Increased Network Excitability Research Articles

Valproate treatment induces age- and sex-dependent neuronal activity changes according to a patch clamp study.

Autism spectrum disorder is a heterogeneous neurodevelopmental disorder characterized by impaired social interactions, restricted, and stereotyped behaviors. The valproic acid model is one of the most recognized and broadly used models in rats to induce core symptoms of this disorder. Comorbidity of epilepsy and autism occurs frequently, due to similar background mechanisms that include the imbalance of excitation and inhibition. In this series of experiments, treatment was performed on rat dams with a single 500mg/kg dose i.p. valproate injection on embryonic day 12.5. Intracellular whole-cell patch clamp recordings were performed on brain slices prepared from adolescent and adult offspring of both sexes on pyramidal neurons of the medial prefrontal cortex and entorhinal cortex. Current clamp stimulation utilizing conventional current step protocols and dynamic clamp stimulation were applied to assess neuronal excitability. Membrane properties and spiking characteristics of layer II-III pyramidal cells were analyzed in both cortical regions. Significant sex-dependent and age-dependent differences were found in several parameters in the control groups. Considering membrane resistance, rheobase, voltage sag slope, and afterdepolarization slope, we observed notable changes mainly in the female groups. Valproate treatment seemed to enhance these differences and increase network excitability. However, it is possible that compensatory mechanisms took place during the maturation of the network while reaching the age-group of 3 months. Based on the results, the expression of the hyperpolarization-activated cyclic nucleotide-gated channels may be appreciably affected by the valproate treatment, which influences fundamental electrophysiological properties of the neurons such as the voltage sag. Remarkable changes appeared in the prefrontal cortex; however, also the entorhinal cortex shows similar tendencies.

Closed-loop cervical epidural stimulation partially restores ipsilesional diaphragm EMG after acute C2 hemisection

Cervical spinal cord injury creates lasting respiratory deficits which can require mechanical ventilation long-term. We have shown that closed-loop epidural stimulation (CL-ES) elicits respiratory plasticity in the form of increased phrenic network excitability (Malone et. al., E Neuro, Vol 9, 0426–21.2021, 2022); however, the ability of this treatment to create functional benefits for breathing function per se after injury has not been demonstrated. Here, we demonstrate in C2 hemisected anesthetized rats, a 20-minute bout of CL-ES administered at current amplitudes below the motor threshold restores paralyzed hemidiaphragm activity in-phase with breathing while potentiating contralesional activity. While this acute bout of stimulation did not elicit the increased network excitability seen in our chronic model, a subset of stimulated animals continued spontaneous ipsilesional diaphragm activity for several seconds after stopping stimulation. These results support the use of CL-ES as a therapeutic to rescue breathing after high cervical spinal cord injury, with the potential to lead to lasting recovery and device independence.

A human in vitro neuronal model for studying homeostatic plasticity at the network level

Mechanisms that underlie homeostatic plasticity have been extensively investigated at single-cell levels in animal models, but are less well understood at the network level. Here, we used microelectrode arrays to characterize neuronal networks following induction of homeostatic plasticity in human induced pluripotent stem cell (hiPSC)-derived glutamatergic neurons co-cultured with rat astrocytes. Chronic suppression of neuronal activity through tetrodotoxin (TTX) elicited a time-dependent network re-arrangement. Increased expression of AMPA receptors and the elongation of axon initial segments were associated with increased network excitability following TTX treatment. Transcriptomic profiling of TTX-treated neurons revealed up-regulated genes related to extracellular matrix organization, while down-regulated genes related to cell communication; also astrocytic gene expression was found altered. Overall, our study shows that hiPSC-derived neuronal networks provide a reliable invitro platform to measure and characterize homeostatic plasticity at network and single-cell levels; this platform can be extended to investigate altered homeostatic plasticity in brain disorders.

M-current modulation of cortical slow oscillations: Network dynamics and computational modeling.

