NIHRDataInsights/HRCSData huggingface dataset:
Title: Glycosylphosphatidylinositol (GPI)-anchoring of calcium channel alpha2delta subunits: function and pharmacology
Funder: Medical Research Council

Abstract: Voltage-gated calcium channels are essential for numerous processes in excitable cells, including transmitter release from neurons. The alpha2delta subunit is an accessory subunit of voltage-gated calcium channels, and four alpha2delta genes have been cloned. The alpha2delta subunit proteins are important for three main reasons: Firstly, they are essential for the correct trafficking and functioning of voltage-gated calcium channels, and may also have other roles, such as cell adhesion. Secondly, many forms of damage to peripheral sensory nerves, both in patients and animal models, may result in long-lasting neuropathic pain, associated with altered expression of many genes in damaged dorsal root ganglion (DRG) neurons, one of which is alpha2delta-1, which is strongly up-regulated. Thirdly, alpha2delta-1 and alpha2delta-2 are the therapeutically-relevant binding sites for the gabapentinoid drugs, gabapentin and pregabalin, which are effective in the treatment of certain forms of epilepsy and in neuropathic pain. We have found that these drugs do not cause acute block of calcium channels, but rather inhibit trafficking of the alpha2-delta subunits both in vitro and in vivo. We have further shown in vitro that trafficking of CaValpha1 subunits is also inhibited. They therefore represent an example of a novel mechanism of action of small molecule therapeutic agents, to inhibit ion channel trafficking. It is therefore imperative that we understand better the mechanisms of alpha2delta processing in cells. Our recent novel and exciting finding that alpha2delta subunits are not, as previously thought, transmembrane proteins, but rather are attached to the external leaflet of the plasma-membrane by a glycosylphosphatidylinositol (GPI)-lipid-anchor, in a similar way to prion-proteins, has opened a new route to understanding their mechanism of action. GPI-anchored proteins are strongly localised in membrane microdomains termed lipid-rafts. We will now examine the trafficking and localization of alpha2delta subunits in cell lines, polarized cells and neurons, in relation to their GPI-anchoring. We will examine the functional consequences and the biochemistry of GPI-anchoring, particularly of the major brain isoforms alpha2delta -1 and alpha2delta-3. We will also examine the consequences of the potential interaction of alpha2delta subunits with other lipid raft-associated proteins, such as prion proteins. Finally, a major objective is to examine the mechanism of action of gabapentinoid drugs in relation to the GPI-anchoring of alpha2delta-1 and -2, since such proteins utilise specific trafficking pathways. This research has the potential to elucidate the mechanism of action of alpha2delta subunits, and the gabapentinoid drugs which bind to them.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Neuronal Pathways of Sleep and Anaesthesia
Funder: Medical Research Council

Abstract: Every year, 230 million patients worldwide are given general anaesthetics, drugs that remove our most precious human attribute ? consciousness. The induction of safe and reversible loss of consciousness is a pillar of modern medicine. It also poses one of the most long-standing puzzles in neuroscience. Anaesthesia is a routine procedure, but we do not really understand how it works. Although it is clear that general anaesthetics act at a relatively small number of targets, finding the links between these receptors and anaesthetic-induced loss of consciousness presents a fascinating and important challenge. In this Programme Grant application, we propose to test and elaborate our hypothesis that anaesthetics exert their sedative/hypnotic effects by recruiting natural sleep pathways. We will combine mouse genetics, electrophysiology, and synthetic chemistry to identify the neuronal pathways that underlie the actions of anaesthetics and sedative drugs whose molecular targets have been established. By exploiting what is known at the molecular level, our research should provide insights into both how natural sleep is regulated, as well as how anaesthetics act at the level of neuronal networks. This should inform strategies for the development of new drugs, including those to treat sleep disorders. The Programme has five related projects: In Project 1 we will measure cortical EEG changes simultaneously with changes in the local field potentials of selected thalamic nuclei during anaesthetic-induced loss of consciousness. This will provide fundamental reference data on thalamocortical connectivity in normal animals; these experiments will be repeated for the novel mice we plan to construct, in which known targets are either deleted, or made anaesthetic-insensitive. In Project 2 we will test the hypothesis that anaesthetics such as propofol exert their major effects by acting directly on GABA-A receptors on thalamic neurons. In Project 3 we will pursue an important recent discovery that the sedative/hypnotic agent dexmedetomidine (DEX) does not, as previously thought, act on adrenergic neurons. We will exploit the highly restricted expression of alpha2A-adrenergic receptors, the target for dexmedetomidine?s sedative actions, to test the idea that this drug acts at specific cortical sites. In Project 4, we will develop some exciting new findings of our own that point to an important role for TASK-3 potassium channels in anaesthesia, and we will test the hypothesis that their high cortical expression is responsible. In Project 5 we will investigate the mechanisms by which the hypothalamic tuberomammillary nucleus is inhibited during both anaesthesia and natural sleep.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: NEUROLOGICAL
Title: Peripheral voltage gated sodium channels in health and disease
Funder: Medical Research Council

