[Show abstract][Hide abstract] ABSTRACT: This study assessed the anticonvulsant and seizure generation effects of carbamazepine (CBZ), oxcarbazepine (OXC) and eslicarbazepine (S-Lic) in wild-type mice. Electrophysiological recordings were made to discriminate potential cellular and synaptic mechanisms underlying anti- and pro-epileptic actions. The anticonvulsant and pro-convulsant effects were evaluated in the MES, the 6-Hz and the Irwin tests. Whole-cell patch-clamp recordings were used to investigate the effects on fast excitatory and inhibitory synaptic transmission in hippocampal area CA1. The safety window for CBZ, OXC and eslicarbazepine (ED50 value against the MES test and the dose that produces grade 5 convulsions in all mice), was 6.3, 6.0 and 12.5, respectively. At high concentrations the three drugs reduced synaptic transmission. CBZ and OXC enhanced excitatory postsynaptic currents (EPSCs) at low, therapeutically-relevant concentrations. These effects were associated with no change in inhibitory postsynaptic currents (IPSCs) resulting in altered balance between excitation and inhibition. S-Lic had no effect on EPSC or IPSC amplitudes over the same concentration range. The CBZ mediated enhancement of EPSCs was blocked by DPCPX, a selective antagonist, and occluded by CCPA, a selective agonist of the adenosine A1 receptor. Furthermore, reduction of endogenous adenosine by application of the enzyme adenosine deaminase also abolished the CBZ- and OXC-induced increase of EPSCs, indicating that the two drugs act as antagonists at native adenosine receptors. In conclusion, CBZ and OXC possess pro-epileptic actions at clinically-relevant concentrations through the enhancement of excitatory synaptic transmission. S-Lic by comparison has no such effect on synaptic transmission, explaining its lack of seizure exacerbation.
[Show abstract][Hide abstract] ABSTRACT: Rett syndrome (RTT) is a genetic disorder characterized by a range of features including cognitive impairment, gait abnormalities and a reduction in purposeful hand skills. Mice harbouring knockout mutations in the Mecp2 gene display many RTT-like characteristics and are central to efforts to find novel therapies for the disorder. As hand stereotypies and gait abnormalities constitute major diagnostic criteria in RTT, it is clear that motor and gait-related phenotypes will be of importance in assessing preclinical therapeutic outcomes. We therefore aimed to assess gait properties over the prodromal phase in a functional knockout mouse model of RTT. In male Mecp2 knockout mice, we observed alterations in stride, coordination and balance parameters at 4 weeks of age, before the onset of other overt phenotypic changes as revealed by observational scoring. These data suggest that gait measures may be used as a robust and early marker of MeCP2-dysfunction in future preclinical therapeutic studies.
PLoS ONE 11/2014; 9(11):e112889. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Functional Mecp2 knockout mice display a range of deficits in skeletal properties including mechanical strength, stiffness and microhardness.•Biomechanical deficits were accompanied by microstructural changes such as decreased trabecular thickness.•Defects in biomechanical and material properties were ameliorated by unsilencing the Mecp2 gene.
[Show abstract][Hide abstract] ABSTRACT: We use a computational model of a hippocampal CA1 pyramidal cell to demonstrate that spine head calcium provides an instantaneous readout at each synapse of the postsynaptic weighted sum of all presynaptic activity impinging on the cell. The form of the readout is equivalent to the functions of weighted, summed inputs used in neural network learning rules. Within a dendritic layer, peak spine head calcium levels are either a linear or sigmoidal function of the number of coactive synapses, with nonlinearity depending on the ability of voltage spread in the dendrites to reach calcium spike threshold. This is strongly controlled by the potassium A-type current, with calcium spikes and the consequent sigmoidal increase in peak spine head calcium present only when the A-channel density is low. Other membrane characteristics influence the gain of the relationship between peak calcium and the number of active synapses. In particular, increasing spine neck resistance increases the gain due to increased voltage responses to synaptic input in spine heads. Colocation of stimulated synapses on a single dendritic branch also increases the gain of the response. Input pathways cooperate: CA3 inputs to the proximal apical dendrites can strongly amplify peak calcium levels due to weak EC input to the distal dendrites, but not so strongly vice versa. CA3 inputs to the basal dendrites can boost calcium levels in the proximal apical dendrites, but the relative electrical compactness of the basal dendrites results in the reverse effect being less significant. These results give pointers as to how to better describe the contributions of pre- and postsynaptic activity in the learning "rules" that apply in these cells. The calcium signal is closer in form to the activity measures used in traditional neural network learning rules than to the spike times used in spike-timing-dependent plasticity.
