Circuit Hyperexcitability Research Articles

Emergent epileptiform activity in spinal sensory circuits drives ectopic bursting in afferent axons and sensory dysfunction after cord injury.

Spinal cord injury leads to hyperexcitability and dysfunction in spinal sensory processing. As hyperexcitable circuits can become epileptiform, we explored whether such activity emerges in a thoracic spinal cord injury (SCI) contusion model of neuropathic pain. Recordings from spinal sensory axons in multiple below-lesion segmental dorsal roots demonstrated that SCI facilitated the emergence of spontaneous ectopic burst spiking in afferent axons, which were correlated across multiple adjacent dorsal roots. Burst frequency correlated with behavioral mechanosensitivity. The same bursting events were recruited by afferent stimulation, and timing interactions with ongoing spontaneous bursts revealed that recruitment was limited by a prolonged post-burst refractory period. Ectopic bursting in afferent axons was driven by GABAA receptor activation, presumably by conversion of subthreshold GABAergic interneuronal presynaptic axoaxonic inhibitory actions to suprathreshold spiking. Collectively, the emergence of stereotyped bursting circuitry with hypersynchrony, sensory input activation, post-burst refractory period, and reorganization of connectivity represent defining features of an epileptiform network. Indeed, these same features were reproduced in naive animals with the convulsant 4-aminopyridine (fampridine). We conclude that spinal cord injury promotes the emergence of epileptiform activity in spinal sensory networks that promote profound corruption of sensory signaling. This includes hyperexcitability and bursting by ectopic spiking in afferent axons that propagate bidirectionally by reentrant central and peripheral projections as well as sensory circuit hypoexcitability during the burst refractory period. More broadly, the work links circuit hyperexcitability to epileptiform circuit emergence, further strengthening it as a conceptual basis to understand features of sensory dysfunction and neuropathic pain.

Entorhinal cortex vulnerability to human APP expression promotes hyperexcitability and tau pathology.

Preventative treatment for Alzheimer's Disease is of dire importance, and yet, cellular mechanisms underlying early regional vulnerability in Alzheimer's Disease remain unknown. In human patients with Alzheimer's Disease, one of the earliest observed pathophysiological correlates to cognitive decline is hyperexcitability. In mouse models, early hyperexcitability has been shown in the entorhinal cortex, the first cortical region impacted by Alzheimer's Disease. The origin of hyperexcitability in early-stage disease and why it preferentially emerges in specific regions is unclear. Using cortical-region and cell-type-specific proteomics coupled with ex vivo and in vivo electrophysiology, we uncovered differential susceptibility to human-specific amyloid precursor protein (hAPP) in a model of sporadic Alzheimer's. Unexpectedly, our findings reveal that early entorhinal hyperexcitability may result from intrinsic vulnerability of parvalbumin (PV) interneurons, rather than the suspected layer II excitatory neurons. This vulnerability of entorhinal PV interneurons is specific to hAPP, as it could not be recapitulated with increased murine APP expression. However, partial replication of the findings could be seen after introduction of a murine APP chimera containing a humanized amyloid-beta sequence. Surprisingly, neurons in the Somatosensory Cortex showed no such vulnerability to adult-onset hAPP expression. hAPP-induced hyperexcitability in entorhinal cortex could be ameliorated by enhancing PV interneuron excitability in vivo. Co-expression of human Tau with hAPP decreased circuit hyperexcitability, but at the expense of increased pathological tau species. This study suggests early disease interventions targeting non-excitatory cell types may protect regions with early vulnerability to pathological symptoms of Alzheimer's Disease and downstream cognitive decline.

Isoflurane-induced neuroinflammation and NKCC1/KCC2 dysregulation result in long-term cognitive disorder in neonatal mice