Abstract: Peripheral voltage-gated sodium channels Nav1.7, Nav1.8 and Nav1.9 are interesting analgesic drug targets. We will examine channel protein distribution of expression within and outside the PNS carefully using epitope tags. We will make Bac-Cre transgenic mice for Nav1.7 and Nav1.9 to examine the function and content of cells that express these channels using cell ablation studies (as we did for Nav1.8). We will focus on Nav1.9+ Nav1.8- neurons that seem to be critical for neuropathic pain. We will examine the wiring patterns of defined sets of neurons in normal and chronic pain states. We will develop function blocking monoclonal antibodies for Nav1.7 that may be clinically useful. We will characterise the microRNAs that are essential for sodium channel expression and determine their mechanism of action. We will continue to work with human geneticists to model sodium- channel related pain, and other syndromes.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Functional anatomy of cognitive training using chunking in working memory in Alzheimers disease
Funder: Medical Research Council

Abstract: BACKGROUND: Working memory (WM) - the ability to hold and use information over short periods of time, is affected early in Alzheimer?s disease (AD). Novel tasks have been developed by my collaborators at the MRC-CBU to investigate strategies that improve WM, such as chunking. Chunking refers to the ability to group information into ?chunks? to improve memory capacity, and involves prefrontal and parietal cortices. In a pilot study I tested 30 patients with early AD on WM tasks requiring chunking. Pilot data demonstrates that individuals with minimal AD used and could learn chunking strategies to improve their WM capacity. AIMS AND OBJECTIVES: This trial uses a novel cognitive training regime based on chunking, a powerful mnemonic strategy, in subjects with early AD. It utilises functional neuroimaging to examine re-organisation of brain activity following cognitive training. The study aims to test the following hypotheses: 1) Training individuals with early AD in the use of chunking strategies will improve their WM capacity. 2) Following training in chunking, improvement in WM capacity will generalise across different modalities of WM tasks and a measure of general cognitive functioning: fluid intelligence. 3) Improvement in WM capacity following cognitive training will be associated with re-organisation of functional activity in the prefrontal cortex (PFC) and posterior parietal cortex (PPC). METHODOLOGY: 30 patients with early AD will be assessed on fluid intelligence and WM tasks, and will perform a verbal WM chunking task whilst undergoing fMRI. They will then be randomised to either a control group or cognitive training group, whereby chunking techniques will be taught and practised for a period of 6 weeks. All subjects will then be reassessed using the same measures of fluid intelligence, WM and fMRI protocol, allowing the above hypotheses to be tested. MEDICAL OPPORTUNITIES: The use of cognitive training in chunking is a novel and important therapeutic approach to preserving memory and quality of life for individuals suffering from AD.


Research activity codes: 6.6 PSYCHOLOGICAL AND BEHAVIOURAL
Health categories: NEUROLOGICAL
Title: Intracraneal Recordings in humans: Study of memory processes and applications to neural prostheses.
Funder: Medical Research Council

Abstract: The focus of the proposed research is to study how individual neurons in human brain encode different cognitive functions. Particularly, I am interested in the mechanisms supporting the formation of episodic memories. Extracellular recordings measuring the activity of single neurons and Local Field Potentials (LFP) are one of the predominant methods for studying the underlying mechanisms supporting brain processes. In rare occasions such recordings are available in humans due to the implantation of electrodes in epileptic patients for clinical reasons. Previous studies involving extracellular recording in humans have shown that neurons in the Medial Temporal Lobe (MTL) of the human brain fire to conscious perception of different sensory stimuli representing abstract concepts and, moreover, those concepts can be decoded from the signal obtained during recordings. This unique opportunity of recording from neurons in humans will allow me to study the neural activity in the MTL during the manipulation of concepts forming sequences, which is a simplistic view of an episodic memory. The experiments will include the monitoring of the subject?s gaze employing an eye-tracker in parallel to the recording of the neural activity by intracranial electrodes. The eyetracker will allow performing unique experiments thanks to the complementary information about patient?s behaviour. I will also investigate the possibility of decoding the sequences presented from the neural activity. This technique will be also useful for the future development of cognitive neural prosthetics to be used by patients suffering from spinal cord injuries or neurodegenerative diseases affecting the peripheral nervous system. I will be spending 12 months at Andersen Lab at the California Institute of Technology, one of the leading laboratories in the field, with the intention of receiving experimental training in monkey neurophysiology and intracranial recordings for neural prosthetics.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Frontal cortical mechanisms and interactions during learning and decision making
Funder: Medical Research Council

