D-serine is a co-agonist at NMDA receptors in the brain but the study of this amino acid is restricted by current techniques. I have designed highly sensitive D-serine biosensors that permit accurate real-time recordings of D-serine in the brain in a selective manner. I demonstrate that these tools are ideal for investigating factors involved in the regulation of this amino acid and the role that D-serine plays in excitotoxic cell death mediated via NMDA receptors. I have established that the extracellular basal concentrations of D-serine in the rat brain are heterogeneous and vary even within brain structures. This suggests that Dserine is an important regulatory constraint for NMDA receptor activation, as receptor response can only be potentiated in regions with low D-serine content. Additionally, I show that these microelectrode biosensors have the potential to be used in vivo to detected extracellular D-serine levels. In addition I have observed real-time activity dependent regulation (both loss and release) of D-serine by ionotropic glutamate receptor agonists AMPA, NMDA and kainate, PAR1-agonist TFFLLRNH2 and high frequency stimulation in vitro in a number of brain areas. A decrease in D-serine concentration is potentially neuroprotective as it suggests a reduction in NMDA receptor activation. However, D-serine release can be observed in regions where the co-agonist site of the NMDA receptor is likely to be already saturated implying an alternative function of D-serine in the brain. These findings indicate multifaceted regulation of this amino acid that is brain-region specific. Finally, I have investigated the role of D-serine release during models of stroke (hypoxia and ischemia) and found that D-serine levels are reduced in brain regions deprived of oxygen. This is ultimately neuroprotective as it will reduce over - excitation at the NMDA receptor during these insults. In the more profound model of stroke, oxygen-glucose deprivation, D-serine is eventually released. This release precedes anoxic depolarisation and could therefore contribute to its initiation via enhanced activation of the NMDA receptor.

This thesis aims to improve our understanding of the macroevolutionary implications of antipredator defences, particularly with regard to how defence impacts biodiversity (including both species and trait diversity). To do this I took a phylogenetic comparative approach and used multiple study systems in an attempt to ensure the generality of my work. I begin by investigating how chemical defence and protective coloration influence ecology by testing for life history and ecological correlates of these defences (Chapter 1). Upon finding evidence for an increased niche space in chemically-defended species, and to some degree in conspicuously-patterned species, I explore whether this leads to increased diversification by increasing speciation rates and/or lowering extinction rates (Chapter 2), as also predicted by escape-and-radiate theory (a major and highly influential framework for the macroevolution of natural enemy interactions). Both conspicuous coloration and chemical defence increased speciation rates, but extinction rates were also raised in chemically-defended lineages, leading to a reduction in net diversification. Macroevolutionary extinction rates may or may not be related to contemporary extinction risk, but if they are then there may be conservation implications by allowing prediction of threat status of species with limited direct information. Consequently, in Chapter 3 I asked whether chemically-defended species are more threatened than those lacking such a defence. In accordance with the macroevolutionary results from Chapter 2, I found that chemical defence is indeed associated with a higher extinction risk even amongst contemporary species. In addition to factors that promote diversity, in this thesis I also investigated convergent evolution as a means of constraining diversity of phenotypic traits, using mimicry as a case study for antipredator defences. Many antipredator defences are convergent to some degree, with examples in the repeated evolution of chemical defences and warning coloration as well as independently derived similarity in protective mimicry. However, methods of quantifying the strength of convergent evolution are lacking, not to mention a conceptual framework to define 'strength' in this context, I began by developing a new method to do this which I called the Wheatsheaf index (Chapter 4). Subsequently, I (in collaboration with a colleague, Amanda Minter) also designed software in the form of an R package (called 'windex') to enable user-friendly implementation of the Wheatsheaf index in a familiar statistical environment to many biologists (Chapter 5). In the final data chapter of this thesis, I apply this method in a case study to explore the patterns of phenotypic convergence that result from the evolution of Batesian and Müllerian mimicry complexes. I find that these two types of protective mimicry are generally characterised by convergence in different broad types of traits, but that the specific traits which converge in a given mimicry complex are less predictable (Chapter 6). Overall, this thesis provides novel insights into the evolutionary patterns and consequences of antipredator defences, develops a framework and methods for the analysis of convergent evolution, and suggests further avenues of research for future studies.