BackgroundThe inhalational anesthetic isoflurane is commonly utilized in clinical practice, particularly in the field of pediatric anesthesia. Research has demonstrated its capacity to induce neuroinflammation and long-term behavioral disorders; however, the underlying mechanism remains unclear [1]. The cation-chloride cotransporters Na+–K+–2Cl−–1 (NKCC1) and K+–2Cl−–2 (KCC2) play a pivotal role in regulating neuronal responses to gamma-aminobutyric acid (GABA) [2]. Imbalances in NKCC1/KCC2 can disrupt GABA neurotransmission, potentially leading to neural circuit hyperexcitability and reduced inhibition following neonatal exposure to anesthesia [3]. Therefore, this study postulates that anesthetics have the potential to dysregulate NKCC1 and/or KCC2 during brain development.MethodsWe administered 1.5% isoflurane anesthesia to neonatal rats for a duration of 4 h at postnatal day 7 (PND7). Anxiety levels were assessed using the open field test at PND28, while cognitive function was evaluated using the Morris water maze test between PND31 and PND34. Protein levels of NKCC1, KCC2, BDNF, and phosphorylated ERK (P-ERK) in the hippocampus were measured through Western blotting analysis. Pro-inflammatory cytokines IL-1β, IL-6, and TNF-α were quantified using ELISA.ResultsWe observed a decrease in locomotion trajectories within the central region and a significantly shorter total distance in the ISO group compared to CON pups, indicating that isoflurane induces anxiety-like behavior. In the Morris water maze (MWM) test, rats exposed to isoflurane exhibited prolonged escape latency onto the platform. Additionally, isoflurane administration resulted in reduced time spent crossing in the MWM experiment at PND34, suggesting long-term impairment of memory function. Furthermore, we found that isoflurane triggered activation of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α; downregulated KCC2/BDNF/P-ERK expression; and increased the NKCC1/KCC2 ratio in the hippocampus of PND7 rats. Bumetadine (NKCC1 specific inhibitors) reversed cognitive damage and effective disorder induced by isoflurane in neonatal rats by inhibiting TNF-α activation, normalizing IL-6 and IL-1β levels, restoring KCC2 expression levels as well as BDNF and ERK signaling pathways. Based on these findings, it can be speculated that BDNF, P-ERK, IL-1β, IL-6 and TNF - α may act downstream of the NKCC1/KCC2 pathway.ConclusionsOur findings provide evidence that isoflurane administration in neonatal rats leads to persistent cognitive deficits through dysregulation of the Cation-Chloride Cotransporters NKCC1 and KCC2, BDNF, p-ERK proteins, as well as neuroinflammatory processes.

Progressive Circuit Hyperexcitability in Mouse Neocortical Slice Cultures with Increasing Duration of Activity Silencing.

Forebrain neurons deprived of activity become hyperactive when activity is restored. Rebound activity has been linked to spontaneous seizures in vivo following prolonged activity blockade. Here, we measured the time course of rebound activity and the contributing circuit mechanisms using calcium imaging, synaptic staining, and whole-cell patch clamp in organotypic slice cultures of mouse neocortex. Calcium imaging revealed hypersynchronous activity increasing in intensity with longer periods of deprivation. While activity partially recovered 3 d after slices were released from 5 d of deprivation, they were less able to recover after 10 d of deprivation. However, even after the longer period of deprivation, activity patterns eventually returned to baseline levels. The degree of deprivation-induced rebound was age-dependent, with the greatest effects occurring when silencing began in the second week. Pharmacological blockade of NMDA receptors indicated that hypersynchronous rebound activity did not require activation of Hebbian plasticity. In single-neuron recordings, input resistance roughly doubled with a concomitant increase in intrinsic excitability. Synaptic imaging of pre- and postsynaptic proteins revealed dramatic reductions in the number of presumptive synapses with a larger effect on inhibitory than excitatory synapses. Putative excitatory synapses colocalizing PSD-95 and Bassoon declined by 39 and 56% following 5 and 10 d of deprivation, but presumptive inhibitory synapses colocalizing gephyrin and VGAT declined by 55 and 73%, respectively. The results suggest that with prolonged deprivation, a progressive reduction in synapse number is accompanied by a shift in the balance between excitation and inhibition and increased cellular excitability.

Female-specific dysfunction of sensory neocortical circuits in a mouse model of autism mediated by mGluR5 and estrogen receptor α

Little is known of the brain mechanisms that mediate sex-specific autism symptoms. Here, we demonstrate that deletion of the autism spectrum disorder (ASD)-risk gene, Pten, in neocortical pyramidal neurons (NSEPten knockout [KO]) results in robust cortical circuit hyperexcitability selectively in female mice observed as prolonged spontaneous persistent activity states. Circuit hyperexcitability in females is mediated by metabotropic glutamate receptor 5 (mGluR5) and estrogen receptor α (ERα) signaling to mitogen-activated protein kinases (Erk1/2) and de novo protein synthesis. Pten KO layer 5 neurons have a female-specific increase in mGluR5 and mGluR5-dependent protein synthesis. Furthermore, mGluR5-ERα complexes are generally elevated in female cortices, and genetic reduction of ERα rescues enhanced circuit excitability, protein synthesis, and neuron size selectively in NSEPten KO females. Female NSEPten KO mice display deficits in sensory processing and social behaviors as well as mGluR5-dependent seizures. These results reveal mechanisms by which sex and a high-confidence ASD-risk gene interact to affect brain function and behavior.

