Silencing of P2X7R by RNA interference in the hippocampus can attenuate morphological and behavioral impact of pilocarpine-induced epilepsy

Abstract

Cell signaling mediated by P2X7 receptors (P2X7R) has been suggested to be involved in epileptogenesis, via modulation of intracellular calcium levels, excitotoxicity, activation of inflammatory cascades, and cell death, among other mechanisms. These processes have been described to be involved in pilocarpine-induced status epilepticus (SE) and contribute to hyperexcitability, resulting in spontaneous and recurrent seizures. Here, we aimed to investigate the role of P2X7R in epileptogenesis in vivo using RNA interference (RNAi) to inhibit the expression of this receptor. Small interfering RNA (siRNA) targeting P2X7R mRNA was injected into the lateral ventricles (icv) 6 h after SE. Four groups were studied: SalineVehicle, Saline-siRNA, Pilo-Vehicle, and Pilo-siRNA. P2X7R was quantified by western blotting and neuronal death assessed by Fluoro-Jade B histochemistry. The hippocampal volume (edema) was determined 48 h following RNAi. Behavioral parameters as latency to the appearance of spontaneous seizures and the number of seizures were determined until 60 days after the SE onset. The Saline-siRNA and Pilo-siRNA groups showed a 43 and 37% reduction, respectively, in P2X7R protein levels compared to respective vehicle groups. Neuroprotection was observed in CA1 and CA3 of the PilosiRNA group compared to Pilo-Vehicle. P2X7R silencing in pilocarpine group reversed the increase in the edema detected in the hilus, suprapyramidal dentate gyrus, CA1, and CA3; reduced mortality rate following SE; increased the time to onset of spontaneous seizure; and reduced the number of seizures, when compared to the Pilo-Vehicle group. Therefore, our data highlights the potential of P2X7R as a therapeutic target for the adjunct treatment of epilepsy.

Mesial Temporal Lobe Epilepsy (MTLE) Drug-Refractoriness Is Associated With P2X7 Receptors Overexpression in the Human Hippocampus and Temporal Neocortex and May Be Predicted by Low Circulating Levels of miR-22

“…Interestingly, P2X7R antagonists reduce the number and duration of spontaneous seizures and gliosis, which effects are maintained beyond treatment cessation (Jimenez-Pacheco et al, 2013 , 2016 ). The association between P2X7R antagonism and seizure suppression was confirmed by other research groups (Mesuret et al, 2014 ; Amhaoul et al, 2016 ; Amorim et al, 2017 ). Considering that increased expression of the P2X7R has been associated with down-modulation of the non-coding microRNA, miR-22, at the post-transcriptional level in a mouse model of status epilepticus (Jimenez-Mateos et al, 2015 ; Engel et al, 2017 ), we now set to compare the expression of P2X7R and miR-22 in epileptic brains and sera, respectively, of MTLE-HS patients.…”

Deletion of P2X7 Receptor Decreases Basal Glutathione Level by Changing Glutamate-Glutamine Cycle and Neutral Amino Acid Transporters

“…Recently, the decrease of GSH level has been shown to induce cognitive decline and neuronal death during aging and neurodegenerative diseases [53,54]. Interestingly, P2X7R is a therapeutic target in the treatment of epilepsy [12][13][14]55]. Indeed, benzoylbenzoyl-ATP (BzATP, an P2X7R agonist) increases GSH concentration in the cerebrum through penicillin-induced epileptiform activity, which is reversed by A-438079, a P2X7R antagonist [56].…”

Beyond Seizure Control: Treating Comorbidities in Epilepsy via Targeting of the P2X7 Receptor

“…Suggesting an effect on the process of epileptogenesis, two studies showed P2X7R antagonism to decrease the mean kindling score in the rat PTZ kindling model [127,134]. Similar results were found using the rat pilocarpine model, where silencing of the P2X7R via siRNA resulted in reduced mortality, increased time to first spontaneous seizure, and reduced the number of spontaneous seizures [135]. In contrast, no effects on seizurefrequency via P2X7R antagonism was observed by Hong et al using a rat model where epilepsy development was induced via lithium-pilocarpine [118].…”