Animals that inhabit unfavourable habitats and experience seasons where the cost of maintenance exceeds the available energy resources have over time developed behavioural and physiological mechanisms to survive. These adaptations include changes in activity, improvement of cold tolerance by using nonshivering thermogenesis (NST), improvement of thermal conductance, reduction of body mass, or acclimation to colder temperatures (reduction of metabolic requirement). In addition some species exhibit heterothermy, in the form of either daily torpor or longer-term hibernation. The southern African hedgehog (Atelerix frontalis) is an excellent candidate to investigate the phenomenon of heterothermy because it is a small insectivore (summer body mass ca. 300 to 400g), burrows, inhabits harsh habitats and is not easy to find during the winter months. In this study I aimed to investigate whether A. frontalis exhibits seasonal differences in metabolic rate and furthermore if this species exhibits heterothermy. The study was carried out in the Northern Cape Province, South Africa. Hedgehogs were hand captured and their metabolic rates were measured using indirect calorimetry. Individuals were implanted with temperature dataloggers for a summer period (November 2009-January 2010) and a winter period (May-August 2009). The summer BMR of adult A. frontalis (0.448 ±0.035 mlO2/g/h, n=4) was significantly lower than their winter BMR (0.811 ±0.073 mlO2/g/h, n=4) and statistical analyses revealed that this was an affect caused by seasonal changes in the ambient environment. Individuals spent up to 84 percent of time during the measurement period torpid (-8°C <Ta<21°C). Body mass appears to be an important factor in determining the pattern of heterothermy (daily torpor versus hibernation) used in this species. To my knowledge the extremely low body temperature (Tb min) of 1.0°C recorded for A. frontalis is the lowest Tb min recorded for a mainland Afrotropical mammal. This species displays classic up-regulation in metabolic rate during winter, resulting in an increase in the energetic requirements of the species. As a result, heterothermy appears to play a significant role in the energy balance of this species during winter, contributing to energy saving. Heterothermy may enable this species to survive in the face of global climate change.

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PART I STUDIES ON HEART MUSCLE LIPASES The study of the role of lipids in supplying the energy requirements of the heart has attracted widespread attention, particularly within the past decade. It is now known that the heart, under normal conditions, oxidizes lipids as its main source of energy. Numerous investigators have studied the in vivo and in vitro uptake and utilization of exogenously supplied lipids in the form of triglycerides, free fatty acids and ketone bodies. However, very few have studied the utilization of endogenous lipids by the working heart. We have examined the relative importance of both endogenous glycogen and triglycerides for supplying the caloric needs of the isolated beating rat heart, and found that under the perfusion conditions used, endogenous glycogen appears to supply the initial source of energy. A lipase in rat cardiac tissue was also examined. The enzyme had a pH optimum near 6.8, and was strongly inhibited by 0.2 M NaF and by 2 x 10⁻⁴M diisopropylfluorophosphate. Most of the activity was found in the nuclear fraction of tissue homogenates. The enzyme hydrolyzed both monoolein and mono-stearin, and possessed much less activity against tripalmitin. The enzyme also rapidly hydrolyzed the monostearin component of Ediolʀ (a commercial coconut oil emulsion widely used in lipase studies), and the implications of these findings are discussed. It was concluded from these studies that a lipase other than lipoprotein lipase exists in rat myocardium. PART II STUDIES ON CYCLIC 3', 5'-NUCLEOTIDE PHOSPHODIESTERASE In recent years, the study of the role of cyclic 3', 5'-adenosine monophosphate (cyclic 3', 5'-AMP) in the regulation of several biological reactions and processes has received widespread attention. The presence of a physiological mechanism for terminating the action of cyclic 3', 5'-AMP in biological systems would therefore be expected. Indeed, an enzyme, cyclic 3', 5'-nucleotide phosphodiesterase has been shown to exist in most mammalian tissues which have been studied for its activity. The central nervous system, particularly the cerebral cortex, possesses a very high activity of this enzyme. In this study, cyclic 3', 5'-nucleotide phosphodiesterase was partially purified from rabbit brain and its properties were studied. The enzyme required Mg⁺⁺ions for activity and was inhibited by 2 x l0⁻⁴M theophylline. Cyclic 3', 5'-dAMP, cyclic 3', 5'-GMP and cyclic 3', 5'-dGMP were hydrolyzed by the brain diesterase at approximately one-half the rate at which cyclic 3', 5'-AMP was hydrolyzed. Little activity against cyclic 3', 5'-CMP, cyclic 3', 5'-dCMP and cyclic 3', 5'-TMP was detected, although cyclic 3', 5'-UMP was hydrolyzed at approximately 13% of the rate at which cyclic 3', 5'-AMP was hydrolyzed. The brain diesterase therefore possessed a high specificity for cyclic 3', 5'-nucleotides with purine bases. Optimum enzyme activity was observed near pH 7.0, and the activity was stimulated about 1.5-fold by 0.06 M imidazole. The Km value of the enzyme with cyclic 3', 5'-AMP as substrate was approximately 0.8 x 10⁻⁴M. The properties of the partially purified phosphodiesterase from brain were thus very similar to the diesterases which have been purified from beef and dog hearts. A study of the intracellular localization of the brain diesterase indicated that about 50% of the activity was located in the 105,000 x g supernate. The microsomal and mitochondrial fractions also contained considerable amounts of diesterase activity, but little activity was located in the nuclear fraction. A survey of cyclic 3', 5'-nucleotide phosphodiesterase activity in several available specimens of the plant kingdom indicated the absence of this enzyme activity in these organisms. However, appreciable levels of diesterase activity were detected in E. coli. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

