Spontaneous Events Research Articles

Early onset epilepsy and sudden unexpected death in epilepsy with cardiac arrhythmia in mice carrying the early infantile epileptic encephalopathy 47 gain-of-function FHF1(FGF12) missense mutation.

Fibroblast growth factor homologous factors (FHFs) are brain and cardiac sodium channel-binding proteins that modulate channel density and inactivation gating. A recurrent de novo gain-of-function missense mutation in the FHF1(FGF12) gene (p.Arg52His) is associated with early infantile epileptic encephalopathy 47 (EIEE47; Online Mendelian Inheritance in Man database 617166). To determine whether the FHF1 missense mutation is sufficient to cause EIEE and to establish an animal model for EIEE47, we sought to engineer this mutation into mice. The Arg52His mutation was introduced into fertilized eggs by CRISPR (clustered regularly interspaced short palindromic repeats) editing to generate Fhf1R52H/F+ mice. Spontaneous epileptiform events in Fhf1R52H/+ mice were assessed by cortical electroencephalography (EEG) and video monitoring. Basal heart rhythm and seizure-induced arrhythmia were recorded by electrocardiography. Modulation of cardiac sodium channel inactivation by FHF1BR52H protein was assayed by voltage-clamp recordings of FHF-deficient mouse cardiomyocytes infected with adenoviruses expressing wild-type FHF1B or FHF1BR52H protein. All Fhf1R52H/+ mice experienced seizure or seizurelike episodes with lethal ending between 12 and 26days of age. EEG recordings in 19-20-day-old mice confirmed sudden unexpected death in epilepsy (SUDEP) as severe tonic seizures immediately preceding loss of brain activity and death. Within 2-53s after lethal seizure onset, heart rate abruptly declined from 572±16bpm to 108±15bpm, suggesting a parasympathetic surge accompanying seizures that may have contributed to SUDEP. Although ectopic overexpression of FHF1BR52H in cardiomyocytes induced a 15-mV depolarizing shift in voltage of steady-state sodium channel inactivation and slowed the rate of channel inactivation, heart rhythm was normal in Fhf1R52H/+ mice prior to seizure. The Fhf1 missense mutation p.Arg52His induces epileptic encephalopathy with full penetrance in mice. Both Fhf1 (p.Arg52His) and Scn8a (p.Asn1768Asp) missense mutations enhance sodium channel Nav 1.6 currents and induce SUDEP with bradycardia in mice, suggesting an FHF1/Nav 1.6 functional axis underlying altered brain sodium channel gating in epileptic encephalopathy.

Design and Demonstration In Vitro of a Mouse-Specific Transcranial Magnetic Stimulation Coil

Transcranial magnetic stimulation (TMS) is a technique used to treat different neurological disorders non-invasively. A pulsed current to a coil generates a magnetic field ( $B$ -field) which induces an electric field ( $E$ -field). Underlying biophysical effects of TMS are unclear. Therefore, animal experiments are needed; however, making small TMS coils suitable for mice is difficult and their field strengths are typically much lower than for human sized coils. Objectives/Hypothesis . We aimed to design and demonstrate a mouse-specific coil that can generate high and focused $E$ -field. Methods . We designed a tapered TMS coil of 50 turns of 0.2 mm diameter copper wire around a 5 mm diameter tapered powdered iron core and discharged a $220~\mu \text{F}$ capacitor at 50 V through it. We measured $B$ -field with a Hall probe and induced $E$ -field with a wire loop. We measured temperature rise with a thermocouple. We applied 1200 pulses of continuous theta burst stimulation (cTBS) and intermittent theta burst stimulation (iTBS) to mouse brain slices and analyzed how spontaneous electrical activity changed. Results . The coil gave maximum $B$ -field of 685 mT at the base of the coil and 340 mT at 2 mm below the coil, and maximum $E$ -field 2 mm below the coil of approximately 10 V/m, at 50 V power supply, with a temperature increase of 20 °C after 1200 pulses of cTBS. We observed no changes in $B$ -field with heating. cTBS reduced frequency of spontaneous population events in mouse brain slices up to 20 min after stimulation and iTBS increased frequency up to 20 min after stimulation. No frequency changes occurred after 20 min. No changes in amplitude of spontaneous events were found. Conclusion . The design generated fields strong enough to modulate brain activity in vitro .

Homeostatic Depression Shows Heightened Sensitivity to Synaptic Calcium.

