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The movement of ions in and out of neurons can exert significant effects on neighboring cells. Here we report several experimentally important consequences of activation of the optogenetic chloride pump, halorhodopsin. We recorded extracellular K+concentration ([K+]extra) in neocortical brain slices prepared from young adult mice (both sexes) which express halorhodopsin in pyramidal cells. Strong halorhodopsin activation induced a pronounced drop in [K+]extrathat persisted for the duration of illumination. Pharmacological blockade of K+channels reduced the amplitude of this drop, indicating that it represents K+redistribution into cells during the period of hyperpolarization. Halorhodopsin thus drives the inward movement of both Cl–directly, and K+secondarily. When the illumination period ended, a rebound surge in extracellular [K+] developed over tens of seconds, partly reflecting the previous inward redistribution of K+, but additionally driven by clearance of Cl–coupled to K+by the potassium-chloride cotransporter, KCC2. The drop in [K+]extraduring light activation leads to a small (2-3 mV) hyperpolarization also of other cells that do not express halorhodopsin. Its activation therefore has both direct and indirect inhibitory effects. Finally, we show that persistent strong activation of halorhodopsin causes cortical spreading depolarizations (CSDs), bothin vitroandin vivo. This novel means of triggering CSDs is unusual, in that the events can arise during the actual period of illumination, when neurons are being hyperpolarized and [K+]extrais low. We suggest that this fundamentally different experimental model of CSDs will open up new avenues of research to explain how they occur naturally.SIGNIFICANCE STATEMENTHalorhodopsin is a light-activated electrogenic chloride pump, which has been widely used to inhibit neurons optogenetically. Here, we demonstrate three previously unrecognized consequences of its use: (1) intense activation leads to secondary movement of K+ions into the cells; (2) the resultant drop in extracellular [K+] reduces excitability also in other, nonexpressing cells; and (3) intense persistent halorhodopsin activation can trigger cortical spreading depolarization (CSD). Halorhodopsin-induced CSDs can occur when neurons are hyperpolarized and extracellular [K+] is low. This contrasts with the most widely used experimental models that trigger CSDs with high [K+]. Both models, however, are consistent with the hypothesis that CSDs arise following net inward ionic movement into the principal neuron population.

Epilepsies are multifaceted neurological disorders characterized by abnormal brain activity, e.g., caused by imbalanced synaptic excitation and inhibition. The neural extracellular matrix (ECM) is dynamically modulated by physiological and pathophysiological activity and critically involved in controlling the brain’s excitability. We used different epilepsy models, i.e. mice lacking the presynaptic scaffolding protein Bassoon at excitatory, inhibitory or all synapse types as genetic models for rapidly generalizing early-onset epilepsy, and intra-hippocampal kainate injection, a model for acquired temporal lobe epilepsy, to study the relationship between epileptic seizures and ECM composition. Electroencephalogram recordings revealed Bassoon deletion at excitatory or inhibitory synapses having diverse effects on epilepsy-related phenotypes. While constitutiveBsnmutants and GABAergic neuron-specific knockouts (BsnDlx5/6cKO) displayed severe epilepsy with more and stronger seizures than kainate-injected animals, mutants lacking Bassoon solely in excitatory forebrain neurons (BsnEmx1cKO) showed only mild impairments. By semiquantitative immunoblotting and immunohistochemistry we show model-specific patterns of neural ECM remodeling, and we also demonstrate significant upregulation of the ECM receptor CD44 in null andBsnDlx5/6cKOmutants. ECM-associated WFA-binding chondroitin sulfates were strongly augmented in seizure models. Strikingly, Brevican, Neurocan, Aggrecan and link protein Hapln1 levels reliably predicted seizure properties across models, suggesting a link between ECM state and epileptic phenotype.

