VictorJuiz/thesis huggingface dataset:
Calcium and chloride dynamics in immature neurons and their role in dendritic growth

Activity-dependent dendritic development is an important process in the maturation of neuronal circuits. The precise morphology of a neuron’s dendritic tree dictates which other cells it is able to interact with and how it will receive and process synaptic information. The aim of this Thesis was to investigate the mechanisms by which dendrites grow and, in particular, how changes in intracellular ion concentrations contribute to these mechanisms. One important activity-dependent signal is calcium as it can translate neuronal activity into morphological changes. Despite this, very little is known about calcium signalling during the period of dendritic development. Using single-cell electroporation of immature CA1 hippocampal pyramidal neurons, I characterised the spatial and temporal properties of local calcium transients in growing dendrites. This revealed a high frequency of transients at shaft filopodia and stable branchpoints, but an almost complete absence from the tips of dendritic branches. Another important factor during development is the intracellular chloride concentration because this regulates neuronal excitability. Prematurely lowering intracellular chloride by expressing the chloride co-transporter KCC2 led to less stable dendritic filopodia and stunted dendritic growth. These effects were independent of local calcium signalling and suggested that chloride regulation itself may be fundamental to normal dendritic growth. To examine this further I developed imaging techniques to measure the spatial and temporal dynamics of chloride in growing dendrites. This work revealed a somatodendritic gradient of increasing intracellular chloride, whereby the highest concentrations were found at sites of growth. Further analysis suggested a close link between local chloride regulation and morphological changes. The dendritic tips that exhibited high intracellular chloride levels and the potential to rapidly modulate these levels, also exhibited the greatest morphological dynamics. These findings have important implications for understanding the mechanisms of dendritic growth and establish the spatiotemporal regulation of chloride as a key parameter.
Photoelectrochemistry

The purpose of this work was to assess the potential of the Ru(II)(bipy)3 complexes as photogalvanic systems, using the criteria obtained from the theory for an ideal photogalvanic solar cell developed by Albery and Archer. To this end two techniques were developed to measure the rate of the thermal back reaction between the high energy photoproducts Ru(III) and Fe(II) The thermal back reaction is a serious loss process in such a cell.
The first technique, the Optical Rotating Disc Electrode, was improved by the design and construction of new apparatus. It allowed measurement of most of the important parameters of a photogalvanic cell and confirmed much of the theory for such a cell. The second technique, Flash Electrolysis, first used by Perone, was developed for use with a transparent disc electrode. Further investigations of the thermal back reaction were made using the Stopped Flow technique.
The results of these experiments on Ru(IIl)(bipy)3 and its derivatives show that the reaction obeys the Marcus theory of electron transfer. However the activation parameters are dominated by "compensation effects" which, at the elevated temperatures of a practical photogalvanic cell (> 60°C) , reduce the differences between different derivatives. The very similar rates of reaction so produced are much too high for the development of an efficient photogalvanic cell. Ru(II) (bipy)3 systems thus provide a useful model for other possible photogalvanic systems buc cannot be of any practical use themselves.
Hate crimes hurt more: Can restorative practices help repair the harms?

The current retributive approach to tackling hate crime, while intuitively grounded in the principle of proportionately, does little to either repair the harms caused by incidents of hate or engender greater levels of acceptance of those deemed as “different”. This thesis therefore explores whether restorative justice, a relatively new theory and practice of criminal justice, is better placed to tackle the causes and consequences of hate victimisation.
The 18 month empirical study, carried out to examine the thesis’ aims, uses a triangulation approach by incorporating observations of restorative justice meetings, semi-structured interviews with victim participants and semi-structured interviews with restorative practitioners who have experience facilitating hate crime cases. The mainly qualitative data collated provides for a detailed evaluation of the various processes found within restorative practices that: 1) helped to alleviate the distress caused by hate victimisation and 2) prevented the recurrence of hate-motivated incidents.
A broad conceptualisation of hate crime was used within the thesis that included “hate incidents”. This allowed me to explore the utility of restorative practices in cases involving serious violence and the more pervasive “low-level”, but nonetheless highly deleterious, non-criminal incidents of hate that are frequently committed against minority group individuals.
There were also several unanticipated findings from the study. First, data emerged which highlighted various aspects of the restorative practice which were unforeseen as being central to the successful application of restorative processes, these are discussed throughout the thesis. Second, great insight was gained into the nature of hate victimisation, helping to unravel some of the complex socio-cultural factors pivotal to both the cause and effect of hate victimisation. It is hoped that these additional findings provide important epistemological advancements in both fields of study.
A study of the Bloom's syndrome protein

