A series of trials was carried out to examine the role of prolactin (PRL) in the control of lactogenesis in New Zealand Romney x Border Leicester ewes. In addition, a study was made of differences in milk yields and plasma PRL concentrations between spring- and autumn-lambing ewes.Daily subcutaneous injections of 2 mg CB154 inhibited PRL secretion and delayed lactogenesis. There were no consistent effects on plasma progesterone or insulin concentrations. CB154 treatment was more effective in reducing milk yield in twin-bearing than in single-bearing ewes when used for 20 days than for 9 days prepartum. The differential effects on milk yield cannot be explained by corresponding effects on plasma PRL or insulin concentrations. Circulating PRL during the period 20 to 10 days prepartum may have an important effect on milk yield in twin- but not single-bearing ewes.Subcutaneous injections of 0.5 mg/kg live weight oPRL, administered on 2 consecutive days peripartum, to ewes treated with CB154 for 7 days prepartum, resulted in milk yields similar to those in control ewes and significantly (P<0.01) greater than those in ewes treated with CB154 alone. This indicated that oPRL prevented the CB154-induced reduction of milk yields and has established that the effect of CB154 on lactogenesis is mediated through suppression of PRL secretion and not by effects on some other hormone.Injection of 10 mg oPRL directly into one mammary gland (via the teat duct) increased milk yields relative to the contralateral, bicarbonate-treated gland in CB154-treated ewes. The intramammary oPRL injection did not raise circulating PRL concentrations. Furthermore, the milk yields of bicarbonate-treated glands in ewes treated with bicarbonate only, did not differ from those of bicarbonate-treated glands in ewes treated with oPRL in the contralateral gland, demonstrating that there were no effects of oPRL, transferred via the circulation from the treated gland, on the contralateral gland. Glands treated with oPRL produced 15% (P<0.05) more milk than the bicarbonate-treated glands during the first 8 days of lactation and the difference was maintained throughout the 8-week lactation period, indicating that the oPRL had effected a permanent change in the ability of the gland to produce milk. It is concluded that PRL acts directly on the mammary gland without the need for a putative intermediate hormone, and that intramammary PRL concentrations during lactogenesis may have long-lasting effects on lactation.The possibility was examined that dietary differences were responsible for seasonal differences in plasma PRL concentrations, milk yields, milk composition, lamb birthweight and lamb growth rate, observed in earlier trials. Mean plasma PRL levels were significantly (P<0.01) higher in spring- (192±38 ng/ml) than in autumn- (71±17 ng/ml) lambing ewes housed indoors under constant photoperiod (18L:6D) and fed the same diet. Milk yields were also significantly (P<0.05) higher in the spring- (2041±114 g/d) than in the autumn- (1563±109 g/d) lambing ewes over the 8 day lactation. Lamb growth rates (adjusted for birthweight, birthrank and sex of lamb) from birth to 8 weeks of age were significantly (P<0.001) higher in spring (282±12 g/d) than in autumn (225±15 g/d). The seasonal differences were confounded with corresponding differences in ewe live weight and it was not possible to determine whether dietary differences contributed significantly to the differences observed.Two routes of oPRL supplementation were used to test the effectiveness of elevating peripheral or local levels of PRL in autumn-lambing ewes which, based on previous results, were expected to have low plasma PRL concentrations and milk yields relative to spring-lambing ewes. Administration of 10 mg supplementary oPRL directly into the gland or subcutaneous injection of 0.5 mg/kg oPRL did not increase the milk yields, or change the composition of milk, compared to controls. These results suggest that the circulating level of PRL, and the intramammary concentration of PRL, in autumn-lambing ewes are not limiting lactogenesis. Because the plasma prolactin concentration in the ewes was unexpectedly high, it was not possible to reach firm conclusions regarding possible effects of supplementary oPRL in ewes with naturally low plasma PRL concentrations. Nevertheless, the results indicate that raising the intramammary concentration of PRL around the time of parturition, in ewes with circulating PRL levels characteristic of normal spring-lambing ewes, does not enhance lactogenesis.It is concluded that PRL is important to the complete initiation of lactogenesis in ewes, that it acts directly on the gland and that it is necessary for establishing the maximum potential of the gland to secrete milk.