Research using membrane capacitance (C

Background Progesterone is essential for a healthy pregnancy. Several small trials have suggested that progesterone therapy may rescue a pregnancy in women with early pregnancy bleeding, which is a symptom that is strongly associated with miscarriage. Objectives (1) To assess the effects of vaginal micronised progesterone in women with vaginal bleeding in the first 12 weeks of pregnancy. (2) To evaluate the cost-effectiveness of progesterone in women with early pregnancy bleeding. Design A multicentre, double-blind, placebo-controlled, randomised trial of progesterone in women with early pregnancy vaginal bleeding. Setting A total of 48 hospitals in the UK. Participants Women aged 16–39 years with early pregnancy bleeding. Interventions Women aged 16–39 years were randomly assigned to receive twice-daily vaginal suppositories containing either 400 mg of progesterone or a matched placebo from presentation to 16 weeks of gestation. Main outcome measures The primary outcome was live birth at ≥ 34 weeks. In addition, a within-trial cost-effectiveness analysis was conducted from an NHS and NHS/Personal Social Services perspective. Results A total of 4153 women from 48 hospitals in the UK received either progesterone (n = 2079) or placebo (n = 2074). The follow-up rate for the primary outcome was 97.2% (4038 out of 4153 participants). The live birth rate was 75% (1513 out of 2025 participants) in the progesterone group and 72% (1459 out of 2013 participants) in the placebo group (relative rate 1.03, 95% confidence interval 1.00 to 1.07; p = 0.08). A significant subgroup effect (interaction test p = 0.007) was identified for prespecified subgroups by the number of previous miscarriages: none (74% in the progesterone group vs. 75% in the placebo group; relative rate 0.99, 95% confidence interval 0.95 to 1.04; p = 0.72); one or two (76% in the progesterone group vs. 72% in the placebo group; relative rate 1.05, 95% confidence interval 1.00 to 1.12; p = 0.07); and three or more (72% in the progesterone group vs. 57% in the placebo group; relative rate 1.28, 95% confidence interval 1.08 to 1.51; p = 0.004). A significant post hoc subgroup effect (interaction test p = 0.01) was identified in the subgroup of participants with early pregnancy bleeding and any number of previous miscarriage(s) (75% in the progesterone group vs. 70% in the placebo group; relative rate 1.09, 95% confidence interval 1.03 to 1.15; p = 0.003). There were no significant differences in the rate of adverse events between the groups. The results of the health economics analysis show that progesterone was more costly than placebo (£7655 vs. £7572), with a mean cost difference of £83 (adjusted mean difference £76, 95% confidence interval –£559 to £711) between the two arms. Thus, the incremental cost-effectiveness ratio of progesterone compared with placebo was estimated as £3305 per additional live birth at ≥ 34 weeks of gestation. Conclusions Progesterone therapy in the first trimester of pregnancy did not result in a significantly higher rate of live births among women with threatened miscarriage overall, but an important subgroup effect was identified. A conclusion on the cost-effectiveness of the PRISM trial would depend on the amount that society is willing to pay to increase the chances of an additional live birth at ≥ 34 weeks. For future work, we plan to conduct an individual participant data meta-analysis using all existing data sets. Trial registration Current Controlled Trials ISRCTN14163439, EudraCT 2014-002348-42 and Integrated Research Application System (IRAS) 158326. Funding This project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 24, No. 33. See the NIHR Journals Library website for further project information.

This technical note introduces adiabatic dynamic causal modelling, a method for inferring slow changes in biophysical parameters that control fluctuations of fast neuronal states. The application domain we have in mind is inferring slow changes in variables (e.g., extracellular ion concentrations or synaptic efficacy) that underlie phase transitions in brain activity (e.g., paroxysmal seizure activity). The scheme is efficient and yet retains a biophysical interpretation, in virtue of being based on established neural mass models that are equipped with a slow dynamic on the parameters (such as synaptic rate constants or effective connectivity). In brief, we use an adiabatic approximation to summarise fast fluctuations in hidden neuronal states (and their expression in sensors) in terms of their second order statistics; namely, their complex cross spectra. This allows one to specify and compare models of slowly changing parameters (using Bayesian model reduction) that generate a sequence of empirical cross spectra of electrophysiological recordings. Crucially, we use the slow fluctuations in the spectral power of neuronal activity as empirical priors on changes in synaptic parameters. This introduces a circular causality, in which synaptic parameters underwrite fast neuronal activity that, in turn, induces activity-dependent plasticity in synaptic parameters. In this foundational paper, we describe the underlying model, establish its face validity using simulations and provide an illustrative application to a chemoconvulsant animal model of seizure activity.

Objective. The development of experimental methodology utilizing graphene micro-transistor arrays to facilitate and advance translational research into cortical spreading depression (CSD) in the awake brain. Approach. CSDs were reliably induced in awake nontransgenic mice using optogenetic methods. High-fidelity DC-coupled electrophysiological mapping of propagating CSDs was obtained using flexible arrays of graphene soultion-gated field-effect transistors (gSGFETs). Main results. Viral vectors targetted channelrhopsin expression in neurons of the motor cortex resulting in a transduction volume 1 mm3. 5-10 s of continous blue light stimulation induced CSD that propagated across the cortex at a velocity of 3.0 0.1 mm min-1. Graphene micro-transistor arrays enabled high-density mapping of infraslow activity correlated with neuronal activity suppression across multiple frequency bands during both CSD initiation and propagation. Localized differences in the CSD waveform could be detected and categorized into distinct clusters demonstrating the spatial resolution advantages of DC-coupled recordings. We exploited the reliable and repeatable induction of CSDs using this preparation to perform proof-of-principle pharmacological interrogation studies using NMDA antagonists. MK801 (3 mg kg-1) suppressed CSD induction and propagation, an effect mirrored, albeit transiently, by ketamine (15 mg kg-1), thus demonstrating this models' applicability as a preclinical drug screening platform. Finally, we report that CSDs could be detected through the skull using graphene micro-transistors, highlighting additional advantages and future applications of this technology. Significance. CSD is thought to contribute to the pathophysiology of several neurological diseases. CSD research will benefit from technological advances that permit high density electrophysiological mapping of the CSD waveform and propagation across the cortex. We report an in vivo assay that permits minimally invasive optogenetic induction, combined with multichannel DC-coupled recordings enabled by gSGFETs in the awake brain. Adoption of this technological approach could facilitate and transform preclinical investigations of CSD in disease relevant models., This work has been funded by the European Union's Horizon 2020 research and innovation programme under Grant Agreement Nos. 785219 and 881603 (Graphene Flagship) and co-funded by the European Regional Development Funds (ERDF) allocated to the Programa operatiu FEDER de Catalunya 2014–2020, with the support of the Secretaria d'Universitats i Recerca of the Departament d'Empresa i Coneixement of the Generalitat de Catalunya for emerging technology clusters devoted to the valorization and transfer of research results (GraphCAT 001-P-001702). RW is funded by a Senior Research Fellowship awarded by the Worshipful Company of Pewterers. EMC was awarded an EMBO Short-Term Fellowship ASTF 8157 to spend time in RW's lab. DR is a Biotechnology and Biological Sciences Research Council (BBSRC) LIDo sponsored PhD student. JAG and EDC acknowledge the Ministerio de Ciencia, Innovación y Universidades, la Agencia Estatal de Investigación (AEI) y el Fondo Europeo de Desarrollo Regional (FEDER/UE) for the FIS2017-85787-R research project. This work has made use of the Spanish ICTS Network MICRONANOFABS partially supported by MICINN and the ICTS 'NANBIOSIS', more specifically by the Micro-NanoTechnology Unit of the CIBER in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN) at the IMB-CNM. Equipment used was partially funded by Fondo Europeo de Desarrollo Regional (FEDER/UE) FICTS14/20-2-23. IMF is funded by the European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No. 665919.

