An investigation into dopamine-melatonin interactions in the rat Corpus striatum and pineal gland: a possible pineal-striatal axis
- Authors: Boyd, Clinton Shane
- Date: 2000
- Subjects: Pineal gland -- Research Melatonin Dopamine -- Physiological effect Dopamine Brain chemistry Rats -- Physiology
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3906 , http://hdl.handle.net/10962/d1003965
- Description: Dysfunction of central dopaminergic systems has been implicated in neuroendocrine, neurodegenerative and psychiatric disorders. Monoamine oxidase and catechol-Omethyltransferase represent the key catabolic enzymes of dopamine, terminating neurotransmission following synaptic release of this catecholamine. Thus, both enzymes have been associated with the pathology of dopaminergic systems and represent therapeutic targets elf enormous clinical importance. Some neuroendocrine and circadian effects of melatonin have been attributed to an antidopamimetic effect of this pineal hormone in the hypothalamus and pituitary. Furthermore, both melatonin and dopamine modulate the behavioural output of the mesencephalic dopaminergic pathways of the basal ganglia, including movement disorders. However, the biochemical basis for the tonic inhibitory effect of melatonin in the nigro-striatal pathway has been poorly delineated. Thus, this study determined whether melatonin influences dopaminergic function in the corpus striatum of the Wistar rat by modulating monoamine oxidase and catecholO- methyltransferase activity. Reciprocally, the putative existence of an intrapineal dopaminergic system was investigated by determining the effect of selective dopaminergic agents, R-( -)apomorphine, haloperidol and dopamine, on indole metabolism of the pineal gland. The akinetic state of drug-induced catalepsy was employed as an animal model of Parkinson's disease to probe the neurotransmitter systems involved in the behavioural effects of melatonin. Indole metabolism was a reliable indicator of state-dependent metabolic fluxes in pineal gland function. These included a robust diurnal and seasonal variation in N-acetylserotonin and melatonin biosynthesis, and photoperiod- and drug-induced alterations of Inftabolism. The predominant changes could be attributed to an effect on serotonin N-acetyltransferase activity and/or the melatoninl5-methoxytryptophol ratio. Pineal 5-methoxyindole biosynthesis was determined primarily by the bioavailability of the corresponding 5-hydroxyindole and its affinity for hydroxyindole-O-methyltransferase. Evidence was found for the negative feedback or paracrine control of pineal indole metabolism by melatonin. A high inter-individual variability was observed in the biosynthesis of N-acetylserotonin and melatonin biosynthesis, and the weight of the pineal glands. Accordingly, the rats could be classified as either high or low capacity producers of these two indoles. R-(-)-apomorphine and dopamine in vitro, but not acute haloperidol in vivo, had dose- and phase-dependent effects on pineal indole metabolism. The predominant effect was a suppression of the scotophase-dependent induction ofN-acetylserotonin and melatonin biosynthesis by dopamine and R-( -)-apomorphine. It is postulated that these agonists inhibited nocturnal N-acetyltransferase activity via postsynaptic pineal D2 or D2-like receptors. The observed modulatory nature of the intrapineal dopaminergic system suggests that dopamine may be involved in the long-term regulation of pineal indole biosynthesis. Several lines of evidence are presented that the activity of striatal monoamine oxidase A and catechol-O-methyltransferase, represented predominantly by the soluble isoform, is statedependent and regulated in vivo by endogenous melatonin. Firstly, both enzymes showed a daynight variation in activity. Secondly, acute and subchronic administration and photoperiod manipulation studies indicated that both exogenous and endogenous melatonin inhibited each enzyme in a chronotypic fashion, with a more robust effect against catechol- -methyltransferase. The intensity of the in vivo effects was critically dependent on the dose, duration, route and the phase-timing of administration during the light dark cycle, and the length of the exposure to constant light. Melatonin in vitro had no effect on basal or Mg2+ -induced catechol-Omethyltransferase activity. Thus, it is proposed that the in vivo effects of the hormone can be attributed to a time-dependent change in the amount of active molecules of this enzyme. In contrast, melatonin and numerous other endogenous indolic compounds were found to be reversible inhibitors of striatal monoamine oxidase A in vitro. Structure-activity modeling revealed that the 5-methoxy moiety on the indole nucleus and substitution of the free primary amine