Synapses and circuits rely on homeostatic forms of regulation in order to transmit meaningful information. The Drosophila melanogaster neuromuscular junction (NMJ) is a well-studied synapse that shows robust homeostatic control of function. Most prior studies of homeostatic plasticity at the NMJ have centered on presynaptic homeostatic potentiation (PHP). PHP happens when postsynaptic muscle neurotransmitter receptors are impaired, triggering retrograde signaling that causes an increase in presynaptic neurotransmitter release. As a result, normal levels of evoked excitation are maintained. The counterpart to PHP at the NMJ is presynaptic homeostatic depression (PHD). Overexpression of the Drosophila vesicular glutamate transporter (VGlut) causes an increase in the amplitude of spontaneous events. PHD happens when the synapse responds to the challenge by decreasing quantal content (QC) during evoked neurotransmissionagain, resulting in normal levels of postsynaptic excitation. We hypothesized that there may exist a class of molecules that affects both PHP and PHD. Impairment of any such molecule could hurt a synapses ability to respond to any significant homeostatic challenge. We conducted an electrophysiology-based screen for blocks of PHD. We did not observe a block of PHD in the genetic conditions screened, but we found loss-of-function conditions that led to a substantial deficit in evoked amplitude when combined with VGlut overexpression. The conditions causing this phenotype included a double heterozygous loss-of-function condition for genes encoding the inositol trisphosphate receptor (IP3R itpr) and ryanodine receptor (RyR). IP3Rs and RyRs gate calcium release from intracellular stores. Pharmacological agents targeting IP3R and RyR recapitulated the genetic losses of these factors, as did lowering calcium levels from other sources. Our data are consistent with the idea that the homeostatic signaling process underlying PHD is especially sensitive to levels of calcium at the presynapse.

Relationship between polypharmacy and adverse events

A retrospective data analysis was performed to investigate the association between polypharmacy and adverse events using three different spontaneous adverse event reporting system databases. Multivariate logistic regression analyses were performed to investigate the association between the number of drugs and adverse events, including hepatic disorders, renal disorders, hypersensitivity, and extrapyramidal syndrome. The results showed that the risk of hepatic and renal disorders increased with the number of drugs. Thus, decreasing the number of drugs may reduce the risk of hepatic and renal disorders. Furthermore, attention should be given to specific drugs that may cause hypersensitivity and extrapyramidal syndrome.

Hyperexcitability of Nucleus Tractus Solitarii Neurons in the Streptozotocin‐Induced Model of Alzheimer's Disease

Besides the stereotypical problems of memory and cognition in patients with Alzheimer's Disease (AD), another highly prevalent symptom is of respiratory origin. We have repeatedly shown blunted respiratory responses to hypoxia in the streptozotocin (STZ) -induced AD model, which is similar to sleep disordered breathing found in nearly 80% of AD patients. Our current study focuses on the electrophysiological changes within the nucleus tractus solitarii (nTS), a brainstem area known to control respiratory function. Using male Sprague-Dawley rats (n = 15 rats, 240±8 g), we induced AD symptoms with microinjections of 2 mg/kg Streptozotocin (STZ) into the lateral cerebral ventricles. Control animals received 0.9 mM citrate buffer as vehicle control (CTL). The disease model was verified using the Morris water maze to test for memory dysfunction. We recorded nTS neuronal responses via whole cell patch clamp. Action potential (AP) discharge upon current-stimulation was higher in STZ-AD when compared to control. Also, spontaneous spike activity (no stimulation) was elevated. These data corroborate earlier results using a higher (2x1.5 mg) STZ dosage. Spontaneous post-synaptic currents in the absence of stimulation resemble nTS network activity. The number of spontaneous events in nTS neurons of the STZ-AD group was surprisingly similar to CTL in the current study. On the other hand, network activity mirrors the hyperexcitability seen from our current injection when using the higher STZ dosage. While there was no appreciable change in resting membrane potential, AP threshold of STZ-AD nTS cells shifted towards a more negative membrane potential, overall contributing to increased excitability in these cells. AP phase plane plots in STZ-AD had a significantly smaller overshoot with a reduced velocity of up and down stroke, possibly indicating altered function of voltage-gated sodium channels. Analysis of transient K+ currents (A-type) revealed a slight shift to higher activation voltages (from -20 mV [CTL] to 0 mV [STZ-AD]) in STZ-AD. This shift favors AP generation and may contribute to a lower spike threshold in STZ-AD. There were no changes in the steady state K+ currents between groups. In summary, nTS cells of the STZ-AD group showed increased excitability that may involve gating properties of voltage-gated sodium channels and A-type potassium channels. The observed neurophysiological changes in the nTS may be part of the neuronal correlate for increased breathing frequency and blunted (ceiling effect) respiratory responses to hypoxia in the STZ-AD model.