Individuals who have Down syndrome (caused by trisomy of chromosome 21), have a greatly elevated risk of early-onset Alzheimer’s disease, in which amyloid-β accumulates in the brain. Amyloid-β is a product of the chromosome 21 gene APP (amyloid precursor protein) and the extra copy or ‘dose’ of APP is thought to be the cause of this early-onset Alzheimer’s disease. However, other chromosome 21 genes likely modulate disease when in three-copies in people with Down syndrome. Here we show that an extra copy of chromosome 21 genes, other than APP, influences APP/Aβ biology. We crossed Down syndrome mouse models with partial trisomies, to an APP transgenic model and found that extra copies of subgroups of chromosome 21 gene(s) modulate amyloid-β aggregation and APP transgene-associated mortality, independently of changing amyloid precursor protein abundance. Thus, genes on chromosome 21, other than APP, likely modulate Alzheimer’s disease in people who have Down syndrome.

Mapping the entire frequency bandwidth of brain electrophysiological signals is of paramount importance for understanding physiological and pathological states. The ability to record simultaneously DC-shifts, infraslow oscillations (, This work has received funding from the European Union’s Horizon 2020 research and innovation programme under Grant Agreement No. 881603 (GrapheneCore3). ICN2 is supported by the Severo Ochoa Centres of Excellence programme, funded by the Spanish Research Agency (AEI, grant no. SEV-2017–0706), and by the CERCA Programme/Generalitat de Catalunya. A.B.C. is supported by the International PhD Programme La Caixa-Severo Ochoa (Programa Internacional de Becas ‘la Caixa’-Severo Ochoa). This work has made use of the Spanish ICTS Network MICRONANOFABS, partially supported by MICINN and the ICTS ‘NANBIOSIS’, more specifically by the Micro-NanoTechnology Unit of the CIBER in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN) at the IMB-CNM. We also acknowledge funding from the Generalitat de Catalunya (2017 SGR 1426), and the 2DTecBio project (FIS2017-85787-R) funded by the Ministerio de Ciencia, Innovación y Universidades of Spain, the Agencia Estatal de Investigación (AEI) and the Fondo Europeo de Desarrollo Regional (FEDER/UE). Part of this work was co-funded by the European Regional Development Funds (ERDF) allocated to the Programa operatiu FEDER de Catalunya 2014–2020, with the support of the Secretaria d’Universitats i Recerca of the Departament d’Empresa i Coneixement of the Generalitat de Catalunya for emerging technology clusters devoted to the valorization and transfer of research results (GraphCAT 001-P-001702). A.B.C. acknowledges that this work has been carried out within the framework of a PhD in Electrical and Telecommunication Engineering at the Universitat Autònoma de Barcelona. R.C.W. is funded by a Senior Research Fellowship awarded by the Worshipful Company of Pewterers. D.R. is a Biotechnology and Biological Sciences Research Council (BBSRC) LIDo sponsored PhD student. We thank M. Walker and L. Lemieux (UCL Queen Square Institute of Neurology) for their comments on the manuscript.

Objectives To assess the cost‐effectiveness of progesterone compared with placebo in preventing pregnancy loss in women with early pregnancy vaginal bleeding. Design Economic evaluation alongside a large multi‐centre randomised placebo‐controlled trial. Setting Forty‐eight UK NHS early pregnancy units. Population Four thousand one hundred and fifty‐three women aged 16–39 years with bleeding in early pregnancy and ultrasound evidence of an intrauterine sac. Methods An incremental cost‐effectiveness analysis was performed from National Health Service (NHS) and NHS and Personal Social Services perspectives. Subgroup analyses were carried out on women with one or more and three or more previous miscarriages. Main outcome measures Cost per additional live birth at ≥34 weeks of gestation. Results Progesterone intervention led to an effect difference of 0.022 (95% CI −0.004 to 0.050) in the trial. The mean cost per woman in the progesterone group was £76 (95% CI −£559 to £711) more than the mean cost in the placebo group. The incremental cost‐effectiveness ratio for progesterone compared with placebo was £3305 per additional live birth. For women with at least one previous miscarriage, progesterone was more effective than placebo with an effect difference of 0.055 (95% CI 0.014–0.096) and this was associated with a cost saving of £322 (95% CI −£1318 to £673). Conclusions The results suggest that progesterone is associated with a small positive impact and a small additional cost. Both subgroup analyses were more favourable, especially for women who had one or more previous miscarriages. Given available evidence, progesterone is likely to be a cost‐effective intervention, particularly for women with previous miscarriage(s). Tweetable abstract Progesterone treatment is likely to be cost‐effective in women with early pregnancy bleeding and a history of miscarriage., Tweetable abstract Progesterone treatment is likely to be cost‐effective in women with early pregnancy bleeding and a history of miscarriage.

