Individuals with schizophrenia have severe cognitive impairments that impact upon their ability to function within society. Better understanding the genetic mechanisms underlying schizophrenia and cognition provides an opportunity for targeted pharmacological intervention. This thesis investigates common and rare genetic variation in schizophrenia and their associations with cognitive ability in schizophrenia cases and healthy controls. Polygenic risk of schizophrenia and bipolar disorder predicted ability on tests of cognitive domains affected in schizophrenia, performance, verbal and full scale IQ in healthy controls. Increased polygenic risk of schizophrenia was robustly associated with lower performance IQ at different training thresholds in two independent cognition samples. There was no consistent association between bipolar polygenic risk and cognition. Common genetic differences between schizophrenia and bipolar disorder were associated with verbal and full scale IQ. I investigated the hypothesis that 155 gene-sets across six biological categories relating to cognition, brain function and structure were enriched for SNPs influencing general cognitive ability. Schizophrenia polygenic pathway scores for gene-sets were not associated with general cognitive ability in schizophrenia patients, or performance IQ in healthy individuals. Separately, neither gene-sets vi nor general categories were enriched for common SNPs showing association with general cognitive ability in schizophrenia cases. Associations between rare CNVs and general cognitive ability were tested in schizophrenia cases. Cases with a known pathogenic CNV performed approximately one standard deviation below other schizophrenia cases on the MATRICS composite score. In addition, increases in the number of genes hit by large (>100kb) and rare (frequency <1%) CNVs were associated with lower general cognitive ability. However, the number of genes hit in gene-sets previously mentioned was not associated with the MATRICS composite score. These findings indicate genetic variation in schizophrenia is associated with cognitive ability in schizophrenia cases and healthy controls, providing direction for future research.

Disease (HD) is an autosomal dominant neurodegenerative disease caused by a CAG expansion on the HD gene on chromosome 4, which gives rise to an expanded polyglutamine (polyQ) tract in the huntingtin protein. HD is characterised primarily by motor abnormalities, but is also commonly associated with cognitive impairments and psychiatric disturbances. Huntingtin (HTT) dynamically shuttles between subcellular compartments, however the mutant huntingtin protein (mHTT) is mislocalised to cell nuclei, where it may interfere with nuclear functions, such as transcription. The phosphorylation of HTT has been implicated with the regulation of its subcellular localisation; however the mechanism by which the mislocalisation of mHTT occurs is currently unknown. The localisation of HTT in an immortalised embryonic striatal cell model of HD (StHdhQ111) was identified as being more nuclear with a longer polyQ length. Additionally, stimulation of StHdhQ111 lines with a panel of growth factors alters the apparent subcellular localisation of HTT in a polyQ length-dependent manner. Aberrant kinase responses to growth factor stimulation were identified by ELISA assay. Inhibition of AKT1 and MEK1 phosphorylation indicated that their activation may be involved in mHTT mislocalisation and immediate-early gene expression. Transcriptional dysregulation is a characteristic feature of HD pathogenesis that may be directly influenced by HTT localisation. Microarray analysis of differential gene expression responses to growth factor stimulation uncovered a potential role for SMAD transcription factor activity and the transforming growth factor-β pathway in HD pathogenesis. Primary embryonic striatal cells from the HdhQ111 and HdhQ150 mouse models of HD showed similarities to the StHdhQ111 cell model in terms of HTT localisation, kinase signalling and gene expression. Cellular dysfunction may therefore be an early HD phenotype that is present from embryogenesis, and potentially alters HD development and progression; aberrant control of kinase signalling may regulate mHTT mislocalisation, which in turn modulates transcriptional dysregulation and contributes to HD pathogenesis.