Abstract: Prefrontal and anterior cingulate cortex (PFC and ACC) play fundamental roles in voluntary or goal-guided behaviour. They are essential for decision making and learning. Changes and biases in such cognitive processes are central to the behavioural alterations seen in psychiatric illnesses. The aim of the current proposal is twofold. First, we intend to uncover the mechanisms of operation of PFC and ACC by using computational approaches to describe the effects of selective brain lesions and patterns of brain activity recorded with functional magnetic resonance imaging (fMRI). Second, we intend to uncover the manner in which these brain areas are able to perform such functions through their interactions with other brain regions. Combined lesion-fMRI studies will address this second aim. The work will be carried out using a macaque model and will employ the Oxford MRC-funded macaque MRI facilities. The focus is on five brain regions. It is hypothesized that ventromedial frontal cortex is essential making value-guided decisions because it rapidly focuses on key choice options; after a lesion it is hypothesized that activity in other brain regions will no longer be correlated with the values of the key choice options that might be taken. It is hypothesized that lateral orbitofrontal cortex and dopamine make distinctive contributions to the learning of stimulus-reward associations; while the former learns precise associations the latter reinforces choices in proportion to their proximity to reward regardless of the exact nature of any contingency. Third, it is hypothesized that frontopolar cortex has a special contribution to make to learning because it mediates counterfactual learning; the ability to learn not from experience but from observation of consequences of choices not taken. Fourth, it is suggested that the anterior cingulate sulcus has a role in assigning value to the component stages of a course of action that determines how primates engage in, start, and stop courses of actions. Fifth, it is suggested that the anterior cingulate gyrus has a role in the valuation of social information that is related not just to the manner in which the social status of conspecifics is perceived but to the manner in which an individual expresses social status. Finally it is hoped that provision of the resources requested will make it possible to continue a programme of work using diffusion weighted MRI to compare anatomical structure in humans and macaques and investigating individual differences in learning induced changes in anatomical connexions.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;4.2 EVALUATION OF MARKERS AND TECHNOLOGIES
Health categories: NEUROLOGICAL
Title: Generating midbrain dopamine neurons from stem cells using novel contact-dependent signalling proteins
Funder: Parkinson's UK

Abstract: During our project investigating novel proteins that are expressed in the developing brain and that might help the conversion of stem cells to dopamine nerve cells, we identified a protein that responds to vitamin D, the vitamin D receptor. We therefore investigated what would happen when we added active vitamin D to developing nerve cells in cell culture, and saw that rather than generating new nerve cells, the active vitamin D increased survival of dopamine nerve cells already present. Therefore we wish to explore precisely whether if we give vitamin D orally to rats, we can protect the dopamine nerve cells in their brains. We will feed extra vitamin D to groups of rats prior to and following injection of a toxin to one side of their brain to partially destroy the dopamine nerve cells on that side. Because the toxin works slowly, we should see if vitamin D can prevent some of the nerve cells from dying. We will look at the animals’ movements by using specialised behavioural tests and at the end investigate how many nerve cells were protected by vitamin D. In running this study we aim to know whether vitamin D in the diet can enhance protection of dopamine nerve cells, and if this does indeed occur, then this effect could be translated fairly rapidly to a clinical trial to see if extra vitamin D intake can improve symptoms for people with Parkinson’s.


Research activity codes: 5.2 CELLULAR AND GENE THERAPIES
Health categories: NEUROLOGICAL
Title: Derivation of conditionally immortalised mouse drG cell lines.
Funder: National Centre for the Replacement, Refinement and Reduction of Animals in Research

Abstract: Rodents are used in academic and pharmaceutical research to obtain cells for culture and tissues for histology. Neuroscience research is particularly dependent on primary cultures of neurons prepared from rodents (mainly rats but increasingly from transgenic mice as well). This is because neurons do not divide and thus do not exist as cell lines. Pain is an important warning system to guard against tissue damage. Pathological pain, however, has no warning value and has huge economical, social and personal costs on society. "Pain" scientists study sensory neurons of the dorsal root ganglia (drG) in order to delineate pain signalling and thus discover new analgesic drug targets. drG neurons are a heterogeneous population of neurons with distinct functional and histochemical properties. However, published drG-derived cell lines do not represent a particular type of drG neurons but rather provide a "generic" sensory neuron environment. They do not represent all functional types of drG neurons and thus do not replace the need for primary cultures to study other sensory neuron types. The aim of this proposal is to generate conditionally immortalised neuronal cell lines from adult mouse drG neurons. We aim to isolate and functionally characterise enough clones to reflect the functional diversity of drG neurons. We plan to use the established H2Kb-ts-A58 Immorto mouse which has been used by many groups to derive conditionally immortalised cell lines from various tissues. However, one drawback of the Immorto mouse is that all cells express the Immorto transgene and therefore a drG culture from it will contain immortalised cells from neuronal, glial and non-neuronal origins we have devised a novel strategy in order to select against the survival and propagation of non-neuronal cell types from Immorto drG cultures. The strategy is based on expressing the bifunctional reporter gene βgeo selectively in neuronal cells of the drG using Cre recombinase. This will enable us to use antibiotic selection to purify clones of neuronal origin only. We believe that the impact on the 3Rs will be considerable and easy to realise. The generated cell line can be deposited and be made available for academic researchers to validate and use in their research. We will also make them available for drug companies to replace acute cultures for in vitro screening and testing experiments.