The use of graphene transistors for transducing neural activity has demonstrated the potential to extend the spatiotemporal resolution of electrophysiological methods to lower frequencies, providing a new tool to understand the role of the infra-slow activity.

OBJECTIVE: Loss of function of the astrocyte-specific protein MLC1 leads to the childhood-onset leukodystrophy "megalencephalic leukoencephalopathy with subcortical cysts" (MLC). Studies on isolated cells show a role for MLC1 in astrocyte volume regulation and suggest that disturbed brain ion and water homeostasis is central to the disease. Excitability of neuronal networks is particularly sensitive to ion and water homeostasis. In line with this, reports of seizures and epilepsy in MLC patients exist. However, systematic assessment and mechanistic understanding of seizures in MLC are lacking. METHODS: We analyzed an MLC patient inventory to study occurrence of seizures in MLC. We used two distinct genetic mouse models of MLC to further study epileptiform activity and seizure threshold through wireless extracellular field potential recordings. Whole-cell patch-clamp recordings and K+ -sensitive electrode recordings in mouse brain slices were used to explore the underlying mechanisms of epilepsy in MLC. RESULTS: An early onset of seizures is common in MLC. Similarly, in MLC mice, we uncovered spontaneous epileptiform brain activity and a lowered threshold for induced seizures. At the cellular level, we found that although passive and active properties of individual pyramidal neurons are unchanged, extracellular K+ dynamics and neuronal network activity are abnormal in MLC mice. INTERPRETATION: Disturbed astrocyte regulation of ion and water homeostasis in MLC causes hyperexcitability of neuronal networks and seizures. These findings suggest a role for defective astrocyte volume regulation in epilepsy. Ann Neurol 2018;83:636-649.

Focal epilepsy involves excessive cortical activity that propagates both locally and distally. Does this propagation follow the same routes as normal cortical activity? We pharmacologically induced focal seizures in primary visual cortex (V1) of awake mice, and compared their propagation to the retinotopic organization of V1 and higher visual areas. We used simultaneous local field potential recordings and widefield imaging of a genetically encoded calcium indicator to measure prolonged seizures (ictal events) and brief interictal events. Both types of event are orders of magnitude larger than normal visual responses, and both start as standing waves: synchronous elevated activity in the V1 focus and in homotopic locations in higher areas, i.e. locations with matching retinotopic preference. Following this common beginning, however, seizures persist and propagate both locally and into homotopic distal regions, and eventually invade all of visual cortex and beyond. We conclude that seizure initiation resembles the initiation of interictal events, and seizure propagation respects the connectivity underlying normal visual processing.Focal cortical seizures result from local and widespread propagation of excitatory activity. Here the authors employ widefield calcium imaging in mouse visual areas to demonstrate that these seizures start as local synchronous activation and then propagate along the connectivity that underlies normal sensory processing.

BACKGROUND\ud Bleeding in early pregnancy is strongly associated with pregnancy loss. Progesterone is essential for the maintenance of pregnancy. Several small trials have suggested that progesterone therapy may improve pregnancy outcomes in women who have bleeding in early pregnancy.\ud \ud METHODS\ud We conducted a multicenter, randomized, double-blind, placebo-controlled trial to evaluate progesterone, as compared with placebo, in women with vaginal bleeding in early pregnancy. Women were randomly assigned to receive vaginal suppositories containing either 400 mg of progesterone or matching placebo twice daily, from the time at which they presented with bleeding through 16 weeks of gestation. The primary outcome was the birth of a live-born baby after at least 34 weeks of gestation. The primary analysis was performed in all participants for whom data on the primary outcome were available. A sensitivity analysis of the primary outcome that included all the participants was performed with the use of multiple imputation to account for missing data.\ud \ud RESULTS\ud A total of 4153 women, recruited at 48 hospitals in the United Kingdom, were randomly assigned to receive progesterone (2079 women) or placebo (2074 women). The percentage of women with available data for the primary outcome was 97% (4038 of 4153 women). The incidence of live births after at least 34 weeks of gestation was 75% (1513 of 2025 women) in the progesterone group and 72% (1459 of 2013 women) in the placebo group (relative rate, 1.03; 95% confidence interval [CI], 1.00 to 1.07; P=0.08). The sensitivity analysis, in which missing primary outcome data were imputed, resulted in a similar finding (relative rate, 1.03; 95% CI, 1.00 to 1.07; P=0.08). The incidence of adverse events did not differ significantly between the groups.\ud \ud CONCLUSIONS\ud Among women with bleeding in early pregnancy, progesterone therapy administered during the first trimester did not result in a significantly higher incidence of live births than placebo. (Funded by the United Kingdom National Institute for Health Research Health Technology Assessment program; PRISM Current Controlled Trials number, ISRCTN14163439. opens in new tab.)