Role of flow reversals in transition to turbulence and relaminarization of pulsatile flows

Abstract

Induction of somatic mutations by low concentrations of tritiated water (HTO): evidence for the possible existence of a dose-rate threshold.

Tritium is a low energy beta emitter and is discharged into the aquatic environment primarily in the form of tritiated water (HTO) from nuclear power plants or from nuclear fuel reprocessing plants. Although the biological effects of HTO exposures at significant doses or dose rates have been extensively studied, there are few reports concerning the biological effects of HTO exposures at very low dose rates. In the present study using a hyper-sensitive assay system, we investigated the dose rate effect of HTO on the induction of mutations. Confluent cell populations were exposed to HTO for a total dose of 0.2 Gy at dose rates between 4.9 mGy/day and 192 mGy/day by incubating cells in medium containing HTO. HTO-induced mutant frequencies and mutation spectra were then investigated. A significant inflection point for both the mutant frequency and mutation spectra was found between 11 mGy/day and 21.6 mGy/day. Mutation spectra analysis revealed that a mechanistic change in the nature of the mutation events occurred around 11 mGy/day. The present observations and published experimental results from oral administrations of HTO to mice suggest that a threshold dose-rate for HTO exposures might exist between 11 mGy/day and 21.6 mGy/day where the nature of the mutation events induced by HTO becomes similar to those seen in spontaneous events.

Patterns of use, effectiveness and safety of gadolinium contrast agents: a European prospective cross-sectional multicentre observational study

BackgroundThe EU gadolinium-based contrast agents (GBCA) market has changed in recent years due to the European Medicines Agency decision to suspend the marketing authorisation of linear GBCA and the marketing authorisation of new generic macrocyclic GBCA. The study aims to understand the patterns of (GBCA) use, and to study the effectiveness and safety of GBCA in routine practice across Europe.MethodsProspective, cross-sectional, multicentre, observational study in patients undergoing contrast-enhanced magnetic resonance. Reported usage patterns included indication, referral and examination details. Assessment of effectiveness included changes in radiological diagnosis, diagnostic confidence and image quality. Safety data were collected by spontaneous patient adverse event (AE) reporting.Results2118 patients were included from 8 centres across 5 European countries between December 2018 and November 2019. Clariscan, Dotarem (gadoteric acid), Gadovist (gadobutrol) and ProHance (gadoteridol) were utilised in 1513 (71.4%), 356 (16.8%), 237 (11.2%) and 12 (0.6%) patients, respectively. Most were performed in CNS-related indications (46.2%). Mean GBCA doses were 0.10 mmol/kg body weight, except for Gadovist (mean 0.12 mmol/kg). GBCA use increased confidence in diagnosis in 96.2% of examinations and resulted in a change in radiological diagnosis in 73.9% of patients. Image quality was considered excellent or good in 96.1% of patients and across all GBCA. Four patients reported AEs (0.19%), with only 1 (0.05%) considered serious.ConclusionsThis European study confirmed that GBCAs are used appropriately in Europe for a wide range of indications. The study demonstrated a significant increase in diagnostic confidence after GBCA use and confirmed the good safety profile of GBCAs, with comparable results for all agents used.

2. Establishing Preventive Strategies Against the Side Effects of Anticancer Drugs Using the Medical Large Databases of Spontaneous Adverse Event Reports

2. Establishing Preventive Strategies Against the Side Effects of Anticancer Drugs Using the Medical Large Databases of Spontaneous Adverse Event Reports

Effect of unplanned spontaneous follicular growth and ovulation on pregnancy outcomes in planned artificial frozen embryo transfer cycles: a propensity score matching study.

