The history of human muscular dystrophy with reference to clinical, histological and biochemical studies is reviewed. The value of the recent discovery of an experimental mouse with hereditary muscular dystrophy with clinical, histological, and biochemical similarities to human muscular dystrophy is discussed. The histology of normal muscle, with special reference to ultrastructure, is also reviewed. For this investigation mouse muscle samples for the electron microscope were fixed in Palades osmic acid solution, embedded in methacrylate or epon and stained with either lead hydroxide or phosphotungstic acid. A method for obtaining one day old muscle samples, while keeping the animals alive until the dystrophic symptoms are noted clinically, is outlined. Light microscope sections were obtained from electron microscope blocks and examined after staining with toluidine blue. Experimental results by light and electron microscope observations showed there to be no histological differences between one day old dystrophic and normal muscle. However one day old muscle showed histological differences compared to older muscle. The chief differences were the smaller size of fibers, random distribution of mitochondria and enlarged nuclei, and the presence of abundant interfibrillar sarcoplasm with a conspicuous granular components. The regular repeating pattern of the endoplasmic reticulum of adult muscle fibers was hot seen. Some "atypical" fibers showed similarities to altered fibers seen in older dystrophic mice. The mitochondria were swollen and vacuolated with few cristae and pale matrix. Endoplasmic reticular components were vacuolated and adjacent myofibrils were disorganized. Atrophy of fibers was first noted at 14 days of age and were conspicuous at 63 days. By 63 days alterations were also noted in mitochondria and endoplasmic reticulum and these changes became more prominent with the progress of the disease. Atrophy of the myofibrils was evident and the Z-bands were often irregular and out of register. Connective tissue also increased greatly. Several miscellaneous structures are also discussed. The histological findings are compared to those found by other workers. Several suggestions put forward by others as to the possible cause of the disease are summarized. On the basis of morphological findings it is suggested that ribonucleo-protein synthesis in the nuclei of dystrophic muscle fibers is increased but that there may be some intermediate stage at fault which prevents the conversion of amino acids to myofilaments. / Medicine, Faculty of / Graduate

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The use of Positive Reinforcement Training (PRT) for co-operation during routine husbandry and laboratory procedures is widely advocated as a means of promoting the welfare of nonhuman primates. However, while research originating in US zoos provide qualitative descriptions of how PRT may be used in the training of a wide variety of species, quantitative data and evidence to support the view that PRT reduces stress predominately comes from laboratory studies of primates whose training may have used other methods. Despite official guidelines, training is rarely carried out in the UK and the educational and wider organisational structures concerning training, present in the US are largely absent. The techniques used in the UK were assessed through detailed observations recorded when four stump-tailed macaques were trained to co-operate during venipuncture. Data recorded during training sessions showed that although food rewards were given, their delivery was slow and inconsistent. A certain amount of coercion was used which violates a principle of PRT which states that co-operation should be voluntary. The macaques showed increasing resistance to the process and a mild but detrimental effect on the subsequent behaviour of the study animals. When training resumed 18 months later there were considerable improvements in the techniques used. The macaques showed a greater willingness to participate and there were no significant changes in their behaviour when training days were compared to those when training did not take place. The behaviour of the macaques during venipuncture was judged to be arising from engineered compliance rather than voluntary co-operation. However, it was concluded that the technique observed, if carried out correctly, was a reasonable compromise between forced restraint and voluntary co-operation given the paucity of evidence showing the effectiveness of PRT for invasive procedures. However, it was also concluded that the use of coercion should be recognised and provide a focus for future refinement. The effectiveness and welfare implications ofPRT was assessed through the training of common marmosets to target and allow in-homecage weighing and to provide urine samples. It was found that the trained animals perfonned reliably and that time invested in training could be recouped through faster data collection. Following a period of training or increased positive contact with humans, observations of marmoset behaviour showed a decrease in stress related behaviours and an increase in allogrooming supporting the view that improved relations with humans had a beneficial effect. Following exposure to a mild stressor, trained marmosets showed no elevation in levels of urinary cortisol or stress related behaviours. Untrained animals showed increased levels of locomoting and selfscratching following exposure to the same stressor. It was concluded that PRT successfully reduced the stress associated with the presence of, and manipulation by, humans. Final recommendations were that training can promote the welfare of nonhuman primates and should be used in UK laboratories to a greater extent than is currently the case. However, the lack of educational opportunities for animal trainers in the UK needs to be addressed. It was also recommended that in light of the growing evidence showing the benefits that can arise from training and good relations with humans, the zero-handling policy practiced in many UK zoos should be reassessed.

