MBD5 regulates NMDA receptor expression and seizures by inhibiting Stat1 transcription.

Epilepsy is considered to result from an imbalance between excitation and inhibition of the central nervous system. Pathogenic mutations in the methyl-CpG binding domain protein 5 gene (MBD5) are known to cause epilepsy. However, the function and mechanism of MBD5 in epilepsy remain elusive. Here, we found that MBD5 was mainly localized in the pyramidal cells and granular cells of mouse hippocampus, and its expression was increased in the brain tissues of mouse models of epilepsy. Exogenous overexpression of MBD5 inhibited the transcription of the signal transducer and activator of transcription 1 gene (Stat1), resulting in increased expression of N-methyl-d-aspartate receptor (NMDAR) subunit 1 (GluN1), 2A (GluN2A) and 2B (GluN2B), leading to aggravation of the epileptic behaviour phenotype in mice. The epileptic behavioural phenotype was alleviated by overexpression of STAT1 which reduced the expression of NMDARs, and by the NMDAR antagonist memantine. These results indicate that MBD5 accumulation affects seizures through STAT1-mediated inhibition of NMDAR expression in mice. Collectively, our findings suggest that the MBD5-STAT1-NMDAR pathway may be a new pathway that regulates the epileptic behavioural phenotype and may represent a new treatment target.

Muscle and skin fibroblast TDP-43 expression, dynamic mutation analysis of NOTCH2NLC and C9orf72 in patients with FOSMN.

BACKGROUND: Facial-onset sensory and motor neuronopathy (FOSMN) syndrome is a rare clinical syndrome in which the etiopathogenesis and disease-causing genes remain unknown. In addition, clinical and molecular pathological studies have rarely been evaluated in a large case series. METHODS: In this study, we present the clinical features and electrodiagnostic findings of the largest cohort of six patients with FOSMN in East Asia to date. Immunofluorescence assessment of TAR DNA-binding protein (TDP)-43 in muscle and skin fibroblasts, detection of GGC trinucleotide repeat expansions in NOTCH2NLC gene, and GGGGCC hexanucleotide repeat expansions in the C9orf72 gene were also performed. RESULTS: All patients exhibited typical symptoms and signs of FOSMN syndrome. Almost all patients showed a delayed or absent blink reflex. Neurogenic damage was found in five patients by electromyography. Two of the five patients with muscle and skin biopsies showed TDP-43-positive inclusions in both the nucleus and cytoplasm of muscular tissue and skin fibroblasts. There were no repeat expansions in the C9orf72 or NOTCH2NLC genes in any of the six patients. CONCLUSIONS: To date, this is the largest FOSMN cohort in East Asia. TDP-43-positive cytoplasmic inclusions in muscle and skin fibroblasts may be a pathologic feature of the disease. The patient's dynamic mutation test showed no GGC trinucleotide repeat expansions in the NOTCH2NLC and GGGGCC hexanucleotide repeat expansions in the C9orf72 gene. Further studies are needed with more patients.

GJB1 mutations c.212T>G and c.311A>C induce apoptosis and inwardly rectifying potassium current changes in X-linked Charcot-Marie-Tooth type 1.

Gap junction beta 1 (GJB1) is the pathogenic gene of X-linked Charcot-Marie-Tooth type 1 (CMTX1), a rare hereditary sensorimotor neuropathy. However, different mutations of GJB1 result in heterogeneous clinical manifestations with only some mutations leading to central nervous system involvement. We previously reported two GJB1 missense mutations: one novel mutation (c.212T > G) found in a CMTX1 family that only manifested as peripheral neuropathy, and another previously reported mutation GJB1(c.311A > C) leading to involvement of the peripheral nerves and cerebral white matter. However, the mechanism by which GJB1 mutations lead to CMTX1 has not been fully characterized. Here, we generated Schwann cells and primary cultured oligodendrocytes with these two mutations, resulting in the Cx32I71S (GJB1 c.212T > G) and Cx32K104T (GJB1 c.311A > C) mutants, to analyze the pathogenic mechanism using cytology, molecular biology, and electrophysiological methods. Both mutants showed abnormal endoplasmic reticulum aggregation, especially the Cx32K104T mutant, leading to an increase in endoplasmic reticulum stress, resulting in apoptosis. Furthermore, whole-cell patch clamp experiments in oligodendrocytes revealed that the Cx32K104T mutant reduced the cell membrane potential and inwardly rectifying potassium currents, which may be a vital element for central involvement. Therefore, our results may provide a new perspective for understanding the pathogenesis of CMTX1.

ERRα regulates synaptic transmission through reactive oxygen species in hippocampal neurons.

Reactive oxygen species (ROS) play multiple roles in synaptic transmission, and estrogen-related receptor α (ERRα) is involved in regulating ROS production. The purpose of our study was to explore the underlying effect of ERRα on ROS production, neurite formation and synaptic transmission. Our results revealed that knocking down ERRα expression affected the formation of neuronal neurites and dendritic spines, which are the basic structures of synaptic transmission and play important roles in learning, memory and neuronal plasticity; moreover, the amplitude and frequency of miniature excitatory postsynaptic currents (mEPSCs) and miniature inhibitory postsynaptic currents (mIPSCs) were decreased. These abnormalities were reversed by overexpression of human ERRα. Additionally, we also found that knocking down ERRα expression increased intracellular ROS levels in neurons. ROS inhibitor PBN rescued the changes in neurite formation and synaptic transmission induced by ERRα knockdown. These results indicate a new possible cellular mechanism by which ERRα affects intracellular ROS levels, which in turn regulate neurite and dendritic spine formation and synaptic transmission.

