PAK1 c.1409 T > a (p. Leu470Gln) de novo variant affects the protein kinase domain, leading to epilepsy, macrocephaly, spastic quadriplegia, and hydrocephalus: Case report and review of the literature.

The p-21-activated kinase 1 (PAK1) protein, encoded by the PAK1 gene, is an evolutionarily conserved serine/threonine-protein kinase that regulates key cellular developmental processes. To date, seven de novo PAK1 variants have been reported to cause the Intellectual Developmental Disorder with Macrocephaly, Seizures, and Speech Delay (IDDMSSD). In addition to the namesake features, other common characteristics include structural brain anomalies, delayed development, hypotonia, and dysmorphic features. Here, we report a de novo PAK1 NM_002576.5: c.1409 T > A variant (p.Leu470Gln) identified by trio genome sequencing (GS) in a 13-year-old boy with postnatal macrocephaly, obstructive hydrocephalus, medically refractory epilepsy, spastic quadriplegia, white matter hyperintensities, profound developmental disabilities, and a horseshoe kidney. This is the first recurrently affected residue identified in the protein kinase domain. Combined assessment of the eight pathogenic PAK1 missense variants reveal that the variants cluster in either the protein kinase or autoregulatory domains. Although interpretation of the phenotypic spectrum is limited by the sample size, neuroanatomical alterations were found more often in individuals with PAK1 variants in the autoregulatory domain. In contrast, non-neurological comorbidities were found more often in individuals with PAK1 variants in the protein kinase domain. Together, these findings expand the clinical spectrum of PAK1-associated IDDMSSD and reveal potential correlations with the affected protein domains.

Neurobehavioral deficits and a progressive ictogenesis in the tetrodotoxin model of epileptic spasms.

OBJECTIVE: Our goal was to determine whether animals with a history of epileptic spasms have learning and memory deficits. We also used continuous (24/7) long-term electroencephalographic (EEG) recordings to evaluate the evolution of epileptiform activity in the same animals over time. METHODS: Object recognition memory and object location memory tests were undertaken, as well as a matching to place water maze test that evaluated working memory. A retrospective analysis was undertaken of long-term video/EEG recordings from rats with epileptic spasms. The frequency and duration of the ictal events of spasms were quantified. RESULTS: Rats with a history of epileptic spasms showed impairment on the three behavioral tests, and their scores on the object recognition memory and matching to place water maze tests indicated neocortical involvement in the observed impaired cognition. Analysis of EEG recordings unexpectedly showed that the ictal events of spasms and their accompanying behaviors progressively increased in duration over a 2-week period soon after onset, after which spasm duration plateaued. At the same time, spasm frequency remained unchanged. Soon after spasm onset, ictal events were variable in wave form but became more stereotyped as the syndrome evolved. SIGNIFICANCE: Our EEG findings are the first to demonstrate progressive ictogenesis for epileptic spasms. Furthermore, in demonstrating cognitive deficits in the tetrodotoxin model, we have met a criterion for an animal model of West syndrome. Animal models will allow in-depth studies of spasm progression's potential role in cognitive regression and may elucidate why early treatment is considered essential for improved neurodevelopmental outcomes in children.

A Role for Insulin-like Growth Factor 1 in the Generation of Epileptic Spasms in a murine model.

OBJECTIVE: Infantile spasms are associated with a wide variety of clinical conditions, including perinatal brain injuries. We have created a model in which prolonged infusion of tetrodotoxin (TTX) into the neocortex, beginning in infancy, produces a localized lesion and reproduces the behavioral spasms, electroencephalogram (EEG) abnormalities, and drug responsiveness seen clinically. Here, we undertook experiments to explore the possibility that the growth factor IGF-1 plays a role in generating epileptic spasms. METHODS: We combined long-term video EEG recordings with quantitative immunohistochemical and biochemical analyses to unravel IGF-1's role in spasm generation. Immunohistochemistry was undertaken in surgically resected tissue from infantile spasms patients. We used viral injections in neonatal conditional IGF-1R knock-out mice to show that an IGF-1-derived tripeptide (1-3)IGF-1, acts through the IGF-1 receptor to abolish spasms. RESULTS: Immunohistochemical methods revealed widespread loss of IGF-1 from cortical neurons, but an increase in IGF-1 in the reactive astrocytes in the TTX-induced lesion. Very similar changes were observed in the neocortex from patients with spasms. In animals, we observed reduced signaling through the IGF-1 growth pathways in areas remote from the lesion. To show the reduction in IGF-1 expression plays a role in spasm generation, epileptic rats were treated with (1-3)IGF-1. We provide 3 lines of evidence that (1-3)IGF-1 activates the IGF-1 signaling pathway by acting through the receptor for IGF-1. Treatment with (1-3)IGF-1 abolished spasms and hypsarrhythmia-like activity in the majority of animals. INTERPRETATION: Results implicate IGF-1 in the pathogenesis of infantile spasms and IGF-1 analogues as potential novel therapies for this neurodevelopmental disorder. ANN NEUROL 2022;92:45-60.