Abnormal gamma phase-amplitude coupling in the parahippocampal cortex is associated with network hyperexcitability in Alzheimer's disease.

While animal models of Alzheimer's disease (AD) have shown altered gamma oscillations (∼40 Hz) in local neural circuits, the low signal-to-noise ratio of gamma in the resting human brain precludes its quantification via conventional spectral estimates. Phase-amplitude coupling (PAC) indicating the dynamic integration between the gamma amplitude and the phase of low-frequency (4-12 Hz) oscillations is a useful alternative to capture local gamma activity. In addition, PAC is also an index of neuronal excitability as the phase of low-frequency oscillations that modulate gamma amplitude, effectively regulates the excitability of local neuronal firing. In this study, we sought to examine the local neuronal activity and excitability using gamma PAC, within brain regions vulnerable to early AD pathophysiology-entorhinal cortex and parahippocampus, in a clinical population of patients with AD and age-matched controls. Our clinical cohorts consisted of a well-characterized cohort of AD patients (n = 50; age, 60 ± 8 years) with positive AD biomarkers, and age-matched, cognitively unimpaired controls (n = 35; age, 63 ± 5.8 years). We identified the presence or the absence of epileptiform activity in AD patients (AD patients with epileptiform activity, AD-EPI+, n = 20; AD patients without epileptiform activity, AD-EPI-, n = 30) using long-term electroencephalography (LTM-EEG) and 1-hour long magnetoencephalography (MEG) with simultaneous EEG. Using the source reconstructed MEG data, we computed gamma PAC as the coupling between amplitude of the gamma frequency (30-40 Hz) with phase of the theta (4-8 Hz) and alpha (8-12 Hz) frequency oscillations, within entorhinal and parahippocampal cortices. We found that patients with AD have reduced gamma PAC in the left parahippocampal cortex, compared to age-matched controls. Furthermore, AD-EPI+ patients showed greater reductions in gamma PAC than AD-EPI- in bilateral parahippocampal cortices. In contrast, entorhinal cortices did not show gamma PAC abnormalities in patients with AD. Our findings demonstrate the spatial patterns of altered gamma oscillations indicating possible region-specific manifestations of network hyperexcitability within medial temporal lobe regions vulnerable to AD pathophysiology. Greater deficits in AD-EPI+ suggests that reduced gamma PAC is a sensitive index of network hyperexcitability in AD patients. Collectively, the current results emphasize the importance of investigating the role of neural circuit hyperexcitability in early AD pathophysiology and explore its potential as a modifiable contributor to AD pathobiology.

Evolutionary conservation of hippocampal mossy fiber synapse properties

Various specialized structural/functional properties are considered essential for contextual memory encoding by hippocampal mossy fiber (MF) synapses. Although investigated to exquisite detail in model organisms, synapses, including MFs, have undergone minimal functional interrogation in humans. To determine the translational relevance of rodent findings, we evaluated MF properties within human tissue resected to treat epilepsy. Human MFs exhibit remarkably similar hallmark features to rodents, including AMPA receptor-dominated synapses with small contributions from NMDA and kainate receptors, large dynamic range with strong frequency facilitation, NMDA receptor-independent presynaptic long-term potentiation, and strong cyclic AMP (cAMP) sensitivity of release. Array tomography confirmed the evolutionary conservation of MF ultrastructure. The astonishing congruence of rodent and human MF core features argues that the basic MF properties delineated in animal models remain critical to human MF function. Finally, a selective deficit in GABAergic inhibitory tone onto human MF postsynaptic targets suggests that unrestrained detonator excitatory drive contributes to epileptic circuit hyperexcitability.