The eggshells of many avian species are spotted in appearance but the functional significance of such maculation is poorly understood. Protoporphyrin, responsible for brownish-red colouring on eggshells, is postulated to reinforce the structural integrity of eggshells under conditions when dietary calcium (Ca) is scarce. Within the context of this hypothesis, this thesis documents the use of Ca supplementation of two common British passerine species, blue (Cyanistes caeruleus) and great tits (Parus major), to explore the relationships between eggshell Ca and protoporphyrin content and visible pigment spotting. It further assesses the diversity of avian eggshell coloration using museum eggshells of 73 British passerine species. Despite low soil Ca availability, females were not necessarily Ca-limited but Ca-supplements may still influence eggshell traits and breeding behaviour, possibly by providing females with more time to invest in other activities. The importance of quantifying eggshell pigment concentrations directly, rather than using a proxy, is highlighted. Finally, this thesis shows that passerine eggshell pigment concentrations are highly phylogenetically conserved, thereby encouraging future studies testing key hypotheses to compare eggshell pigmentation of closely related species. This phylogenetic association may be essential to explain the functional significance of eggshell coloration of avian species.

This study reviews and evaluates the current status of management planning and policy in Protected Areas (PAs) in Bangladesh via a case study of two Wildlife Sanctuaries and one National Park. Using a mixed method approach, the research evaluates the perceptions and attitudes of local residents and other stakeholders towards the effectiveness of protected area planning and management, specifically co-management plans for the case study areas. The research was based on semi-structured and in-depth interviews with key informants, focus group discussions, and questionnaire surveys of village residents. The study revealed a complex socio-economic context characterised by poverty; within this, a diverse range of stakeholders exist whose interests in, and perceptions of, protected area management do not necessarily coincide with those of the Forest Department officials, leading to situations of conflict and difficulty for the Forest Department in enforcing the law over areas of forest in Bangladesh, a situation that has not been helped by the limited availability of manpower and modern equipment. It is concluded that the co-management approach, by taking into account the interests, wishes, and aspirations of the local communities, holds out better prospects of protecting the forest, meeting the objectives of the protected area management plans, and development ambitions of local communities. However, the findings suggest that community participation needs to be improved if effective forms of co-management are to be achieved, in turn improving the chances of conserving the forest for future generations while permitting the present generation to pursue sustainable livelihoods. Based on this study, it is clear that not only is action required to increase the participation of the local community, but human resource development is required to produce parallel institutional capacity building within the Forest Department. Moreover, programmes designed to support the generation of alternative livelihood opportunities are required to reduce dependency on forest resources. All of these lines of development are essential to increase the capacity of the local communities and officials to work together to develop and implement the management objectives of the National Parks and Wildlife Sanctuaries. However, without genuine political will to act it will be difficult to achieve the aforementioned ambitions/objectives.