Bloom's syndrome is a rare autosomal recessive disorder characterised by an early onset of cancer of many types, erythematous lesions on sun-exposed skin, retarded growth, immunodeficiency and sub- or infertility. Cells from Bloom's syndrome patients have replication defects and an abnormally unstable genome manifested in chromosomal breaks and deletions and in an increased mutation rate. Most characteristically, these cells show elevated levels of sister-chromatid exchanges which probably result from homologous recombination events. Since the cells are not hypersensitive to DNA damaging agents, the defect is unlikely to be in one of the common DNA repair pathways. The gene mutated in Bloom's syndrome, BLM, was cloned in 1995 and found to encode a helicase from the RecQ family. This family is named after its E. coli member, RecQ, and includes at least five human genes. Three of these are mutated in inherited disorders; Bloom's syndrome, Werner's syndrome and Rothmund-Thomson syndrome.
In my DPhil project, I have investigated the enzymatic properties of the BLM protein. I have purified the protein in recombinant form and shown that it is a DNA-dependent ATPase and an ATP-dependent helicase with 3'-5' polarity. It binds and unwinds a variety of DNA structures, with a preference for tetraplex (G4)-DNA, Holliday junctions (recombination intermediates) and internal DNA bubbles. Furthermore, it is capable of branch migration, an activity distinct from its helicase activity. BLM forms oligomeric rings with fourfold and sixfold symmetry, both in a cell extract and as purified protein.
These results, in combination with the cellular phenotype of Bloom's syndrome and with evidence from the analysis of other RecQ homologues in model organisms such as yeast and E. coli, point to a role for BLM in somatic recombination (recombinational repair). Models for this function are discussed in this thesis.
Investigating the presynaptic control of striatal dopamine release

Dopamine (DA) is a key neuromodulator in the striatum, and is important for action selection and reinforcement learning. Dysfunctions in striatal DA signalling contribute to numerous disorders including Parkinson’s disease (PD) and drug addiction.
Midbrain DA neurons switch from low to high frequency firing in response to reward-related events, which is proposed to increase striatal DA release. However, in addition to DA neuron firing pattern, striatal DA signalling depends upon the short-term plasticity of DA release, which is controlled by presynaptic and local network factors.
This thesis uses fast-scan cyclic voltammetry, in murine striatal slices, to detect subsecond changes in extracellular DA, and investigate the presynaptic control of striatal DA release and release plasticity. Acetylcholine from striatal cholinergic interneurons, acting at nicotinic receptors (nAChRs) on DA terminals, is one factor that strongly influences DA release. This thesis particularly explores how presynaptic factors interact with nAChRs to control DA release.
Firstly, release probability, a key determinant of release plasticity at many synapses, was found to be only weakly related to DA release plasticity, and only when nAChRs are inactive. Secondly, a direct role of the DA uptake transporter (DAT) in controlling DA release plasticity was identified, when nAChRs are inactive. Thirdly, regional differences were identified in the role of the DAT in controlling DA release via control of D2 receptor activation, when nAChRs are active. Finally, mutant α-synuclein, which causes PD in humans, was found to only subtly affect striatal DA release.
These data suggest that the control of striatal DA release differs substantially from other central transmitters. Release probability and α-synuclein play only minor roles, but nAChRs and the DAT significantly control DA release plasticity. These findings review our understanding of striatal DA release and may have implications for understanding the actions of drugs of abuse and early PD pathogenesis.
The axial capacity of driven piles in clay