Does unplanned spontaneous follicular growth and ovulation affect clinical outcomes after planned artificial frozen-thawed embryo transfer (AC-FET) cycles? AC-FET and spontaneous follicular growth and ovulation events resulted in notably better pregnancy outcomes with a significantly higher implantation rate (IR), clinical pregnancy rate (CPR), ongoing pregnancy rate (OPR) and live birth rate (LBR) and a significantly lower miscarriage rate. The AC-FET protocol without GnRH agonist administration is associated with a low incidence of follicular growth and ovulation. In the literature, authors often refer to these types of cycles with concern due to possibly impaired FET outcomes. However, the real impact of such cycles has yet to be elucidated due to the lack of existing data. This was a retrospective clinical study involving 2256 AC-FET cycles conducted between January 2017 and August 2019. Propensity score (PS) matching was used to control for confounding variables. Subjects were divided into two groups: a study group: cycles with spontaneous follicular growth and ovulation (the maximum diameter of follicles in any ovary was ≥14mm and ovulation was confirmed by consecutive ultrasound examinations) and a control group featuring cycles without growing follicles (the maximum diameter of follicles in both ovaries were <10mm). The study group was matched by PS with the control group at a ratio of 1:2. The study group consisted of 195 patients before PS matching and 176 patients after matching. The numbers of participants in the control group before and after PS matching were 2061 and 329, respectively. This analysis showed that patient age (adjusted odds ratio [aOR] 1.05; 95% CI 1.01-1.09; P=0.010) and basal FSH level (aOR 1.06; 95% CI 1.01-1.11; P=0.012) were significantly and positively related with the spontaneous follicular growth and ovulation event. In addition, this event was negatively correlated with BMI (aOR 0.92; 95% CI 0.87-0.97; P=0.002), AMH level (aOR 0.66; 95% CI 0.59-0.74; P<0.001) and a high starting oestrogen dose (aOR 0.53; 95% CI 0.38-0.76 for 6mg vs. 4mg; P<0.001). Baseline characteristics were similar between groups after PS matching. Patients in the study group had a significantly higher IR (28.8% vs. 21.8%, P=0.016), CPR (44.9% vs. 33.4%, P=0.011), OPR (39.2% vs. 26.1%, P=0.002) and LBR (39.2% vs. 24.9%, P=0.001) and a lower miscarriage rate (12.7% vs. 25.5%, P=0.030), compared with those in the control group. This was a retrospective study carried out in a single centre and was therefore susceptible to bias. In addition, we only analysed patients with normal ovulation patterns and excluded those with follicular growth but without ovulation. Further studies remain necessary to confirm our results. It is not necessary to cancel cycles that experience spontaneous follicular growth and ovulation. Our data support promising clinical outcomes after this event. Our findings are important as they can better inform clinicians and patients. This research was supported by National Natural Science Foundation of China (grant no. 81701507, 81801404, 81871210, 82071648), Natural Science Foundation of Jiangsu Province (grant no. BK20171126, BK20201123) and Jiangsu Province '333' project. The authors declare that they have no competing interests. N/A.

Resolving Energy Losses Caused by End-Users in Electrical Grid Systems

This study utilises the Pareto approach to highlight the energy losses that mainly originate from the phenomena of tiny, initiated events created by end-users of electricity in Australia. Simulation modelling was applied through two stages to examine residential households’ electricity consumption behaviour in New South Wales, Australia. Stage one analysis applied Hierarchical agglomerative clustering and a dendrogram to denote the respective Euclidean distance between the different clusters. Heat maps and threshold value area charts were used to compare the mean power demand for six respective clusters. Stage two used ‘sensitivity analysis’ to investigate how uncertainty in the electricity demand can be allocated to the uncertainty of energy losses. The findings envision practical solutions to dealing with the variability of energy losses and the proposal to set new demand-side strategies associated with individuals. Retail prices of electricity in Australia have risen by roughly 60% since 2007. The research contributes to knowledge about the roots of energy losses in Australia, creating a $210M cost value. Energy losses are of significant economic value, while also impacting energy security. The first limitation of this study is using approaches from complexity theory to grasp the philosophical issues behind the research design and clarifying which insights suit what kind of evidence, thus identifying the data that needed to be collected. The second limitation is that this study’s methodology used a mostly quantitative approach that describes and explains a complex phenomenon in depth more than exploring and confirming that phenomenon. The third and final limitation is that this study’s context is also limited regarding selected sample criteria. The context is limited to a particular demographic area in New South Wales (NSW) in Australia and is also limited to residential houses (not industrial or commercial), which was opposed by data availability and access. The research draws on ‘peak and off-peak’ scales of electricity demand cause energy losses. The research shows the role of the phenomena of spontaneous emergence as a non-linked constraint which is the main issue that splits the optimal solution into pieces and significantly complicates the solution task. Demand side management (DSM) of electricity can be improved from this to construct new demand-side strategies. The study is structured around understanding the consequences of the scalability of events and the clustering dynamic of non-linearity through relevance complexity concepts exclusive to spontaneous emergence (SE), power laws (PLs), Paretian approach (PA), and tiny initiated events (TIEs). We examined the issues of the spontaneous emergence of non-linear, dynamic behaviour involved in the electricity demand of end-users on the basis of pushing individual systems of end-users to the edge of self-organised criticality (SOC). Revising the demand system’s complexity has value in constituting a core domain of interest in what is new in the field of demand side management (DSM), thus contributing to understanding end-users’ behaviour-driven energy losses from both theoretical and empirical perspectives.

