The mechanisms that extend lifespan in humans are poorly understood. Here we show that extended longevity in humans is associated with a distinct transcriptome signature in the cerebral cortex that ...is characterized by downregulation of genes related to neural excitation and synaptic function. In Caenorhabditis elegans, neural excitation increases with age and inhibition of excitation globally, or in glutamatergic or cholinergic neurons, increases longevity. Furthermore, longevity is dynamically regulated by the excitatory-inhibitory balance of neural circuits. The transcription factor REST is upregulated in humans with extended longevity and represses excitation-related genes. Notably, REST-deficient mice exhibit increased cortical activity and neuronal excitability during ageing. Similarly, loss-of-function mutations in the C. elegans REST orthologue genes spr-3 and spr-4 elevate neural excitation and reduce the lifespan of long-lived daf-2 mutants. In wild-type worms, overexpression of spr-4 suppresses excitation and extends lifespan. REST, SPR-3, SPR-4 and reduced excitation activate the longevity-associated transcription factors FOXO1 and DAF-16 in mammals and worms, respectively. These findings reveal a conserved mechanism of ageing that is mediated by neural circuit activity and regulated by REST.
Repetitive transcranial magnetic stimulation (rTMS) is a widely-used method for modulating cortical excitability in humans, by mechanisms thought to involve use-dependent synaptic plasticity. For ...example, when low frequency rTMS (LF rTMS) is applied over the motor cortex, in humans, it predictably leads to a suppression of the motor evoked potential (MEP), presumably reflecting long-term depression (LTD) -like mechanisms. Yet how closely such rTMS effects actually match LTD is unknown. We therefore sought to (1) reproduce cortico-spinal depression by LF rTMS in rats, (2) establish a reliable animal model for rTMS effects that may enable mechanistic studies, and (3) test whether LTD-like properties are evident in the rat LF rTMS setup. Lateralized MEPs were obtained from anesthetized Long-Evans rats. To test frequency-dependence of LF rTMS, rats underwent rTMS at one of three frequencies, 0.25, 0.5, or 1 Hz. We next tested the dependence of rTMS effects on N-methyl-D-aspartate glutamate receptor (NMDAR), by application of two NMDAR antagonists. We find that 1 Hz rTMS preferentially depresses unilateral MEP in rats, and that this LTD-like effect is blocked by NMDAR antagonists. These are the first electrophysiological data showing depression of cortical excitability following LF rTMS in rats, and the first to demonstrate dependence of this form of cortical plasticity on the NMDAR. We also note that our report is the first to show that the capacity for LTD-type cortical suppression by rTMS is present under barbiturate anesthesia, suggesting that future neuromodulatory rTMS applications under anesthesia may be considered.
DEPDC5 is a newly identified epilepsy-related gene implicated in focal epilepsy, brain malformations, and Sudden Unexplained Death in Epilepsy (SUDEP). In vitro, DEPDC5 negatively regulates amino ...acid sensing by the mTOR complex 1 (mTORC1) pathway, but the role of DEPDC5 in neurodevelopment and epilepsy has not been described. No animal model of DEPDC5-related epilepsy has recapitulated the neurological phenotypes seen in patients, and germline knockout rodent models are embryonic lethal. Here, we establish a neuron-specific Depdc5 conditional knockout mouse by cre-recombination under the Synapsin1 promotor. Depdc5flox/flox-Syn1Cre (Depdc5cc+) mice survive to adulthood with a progressive neurologic phenotype that includes motor abnormalities (i.e., hind limb clasping) and reduced survival compared to littermate control mice. Depdc5cc+ mice have larger brains with increased cortical neuron size and dysplastic neurons throughout the cortex, comparable to the abnormal neurons seen in human focal cortical dysplasia specimens. Depdc5 results in constitutive mTORC1 hyperactivation exclusively in neurons as measured by the increased phosphorylation of the downstream ribosomal protein S6. Despite a lack of increased mTORC1 signaling within astrocytes, Depdc5cc+ brains show reactive astrogliosis. We observed two Depdc5cc+ mice to have spontaneous seizures, including a terminal seizure. We demonstrate that as a group Depdc5cc+ mice have lowered seizure thresholds, as evidenced by decreased latency to seizures after chemoconvulsant injection and increased mortality from pentylenetetrazole-induced seizures. In summary, our neuron-specific Depdc5 knockout mouse model recapitulates clinical, pathological, and biochemical features of human DEPDC5-related epilepsy and brain malformations. We thereby present an important model in which to study targeted therapeutic strategies for DEPDC5-related conditions.