Cell‐type specific alterations in intrinsic excitability and neurotransmission in short‐term virally induced model of Alzheimer’s Disease

AbstractBackgroundAlzheimer’s Disease (AD) is a neurodegenerative cascade resulting in neuronal dysfunction and memory loss, classically characterized by the deposition of amyloid plaques throughout the brain. Although deposition of these proteins correlates with cognitive loss in AD, these aggregates may not be causally related to initial neuronal and circuit pathology in AD. Interestingly, evidence in AD patients and mouse models has reported cortical circuit hyperexcitability prior to plaque pathology. The cell and molecular mechanisms underlying hyperexcitability in early‐stage AD remain unclear. In healthy brains, normal circuit operations are maintained through a fine‐tuned balance between excitatory (pyramidal) and inhibitory neuron activity. In several APP‐expressing mouse models, pyramidal and inhibitory neuron physiology are differentially affected and also evolve with disease severity. However, these parameters have not been explored in an adult‐onset APP model.MethodWe examined rapidly emerging physiological changes in cortical PV interneurons and neighboring pyramidal neurons in an adult‐onset AD model. We analyzed contributions of overexpressed wild‐type human amyloid precursor protein (WT hAPP) using an AAV viral approach. We fluorescently targeted cell types for whole‐cell patch‐clamp electrophysiology using a PV‐specific enhancer labeling technique or an excitatory neuron‐specific AAV. Lastly, we optogenetically stimulated PV interneurons to assess neurotransmission onto neighboring pyramidal cells.ResultWe report alterations in cortical PV interneuron and pyramidal neuron intrinsic excitability, as well as PV‐to‐pyramidal neurotransmission, after short‐term induction of hAPP virally. Interestingly, each of these two neuronal subtypes displayed unique changes suggesting different cell‐type‐specific molecular alterations.ConclusionThis breakdown of APP processing and its effects on PV interneuron and pyramidal neurons provides potential insights into early cell‐type‐specific mechanisms of the pathogenesis of Alzheimer’s Disease, thus providing specific interventions to ameliorate early symptoms and potentially slow the progression towards severe disease.

Enhancing GAT-3 in thalamic astrocytes promotes resilience to brain injury in rodents.

Inflammatory processes induced by brain injury are important for recovery; however, when uncontrolled, inflammation can be deleterious, likely explaining why most anti-inflammatory treatments have failed to improve neurological outcomes after brain injury in clinical trials. In the thalamus, chronic activation of glial cells, a proxy of inflammation, has been suggested as an indicator of increased seizure risk and cognitive deficits that develop after cortical injury. Furthermore, lesions in the thalamus, more than other brain regions, have been reported in patients with viral infections associated with neurological deficits, such as SARS-CoV-2. However, the extent to which thalamic inflammation is a driver or by-product of neurological deficits remains unknown. Here, we found that thalamic inflammation in mice was sufficient to phenocopy the cellular and circuit hyperexcitability, enhanced seizure risk, and disruptions in cortical rhythms that develop after cortical injury. In our model, down-regulation of the GABA transporter GAT-3 in thalamic astrocytes mediated this neurological dysfunction. In addition, GAT-3 was decreased in regions of thalamic reactive astrocytes in mouse models of cortical injury. Enhancing GAT-3 in thalamic astrocytes prevented seizure risk, restored cortical states, and was protective against severe chemoconvulsant-induced seizures and mortality in a mouse model of traumatic brain injury, emphasizing the potential of therapeutically targeting this pathway. Together, our results identified a potential therapeutic target for reducing negative outcomes after brain injury.

Enhanced motor cortex output and disinhibition in asymptomatic female mice with C9orf72 genetic expansion.

Information and action coding by cortical circuits relies on a balanced dialogue between excitation and inhibition. Circuit hyperexcitability is considered a potential pathophysiological mechanism in various brain disorders, but the underlying deficits, especially at early disease stages, remain largely unknown. We report that asymptomatic female mice carrying the chromosome 9 open reading frame 72 (C9orf72) repeat expansion, which represents a high-prevalence genetic abnormality for human amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) spectrum disorder, exhibit abnormal motor cortex output. The number of primary motor cortex (M1) layer 5 pyramidal neurons is reduced in asymptomatic mice, with the surviving neurons receiving a decreased inhibitory drive that results in a higher M1 output, specifically during high-speed animal locomotion. Importantly, using deep-learning algorithms revealed that speed-dependent M1 output predicts the likelihood of C9orf72 genetic expansion. Our data link early circuit abnormalities with a gene mutation in asymptomatic ALS/FTLD carriers.