An investigation was carried out into the relationship between environment and reproduction in the freshwater triclad species. By studying two populations occupying adjacent habitats, under varying conditions of temperature, flow rate and resource availability, the effects of habitat variability on the di~ribution~ density, size structure and the levels of se>:ual and ase>:ual reproduction within each population were assessed. The population occupying a habitat which was characterised by its eurythermic temperature regime, high flow rates and complex macroinvertebrate community, occurred at low densities. Within this population, individuals were larger, on average, than in the adjacent population, and sexual reproduction occurred at high levels throughout the year, with asexual reproduction (by binary and multiple fission) also occurring throughout the yea~, but at lower levels. The seasonal nature of this habitat was reflected within the popUlation of , which exhibited spring peaks in density, followed by summer peaks in the level of sexual reproduction. In contrast, the adjacent population occupied a habitat which was characterised by its stenothermic temperature regime, low flow rates and a less complete macroinvertebrate community, and occurred at much higher densities. Within this population, individuals were smaller, on average, than in the adjacent population, and sexual reproduction was virtually absent, wi th asexual reproduction (by binary and multiple fission) occurring throughout the year at appreciable levels. The lack of seasonality within this habitat was similarly reflected in the lack of any seasonal fluctuations in density, or level of (in this case asexual) reproduction within the population. The level of food availability varied seasonally in both habitats, however, and was generally similar, in terms of biomass, in bath areas. A hypothesis was presented which of triclads within each habitat related the density to the total food availability, measured as stream drift. It was suggested that at low population densities, the relatively higher levels of net resource availability per i ndi vi dual favoured the occurrence of sexual reproduction, in contrast with situations of high population density, where the relatively lower levels of net resource availability per individual inhibited the occurrence of sexual reproduction, thus favouring asexual reproduction. This hypothesis was supported by the results of laboratory investigations in other studies, together with the evidence gained from field observations from this study, particularly the observation that in the low dem::.i ty population, seasonal cycles in the level of sexual reproduction were out of phase with seasonal cycles of a similar nature in the level of popUlation denSity. Further evidence, from the results of .a field manipulation experiment, was presented which supported the hypothesis. In an area of high popUlation denSity, density was reduced artificially, resulting in a significant increase in the level of sexual reproduction within the population. population, It was concluded that in the high density intense intraspecific competition for food resulted in a low net level of food availability per individual, which in turn inhibited the process of sexualisation in triclads from that area. The results from these two populations of ~~~!e~~~, indicating that net food availability controls the occurrence of sexual reproduction in this species, are in marked contrast to the findings of previous studies, in which habitat temperature is implicated as the dominant environmental influence on this process.

Lampreys (Petromyzontiformes) are an ancient vertebrate group, comprising 40 currently recognised species that range throughout the Northern and Southern Hemispheres. Despite a conserved morphology, lampreys nevertheless express a diverse range of life history strategies. Unusually for vertebrates larval lampreys are filter-feeding organisms prior to undergoing an extensive anatomical reorganisation, and the adoption of either a parasitic or a non-parasitic adult life. Parasitic lampreys consume the flesh and blood of actinopterygian fishes, either in marine or freshwater environments, while non-parasitic lampreys do not feed following their metamorphosis from the larval form. Morphological and genetic similarities between pairs of parasitic and non-parasitic lampreys have led to taxonomic confusion regarding the specific status of many non-parasitic forms, and the suggestion that the loss of the trophic adult phenotype is the result of a single species capable of producing alternative life history strategies. In this thesis it is argued that at least some paired species of lampreys do not comprise two distinct evolutionary lineages; rather, that non-parasitic lampreys represent one extreme in a continuum of life history variation expressed by a parasitic species. Some lamprey species, such as the European river lamprey Lampetra fluviatilis, are morphologically variable, exhibiting divergent phenotypes in response to ecological pressures, such as alternative foraging environments. Loch Lomond, Scotland contains a population of L. fluviatilis that feeds exclusively in the lake and exhibits a reduced body size and an overall morphology distinct from the typical anadromous form. Its foraging strategy indicates that it may be capable of switching hosts in the face of declining numbers of a presumed favoured and formerly abundant host, suggesting a certain amount of plasticity in its trophic ecology that may have ensured its survival in this freshwater lake. This freshwater-resident form, as well as anadromous L. fluviatilis and the non-parasitic species L. planeri, were found to spawn in a single river system within the Loch Lomond basin, and this site is crucial for the continued presence of this life history variant in Loch Lomond. The appearance of sexually mature specimens of three discrete phenotypes in this river, each representing an alternative life history strategy that may, or may not, belong to a single species, provides a crucial opportunity to test the strength of assortative mating between lamprey species pairs. Within this system the strength of assortative mating was found to be weak, and points to the possibility that freshwater-resident L. fluviatilis are mitigating gene flow between large anadromous parasitic L. fluviatilis, and small, non-parasitic L. planeri. As well as weak behavioural isolation, inter-specific sneak male mating tactics were documented among these populations, and represents the first time this phenomenon has been observed between paired lamprey species. Such behaviour indicates a lack of species-specific cues acting between L. fluviatilis and L. planeri, and suggests that hybrid offspring could be common in some systems. Testing hybrid viability (survivorship) between Loch Lomond’s two L. fluviatilis life history strategies and the sympatric L. planeri revealed no post-zygotic barriers to gene flow, at least in the form of gamete incompatibility. Perhaps more convincingly though, when comparing traditional morphometrics and body shape variation, as well as mitochondrial DNA sequences, between L. fluviatilis expressing different foraging strategies with populations of L. planeri, no robust species specific differentiation was observed. In fact, species delimitation between L. fluviatilis and L. planeri appears to be related solely to overall body size, which is itself a function of life history strategy. However, life history strategy was not correlated with current species designation as relationships among mtDNA haplotypes indicate non-parasitic populations have evolved independently multiple times throughout the geographic range of L. fluviatilis in Europe. Therefore, L. planeri should not be considered as a distinct species, either morphologically or genetically. Instead, L. fluviatilis appears capable of expressing a range of life history strategies; from parasitic anadromous populations through to non-parasitic stream-resident populations. The overall research approach employed in this thesis, i.e., the combination of ecological, behavioural, taxonomic and molecular studies, could be used to robustly examine the evolutionary ecology of parasitic and non-parasitic lampreys elsewhere.

