Background Operant conditioning of one’s slow cortical potential (SCP) or sensorimotor rhythm (SMR) can be used to control epilepsy or to manipulate external devices, as applied in BCI (Brain-Computer Interface). To be practical, a BCI-system should use as less channels as possible. For this purpose, a wireless biofeedback system was developed that allows feedback of a single EEG-channel in discrete trials. The commonly accepted view that both the SCP and SMR are a reflection of central arousal suggests a functional relationship between SCP and SMR networks. Methods A training was performed that aimed to teach 19 participants to control their SCP (n=9) or SMR (n=10) over vertex. Participants received 20 neurofeedback sessions, each comprising of 96 trials in which they had to decrease cortical arousal (SCP positivity/SMR enhancement) and 64 trials in which they had to increase cortical arousal (SCP negativity/SMR suppression). In a trial, participants were required to exceed an individual threshold level of the feedback parameter relative to a 500 ms pre-feedback baseline and hold this level for 2 seconds (SCP) or 0.5 seconds (SMR) in order to obtain reinforcement. Results Overall, 10 of the total of 19 participants achieved control over their EEG. In the SCP-trained group, 4 out of 9 participants were able to increase the differentiation between their SCP responses on positivity- required vs. negativity-required trials over the course of the experiment. Improvements in control over the SMR in suppression-required and enhancement-required trials were acquired by respectively 3 and 4 of the 10 SMR-trained participants. These SMR-trained responders did not show differentiation between their SMR responses in enhancement-required vs. suppression-required trials. Interestingly, the SMR responders did show a differentiation in their SCP response while trained on SMR. Conclusions It can be concluded from this experiment that, with the proposed method, a number of the participants are able to acquire control over their SCP or SMR. For SMR, however, bidirectional control is very difficult to achieve with the present training procedure. Furthermore, SCP positivity and SMR enhancement are easier to learn compared to their counterparts. The observed SCP differentiation while training SMR and absence of equivalent SMR changes while training SCP suggest that SMR training modulates the central arousal system, whereas SCP training invokes local effects.

Stroke is a major cause of death and disability in the UK. Few treatments exist and those that do, such as thrombolysis (‘clot-busting’ treatment) must be given urgently and are not risk-free. Large scale randomised controlled trials are crucial for the development of safe, effective, acute interventions, but progress has been limited, ostensibly due to ethical and regulatory difficulties. Theoretical work in this area has focussed primarily upon the requirement for prospective informed consent, but has also considered potential conflicts of interests inherent in the dual role of clinicianresearchers, and the notion that research and clinical practice are, can be, and should be conducted separately. Empirical evidence on this topic is lacking. By providing such evidence, this study examines claims made in the literature regarding the difficulties encountered or perceived in conducting emergency research. It also explores whether, how, and to what effect, the distinction between research and clinical activity advocated in the bioethical literature is maintained. Methods Ethnographic methods were employed, including participant observation, semistructured interviews, and audio-recording of research consent interactions in an acute stroke unit. Data were analysed drawing upon constant comparative and framework methods. Results and conclusion Whilst providing empirical evidence supporting some of the theoretical and conceptual literature, the data also furnish a detailed account of pragmatic issues encountered and managed daily by healthcare professionals in the acute stroke environment. Whilst attempts were made at the study site to separate, at least in part, clinical and research activity, it was observed that absolute separation of clinical activities is neither attainable, sustainable, nor desirable. Placement of research nurses within the clinical environment may promote transparency and greater understanding of their role, whilst simultaneously demystifying research concepts. Ultimately this may promote closer working relationships, contributing to enhanced recruitment, retention and management of research participants.

Voice tremor is associated with Parkinson’s disease (PD), however little is known about the precise characteristics of PD voice tremor, optimum methods of evaluation or possible relationships with other speech, voice, and disease variables. The question of possible differences between voice tremor in people with PD (pwPD) and neurologically healthy ageing people has not been addressed. Thirty pwPD ‘off-medication’ and twenty eight age-sex matched neurologically healthy controls were evaluated for voice tremor features using acoustic measurement, auditory perceptual voice rating, and nasendoscopic vocal tract examination. Speech intelligibility, severity of voice impairment, voice disability and disease variables (duration, disability, motor symptom severity, phenotype) were measured and examined for relationship with acoustic voice tremor measures. Results showed that pwPD were more likely to show greater auditory perceived voice instability and a greater magnitude of frequency and amplitude tremor in comparison to controls, however without statistical significance. PwPD had a higher rate of amplitude tremor than controls (p<0.05). Judged from ‘silent’ video recordings of nasendoscopic examination, pwPD had a greater amount of tremor in the palate, tongue, and global larynx (vertical dimension) than controls during rest breathing, sustained /s/, /a/ and /i/ (p<0.05). Acoustic voice tremor did not relate significantly to other speech and voice variables. PwPD had a significantly higher voice disability than controls (p<0.05), though this was independent of voice tremor. The magnitude of frequency tremor was positively associated with disease duration (p<0.05). A lower rate of amplitude tremor was associated with an increase in motor symptoms severity (p<0.05). Acoustic voice tremor did not relate in any significant way to PD disability or phenotype. ii PD voice tremor is characterised by auditory perceived instability and tremor, a mean amplitude tremor of 4.94 Hz, and tremor in vocal tract structures. Acoustic analysis and nasendoscopy proved valuable adjunctive tools for characterising voice tremor. Voice tremor is not present in all people with PD, but does appear to increase with disease duration. However pwPD examined here represent a relatively mild group with relatively short disease duration. Further work will look at people with more severe disease symptomatology and longer duration.