[Show abstract][Hide abstract] ABSTRACT: Numerous experimental models have been developed to reiterate endophenotypes of Rett syndrome, a neurodevelopmental disorder with a multitude of motor, cognitive and vegetative symptoms. Here, female Mecp2(Stop) mice (Guy et al., 2007) were characterised at mild symptomatic conditions in tests for anxiety (open field, elevated plus maze) and home cage observation systems for food intake, locomotor activity and circadian rhythms. Aged 8-9 months, Mecp2(Stop) mice presented with heightened body weight, lower overall activity in the open field, but no anxiety phenotype. Although home cage activity scans conducted in two different observation systems, PhenoMaster and PhenoTyper, confirmed normal circadian activity, they revealed severely compromised habituation to a novel environment in all parameters registered including those derived from a non-linear decay model such as initial exploration maximum, decay half-life of activity and span, as well as plateau. Furthermore, overall activity was significantly reduced in nocturnal periods due to reductions in both fast ambulatory movements, but also a slow lingering. In contrast, light-period activity profiles during which the amount of sleep was highest remained normal in Mecp2(Stop) mice. These data confirm the slow and progressive development of Rett-like symptoms in female Mecp2(Stop) mice resulting in a prominent reduction of overall locomotor activity, while circadian rhythms are maintained. Alterations in the time-course of habituation may indicate deficiencies in cognitive processing.
Behavioural brain research 05/2013; · 3.22 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Rett syndrome (RTT) is a disorder with a pronounced neurological phenotype and is caused mainly by mutations in the X-linked gene MECP2. A common feature of RTT is an abnormal EEG and a propensity for seizures. In the current study we aimed to assess brain network excitability and seizure propensity in a mouse model of RTT. Mice in which Mecp2 expression was silenced (Mecp2(stop/y)) showed a higher seizure score (mean = 6 ± 0.8 compared to 4 ± 0.2 in wild-type, WT) and more rapid seizure onset (median onset = 10 mins in Mecp2(stop/y) and 32 mins in WT) when challenged with the convulsant drug kainic acid (25mg/Kg). Hippocampal slices from Mecp2(stop/y) brain displayed no spontaneous field potential activities under control conditions but showed higher power gamma frequency field potential oscillations compared to WT in response to kainic acid (400 nM) in vitro. Brain slices challenged with the GABA(A) receptor antagonist bicuculline (0.1-10μM) and the potassium channel blocker 4-aminopyridine (1-50μM) also revealed differences between genotypes with hippocampal circuits from Mecp2(stop/y) mouse slices showing enhanced epileptiform burst duration and frequency. In contrast to these network level findings, single cell analysis of pyramidal cells by whole-cell patch clamp recording revealed no detectable differences in synaptic or biophysical properties between MeCP2-containing and MeCP2-deficient neurons. These data support the proposal that loss of MeCP2 alters network level excitability in the brain to promote epileptogenesis.
[Show abstract][Hide abstract] ABSTRACT: Typical Rett syndrome (RTT) is a pediatric disorder caused by loss-of-function mutations in the methyl-CpG binding protein 2 (MECP2) gene. The demonstrated reversibility of RTT-like phenotypes in mice suggests that MECP2 gene replacement is a potential therapeutic option in patients. We report improvements in survival and phenotypic severity in Mecp2-null male mice after neonatal intracranial delivery of a single-stranded (ss) AAV9/chicken β-actin (CBA)-MECP2 vector. Median survival was 16.6 weeks for MECP2-treated versus 9.3 weeks for green fluorescent protein (GFP)-treated mice. ssAAV9/CBA-MECP2-treated mice also showed significant improvement in the phenotype severity score, in locomotor function, and in exploratory activity, as well as a normalization of neuronal nuclear volume in transduced cells. Wild-type (WT) mice receiving neonatal injections of the same ssAAV9/CBA-MECP2 vector did not show any significant deficits, suggesting a tolerance for modest MeCP2 overexpression. To test a MECP2 gene replacement approach in a manner more relevant for human translation, a self-complementary (sc) adeno-associated virus (AAV) vector designed to drive MeCP2 expression from a fragment of the Mecp2 promoter was injected intravenously (IV) into juvenile (4-5 weeks old) Mecp2-null mice. While the brain transduction efficiency in juvenile mice was low (~2-4% of neurons), modest improvements in survival were still observed. These results support the concept of MECP2 gene therapy for RTT.Molecular Therapy (2012); doi:10.1038/mt.2012.200.