Huntington’s disease (HD) is an inherited neurodegenerative condition characterised by progressive motor, cognitive and psychiatric symptoms. The most frequent cause of death is respiratory failure, yet little is known about respiratory function through the progression of the disease or the underlying causes of respiratory failure. A thorough exploration of the relevant literature led to the development of a conceptual framework for respiratory failure in people with HD. Within this framework respiratory failure was characterised as type 1 hypoxaemic and type2 hypercapnic failure and further evaluated through (i) an observational study to investigate respiratory function in people with HD, and (ii) the benefit and feasibility of inspiratory muscle training in people with HD. In order to develop understanding of potential underlying causes of type 1 hypoxaemic and type 2 hypercapnic respiratory failure, the observation study aimed to investigate if there was a difference in respiratory function between healthy controls and people with HD at different stages of the disease, and to explore factors that may influence or be influenced by respiratory function. The framework was further evaluated through the intervention study which investigated the feasibility and benefit of inspiratory muscle training in people with HD as a method of increasing capacity of the respiratory system. Method In the observation study 67 people with HD and 39 healthy control participants underwent a series of measurements of respiratory function based on underlying causes of type 1 hypoxaemic respiratory failure and type 2 hypercapnic respiratory failure. These included measurement of lung volume, respiratory muscle strength and endurance. Exercise capacity, physical activity, swallow and posture as potential influencing factors were also measured in people with HD. In the intervention study 20 people with HD were randomly allocated either to inspiratory muscle training at 50% of maximal inspiratory pressure, or to training against a load suggested to have no effect, completed in the home. The training protocol was 30 breaths, twice daily for six weeks, which was preceded by a habituation period of one week. Sniff nasal inspiratory pressure, peak cough flow and 30 second sit to stand were measured before and after the intervention. The programme was supported by alternate weekly phone calls and home visits. Results All measures of respiratory function, except FEV1/FVC were significantly decreased (p <0.001) in people with manifest HD compared to healthy control participants and people with pre-manifest HD. There was no difference between healthy control participants and people with pre-manifest HD. Respiratory function demonstrated a significant linear decline with disease progression measured by the total functional capacity scale (p<0.001). In particular, peak cough flow was abnormal at the middle stage of the disease. Exercise capacity, physical activity, swallow and posture were significantly related to respiratory function in people with manifest HD (p range 0.016-0.001). In people with manifest HD, exercise capacity was 27.73% ±26.29 predicted and swallow capacity was abnormal in 84.80% of participants. In the intervention study, five participants completed the intervention arm and 7 completed the sham arm. Adherence to the inspiratory muscle training programme ranged from 37-100% across both groups, with mean adherence rates of 70.67% ±26.35 and 74.53% ±21.03 for intervention and sham groups respectively. There was no difference in inspiratory muscle strength, peak cough flow or 30 second sit to stand as a result of the intervention. Participants and their carers identified carer support as a key enabler and life events as a barrier for carrying out the exercises. Conclusion The findings from this study indicate that people with HD are susceptible to type 1 hypoxaemic respiratory failure and predisposed to type 2 hypercapnic respiratory failure due to increased elastic and resistive loads and decreased capacity of respiratory muscles. The risk of type 1 hypoxaemic respiratory failure is high due to decreased swallow capacity and concomitant decreased cough efficacy. Decreased lung volume leading to hypoventilation may be impact on both type 1 hypoxaemic respiratory failure and lead on to type 2 hypercapnic respiratory failure. The predisposition to type 2 hypercapnic respiratory failure is due to decreased respiratory muscle capacity and increased elastic and resistive load. The study also highlighted the complex relationship between respiratory function, exercise capacity and physical activity. Although inspiratory muscle strength, cough efficacy and functional activity remained unchanged in this small sample, the results of the intervention study suggest that inspiratory muscle training is feasible in people with HD. Further studies should use protocols that are directly related to the primary outcome measure e.g. a power based protocol to improve cough efficacy or an endurance based protocol to improve physical activity. A model of respiratory failure in people with HD incorporating both type 1 hypoxaemic and type 2 hypercapnic respiratory failure can be proposed based from the findings of the studies that informs future research and clinical management of people with HD.

Absence seizures (ASs) are the predominant form of seizure featuring in the idiopathic generalised epilepsies, and are the only seizure type of childhood absence epilepsy. They are characterised by behavioural arrest, impairment of consciousness and an electrographic signature of spike-and-wave discharges (SWDs) and are associated with psychosocial and cognitive impairment of development. The seizures are known to arise in the thalamocortical network, but the firing dynamics of thalamic neurones during seizure is not known. In vivo and in vitro studies have yielded contradictory results, suggesting predominant silence and regular burst firing respectively, but no studies have previously recorded from intact, single thalamic neurones in a freely moving model of absence epilepsy. In this thesis it has been shown that, in Genetic Absence Epilepsy Rats from Strasbourg, thalamocortical (TC) neurones are mostly either silent or fire single spikes irregularly but synchronously during AS. T-type calcium channel-mediated bursts in neurones of the reticular thalamic nucleus (nRT) were frequently observed during full seizure expression. These cells expressed varied firing patterns ranging from regular burst firing to predominant silence, with similarly varying degrees of synchrony. It is also suggested that the nRT burst firing observed may be required for seizure generation. T-type calcium channel-mediated burst firing of TC neurones is neither necessary for, nor commonly observed in, the full generation or propagation of absence seizures These results suggest that TC neurones are predominantly silent during AS. This is compatible with the idea of a cortical seizure initiator and driver, as suggested by the cortical initiation site and cortical abnormalities observed in multiple experimental AS models. The observations herein also confirm that the temporal relationship between thalamic firing and SWDs previously observed in anaesthetised animals is maintained in the freely moving condition, but suggest that there is a greater incidence of asynchronous thalamic activity during AS (particularly of nRT neurones) than previously suggested. The firing dynamics of thalamic neurones observed are a crucial step towards understanding TC network activity during AS, and provide a significant insight into the role of the thalamus in alterations of sensation, movement, and consciousness associated with these seizures.