Freshwater ostracods are potentially useful for environmental monitoring and, since their calcified valves may be preserved in lake sediments, are also valuable in palaeolimnological studies of environmental pollution such as eutrophication and surface water acidification. To establish a data-base of ostracod ecology, biannual survey work in 1989-1990 was performed in 75 lakes and tarns of a wide range of physico-chemical characteristics in the Lake District in Cumbria, ranging from large eutrophic lakes such as Windermere and Ullswater to tiny, upland acidic tarns. pH ranged from 4.3 (Black Pool) to 8.0 (Browns Tarn). Littoral margin samples were taken from all 75 sites and yielded 31 ostracod species, of which 8 were new to the Lake District fauna. The collection of multiple littoral samples from two sites demonstrated that a single sample produced an adequate faunal representation if it encompassed a variety of microhabitats. Statistical analysis, incorporating a multistage, multivariate technique, has shown that 18 species make up over 99% of the data set, and that 5 species, Cvpria ophthalmica, Cvclocypris ovum. Metacvpris cordata. Candona Candida and Cvpridopsis vidua. dominate the community in 71% of the sites containing ostracods, forming eight distinct assemblage groups. No ostracods were found in 13 of the 75 sites, 11 of which were acidic, having a pH of below 5.7. Equations were derived to predict both species distribution and diversity. Important predictors of community structures were shown to be pH, [Ca] 2+ , [Mg] 2+ , substrate, lake size and altitude. The equations were tested by further sampling of additional sites in the Lake District. Predictions of total species number and density generally provided an excellent fit to the observed data, although individual species predictions were poorer, especially in alkaline conditions. Substrate was not included in the analysis, due to quantitative difficulties, but this factor must be included in future predictive models as it was shown to be an important parameter in determining distribution. Deep-water sampling was carried out in 6 lakes. 10 species were collected, including Candona necrlecta, which was absent in the littoral samples. 9 species contributed to over 99% of the data set, and 2, Cypria ophthalmica and Candona Candida dominated the community in 75% of the sites containing ostracods. Community structure was predominantly determined by water depth (together with the associated temperature effect) and substrate. The sex ratio of Cyclocvpris ovum was shown be biased towards the female only at high alkalinities. Other species had sex ratios biased towards the male or female, the values independent of water quality. Large, swimming ostracod species were absent in sites containing fish. A series of laboratory experiments using three species of Ostracoda, (Cvpria ophthalmica. Cvpricercus fuscatus. and Eucvpris virens), and a predator (Gasterosteus aculeatus) correlated increasing ostracod size with an increased rate of predation, suggesting that predation could limit ostracod distribution. From the results of principal component analysis, it was concluded that the main characteristics that chemically differentiate the sites are calcium, magnesium, hydrogen and sodium ion concentrations. Toxicity tests were used to expose selected species to a wide range of calcium, magnesium, sodium and aluminium concentrations, at both neutral and acidic pH levels. Aluminium was selected as it has been highlighted as a major factor in the toxicity of acid waters. All species tolerated a wider spectrum of ionic concentrations than those in which they were recorded in the field, although the order of species survival in the experiments was similar to that found in the Lake District. It is suggested that whilst adult Ostracoda do not suffer from the acute toxicity of pH or aluminium, they may be unable to successfully reproduce in harsh environmental conditions. The waters of the English Lake District are not particularly species-rich due primarily to low alkalinity and low levels of dissolved cations, but also because they are cold. Only in small, ion-enriched pools is ostracod density sufficiently high to warrant their consideration as important detritivorous contributors in the cycling of nutrients. The Lake District fauna is compared with those recorded in other parts of Britain and Europe.

