NIHRDataInsights/HRCSData huggingface dataset:
Title: Investigating a neuronal subcellular transcriptome by the novel technique of RNA TU-tagging, in a normal and ALS-related mouse model.
Funder: Medical Research Council

Abstract: Coding and non-coding RNAs are transported to different subcellular localisations for local translation and regulation. Localised 'RNA metabolism' is important in polarised cells such as neurons, but techniques to study RNA localisation are not able to isolate subcellular transcriptomes in vivo. Motor neurons (MNs) have cell bodies in the spinal cord, numerous arborizing dendrites and they connect to muscle fibres at the neuromuscular junction (NMJ) through an axon that can exceed 1 meter in length. We are interested in MNs because of our research into Amyotrophic Lateral Sclerosis (ALS), a fatal neurodegenerative disorder that is characterized by the progressive loss of motor neurons that 'die back' from NMJs. ALS-causative mutations in RNA-binding and -transport genes (TARDBP 'TDP-43', FUS), and in C9ORF72 that induce RNA foci - have highlighted aberrant subcellular RNA metabolism in ALS pathogenesis. Thus there is a clear need to identify the neuronal subcellular transcriptome in vivo to understand normal function, and dysfunction in our paradigm disease, ALS. We will use a new technique, 'TU-tagging' (already successful in mouse) to identify the subcellular transcriptome of three compartments of adult motor neurons in normal mouse and in a mouse with a mutation in Tdp-43. Our preliminary data from a Tdp-43 mutant we are characterising shows disruption of many downstream genes including b-synuclein and tau. TU-tagging works through the cell-specific expression of an enzyme (UPRT) and treatment with a drug (4TU), allowing incorporation of thio-uridine into RNA in specific cell types. Thio-uridine RNA is biotinylated and then pulled-down. We will use TU-tagging to identify RNA from (1) MN cell bodies and dendrites, (2) MN axons, (3) NMJs. This will allow us insight in the biology of MNs, axons and NMJs and their susceptibility in disease. TU-tagging is applicable to all models of normal function and disease, and of neuronal injury.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Amyotrophic Lateral Sclerosis and the DNA Damage Response
Funder: Medical Research Council

Abstract: Amyotrophic Lateral Sclerosis (ALS) is caused by premature/accelerated degeneration of motor neurones. Whereas some cases of ALS are sporadic and of unknown molecular cause, others are due to hereditary dominant mutations in one of approximately eight genes. It has emerged recently that several of the proteins encoded by these genes are involved in regulating mRNA transcription, splicing, and stability, including Fused-in-Sarcoma/Translocated-in-Sarcoma (FUS/TLS) and TAR DNA-binding protein 43 (TDP-43). This raises the possibility that, in some cases of ALS, neuronal cell death may involve loss-of-function defects in RNA processing. However, how and why defects in RNA processing might result in the neurodegenerative pathology that typifies ALS is unclear. We now present preliminary evidence suggesting that FUS and TDP-43 are components of the cellular response to oxidative DNA damage, a process known to be important to slow or prevent neurodegeneration. We show that the FUS/TLS and TDP-43 proteins are redistributed in response to DNA damage, with FUS rapidly accumulating at sites of DNA damage and TDP-43 rapidly expelled. We show that this redistribution is an active process that is regulated by established DNA damage signalling proteins, supporting the idea that RNA processing is a bona fide component of the cellular DNA damage response. Based on these observations, we propose that RNA processing by FUS/TLS and TDP-43 is required to ensure that the stability of nascent pre-mRNA and the progression of RNA polymerase is properly managed and controlled during transcription in the presence of DNA lesions, and that mutation of these and likely other RNA processing factors results in dysfunctional gene expression at sites of DNA damage and consequently progressive neuronal cell death. We now plan to address this hypothesis directly, using a combination of molecular and cellular approaches.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Structural Basis of Sigma-1 Receptor Ligand Interactions and Signalling
Funder: Medical Research Council