This thesis encompasses a range of experiments designed to characterise and validate a method of desensitising rodent hosts in the neonatal period to human tissue in order to promote the survival of human striatal grafts in the adult host. Thus, the successful application of this method is important to allow the preclinical testing of potential human donor cells for therapeutic transplantation, specifically in neurological disease. Demonstrating safety and functionality of transplanted human cells in rodent hosts requires long term assessment of surviving grafts, for which current immune suppression methods are insufficient. The experiments presented in this thesis were therefore designed to determine the optimum parameters of a previously described method of desensitising rats to human tissue and to validate this method in mice. The findings provide further support for the neonatal desensitisation method in rat hosts, and suggest the potential for use of non-neural tissue types for desensitisation of neonates. The data presented in this thesis also has implications for the mechanisms underlying the success of the method in the rat. However interpretation was difficult as graft survival was generally poor and even mouse to mouse allografts did not survive to the level expected. Thus this highlights the need to reassess standard immunosuppression protocols in mice, and determine what differs between the rat and mouse rejection response to xenografts.

Disease loci underlying complex disorders such as schizophrenia(SCZ)are likely to exist as variants of multiple typesbeyond the single nucleotide polymorphisms(SNPs) typically surveyed in genome-­‐wide association studies(GWAS). Large copy number variants (CNV), small insertions and deletions (INDELS),rare compound heterozygous mutations and other classes of genetic variation are all expected to contribute to disease risk. A true understanding of disease genetics cannot be attained without careful consideration of each class of variation and how they are related. Additionally, it is often the case that no or few single variants have strong enough effect sizes to attain the level of statistical significance required to offset the large number of tests performed in a genome-­‐wide single locus survey of any variant type. It has been shown that a cumulative burden of risk alleles is correlated with disease risk, indicating true biological signal within those variants. However, this approach merely implicates that class of variation across the genome and does not refine variants to a subset of risk regions or genes. Defining sets of genes within biological pathways and testing them as a group will allow for both the power of aggregating loci of small effect and interpretation of the underlying biology. This dissertation assesses the role of multiple classes of variation from different technologies in risk to schizophrenia with a particular focus on brain related biological pathways.

Genome-wide association studies have identified TCF4 (transcription factor 4) as a susceptibility gene for schizophrenia. In addition, rare TCF4 mutations cause Pitt Hopkins syndrome (PTHS), a severe form of intellectual disability associated with characteristic facial features, developmental delay and autonomic dysfunction. TCF4 belongs to the basic helix-loop-helix family (bHLH) of transcription factors that play a central role in development, however the precise function of this gene in the brain is unknown. In this study, I use molecular and cellular techniques to improve our understanding of TCF4 function in the brain. Modeling of PTHS-associated missense mutations in transfected cells showed that TCF4 mutations affecting the DNA-binding domain cause mislocalisation of the mutant protein. DNA-binding domain mutations also impaired dimerisation and attenuated transcriptional activity at the NRXN1 and CNTNAP2 promoters. TCF4 mutations affecting other domains of the protein had context-specific deficits in dimerisation and transactivation under the same conditions. Microarray analysis of SH-SY5Y cells where all TCF4 isoforms had been knocked down identified gene expression changes affecting cellular processes including epithelial-to-mesenchymal transition, apoptosis and neurodevelopment. However, isoform-specific knockdown experiments showed that TCF4-A and TCF4-B isoforms affect distinct biological processes such as the cell cycle, chromatin modification (TCF4-B), cell adhesion and cytoskeletal remodeling (TCF4-A). Finally, mass spectrometry was used to identify TCF4-associated proteins in SHSY5Y cells. These experiments showed that TCF4-B and TCF4-A co-purified with proteins involved in chromatin organization, cell cycle control and RNA splicing. In addition to the bHLH factors HAND2 and TWIST2, TCF4 co-purified with components of the nuclear co-repressor complex. TCF4 also has multiple phosphorylation sites in both activation domains suggesting that TCF4 function may be regulated by kinase signaling. In conclusion, these data provide a mechanistic insight into the function of TCF4 that may advance our understanding of disease processes in PTHS and schizophrenia.