The slow oscillation is a synchronized network activity expressed by the cortical network in slow wave sleep and under anesthesia. Waking up requires a transition from this synchronized brain state to a desynchronized one. Cholinergic innervation is critical for the transition from slow-wave-sleep to wakefulness, and muscarinic action is largely exerted through the muscarinic-sensitive potassium current (M-current) block. We investigated the dynamical impact of blocking the M-current on slow oscillations, both in cortical slices and in a cortical network computational model. Blocking M-current resulted in an elongation of Up states (by four times) and in a significant firing rate increase, reflecting an increased network excitability, albeit no epileptiform discharges occurred. These effects were replicated in a biophysical cortical model, where a parametric reduction of the M-current resulted in a progressive elongation of Up states and firing rate. All neurons, and not only those modeled with M-current, increased their firing rates due to network recurrency. Further increases in excitability induced even longer Up states, approaching the microarousals described in the transition towards wakefulness. Our results bridge an ionic current with network modulation, providing a mechanistic insight into network dynamics of awakening.

Hippocampal mossy cells exhibit some of the earliest signs of increased excitability in the Tg2576 model of Alzheimer’s disease neuropathology

AbstractBackgroundAlzheimer’s disease (AD) is a neurodegenerative illness characterized by progressive accumulation of amyloid beta (Aβ) and neurofibrillary tangles, with cognitive impairment and altered neural activity. Hyperexcitability in the early stages of AD contribute to Aβ accumulation and cognitive impairment, aggravating the progression of AD. However, the hyperexcitability origin is not clear. This study aimed to test whether mossy cells (MCs), an excitatory cell of the hippocampal dentate gyrus, show increased excitability at early stages of AD and contribute to the increased network excitability generation. Indeed, alterations of MCs contribute to hyperexcitability and cognitive impairment in epilepsy. However, the role of MCs in AD has not been substantially explored.MethodsIntrinsic and synaptic properties of MCs and granule cells (GCs) from WT and Tg2576 mice at early ages (1‐2 m.o.) were characterized by whole‐cell patch‐clamp recordings. Synaptic properties included the frequency and amplitude of spontaneous excitatory postsynaptic potentials (EPSPs) and excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs). Deterioration in MCs morphology was evaluated using Nissl staining and GluR2/3 labeling by light‐ and confocal microscopy. Aβ deposition was evaluated using the McSA1 antibody.ResultsTg2576 GCs did not have any significant difference in their intrinsic properties, as we shown previously in mice ∼3 m.o. However, an enhanced excitatory and inhibitory input to GCs, depicted by augmented IPSC (7.16 vs 14.04 events/s) and NMDA‐mediated EPSC frequencies (0.81 vs 1.41 events/s) were found. Interestingly, Tg2576 MCs had an augmented EPSP frequency (5.75 vs 9.44 events/s), and their intrinsic properties showed a depolarized RMP (‐72.88 vs ‐58.36 mV), and reduced rheobase (145.56 vs 47.14 pA), AP amplitude (98.14 vs 76.66 mV), time‐to‐peak (552.75 vs 266.16 ms) and maximum rise (171.44 vs 88.68 mV/ms) and decay slopes (‐61.17 vs ‐42.38 mV/ms). The correlation between #APs and current injected showed Tg2576 MCs fired significantly more APs (SEZD = 0.34; z = 2.48). Tg2576 MCs showed robust intracellular Aβ aggregation without any significant morphological change.ConclusionsMCs changes in excitability and early accumulation of Aβ suggest that MCs could be the cause of increased excitability occurring later in GCs. In this manner, MCs could be an important contributor to AD.

Differential Modes of Action of α1- and α1γ2-Autoantibodies Derived from Patients with GABAAR Encephalitis.