Brain state-dependent high-frequency activity as a biomarker for abnormal neocortical networks in an epileptic spasms animal model.

OBJECTIVE: Epileptic spasms are a hallmark of a severe epileptic state. A previous study showed neocortical up and down states defined by unit activity play a role in the generation of spasms. However, recording unit activity is challenging in clinical settings, and more accessible neurophysiological signals are needed for the analysis of these brain states. METHODS: In the tetrodotoxin model, we used 16-channel microarrays to record electrophysiological activity in the neocortex during interictal periods and spasms. High-frequency activity (HFA) in the frequency range of fast ripples (200-500 Hz) was analyzed, as were slow wave oscillations (1-8 Hz), and correlated with the neocortical up and down states defined by multiunit activity (MUA). RESULTS: HFA and MUA had high temporal correlation during interictal and ictal periods. Both increased strikingly during interictal up states and ictal events but were silenced during interictal down states and preictal pauses, and their distributions were clustered at the peak of slow oscillations in local field potential recordings. In addition, both HFA power and MUA firing rates were increased to a greater extent during spasms than interictal up states. During non-rapid eye movement sleep, the HFA rhythmicity faithfully followed the MUA up and down states, but during rapid eye movement sleep when MUA up and down states disappeared the HFA rhythmicity was largely absent. We also observed an increase in the number of HFA down state minutes prior to ictal onset, consistent with the results from analyses of MUA down states. SIGNIFICANCE: This study provides evidence that HFA may serve as a biomarker for the pathological up states of epileptic spasms. The availability of HFA recordings makes this a clinically practical technique. These findings will likely provide a novel approach for localizing and studying epileptogenic neocortical networks not only in spasms patients but also in other types of epilepsy.

Acthar® Gel (repository corticotropin injection) dose-response relationships in an animal model of epileptic spasms.

Studies were undertaken to evaluate the effectiveness of Acthar® Gel (repository corticotropin injection [RCI]) in the tetrodotoxin (TTX) model of early-life-induced epileptic spasms. Repository corticotropin injection (RCI) is widely used in the United States to treat infantile spasms. A major component of RCI is N25 deamidated ACTH. Additionally, we hoped to provide some insight into the possible role circulating corticosteroids play in spasm cessation by comparing the RCI dose-response relationships for spasm suppression to RCI-induced corticosterone release from the adrenal gland. Spasms were induced by chronic TTX infusion into the neocortex beginning on postnatal day 11. Repository corticotropin injection (RCI) dosages were between 8 and 32 IU/kg/day. Drug titration protocols were used, and comparisons were made to injections of a vehicle gel. Video/EEG recordings (24/7) monitored the drug's effects continuously for up to 2 months. Tetrodotoxin (TTX)-infused control rats were monitored for the same period of time. In separate experiments, the same dosages of RCI were given to rats and 1 h later plasma was collected and assayed for corticosterone. A parallel study compared the effects of 1-day and 10-day RCI treatments on circulating corticosterone. Results showed that RCI was ineffective at dosages of 8, 12, and 16 IU/kg/day but eliminated spasms in 66% of animals treated with 24 or 32 IU/kg/day. Treating animals with 32 IU/kg/day alone produced the same degree of spasms suppression as observed during the titration protocols. In rats that had hypsarrhythmia-like activity, RCI eliminated this abnormal interictal EEG pattern in all rats that became seizure-free. In terms of plasma corticosterone, 1- and 10-day treatments with RCI produced similar increases in this hormone and the levels increased linearly with increasing dosages of RCI. This stood in sharp contrast to the sigmoid-like dose-response curve for decreases in spasm counts. Our results further validate the TTX model as relevant for the study of infantile spasms. The model should be useful for investigating how RCI acts to eliminate seizures and hypsarrhythmia. Dose-response results suggest that either very high concentrations of circulating corticosteroids are required to abolish spasms or RCI acts through a different mechanism.

