The Role of NMDA Receptor Subtypes in Short-Term Plasticity in the Rat Entorhinal Cortex

We have previously shown that spontaneous release of glutamate in the entorhinal cortex (EC) is tonically facilitated via activation of presynaptic NMDA receptors (NMDAr) containing the NR2B subunit. Here we show that the same receptors mediate short-term plasticity manifested by frequency-dependent facilitation of evoked glutamate release at these synapses. Whole-cell patch-clamp recordings were made from layer V pyramidal neurones in rat EC slices. Evoked excitatory postsynaptic currents showed strong facilitation at relatively low frequencies (3 Hz) of activation. Facilitation was abolished by an NR2B-selective blocker (Ro 25-6981), but unaffected by NR2A-selective antagonists (Zn2+, NVP-AAM077). In contrast, postsynaptic NMDAr-mediated responses could be reduced by subunit-selective concentrations of all three antagonists. The data suggest that NMDAr involved in presynaptic plasticity in layer V are exclusively NR1/NR2B diheteromers, whilst postsynaptically they are probably a mixture of NR1/NR2A, NR1/NR2B diheteromers and NR1/NR2A/NR2B triheteromeric receptors

Convergent metabotropic signalling pathways inhibit SK channels to promote synaptic plasticity in the hippocampus

Hebbian synaptic plasticity at hippocampal Schaffer collateral synapses is tightly regulated by postsynaptic SK channels that restrict NMDA receptor activity. SK channels are themselves modulated by G-protein-coupled signalling pathways, but it is not clear under what conditions these are activated to enable synaptic plasticity. Here, we show that muscarinic M1 receptor (M1R) and type 1 metabotropic glutamate receptor (mGluR1) signalling pathways, which are known to inhibit SK channels and thereby disinhibit NMDA receptors, converge to facilitate spine calcium transients during the induction of long-term potentiation (LTP) at hippocampal Schaffer collateral synapses onto CA1 pyramidal neurons of male rats. Furthermore, mGluR1 activation is required for LTP induced by reactivated place cell firing patterns that occur in sharp wave ripple events during rest or sleep. In contrast, M1R activation is required for LTP induced by place cell firing patterns during exploration. Thus, we describe a common mechanism that enables synaptic plasticity during both encoding and consolidation of memories within hippocampal circuits

Cortactin regulates endo-lysosomal sorting of AMPARs via direct interaction with GluA2 subunit

Abstract AMPA receptor (AMPAR) trafficking is a key determinant of synaptic strength and synaptic plasticity. Under basal conditions, constitutive trafficking maintains surface AMPARs by internalization into the endosomal system, where the majority are sorted and targeted for recycling back to the plasma membrane. NMDA receptor (NMDAR)-dependent Long-Term Depression (LTD) is characterised by a reduction in synaptic strength, and involves endosomal sorting of AMPARs away from recycling pathways to lysosomes. The mechanisms that determine whether AMPARs are trafficked to lysosomes or to recycling endosomes, especially in response to NMDAR stimulation, are unclear. Here, we define a role for the actin-regulatory protein cortactin as a mediator of AMPAR endosomal sorting by direct interaction with the GluA2 subunit. Disrupting GluA2-cortactin binding in neurons causes the targeting of GluA2/A3-containing receptors to lysosomes and their consequent degradation, resulting in a loss of surface and synaptic GluA2 under basal conditions and an occlusion of subsequent LTD expression. Furthermore, we show that NMDAR stimulation causes a dissociation of endogenous cortactin from GluA2 via tyrosine phosphorylation of cortactin. These results demonstrate that cortactin maintains GluA2/A3 levels by directing receptors away from lysosomes, and that disrupting GluA2-cortactin interactions to target GluA2/A3 to lysosomes is an essential component of LTD expression

Cortactin regulates endo-lysosomal sorting of AMPARs via direct interaction with GluA2 subunit