Neuroimaging techniques applied to a variety of organisms-from zebrafish, to rodents to humans-can offer valuable insights into neuronal network properties and their dysfunction in epilepsy. A wide range of imaging methods used to monitor neuronal circuits and networks during evoked seizures in animal models and advances in functional magnetic resonance imaging (fMRI) applied to patients with epilepsy were discussed during the XIV Workshop on Neurobiology of Epilepsy (XIV WONOEP) organized in 2017 by the Neurobiology Commission of the International League Against Epilepsy (ILAE). We review the growing number of technological approaches developed, as well as the current state of knowledge gained from studies applying these advanced imaging approaches to epilepsy research.

Refractory focal neocortical epilepsy is a devastating disease for which there is frequently no effective treatment. Gene therapy represents a promising alternative, but treating epilepsy in this way involves irreversible changes to brain tissue, so vector design must be carefully optimized to guarantee safety without compromising efficacy. We set out to develop an epilepsy gene therapy vector optimized for clinical translation. The gene encoding the voltage-gated potassium channel Kv1.1, KCNA1, was codon-optimized for human expression and mutated to accelerate the channels’ recovery from inactivation. For improved safety, this engineered potassium channel (EKC) gene was packaged into a non-integrating lentiviral vector under the control of a cell type-specific CAMK2A promoter. In a blinded, randomized, placebo-controlled pre-clinical trial, the EKC lentivector robustly reduced seizure frequency in a rat model of focal neocortical epilepsy characterized by discrete spontaneous seizures. This demonstration of efficacy in a clinically relevant setting, combined with the improved safety conferred by cell type-specific expression and integration-deficient delivery, identify EKC gene therapy as ready for clinical translation in the treatment of refractory focal epilepsy.

SummaryObjectiveTo characterize a rat model of focal neocortical epilepsy for use in developing novel therapeutic strategies in a type of epilepsy that represents a significant unmet need.MethodsIntracortical tetanus toxin (TeNT) injection was used to induce epilepsy in rats. Seizures and their behavioural manifestations were evaluated with continuous video-electrocorticography telemetry.ResultsTeNT injection into rat primary visual cortex induced focal neocortical epilepsy without preceding status epilepticus. The latency to first seizure ranged from 3 to 7 days. Seizure duration was bimodal, with both short (approximately 30s) and long-lasting (>100s) seizures occurring in the same animals. Seizures were accompanied by non-motor features such as behavioural arrest, or motor seizures with or without evolution to generalized tonic-clonic seizures. Seizures were commoner during the sleep phase of a light-dark cycle. Seizure occurrence was not random, and tended to cluster with significantly higher probability of recurrence within 24 hours of a previous seizure. Across animals, the number of seizures in the first week could be used to predict the number of seizures in the following 22 days.SignificanceThe TeNT model of visual cortical epilepsy is a robust model of acquired focal neocortical epilepsy, and is well suited for preclinical evaluation of novel anti-epileptic strategies. We provide here a detailed analysis of the epilepsy phenotype, seizure activity, electrographic features, and the semiology. In addition we provide a predictive framework that can be used to reduce variation and consequently animal use in pre-clinical studies of potential treatments.Key PointsTetanus toxin injection into rat visual cortex induces focal cortical epilepsy.Electrographic seizures were associated with non-motor and motor features with or without evolution to generalized tonic-clonic seizures.Seizures could not be provoked by intermittent photic stimulation.Seizures were clustered in time and exhibited a circadian variation in frequency.The number of seizures in first week after seizure onset could be used to predict the total number of seizures in the following 3 weeks.

Pharmaceutical treatment can be inadequate, non-effective, or intolerable for many people suffering from a neuronal channelopathy. Development of novel treatment options, particularly those with the potential to be curative is warranted. Gene therapy approaches can permit cell-specific modification of neuronal and circuit excitability and have been investigated experimentally as a therapy for numerous neurological disorders, with clinical trials for several neurodegenerative diseases ongoing. Channelopathies can arise from a wide array of gene mutations; however they usually result in periods of aberrant network excitability. Therefore gene therapy strategies based on up or downregulation of genes that modulate neuronal excitability may be effective therapy for a wide range of neuronal channelopathies. As many channelopathies are paroxysmal in nature, optogenetic or chemogenetic approaches may be well suited to treat the symptoms of these diseases. Recent advances in gene-editing technologies such as the CRISPR-Cas9 system could in the future result in entirely novel treatment for a channelopathy by repairing disease-causing channel mutations at the germline level. As the brain may develop and wire abnormally as a consequence of an inherited or de novo channelopathy, the choice of optimal gene therapy or gene editing strategy will depend on the time of intervention (germline, neonatal or adult). This article is part of the Special Issue entitled 'Channelopathies.'

