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Introduction: Colorectal cancer is a major clinical and public health problem. Early stage colorectal cancer is amenable to surgical intervention with a high cure rate. Screening for the disease has now been rolled out throughout the UK, although current screening modality of faecal occult blood testing (FOBT) is suboptimal. Specific genetic alterations found in primary tumours, including mutations in proto-oncogenes, microsatellite instability and loss of heterozygosity (LOH), have been shown to be detectable in the host plasma. Colorectal cancer is an excellent paradigm in which to investigate the application of assays to detect tumour-related plasma DNA. There is substantial data regarding specific mutations and their frequency in colorectal cancer tissue and adenomas. The ability to detect early stage colorectal neoplasia using DNA plasma assays holds considerable promise as a non-invasive screening modality. Materials and Methods: Assay performance was assessed using colorectal cancer cell lines and archived material from genetic studies. Assays were then applied to a prospective cohort of 124 colorectal neoplasia cases and controls. Plasma DNA was quantified using a sensitive DNA binding fluorescent dye and detection platform. A well-characterised tumour-specific mutation was used as the target for the first assay. This mutation was in a poly A tract of the transforming growth factor beta receptor II (TGFpRII) gene and was assessed using a restriction fragment length polymorphism (RFLP) assay. Microsatellite analysis was performed using 3 polymorphic markers relevant to colorectal neoplasia in matched tumour, normal and plasma DNA samples. A novel assay was developed exploiting real-time fluorescent PCR amplification of single nucleotide polymorphisms in the adenomatous polyposis coli (APC) gene as a means to detect tumour-specific allelic imbalance. Assay performance was determined in spiking experiments, and performance assessed in clinical samples. A further iteration of the assay was developed to quantify tumour specific alleles in plasma by counting individual alleles PCR amplified from plasma DNA of cases and controls. Results: Quantification of total plasma DNA revealed a significant difference between cases and controls (area under the receiver operator curve: 0.7). Despite intensive efforts to overcome the problem, the TGFpRII RFLP assay was affected by technical difficulties when applied to plasma DNA. The required high number of PCR cycles introduced artefact and limited its applicability. Microsatellite analysis demonstrated LOH in matched tumour and plasma samples with maximal sensitivity of 67% but specificity was only 32%. The quantitative PCR assay to detect APC LOH was able to detect 3-5ng of homozygous DNA introduced into 1 ml of heterozygous plasma, and accurately quantified LOH in tumour tissue. However, it was not able to discriminate cases from controls by assessment of plasma DNA. Counting of alleles in plasma DNA from a test case demonstrated matching LOH to that seen in the primary tumour. Allele counting of a further cohort suggested that this approach has a low false positive rate. Discussion: Analysis of plasma DNA is technically challenging due to a low abundance of partially fragmented mutant DNA sequences admixed with normal DNA in plasma with DNAases and PCR inhibitors. However total quantity of plasma DNA was higher in cancer cases compared to controls with good specificity for cancer at high DNA concentrations. Fluorescent microsatellite analysis of plasma DNA demonstrated encouraging overall sensitivity but poor specificity that waslikely at least partly a reflection of low DNA abundance and hence sampling error within aliquots of plasma DNA. A quantitative PCR approach was developed and validated with relevant levels of in vitro sensitivity, and improved discrimination of LOH in some clinical samples. Adaptation of this approach to count alleles individually is labour intensive and expensive but appears to address issues of sampling error due to abundance and hence improves specificity. These data, whilst highlighting some of the technical challenges in the field, demonstrate associations of plasma DNA in colorectal neoplasia that warrant further investigation.

This thesis is about the use of the term social exclusion in contemporary Scotland and how it has given rise to the idea of 'excluded spaces' in political and academic commentaries on deprived neighbourhoods. It argues that, despite criticism, the term offers a useful way of re-assessing disadvantaged places, such that space (for example, in the guise of socio-spatial segregation) should be considered not only an outcome but also an input into processes of exclusion. This is illustrated through exploration of the reproduction of the frequently negative place-images surrounding two deprived neighbourhoods in urban Scotland. The thesis explores how such representation may stigmatise residents therein, ultimately resulting in the production of landscapes of exclusion. Use of a range of qualitative methods (though primarily biographical interviews) demonstrates how negative place-images are constructed and remain resilient to change while also revealing the concrete outcomes of such 'imagined' geographies. Overall, the thesis makes three main points. First, that the construction of 'imagined' and 'real' places are fluid and dynamic processes and differences between 'images' and 'realities' of places are neither as clear cut, nor as 'true' or 'inaccurate' as might be assumed. Second, that 'real' and 'imagined' representations of place are reproduced through practices in, and attitudes towards, place. And third, that migration processes are influenced by, and influential to, the construction of social and place identities. Construction of socio-spatial stereotypes reveals space to be both an outcome and input into processes of exclusion while also demonstrating how many of the taken-for-granted assumptions about disadvantaged neighbourhoods might be considered place-myths.

The microinjection of noradrenaline (NA) into the hypothalamic paraventricular nucleus (PVN) of the rat results in feeding. This response was shown; contrary to previous reports; to be mediated through both a-1 and a-2 NA receptors. Selective blockade of these two receptor sub-types, in conjunction with re-uptake blockade was used to examine the individual contributions of each receptor type to the whole response. It is suggested that the previously reported a-2 receptor specificity of the response to microinjected NA is a result of the location of these receptors. The post-synaptic a-1 receptor being located close to the pre-synaptic re-uptake mechanism, whilst the post-synaptic a-2 receptor is located outside the synapse and thus away from the re-uptake mechanism. The re-uptake mechanism acts to create a concentration difference of microinjected NA between the two receptor sub-types, resulting in a higher concentration and thus a preferential action at a-2 receptors. The involvement of the paraventricular NA system in stress induced eating was examined using a tail pinch procedure. Microinjection of NA antagonists into PVN prior to the onset of the pinch had no effect on the duration or latency of the eating response, thus there was no evidence for the involvement of this system in tail pinch elicited feeding. Further to the suggestion that the NA a-2 receptor is extra-synaptic whilst a-1 is intrasynaptic, the actions of NA were examined at a second site. NA microinjected into the ventral striatum elicited a vigorous locomotor response, although the origins of this showed a clear priming effect. However, this response was unaffected by prior microinjection of NA a-antagonists, preventing an analysis of receptor involvement comparable with that performed in PVN.