Abstract AMPA receptor (AMPAR) trafficking is a key determinant of synaptic strength and synaptic plasticity. Under basal conditions, constitutive trafficking maintains surface AMPARs by internalization into the endosomal system, where the majority are sorted and targeted for recycling back to the plasma membrane. NMDA receptor (NMDAR)-dependent Long-Term Depression (LTD) is characterised by a reduction in synaptic strength, and involves endosomal sorting of AMPARs away from recycling pathways to lysosomes. The mechanisms that determine whether AMPARs are trafficked to lysosomes or to recycling endosomes, especially in response to NMDAR stimulation, are unclear. Here, we define a role for the actin-regulatory protein cortactin as a mediator of AMPAR endosomal sorting by direct interaction with the GluA2 subunit. Disrupting GluA2-cortactin binding in neurons causes the targeting of GluA2/A3-containing receptors to lysosomes and their consequent degradation, resulting in a loss of surface and synaptic GluA2 under basal conditions and an occlusion of subsequent LTD expression. Furthermore, we show that NMDAR stimulation causes a dissociation of endogenous cortactin from GluA2 via tyrosine phosphorylation of cortactin. These results demonstrate that cortactin maintains GluA2/A3 levels by directing receptors away from lysosomes, and that disrupting GluA2-cortactin interactions to target GluA2/A3 to lysosomes is an essential component of LTD expression

Opportunities and challenges of delivering digital clinical trials: lessons learned from a randomised controlled trial of an online behavioural intervention for children and young people

Background: Despite being the gold standard of research to determine effectiveness, randomised controlled trials (RCTs) often struggle with participant recruitment, engagement and retention. These issues may be exacerbated when recruiting vulnerable populations, such as participants with mental health issues. We aimed to update understanding of the scope of these problems in trials of health technology, and identify possible solutions through reflecting on experiences from an exemplar trial (Online Remote Behavioural Intervention for Tics; ORBIT).Method: We extracted anonymised data on recruitment, retention and requests for more funding and time from trials funded by the largest funder of health technology trials in the UK (the National Institute of Health Research Health Technology Assessment) between 2010-2020, and compared these with data from a recent, successful trial (ORBIT). ORBIT aimed to assess the clinical- and cost-effectiveness of blended online and human behavioural therapy for tics in young people. Many of the trial procedures, including recruitment, the intervention and data collection, were undertaken online. Results: Data were extracted on 51 trials conducted between 2010 and 2020. 60% of trials failed to reach their original recruitment target and only 44% achieved their follow-up in the specified time frame. In contrast, ORBIT recruited to target and achieved 90% follow up. We posit that these achievements are related to a) judicious use of digital technology for trial procedures and b) adequate numbers of highly trained and motivated trial staff. We provide details of both these to help other research teams plan and cost for successful trials. Conclusion: An approach combining human and online methods may be advantageous in facilitating trial delivery, particularly in paediatric mental health services. Given the importance of successful clinical trials in advancing healthcare delivery and the waste of human and economic resources associated with unsuccessfully delivered trials, it is imperative that trials are appropriately costed and future research focusses on improving trial design and delivery. Trial registration: The ORBIT Trial is registered with ISRTCN (ISRCTN70758207) and clinicaltrials.gov (NCT03483493)

The Blood of Healthy Individuals Exhibits CD8 T Cells with a Highly Altered TCR Vb Repertoire but with an Unmodified Phenotype