Abnormal larval neuromuscular junction morphology and physiology in Drosophila prickle isoform mutants with known axonal transport defects and adult seizure behavior

Previous studies have demonstrated the striking mutational effects of the Drosophila planar cell polarity gene prickle (pk) on larval motor axon microtubule-mediated vesicular transport and on adult epileptic behavior associated with neuronal circuit hyperexcitability. Mutant alleles of the prickle-prickle (pkpk ) and prickle-spiny-legs (pksple ) isoforms (hereafter referred to as pk and sple alleles, respectively) exhibit differential phenotypes. While both pk and sple affect larval motor axon transport, only sple confers motor circuit and behavior hyperexcitability. However, mutations in the two isoforms apparently counteract to ameliorate adult motor circuit and behavioral hyperexcitability in heteroallelic pkpk/pkspl e flies. We have further investigated the consequences of altered axonal transport in the development and function of the larval neuromuscular junction (NMJ). We uncovered robust dominant phenotypes in both pk and sple alleles, including synaptic terminal overgrowth (as revealed by anti-HRP and -Dlg immunostaining) and poor vesicle release synchronicity (as indicated by synaptic bouton focal recording). However, we observed recessive alteration of synaptic transmission only in pk/pk larvae, i.e. increased excitatory junctional potential (EJP) amplitude in pk/pk but not in pk/+ or sple/sple. Interestingly, for motor terminal excitability sustained by presynaptic Ca2+ channels, both pk and sple exerted strong effects to produce prolonged depolarization. Notably, only sple acted dominantly whereas pk/+ appeared normal, but was able to suppress the sple phenotypes, i.e. pk/sple appeared normal. Our observations contrast the differential roles of the pk and sple isoforms and highlight their distinct, variable phenotypic expression in the various structural and functional aspects of the larval NMJ.

Biophysical Kv3 channel alterations dampen excitability of cortical PV interneurons and contribute to network hyperexcitability in early Alzheimer's.

In Alzheimer's disease (AD), a multitude of genetic risk factors and early biomarkers are known. Nevertheless, the causal factors responsible for initiating cognitive decline in AD remain controversial. Toxic plaques and tangles correlate with progressive neuropathology, yet disruptions in circuit activity emerge before their deposition in AD models and patients. Parvalbumin (PV) interneurons are potential candidates for dysregulating cortical excitability as they display altered action potential (AP) firing before neighboring excitatory neurons in prodromal AD. Here, we report a novel mechanism responsible for PV hypoexcitability in young adult familial AD mice. We found that biophysical modulation of Kv3 channels, but not changes in their mRNA or protein expression, were responsible for dampened excitability in young 5xFAD mice. These K+ conductances could efficiently regulate near-threshold AP firing, resulting in gamma-frequency-specific network hyperexcitability. Thus, biophysical ion channel alterations alone may reshape cortical network activity prior to changes in their expression levels. Our findings demonstrate an opportunity to design a novel class of targeted therapies to ameliorate cortical circuit hyperexcitability in early AD.

FMRP Sustains Presynaptic Function via Control of Activity-Dependent Bulk Endocytosis.

Synaptic vesicle (SV) recycling is essential for the maintenance of neurotransmission, with a number of neurodevelopmental disorders linked to defects in this process. Fragile X syndrome (FXS) results from a loss of fragile X mental retardation protein (FMRP) encoded by the FMR1 gene. Hyperexcitability of neuronal circuits is a key feature of FXS, therefore we investigated whether SV recycling was affected by the absence of FMRP during increased neuronal activity. We revealed that primary neuronal cultures from male Fmr1 knock-out (KO) rats display a specific defect in activity-dependent bulk endocytosis (ADBE). ADBE is dominant during intense neuronal activity, and this defect resulted in an inability of Fmr1 KO neurons to sustain SV recycling during trains of high-frequency stimulation. Using a molecular replacement strategy, we also revealed that a human FMRP mutant that cannot bind BK channels failed to correct ADBE dysfunction in KO neurons, however this dysfunction was corrected by BK channel agonists. Therefore, FMRP performs a key role in sustaining neurotransmitter release via selective control of ADBE, suggesting intervention via this endocytosis mode may correct the hyperexcitability observed in FXS.SIGNIFICANCE STATEMENT Loss of fragile X mental retardation protein (FMRP) results in fragile X syndrome (FXS), however whether its loss has a direct role in neurotransmitter release remains a matter of debate. We demonstrate that neurons lacking FMRP display a specific defect in a mechanism that sustains neurotransmitter release during intense neuronal firing, called activity-dependent bulk endocytosis (ADBE). This discovery provides key insights into mechanisms of brain communication that occur because of loss of FMRP function. Importantly it also reveals ADBE as a potential therapeutic target to correct the circuit hyperexcitability observed in FXS.