Autoantibodies against central nervous system proteins are increasingly being recognized in association with neurologic disorders. Although a growing number of neural autoantibodies have been identified, a causal link between specific autoantibodies and disease symptoms remains unclear, as most studies use patient-derived CSF-containing mixtures of autoantibodies. This raises questions concerning mechanism of action and which autoantibodies truly contribute to disease progression. To address this issue, monoclonal autoantibodies were isolated from a young girl with a range of neurologic symptoms, some of which reacted with specific GABAA receptor (GABAAR) subunits, α1-subunit and α1γ2-subunit, which in this study we have characterized in detail using a combination of cellular imaging and electrophysiological techniques. These studies in neurons from wild-type mice (C57BL/6J; RRID:IMSR_JAX:000664) of mixed-sex revealed that the α1 and α1γ2 subunit-specific antibodies have differential effects on the GABAA receptor. Namely, the α1-antibody was found to directly affect GABAA receptor function on a short time scale that diminished GABA currents, leading to increased network excitability. On longer time scales those antibodies also triggered a redistribution of the GABAA receptor away from synapses. In contrast, the α1γ2-antibody had no direct effect on GABAA receptor function and could possibly mediate its effect through other actors of the immune system. Taken together, these data highlight the complexity underlying autoimmune disorders and show that antibodies can exert their effect through many mechanisms within the same disease.

Synaptic dysregulation and hyperexcitability induced by intracellular amyloid beta oligomers.

Intracellular amyloid beta oligomer (iAβo) accumulation and neuronal hyperexcitability are two crucial events at early stages of Alzheimer's disease (AD). However, to date, no mechanism linking iAβo with an increase in neuronal excitability has been reported. Here, the effects of human AD brain‐derived (h‐iAβo) and synthetic (iAβo) peptides on synaptic currents and action potential firing were investigated in hippocampal neurons. Starting from 500 pM, iAβo rapidly increased the frequency of synaptic currents and higher concentrations potentiated the AMPA receptor‐mediated current. Both effects were PKC‐dependent. Parallel recordings of synaptic currents and nitric oxide (NO)‐associated fluorescence showed that the increased frequency, related to pre‐synaptic release, was dependent on a NO‐mediated retrograde signaling. Moreover, increased synchronization in NO production was also observed in neurons neighboring those dialyzed with iAβo, indicating that iAβo can increase network excitability at a distance. Current‐clamp recordings suggested that iAβo increased neuronal excitability via AMPA‐driven synaptic activity without altering membrane intrinsic properties. These results strongly indicate that iAβo causes functional spreading of hyperexcitability through a synaptic‐driven mechanism and offers an important neuropathological significance to intracellular species in the initial stages of AD, which include brain hyperexcitability and seizures.

LGI1 downregulation increases neuronal circuit excitability.

Leucine-rich glioma-inactivated 1 (LGI1) is a secreted transsynaptic protein that interacts presynaptically with Kv1.1 potassium channels and a disintegrin and metalloprotease (ADAM) protein 23, and postsynaptically influences α-amino-3-hydroxy-5-methylisoxazole-4-propionate receptors through a direct link with the ADAM22 cell adhesion protein. Haploinsufficiency of LGI1 or autoantibodies directed against LGI1 are associated with human epilepsy, generating the hypothesis that a subacute reduction of LGI1 is sufficient to increase network excitability. We tested this hypothesis in ex vivo hippocampal slices and in neuronal cultures, by subacutely reducing LGI1 expression with shRNA. Injection of shRNA-LGI1 in the hippocampus increased dentate granule cell excitability and low-frequency facilitation of mossy fibers to CA3 pyramidal cell neurotransmission. Application of the Kv1 family blocker, α-dendrotoxin, occluded this effect, implicating the involvement of Kv1.1. This subacute reduction of LGI1 was also sufficient to increase neuronal network activity in neuronal primary culture. These results indicate that a subacute reduction in LGI1 potentiates neuronal excitability and short-term synaptic plasticity, and increases neuronal network excitability, opening new avenues for the treatment of limbic encephalitis and temporal lobe epilepsies.

An Astrocytic Influence on Impaired Tonic Inhibition in Hippocampal CA1 Pyramidal Neurons in a Mouse Model of Rett Syndrome.