Parkinson’s disease (PD) is a common neurodegenerative condition with multiple associated non-motor symptoms. Of these, dementia is a frequent debilitating complication of the disorder, with significant morbidity and mortality. Some forms of mild cognitive impairment in PD (PD-MCI) may represent a pre-dementia state and certain clinical, laboratory and neurophysiological parameters may increase the accuracy of prediction of cognitive decline. If validated, these markers would offer the opportunity for disease modification and therapeutic intervention at a critical early stage of the illness, when the viable neuronal population is greater. The key aim of this thesis was to characterise cognitive impairment in PD in a cohort of newly diagnosed cases, and evaluate how a panel of biomarkers correlated with cognitive phenotypes to predict risk of future cognitive decline. The main findings were that PD-MCI was common, and was associated with a distinct clinical phenotype. Memory impairment was the most common single domain affected, although the majority of those with PD-MCI were classified as nonamnestic single domain subtype. A significant correlation was found between pattern recognition memory, sensitive to temporal lobe impairments, and cerebrospinal amyloid-β 1-42 levels, thought to represent amyloid-β metabolism and deposition Both amyloid-β 1-42 and 1-40 levels were significantly lower in those with impaired cognition. In addition, short latency afferent inhibition, a neurophysiological in vivo non-invasive measurement of cholinergic function, was also reduced in participants with mild cognitive impairment. These findings suggest that cholinergic dysfunction and amyloid deposition may contribute to the underlying pathophysiology of early PD- MCI. The major conclusion from this thesis is that PD-MCI is heterogeneous and more frequent than previously reported in early disease. This is associated with abnormalities of amyloid processing and cholinergic dysfunction, and may highlight those at risk of developing dementia. Longitudinal assessment of these individuals will enable us to determine and better model those measures predictive of cognitive decline at an early disease stage.

Alzheimer's disease (AD) is the most common form of dementia in the elderly and is characterized as a 'progressive neurodegenerative disorder' with neuronal amyloid deposition and impaired cognitive function. Impaired insulin signaling is one of the many similarities between AD and type 2 diabetes. Currently no effective treatment is on the market for AD and hence novel therapeutic strategies are needed. In this study, effects of GLP-l analogue, liraglutide on neuroinflammation, stem cell proliferation, differentiation and cognition were investigated. Chronic treatment of liraglutide enhanced neurogenesis in 3, 6, 12, 15 months old APP IPS 1 mice and exerted beneficial effects on different pathological hallmarks (amyloid and dense core plaque, neurogenesis, synaptic density and inflammation) in 7 months old APP/PS 1 mice. Liraglutide administration reduced mean activated microglia load, reactive astrocyte load, cytokines and nitrite levels in the brains of mouse model of neuroinflammation (irradiated mouse brain) indicating that liraglutide has direct role in reducing neuroinflammation. Liraglutide also increased the stem cell pool, progenitor cell proliferation and differentiation, enhanced LTP, recognition memory in irradiated mouse brain. These observations highlight the immense potential of liraglutide as a therapeutic agent for the treatment of neurodegenerative disorders such as Alzheimer's disease.