Hyperexcitability and Homeostasis in Fragile X Syndrome.

Fragile X Syndrome (FXS) is a leading inherited cause of autism and intellectual disability, resulting from a mutation in the FMR1 gene and subsequent loss of its protein product FMRP. Despite this simple genetic origin, FXS is a phenotypically complex disorder with a range of physical and neurocognitive disruptions. While numerous molecular and cellular pathways are affected by FMRP loss, there is growing evidence that circuit hyperexcitability may be a common convergence point that can account for many of the wide-ranging phenotypes seen in FXS. The mechanisms for hyperexcitability in FXS include alterations to excitatory synaptic function and connectivity, reduced inhibitory neuron activity, as well as changes to ion channel expression and conductance. However, understanding the impact of FMR1 mutation on circuit function is complicated by the inherent plasticity in neural circuits, which display an array of homeostatic mechanisms to maintain activity near set levels. FMRP is also an important regulator of activity-dependent plasticity in the brain, meaning that dysregulated plasticity can be both a cause and consequence of hyperexcitable networks in FXS. This makes it difficult to separate the direct effects of FMR1 mutation from the myriad and pleiotropic compensatory changes associated with it, both of which are likely to contribute to FXS pathophysiology. Here we will: (1) review evidence for hyperexcitability and homeostatic plasticity phenotypes in FXS models, focusing on similarities/differences across brain regions, cell-types, and developmental time points; (2) examine how excitability and plasticity disruptions interact with each other to ultimately contribute to circuit dysfunction in FXS; and (3) discuss how these synaptic and circuit deficits contribute to disease-relevant behavioral phenotypes like epilepsy and sensory hypersensitivity. Through this discussion of where the current field stands, we aim to introduce perspectives moving forward in FXS research.

Stepwise disassembly of GABAergic synapses during pathogenic excitotoxicity.

SUMMARYGABAergic synaptic inhibition controls neuronal firing, excitability, and synaptic plasticity to regulate neuronal circuits. Following an acute excitotoxic insult, inhibitory synapses are eliminated, reducing synaptic inhibition, elevating circuit excitability, and contributing to the pathophysiology of brain injuries. However, mechanisms that drive inhibitory synapse disassembly and elimination are undefined. We find that inhibitory synapses are disassembled in a sequential manner following excitotoxicity: GABAARs undergo rapid nanoscale rearrangement and are dispersed from the synapse along with presynaptic active zone components, followed by the gradual removal of the gephyrin scaffold, prior to complete elimination of the presynaptic terminal. GABAAR nanoscale reorganization and synaptic declustering depends on calcineurin signaling, whereas disassembly of gephyrin relies on calpain activation, and blockade of both enzymes preserves inhibitory synapses after excitotoxic insult. Thus, inhibitory synapse disassembly occurs rapidly, with nanoscale precision, in a stepwise manner and most likely represents a critical step in the progression of hyperexcitability following excitotoxicity.

Levetiracetam Treatment Normalizes Levels of Presynaptic Endocytosis Machinery and Restores Nonamyloidogenic APP Processing in App Knock-in Mice.