•Conditional neuron-specific Depdc5 knockout mice (Depdc5cc+) have dysplastic cortical neurons.•Similar to other mTORopathy models, Depdc5cc+ mice have larger brains and increased neuron size.•Neuronal Depdc5 loss results in neuronal mTORC1 hyperactivation and evidence of gliosis.•Depdc5cc+ mice demonstrate a decreased seizure threshold, and some display seizure.•Depdc5cc+ mice have increased mortality, likely due in part to terminal seizures.
Angelman syndrome (AS) is a neurodevelopmental disorder arising from loss-of-function mutations in the maternally inherited copy of the UBE3A gene, and is characterized by an absence of speech, ...excessive laughter, cognitive delay, motor deficits, and seizures. Despite the fact that the symptoms of AS occur in early childhood, behavioral characterization of AS mouse models has focused primarily on adult phenotypes. In this report we describe juvenile behaviors in AS mice that are strain-independent and clinically relevant. We find that young AS mice, compared with their wild-type littermates, produce an increased number of ultrasonic vocalizations. In addition, young AS mice have defects in motor coordination, as well as abnormal brain activity that results in an enhanced seizure-like response to an audiogenic challenge. The enhanced seizure-like activity, but not the increased ultrasonic vocalizations or motor deficits, is rescued in juvenile AS mice by genetically reducing the expression level of the activity-regulated cytoskeleton-associated protein, Arc . These findings suggest that therapeutic interventions that reduce the level of Arc expression have the potential to reverse the seizures associated with AS. In addition, the identification of aberrant behaviors in young AS mice may provide clues regarding the neural circuit defects that occur in AS and ultimately allow new approaches for treating this disorder.
Significance Angelman syndrome (AS) is a human neurodevelopmental disorder caused by mutation of a specific gene, UBE3A . Studies of behavior in adult mouse models of AS reveal abnormalities similar to those observed in humans with AS. Because AS affects children, we hypothesized that it might be helpful to study this disorder using juvenile mice. We found that young AS mice display aberrant communication and motor behaviors and increased brain activity. Reducing the expression of the synaptic protein ARC reverses abnormal brain activity in AS mice, but has no effect on communication and motor behaviors in these mice. These findings suggest new approaches for identifying the neural circuits that are defective in AS, and for developing therapies for treating this disorder.
Promising results in adult neurologic and psychiatric disorders are driving active research into transcranial brain stimulation techniques, particularly transcranial magnetic stimulation (TMS) and ...transcranial direct current stimulation (tDCS), in childhood and adolescent syndromes. TMS has realistic utility as an experimental tool tested in a range of pediatric neuropathologies such as perinatal stroke, depression, Tourette syndrome, and autism spectrum disorder (ASD). tDCS has also been tested as a treatment for a number of pediatric neurologic conditions, including ASD, attention-deficit/hyperactivity disorder, epilepsy, and cerebral palsy. Here, we complement recent reviews with an update of published TMS and tDCS results in children, and discuss developmental neuroscience considerations that should inform pediatric transcranial stimulation.