Typescript (photocopy). / Digitized by Kansas Correctional Industries

During peak lactation female mammals reach a limit in their maximal sustained energy intake (SusEI). The causes of such limits is disputed. In this thesis, I examined the causes of the limits on SusEI at peak lactation, and then explored the consequences of such limits for reproductive performance. Finally I tested a possible physiological mechanism that may underpin the trade-off between reproduction and somatic protection (the oxidative stress theory). To answer these questions, I studied reproductive performance and oxidative stress in two lines of mice previously selected for high and low food intake (MH and ML, respectively). I found that these mice reached a plateau in their food intake around day 13 of lactation. In support of the heat dissipation limits theory, reproductive performance in the MH mice was significantly higher than that of the ML mice. Oxidative damage is expected to be higher among lactating individuals. Moreover, lactating mice with greater reproductive performance are also predicted to experience more oxidative damage. By measuring multiple-markers of oxidative damage and protection in different tissues, I found that lactation resulted in reduced oxidative damage in both brain and serum. Additionally, it did not increase oxidative damage to proteins and DNA in liver. Moreover, multiple measures of oxidative stress in the mammary gland were not significantly different between mice with different reproductive effort. Furthermore, I found that lactating mice with greater reproductive performance (litter size and litter mass) had reduced protein damage in their livers and upregulated protection (HSP70) in their brains. These results were inconsistent with the oxidative stress theory. Finally, I employed a novel approach to assess oxidative stress differences with metabolomics analysis. I found that lactation resulted in significant differences in the metabolome. By focusing on the metabolites that are related to vi oxidative stress, I found that most of these metabolites measured in livers and brains were not affected by lactation which provides more evidence against the oxidative stress theory. My results provide support for the heat dissipation theory as a mechanism explaining the limits on reproductive performance. Moreover it provides comprehensive information against oxidative stress as a mediator of life history trade-offs.

The Siberian hamster (Phodopus sungorus) undergoes a suite of physiological changes in response to short day (SD) photoperiod which includes a marked reduction in body mass (up to 40%). This altered physiology can be reversed by a return to long day (LD) photoperiod and is driven by changes hypothalamic gene expression. Additionally, stimulation of weight regain occurs through spontaneous exercise when hamsters are provided with a running wheel (RW), despite intact photoperiod appropriate hypothalamic gene expression. The foundation hypothesis for this investigation was that the change in body weight in both paradigms is underpinned by an alteration of the growth hormone (GH) axis. Pasireotide, a somatostatin agonist, was utilised to inhibit GH secretion from the pituitary in both paradigms. Measurement of body mass, mass of internal organs, body composition by magnetic resonance imaging, hormonal analysis and in situ hybridization were used to determine the effect of a blockade of GH secretion by pasireotide. Pasireotide suppressed the GH axis in Siberian hamsters; with reduced circulating insulin-like growth factor-1 and altered hypothalamic gene expression of somatostatin (srif) and growth hormone – releasing hormone (ghrh) consistent with an inhibition of pituitary GH secretion. Pasireotide treatment inhibited RW and LD stimulated growth, and when administered to LD hamsters caused weight loss in a similar manner to that which occurs in SD and accompanied by testicular atrophy. In addition, pasireotide increased the incidence of torpor and increased bout length of this hypometabolic state in sedentary SD hamsters. In conclusion, evidence is provided for the hypothalamic – pituitary – growth hormone axis in the determination of photoperiod and RW induced body weight changes. Furthermore, the data show evidence for a novel muscle – brain pathway and evidence for a neuroendocrine pathway involved in torpor induction.

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

published_or_final_version / Pathology / Doctoral / Doctor of Philosophy

published_or_final_version / Obstetrics and Gynaecology / Master / Master of Philosophy