Remote and Long-Term Self-Monitoring of Electroencephalographic and Noninvasive Measurable Variables at Home in Patients With Epilepsy (EEG@HOME): Protocol for an Observational Study.

BackgroundEpileptic seizures are spontaneous events that severely affect the lives of patients due to their recurrence and unpredictability. The integration of new wearable and mobile technologies to collect electroencephalographic (EEG) and extracerebral signals in a portable system might be the solution to prospectively identify times of seizure occurrence or propensity. The performances of several seizure detection devices have been assessed by validated studies, and patient perspectives on wearables have been explored to better match their needs. Despite this, there is a major gap in the literature on long-term, real-life acceptability and performance of mobile technology essential to managing chronic disorders such as epilepsy.ObjectiveEEG@HOME is an observational, nonrandomized, noninterventional study that aims to develop a new feasible procedure that allows people with epilepsy to independently, continuously, and safely acquire noninvasive variables at home. The data collected will be analyzed to develop a general model to predict periods of increased seizure risk.MethodsA total of 12 adults with a diagnosis of pharmaco-resistant epilepsy and at least 20 seizures per year will be recruited at King’s College Hospital, London. Participants will be asked to self-apply an easy and portable EEG recording system (ANT Neuro) to record scalp EEG at home twice daily. From each serial EEG recording, brain network ictogenicity (BNI), a new biomarker of the propensity of the brain to develop seizures, will be extracted. A noninvasive wrist-worn device (Fitbit Charge 3; Fitbit Inc) will be used to collect non-EEG biosignals (heart rate, sleep quality index, and steps), and a smartphone app (Seer app; Seer Medical) will be used to collect data related to seizure occurrence, medication taken, sleep quality, stress, and mood. All data will be collected continuously for 6 months. Standardized questionnaires (the Post-Study System Usability Questionnaire and System Usability Scale) will be completed to assess the acceptability and feasibility of the procedure. BNI, continuous wrist-worn sensor biosignals, and electronic survey data will be correlated with seizure occurrence as reported in the diary to investigate their potential values as biomarkers of seizure risk.ResultsThe EEG@HOME project received funding from Epilepsy Research UK in 2018 and was approved by the Bromley Research Ethics Committee in March 2020. The first participants were enrolled in October 2020, and we expect to publish the first results by the end of 2022.ConclusionsWith the EEG@HOME study, we aim to take advantage of new advances in remote monitoring technology, including self-applied EEG, to investigate the feasibility of long-term disease self-monitoring. Further, we hope our study will bring new insights into noninvasively collected personalized risk factors of seizure occurrence and seizure propensity that may help to mitigate one of the most difficult aspects of refractory epilepsy: the unpredictability of seizure occurrence.International Registered Report Identifier (IRRID)PRR1-10.2196/25309

Neuroleptic malignant syndrome associated with long-acting injectable versus oral second-generation antipsychotics: Analyses based on a spontaneous reporting system database in Japan