Tetanus toxin provides a reliable means of inducing recurrent spontaneous epileptic seizures in the hippocampus and neocortex. The hippocampal tetanus toxin model differs from other methods of inducing spontaneous limbic seizures in that it does not require status epilepticus, and it does not have major cell loss as an early component of the syndrome. Indeed, it has proved useful in showing that seizures can occur in the absence of gross pathology, and that long-term behavioral complications of seizures need not depend on neuronal loss, or ongoing epileptic activity. The characteristics of the neocortical models depend upon the area into which the toxin is introduced; interictal activity predominantly follows injection into sensory areas, but epilepsia partialis continua can occur, following injection into motor cortex. The frequent epileptiform abnormalities/seizures in these models make them ideal for testing new therapies. However, variation in the potency of commercially available toxin can prove problematic.

Focal epilepsy involves excessive cortical activity that propagates both locally and distally. Does this propagation follow the same routes as normal cortical activity? We pharmacologically induced focal seizures in primary visual cortex (V1) of awake mice, and compared their propagation to the retinotopic organization of V1 and higher visual areas. We used simultaneous local field potential recordings and widefield imaging of a genetically encoded calcium indicator to measure prolonged seizures (ictal events) and brief interictal events. Both types of event are orders of magnitude larger than normal visual responses, and both start as standing waves: synchronous elevated activity in the V1 focus and in homotopic locations in higher areas, i.e. locations with matching retinotopic preference. Following this common beginning, however, seizures persist and propagate both locally and into homotopic distal regions, and eventually invade all of visual cortex and beyond. We conclude that seizure initiation resembles the initiation of interictal events, and seizure propagation respects the connectivity underlying normal visual processing.Focal cortical seizures result from local and widespread propagation of excitatory activity. Here the authors employ widefield calcium imaging in mouse visual areas to demonstrate that these seizures start as local synchronous activation and then propagate along the connectivity that underlies normal sensory processing.

This article aims to explore what is understood by the term 'research impact' and to provide a comprehensive assimilation of available literature and information, drawing on global experiences to understand the potential for methods and frameworks of impact assessment being implemented for UK impact assessment. We take a more focused look at the impact component of the UK Research Excellence Framework taking place in 2014 and some of the challenges to evaluating impact and the role that systems might play in the future for capturing the links between research and impact and the requirements we have for these systems. Copyright The Author 2013. Published by Oxford University Press., Oxford University Press.

Background and purpose: P2X receptors are widely expressed in cells of the immune system with varying functions. This study sought to characterize P2X receptor expression in the LAD2 human mast cell line and human lung mast cells (HLMCs). Experimental approach: Reverse transcriptase polymerase chain reaction (RT-PCR) and patch clamp studies were used to characterize P2X expression in mast cells using a range of pharmacological tools. Key results: RT-PCR revealed P2X1, P2X4 and P2X7 transcripts in both cell types; mRNA for P2X6 was also detected in LAD2 cells. Under whole-cell patch clamp conditions, rapid application of ATP (1–1000 µM) to cells clamped at −60 mV consistently evoked inward currents in both types of cells. Brief application of ATP (1 s) evoked a rapidly desensitizing P2X1-like current in both cell types. This current was also elicited by αβmethylene ATP (10 µM, 94% cells, n= 31) and was antagonized in LAD2 cells by NF 449 (1 µM) and pyridoxal phosphate-6-azo(benzene-2,4-disulphonic acid) (1–10 µM). A P2X7-like non-desensitizing current in response to high concentrations of ATP (1–5 mM) was also seen in both cell types (96% LAD2, n= 24; 54% HLMCs, n= 24) which was antagonized by AZ11645373 (1 µM). P2X7-like responses were also evoked in LAD2 cells by 2′(3′)-0-(4-benzoylbenzoyl)ATP (300 µM). A P2X4-like current was evoked by 100 µM ATP (80% LAD2, n= 10; 21% HLMCs, n= 29), the amplitude and duration of which was potentiated by ivermectin (3 µM). Conclusion and implications: Our data confirmed the presence of functional P2X1, P2X4 and P2X7 receptors in LAD2 cells and HLMCs.

Background Novel treatments for drug-resistant epilepsy are required. New method Optogenetics is a combination of optical and genetic methods used to control the activity of specific populations of excitable cells using light with high temporal and spatial resolution. Derived from microbial organisms, ‘opsin’ genes encode light-activated ion channels and pumps. Opsins can be genetically targeted to well-defined neuronal populations in mammalian brains using viral vectors. When exposed to light of an appropriate wavelength, the excitability of neurons can be increased or decreased optically on a millisecond timescale. Comparison with existing method(s) Alternative treatments for drug-resistant epilepsy such as vagal, cortical or subcortical stimulation, focal cooling, callosotomy, or ketogenic diet have met with limited success, whereas optogenetic approaches have shown considerable pre-clinical promise. Conclusions Several groups have reported that optogenetic approaches successfully attenuated epileptiform activity in different rodent models of epilepsy, providing proof of the principle that this approach may translate to an effective treatment for epilepsy patients. However, further studies are required to determine the optimal opsin, in which types (or subtypes) of neurons it should be expressed, and what are the most efficient temporal profiles of photostimulation. Although invasive due to the need to inject a viral vector into the brain and implant a device to deliver light to opsin-transduced neurons, this approach has the potential to be effective in suppressing spontaneous seizures while avoiding the side-effects of anti-epileptic drugs (AEDs) or the need to permanently excise regions of the brain. Optogenetic approaches may treat drug-refractory epilepsies.