Excessive extracellular glutamate after traumatic brain injury (TBI) contributes to excitotoxic cell death and likely to post-traumatic epilepsy. Glutamate transport is the only known mechanism of ...extracellular glutamate clearance, and glutamate transporter 1 (GLT-1) is the major glutamate transporter of the mammalian brain. We tested, by immunoblot, in the rat lateral fluid percussion injury TBI model whether GLT-1 expression is depressed in the cortex after TBI, and whether GLT-1 expression after TBI is restored after treatment with ceftriaxone, a well-tolerated β-lactam antibiotic previously shown to enhance GLT-1 expression in noninjured animals. We then tested whether treatment with ceftriaxone mitigates the associated regional astrogliosis, as reflected by glial fibrillary acid protein (GFAP) expression, and also whether ceftriaxone treatment mitigates the severity of post-traumatic epilepsy. We found that 7 days after TBI, GLT-1 expression in the ipsilesional cortex was reduced by 29% (n=7/group; p<0.01), relative to the contralesional cortex. However, the loss of GLT-1 expression was reversed by treatment with ceftriaxone (200 mg/kg, daily, intraperitoneally). We found that ceftriaxone treatment also decreased the level of regional GFAP expression by 43% in the lesioned cortex, relative to control treatment with saline (n=7 per group; p<0.05), and, 12 weeks after injury, reduced cumulative post-traumatic seizure duration (n=6 rats in the ceftriaxone treatment group and n=5 rats in the saline control group; p<0.001). We cautiously conclude that our data suggest a potential role for ceftriaxone in treatment of epileptogenic TBI.
GLT-1 (EAAT2; slc1a2) is the major glutamate transporter in the brain, and is predominantly expressed in astrocytes, but at lower levels also in excitatory terminals. We generated a conditional GLT-1 ...knock-out mouse to uncover cell-type-specific functional roles of GLT-1. Inactivation of the GLT-1 gene was achieved in either neurons or astrocytes by expression of synapsin-Cre or inducible human GFAP-CreERT2. Elimination of GLT-1 from astrocytes resulted in loss of ∼80% of GLT-1 protein and of glutamate uptake activity that could be solubilized and reconstituted in liposomes. This loss was accompanied by excess mortality, lower body weight, and seizures suggesting that astrocytic GLT-1 is of major importance. However, there was only a small (15%) reduction that did not reach significance of glutamate uptake into crude forebrain synaptosomes. In contrast, when GLT-1 was deleted in neurons, both the GLT-1 protein and glutamate uptake activity that could be solubilized and reconstituted in liposomes were virtually unaffected. These mice showed normal survival, weight gain, and no seizures. However, the synaptosomal glutamate uptake capacity (Vmax) was reduced significantly (40%). In conclusion, astrocytic GLT-1 performs critical functions required for normal weight gain, resistance to epilepsy, and survival. However, the contribution of astrocytic GLT-1 to glutamate uptake into synaptosomes is less than expected, and the contribution of neuronal GLT-1 to synaptosomal glutamate uptake is greater than expected based on their relative protein expression. These results have important implications for the interpretation of the many previous studies assessing glutamate uptake capacity by measuring synaptosomal uptake.
Autism spectrum disorder (ASD) is a clinically and biologically heterogeneous condition characterized by social, repetitive, and sensory behavioral abnormalities. No treatments are approved for the ...core diagnostic symptoms of ASD. To enable the earliest stages of therapeutic discovery and development for ASD, robust and reproducible behavioral phenotypes and biological markers are essential to establish in preclinical animal models. The goal of this study was to identify electroencephalographic (EEG) and behavioral phenotypes that are replicable between independent cohorts in a mouse model of ASD. The larger goal of our strategy is to empower the preclinical biomedical ASD research field by generating robust and reproducible behavioral and physiological phenotypes in animal models of ASD, for the characterization of mechanistic underpinnings of ASD-relevant phenotypes, and to ensure reliability for the discovery of novel therapeutics. Genetic disruption of the
gene, a scaffolding protein involved in the stability of the postsynaptic density in excitatory synapses, is thought to be responsible for a relatively large number of cases of ASD. Therefore, we have thoroughly characterized the robustness of ASD-relevant behavioral phenotypes in two cohorts, and for the first time quantified translational EEG activity in
null mutant mice.