Rett syndrome (RTT) is a severe neurodevelopmental disease caused by mutations in the methyl-CpG binding protein 2 (MECP2) gene. Although altered interneuron development and function are clearly demonstrated in RTT mice, a particular mode of inhibition, tonic inhibition, has not been carefully examined. We report here that tonic inhibition is significantly reduced in pyramidal neurons in the CA1 region of the hippocampus in mice where Mecp2 is deleted either in all cells or specifically in astrocytes. Since no change is detected in the level of GABA receptors, such a reduction in tonic inhibition is likely a result of decreased ambient GABA level in the extracellular space. Consistent with this explanation, we observed increased expression of a GABA transporter, GABA transporter 3 (GAT3), in the hippocampus of the Mecp2 KO mice, as well as a corresponding increase of GAT3 current in hippocampal astrocytes. These phenotypes are relevant to RTT because pharmacological blockage of GAT3 can normalize tonic inhibition and intrinsic excitability in CA1 pyramidal neurons, and rescue the phenotype of increased network excitability in acute hippocampal slices from the Mecp2 KO mice. Finally, chronic administration of a GAT3 antagonist improved a composite symptom score and extended lifespan in the Mecp2 KO mice. Only male mice were used in this study. These results not only advance our understanding of RTT etiology by defining a new neuronal phenotype and revealing how it can be influenced by astrocytic alterations, but also reveal potential targets for intervention.SIGNIFICANCE STATEMENT Our study reports a novel phenotype of reduced tonic inhibition in hippocampal CA1 pyramidal neurons in the Rett syndrome mice, reveal a potential mechanism of increased GABA transporter expression/activity in the neighboring astrocytes, describe a disease-relevant consequence in hyperexcitability, and provide preliminary evidence that targeting this phenotype may slow down disease progression in Rett syndrome mice. These results help our understanding of the disease etiology and identify a new therapeutic target for treating Rett syndrome.

Hyperexcitability and seizures in the THY-Tau22 mouse model of tauopathy

Epileptic seizures constitute a significant comorbidity of Alzheimer's disease (AD), which are recapitulated in transgenic mouse models of amyloidogenesis. Here, we sought to evaluate the potential role of tau pathology regarding seizure occurrence. To this end, we performed intra-hippocampal electroencephalogram (EEG) recordings and PTZ (pentylenetetrazol) seizure threshold tests in THY-Tau22 transgenic mice of AD-like tau pathology. We demonstrate that despite a lack of spontaneous epileptiform activity in Tau22 mice, the animals display increased PTZ-induced seizure susceptibility and mortality. The increased propensity for induced seizures in THY-Tau22 mutants correlates with astrogliosis and increased expression of adenosine kinase, consistent with increased network excitability. These data support an impact of tau pathology toward AD-associated seizures and suggest that tau pathology may contribute to seizure generation in AD independent of Aβ pathology.

A supervised machine learning approach to characterize spinal network function.

Spontaneous activity is a common feature of immature neuronal networks throughout the central nervous system and plays an important role in network development and consolidation. In postnatal rodents, spontaneous activity in the spinal cord exhibits complex, stochastic patterns that have historically proven challenging to characterize. We developed a software tool for quickly and automatically characterizing and classifying episodes of spontaneous activity generated from developing spinal networks. We recorded spontaneous activity from in vitro lumbar ventral roots of 16 neonatal [postnatal day (P)0-P3] mice. Recordings were DC coupled and detrended, and episodes were separated for analysis. Amplitude-, duration-, and frequency-related features were extracted from each episode and organized into five classes. Paired classes and features were used to train and test supervised machine learning algorithms. Multilayer perceptrons were used to classify episodes as rhythmic or multiburst. We increased network excitability with potassium chloride and tested the utility of the tool to detect changes in features and episode class. We also demonstrate usability by having a novel experimenter use the program to classify episodes collected at a later time point (P5). Supervised machine learning-based classification of episodes accounted for changes that traditional approaches cannot detect. Our tool, named SpontaneousClassification, advances the detail in which we can study not only developing spinal networks, but also spontaneous networks in other areas of the nervous system. NEW & NOTEWORTHY Spontaneous activity is important for nervous system network development and consolidation. Our software uses machine learning to automatically and quickly characterize and classify episodes of spontaneous activity in the spinal cord of newborn mice. It detected changes in network activity following KCl-enhanced excitation. Using our software to classify spontaneous activity throughout development, in pathological models, or with neuromodulation, may offer insight into the development and organization of spinal circuits.

Mechanism and consequence of abnormal calcium homeostasis in Rett syndrome astrocytes.