Abstract: The Sigma-1 Receptor (S1R) is a ligand-regulated protein chaperone found at the mitochondria associated ER membrane. Upon activation, S1R binds to the IP3 receptor to regulate inter-organelle calcium signalling. Further activation results in re-localisation of S1R to the plasma membrane where it regulates voltage-gated ion channels. S1R is highly expressed in tissues of the central nervous system, and S1R dysfunction is implicated in pain, amnesia, schizophrenia, depression, stroke, Alzheimer's disease, and addiction. S1R activity is regulated by a large number of drugs, but little is known about the molecular basis of its ligand binding or signalling. No bacterial homologs of S1R have been discovered, and the lack of molecular-level information about S1R function arises largely from the difficulties in producing and stabilising eukaryotic membrane proteins. We have developed a multi-pronged approach to studying S1R structure and interactions in which information from solution NMR studies of bacterially expressed S1R constructs is combined with biophysical and in cell fluorescence studies of mammalian expressed S1R. Details of residues and regions of S1R responsible for ligand and cholesterol binding will be determined by solution NMR on S1R truncation mutants. This information will be validated and extended by studies of full length S1R that is expressed into mammalian cell membranes.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: NEUROLOGICAL
Title: Novel signalling pathways regulating nociception
Funder: Medical Research Council

Abstract: Although effective treatments for some types of pain are available, they are non-ideal: drugs targeting the CNS are often associated with issues such as addiction and tolerance. Those currently available which target peripheral sensory neurones can lead to complete sensory loss, and can also affect motor neurones. We suggest exploring alternative targets within peripheral nociceptive pathways which could normalize rather than block afferent sensory activity. Our programme of study will therefore focus on a novel paradigm (discovered by the applicants) concerning regulation of two specific ion channels expressed in sensory neurons, the M-type K+ channel and the T-type Ca2+ channel. Both regulate sensory neuron excitability by setting the resting membrane potential and firing threshold without directly driving action potential propagation. Our current research indicates that both these channels are physiologically modulated by endogenous reactive oxygen species and gasotransmitters, particularly carbon monoxide (CO). We will build upon substantial pilot studies to characterise the intracellular signalling network through which M- and T-type channels are regulated by gasotransmitters and ROS, and to identify physiological triggers through which such pathways can control channel activity. This will be achieved using a multidisciplinary approach involving patch-clamp electrophysiology and confocal imaging, supported by genetic and pharmacological interventions. Finally we shall investigate the contribution of such regulation to the control of afferent sensory activity both in vitro and in vivo using established behavioural tests and models of acute and chronic pain. Our results will provide a novel approach to pain management in which sensory nerve activity can be normalised rather than fully inhibited, thus providing pain relief without sensory blockade.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;5.1 PHARMACEUTICALS
Health categories: NEUROLOGICAL
Title: Bayesian models of expression in the transcriptome for clinical RNA-seq
Funder: Medical Research Council

Abstract: Background -- RNA-Seq technology is enabling investigation of gene expression at the transcript level, including the identification of alternatively spliced isoforms In Motor Neurone Disease, alternative splicing has been strongly implicated as a pathogenic mechanism. RNA-Seq for clinical data and MND in particular requires the development of new statistical methodologies to tackle challenges specific to such data. Bayesian statistical methods for RNA-seq are desirable to deal with the uncertainty in quantifying transcript expression, but existing approaches are prohibitively slow for big data. Aims & Objectives -- 1) To develop practical algorithms for transcript quantification from RNA-Seq in the Bayesian statistical framework. 2) To build statistical models *around* the transcript quantification problem, addressing problems specific to clinical data. 3) To use the developed algorithms to investigate the effects of splicing in Motor Neurone Disease Methodology -- The Bayesian statistical framework will be the cornerstone of the project. Whilst Bayesian methods are often computationally demanding, I shall make use of approximate posterior inference. I'll build on recent work in this area to make fast algorithms for the analysis of RNA-Seq data. I'll collaborate closely with clinical and wet-lab staff in the SITraN neuroscience facility, giving my work immediate impact on research into MND. Scientific opportunities -- The quantification of transcripts in RNA-Seq bears a close resemblance to Latent Dirichlet Allocation (LDA), a statistical model used for the analysis of text corpora. Investigation of this link will enable the transfer of knowledge from this field to enable statistical advances for processing RNA-Seq.


