Functional Graphene for Peritumoral Brain Microenvironment Modulation Therapy in Glioblastoma.

Peritumoral brain invasion is the main target to cure glioblastoma. Chemoradiotherapy and targeted therapies fail to combat peritumoral relapse. Brain inaccessibility and tumor heterogeneity explain this failure, combined with overlooking the peritumor microenvironment. Reduce graphene oxide (rGO) provides a unique opportunity to modulate the local brain microenvironment. Multimodal graphene impacts are reported on glioblastoma cells in vitro but fail when translated in vivo because of low diffusion. This issue is solved by developing a new rGO formulation involving ultramixing during the functionalization with polyethyleneimine (PEI) leading to the formation of highly water-stable rGO-PEI. Wide mice brain diffusion and biocompatibility are demonstrated. Using an invasive GL261 model, an anti-invasive effect is observed. A major unexpected modification of the peritumoral area is also observed with the neutralization of gliosis. In vitro, mechanistic investigations are performed using primary astrocytes and cytokine array. The result suggests that direct contact of rGO-PEIUT neutralizes astrogliosis, decreasing several proinflammatory cytokines that would explain a bystander tumor anti-invasive effect. rGO also significantly downregulates several proinvasive/protumoral cytokines at the tumor cell level. The results open the way to a new microenvironment anti-invasive nanotherapy using a new graphene nanomaterial that is optimized for in vivo brain delivery.

Randomized clinical trials of oral vitamin D supplementation in need of a paradigm change: The vitamin D autacoid paradigm.

Epidemiological studies highlight the negative correlation between vitamin D levels and the incidence of many non-skeletal diseases including inflammatory diseases, cancer, and metabolic and neurological disorders. However, most randomized controlled trials (RCTs) with oral vitamin D supplementation give mixed results or are inconclusive. It has been said that "discovery commences with the awareness of anomaly". The "anomaly" between our preclinical and clinical data provides the opportunity to propose an alternative paradigm to the vitamin D endocrine system: the vitamin D autacoid paradigm. In the vitamin D autacoid paradigm, the extra-skeletal effects of vitamin D depend on the tissue reserves of vitamin D metabolites. These vitamin D autacoid systems are inducible oscillatory ecosystems in which 1,25D is produced, acts and is inactivated locally. In the vitamin D autacoid paradigm, attaining adequacy of vitamin D in the systemic circulation is necessary but not sufficient; we must also ensure the repletion of the tissue stores. The co-existence of two different vitamin D systems, endocrine and autacoid, with different functions and regulations leads to "significant shifts in the criteria determining the legitimacy both of problems and of proposed solutions". With respect to our clinical trials of vitamin D supplementation for unconventional effects, the proposed solution is administering and quantifying vitamin D metabolites directly to the target tissue.

Activation of GABAA receptors controls mesiotemporal lobe epilepsy despite changes in chloride transporters expression: In vivo and in silico approach.

Mesiotemporal lobe Epilepsy (MTLE), the most frequent form of focal epilepsy, is often drug-resistant. Enriching the epileptic focus with GABA-releasing engineered cells has been proposed as a strategy to prevent seizures. However, ex vivo data from animal models and MTLE patients suggest that, due to changes in chloride homeostasis, GABAA receptor activation is depolarizing and partly responsible for focal interictal discharges and seizure initiation. To understand how these two contradictory aspects of GABAergic neurotransmission coexist in MTLE, we used an established mouse model of MTLE presenting hippocampal sclerosis and recurrent hippocampal paroxysmal discharges (HPDs) 30-40days after a unilateral injection of kainate in the dorsal hippocampus. We first showed that injections of GABAA receptor agonists either systemically or directly into hippocampus suppressed HPDs. Western-blotting and immunostaining revealed that levels of α1, α3 and γ2 GABAA receptor subunits were increased in epileptic mice, compared to saline controls, while levels of R1 and R2 GABAB receptor subunits but also NR1, NR2A and NR2B NMDA receptor subunits and GluR1 and GluR2 AMPA receptor subunits were decreased. In addition, we showed that the expression of the transporter NKCC1, which load neurons with chloride, was increased, whereas KCC2, a chloride extruder, was decreased and that HPDs were suppressed by injection of blockers of NKCC1. These different changes were integrated in a numerical model, and in silico simulations supported the notion that chloride imbalance impair local inhibitory control of pyramidal neurons' activity in this model of MTLE. However, our numerical model also suggested that lasting activation of these receptors restore physiological intracellular chloride concentrations and suppress HPDs. Overall, our study suggests that activation of GABAA receptor remains an effective antiepileptic strategy to suppress focal seizures in MTLE, and demonstrates that modeling and simulation studies provide new insights about the cellular and synaptic mechanisms of this disease.