The main objective of this PhD was to provide fundamental information regarding some metabolic aspects of selenium metabolism in cats and dogs. The total selenium content of a range of commercially available petfoods was analysed using a fluorometric method. The petfoods contained a wide range of selenium concentrations, with up to 6 µg Se/g DM found in cat foods. Mean concentrations of selenium in dog and cat foods were 0.40 and 1.14 µg Se/g DM respectively. All petfoods analysed met the recommended current minimum dietary selenium requirements. The use of blood parameters for the assessment of selenium status was investigated in a study in which cats were fed inorganic and organic selenium supplemented at concentrations of up to 2.0 µg Se/g DM for 32 days. Plasma selenium concentrations reflected dietary selenium intakes, however there were no differences between the different levels of supplementation. Whole blood selenium concentrations showed less distinct patterns and were thought to be a more useful indicator of longer term selenium status. Activities of glutathione peroxidase in plasma and whole blood showed no response and the response of cats to supplementation of the different forms of selenium were similar. In the same study, faecal and urinary excretion (µg/kg BW/d) were measured and apparent absorption and retention were estimated during the last seven days of the 32 day trial. Faecal excretion of selenium remained constant whereas urinary excretion of selenium increased with increased dietary intake. The form of selenium had no effect on excretion or apparent absorption however there was a trend in which more selenium was retained in cats fed organic selenium. A study was conducted with cats and dogs fed high levels (10 µg Se/g DM) of inorganic and organic selenium for 21 days to determine whether there were species differences in their metabolic response. Cats and dogs exhibited the same pattern of response, however cats showed higher plasma selenium levels, lower levels in liver and excreted more selenium compared to dogs. It was concluded from this data that cats and dogs differ in their metabolism of selenium. The effect of heat processing on the addition of inorganic and organic selenium to petfoods was investigated in cats fed 3.0 µg Se/g DM for 11 days. Apparent absorption was higher in cats fed inorganic selenium added after processing, whilst less selenium of organic origin was excreted in the urine when added after processing. These preliminary results suggest heat processing may decrease the apparent availability and utilisation of selenium in petfoods.

Slaughter by ventral neck incision (VNI) is performed on some animals without prior stunning in New Zealand and other countries. A single incision with a razor sharp blade is made in the ventral aspect of the neck, sectioning both carotid arteries and jugular veins, though, not the vertebral arteries. There are a number of potential welfare concerns surrounding slaughter by VNI including pain due to the incision, which may lead to distress during the time before loss of consciousness. The aims of this thesis were to identify cortical responses indicative of noxious stimulation due to slaughter by VNI using analysis of the electroencephalogram (EEG) power spectrum and to investigate the effects of non-penetrative captive bolt (NPCB) stunning on these cortical responses. The studies utilised adaptations of a minimal anaesthesia model, which has been validated in a range of mammalian species. Surgical dehorning was used as a validation technique for this methodology in cattle and demonstrated a ‘typical’ EEG response to noxious stimulation. Cattle slaughtered by VNI without prior stunning produced specific responses in the EEG that strongly indicated responses to noxious stimulation. Causation was investigated in cattle where blood flow through the brain remained intact during neck tissue incision (NTI) or the major blood vessels of the neck were isolated and transected independently of other neck tissues (BVT). The response to neck incision in intact animals was principally due to the noxious sensory input due to incision of neck tissues and not mainly as a result of loss of blood flow through the brain. NPCB stunning produced states of cortical activity that were incompatible with the maintenance of sensibility and pain perception. Experimental examination of the time to onset of undoubted insensibility was attempted in cattle subsequent to a pilot study in sheep. The generation of somatosensory-evoked potentials was problematic in cattle. The conclusions of this thesis are that incision of neck tissues during slaughter without prior stunning constitutes a substantial noxious stimulus. Were an animal conscious, this stimulus would be perceived as painful until the onset of hypoxiainduced insensibility. This would represent a significant compromise to animal welfare.