The in vivo biosynthesis of deoxyribonucleic acid (DNA) from labelled thymidine has been investigated in rat intestinal mucosa. The DNA preparations were fractionated by column chromatography and the fractions were assayed for radioactivity by liquid scintillation counting methods. In the first experiments the DNA was isolated from the intestinal mucosa of rats which had received H³-thymidine 5, 10 or 20 minutes or 24 hours before sacrifice. When the macromolecules were fractionated on ECTEOLA-cellulose the results obtained were inconclusive because no definits pattern of incorporation of radioactivity was observed in the fractions. Chromatography on ECTEOLA-cellulose was considered unsatisfactory, because of the variations in the elution patterns of DNA preparations from experiment to experiment and evidence indicating degradation of DNA during the fractionation procedure. In subsequent experiments fractionation on methylated albumin-kieselguhr (MAK) columns was employed and double labelling experiments were carried out. The animals were injected intravenously with H³-thymidine and 24 hours later with C¹⁴-thymidine. The rats were killed 20 or 40 minutes after the second injection and the double labelled DNA was isolated from the intestinal mucosa. On fractionation by MAK columns reproducible elution patterns were obtained even after storage of the DNA solutions. The main DNA peak was always eluted at the same range of sodium chloride concentration and 95-97 percent of the radioactivity was eluted in this peak. Each subfraction comprising the main peak was examined for H³ and C¹⁴ activity. By studying the H³/C¹⁴ ratios of the fractions newly synthesized material could be compared with older, presumably stabilized DNA. When the animals were exposed to the C¹⁴-labelled thymidine for 40 minutes the H³/C¹⁴ ratios of the subfractions were constant, indicating no metabolic differences between the newly synthesized DNA (C¹⁴-labelled and the "old" (H³-labelled) DNA. However, when the time of exposure to the C¹⁴-labelled precursor in vivo was 20 minutes, the H³ /C¹⁴ ratios of subtractions increased as the sodium chloride concentration of the eluant increased. These results indicated some metabolic differences amongst these fractions. Stepwise enzymatic degradation by snake venom phosphodiesterase of the double labelled DNA preparations, and the main peak obtained after MAK chromatography, indicated the incorporation of thymidine into newly synthesized and "old" DNA occurred well within the chain. / Medicine, Faculty of / Biochemistry and Molecular Biology, Department of / Graduate