of these compounds were the principal determinants of the potency and time-dependency of inhibition. Thus melatonin most likely has a direct inhibitory effect in vivo at the level of the active site of monoamine oxidase A. Exogenous melatonin alone had no cataleptogenic potential whereas a variety of behavioural responses were observed following intraperitoneal administration of y-hydroxybutyrate. The latter responses were state-dependent with day-night variations in intensity. Furthermore, yhydroxybutyrate stimulated melatonin biosynthesis during the photophase both in vitro and in vivo. These results point to a possible involvement of melatonin in the behavioural and neurochemical effects of y-hydroxybutyrate. Thus the general conclusion is that dopamine and melatonin display functional antagonism at the level of the pineal gland and corpus striatum of the Wistar rats. Therefore melatonin may be an important homeostatic modulator of dopaminergic neurotransmission throu~out the central nervous system. Furthermore, the putative existence of a functional pineal-striatal axis would greatly strengthen the argument for a holistic concept of brain homeostasis. The ability of endogenous melatonin to regulate monoamine oxidase A and catechol-O-methyltransferase may represent an alternative strategy for the treatment of disorders associated with these enzymes.
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Genetic variation within and between some rare and common taxa of Cape Proteaceae and the implications for their conservation
- Authors: Brown, Susan Ann
- Date: 2000
- Subjects: Proteaceae -- South Africa Nature conservation -- South Africa Plant conservation -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3905 , http://hdl.handle.net/10962/d1003964
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Studies of the population structure and generic diversity of domesticated and "wild" ostriches (Struthio camelus)
- Authors: Bezuidenhout, Cornelius Carlos
- Date: 2000
- Subjects: Ostriches Ostriches -- Genetics Ostriches -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3907 , http://hdl.handle.net/10962/d1003966
- Description: DNA sequencing and restriction fragment length polymorphism analysis (RFLP) of polymerase chain reaction (PCR) amplified mitochondrial DNA fragments, and random amplified polymorphic DNA sequence (RAPD) analysis were techniques evaluated in this study for applicability in the investigation various aspects of genetic diversity within the ostrich (Struthio camelus). The genetic aspects that were investigated were (i) relationships between ostrich subspecies, (ii) genetic variability between and within domesticated populations of southern African ostriches (Struthio camelus australis), (iii) linking egg production in domesticated ostriches to RAPD profiles, and (iv) determining the zygosity of twin ostriches. In the first part of this study DNA sequencing and the polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) methods were evaluated for resolving genetic differences in the small mtDNA fragments ofthe ostrich. DNA sequencing ofPCR amplified 450 bp 12S rRNA gene fragments of representatives from the southern African population ostrich (S.c. australis) did not reveal any differences between the populatiohs from different geographical areas, representing ostrich lineages with different breeding histories. The PCRRFLP analysis ofmtDNA fragments (450 bp 12S rRNA gene fragment and 550 bp D-loop region) also did not reveal any genetic variability between the domesticated s.,c. australis populations included in this study. PCR-RFLP analysis of a 450 bp 12S rRNA gene fragment, however, showed differences between the subspecies s.c. australis and s.c. molybdophanes. The proportion of shared fragments (F) between these two subspecies was 0.286 and nucleotide sequence divergence estimated at 8.9 %. Divergence time between these two subspecies was estimated at 4.5 million years ago. The data presented from this study are comparable to the data from a previous study in which the entire mitochondrial genome and a larger number of restriction enzymes were used. The PCR-RFLP method thus demonstrated its usefulness for genetic studies of ostriches at thesubspecies level. The sequences used in this study could not reveal any markers that were useful for genetic studies of ostriches at the population level. In the second part of the study the RAPD method was evaluated for application in the genetic studies of ostriches. RAPD profiles, based on three RAPD primers, revealed differences between three subspecies of ostriches and indicated relationships between these subspecies that are consistent with observations from other studies. The numerical analysis of pooled and individual primer data demonstrated that