Neocortical Slow Oscillations Implicated in the Generation of Epileptic Spasms.

OBJECTIVE: Epileptic spasms are a hallmark of severe seizure disorders. The neurophysiological mechanisms and the neuronal circuit(s) that generate these seizures are unresolved and are the focus of studies reported here. METHODS: In the tetrodotoxin model, we used 16-channel microarrays and microwires to record electrophysiological activity in neocortex and thalamus during spasms. Chemogenetic activation was used to examine the role of neocortical pyramidal cells in generating spasms. Comparisons were made to recordings from infantile spasm patients. RESULTS: Current source density and simultaneous multiunit activity analyses indicate that the ictal events of spasms are initiated in infragranular cortical layers. A dramatic pause of neuronal activity was recorded immediately prior to the onset of spasms. This preictal pause is shown to share many features with the down states of slow wave sleep. In addition, the ensuing interictal up states of slow wave rhythms are more intense in epileptic than control animals and occasionally appear sufficient to initiate spasms. Chemogenetic activation of neocortical pyramidal cells supported these observations, as it increased slow oscillations and spasm numbers and clustering. Recordings also revealed a ramp-up in the number of neocortical slow oscillations preceding spasms, which was also observed in infantile spasm patients. INTERPRETATION: Our findings provide evidence that epileptic spasms can arise from the neocortex and reveal a previously unappreciated interplay between brain state physiology and spasm generation. The identification of neocortical up states as a mechanism capable of initiating epileptic spasms will likely provide new targets for interventional therapies. ANN NEUROL 2021;89:226-241.

Stxbp1/Munc18-1 haploinsufficiency impairs inhibition and mediates key neurological features of STXBP1 encephalopathy.

Mutations in genes encoding synaptic proteins cause many neurodevelopmental disorders, with the majority affecting postsynaptic apparatuses and much fewer in presynaptic proteins. Syntaxin-binding protein 1 (STXBP1, also known as MUNC18-1) is an essential component of the presynaptic neurotransmitter release machinery. De novo heterozygous pathogenic variants in STXBP1 are among the most frequent causes of neurodevelopmental disorders including intellectual disabilities and epilepsies. These disorders, collectively referred to as STXBP1 encephalopathy, encompass a broad spectrum of neurologic and psychiatric features, but the pathogenesis remains elusive. Here we modeled STXBP1 encephalopathy in mice and found that Stxbp1 haploinsufficiency caused cognitive, psychiatric, and motor dysfunctions, as well as cortical hyperexcitability and seizures. Furthermore, Stxbp1 haploinsufficiency reduced cortical inhibitory neurotransmission via distinct mechanisms from parvalbumin-expressing and somatostatin-expressing interneurons. These results demonstrate that Stxbp1 haploinsufficient mice recapitulate cardinal features of STXBP1 encephalopathy and indicate that GABAergic synaptic dysfunction is likely a crucial contributor to disease pathogenesis.

PTEN Loss Increases the Connectivity of Fast Synaptic Motifs and Functional Connectivity in a Developing Hippocampal Network.

Changes in synaptic strength and connectivity are thought to be a major mechanism through which many gene variants cause neurological disease. Hyperactivation of the PI3K-mTOR signaling network, via loss of function of repressors such as PTEN, causes epilepsy in humans and animal models, and altered mTOR signaling may contribute to a broad range of neurological diseases. Changes in synaptic transmission have been reported in animal models of PTEN loss; however, the full extent of these changes, and their effect on network function, is still unknown. To better understand the scope of these changes, we recorded from pairs of mouse hippocampal neurons cultured in a two-neuron microcircuit configuration that allowed us to characterize all four major connection types within the hippocampus. Loss of PTEN caused changes in excitatory and inhibitory connectivity, and these changes were postsynaptic, presynaptic, and transynaptic, suggesting that disruption of PTEN has the potential to affect most connection types in the hippocampal circuit. Given the complexity of the changes at the synaptic level, we measured changes in network behavior after deleting Pten from neurons in an organotypic hippocampal slice network. Slices containing Pten-deleted neurons showed increased recruitment of neurons into network bursts. Importantly, these changes were not confined to Pten-deleted neurons, but involved the entire network, suggesting that the extensive changes in synaptic connectivity rewire the entire network in such a way that promotes a widespread increase in functional connectivity.SIGNIFICANCE STATEMENT Homozygous deletion of the Pten gene in neuronal subpopulations in the mouse serves as a valuable model of epilepsy caused by mTOR hyperactivation. To better understand how gene deletions lead to altered neuronal activity, we investigated the synaptic and network effects that occur 1 week after Pten deletion. PTEN loss increased the connectivity of all four types of hippocampal synaptic connections, including two forms of increased inhibition of inhibition, and increased network functional connectivity. These data suggest that single gene mutations that cause neurological diseases such as epilepsy may affect a surprising range of connection types. Moreover, given the robustness of homeostatic plasticity, these diverse effects on connection types may be necessary to cause network phenotypes such as increased synchrony.