Astrocytes play an important role in Rett syndrome (RTT) disease progression. Although the non-cell-autonomous effect of RTT astrocytes on neurons was documented, cell-autonomous phenotypes and mechanisms within RTT astrocytes are not well understood. We report that spontaneous calcium activity is abnormal in RTT astrocytes in vitro, in situ, and in vivo. Such abnormal calcium activity is mediated by calcium overload in the endoplasmic reticulum caused by abnormal store operated calcium entry, which is in part dependent on elevated expression of TRPC4. Furthermore, the abnormal calcium activity leads to excessive activation of extrasynaptic NMDA receptors (eNMDARs) on neighboring neurons and increased network excitability in Mecp2 knockout mice. Finally, both the abnormal astrocytic calcium activity and the excessive activation of eNMDARs are caused by Mecp2 deletion in astrocytes in vivo. Our findings provide evidence that abnormal calcium homeostasis is a key cell-autonomous phenotype in RTT astrocytes, and reveal its mechanism and consequence.

Acute Changes in Electrophysiological Properties of Cortical Regular-Spiking Cells Following Seizures in a Rat Lithium–Pilocarpine Model

Profound alterations in both the synaptic and intrinsic membrane properties of neurons that increase the neuronal network excitability are found in epileptic tissue. However, there are still uncertainties regarding the kind of changes in the intrinsic membrane properties occurring during epileptogenesis. Epileptogenesis is typically triggered by the initial brain-damaging insult, and status epilepticus (SE) is one of such insults. In the present study, we explored the acute changes in the intrinsic membrane properties of pyramidal cells one day after SE in a rat lithium–pilocarpine model. Using whole-cell patch-clamp recording and the dynamic-clamp technique, we investigated the properties of regular-spiking neurons in the entorhinal cortex (EC) and the medial prefrontal cortex (PFC), two areas differentially affected by SE. We found that one day after SE: (1) the intrinsic membrane properties of EC neurons are significantly altered, while the properties of PFC neurons are mostly unchanged; (2) the input resistance and membrane time constant of regular-spiking neurons are reduced due to enhanced leak current; (3) the active membrane properties of neurons are mostly unaffected; and (4) changes in the passive membrane properties diminish the intrinsic neuronal excitability. Therefore, our results suggest that the acute changes in the intrinsic membrane properties of entorhinal neurons following pilocarpine-induced SE do not contribute to network hyperexcitability. In contrast, at the early stage of epileptogenesis, protective homeostatic plasticity of intrinsic membrane properties is observed in the EC; it reduces the neuronal excitability in response to increased network excitability.

New functions of Semaphorin 3E and its receptor PlexinD1 during developing and adult hippocampal formation

The development and maturation of cortical circuits relies on the coordinated actions of long and short range axonal guidance cues. In this regard, the class 3 semaphorins and their receptors have been seen to be involved in the development and maturation of the hippocampal connections. However, although the role of most of their family members have been described, very few data about the participation of Semaphorin 3E (Sema3E) and its receptor PlexinD1 during the development and maturation of the entorhino-hippocampal (EH) connection are available. In the present study, we focused on determining their roles both during development and in adulthood. We determined a relevant role for Sema3E/PlexinD1 in the layer-specific development of the EH connection. Indeed, mice lacking Sema3E/PlexinD1 signalling showed aberrant layering of entorhinal axons in the hippocampus during embryonic and perinatal stages. In addition, absence of Sema3E/PlexinD1 signalling results in further changes in postnatal and adult hippocampal formation, such as numerous misrouted ectopic mossy fibers. More relevantly, we describe how subgranular cells express PlexinD1 and how the absence of Sema3E induces a dysregulation of the proliferation of dentate gyrus progenitors leading to the presence of ectopic cells in the molecular layer. Lastly, Sema3E mutant mice displayed increased network excitability both in the dentate gyrus and the hippocampus proper.

Electrophysiological evidence for long-axis intrinsic diversification of the hippocampus.

