P2X7 Receptor-Dependent microRNA Expression Profile in the Brain Following Status Epilepticus in Mice

Abstract

The ionotropic ATP-gated P2X7 receptor is an important contributor to inflammatory signaling cascades via the release of Interleukin-1β, as well as having roles in cell death, neuronal plasticity and the release of neurotransmitters. Accordingly, there is interest in targeting the P2X7 receptor for the treatment of epilepsy. However, the signaling pathways downstream of P2X7 receptor activation remain incompletely understood. Notably, recent studies showed that P2X7 receptor expression is controlled, in part, by microRNAs (miRNAs). Here, we explored P2X7 receptor-dependent microRNA expression by comparing microRNA expression profiles of wild-type (wt) and P2X7 receptor knockout mice before and after status epilepticus. Genome-wide microRNA profiling was performed using hippocampi from wt and P2X7 receptor knockout mice following status epilepticus induced by intra-amygdala kainic acid. This revealed that the genetic deletion of the P2X7 receptor results in distinct patterns of microRNA expression. Specifically, we found that in vehicle-injected control mice, the lack of the P2X7 receptor resulted in the up-regulation of 50 microRNAs and down-regulation of 35 microRNAs. Post-status epilepticus, P2X7 receptor deficiency led to the up-regulation of 44 microRNAs while 13 microRNAs were down-regulated. Moreover, there was only limited overlap among identified P2X7 receptor-dependent microRNAs between control conditions and post-status epilepticus, suggesting that the P2X7 receptor regulates the expression of different microRNAs during normal physiology and pathology. Bioinformatic analysis revealed that genes targeted by P2X7 receptor-dependent microRNAs were particularly overrepresented in pathways involved in intracellular signaling, inflammation, and cell death; processes that have been repeatedly associated with P2X7 receptor activation. Moreover, whereas genes involved in signaling pathways and inflammation were common among up- and down-regulated P2X7 receptor-dependent microRNAs during physiological and pathological conditions, genes associated with cell death seemed to be restricted to up-regulated microRNAs during both physiological conditions and post-status epilepticus. Taken together, our results demonstrate that the P2X7 receptor impacts on the expression profile of microRNAs in the brain, thereby possibly contributing to both the maintenance of normal cellular homeostasis and pathological processes.

Circulating P2X7 Receptor Signaling Components as Diagnostic Biomarkers for Temporal Lobe Epilepsy

“…Both genotypes experienced similar seizures during status epilepticus ( Figure 4 B), in line with previous studies using the same transgenic P2X7 −/− mouse model [ 34 ]. This suggests differences in cytokine profiles are not due to differences in seizure severity between genotypes.…”

“…Differences in seizure severity and epilepsy development between genotypes may also compensate changes in cytokine expression or increase differences between genotypes. However, as reported previously [34], both wt and P2X7 −/− mice experienced similar seizures during status epilepticus. Whether this is the case for epilepsy development remains to be established.…”

“…Taken together, these studies suggest the P2X7R may negatively regulate plasma levels of the cytokine KC/GRO, and that this cytokine may therefore represent a potential peripheral biomarker of P2X7R activation. Both genotypes experienced similar seizures during status epilepticus (Figure 4B), in line with previous studies using the same transgenic P2X7 −/− mouse model [34]. This suggests differences in cytokine profiles are not due to differences in seizure severity between genotypes.…”

“…Procedures were reviewed and approved by the Research Ethics Committee of the Royal College of Surgeons in Ireland (RCSI) (REC 1322) and Health Products Regulatory Authority (AE19127/P038; AE19127/P057). Mice used for our studies included 8–12-week-old male C57BL/6 OlaHsd wildtype mice, male C56BL/6N-P2rx7 tm1d(EUKOMM)wtsi P2X7 knock-out (KO, P2X7 −/− ) mice [ 34 ], and heterozygous male FVB/NJ mP2X7-EGFP BAC transgenic mice (FVB/N-Tg(RP24-114E20-P2X7/StrepHisEGFP)) which overexpress the P2X7 protein C-terminally fused to the enhanced green fluorescent protein (EGFP) expressed under the control of a BAC-derived P2rx7 promoter (P2X7R-EGFP mice) [ 21 ]. For our studies, we used Line 17 from the initial generation of P2X7R-EGFP mice.…”

Detection and Functional Evaluation of the P2X7 Receptor in hiPSC Derived Neurons and Microglia-Like Cells

“…This ATP serves as a damage-associated molecular pattern (DAMP) which is detected by microglial cells via their purinergic receptors. From amongst both ionotropic (P2X) and metabotropic (P2Y) ATP-gated purinergic receptors ( Huang et al, 2019 ), the P2X7R has attracted most attention due to its unique characteristics such as its low affinity to ATP ( Surprenant et al, 1996 ) and the suggested involvement in a broad range of neurodegenerative diseases such as AD ( Martin et al, 2019 ; Francistiová et al, 2020 ), epilepsy ( Engel et al, 2012 ; Conte et al, 2020 ; Morgan et al, 2020 ), schizophrenia ( Calovi et al, 2020 ), Huntington’s disease ( Ollà et al, 2020 ) and many others. Moreover, another property of the P2X7R is its ability to induce macropore formation upon exposure to high concentrations of ATP ( Di Virgilio et al, 2018 ).…”