Glucose-6-phosphate dehydrogenase alleviates epileptic seizures by repressing reactive oxygen species production to promote signal transducer and activator of transcription 1-mediated N-methyl-d-aspartic acid receptors inhibition.

The pathogenesis of epilepsy remains unclear; however, a prevailing hypothesis suggests that the primary underlying cause is an imbalance between neuronal excitability and inhibition. Glucose-6-phosphate dehydrogenase (G6PD) is a key enzyme in the pentose phosphate pathway, which is primarily involved in deoxynucleic acid synthesis and antioxidant defense mechanisms and exhibits increased expression during the chronic phase of epilepsy, predominantly colocalizing with neurons. G6PD overexpression significantly reduces the frequency and duration of spontaneous recurrent seizures. Furthermore, G6PD overexpression enhances signal transducer and activator of transcription 1 (STAT1) expression, thus influencing N-methyl-d-aspartic acid receptors expression, and subsequently affecting seizure activity. Importantly, the regulation of STAT1 by G6PD appears to be mediated primarily through reactive oxygen species signaling pathways. Collectively, our findings highlight the pivotal role of G6PD in modulating epileptogenesis, and suggest its potential as a therapeutic target for epilepsy.

Activation of TLR7-mediated autophagy increases epileptic susceptibility via reduced KIF5A-dependent GABAA receptor transport in a murine model.

The pathophysiological mechanisms underlying epileptogenesis are poorly understood but are considered to actively involve an imbalance between excitatory and inhibitory synaptic transmission. Excessive activation of autophagy, a cellular pathway that leads to the removal of proteins, is known to aggravate the disease. Toll-like receptor (TLR) 7 is an innate immune receptor that regulates autophagy in infectious and noninfectious diseases. However, the relationship between TLR7, autophagy, and synaptic transmission during epileptogenesis remains unclear. We found that TLR7 was activated in neurons in the early stage of epileptogenesis. TLR7 knockout significantly suppressed seizure susceptibility and neuronal excitability. Furthermore, activation of TLR7 induced autophagy and decreased the expression of kinesin family member 5 A (KIF5A), which influenced interactions with γ-aminobutyric acid type A receptor (GABAAR)-associated protein and GABAARß2/3, thus producing abnormal GABAAR-mediated postsynaptic transmission. Our results indicated that TLR7 is an important factor in regulating epileptogenesis, suggesting a possible therapeutic target for epilepsy.

KCTD13-mediated ubiquitination and degradation of GluN1 regulates excitatory synaptic transmission and seizure susceptibility.

Temporal lobe epilepsy (TLE) is the most common and severe form of epilepsy in adults; however, its underlying pathomechanisms remain elusive. Dysregulation of ubiquitination is increasingly recognized to contribute to the development and maintenance of epilepsy. Herein, we observed for the first time that potassium channel tetramerization domain containing 13 (KCTD13) protein, a substrate-specific adapter for cullin3-based E3 ubiquitin ligase, was markedly down-regulated in the brain tissue of patients with TLE. In a TLE mouse model, the protein expression of KCTD13 dynamically changed during epileptogenesis. Knockdown of KCTD13 in the mouse hippocampus significantly enhanced seizure susceptibility and severity, whereas overexpression of KCTD13 showed the opposite effect. Mechanistically, GluN1, an obligatory subunit of N-methyl-D-aspartic acid receptors (NMDARs), was identified as a potential substrate protein of KCTD13. Further investigation revealed that KCTD13 facilitates lysine-48-linked polyubiquitination of GluN1 and its degradation through the ubiquitin-proteasome pathway. Besides, the lysine residue 860 of GluN1 is the main ubiquitin site. Importantly, dysregulation of KCTD13 affected membrane expression of glutamate receptors and impaired glutamate synaptic transmission. Systemic administration of the NMDAR inhibitor memantine significantly rescued the epileptic phenotype aggravated by KCTD13 knockdown. In conclusion, our results demonstrated an unrecognized pathway of KCTD13-GluN1 in epilepsy, suggesting KCTD13 as a potential neuroprotective therapeutic target for epilepsy.

Beclin1 Deficiency Suppresses Epileptic Seizures.

Epilepsy is a common disease of the nervous system. Autophagy is a degradation process involved in epilepsy, and in turn, seizures can activate autophagy. Beclin1 plays a critical role in autophagy and participates in numerous physiological and pathological processes. However, the mechanism underlying the effect of Beclin1 on epilepsy remains unclear. In this study, we detected increased expression of Beclin1 in brain tissues from patients with temporal lobe epilepsy (TLE). Heterozygous disruption of beclin1 decreased susceptibility to epilepsy and suppressed seizure activity in two mouse epilepsy models. We further illustrated for the first time that heterozygous disruption of beclin1 suppresses excitatory synaptic transmission, which may be caused by a decreased dendritic spine density. These findings suggest for the first time that the regulation of Beclin1 may serve as a strategy for antiepileptic therapy. In addition, Beclin1 participates in synaptic transmission, and the development of dendritic spines may be a biological function of Beclin1 independent of its role in autophagy.