Research activity codes: 1.4 METHODOLOGIES AND MEASUREMENTS
Health categories: NEUROLOGICAL
Title: Abeta-mediated toxicity in Alzheimer's disease: delineating mechanisms of internalisation, cell-cell transmission and synaptic dysfunction
Funder: Medical Research Council

Abstract: Alzheimer's disease is characterised by the abnormal association of Abeta (Ab) peptide to form oligomeric species, with catastrophic consequences for neuronal function and survival. However, clear understanding of the mechanisms by which Ab exerts toxicity remains elusive. Specifically, it is not known how Ab is transmitted around neural networks, how it internalises/accumulates in neurons, the nature of its cellular/molecular targets, or the mechanisms which underlie its disruption of critical neuronal synaptic processes. This proposal will exploit established and novel biochemical, imaging and ultrastructural approaches in cultured and native hippocampal neurons to examine these issues, providing fundamental new insights into the toxic mechanisms of Ab action. A key aim is to translate our recent findings from neuroblastoma cells, defining a pathway of Ab internalisation, into primary neurons. Ab will be tracked alongside cellular/organelle markers to examine endocytic processes and targeting to specific compartments, and followed by ultrastructural investigation. Ab transmission around neuronal networks will be assayed with genetically-encoded fusion constructs providing insight into the spreading of Ab pathology in Alzheimer's patients. We will also examine Ab consequences on presynaptic function - specifically dynamic properties of vesicle pools. Our pilot data indicates striking Ab-induced effects on functional pool partitioning, lateral vesicle trafficking and vesicle endocytic efficiency. Alongside culture experiments we will use an established transgenic model to explore synapse-specific presynaptic deficits in acute slice. This approach also permits a function-ultrastructure readout of recycling vesicles in native tissue, offering unique insights into Ab-induced vesicle pool re-organisation. Our findings will shed new light on pivotal steps in the pathway of Ab action, providing important potential targets for the rational development of therapeutics.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Novel MRI Techniques for Brain Banking and Motor Neuron Disease Research
Funder: Medical Research Council

Abstract: Magnetic resonance imaging (MRI) has enormous potential as a biomarker in neurodegenerative disease, in particular techniques that are sensitive to microstructure show great promise for phenotyping and monitoring disease progression. However, these measures currently lack specificity. We propose to combine post-mortem MRI with histopathology in the same tissue to aid in the interpretation of in-vivo MRI measures. We will focus on motor neuron disease (MND), an area of expertise in Oxford for which MRI has considerable potential as a biomarker. MRI can provide various types of "contrast" (analogous to different tissue stains) with complementary information about anatomy and tissue composition. We will acquire images with a broad range of contrasts, with particular focus on diffusion MRI, a powerful method that presents particular challenges in post-mortem tissue. We will build on our recent advances in MRI software and cutting-edge hardware to improve signal strength by a factor of 5-6 and enable unprecedented spatial detail. We will also explore recent MRI techniques that aim to provide more biologically-meaningful information. We will conduct a proof-of-concept study in MND, which provides an ideal testbed for these methods as a relatively "clean" pathology in which pre- and post-mortem MRI scans are available. A range of MRI scans will be acquired in MND and control brains, the latter being a crucial resource for future expansion into other diseases and conditions. We will demonstrate the potential of MRI-to-histology comparisons in neurodegenerative disease. Specifically, we will investigate whether histological staining supports the use of MRI as an early marker of MND and the related condition frontotemporal dementia. Finally, we will develop a basic prototype database for on-line data exploration and distribution, linked in to the Oxford Brain Bank and freely accessible to the neuroscience community.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: NEUROLOGICAL
Title: A magnetoencephalographic study into the pathophysiological substrates of cognitive and motor symptoms and treatment effects in parkinsonian syndromes
Funder: Medical Research Council