A humoral factor elaborated by tumor tissue has been suggested as the etiological agent which causes the systemic effects accompanying malignant disease. Because of the important role of the nucleic acids in the metabolism of the cell, it was postulated that the tumor factor might produce the systemic effects by altering the nucleic acid metabolism of the host tissues. With these considerations in mind, a study has been made of the effect of several transplantable tumors on the incorporation of formate-C¹⁴ and tritiated thymidine into the nucleic acids of the host tissues. Studies on the incorporation of thymidine into the host tissues have been emphasized since this compound is a specific precursor of DNA thymine. Liquid scintillation counting methods were developed in order to assay the radioactivity of the tritium-labelled thymine. Methods for the liquid scintillation counting of carbon-¹⁴ labelled purines and pyrimidines were also established. The presence of the ascitic and subcutaneous forms of the Ehrlich tumor was found to have little effect on the incorporation of formate-C¹⁴ into the nucleic acids of the host tissues. In contrast an increased uptake of tritiated thymidine by the DNA of the host tissues was observed in animals bearing the Ehrlich ascites tumor, Novikoff hepatoma and the Walker 256 carcinosarcoma. This effect was particularly striking in the case of the liver and spleen of animals bearing the Walker 256 tumor. Other investigators have isolated a substance known as toxohormone from tumor tissue which has been shown to produce certain systemic effects similar to those of tumor tissue. It was postulated that the increased incorporation of thymidine into the DNA of the host tissues might be the result of the action of toxohormone. In order to test this hypothesis, the effect of toxohormone on the incorporation of tritiated thymidine into the DNA of rat liver and spleen was studied. Crude toxohormone caused an increased uptake of thymidine by the DNA of spleen, but the results obtained for liver were equivocal. A highly purified fraction of toxohormone was prepared by ion-exchange chromatography and gel filtration. However this fraction had no effect on the incorporation of thymidine into the DNA of rat liver and spleen. These results suggested that tumor tissue might contain a factor which stimulates DNA synthesis and which could be isolated in company with crude toxohormone. / Medicine, Faculty of / Biochemistry and Molecular Biology, Department of / Graduate

Excretion of radioiodide by Gasterosteus aculeatus has been shown to vary directly with increases in salinity and inversely with size of fish. Erroneous excretion values may be produced by an apparent laboratory diuresis which seems to vary with season and salinity. During the initial stages of sexual maturation in sea water temporary changes in excretion occur which result in an increased retention of radioiodide by the fish. These changes show a greater development at lower salinities. The demonstrated dependence of thyroid uptake upon available radioiodide make this parameter an unreliable estimate of thyroid activity. The conversion ratio, except for errors which occur under conditions where the rate of radioiodide excretion is not constant over the period of the measurements, seems to be a valid estimate of thyroid activity. Technical factors, such as the binding of inorganic radioiodide to precipitated protein, can also produce erroneous conversion ratio values. Differences in the protein binding of inorganic radioiodide suggest that changes in blood proteins occur during sexual maturation. Although a seasonal increase in thyroid activity occurs there is no correlation between this and the increase in radioiodide retention. There is therefore no reason to believe that the thyroid is responsible for the observed changes in retention of radioiodide. / Science, Faculty of / Zoology, Department of / Graduate

The present thesis is a study of the factors that affect the assessment of rate and efficiency of gain in Yorkshire swine in the pre and post-weaning stages of growth. Calculations from metabolism data for growing swine and measurement of the milk production of Yorkshire sows were used to demonstrate the inability of a sow to produce sufficient milk (energy) for her suckling young about 12 to 20 days post-farrowing. A well balanced high energy creep ration was used in a commercial swine herd to overcome the energy debt to suckling piglets and provide for attainment of maximum growth, and hence, reduce the time required by them to reach 200 pounds. The affect of subnormal hemoglobin levels on pre-weaning growth rates and weaning weights of suckling piglets was investigated. The need of a continuous supply of iron and energy (creep ration) for suckling piglets to produce normal hemoglobin levels and to permit them to grow at their genetic potential was demonstrated. Energy content of gain and resting energy metabolism data were used to calculate post-weaning feed requirements of four experimental litters that were 12½ percent inbred. The post-weaning growth studies show uniformity of litter averages for several economic characters of swine, but variations within litters were high as shown by large standard deviations. The relationship between post-weaning rate of gain, feed efficiency and dressed carcass is discussed. The results demonstrate the possibility of rapidly improving rate and efficiency of gain in swine if selection is based on the performance of individual animals. / Land and Food Systems, Faculty of / Graduate