The elongated structure of the hippocampus is critically involved in brain functions of profound importance. The segregation of functions along the longitudinal (septotemporal or dorsoventral) axis of the hippocampus is a slowly developed concept and currently is a widely accepted idea. The segregation of neuroanatomical connections along the hippocampal long axis can provide a basis for the interpretation of the functional segregation. However, an emerging and growing body of data strongly suggests the existence of endogenous diversification in the properties of the local neural network along the long axis of the hippocampus. In particular, recent electrophysiological research provides compelling evidence demonstrating constitutively increased network excitability in the ventral hippocampus with important implications for the endogenous initiation and propagation of physiological hippocampal oscillations yet, under favorable conditions it can also drive the local network towards hyperexcitability. In addition, important specializations in the properties of dorsal and ventral hippocampal synapses may support an optimal signal processing that contributes to the effective execution of the distinct functional roles played by the two hippocampal segments.

Increased gyrification and aberrant adult neurogenesis of the dentate gyrus in adult rats.

A remarkable example of maladaptive plasticity is the development of epilepsy after a brain insult or injury to a normal animal or human. A structure that is considered central to the development of this type of epilepsy is the dentate gyrus (DG), because it is normally a relatively inhibited structure and its quiescence is thought to reduce hippocampal seizure activity. This characteristic of the DG is also considered to be important for normal hippocampal-dependent cognitive functions. It has been suggested that the brain insults which cause epilepsy do so because they cause the DG to be more easily activated. One type of brain insult that is commonly used is induction of severe seizures (status epilepticus; SE) by systemic injection of a convulsant drug. Here we describe an alteration in the DG after this type of experimental SE that may contribute to chronic seizures that has not been described before: large folds or gyri that develop in the DG by 1month after SE. Large gyri appeared to increase network excitability because epileptiform discharges recorded in hippocampal slices after SE were longer in duration when recorded inside gyri relative to locations outside gyri. Large gyri may also increase excitability because immature adult-born neurons accumulated at the base of gyri with time after SE, and previous studies have suggested that abnormalities in adult-born DG neurons promote seizures after SE. In summary, large gyri after SE are a common finding in adult rats, show increased excitability, and are associated with the development of an abnormal spatial distribution of adult-born neurons. Together these alterations may contribute to chronic seizures and associated cognitive comorbidities after SE.

Does rapid and physiological astrocyte-neuron signalling amplify epileptic activity?

The hippocampus is a key brain region in the pathophysiology of mesial temporal lobe epilepsy. Long-term changes of its architecture and function on the network and cellular level are well documented in epilepsy. Astrocytes can control many aspects of neuronal function and their long-term alterations over weeks, months and years play an important role in epilepsy. However, a pathophysiological transformation of astrocytes does not seem to be required for astrocytes to contribute to epileptic activity. Some of the properties of physiological astrocyte-neuron communication could allow these cells to exacerbate or synchronize neuronal firing on shorter time scales of milliseconds to minutes. Therefore, these astrocyte-neuron interactions are increasingly recognized as potential contributors to epileptic activity. Fast and reciprocal communication between astrocytes and neurons is enabled by a diverse set of mechanisms that could both amplify and counteract epileptic activity. They may thus promote or cause development of epileptic activity or inhibit it. Mechanisms of astrocyte-neuron interactions that can quickly increase network excitability involve, for example, astrocyte Ca2+ and Na+ signalling, K+ buffering, gap junction coupling and metabolism. However, rapid changes of astrocyte neurotransmitter uptake and morphology may also underlie or support development of network hyperexcitability. The temporal characteristics of these interactions, their ability to synchronize neuronal activity and their net effect on network activity will determine their contribution to the emergence or maintenance of epileptic activity.

Increased Network Excitability Due to Altered Synaptic Inputs to Neocortical Layer V Intact and Axotomized Pyramidal Neurons after Mild Traumatic Brain Injury.