Abstract: Here I will test two hypotheses. First, that different clinical impairments in patients with Parkinson's Disease (PD) relate to distinct basal ganglia-cortical circuits identifiable through their preferred frequency of interaction and their topography. Second, that Deep Brain Stimulation (DBS) improves these impairments by modulating coupling across these circuits. To this end I will record local field potentials (LFPs) from electrodes implanted in the subthalamic nucleus (STN) of post-surgical PD patients whilst simultaneously performing magnetoencephalography (MEG). Using source reconstruction techniques I will reconstruct electrical activity from cortical areas and compute spectral coherence with electrical activity from the STN. I propose two experiments. In the first I will employ the Posner task in order to determine whether the previously defined alpha frequency STN-cortical network is involved in orienting attention to visuo-spatial cues and is dependent on dopamine. In the second experiment I will perform DBS at therapeutically effective and non-effective frequencies in order to determine specifically how STN-cortical networks are influenced by DBS. In order to facilitate this, we have developed a special amplifier that allows simultaneous stimulation and recording of LFP signals. Furthermore, we will also rely on a number of existing engineering methods to remove the stimulation artifact from the MEG channels prior to performing source reconstruction. Finally, by correlating motor and cognitive parameters with patterns of STN-cortical coupling I will gain valuable insight into: 1) the involvement of specific STN-cortical loops in mediating the symptoms of PD and 2) the therapeutic mechanisms of DBS.


Research activity codes: 5.6 PSYCHOLOGICAL AND BEHAVIOURAL;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: How do nitrergic interneurons control corticostriatal communication?
Funder: Medical Research Council

Abstract: The striatum is the main input nucleus of the basal ganglia and it is crucially involved in motor control and action selection. A massive glutamatergic input is sent from most areas of the cortex to the striatum. Corticostriatal synapses are of exceptional importance, as they are the cellular substrate of reward-mediated learning and display a form of long-term synaptic plasticity that requires not only the co-activation of pre- and postsynaptic neurons, but also release of dopamine. Corticostriatal communication is also controlled by striatal interneurons. In particular, a major role is likely to be played by the striatal interneurons that express nitric oxide synthase (NOS) and produce nitric oxide (NO). Long-term plasticity induced by tetanic stimulation of corticostriatal fibres can only be expressed if the NO signalling cascade is activated. Nevertheless, the nitrergic control of corticostriatal communication is still poorly understood. Previous experiments have used exogenous ligands to interfere with the NO signalling cascade. However, understanding the role played by endogenous NO in corticostriatal communication requires selective and controlled activation of NOS interneurons. To achieve this, we will use paired recordings and optogenetic activation of nitrergic interneurons, in combination with activation of corticostriatal fibres, using transgenic animals that express either green fluorescent protein or channelrodopsin in NOS interneurons. These experiments will reveal how endogenous NO controls corticostriatal communication on a short- and long-term basis. This will substantially improve our understanding of how the striatum processes cortical information in the normal brain and will suggest new pharmacological approaches for the treatment of diseases in which corticostriatal communication is impaired, including drug addiction and Parkinson's disease.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: NEUROLOGICAL
Title: Brain mechanisms underlying reading improvement in post-stroke central alexia
Funder: Medical Research Council

Abstract: We will examine therapy-induced changes in the brain networks supporting reading in patients being treated for central alexia. Our current network model of reading in controls and patients with pure alexia, will be developed to account for central alexia using data acquired from magnetoencephalography (MEG). We will test how well the effects of behavioural therapy, on performance and neural mechanisms, are influenced by tDCS, using a randomized, double-blind, crossover design. Therapeutic intervention will involve an adaptation of a computer-based, cross-modal reading program 'iReadMore' that has already been shown to improve reading speeds in patients with pure alexia. There are theoretical and practical reasons to support the use of this therapy in the larger population of patients with central alexia. If 'iReadMore' proves effective, we will make it available for use via a "web app", meeting an important, widespread, unmet clinical need.