The Impact of Electrographic Seizures on Developing Hippocampal Dendrites Is Calcineurin Dependent.

Neurobehavioral abnormalities are commonly associated with intractable childhood epilepsy. Studies from numerous labs have demonstrated cognitive and socialization deficits in rats and mice that have experienced early-life seizures. However, the cellular and molecular mechanisms underlying these effects are unknown. Previously, experiments have shown that recurrent seizures in infancy suppress the growth of hippocampal dendrites at the same time they impair learning and memory. Experiments in slice cultures have also demonstrated dendrite growth suppression. Here, we crossed calcineurin B1 (CaNB1) floxed and Thy1GFP-M mice to produce mice that were homozygous for the both the floxed CaNB1 and the Thy1GFP-M transgene. Littermates that were homozygous for wild-type CaNB1 and Thy1GFP-M served as controls. Hippocampal slice cultures from these mice were transfected with an AAV/hSyn-mCherry-Cre virus to eliminate CaNB1 from neurons. Immunohistochemical results showed that CaNB1 was eliminated from at least 90% of the transfected CA1 pyramidal cells. Moreover, the CaN-dependent nuclear translocation of the CREB transcription coactivator, CREB-regulated transcriptional coactivator 1 (CRTC1), was blocked in transfected neurons. Cell attach patch recordings combined with live multiphoton imaging demonstrated that the loss of CaNB1 did not prevent neurons from fully participating in electrographic seizure activity. Finally, dendrite reconstruction showed that the elimination of CaNB1 prevented seizure-induced decreases in both dendrite length and branch number. Results suggest that CaN plays a key role in seizure-induced dendrite growth suppression and may contribute to the neurobehavioral comorbidities of childhood epilepsy.

Activation of extracellular regulated kinase and mechanistic target of rapamycin pathway in focal cortical dysplasia.

Neuropathology of resected brain tissue has revealed an association of focal cortical dysplasia (FCD) with drug-resistant epilepsy (DRE). Recent studies have shown that the mechanistic target of rapamycin (mTOR) pathway is hyperactivated in FCD as evidenced by increased phosphorylation of the ribosomal protein S6 (S6) at serine 240/244 (S(240/244) ), a downstream target of mTOR. Moreover, extracellular regulated kinase (ERK) has been shown to phosphorylate S6 at serine 235/236 (S(235/236) ) and tuberous sclerosis complex 2 (TSC2) at serine 664 (S(664) ) leading to hyperactive mTOR signaling. We evaluated ERK phosphorylation of S6 and TSC2 in two types of FCD (FCD I and FCD II) as a candidate mechanism contributing to mTOR pathway dysregulation. Tissue samples from patients with tuberous sclerosis (TS) served as a positive control. Immunostaining for phospho-S6 (pS6(240/244) and pS6(235/236) ), phospho-ERK (pERK), and phospho-TSC2 (pTSC2) was performed on resected brain tissue with FCD and TS. We found increased pS6(240/244) and pS6(235/236) staining in FCD I, FCD II and TS compared to normal-appearing tissue, while pERK and pTSC2 staining was increased only in FCD IIb and TS tissue. Our results suggest that both the ERK and mTOR pathways are dysregulated in FCD and TS; however, the signaling alterations are different for FCD I as compared to FCD II and TS.

Reversal of phenotypes in MECP2 duplication mice using genetic rescue or antisense oligonucleotides.