Long-acting injectable antipsychotics (LAI-APs) remain underutilized. One reason is the concern that LAI-APs might cause serious adverse events such as neuroleptic malignant syndrome (NMS) and lead to prolonged symptoms compared with oral treatment. Because the risk of NMS associated with LAI second-generation antipsychotics (LAI-SGAs) remains unclear, we compared reporting frequency, time to onset, and mortality of NMS between LAI- and oral SGAs using data from a Japanese spontaneous adverse event reporting database between April 2004 and September 2019. Of 5791 patients reporting adverse events due to LAI-SGAs or the equivalent oral SGAs, 768 (13%) developed NMS. LAI aripiprazole and LAI paliperidone were associated with a significantly lower reporting frequency of NMS than the equivalent oral SGAs (adjusted reporting odds ratio [95% confidence interval]: 0.35 [0.19–0.63] and 0.40 [0.27–0.59], respectively). Between 42% and 62% of the NMS associated with LAI- and oral SGAs other than LAI risperidone occurred within 30 days after initiation. The proportion of mortality due to NMS associated with oral aripiprazole was 13.1% and no deaths occurred in patients with NMS associated with LAI aripiprazole. The proportions of mortality due to NMS associated with oral risperidone/paliperidone, LAI risperidone, and LAI paliperidone were 8.8%, 4.2%, and 3.4%, respectively. Our findings showed that LAI-SGAs were not associated with a higher reporting frequency and mortality of NMS compared with oral SGAs, although clinicians need to closely monitor the occurrence of NMS not only during oral SGA treatment, but also, and in particular, in the early stage of LAI-SGA treatment.

Genomic and molecular evolutionary dynamics of transcriptional response regulator genes in bacterial species of the Harveyi clade of Vibrio.

Transcriptional response regulators (TRR) are the most abundant signal transducers in prokaryotic systems that mediate intracellular changes in response to environmental signals. They are involved in a wide range of biological processes that allow bacteria to persist in particular habitats. There is strong evidence that the bacterial habitat and their lifestyle influence the size of their TRR genetic repertoire. Therefore, it would be expected that the evolution of bacterial genomes could be linked to natural selection processes. To test this hypothesis, we explored the evolutionary dynamics of TRR genes of the widely studied Harveyi clade of the genus Vibrio at the molecular and genomic levels. Our results suggest that the TRR genetic repertoire of the species belonging to the Harveyi clade is a product of genomic reduction and expansion. The gene loss and gains that drive their genomic reduction and expansion could be attributed to natural selection and random genetic drift. It seems that natural selection acts to maintain the ancestral state of core TRR genes (shared by all species) by purifying processes and could be driving the loss of some accessory (found in certain species) genes through the diversification of sequences. The neutrality observed in gene gain could be attributed to spontaneous events as horizontal gene transfer driven by stochastic events as occurs in random genetic drift.

The Accessory Helix of Complexin Stabilizes a Partially Unzippered State of the SNARE Complex and Mediates the Complexin Clamping Function In Vivo.

Spontaneous synaptic transmission is regulated by the protein complexin (Cpx). Cpx binds the SNARE complex, a coil-coiled four-helical bundle that mediates the attachment of a synaptic vesicle (SV) to the presynaptic membrane (PM). Cpx is thought to clamp spontaneous fusion events by stabilizing a partially unraveled state of the SNARE bundle; however, the molecular detail of this mechanism is still debated. We combined electrophysiology, molecular modeling, and site-directed mutagenesis in Drosophila to develop and validate the atomic model of the Cpx-mediated clamped state of the SNARE complex. We took advantage of botulinum neurotoxins (BoNTs) B and G, which cleave the SNARE protein synaptobrevin (Syb) at different sites. Monitoring synaptic depression on BoNT loading revealed that the clamped state of the SNARE complex has two or three unraveled helical turns of Syb. Site-directed mutagenesis showed that the Cpx clamping function is predominantly maintained by its accessory helix (AH), while molecular modeling suggested that the Cpx AH interacts with the unraveled C terminus of Syb and the SV lipid bilayer. The developed molecular model was employed to design new Cpx poor-clamp and super-clamp mutations and to tested the predictions in silico employing molecular dynamics simulations. Subsequently, we generated Drosophila lines harboring these mutations and confirmed the poor-clamp and super-clamp phenotypes in vivo. Altogether, these results validate the atomic model of the Cpx-mediated fusion clamp, wherein the Cpx AH inserts between the SNARE bundle and the SV lipid bilayer, simultaneously binding the unraveled C terminus of Syb and preventing full SNARE assembly.