Research activity codes: 5.6 PSYCHOLOGICAL AND BEHAVIOURAL
Health categories: NEUROLOGICAL
Title: Regulation of microglial proliferation and its contribution to chronic neurodegeneration
Funder: Medical Research Council

Abstract: A growing body of evidence suggests that systemic inflammation may contribute to the progression of chronic neurodegeneration and the exacerbation of symptoms We have proposed that signals generated during neurodegeneration prime microglia and exaggerate their response to systemic inflammation. Here we will characterise and target one of the most prominent innate immune activities: the control of microglial proliferation and priming. We hypothesise that priming and proliferation of macrophages and microglia in the brain are intimately related. We will use the delivery of novel RGB viral vectors (collaborators in Hamburg) to the bone marrow using X-ray guidance (collaborators in Southampton), to identify the recruitment of BM cells to the perivascular, meningeal or microglia populations. We will do this in naïve animals, in animals with neurodegeneration, those with systemic inflammation and when these are combined. We will investigate how neurodegeneration in combination with systemic inflammation drives proliferation of recruited or local macrophage populations focussing on the induction of the mitogenic signalling pathways. We will use animal models of neurodegeneration: the ME7 model of prion disease and the APP PS1 transgenic model of Alzheimer's disease. We will correlate our findings with the study of human tissue from AD patients who have died with or without systemic inflammation. To address the contribution of the CSF1R pathway to proliferation and priming we will use an inhibitor of the CSF1R tyrosine kinase activity and assess the impact using behavioural, cellular, molecular and histopathological assays. The proposed multidisciplinary and collaborative research plan will provide novel insights into the understanding of the innate immune response in chronic neurodegeneration and characterize microglial proliferation, identify molecular targets and design and analyse therapeutic approaches to control the progression of neurodegenerative diseases.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Enhancing functional recovery after spinal cord injuries with combinatorial treatments
Funder: Medical Research Council

Abstract: We have previously demonstrated that in rats with a complete thoracic spinal cord transection epidural electrical stimulation (ES) of the lumbar spinal cord can restore locomotor function, especially when combined with daily locomotor training. This approach has been successfully applied to a human patient with a severe incomplete spinal cord injury (SCI), who regained independent standing ability and voluntary control of lower limb muscles while under ES. However, this patient did not recover independent walking. Other factors may prevent further functional recovery. Our preliminary observations strongly suggest that rehabilitation upregulates expression of chondroitin sulfate proteoglycans in the lumbar spinal cord, especially in perineuronal nets around motoneurones, thereby inhibiting synaptic plasticity and making rehabilitation less effective. It has been shown that treatment with chondroitinase ABC (ChABC) opens a window during which rehabilitation could be dramatically more effective by removing perineuronal nets. We propose to test whether digestion of perineuronal nets by ChABC can enhance the effects of ES and locomotor training in a severe contusion model of SCI. ChABC will be administered via a lentival vector delivered at the time of lesion or 28 days post-lesion. Trained rats will be supported in a quadrupedal position and step for 30 min daily under ES. Motor and sensory behavioural tests will determine the extent of functional recovery. A combination of anatomical, physiological and histological analyses will determine underlying mechanisms ES is already available to patients, while ChABC is in late preclinical development. Finding out the parameters for combining rehabilitation, ES and ChABC is therefore a precursor to a potentially dramatically successful treatment for human patients.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT;5.2 CELLULAR AND GENE THERAPIES
Health categories: NEUROLOGICAL