Copy number variations have been frequently associated with developmental delay, intellectual disability and autism spectrum disorders. MECP2 duplication syndrome is one of the most common genomic rearrangements in males and is characterized by autism, intellectual disability, motor dysfunction, anxiety, epilepsy, recurrent respiratory tract infections and early death. The broad range of deficits caused by methyl-CpG-binding protein 2 (MeCP2) overexpression poses a daunting challenge to traditional biochemical-pathway-based therapeutic approaches. Accordingly, we sought strategies that directly target MeCP2 and are amenable to translation into clinical therapy. The first question that we addressed was whether the neurological dysfunction is reversible after symptoms set in. Reversal of phenotypes in adult symptomatic mice has been demonstrated in some models of monogenic loss-of-function neurological disorders, including loss of MeCP2 in Rett syndrome, indicating that, at least in some cases, the neuroanatomy may remain sufficiently intact so that correction of the molecular dysfunction underlying these disorders can restore healthy physiology. Given the absence of neurodegeneration in MECP2 duplication syndrome, we propose that restoration of normal MeCP2 levels in MECP2 duplication adult mice would rescue their phenotype. By generating and characterizing a conditional Mecp2-overexpressing mouse model, here we show that correction of MeCP2 levels largely reverses the behavioural, molecular and electrophysiological deficits. We also reduced MeCP2 using an antisense oligonucleotide strategy, which has greater translational potential. Antisense oligonucleotides are small, modified nucleic acids that can selectively hybridize with messenger RNA transcribed from a target gene and silence it, and have been successfully used to correct deficits in different mouse models. We find that antisense oligonucleotide treatment induces a broad phenotypic rescue in adult symptomatic transgenic MECP2 duplication mice (MECP2-TG), and corrected MECP2 levels in lymphoblastoid cells from MECP2 duplication patients in a dose-dependent manner.

Loss of MeCP2 in Parvalbumin-and Somatostatin-Expressing Neurons in Mice Leads to Distinct Rett Syndrome-like Phenotypes.

Inhibitory neurons are critical for proper brain function, and their dysfunction is implicated in several disorders, including autism, schizophrenia, and Rett syndrome. These neurons are heterogeneous, and it is unclear which subtypes contribute to specific neurological phenotypes. We deleted Mecp2, the mouse homolog of the gene that causes Rett syndrome, from the two most populous subtypes, parvalbumin-positive (PV+) and somatostatin-positive (SOM+) neurons. Loss of MeCP2 partially impairs the affected neuron, allowing us to assess the function of each subtype without profound disruption of neuronal circuitry. We found that mice lacking MeCP2 in either PV+ or SOM+ neurons have distinct, non-overlapping neurological features: mice lacking MeCP2 in PV+ neurons developed motor, sensory, memory, and social deficits, whereas those lacking MeCP2 in SOM+ neurons exhibited seizures and stereotypies. Our findings indicate that PV+ and SOM+ neurons contribute complementary aspects of the Rett phenotype and may have modular roles in regulating specific behaviors.

Vigabatrin therapy implicates neocortical high frequency oscillations in an animal model of infantile spasms.

Abnormal high frequency oscillations (HFOs) in EEG recordings are thought to be reflections of mechanisms responsible for focal seizure generation in the temporal lobe and neocortex. HFOs have also been recorded in patients and animal models of infantile spasms. If HFOs are important contributors to infantile spasms then anticonvulsant drugs that suppress these seizures should decrease the occurrence of HFOs. In experiments reported here, we used long-term video/EEG recordings with digital sampling rates capable of capturing HFOs. We tested the effectiveness of vigabatrin (VGB) in the TTX animal model of infantile spasms. VGB was found to be quite effective in suppressing spasms. In 3 of 5 animals, spasms ceased after a daily two week treatment. In the other 2 rats, spasm frequency dramatically decreased but gradually increased following treatment cessation. In all animals, hypsarrhythmia was abolished by the last treatment day. As VGB suppressed the frequency of spasms, there was a decrease in the intensity of the behavioral spasms and the duration of the ictal EEG event. Analysis showed that there was a burst of high frequency activity at ictal onset, followed by a later burst of HFOs. VGB was found to selectively suppress the late HFOs of ictal complexes. VGB also suppressed abnormal HFOs recorded during the interictal periods. Thus VGB was found to be effective in suppressing both the generation of spasms and hypsarrhythmia in the TTX model. Vigabatrin also appears to preferentially suppress the generation of abnormal HFOs, thus implicating neocortical HFOs in the infantile spasms disease state.

Catastrophic childhood epilepsy: a recent convergence of basic and clinical neuroscience.