Astrocytic mitochondria in adult mouse brain slices show spontaneous calcium influx events with unique properties

Astrocytes govern critical aspects of brain function via spontaneous calcium signals in their soma and processes. A significant proportion of these spontaneous astrocytic calcium events are associated with mitochondria, however, the extent, sources, or kinetics of astrocytic mitochondrial calcium influx have not been studied in the adult mouse brain. To measure calcium influx into astrocytic mitochondria in situ, we generated an adeno-associated virus (AAV) with the astrocyte-specific GfaABC1D promoter driving expression of the genetically encoded calcium indicator, GCaMP6f tagged to mito7, a mitochondrial matrix targeted signal sequence. Using this construct, we observed AAV-mediated expression of GCaMP6f in adult mouse astrocytic mitochondria that co-localized with MitoTracker deep red (MTDR) in the dorsolateral striatum (DLS) and in the hippocampal stratum radiatum (HPC). Astrocytic mitochondria co-labeled with MTDR and GCaMP6f displayed robust, spontaneous calcium influx events in situ, with subcellular differences in calcium influx kinetics between somatic, branch, and branchlet mitochondria, and inter-regional differences between mitochondria in DLS and HPC astrocytes. Calcium influx into astrocytic mitochondria was strongly dependent on endoplasmic reticulum calcium stores, but did not require the mitochondrial calcium uniporter, MCU. Exposure to either glutamate, D1 or D2 dopamine receptor agonists increased calcium influx in some mitochondria, while simultaneously decreasing calcium influx in other mitochondria from the same astrocyte. These findings show that astrocytic mitochondria possess unique properties with regard to their subcellular morphology, mechanisms of calcium influx, and responses to neurotransmitter receptor agonists. Our results have important implications for understanding the role of astrocytic mitochondria during pathological processes.

Autonomous nervous system regulation of pain in children with cerebral palsy

ABSTRACT Aim: Children with cerebral palsy (CP) have increased pain sensitivity and recurrent pain episodes; however, pain is underreported in children with intellectual impairment. Cardiac autonomic regulation is imbalanced in chronic pain conditions and neurological disorders. This study aims at exploring the autonomous nervous system regulation of pain in children with CP compared with typically developing peers (TDP).Method: Heart rate variability (HRV) was recorded during 24 hours in 26 children with CP and 26 TDP, and examined offline at baseline (sleeping, seated rest) and during spontaneous pain events. Pain and fatigue, HRV indices (linear indices on time – IBI, SDNN, RMSSD – and frequency domains – high, low, and very low frequency – and non-linear indices – Hurst coefficient and multiscale entropy) were computed.Results: Children with CP showed comparable HRV during daily conditions and similar reductions after pain events than their TDP, regardless of their level of intellectual impairment. Interpretation: Children with CP have an intact autonomic regulation in acute pain events. HRV could be an accurate pain biomarker in children with CP and intellectual disability. What this paper adds: Autonomic regulation in acute pain is efficient in children with cerebral palsy. Heart rate variability indices can be reliable pain biomarkers in intellectual impairment.

Correlations Between SGLT-2 Inhibitors and Acute Renal Failure by Signal Detection Using FAERS: Stratified Analysis for Reporting Country and Concomitant Drugs.

Previous studies have shown conflicting observations regarding the correlation between sodium-glucose-cotransporter-2 inhibitors (SGLT2i) and acute renal failure. Although wide use has contributed to the accumulation of safety information on SGLT2i, the examination of the countries reporting cases of SGLT2i use and influence of concomitant drugs has been insufficient in studies using spontaneous adverse event reporting databases. We aimed to re-examine the correlation between SGLT2i and acute renal failure using the latest United States Food and Drug Administration's Adverse Event Reporting System (FAERS) records and to conduct a stratified analysis for the reporting countries (Japan or other countries), as well as the concomitant use of drugs such as angiotensin-converting enzyme inhibitors (ACEis) and angiotensin II receptor blockers (ARBs) with SGLT2i. The reporting odds ratio (ROR) and 95% confidence interval (CI) for cases recorded on FAERS from January 2013 to March 2020 were calculated. We then limited the cases to patients using SGLT2i and receiving treatment for diabetes mellitusand then calculated the ROR. A stratified analysis was performed for reporting countries (Japan or other countries), and the presence or absence of concomitant use of an angiotensin-converting enzyme inhibitor (ACEi) or angiotensin II receptor blocker (ARB) to examine their influence on the correlation between SGLT2i and acute renal failure. Of the 5,337,069 cases of adverse events recorded on FAERS, 410,569 were cases in which patients had received treatment for diabetes. The ROR for SGLT2i calculated from the total analysis subjects was 4.16 (95% CI 4.01-4.31), suggesting its correlation with acute renal failure. Similar results were obtained for the cases in which patients had received treatment for diabetes. However, the stratified analysis of these diabetes-treatment cases for reporting countries showed no correlation between SGLT2i and acute renal failure in cases reported in Japan with ROR 0.58 (95% CI 0.49-0.69). In contrast, a correlation was suggested in cases reported in countries other than Japan with ROR 1.91 (95% CI 1.83-1.98). Moreover, the stratified analysis for the concomitant use of an ACEi or ARB showed that the ROR tended to be low in the cases with one of these drugs. Examination with the signal detection method using FAERS suggested the correlation between SGLT2i and the onset of acute renal failure. However, when focusing on the cases reported in Japan, such a correlation was not suggested. In addition, this study indicated that the signal of acute renal failure tends to be reduced in cases with the concomitant use of either an ACEi or ARB. Through this study we suggest that patients should be closely monitored when they take SGLT2i without an ACEi or ARB.