Toxic amyloid-beta (Aβ) peptides, produced by sequential proteolytic cleavage of the amyloid precursor protein (APP), play a key role in the initial stage of Alzheimer’s disease (AD). Increasing evidence indicates that Aβ42 induces neuronal circuit hyperexcitability in the early stages of AD pathology. As a result, researchers have investigated treatments that modulate the excitatory/inhibitory imbalance as potential AD therapies. For example, levetiracetam, an atypical antiepileptic drug used to quell hyperexcitability, has garnered recent interest in the AD field, even though its exact mechanism(s) of action remains elusive. Here, we show that in APP knock-in mouse models of amyloid pathology, chronic levetiracetam administration decreases cortical Aβ42 levels and lowers the amyloid plaque burden. In addition, using multiplexed tandem mass tag-quantitative mass spectrometry-based proteomic analysis, we determined that chronic levetiracetam administration selectively normalizes levels of presynaptic endocytic proteins. Finally, we found that levetiracetam treatment selectively lowers beta carboxyl-terminal fragment levels, while the abundance of full-length APP remains unchanged. In summary, this work reports that chronic treatment with levetiracetam serves as a useful therapeutic in AD by normalizing levels of presynaptic endocytic proteins and altering APP cleavage preference, leading to a decrease in both Aβ42 levels and the amyloid plaque burden. These novel findings provide novel evidence for the previously documented therapeutic value of levetiracetam to mitigate AD pathology.

Partial Activation of TrkB Receptors Corrects Interneuronal Calcium Channel Dysfunction and Reduces Epileptogenic Activity in Neocortex following Injury.

Decreased GABAergic inhibition due to dysfunction of inhibitory interneurons plays an important role in post-traumatic epileptogenesis. Reduced N-current Ca2+ channel function in GABAergic terminals contributes to interneuronal abnormalities and neural circuit hyperexcitability in the partial neocortical isolation (undercut, UC) model of post-traumatic epileptogenesis. Because brain-derived neurotrophic factor (BDNF) supports the development and maintenance of interneurons, we hypothesized that the activation of BDNF tropomyosin kinase B (TrkB) receptors by a small molecule, TrkB partial agonist, PTX BD4-3 (BD), would correct N channel abnormalities and enhance inhibitory synaptic transmission in UC cortex. Immunocytochemistry (ICC) and western blots were used to quantify N- and P/Q-type channels. We recorded evoked (e)IPSCs and responses to N and P/Q channel blockers to determine the effects of BD on channel function. Field potential recordings were used to determine the effects of BD on circuit hyperexcitability. Chronic BD treatment 1) upregulated N and P/Q channel immunoreactivity in GABAergic terminals; 2) increased the effects of N or P/Q channel blockade on evoked inhibitory postsynaptic currents (eIPSCs); 3) increased GABA release probability and the frequency of sIPSCs; and 4) reduced the incidence of epileptiform discharges in UC cortex. The results suggest that chronic TrkB activation is a promising approach for rescuing injury-induced calcium channel abnormalities in inhibitory terminals, thereby improving interneuronal function and suppressing circuit hyperexcitability.

Altered dendritic spine function and integration in a mouse model of fragile X syndrome

Cellular and circuit hyperexcitability are core features of fragile X syndrome and related autism spectrum disorder models. However, the cellular and synaptic bases of this hyperexcitability have proved elusive. We report in a mouse model of fragile X syndrome, glutamate uncaging onto individual dendritic spines yields stronger single-spine excitation than wild-type, with more silent spines. Furthermore, fewer spines are required to trigger an action potential with near-simultaneous uncaging at multiple spines. This is, in part, from increased dendritic gain due to increased intrinsic excitability, resulting from reduced hyperpolarization-activated currents, and increased NMDA receptor signaling. Using super-resolution microscopy we detect no change in dendritic spine morphology, indicating no structure-function relationship at this age. However, ultrastructural analysis shows a 3-fold increase in multiply-innervated spines, accounting for the increased single-spine glutamate currents. Thus, loss of FMRP causes abnormal synaptogenesis, leading to large numbers of poly-synaptic spines despite normal spine morphology, thus explaining the synaptic perturbations underlying circuit hyperexcitability.

Sevoflurane-Induced Dysregulation of Cation-Chloride Cotransporters NKCC1 and KCC2 in Neonatal Mouse Brain.