[Show abstract][Hide abstract] ABSTRACT: Rett syndrome is a neurological disorder caused by mutation of the X-linked MECP2 gene. Mice lacking functional Mecp2 display a spectrum of Rett syndrome-like signs, including disturbances in motor function and abnormal patterns of breathing, accompanied by structural defects in central motor areas and the brainstem. Although routinely classified as a neurodevelopmental disorder, many aspects of the mouse phenotype can be effectively reversed by activation of a quiescent Mecp2 gene in adults. This suggests that absence of Mecp2 during brain development does not irreversibly compromise brain function. It is conceivable, however, that deep-seated neurological defects persist in mice rescued by late activation of Mecp2. To test this possibility, we have quantitatively analysed structural and functional plasticity of the rescued adult male mouse brain. Activation of Mecp2 in ∼70% of neurons reversed many morphological defects in the motor cortex, including neuronal size and dendritic complexity. Restoration of Mecp2 expression was also accompanied by a significant improvement in respiratory and sensory-motor functions, including breathing pattern, grip strength, balance beam and rotarod performance. Our findings sustain the view that MeCP2 does not play a pivotal role in brain development, but may instead be required to maintain full neurological function once development is complete.
[Show abstract][Hide abstract] ABSTRACT: Rett syndrome (RTT), a neurodevelopmental condition characterized by delayed-onset loss of spoken language and the development of distinctive hand stereotypies, affects approximately 1 in 10,000 live female births. Clinical diagnosis has been based on symptoms such as loss of acquired purposeful hand skills, autistic behaviors, motor dysfunctions, seizure disorders, and gait abnormalities. RTT is a genetic disease and is caused almost exclusively by mutations in the X-linked gene, MECP2, to produce a phenotype that is thought to be primarily of neurological origin. Clinical reports show RTT patients to have a smaller brain volume, especially in the cerebral hemispheres, and alterations in various neurotransmitter systems, including acetylcholine, dopamine, serotonin, glutamate, substance P, and various trophic factors. Because of its monogenetic characteristic, disruption of Mecp2 is readily recapitulated in mice to produce a prominent RTT-like phenotype and provide an excellent platform for understanding the pathogenesis of RTT. As shown in human studies, Mecp2 mutants also display subtle alterations in neuronal morphology, including smaller cortical neurons with a higher-packing density and reduced dendritic complexity. Neurophysiological studies in Mecp2-mutant mice consistently report alterations in synaptic function, notably, defects in synaptic plasticity. These data suggest that RTT might be regarded as a synaptopathy (disease of the synapse) and thus potentially amenable to rational therapeutic intervention.
[Show abstract][Hide abstract] ABSTRACT: Caspase 3 activation has been linked to the acute neurotoxic effects of central nervous system damage, as in traumatic brain injury or cerebral ischaemia, and also to the early events leading to long-term neurodegeneration, as in Alzheimer's disease. However, the precise mechanisms activating caspase 3 in neuronal injury are unclear. RhoB is a member of the Rho GTPase family that is dramatically induced by cerebral ischaemia or neurotrauma, both in preclinical models and clinically. In the current study, we tested the hypothesis that RhoB might directly modulate caspase 3 activity and apoptotic or necrotic responses in neurons. Over-expression of RhoB in the NG108-15 neuronal cell line or in cultured corticohippocampal neurons elevated caspase 3 activity without inducing overt toxicity. Cultured corticohippocampal neurons from RhoB knockout mice did not show any differences in sensitivity to a necrotic stimulus - acute calcium ionophore exposure - compared with neurons from wild-type mice. However, corticohippocampal neurons lacking RhoB exhibited a reduction in the degree of DNA fragmentation and caspase 3 activation induced by the apoptotic agent staurosporine, in parallel with increased neuronal survival. Staurosporine induction of caspase 9 activity was also suppressed. RhoB knockout mice showed reduced basal levels of caspase 3 activity in the adult brain. These data directly implicate neuronal RhoB in caspase 3 activation and the initial stages of programmed cell death, and suggest that RhoB may represent an attractive target for therapeutic intervention in conditions involving elevated caspase 3 activity in the central nervous system.