Schizophrenia is a highly heritable, common psychiatric disorder. Although onset generally occurs during adolescence, multiple lines of evidence point to a neurodevelopmental insult that occurs many years prior to the presentation of symptoms. Many different approaches have been used to elucidate the genetic risk factors and their impact; however, few unequivocal facts have been established. With a considerable amount of data publically available, integrative approaches look to leverage multiple data sources to identify coherent themes. This thesis investigates the neurodevelopmental hypothesis of schizophrenia by incorporating results from genome-wide association studies and copy number variation studies with gene expression datasets, with the overall aim of identifying functional pathways that may be disrupted in the aetiology of the disorder. This study used foetal and developmental expression datasets of the human brain and statistical approaches to characterise the expression profiles of schizophrenia risk genes. Both spatial profiles in the mid-foetal brain and temporal profiles across development were considered. Data from genome-wide association studies and copy number variation studies were used to test for an enrichment of risk genes; in addition the genetic overlap with bipolar disorder identified through genome-wide association studies was used for validation. Gene sets with a common expression profile enriched for schizophrenia variants were used to identify biological pathways and assessed for their polygenic contribution to schizophrenia risk. The results of this thesis converged on a common developmental expression profile for schizophrenia risk genes. Genes identified with this profile were shown to harbour multiple, common risk variants for schizophrenia and were implicated in epigenetic processes relating to the regulation of gene transcription. Together this suggests that schizophrenia associated genes are involved in brain development, particularly during foetal stages, and may play a role in the regulation of this process.

The diagnosis and management of normal pressure hydrocephalus (NPH) remains controversial. The aim of this prospective study was to assess the usefulness of clinical and radiological criteria together with supplemental neuropsychological and gait tests, cerebrospinal fluid (CSF) hydrodynamic studies and external lumbar drainage (ELD) in identifying those who may respond to a shunt and to compare the outcomes with the non-shunted patients at one year. Method: Forty patients with a clinical diagnosis of NPH were prospectively studied according to a fixed management protocol. Resistance to CSF (Rcsf) was measured using a lumbar infusion study and an ELD was used to determine improvement in neuropsychological and gait tests following CSF drainage. Based on specific criteria those who showed improvement were shunted. Clinical and radiological outcomes were assessed at one year in all patients. Results: Twenty three (57.5%) patients were shunted. Improvement was observed in 74% of shunted patients, while 17% did not improve and 9% deteriorated following surgery. Age, etiology, presentation, duration of symptoms and presence of co-morbid factors were unrelated to outcome. Improvement was found in 63% of shunted patients with RCSF of 12 mmHg/ml/min or higher. The sensitivity was 64% with a positive predictive value of 68%. Both Rcsf testing and ELD enhanced the positive predictive outcome of shunt operation. Using the non-shunted patients as controls, the mean difference between the two groups over time differed significantly in all the neuropsychological tests and some gait test. Conclusions: No single test was able to predict overall success with shunting but the results showed that a high percentage of improvement could be achieved by using continuous ELD and a rigorous protocol. Greater improvements were observed in cognitive and gait function than in sphincter control. An Rcsf of 12mmHg/ml/min or more was related to better outcomes. Consideration was given to the significant limitations in the study design and outcome measures.

Depression is a significant global problem and is among the leading causes of disability worldwide. Depression in children and adolescents is associated with wide-ranging impairments and often marks the beginning of a lifelong, chronic illness. Early treatment and prevention of depression is therefore a major public health concern. Parental depression is one of the most consistently identified risk factors for depression in young people. Although depression is a highly heterogeneous disorder, most studies examining cross-generational depression risk have simply dichotomised parents into ‘depressed’ and ‘non-depressed’ groups and clinical characteristics beyond diagnostic status are rarely presented. In this thesis I examine how differences in clinical features of parental depression including variations in depression course, severity, timing and symptom manifestation differentially relate to offspring depression risk. Data were drawn from the Early Prediction of Adolescent Depression study. A three-wave longitudinal study of the offspring of 337 parents with a history of recurrent unipolar depression. Within this high risk group of offspring, specific clinical features of parental depression were identified that may serve as useful markers of current and/or future offspring depression risk. These included a recent episode of clinical depression, an episode involving severe impairment or hospitalisation and symptoms of appetite or weight loss. In addition, findings from this thesis highlight that there is considerable variability in the course of parent depression over time and suggest that any persistent symptoms of depression in parents, even those at low levels, may be clinically important in indexing offspring risk for depression symptoms. vi Findings highlight the importance of considering clinical characteristics of depression in parents beyond diagnostic status when examining cross-generational depression risk. The identification of subgroups of offspring who are at greatest risk can help ensure that clinical services and preventative interventions are targeted to those with greatest need.