Research activity codes: 2.6 RESOURCES AND INFRASTRUCTURE (AETIOLOGY);5.9 RESOURCES AND INFRASTRUCTURE (TREATMENT DEVELOPMENT)
Health categories: NEUROLOGICAL
Title: A chronic model of epilepsy in organotypic brain slice cultures of the rat entorhinal cortex
Funder: National Centre for the Replacement, Refinement and Reduction of Animals in Research

Abstract: Epilepsy is a serious neurological disorder affecting around 50 million people world wide. Research into basic mechanisms of the disorder relies heavily on the use of animals. A favoured approach in this area is to establish chronic epileptic conditions in vivo and then use brain tissue from these animals in in vitro experiments to determine changes in neuronal networks that accompany overt chronic seizures. The in vivo chronic models involve instigating a severe acute seizure in the experimental animal, which is followed after a latent period of weeks to months by establishment of chronic spontaneous recurrent seizures. However, determination of changes underlying the development of the chronic condition is difficult, as the approach dictates that longitudinal changes in synaptic networks over the development phase cannot be conducted in a single animal. Individual animals must be sacrificed at arbitrary time points without knowledge of where that animal is in the chronic development process. This makes for a high degree of variability in the data, and is extremely expensive in terms of the total number of animals required. The current application seeks to development an in vitro model of the whole process in tissue from individual animals. Brain slices will be prepared from normal rats, and maintained in long-term organotypic cultures. After a period of stabilisation they will be subjected to a similar acute insult to that induced in the in vivo models, and thereafter monitored (electrophysiologically and histologically) at regular intervals to establish changes occurring during the period leading to appearance of chronic electrographic seizures activity. Since multiple slices can be prepared from a single animal, these can be used to monitor longitudinal changes in network activity in that animal, with untreated slices from the same animal used a parallel controls. Such an approach has the major advantages of eliminating the need for the severe and stressful acute induction phase in the animals, and dramatically reducing the number of animals needed to establish time-dependent changes in networks leading to chronic seizures. We will develop this model in organotypic slices taken from young adult animals and once the model is established we will use it to investigate changes in glutamate and GABA transmission that underlie the development of chronic seizures.


Research activity codes: 2.6 RESOURCES AND INFRASTRUCTURE (AETIOLOGY);2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL
Title: Development of cell based assays as replacement assays for Botulinum toxins and antitoxins
Funder: National Centre for the Replacement, Refinement and Reduction of Animals in Research

Abstract: Potency testing of Botulinum toxin and antitoxin therapies is entirely dependent on mouse lethality bioassay where groups of mice are exposed to the toxin by intra-peritoneal injection and monitored over 72 or 96 hours. Due to a lethal endpoint and the large number of laboratory animals used, the mouse LD50 assay is considered inhumane and new alternative methods required to refine, replace and to reduce animal use. More humane non lethal local flaccid paralysis assays have been developed which are approximately 10 times more sensitive than the lethality method but still require large numbers of animals The use of primary cells is limited because of the continued requirement for animals, time-consuming dissections and potential contamination with non-neuronal components. One approach to avoid these limitations is to develop sensitive human cell lines models. To date, cell based approaches using established cell lines have failed to provide a sensitive functional endpoint and new human cell based assay still eagerly required in new therapy strategies . In order to develop a suitable human cell model, human neuroblastoma cell lines have been established in culture at NIBSC and routinely differentiated into mature neurons. The suitability of differentiated SH-SY5Y cells for detecting high picomolar levels of Botulinum type A has already been evidenced using neurotransmitter release measurements. Application of a micro-electrode arrays (MEAs) platform as a potential functional neurotoxicity assay will be implemented in parallel with imaging of synaptic vesicle recycling, immunodetection and neurotransmitter release measurements. This new approach is entirely non animal orientated and will not rely on any animal experimentation widely used in other applications and strategies. In vitro functional neurotoxicity assay offers the potential of entire animal replacement because of its mode of action involving all key steps of Botulinum intoxication.


Research activity codes: 5.9 RESOURCES AND INFRASTRUCTURE (TREATMENT DEVELOPMENT)
Health categories: NEUROLOGICAL