CD8 T cell clonal expansions (TCE) have been observed in elderly, healthy individuals as well in old mice, and have been associated with the ageing process. Both chronic latent and non-persistent viral infections have been proposed to drive the development of distinct non-functional and functional TCE respectively. Biases in TCR Vβ repertoire diversity are also recurrently observed in patients that have undergone strong immune challenge, and are preferentially observed in the CD8 compartment. Healthy adults can also exhibit CD8 T cells with strong alterations of their CDR3 length distribution. Surprisingly, no specific investigations have been conducted to analyze the CD8 T cell repertoire in normal adults, to determine if such alterations in TCR Vβ repertoire share the features of TCE. In this study, we characterized the phenotype and function of the CD8 population in healthy individuals of 25–52 years of age. All but one of the EBV-positive HLA-B8 healthy volunteers that were studied were CMV-negative. Using a specific unsupervised statistical method, we identified Vβ families with altered CDR3 length distribution and increased TCR Vβ/HPRT transcript ratios in all individuals tested. The increase in TCR Vβ/HPRT transcript ratio was more frequently associated with an increase in the percentage of the corresponding Vβ+ T cells than with an absence of modification of their percentage. However, in contrast with the previously described TCE, these CD8+ T cells were not preferentially found in the memory CD8 subset, they exhibited normal effector functions (cytokine secretion and cytotoxic molecule expression) and they were not reactive to a pool of EBV/CMV/Flu virus peptides. Taken together, the combined analysis of transcripts and proteins of the TCR Vβ repertoire led to the identification of different types of CD8+ T cell clone expansion or contraction in healthy individuals, a situation that appears more complex than previously described in aged individuals

Investigating a therapist-guided, parent-assisted remote digital behavioural intervention for tics in children and adolescents: 'Online Remote Behavioural Intervention for Tics' (ORBIT) trial: protocol of an internal pilot study and single randomised controlled trial

IntroductionTourette syndrome and chronic tic disorder are common, disabling childhood-onset conditions. Guidelines recommend that behavioural therapy should be offered as first-line treatment for children with tics. However, there are very few trained behaviour therapists for tics and many patients cannot access appropriate care. This trial investigates whether an internet-delivered intervention for tics can reduce severity of symptoms.Method and analysisThis parallel-group, single-blind, randomised controlled superiority trial with an internal pilot will recruit children and young people (aged 9-17 years) with tic disorders. Participants will be randomised to receive 10-weeks of either online, remotely-delivered, therapist-supported exposure response prevention (ERP) behavioural therapy for tics, or online, remotely delivered, therapist-supported education about tics and co-occurring conditions. Participants will be followed-up mid-treatment, and 3-, 6-, 12-, and 18-month post-randomisation.The primary outcome is reduction in tic severity as measured on the Yale Global Tic SeverityScale (YGTSS) total tic severity score. Secondary outcomes include a cost-effectiveness analysis and estimate of the longer-term impact on patient outcomes and healthcare services.An integrated process evaluation will analyse quantitative and qualitative data in order to fully explore the implementation of the intervention and identify barriers and facilitators to implementation. The trial is funded by the National Institute of Health Research (NIHR),Health Technology Assessment (16/19/02).Ethics and disseminationThe findings from the study will inform clinicians, healthcare providers and policy makers about the clinical and cost-effectiveness of an internet delivered treatment for children and young people with tics. The results will be submitted for publication in peer-reviewed journals. The study has received ethical approval from North West Greater Manchester Research Ethics Committee (Ref: 18/NW/0079).Trial registration: ISRCTN70758207 and ClinicalTrials.gov (NCT03483493)

Effects of antiplatelet therapy on stroke risk by brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases: subgroup analyses of the RESTART randomised, open-label trial

Background Findings from the RESTART trial suggest that starting antiplatelet therapy might reduce the risk of recurrent symptomatic intracerebral haemorrhage compared with avoiding antiplatelet therapy. Brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases (such as cerebral microbleeds) are associated with greater risks of recurrent intracerebral haemorrhage. We did subgroup analyses of the RESTART trial to explore whether these brain imaging features modify the effects of antiplatelet therapy

Genetic mechanisms of critical illness in COVID-19.