the subspecies s.c. australis is more closely related to s.c. massaicus than to s.c. molybdophanes. RAPD marker differences between s.c. molybdophanes on the one hand, and s.c. massaicus and s.c. australis on the other is also consistent with observations from studies that proposed separate specie~ status for s.c. molybdophanes. RAPD analysis by five primers revealed geographic variation between s.c. australis populations. The clustering patterns observed in the dendrograms and Neighbour Joining Trees generated by computer programs showed trends of separating ostric1;t populations into geographical groups, possibly reflecting their different breeding histories. In the RAPD profiles of the inbred population, band-sharing was generally greater than in the outbreeding group. RAPD analysis thus showed that it may be a useful method in the population studies of domesticated S. c. australis. RAPDs also generated data that grouped ostriches according to trends in egg production capabilities. Analysis ofRAPD profiles by computer software showed a Neighbour Joining Tree and a dendrogram that predominantly grouped ostriches into clusters associated with either good or poor egg production. Evidence supporting the suitability of RAPDs as a tool in breeding programmes of ostriches was thus provided by this study. RAPDs also provided data, demonstrating that two sets of ostrich twins were non-identical twins. It was demonstrated by this study that RAPDs analysis may be a useful technique for applying to (1) systematic (2) population (3) breeding and (4) twin studies of ostriches (Struthio camelus).
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Sulphide-enhanced hydrolysis of primary sewage sludge : implications for the bioremediation of sulphate-enriched wastewaters
- Authors: Whittington-Jones, Kevin John
- Date: 2000
- Subjects: Bioremediation Sewage sludge Hydrolysis
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3910 , http://hdl.handle.net/10962/d1003969
- Description: The potential application of sulphate reducing bacteria for the bioremediation of acid mine drainage has already been recognised, and offers significant financial advantages over conventional chemical treatment approaches. Although the technology has been demonstrated successfully on both small- and large-scale, it’s extensive implementation has been constrained by the provision of suitable and cost effective electron donor and carbon sources. Primary sewage sludge is readily available in large quantities, but the slow rate of solubilization and low yield of soluble products do not apparently favour its use for this application. A number of pre-treatment steps have been introduced in an attempt to improve the yield and rates under methanogenic conditions. However, although early work suggested that degradation of lignocellulose and proteins may be more rapid under sulphate reducing conditions, the fate of primary sewage sludge under these conditions has been ignored. It was proposed that by combining the hydrolysis of primary sewage sludge and biological sulphate reduction, in a settling sludge bed, both processes would be enhanced. The aim of this study was to test this hypothesis on laboratory- and pilot-scale, and attempt to elucidate the underlying mechanism involved. The solubilization of primary sewage sludge was enhanced in the presence of sulphate reduction in continuous laboratory-scale reactors. Particulate matter accumulated in the bed of non-sulphidogenic systems, but not in sulphidogenic ones. This was attributed to increased solubilization and the smaller average floc size in the latter. Solubilization occurred within the settling sludge bed of the reactors, and offered a possible explanation for the better performance of the multiple- over single-stage reactor. A pilot-scale Falling Sludge Bed Reactor was constructed at Grootvlei Gold Mine, Springs, South Africa, and resulted in the solubilization of more than 70% of the influent primary sewage sludge. The system was also found to be highly resilient to severe perturbations, and returned rapidly to steady-state. Flask studies revealed that the hydrolysis of both proteins and complex carbohydrates was accelerated in the presence of biological sulphate reduction or sulphide. A study of the enzymology of sludge digestion revealed that sulphate reduction had little direct effect on the activity of the hydrolytic enzymes, but that reactor design was critical in the prevention of washout of these enzymes. Finally, a descriptive model was developed to explain the enhanced hydrolysis of primary sewage sludge. The model incorporated the effect of sulphidogenesis on floc fracture and reflocculation, and likely implications for mass transfer limitations.
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