The cation-chloride cotransporters Na+-K+-2Cl--1 (NKCC1) and K+-2Cl--2 (KCC2) critically regulate neuronal responses to gamma-aminobutyric acid (GABA). NKCC1 renders GABA excitatory in immature neurons while expression of KCC2 signals GABA maturation to its inhibitory role. Imbalances in NKCC1/KCC2 alter GABA neurotransmission, which may contribute to hyperexcitability and blunted inhibition in neurocircuitry after neonatal exposure to anesthesia. Thus, we hypothesized that anesthetics may dysregulate NKCC1 and/or KCC2 in developing brain. We exposed postnatal day (PND) 7 mice to sevoflurane or carrier gases and assessed NKCC1 and KCC2 expression across three brain regions 6 h and 24 h after initial exposure. To test differences in behavior, we challenged pups receiving sevoflurane or carrier gases on PND7 with propofol on PND8 and recorded parameters of anesthesia induction and maintenance. Sevoflurane exposure increased cortical NKCC1 at 6 h (p = 0.03) and decreased cortical and hippocampal KCC2 at 24 h (p = 0.009 and p = 0.007, respectively). NKCC1/KCC2 ratio was significantly increased at both 6 h (p = 0.02) and 24 h (p = 0.03) in cortex and at 24 h (p = 0.02) in hippocampus. After propofol challenge on PND8, pups previously exposed to sevoflurane on PND7 regained righting reflex significantly faster than their non-exposed cohort (p < 0.001). Disturbing NKCC1/KCC2 balance may underlie circuit hyperexcitability and contribute to neurodevelopmental impairments we have observed in previous studies of neonatal anesthesia exposure. Human infants previously exposed to anesthesia may require higher concentrations of anesthetic drugs, potentially compounding their susceptibility for neurodevelopmental sequalae.

Reigning in Excitatory Signaling in CDKL5 Deficiency.

AMPA Receptor Dysregulation and Therapeutic Interventions in a Mouse Model of CDKL5 Deficiency Disorder Yennawar M, White RS, Jensen FE. J Neurosci. 2019;39(24):4814-4828. doi:10.1523/JNEUROSCI.2041-18.2019Pathogenic mutations in cyclin-dependent kinase-like 5 (CDKL5) result in CDKL5 deficiency disorder (CDD), a rare disease marked by early-life seizures, autistic behaviors, and intellectual disability. Although mouse models of CDD exhibit dendritic instability and alterations in synaptic scaffolding proteins, studies of glutamate receptor levels and function are limited. Here, we used a novel mouse model of CDD, the Cdkl5R59X knock-in mouse (R59X), to investigate changes in synaptic glutamate receptor subunits and functional consequences. Male mice were used for all experiments to avoid the confounding effects of X-inactivation that would be present in female heterozygous mice. We showed that adult male R59X mice recapitulated the behavioral outcomes observed in other mouse models of CDD, including social deficits and memory and learning impairments, and exhibited decreased latency to seizure upon pentylenetetrazol (PTZ) administration. Furthermore, we observed a specific increase in GluA2-lacking AMPA receptors (AMPARs) in the adultR59X hippocampus, which is accompanied electrophysiologically by increased rectification ratio of AMPAR excitatory postsynaptic currents and elevated early-phase long-term potentiation (LTP). Finally, we showed that acute treatment with the GluA2-lacking AMPAR blocker IEM-1460 decreased AMPAR currents and rescued social deficits, working memory impairments, and seizure behavior latency in R59X mice. Altered NMDAR Signaling Underlies Autistic-Like Features in Mouse Models of CDKL5 Deficiency Disorder Tang S, Terzic B, Wang IJ, et al. Nat Commun. 2019;10(1):2655.Cyclin-dependent kinase-like 5 (CDKL5) deficiency disorder (CDD) is characterized by epilepsy, intellectual disability, and autistic features, and CDKL5-deficient mice exhibit a constellation of behavioral phenotypes reminiscent of the human disorder. We previously found that CDKL5 dysfunction in forebrain glutamatergic neurons results in deficits in learning and memory. However, the pathogenic origin of the autistic features of CDD remains unknown. Here, we find that selective loss of CDKL5 in GABAergic neurons leads to autistic-like phenotypes in mice accompanied by excessive glutamatergic transmission, hyperexcitability, and increased levels of postsynaptic N-methyl-D-aspartate (NMDA) receptors. Acute, low-dose inhibition of NMDAR signaling ameliorates autistic-like behaviors in GABAergic knockout mice, as well as a novel mouse model bearing a CDD-associated nonsense mutation, CDKL5 R59X, implicating the translational potential of this mechanism. Together, our findings suggest that enhanced NMDAR signaling and circuit hyperexcitability underlie autistic-like features in mouse models of CDD and provide a new therapeutic avenue to treat CDD-related symptoms.