Host-mediated lung inflammation is present1, and drives mortality2, in the critical illness caused by coronavirus disease 2019 (COVID-19). Host genetic variants associated with critical illness may identify mechanistic targets for therapeutic development3. Here we report the results of the GenOMICC (Genetics Of Mortality In Critical Care) genome-wide association study in 2,244 critically ill patients with COVID-19 from 208 UK intensive care units. We have identified and replicated the following new genome-wide significant associations: on chromosome 12q24.13 (rs10735079, P = 1.65 × 10-8) in a gene cluster that encodes antiviral restriction enzyme activators (OAS1, OAS2 and OAS3); on chromosome 19p13.2 (rs74956615, P = 2.3 × 10-8) near the gene that encodes tyrosine kinase 2 (TYK2); on chromosome 19p13.3 (rs2109069, P = 3.98 ×  10-12) within the gene that encodes dipeptidyl peptidase 9 (DPP9); and on chromosome 21q22.1 (rs2236757, P = 4.99 × 10-8) in the interferon receptor gene IFNAR2. We identified potential targets for repurposing of licensed medications: using Mendelian randomization, we found evidence that low expression of IFNAR2, or high expression of TYK2, are associated with life-threatening disease; and transcriptome-wide association in lung tissue revealed that high expression of the monocyte-macrophage chemotactic receptor CCR2 is associated with severe COVID-19. Our results identify robust genetic signals relating to key host antiviral defence mechanisms and mediators of inflammatory organ damage in COVID-19. Both mechanisms may be amenable to targeted treatment with existing drugs. However, large-scale randomized clinical trials will be essential before any change to clinical practice

BHPR research: qualitative1. Complex reasoning determines patients' perception of outcome following foot surgery in rheumatoid arhtritis

Background: Foot surgery is common in patients with RA but research into surgical outcomes is limited and conceptually flawed as current outcome measures lack face validity: to date no one has asked patients what is important to them. This study aimed to determine which factors are important to patients when evaluating the success of foot surgery in RA Methods: Semi structured interviews of RA patients who had undergone foot surgery were conducted and transcribed verbatim. Thematic analysis of interviews was conducted to explore issues that were important to patients. Results: 11 RA patients (9 ♂, mean age 59, dis dur = 22yrs, mean of 3 yrs post op) with mixed experiences of foot surgery were interviewed. Patients interpreted outcome in respect to a multitude of factors, frequently positive change in one aspect contrasted with negative opinions about another. Overall, four major themes emerged. Function: Functional ability & participation in valued activities were very important to patients. Walking ability was a key concern but patients interpreted levels of activity in light of other aspects of their disease, reflecting on change in functional ability more than overall level. Positive feelings of improved mobility were often moderated by negative self perception ("I mean, I still walk like a waddling duck”). Appearance: Appearance was important to almost all patients but perhaps the most complex theme of all. Physical appearance, foot shape, and footwear were closely interlinked, yet patients saw these as distinct separate concepts. Patients need to legitimize these feelings was clear and they frequently entered into a defensive repertoire ("it's not cosmetic surgery; it's something that's more important than that, you know?”). Clinician opinion: Surgeons' post operative evaluation of the procedure was very influential. The impact of this appraisal continued to affect patients' lasting impression irrespective of how the outcome compared to their initial goals ("when he'd done it ... he said that hasn't worked as good as he'd wanted to ... but the pain has gone”). Pain: Whilst pain was important to almost all patients, it appeared to be less important than the other themes. Pain was predominately raised when it influenced other themes, such as function; many still felt the need to legitimize their foot pain in order for health professionals to take it seriously ("in the end I went to my GP because it had happened a few times and I went to an orthopaedic surgeon who was quite dismissive of it, it was like what are you complaining about”). Conclusions: Patients interpret the outcome of foot surgery using a multitude of interrelated factors, particularly functional ability, appearance and surgeons' appraisal of the procedure. While pain was often noted, this appeared less important than other factors in the overall outcome of the surgery. Future research into foot surgery should incorporate the complexity of how patients determine their outcome Disclosure statement: All authors have declared no conflicts of interes