Differential Effects of Antiepileptic Drugs on Focal Seizures in the Intrahippocampal Kainate Mouse Model of Mesial Temporal Lobe Epilepsy.

AIMS: Mesial temporal lobe epilepsy (MTLE) is the most common form of drug-refractory epilepsy. Most of the morphological and electrophysiological features of human MTLE can be reproduced in a mouse by a unilateral intrahippocampal injection of kainate (MTLE mouse model). The effects of antiepileptic drugs (AEDs) on the occurrence of recurrent focal hippocampal seizures in this model remain to be specified. Here, we addressed the pharmacological reactivity of this model to the most commonly used AEDs. METHODS: Using depth electroencephalographical (EEG) recordings, we tested the dose-response effects of acute injection of nine AEDs on the occurrence of hippocampal paroxysmal discharges (HPDs) as well as on ictal and interictal power spectra in the MTLE mouse model. RESULTS: Valproate, carbamazepine, and lamotrigine dose dependently suppressed HPDs and modified the general behavior and/or EEG activity. Levetiracetam and pregabalin suppressed HPDs at high doses but without any behavioral nor interictal EEG changes. Finally, phenobarbital, tiagabine, vigabatrin, and diazepam suppressed HPDs in a dose-dependent manner at doses devoid of obvious behavioral effects. CONCLUSION: The MTLE mouse model displays a differential sensitivity to AEDs with a greater efficacy of drug that facilitates GABAergic transmission. This model provides an efficient tool to identify new treatment for drug-resistant forms of focal epilepsies.

Long-term modifications of epileptogenesis and hippocampal rhythms after prolonged hyperthermic seizures in the mouse.

Complex febrile seizures are often reported in the history of patients with mesio-temporal lobe epilepsy (MTLE) but their role in its physiopathology remains controversial. We postulated that prolonged hyperthermic seizures might, as a "single-hit", modify the hippocampal rhythms, facilitate epileptogenesis and influence subsequent epilepsy when a second-hit already exists or subsequently occurs. To test this hypothesis, we examined the effects of hyperthermic seizures (30min at 40-41°C) at postnatal day 10 on hippocampal activity in C57BL/6J mice in comparison to their littermates in sham conditions (22°C), with or without another insult. Using local field potential, we observed an asymmetry in the hippocampal susceptibility to seize in hyperthermic conditions. When these mice were adult, an asymmetrical increase of low frequency power was also recorded in the hippocampus when compared to sham animals. Using two different "two-hit" protocols, no increase in seizures or hippocampal discharge frequency or duration was observed, either in mice with a genetic CA3 dysplasia (Dcx knockout), or in mice injected with kainate into the dorsal hippocampus at P60. However, in the latter condition, which is reminiscent of MTLE, the hyperthermic seizures accelerated epileptogenesis and decreased the power in the high frequency gamma band, as well as decreasing the coherence between hippocampi and the involvement of the contralateral hippocampus during hippocampal paroxysmal discharges. Our data suggest that a single episode of prolonged hyperthermic seizures does not induce per se, but accelerates epileptogenesis and could lead to an asymmetrical dysfunction in the hippocampal rhythmicity in both physiological and pathological conditions.

Synchrotron X-ray interlaced microbeams suppress paroxysmal oscillations in neuronal networks initiating generalized epilepsy.

Radiotherapy has shown some efficacy for epilepsies but the insufficient confinement of the radiation dose to the pathological target reduces its indications. Synchrotron-generated X-rays overcome this limitation and allow the delivery of focalized radiation doses to discrete brain volumes via interlaced arrays of microbeams (IntMRT). Here, we used IntMRT to target brain structures involved in seizure generation in a rat model of absence epilepsy (GAERS). We addressed the issue of whether and how synchrotron radiotherapeutic treatment suppresses epileptic activities in neuronal networks. IntMRT was used to target the somatosensory cortex (S1Cx), a region involved in seizure generation in the GAERS. The antiepileptic mechanisms were investigated by recording multisite local-field potentials and the intracellular activity of irradiated S1Cx pyramidal neurons in vivo. MRI and histopathological images displayed precise and sharp dose deposition and revealed no impairment of surrounding tissues. Local-field potentials from behaving animals demonstrated a quasi-total abolition of epileptiform activities within the target. The irradiated S1Cx was unable to initiate seizures, whereas neighboring non-irradiated cortical and thalamic regions could still produce pathological oscillations. In vivo intracellular recordings showed that irradiated pyramidal neurons were strongly hyperpolarized and displayed a decreased excitability and a reduction of spontaneous synaptic activities. These functional alterations explain the suppression of large-scale synchronization within irradiated cortical networks. Our work provides the first post-irradiation electrophysiological recordings of individual neurons. Altogether, our data are a critical step towards understanding how X-ray radiation impacts neuronal physiology and epileptogenic processes.