Causes of morbidity and mortality were investigated for 108 green turtles (Chelonia mydas) stranded in south-east Queensland between 1990 and 1996. This study was undertaken as part of a broader carcass salvage program for south Queensland, and within the context of a population study of C. mydas in the Moreton Bay feeding ground. Accurate pathological characterisation of disease in C. mydas was achieved by detailed necropsy and histological examination. Varied inflammatory responses and degenerative changes were observed in stranded C. mydas. Supportive disciplines of microbiology, parasitology, and clinical chemistry were used to elucidate aetiology and pathogenesis of selected conditions. Heavy metal and pesticide levels were assessed in a sub-sample of turtles. Direct anthropogenic causes (including trauma, foreign body ingestion and drowning) accounted for 34% of mortalities of C. mydas in this study. The majority of the trauma cases were turtles with skull fractures resulting from blunt impacts. The remainder had boat propeller injuries, or miscellaneous trauma. Almost half of the turtles with lethal boat propeller damage had evidence of pre-existing disease which may well have predisposed them to boat strike, emphasising the importance of full necropsy examination, even when the cause of death appears obvious. Fishing line was the only ingested foreign body consistently implicated in the production of fatal intestinal obstruction. Marine turtle fibropapillomatosis, a panzootic viral disease which is considered to involve some indirect anthropogenic factors, accounted for 7% of mortalities. The findings in this study were consistent with much of the previously described pathology of this condition. Naturally-occurring diseases (for which human influences are unknown) accounted for the remaining 59% of strandings. Coccidiosis, caused by Caryospora cheloniae, was recorded for the first time in wild C. mydas. It occurred both as an epizootic (in 1991) and as sporadic cases. A variety of manifestations, including disseminated and enteric forms, were recognised. Infection with a Cryptosporidium-like protozoan appeared to occur concurrently with coccidiosis in one turtle in this study. Attempts to establish experimental coccidial infections in hatchling C. mydas were unsuccessful. Infections with cardiovascular (spirorchid) flukes were almost universal in stranded C. mydas in this study. They ranged from mild, incidental findings (such as occasional fluke vii egg granulomas evident microscopically in otherwise normal tissues) to a variety of severe changes, including thrombosis, which were likely to have produced morbidity. The present study clarified the range of cardiovascular lesions associated with spirorchidiasis, including the sequence of thrombus resolution and exteriorisation from vessels. In some cases spirorchid vasculitis was associated with fatal disseminated bacterial infections. Other sporadic, naturally-occurring diseases included mycotic pneumonia, bacterial meningoencephalitis and a miscellany of gastrointestinal conditions, including chronic intestinal tympany and obstipation, for which the underlying cause could not always be determined. Evidence indicated that gastrointestinal motility in C. mydas was prone to both direct and indirect disturbance and that tympany and obstipation could be final common outcomes of a range of insults. Eighteen abnormally buoyant turtles were examined during this study. The cause could usually be ascribed to an underlying disease, including (in decreasing order of frequency) trapped internal gas, usually intestinal; neurological disease such as traumatic brain injuries; and pulmonary disease. In two cases, no underlying cause was detected. Trace metal (arsenic, cadmium, mercury, selenium and zinc) concentrations were determined in the livers and kidneys of 50 turtles of mixed species (mostly C. mydas). These results were considered to provide baseline data for sea turtles in SE Qld. This study offered the largest dataset available for some metals in C. mydas, and provided evidence of high background levels of cadmium as a normal feature for the species. Some unusual age–related trends in metal accumulation were detected. Concentrations of cadmium, zinc and selenium in the kidney decreased with increasing age, whereas zinc concentrations in the liver tended to increase. Determining the impact of disease on wildlife populations is an increasingly necessary task, which will require multidisciplinary teams. Necropsy surveys like the present study are an essential component of the growing field of conservation medicine. In addition to providing data relevant to management, such as the relative proportions of anthropogenic and naturally-occurring mortalities, necropsy surveys can identify a range of endemic pathogens, and help to collect prevalence data for determining disease impacts at the population level.

Slaughter by ventral neck incision (VNI) is performed on some animals without prior stunning in New Zealand and other countries. A single incision with a razor sharp blade is made in the ventral aspect of the neck, sectioning both carotid arteries and jugular veins, though, not the vertebral arteries. There are a number of potential welfare concerns surrounding slaughter by VNI including pain due to the incision, which may lead to distress during the time before loss of consciousness. The aims of this thesis were to identify cortical responses indicative of noxious stimulation due to slaughter by VNI using analysis of the electroencephalogram (EEG) power spectrum and to investigate the effects of non-penetrative captive bolt (NPCB) stunning on these cortical responses. The studies utilised adaptations of a minimal anaesthesia model, which has been validated in a range of mammalian species. Surgical dehorning was used as a validation technique for this methodology in cattle and demonstrated a ‘typical’ EEG response to noxious stimulation. Cattle slaughtered by VNI without prior stunning produced specific responses in the EEG that strongly indicated responses to noxious stimulation. Causation was investigated in cattle where blood flow through the brain remained intact during neck tissue incision (NTI) or the major blood vessels of the neck were isolated and transected independently of other neck tissues (BVT). The response to neck incision in intact animals was principally due to the noxious sensory input due to incision of neck tissues and not mainly as a result of loss of blood flow through the brain. NPCB stunning produced states of cortical activity that were incompatible with the maintenance of sensibility and pain perception. Experimental examination of the time to onset of undoubted insensibility was attempted in cattle subsequent to a pilot study in sheep. The generation of somatosensory-evoked potentials was problematic in cattle. The conclusions of this thesis are that incision of neck tissues during slaughter without prior stunning constitutes a substantial noxious stimulus. Were an animal conscious, this stimulus would be perceived as painful until the onset of hypoxiainduced insensibility. This would represent a significant compromise to animal welfare.