Advances in understanding the genetics and underlying pathology of the catastrophic childhood epilepsies are pointing toward treatments.

How is homeostatic plasticity important in epilepsy?

Maintaining physiological variables within narrow operating limits by homeostatic mechanisms is a fundamental property of most if not all living cells and organisms. In recent years, research from many laboratories has shown that the activity of neurons and neural circuits are also homeostatically regulated. Here, we attempt to apply concepts of homeostasis in general, and more specifically synaptic homeostatic plasticity, to the study of epilepsy. We hypothesize that homeostatic mechanisms are actively engaged in the epileptic brain. These processes attempt to re-establish normal neuronal and network activity, but are opposed by the concurrent mechanisms underlying epileptogenesis. In forms of intractable epilepsy, seizures are so frequent and intense that homeostatic mechanisms are unable to restore normal levels of neuronal activity. In such cases, we contend that homeostatic plasticity mechanisms nevertheless remain active. However, their continuing attempts to reset neuronal activity become maladaptive and results in dyshomeostasis with neurobehavioral consequences. Using the developing hippocampus as a model system, we briefly review experimental results and present a series of arguments to propose that the cognitive neurobehavioral comorbidities of childhood epilepsy result, at least in part, from unchecked homeostatic mechanisms.

Early-life seizures result in deficits in social behavior and learning.

Children with epilepsy show a high co-morbidity with psychiatric disorders and autism. One of the critical determinants of a child's behavioral outcome with autism and cognitive dysfunction is the age of onset of seizures. In order to examine whether seizures during postnatal days 7-11 result in learning and memory deficits and behavioral features of autism we administered the inhalant flurothyl to induce seizures in C57BL/6J mice. Mice received three seizures per day for five days starting on postnatal day 7. Parallel control groups consisted of similarly handled animals that were not exposed to flurothyl and naïve mice. Subjects were then processed through a battery of behavioral tests in adulthood: elevated-plus maze, nose-poke assay, marble burying, social partition, social chamber, fear conditioning, and Morris water maze. Mice with early-life seizures had learning and memory deficits in the training portion of the Morris water maze (p<0.05) and probe trial (p<0.01). Mice with seizures showed no differences in marble burying, the nose-poke assay, or elevated plus-maze testing compared to controls. However, they showed a significant difference in the social chamber and social partition tests. Mice with seizures during postnatal days 7-11 showed a significant decrease in social interaction in the social chamber test and had a significant impairment in social behavior in the social partition test. Together, these results indicate that early life seizures result in deficits in hippocampal-dependent memory tasks and produce long-term disruptions in social behavior.

Loss of mTOR repressors Tsc1 or Pten has divergent effects on excitatory and inhibitory synaptic transmission in single hippocampal neuron cultures.

The Pten and Tsc1 genes both encode proteins that repress mechanistic target of rapamycin (mTOR) signaling. Disruption of either gene in the brain results in epilepsy and autism-like symptoms in humans and mouse models, therefore it is important to understand the molecular and physiological events that lead from gene disruption to disease phenotypes. Given the similar roles these two molecules play in the regulation of cellular growth and the overlap in the phenotypes that result from their loss, we predicted that the deletion of either the Pten or Tsc1 gene from autaptic hippocampal neurons would have similar effects on neuronal morphology and synaptic transmission. Accordingly, we found that loss of either Pten or Tsc1 caused comparable increases in soma size, dendrite length and action potential properties. However, the effects of Pten and Tsc1 loss on synaptic transmission were different. Loss of Pten lead to an increase in both excitatory and inhibitory neurotransmission, while loss of Tsc1 did not affect excitatory neurotransmission and reduced inhibitory transmission by decreasing mIPSC amplitude. Although the loss of Pten or Tsc1 both increased downstream mTORC1 signaling, phosphorylation of Akt was increased in Pten-ko and decreased in Tsc1-ko neurons, potentially accounting for the different effects on synaptic transmission. Despite the different effects at the synaptic level, our data suggest that loss of Pten or Tsc1 may both lead to an increase in the ratio of excitation to inhibition at the network level, an effect that has been proposed to underlie both epilepsy and autism.

The effects of early-life seizures on hippocampal dendrite development and later-life learning and memory.