Membrane proteins in Xenopus laevis embryos were analysed on 1- and 2-dimensional gels. Proteins showing temporal variation in their expression in the membrane during Xenopus development were detected. The overall range of membrane proteins present between fertilisation and late neurula stage embryos was found to stay essentially constant, with few proteins showing temporal variation. Microinjection of 30 S-methionine into Xenopus laevis embryos was used to investigate membrane protein synthesis. Dramatic changes in the composition of proteins synthesised during Xenopus laevis early development were noted, with most significant changes observed at the gastrula stage following the mid-blastula transition. Cell membrane variation in Xenopus laevis embryos was studied using cell surface labelling with 125 Iodine. Two proteins specific to labelling of disaggregated embryos were detected and these are thought to be specific to intercellular membranes. Cell surface labelling with 125 Iodine was also used to characterise overall changes in the cell membrane during Xenopus laevis early development. Histo- autoradiography of 125 Iodine surface labelled embryos was used to confirm the origin of cleavage furrow membrane in the dividing embryo, and the source of ciliated cells from the double layered epidermis in Xenopus laevis neurulae. 2B12, a monoclonal antibody raised against adult Xenopus laevis brain tissue, was characterised on embryo and tissue sections using indirect immunofluoresence. Biochemical analysis of 2B12 activity has indicated that the antigen may be a ganglioside, specific to myelin and other nervous tissue membrane.

Homologues of developmentally important regulatory genes from other species have been found to be expressed in early amphibian development. The Drosophila dorso-ventral polarity .gene, dorsal, is involved in the formation of ventral structures and dorsal- mutants arise from a default state where only dorsal structures are formed. A Xenopus homologue of dorsal may play a crucial role in interpreting some of the earliest embryonic signals.

The main aim of this work was to identify the important cis-acting regulatory sequences, and the trans-acting factors with which they interact, which are required for the tissue-specific expression of the Xenopus borealis skeletal actin gene. All sequences necessary and sufficient for the correct spatial and temporal expression of the Xenopus borealis skeletal actin gene are located in a 156 bp fragment of the gene that spans from nucleotide residues -197 to -42 in its promoter. This region of the skeletal actin promoter contains three imperfect repeats of a CArG sequence motif that has been demonstrated to be important in the expression of other sarcomeric actin genes. Deletion analysis of the promoter of the Xenopus borealis skeletal actin gene, using Xenopus micro-injection techniques as a transient assay system for promoter activity, have identified that CArG box3 is essential for skeletal actin gene expression. By using band shift assays I have demonstrated that, under my assay conditions, CArG box2 is unable to bind any proteins in vitro . Conversely, the CArG boxl sequence exhibits two binding activities on band shift analysis. One of these is antigenically related to the transcription factor SRF, whilst the second appears to be distinct from this protein. CArG box3 also interacts with a protein in vitro. Although this sequence exhibits a similar shift to that of the CArG boxl/SRF complex on band shift analysis, my experiments suggest that this protein is distinct from SRF. A combination of the CArG boxl and CArG box3 motifs is unable to confer muscle-specific gene expression on a heterologous promoter. Furthermore, I have identified an upstream regulatory element (URE) in the Xenopus borealis skeletal actin gene promoter that spans from nucleotides -197 to -167 that is required for the expression of the gene, at least when sequences between nucleotide -42 and +28 are absent. The URE of the Xenopus borealis skeletal actin gene is capable of interacting with a trans-acting factor(s) in vitro. In addition to this a further region of the gene which spans from nucleotide residues -83 to -42 is also capable of interacting with a factor(s) in vitro. The mechanisms by which these multiple regulatory elements control the tissue-specific expression of the Xenopus borealis skeletal actin gene will be discussed.