Mild traumatic brain injury (mTBI) can produce long lasting cognitive dysfunction. There is typically no cell death and only diffuse structural injury after mTBI. Thus, functional changes in intact neurons may contribute to symptoms. We have previously shown altered intrinsic properties of axotomized and intact neurons within 2 d after a central fluid percussion injury in mice expressing yellow fluorescent protein (YFP) that allow identification of axonal state prior to recording. Here, whole-cell patch clamp recordings were used to examine synaptic properties of YFP(+) layer V pyramidal neurons. An increased frequency of spontaneous and miniature excitatory postsynaptic currents (EPSCs) was recorded from axotomized neurons at 1 d and intact neurons at 2 d after injury, likely reflecting an increased number of afferents. This also was reflected in the increased amplitude of the EPSC evoked by local extracellular stimulation for all neurons from injured cortex and increased likelihood of producing an action potential for intact cells. Field potentials recorded in superficial layers after online deep layer stimulation contained a single negative peak in controls but multiple negative peaks in injured tissue. The amplitude of this evoked negativity was significantly larger than controls over a series of stimulus intensities at both the 1 d and 2 d survival times. Interictal-like spikes never occurred in the field potential recordings from controls but were observed in 20-80% of stimulus presentations in injured cortex. Together, these results suggest an overall increase in network excitability and the production of particularly powerful (intact) neurons that have both increased intrinsic and synaptic excitability.

MiR-134-dependent regulation of Pumilio-2 is necessary for homeostatic synaptic depression.

Neurons employ a set of homeostatic plasticity mechanisms to counterbalance altered levels of network activity. The molecular mechanisms underlying homeostatic plasticity in response to increased network excitability are still poorly understood. Here, we describe a sequential homeostatic synaptic depression mechanism in primary hippocampal neurons involving miRNA-dependent translational regulation. This mechanism consists of an initial phase of synapse elimination followed by a reinforcing phase of synaptic downscaling. The activity-regulated microRNA miR-134 is necessary for both synapse elimination and the structural rearrangements leading to synaptic downscaling. Results from miR-134 inhibition further uncover a differential requirement for GluA1/2 subunits for the functional expression of homeostatic synaptic depression. Downregulation of the miR-134 target Pumilio-2 in response to chronic activity, which selectively occurs in the synapto-dendritic compartment, is required for miR-134-mediated homeostatic synaptic depression. We further identified polo-like kinase 2 (Plk2) as a novel target of Pumilio-2 involved in the control of GluA2 surface expression. In summary, we have described a novel pathway of homeostatic plasticity that stabilizes neuronal circuits in response to increased network activity.

Enhanced in vitro CA1 network activity in a sodium channel β1(C121W) subunit model of genetic epilepsy

A NaV β1(C121W) mouse model of human genetic epilepsy has enhanced neuronal excitability and temperature sensitivity attributed to a decreased threshold for action potential firing in the axon initial segment. To investigate the network consequences of this neuronal dysfunction and to establish a genetic disease state model we developed an in vitro assay to investigate CA1 network properties and antiepileptic drug sensitivity. CA1 network oscillations were induced by tetanic stimulation and average number of spikes, interspike interval (ISI), duration, and latency were measured in slices from control and NaV β1(C121W) heterozygous mice in the presence and absence of retigabine or carbamazepine. Retigabine was also tested in a thermogenic seizure model. Oscillations were reliably induced by tetanic stimulation and were maintained after severing connections between CA3 and CA1, suggesting a local recurrent circuit. Blocking α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), γ-aminobutyric acid receptor A (GABAA ), Ih , and T-type Ca(2+) channels/receptors reduced the number of spikes. Slices from NaV β1(C121W) heterozygous mice displayed several hallmarks of increased network excitability including increases in duration of the oscillation, the number and frequency of spikes and a decrease in their onset latency. The effect of genotype on network excitability was temperature sensitive, as it was seen only at elevated temperatures. Carbamazepine and retigabine were more effective in reducing network excitability in slices from NaV β1(C121W) heterozygous mice. Retigabine appeared to be more effective in suppressing time to thermogenic seizures in NaV β1(C121W) heterozygous mice compared to wild-type (WT) controls. Hippocampal networks of the NaV β1(C121W) heterozygous mouse model of genetic epilepsy show enhanced excitability consistent with earlier single neuron studies bridging important scales of brain complexity relevant to seizure genesis. Altered pharmacosensitivity further suggests that genetic epilepsy models may be useful in the development of novel antiepileptic drugs that target disease state pathology. A PowerPoint slide summarizing this article is available for download in the Supporting Information section here.