and mEC. CA3 FR/R ratios were higher in the absence of mEC. Moreover, SE (vs N) slices showed significantly higher FR/R ratios independently of the presence of mEC. Taken together, these findings suggest lasting effects of immature SE in network dynamics governing hippocampal-entorhinal communication which may impact adult cognitive, behavioral and/or seizure threshold sequalae. HIGHLIGHTS: Early-life Status Epilepticus (SE) impacts on the adult hippocampal – entorhinal communication in the in vitro 4-AP model Post-SE CA3 output decreases in HFO power with no change in interictal discharge frequency Post-SE mEC output increases both in HFO power and interictal discharge frequency Interictal HFO dynamics in CA3-mEC change upon the connectivity state of the two areas and priorhistory of early-life SE

Epilepsy is a severe neurological disease manifested by spontaneous recurrent seizures due to abnormal hyper-synchronisation of neuronal activity. Epilepsy affects about 1% of the population and up to 40% of patients experience seizures that are resistant to currently available drugs, thus highlighting an urgent need for novel treatments. In this regard, anti-inflammatory drugs emerged as potential therapeutic candidates. In particular, specific molecules apt to resolve the neuroinflammatory response occurring in acquired epilepsies have been proven to counteract seizures in experimental models, and in humans. One candidate investigational molecule has been recently identified as the lipid mediator n-3 docosapentaenoic acid-derived protectin D1(PD1 n-3DPA ) which significantly reduced seizures, cell loss and cognitive deficit in a mouse model of acquired epilepsy. However, the mechanisms that mediate PD1 n-3DPA effect remain elusive. We here addressed whether PD1 n-3DPA has direct effects on neuronal activity independent on its anti-inflammatory action. We incubated therefore hippocampal slices with PD1 n-3DPA and investigated its effect on excitatory and inhibitory synaptic inputs to the CA1 pyramidal neurons. We demonstrate that inhibitory drive onto the perisomatic region of the pyramidal neurons is increased by PD1 n-3DPA , and this effect is mediated by pertussis toxin-sensitive G-protein coupled receptors. Our data indicate that PD1 n-3DPA acts directly on inhibitory transmission, most likely at presynaptic site of inhibitory synapses as also supported by oocyte and immunohistochemical experiments. Thus, in addition to its anti-inflammatory effects, PD1 n-3DPA anti-seizure and neuroprotective effects may be mediated by its direct action on neuronal excitability by modulating their synaptic inputs.