European Journal of Neuroscience 11/2011; 34(11):1737-46. · 3.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mutations in the X-linked gene MECP2 (methyl CpG-binding protein 2) are the primary cause of the neurodevelopmental disorder RTT (Rett syndrome), and are also implicated in other neurological conditions. The expression product of this gene, MeCP2, is a widely expressed nuclear protein, especially abundant in mature neurons of the CNS (central nervous system). The major recognized consequences of MECP2 mutation occur in the CNS, but there is growing awareness of peripheral effects contributing to the full RTT phenotype. MeCP2 is classically considered to act as a DNA methylation-dependent transcriptional repressor, but may have additional roles in regulating gene expression and chromatin structure. Knocking out Mecp2 function in mice recapitulates many of the overt neurological features seen in RTT patients, and the characteristic postnatally delayed onset of symptoms is accompanied by aberrant neuronal morphology and deficits in synaptic physiology. Evidence that reactivation of endogenous Mecp2 in mutant mice, even at adult stages, can reverse aspects of RTT-like pathology and result in apparently functionally mature neurons has provided renewed hope for patients, but has also provoked discussion about traditional boundaries between neurodevelopmental disorders and those involving dysfunction at later stages. In the present paper we review the neurobiology of MeCP2 and consider the various genetic (including gene therapy), pharmacological and environmental interventions that have been, and could be, developed to attempt phenotypic rescue in RTT. Such approaches are already providing valuable insights into the potential tractability of RTT and related conditions, and are useful pointers for the development of future therapeutic strategies.
[Show abstract][Hide abstract] ABSTRACT: Synaptic plasticity is the ability of synaptic connections between neurons to be strengthened or weakened; a process that is central to the information processing within the brain and which plays a particularly important role in enabling higher cognitive processes [1,2]. Its role in disease is becoming increasingly clear across a wide spectrum of CNS disorders. Thus, for example, dysfunctional synaptic plasticity has been reported in neurodegenerative disorders such as Alzheimer's Disease (AD) as well as in schizophrenia and in a range of disorders associated with learning disabilities . Moreover, maladaptive plasticity processes in response to specific external challenges are believed to underlie disorders such as addiction and post-traumatic stress disorder (PTSD). The molecular basis of normal and disease plasticity is rapidly being unravelled such that synaptic plasticity now provides a unique platform from which to launch the hunt for highly innovative drugs to treat CNS disease by either, firstly, rectifying identifiable abnormalities in these processes, or secondly, utilizing these processes as a vehicle to rectify, or bypass, other mechanisms underlying disease. In this respect, recent advances have been made in studying synaptic plasticity in humans at the molecular through to clinical level and these approaches now provide a real opportunity to test synaptic plasticity as a treatment paradigm for a wide variety of CNS disorders.
Current Opinion in Pharmacology 07/2011; 11(5):540-8. · 4.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Proteinase-activated receptor-2 (PAR2) is widely expressed in the CNS under normal physiological conditions. However, its potential role in modulating neuronal excitability and synaptic transmission remains to be determined. Here, we have investigated whether PAR2 activation modulates synaptic activity in the hippocampus.
PAR2 activation and its effect on the hippocampus were examined in rat primary cultures and acute slices using whole cell patch clamp and standard extracellular recordings, respectively.
PAR2 activation leads to a depolarization of hippocampal neurones and a paradoxical reduction in the occurrence of synaptically driven spontaneous action potentials (APs). PAR2-induced neuronal depolarization was abolished following either the inhibition of astrocytic function or antagonism of ionotropic glutamate receptors whilst the PAR2-induced decrease in AP frequency was also reduced when astrocytic function was inhibited. Furthermore, when examined in acute hippocampal slices, PAR2 activation induced a profound long-term depression of synaptic transmission that was dependent on NMDA receptor activation and was sensitive to disruption of astrocytic function.
These novel findings show that PAR2 activation indirectly inhibits hippocampal synaptic activity and indicate that these receptors may play an active role in modulating normal physiological CNS function, in addition to their role in pathophysiological disorders.