Hemodialysis-related low thrombogenicity measured by total thrombus-formation analysis system in patients undergoing percutaneous coronary intervention.

BackgroundEstablished antithrombotic therapies can increase bleeding risk, especially in hemodialysis (HD) patients. The Total Thrombus-formation Analysis System (T-TAS) is useful for evaluating thrombogenicity. The aim of this study was to examine the relationship between HD and thrombogenicity, or bleeding events in patients undergoing percutaneous coronary intervention (PCI). MethodsIn this retrospective cohort study, 300 patients undergoing elective PCI were enrolled between April 2017 and March 2019. Blood samples obtained on the day of PCI were analyzed with T-TAS to compute the thrombus formation area under the curve (AUC; PL18-AUC10 for platelet chip; AR10-AUC30 for atheroma chip). The patients were divided into three groups according to estimated glomerular filtration rate (eGFR): 33 HD patients, 124 non-HD patients with eGFR <60 mL/min/1.73m2, and 143 non-HD patients with eGFR ≥60. We examined the thrombogenicity and spontaneous bleeding events within 1-year post-PCI. ResultsHD was significantly associated with both low PL18-AUC10 and AR10-AUC30 levels determined by T-TAS. Bleeding events defined by the Bleeding Academic Research Consortium criteria types 2, 3, or 5 occurred during follow-up in 27 patients (9.0%): 7 in HD, 10 in non-HD with eGFR <60, and 10 in non-HD with eGFR ≥60. Both T-TAS parameters in the patients with bleeding were lower compared with those in the patients without bleeding, and HD was significantly associated with 1-year bleeding events. ConclusionsThe results suggested that HD patients undergoing PCI might be a predictor for low thrombogenicity measured by T-TAS and 1-year bleeding risk.

Impact of double heterozygosity for Factor V Leiden and Prothrombin G20210A on the thrombotic phenotype

IntroductionBecause of the rarity of double heterozygosity for Factor V Leiden (FVL) and Prothrombin (FII) G20210A, little is known about the thrombotic phenotype in double heterozygotes. Material and methodsIn a retrospective cohort study of patients referred for a thrombophilia work-up, we investigated whether double heterozygotes (n = 138) exhibit a more severe thrombotic phenotype compared with single FVL or FIIG20210A heterozygotes, single FVL homozygotes, or wildtype carriers. ResultsThe risk of venous thromboembolism (VTE) was higher for female but not male double heterozygotes compared with single heterozygotes (FVL: 2.51, 95%CI 1.55–4.08, FIIG20210A: 1.75, 95%CI 1.14–2.68) and wildtype carriers (HR 2.53, 95%CI 1.58–4.05) but not compared with FVL homozygotes (HR 1.31, 95%CI 0.94–1.83). Female double heterozygotes developed VTE nearly a decade earlier than wildtype carriers and FVL heterozygotes (mean 44.2 vs. 52.6 and 52.2 years), most often in association with oral contraceptives. Spontaneous VTE and arterial thromboembolic events were not more frequent in double heterozygotes compared with the other genotype groups.Deep vein thrombosis (DVT) of the lower limb was the predominant VTE location in double heterozygotes, atypical vein thrombosis was rare. A phenomenon that has been described as the FVL paradox, a higher proportion of isolated DVT than pulmonary embolism, was also found for double heterozygotes. ConclusionThe thrombotic phenotype in double heterozygotes resembles the appearance of the thrombotic phenotype in FVL carriers but the thrombotic risk is aggravated by women-specific risk factors.