Repeated, efficient, and extensive use of prototypes is a vital activity that can make the difference between successful and unsuccessful entry of new products into the competitive world market. In this respect, physical prototyping can prove to be very lengthy and expensive, especially if modifications resulting from design reviews involve tool redesign. The availability and affordability of advanced computer technology has paved the way for increasing utilization of prototypes that are digital and created in computer-based environments, i.e. they are virtual as opposed to being physical. The technology for using virtual prototypes was pioneered and adopted initially by large automotive and aerospace industries. Small-to-medium enterprises (SMEs) in the manufacturing industry also need to take virtual prototyping (VP) technology more seriously in order to exploit the benefits. VP is becoming very advanced and may eventually dominate the product development process. However, physical prototypes will still be required for the near future, albeit less frequently. This paper presents a general survey of the available VP techniques and highlights some of the most important developments and research issues while providing sources for further reference. The purpose of the paper is to provide potential SME users with a broad picture of the field of VP and to identify issues and information relevant to the deployment and implementation of VP technology.

Epilepsy represents a major burden to society, not least because approximately 25% of patients do not respond satisfactorily to antiepileptic medication, and only a minority with pharmacoresistant epilepsy are eligible for potentially curative surgery. Several studies have explored gene therapy as a treatment strategy. The translation of scientific breakthroughs into the clinic faces several challenges, including the validation of experimental models of human pharmacoresistant epilepsy, establishment of sensitive and specific measures of therapeutic efficacy, and evaluation of the long-term safety of gene therapy. On the basis of successful reports of gene therapy in experimental models of epilepsy, a roadmap toward clinical trials is proposed.

Two-dimensional ultrasonic measuring systems have found applications in a number of fields including robotics and process control. To provide two-dimensional ranging, an array of receiving and/or transmitting elements are required, which have traditionally required spacing at sub-λ/2 intervals. The λ/2 limitation has been removed by new processing methods but the process still remains computationally intensive. This paper describes a method of reducing the computational effort significantly whilst increasing the accuracy of measurement. In order to optimise the system further, an analysis of the effect of different receiving element distributions within the array on systematic and random errors is also presented.

Beam-forming techniques are widely used to determine 2D and 3D position of targets. One of the main restrictions is the requirement for λ/2 distance between element to avoid ambiguities due to sidelobes. This requirement can be eliminated when short pulses are used and the beam-forming process is applied to the envelope of the received signals only. In airborne ultrasound, the information available from the envelope received signals, is the time of flight. To calculate 3D position of targets, a non-linear equation system has to be solved. The solution to this system can be found by several numerical methods, mainly iterative processes. In this paper, a new 3D ultrasonic 3D target location system is presented which uses the Broyden method in conjunction with coarse beam forming. The operation of the system has been demonstrated using simulated data based on the characteristics of the 2D ultrasonic target location system developed at Nottingham University.

In this article the performance of a new type of lens is compared with traditional meniscus lenses for CO2 laser cutting of medium (>5 mm) section stainless steel. Dual Focus™ lenses produce two focus spots, one above the other. This type of optic was found to be capable of higher cutting speeds and better quality cuts. A phenomenological model is presented which explains the superior performance of Dual Focus lenses. The model concentrates upon the fact that during high speed cutting of medium section metals, the laser does not irradiate the lower portion of the cut zone. This lower part of the cut is heated by the melt flowing over it, which has been previously heated by the laser. This melt preheating involves narrower kerfs and higher cut speeds if a dual focus lens is used.

Gene therapy in human disease has expanded rapidly in recent years with the development of safer and more effective viral vectors, and presents a novel approach to the treatment of epilepsy. Studies in animals models have demonstrated that overexpression of inhibitory peptides can modify seizure threshold, prevent the development of epilepsy, and modify established epilepsy. More recently there has been a flurry of studies using optogenetics in which light-activated channels expressed in neurons can transiently change neuronal excitability on exposure to light, thereby enabling the development of closed loop systems to detect and stop seizure activity. The treatment of status epilepticus presents its own challenges. Because of both the delay in gene expression following transfection and also the necessity of using focal transfection, there are a limited number of situations in which gene therapy can be used in status epilepticus. One such condition is epilepsia partialis continua (EPC). We have used gene therapy in a model of EPC and have shown that we can "cure" the condition. Recent evidence suggesting that gene therapy targeting subcortical regions can modify generalized or more diffuse epilepsies, indicates that the range of situations in status epilepticus in which gene therapy could be used will expand.