Severe childhood epilepsy is commonly associated with intellectual developmental disabilities. The reasons for these cognitive deficits are likely multifactorial and will vary between epilepsy syndromes and even among children with the same syndrome. However, one factor these children have in common is the recurring seizures they experience - sometimes on a daily basis. Supporting the idea that the seizures themselves can contribute to intellectual disabilities are laboratory results demonstrating spatial learning and memory deficits in normal mice and rats that have experienced recurrent seizures in infancy. Studies reviewed here have shown that seizures in vivo and electrographic seizure activity in vitro both suppress the growth of hippocampal pyramidal cell dendrites. A simplification of dendritic arborization and a resulting decrease in the number and/or properties of the excitatory synapses on them could help explain the observed cognitive disabilities. There are a wide variety of candidate mechanisms that could be involved in seizure-induced growth suppression. The challenge is designing experiments that will help focus research on a limited number of potential molecular events. Thus far, results suggest that growth suppression is NMDA receptor-dependent and associated with a decrease in activation of the transcription factor CREB. The latter result is intriguing since CREB is known to play an important role in dendrite growth. Seizure-induced dendrite growth suppression may not occur as a single process in which pyramidal cells dendrites simply stop growing or grow slower compared to normal neurons. Instead, recent results suggest that after only a few hours of synchronized epileptiform activity in vitro dendrites appear to partially retract. This acute response is also NMDA receptor dependent and appears to be mediated by the Ca(+2)/calmodulin-dependent phosphatase, calcineurin. An understanding of the staging of seizure-induced growth suppression and the underlying molecular mechanisms will likely prove crucial for developing therapeutic strategies aimed at ameliorating the intellectual developmental disabilities associated with intractable childhood epilepsy.

Multiple roles for mammalian target of rapamycin signaling in both glutamatergic and GABAergic synaptic transmission.

The mammalian target of rapamycin (mTOR) signaling pathway in neurons integrates a variety of extracellular signals to produce appropriate translational responses. mTOR signaling is hyperactive in neurological syndromes in both humans and mouse models that are characterized by epilepsy, autism, and cognitive disturbances. In addition, rapamycin, a clinically important immunosuppressant, is a specific and potent inhibitor of mTOR signaling. While mTOR is known to regulate growth and synaptic plasticity of glutamatergic neurons, its effects on basic parameters of synaptic transmission are less well studied, and its role in regulating GABAergic transmission is unexplored. We therefore performed an electrophysiological and morphological comparison of glutamatergic and GABAergic neurons in which mTOR signaling was either increased by loss of the repressor Pten or decreased by treatment with rapamycin. We found that hyperactive mTOR signaling increased evoked synaptic responses in both glutamatergic and GABAergic neurons by ∼50%, due to an increase in the number of synaptic vesicles available for release, the number of synapses formed, and the miniature event size. Prolonged (72 h) rapamycin treatment prevented these abnormalities and also decreased synaptic transmission in wild-type glutamatergic, but not GABAergic, neurons. Further analyses suggested that hyperactivation of the mTOR pathway also impairs presynaptic function, possibly by interfering with vesicle fusion. Despite this presynaptic impairment, the net effect of Pten loss is enhanced synaptic transmission in both GABAergic and glutamatergic neurons, which has numerous implications, depending on where in the brain mutations of an mTOR suppressor gene occur.

Impact of seizures on developing dendrites: implications for intellectual developmental disabilities.

Childhood epilepsy can be severe and even catastrophic. In these instances, cognition can be impaired-leading to long-term intellectual disabilities. One factor that could potentially cause cognitive deficits is the frequent seizures that characterize intractable epilepsy. However, it has been difficult to separate the effects seizures may have from those of preexisting neuropathologies and/or the effects of ongoing anticonvulsant therapies. Therefore, important questions are: Do early life seizures produce the learning deficits? And if they do, how do they do it? Results from recent animal models studies reviewed here show that recurrent seizures in infancy stop the growth of CA1 hippocampal dendrites. We speculate that the molecular mechanisms responsible for seizure-induced growth suppression are homeostatic/neuroprotective, used by the developing nervous system in an attempt to limit neuronal and network excitability and prevent the continued generation of seizures. However, by preventing the normal growth of dendrites, there is a reduction in CA1 glutamatergic synapses that supports long-lasting forms of synaptic plasticity thought to be the cellular basis of learning and memory. Therefore, dendrite growth suppression would reduce the neuroanatomic substrates for learning and memory, and in so doing could contribute in important ways to spatial learning and memory deficits that may be relevant to the cognitive deficits associated with childhood epilepsy.