British Journal of Pharmacology 03/2011; 163(5):984-94. · 5.07 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Rett syndrome (RTT), a disorder caused almost exclusively by mutations in the X-linked gene, MECP2, has a phenotype thought to be primarily of neurological origin. Disruption of Mecp2 in mice results in a prominent RTT-like phenotype. One of the consequences of MeCP2 absence in the brain is altered functional and structural plasticity. We aimed to characterize synaptic effects related to plasticity in the hippocampus further and establish whether plasticity defects are amenable to pharmacological reversal. Using male mice in which Mecp2 expression was prevented by a stop cassette, we assessed synaptic plasticity in area CA1 at different phenotypic stages, scoring the mice weekly for overt RTT-like signs. Strongly symptomatic Mecp2(stop/y) mice displayed reduced long-term potentiation (LTP, 40.2±1.6% of wild-type), post-tetanic potentiation (PTP, 45±18.8% of wild-type) and paired-pulse facilitation (PPF, 78±0.1% of wild type) (all P<0.05), the impairment increasing with symptom severity score. These plasticity impairments were absent in presymptomatic mice. Repeated high frequency stimulation revealed pronounced LTP saturation in symptomatic Mecp2(stop/y) mice, suggesting an LTP 'ceiling' effect. Bath application of the weak NMDA receptor blocker memantine (1 μM) resulted in partial restoration of a short-term plasticity component. These data support that idea that progressive functional synaptic impairment is a key feature in the RTT brain and demonstrate the potential for the pharmacological restoration of plasticity function.
[Show abstract][Hide abstract] ABSTRACT: The hippocampus plays an important role in the encoding and retrieval of spatial and non-spatial memories. Much is known about the anatomical, physiological and molecular characteristics as well as the connectivity and synaptic properties of various cell types in the hippocampal circuits , but how these detailed properties of individual neurons give rise to the encoding and retrieval of memories remains unclear. Computational models play an instrumental role in providing clues on how these processes may take place. Here, we present three computational models of the region CA1 of the hippocampus at various levels of detail. Issues such as retrieval of memories as a function of cue loading, presentation frequency and learning paradigm, memory capacity, recall performance, and theta phase precession in the presence of dopamine neuromodulation and various types of inhibitory interneurons are addressed. The models lead to a number of experimentally testable predictions that may lead to a better understanding of the biophysical computations in the hippocampus.
[Show abstract][Hide abstract] ABSTRACT: Mutations in the X-linked MECP2 gene are the primary cause of the severe autism spectrum disorder RTT (Rett syndrome). Deletion of Mecp2 in mice recapitulates many of the overt neurological features seen in humans, and the delayed onset of symptoms is accompanied by deficits in neuronal morphology and synaptic physiology. Recent evidence suggests that reactivation of endogenous Mecp2 in young and adult mice can reverse aspects of RTT-like pathology. In the current perspective, we discuss these findings as well as other genetic, pharmacological and environmental interventions that attempt phenotypic rescue in RTT. We believe these studies provide valuable insights into the tractability of RTT and related conditions and are useful pointers for the development of future therapeutic strategies.
Biochemical Society Transactions 04/2010; 38(2):498-506. · 2.59 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Actin-rich dendritic spines are the locus of excitatory synaptic transmission and plastic events such as long-term potentiation (LTP). Morphological plasticity of spines accompanies activity-dependent changes in synaptic strength. Several Rho GTPase family members are implicated in regulating neuronal and, in particular, spine structure via actin and the actin-binding protein cofilin. However, despite expression in hippocampus and cortex, its ability to modulate actin-regulatory proteins, and its induction during aging, RhoB has been relatively neglected. We previously demonstrated that LTP is associated with specific RhoB activation. Here, we further examined its role in synaptic function using mice with genetic deletion of the RhoB GTPase (RhoB(-/-) mice). Normal basal synaptic transmission accompanied reduced paired-pulse facilitation and post-tetanic potentiation in the hippocampus of RhoB(-/-) mice. Early phase LTP was significantly reduced in RhoB(-/-) animals, whereas the later phase was unaffected. In wild-type mice (RhoB(+/+)), Western blot analysis of potentiated hippocampus showed significant increases in phosphorylated cofilin relative to nonpotentiated slices, which were dramatically impaired in RhoB(-/-) slices. There was also a deficit in phosphorylated Lim kinase levels in the hippocampus from RhoB(-/-) mice. Morphological analysis suggested that lack of RhoB resulted in increased dendritic branching and decreased spine number. Furthermore, an increase in the proportion of stubby relative to thin spines was observed. Moreover, spines demonstrated increased length along with increased head and neck widths. These data implicate RhoB in cofilin regulation and dendritic and spine morphology, highlighting its importance in synaptic plasticity at a structural and functional level.
Journal of Neuroscience 03/2010; 30(9):3508-17. · 6.75 Impact Factor