Energy demands for osmotic regulation and the possible osmoregulatory role of the thyroid gland were investigated in the euryhaline starry flounder, Platichthys stellatus. Using a melting-point technique, it was established that flounder could regulate body fluid concentration independent of widely divergent environmental salinities. Small flounder experienced more rapid disturbances of body fluid concentration than large flounder after abrupt salinity alterations. The standard metabolic rate of flounder adapted to fresh water was consistently and significantly less than that of marine flounder. In supernormal salinities standard metabolic rate was significantly greater than in normal sea water. These findings agree with the theory that energy demands for active electrolyte transport are greater in sea water than fresh water. Thyroid activity was studied in flounder adapted to fresh water and salt water. Correlative with the higher metabolic rate of small flounder was the more rapid turnover and excretion of radioiodine and greater thyroid uptake of small than large flounder. Percentage uptake of radioiodine by the thyroid was shown to be an insensitive and inaccurate criterion for evaluating thyroid activity in different salinities because removal rates of radioiodine from the body and blood differed between fresh water and marine flounder. Using thyroid clearance of radioiodine from the blood as a measure of activity, salt water flounder were shown to have much greater thyroid clearance rates and, hence, more active thyroid glands than flounder adapted to fresh water. The greater activity of the thyroid of marine flounder correlates with greater oxygen demands in sea water and suggests a direct or adjunctive osmoregulatory role of the thyroid gland of fish. / Science, Faculty of / Zoology, Department of / Graduate

A series of three balance experiments were conducted to study the effect of the addition of animal fat to the diet on the utilization of calcium by growing chicks. In two experiments, the chicks were fed diets containing four levels of calcium, 0.75%, 1.0%, 1.25% and 1.50%, with a calcium: phosphorus ratio of 1.7:1. A fifth treatment was fed in which the top calcium level was adjusted to provide a calcium:phosphorus ratio of 2.5:1. All diets fed contained an excess of Vitamin D. Each of the five mineral treatments was fed as a control diet, to which no supplemental fat was added, and as a high fat diet, in which 8% of ground cellulose was replaced by 8% animal fat. In one experiment the diets contained 24% protein. In the other, the protein level was reduced to 20.6%. In the third experiment, to clarify the effect of adjusting the calcium:phosphorus ratio in the diet, three levels of calcium were fed with calcium:phosphorus ratios of approximately 1.5:1 and 2:1. As in the previous experiments, the different mineral levels were fed as low fat diets, and as high fat diets containing 8% animal fat. The diets fed in this experiment contained 23.6% protein. Criteria of calcium utilization used in all these experiments were rate of growth, feed efficiency, calcium balance (% of calcium retained) and calcium retention (calcium retained per 100 grams of gain in body weight). In addition, in the first two experiments, the percentage of bone ash was determined on a sample of chicks from each group at the conclusion of the experiment. It was found that: 1. The calcium requirement for growth and feed efficiency was not affected by the addition of 8% animal fat to the diet. The calcium requirement for maximum growth when the diet contained 24% protein appeared to be between 1.0 and 1.25% of the diet. All levels of calcium fed provided equal growth with diets containing 20.7% protein. 2. The addition of 8% fat to the diet significantly impaired bone calcification when the diet contained 24% protein. No impairment was evident at the 20.7% protein level. Diets containing 1% calcium provided calcification equal to that provided by diets containing 1.25% or 1.5% calcium. 3. Calcium balance (% of calcium retained) was found to be influenced excessively by differences in feed efficiency. A more useful index of calcium utilization was calcium retention, expressed in terms of body weight. 4. The addition of 8% animal fat to the diet impaired calcium retention (expressed as calcium retained per unit gain in weight) when the diet contained 1% to 1.5% calcium. In the case of the low fat diets, calcium retention rose as the level of calcium in the diet was increased to 1.25%, and remained constant as the calcium level was increased further to 1.5%. When fat was added to the diet, calcium retention rose more slowly as the calcium level was increased, and failed to reach the maximum retention of the low fat diets. The degree to which calcium retention was reduced was not sufficient to affect growth, in the presence of vitamin D above the usual allowance. 5. The addition of 8% animal fat to diets containing 24% protein increased the rate of growth and improved feed efficiency. The addition of fat to diets containing 20.7% protein did not affect the rate of growth and had little, if any, effect on feed efficiency. 6. Adjusting the calcium:phosphorus ratio of the diet within the limits tested did not affect growth, calcification, or calcium retention. / Land and Food Systems, Faculty of / Graduate