Diabetic peripheral neuropathy affecting the lower limbs is a common debilitating complication of diabetes mellitus. A significant proportion of affected patients are plagued by severe intractable painful symptoms. In addition to this, the insensitive foot is prone to deformity, trauma and resultant foot ulceration. This thesis deals with the management of diabetic foot problems, with particular reference to painful diabetic neuropathy and foot ulceration. The treatment of painful diabetic neuropathy is far from satisfactory and present therapeutic agents are not without undesirable side effects. The first study (Chapter 3) examines the relationship between blood glucose excursions and pain in patients with symptomatic diabetic neuropathy. Twenty type 1 diabetic patients with peripheral neuropathy (10 painful and 10 painless) wore a continuous glucose monitoring system (CGMS) for 3 days. Symptomatic patients kept a daily pain score diary. Measures of glycaemic stability, the mean amplitude of glycaemic excursions (MAGE) and the M-value were calculated. The study demonstrated that patients with painful neuropathy have greater glucose flux and possibly poorer diabetes control, compared to patients with painless neuropathy. The use of electrical stimulation therapy such as transcutaneous electrical nerve stimulation (TENS) and percutaneous electrical nerve stimulation (PENS), have been shown to provide some benefit when used to treat painful diabetic neuropathy. In a previous open-labelled study, pulsed-dose electrical current delivered through stocking electrodes was shown to produce an 80% reduction in painful symptoms. In the second study (Chapter 4) a double blind, controlled crossover study was carried out to assess the efficacy of pulsed-dose electrical current delivered through stocking electrodes. Thirty patients with painful diabetic neuropathy were randomised to wear silver plated stocking electrodes for 8 hours a night for 6 weeks (pulsed electric current of 50 micro amps delivered by a microcomputer). The control, identical stockings received an insignificant current (5 micro amps). Pre-treatment, weekly and end-of-treatment pain and sleep-disturbance scores were recorded. This study demonstrated that although symptomatic relief occurred, this form of treatment was no more effective than control in the treatment of painful diabetic neuropathy, suggesting that placebo may play a significant role in electro-analgesia. For adequate management of foot ulcers a systematic approach is required. A foot ulcer classification system should aid in planning treatment strategies, monitoring treatment effectiveness, predicting clinical outcomes, and improving communication among healthcare providers. The third study (Chapter 5) examines wound classification systems and factors, which affect the outcome of diabetic foot ulcers. Diabetic patients with new foot ulcers presenting during a 12-month period, had demographics and ulcer characteristics recorded at presentation. Ulcers were followed up until an outcome was noted. This study demonstrated that ulcer area, a measure of ulcer size, predicts the outcome of foot ulcers and that its inclusion into a diabetic foot classification system will make that system a better predictor of outcome. In the fourth study (Chapter 6), two commonly used foot ulcer classification systems are compared as predictors of clinical outcome. Both the Wagner system (grade) and the University of Texas system (grade and stage) were applied to new foot ulcers at presentation, and ulcers were followed up until an outcome was noted. The study revealed that increasing stage, regardless of grade, is associated with increased risk of amputation and prolonged ulcer healing time. The University of Texas system, which combines grade and stage, is a better predictor of outcome. Therefore, strict glucose control should be the first step in the struggle for pain control in patients with painful diabetic neuropathy before other forms of therapy are employed. Additionally, the use of a robust, fully descriptive foot ulcer classification system, such as the University of Texas system should be employed in the management of diabetic foot ulcers. A systematic approach to foot care will aid in reducing the high incidence of lower limb amputations.

As Blood-brain barrier (BBB) breakdown is central to inflammatory lesion formation, it presents a potential target in the formulation of putative therapeutic agents in MS. The action of natalizumab, a monoclonal antibody acting at the BBB, is investigated through a phase III monotherapy trial (AFFIRM) and associated substudies. Subtle BBB disruption from non-inflamed lesions, which could contribute to axonal damage through leakage of inflammatory cells and associated mediators into surrounding parenchyma, is also studied. Introductory chapters (1-3) provide a brief overview of MS, clinical trials, magnetic resonance imaging (MRI), the BBB and natalizumab. Chapter four describes MRI results of AFFIRM- a 2 year multi-centre trial involving 942 patients. Compared with placebo, natalizumab reduced number of gadolinium (Gd)- enhancing lesions by 92%, new/enlarging T2-hyperintense lesions by 83%, and new T1- hypointense lesions by 76%. Chapter five describes a 57 patient AFFIRM trial substudy in which the influence of natalizumab on segmental atrophy was investigated. Atrophy was predominant in grey matter (GM) and was independent of lesion load. Fluctuations in white matter (WM) volume followed changes in inflammatory lesion load. Atrophy was not influenced by natalizumab. The effect of natalizumab on subtle BBB disruption (inferred by measuring the post-Gd %change in T1 weighted signal intensity) is studied in chapter 6. This AFFIRM substudy involved 40 patients (27 on natalizumab, 13 on placebo.) Although subtle BBB leakage was consistently detected in non-visibly enhancing lesions, natalizumab did not influence the degree of leakage. Chapter 7 describes a cross-sectional study which utilised post-Gd change in R1 (1/T1) as a marker BBB leakage. 19 patients (10 RRMS, 9 SPMS) were involved in this study. The subtle leakage observed from non-visibly enhancing lesions was distinct from leakage from visibly enhancing lesions. This was sustained over 60 minutes, greater in smaller lesions and in size-adjusted T1 hypointense lesions.