Slaughter by ventral neck incision (VNI) is performed on some animals without prior stunning in New Zealand and other countries. A single incision with a razor sharp blade is made in the ventral aspect of the neck, sectioning both carotid arteries and jugular veins, though, not the vertebral arteries. There are a number of potential welfare concerns surrounding slaughter by VNI including pain due to the incision, which may lead to distress during the time before loss of consciousness. The aims of this thesis were to identify cortical responses indicative of noxious stimulation due to slaughter by VNI using analysis of the electroencephalogram (EEG) power spectrum and to investigate the effects of non-penetrative captive bolt (NPCB) stunning on these cortical responses. The studies utilised adaptations of a minimal anaesthesia model, which has been validated in a range of mammalian species. Surgical dehorning was used as a validation technique for this methodology in cattle and demonstrated a ‘typical’ EEG response to noxious stimulation. Cattle slaughtered by VNI without prior stunning produced specific responses in the EEG that strongly indicated responses to noxious stimulation. Causation was investigated in cattle where blood flow through the brain remained intact during neck tissue incision (NTI) or the major blood vessels of the neck were isolated and transected independently of other neck tissues (BVT). The response to neck incision in intact animals was principally due to the noxious sensory input due to incision of neck tissues and not mainly as a result of loss of blood flow through the brain. NPCB stunning produced states of cortical activity that were incompatible with the maintenance of sensibility and pain perception. Experimental examination of the time to onset of undoubted insensibility was attempted in cattle subsequent to a pilot study in sheep. The generation of somatosensory-evoked potentials was problematic in cattle. The conclusions of this thesis are that incision of neck tissues during slaughter without prior stunning constitutes a substantial noxious stimulus. Were an animal conscious, this stimulus would be perceived as painful until the onset of hypoxiainduced insensibility. This would represent a significant compromise to animal welfare.

The lens of the eye is a vital tissue in the visual system, responsible for the collection and focusing of light on to the retina. Comprised of epithelial cells at differing stages of differentiation, the transparency of the lens is dependent on the highly ordered crystalline structure of lens proteins. The lens consists of several proteins including crystallins (α, β, γ) that make up 90% of the soluble protein, and the lens cytoskeletal proteins. Cytoskeletal proteins contribute only a fraction of the total lens protein, but are thought to play an important role in the establishment and maintenance of transparency. Calpain-induced degradation of these proteins may be involved in the development of cataracts. This has been an area of research at Lincoln University where a flock of sheep genetically predisposed to cataract maintained as a cataract development model. The aim of this research was to investigate the distribution of cytoskeletal proteins in the lens, and to examine the effects of calpain proteolysis on these proteins, with the goal of establishing the role of the lens cytoskeletal proteins in the ovine cataract model. A combination of techniques was used including immunohistochemistry, which required the development of a specific protocol for ovine lenses. Cytoskeletal proteins were identified using immunohistochemistry in lens tissue sections and exhibited characteristic distributions. Actin displayed preferential distribution in the short sides of the fibre cells in the cortex of the lens but was absent in the lens nucleus, while spectrin in the cortex and nucleus was associated with the fibre cell membrane. Filensin was observed in the outer cortex of lens sections associated with the fibre cell membrane and cytoplasm, although the pattern of localisation was indistinct due to the abundance of filensin breakdown products. Vimentin displayed membrane and cytoplasmic association in the outer cortex that diminished toward the lens nucleus, with membrane associated vimentin only persisting in the deeper regions of the cortex and nucleus. Additionally, the effect of novel calpain inhibitors (Cat0059 and Cat811) in preventing proteolysis of lens cytoskeletal protein was investigated and compared with calpain inhibitors developed elsewhere (SJA6017). The inhibitors were tested at between 10 and 0.1 μM (100 nM). All inhibitors were effective at 10 μM. SJA6017 provided significant protection to vimentin at 1 μM. Cat0059 was found to protect spectrin and filensin at 1 μM, but not vimentin, while inhibitor Cat811 was found to protect spectrin only. SJA6017 added to assays at 100 nM offered significant protection to spectrin, and Cat0059 was found to protect filensin and spectrin to a significant degree at 100 nM, indicating the novel inhibitors were comparable to those developed elsewhere in terms of their effectiveness. Taken together, the evidence presented in this thesis shows the cytoskeletal proteins as crucial elements in the lens. Their pervasive presence coupled with evidence that lens cytoskeletal proteins are sensitive to calpain-induced proteolysis that is inhibited with novel calpain inhibitors suggests that the lens cytoskeletal proteins may be useful targets in cataract prevention for future research.