Neocortical epilepsy is frequently drug-resistant. Surgery to remove the epileptogenic zone is only feasible in a minority of cases, leaving many patients without an effective treatment. We report the potential efficacy of gene therapy in focal neocortical epilepsy using a rodent model in which epilepsy is induced by tetanus toxin injection in the motor cortex. By applying several complementary methods that use continuous wireless electroencephalographic monitoring to quantify epileptic activity, we observed increases in high frequency activity and in the occurrence of epileptiform events. Pyramidal neurons in the epileptic focus showed enhanced intrinsic excitability consistent with seizure generation. Optogenetic inhibition of a subset of principal neurons transduced with halorhodopsin targeted to the epileptic focus by lentiviral delivery was sufficient to attenuate electroencephalographic seizures. Local lentiviral overexpression of the potassium channel Kv1.1 reduced the intrinsic excitability of transduced pyramidal neurons. Coinjection of this Kv1.1 lentivirus with tetanus toxin fully prevented the occurrence of electroencephalographic seizures. Finally, administration of the Kv1.1 lentivirus to an established epileptic focus progressively suppressed epileptic activity over several weeks without detectable behavioral side effects. Thus, gene therapy in a rodent model can be used to suppress seizures acutely, prevent their occurrence after an epileptogenic stimulus, and successfully treat established focal epilepsy.

Moiré contouring is a technique that enables the surface topography of an object to be measured in a single view and has a wide range of potential applications in manufacturing. However, to be useful, it is necessary to be able to calibrate the system and establish the traceability of the calibration. In this paper, the factors that determine the accuracy of the technique are identified, methods of calibration proposed and attainable accuracies estimated. These include the optical magnification, phase measurement and sensitivity, all of which contribute to measurement errors in the technique. The methods are illustrated with results from a digital moiré system, and the improvements required to obtain maximum accuracy are outlined.

An investigation into methods of producing ultrasonic phased arrays resonating at 100 kHz using capacitive transducers is described here. The backplate texture is one of the most critical parameters determining the performance of these transducers, and transducers arrays were produced using a variety of methods to generate an appropriate backplate texture, including etching, chemical machining, silk-screening, shot-peening and machining. The most successful method involved the use of a proprietary conductive plant, and several arrays with both λ and λ 2 element spacings were produced and tested. The transducers can detect targets at distances of up to 1.8m and two-dimensional range maps were constructed from the received signals using digital beam-forming.

Amyloid-β protein (Aβ) is thought to play a central pathogenic role in Alzheimer's disease. Aβ can impair synaptic transmission, but little is known about the effects of Aβ on intrinsic cellular properties. Here we compared the cellular properties of CA1 hippocampal pyramidal neurons in acute slices from preplaque transgenic (Tg+) CRND8 mice and wild-type (Tg-) littermates. CA1 pyramidal neurons from Tg+ mice had narrower action potentials with faster decays than neurons from Tg- littermates. Action potential-evoked intracellular Ca(2+) transients in the apical dendrite were smaller in Tg+ than in Tg- neurons. Resting calcium concentration was higher in Tg+ than in Tg- neurons. The difference in action potential waveform was eliminated by low concentrations of tetraethylammonium ions and of 4-aminopyridine, implicating a fast delayed-rectifier potassium current. Consistent with this suggestion, there was a small increase in immunoreactivity for Kv3.1b in stratum radiatum in Tg+ mice. These changes in intrinsic properties may affect information flow through the hippocampus and contribute to the behavioral deficits observed in mouse models and patients with early-stage Alzheimer's disease.

An ultrasonic system which can measure range and bearing of targets within an area of 1m2 in a processing time of l0ms with a range resolution of 1.5mm and a bearing resolution of 1° has been developed. The operating range of the system can be increased using lower frequency arrays. The systems have a range of potential applications in guidance of automatic vehicles for collision avoidance, target location and navigation.

The capacitive transducer can be used to transmit ultrasound more effectively into air than the more conventional piezoelectric transducer because of its better impedance matching. The behaviour of such transducers is not, however, well understood. An extensive investigation of the way in which the performance of such transducers is affected by variations in backplate, membrane and drive voltage is reported and significant parameters for the construction of such transducers are defined.

The capacitive ultrasonic transducer is particularly suited to the propagation of ultrasound in air because the impedance mismatch is considerably less than with the piezo-electric transducer. However, its behaviour is very complex and not well understood. This paper describes a detailed investigation of its properties and outlines the progress which has been made in its construction.

We report our experience with the use of Argon laser photocoagulation for the treatment of recurrent trichiasis. A total of 28 lids (of 21 patients) with aberrant lashes were treated in our study. Each lash was treated with a beam of 50-200 micron (spot size), 0.2 s (time), and 1-1.2 Watt (power). Neither infiltration nor topical anaesthesia was used. The mean follow-up time was 6.57 months. Successful treatment with no evidence of recurrence was achieved in 67.9% of lids after one to two laser sessions. The remaining 32.1% of the lids required three to four sessions to achieve no recurrence of the condition. There was a significant correlation between the number of aberrant lashes per lid and the number of required laser sessions. Complete healing of the treated areas occurred within six weeks after treatment, with no vascularisation or distortion of the lid margin. Argon laser treatment appears to be a safe and effective alternative to the other recognised methods of therapy in selected cases.

Capacitive transducers provide a means of coupling ultrasound efficiently into air. Many investigations of these transducers have been reported but none have provided a reliable means of producing a transducer resonating at a specific frequency. The authors describe the performance of capacitive transducers constructed with v-groove backplates and membranes of varying thicknesses, and give design criteria for producing transducers at well defined frequencies.