There is evidence for a role for epigenetic mechanisms in Alzheimer's disease (AD), the most common age-dependent neurodegenerative disorder. The most studied epigenetic mark DNA methylation -the addition of a methyl group to cytosines located in CpG dinucleotides (5mC) - is known to change with aging and may reflect subtle changes in gene expression. Recently a second type of modified cytosine - a hydroxylated and methylated form (5hmC) - has been detected in the brain and maybe linked to the regulation of gene expression. Case-control differences in post-mortem brain DNA methylation were sought by examining both global DNA methylation and DNA methylation of two candidate genes relating to AD risk factors. Simultaneous assessment of 5mC and 5hmC methylation at a global level indicate hypomethylation of 5mC and hypermethylation of 5hmC in AD brain relative to controls, consistent with the notion that 5hmC serves as an intermediary form for demethylation of 5mC. Age was separately associated with a decrease in LINEl methylation and an increase in 5hmC methylation. The comorbidity of depression in AD was explored by assessing the methylation status of the serotonin transporter (SERT) gene promoter across several brain areas and showed tentative associations of disease with SERT CpG methylation. These measurable differences are very small and unlikely to represent any biological plausibility. In a subset of AD patients with additional clinical and behavioural measures there was no effect of SERT 5HTTLPR genotype on DNA methylation. The hypothesis that amyloid- deposition in brain is a consequence of amyloid precursor protein (APP) gene over-expression was examined by measuring DNA methylation across the APP gene region. AD status associates with methylation levels of several CpG sites within the 5' region CGI shore and exon 5 of the APP gene. However there are no co-occurring separate associations of total APP protein levels at these CpG sites. This study demonstrates the utility of the Fluidigm microfluidics platform to generate highly parallel bisulphite sequencing/base­ pair resolution CpG data.

Amyotrophic Lateral Sclerosis (ALS), also known as Lou Gehrig’s disease, typically leads to death within 3-5 years of symptom onset. Understanding what causes ALS has been a challenge, but more research in this area, enhanced by advanced technology like high-throughput next generation sequencing, is paving the way for better information and direction. The volume of data generated by genetics researchers has dramatically increased, largely because of increased opportunities for collaboration. ALSoD, a widely used online genetics database for collating, analysing and integrating ALS data, has been updated with analytics tools and is able to portray the data graphically to users. Mutations and other gene variants have been mapped to genomic coordinates, and the inclusion of dbSNP ids has been implemented to facilitate the integration of data from numerous public sources. To increase the usability and functionality of ALSoD, population frequency of each variant found in the 1000 Genome Project and Exome Variation Server (EVS) databases is displayed. To contribute to a better understanding of the pathogenesis of ALS, links to information on animal models are also available. Furthermore, ALSoD can now be viewed on mobile devices and for Android platforms a mobile app is also available.

Osteopontin (OPN), a glycosylated phosphoprotein, is down regulated in remaining dopaminergic neurones in Parkinson’s disease. It has protective properties including anti-inflammatory and anti-apoptotic effects. Therefore, it was hypothesised that OPN treatment protects dopaminergic neurons from toxin induced cell death, and its endogenous expression in cells confers intrinsic protection. Consequently, OPN was investigated for neuroprotective effects in toxin models of dopaminergic cell death using SH-SY5Y and N1E-115 cell lines with different OPN expression phenotypes, primary E14 ventral meseneephalic (VM) culture and rats. -- Cell lines expressed receptors linked to the pro-survival effects of OPN (Igαv, Igβ3, Igβ1 and CD44). However, endogenous OPN expression did not affect vulnerability of cell lines to the toxins MPP+ or H2O2 and exogenous pre-treatment with OPN did not protect them from toxin-induced cell death. By contrast, pre-treatment of VM cultures with OPN protected them against MPP+-induced dopaminergic cell loss. Importantly, a 15-mer peptide fragment of OPN containing the ROD integrin binding site retained protective properties of OPN. Further, treatment with the integrin receptor antagonists, RGDS and GRGDSPK, prevented OPN’s protective effect, suggesting a role for integrins in the protective effect. OPN induced an increase in the number of microglia in VM cultures but the role of glial cells in OPN’s protective effect is not fully clear. In the LPS lesioned rats, there was an up-regulation of the expression of Igαv, Igβ3, Igβ1 and CD44 receptors and prior intra-nigral OPN injection protected dopaminergic cells of the substantia nigra against LPS toxicity. Immunohistochemical investigation revealed that OPN significantly decreased the inflammatory microgliosis produced by LPS.