In vivo physiology and behavioral assays were conducted in two independently bred and tested full cohorts of
null mutant (
KO) and wildtype littermate control (WT) mice. EEG was recorded via wireless implanted telemeters for 7 days of baseline followed by 20 min of recording following pentylenetetrazol (PTZ) challenge. Behaviors relevant to the diagnostic and associated symptoms of ASD were tested on a battery of established behavioral tests. Assays were designed to reproduce and expand on the original behavioral characterization of
KO mice. Two or more corroborative tests were conducted within each behavioral domain, including social, repetitive, cognitive, anxiety-related, sensory, and motor categories of assays.
Relative to WT mice,
KO mice displayed a dramatic resistance to PTZ seizure induction and an enhancement of gamma band oscillatory EEG activity indicative of enhanced inhibitory tone. These findings replicated in two separate cohorts. Behaviorally,
mice exhibited repetitive grooming, deficits in aspects of reciprocal social interactions and vocalizations, and reduced open field activity, as well as variable deficits in sensory responses, anxiety-related behaviors, learning and memory.
Robust animal models and quantitative, replicable biomarkers of neural dysfunction are needed to decrease risk and enable successful drug discovery and development for ASD and other neurodevelopmental disorders. Complementary to the replicated behavioral phenotypes of the
mutant mouse is the new identification of a robust, translational in vivo neurophysiological phenotype. Our findings provide strong evidence for robustness and replicability of key translational phenotypes in
mutant mice and support the usefulness of this mouse model of ASD for therapeutic discovery.
We present an update to the status of research on succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD), a rare disorder of GABA metabolism. This is an unusual disorder featuring the ...accumulation of both GABA and its neuromodulatory analog, gamma-hydroxybutyric acid (GHB), and recent studies have advanced the potential clinical application of NCS-382, a putative GHB receptor antagonist. Animal studies have provided proof-of-concept that enzyme replacement therapy could represent a long-term therapeutic option. The characterization of neuronal stem cells (NSCs) derived from
aldehyde dehydrogenase 5a1
−/−
(
aldh5a1
−/−
) mice, the murine model of SSADHD, has highlighted NSC utility as an in vitro system in which to study therapeutics and associated toxicological properties. Gene expression analyses have revealed that transcripts encoding GABA
A
receptors are down-regulated and may remain largely immature in
aldh5a1
−/−
brain, characterized by excitatory as opposed to inhibitory outputs, the latter being the expected action in the mature central nervous system. This indicates that agents altering chloride channel activity may be therapeutically relevant in SSADHD. The most recent therapeutic prospects include mTOR (mechanistic target of rapamycin) inhibitors, drugs that have received attention with the elucidation of the effects of elevated GABA on autophagy. The outlook for novel therapeutic trials in SSADHD continues to improve.
Drugs targeting metabotropic glutamate receptor 5 (mGluR5) have therapeutic potential in autism spectrum disorders (ASD), including tuberous sclerosis complex (TSC). The question whether inhibition ...or potentiation of mGluR5 could be beneficial depends, among other factors, on the specific indication. To facilitate the development of mGluR5 treatment strategies, we tested the therapeutic utility of mGluR5 negative and positive allosteric modulators (an mGluR5 NAM and PAM) for TSC, using a mutant mouse model with neuronal loss of Tsc2 that demonstrates disease-related phenotypes, including behavioral symptoms of ASD and epilepsy. This model uniquely enables the in vivo characterization and rescue of the electrographic seizures associated with TSC. We demonstrate that inhibition of mGluR5 corrects hyperactivity, seizures, and elevated de novo synaptic protein synthesis. Conversely, positive allosteric modulation of mGluR5 results in the exacerbation of hyperactivity and epileptic phenotypes. The data suggest a meaningful therapeutic potential for mGluR5 NAMs in TSC, which warrants clinical exploration and the continued development of mGluR5 therapies.
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