An instrumented model pile was used to investigate the fundamental behaviour in clay soils of driven cylindrical steel piles used for offshore structures. Four test-bed sites were chosen; two in stiff heavily consolidated clays, and two in normally/lightly overconsolidated clays. Data from these sites confirm that a residual shear surface is formed along the pile during installation, the location of which relative to the shaft surface appears to depend on the shaft roughness. Comparisons with other site investigation data and cavity expansion theoretical predictions indicate that stress relief immediately behind the pile tip during driving gives rise to total radial stresses and pore pressures measured on the pile shaft which are lower than predicted. This stress relief is particulary severe in the stiffer clays. The data did however show that the installation total radial stresses and pore pressures are governed by the initial in-situ stresses and undrained shear strength as is predicated by the theory. During reconsolidation, pore pressures close to the instrument rise initially in all clays, and radial effective stresses drop. The slow recovery in radial effective stress during the later stages of reconsolidation was in some cases insufficient to return it to levels recorded during installation. However, the generation of negative pore pressures during undrained loading increased the radial effective stress and shaft friction at failure. This effect is particularly important in the normally consolidated clays, and is responsible for the set-up of shaft capacity seen in such clays, which might not be observed if the loading were drained. The observed behaviour was therefore quite different from the monotonic increase in radial effective stress during reconsolidation, followed by decrease during undrained loading which was expected from a review of current theory.
The evolution of international inequality

Within the contemporary international order, deep structural inequalities coexist alongside a nominally pluralistic society of states that grants international personality to politically organised communities. Asymmetric interactions between distinct political communities have shaped the development of the international system from the colonial era to the present phase of global economic integration. Rising interdependence, problems of unequal development and the democratic mobilisation of peoples around the world have generated moral claims regarding the injustice of global inequality.
In this context the international politics of inequality have taken the format of challenges by the political representatives of the global South to the dominance of the advanced industrialised North. The normative dimensions of this process can be understood through a focus on this process of political argument between unequals. Political argument is contestation over the principles appropriate to govern a sphere of social interaction. The thesis seeks to vindicate the notion that the challenges by the global South have given rise to a dynamic of political argument within a norm-governed international society. Changes in patterns of normative belief, material power and forms of political organisation have historically shaped North-South relations. Therefore, through the analysis of particular episodes of North-South argument, the thesis attempts to provide insights into the moral limits and possibilities of an evolving international society.
Analysing the organised attempts to challenge inequality on the part of the representatives of the global South, the thesis seeks to advance the position the tensions generated by claims over inequality might provide the nucleus for the incorporation of egalitarian concerns into the operation of international society. Through participation in common practices of statehood, the peoples of the global South possess at least some ability to challenge structural inequalities and thus the potential to expand the moral limits of international society.
Studies on an N-terminal nucleophile hydrolase and enzymes of clavulanic acid biosynthesis

(3R,5R)-Clavulanic acid is a clinically important inhibitor of Class A β-lactamases. Progress has been made in to establishing the steps of clavulanic acid biosynthesis leading to (3S,5S)-clavaminic acid. However, the mechanism by which (3S,5S)-clavaminic acid is converted to the penultimate intermediate (3R,5R)-clavaldehyde remains elusive. It is believed that the products of the later genes (orf10-orf18) of the clavulanic acid biosynthesis gene cluster are probably involved in this conversion.
Part I of this thesis describes biochemical and structural studies carried out on OAT2, a member of N-terminal nucleophile (Ntn) hydrolase superfamily of enzymes. OAT2 has been characterised to be an ornithine acetyl transferase (OAT) and is involved in clavulanic acid biosynthesis. OAT2 catalyses the reversible transfer of the acetyl group between N-acetyl-L-ornithine and L-glutamate. It was found that this reaction is catalysed via the formation of an acyl-enzyme intermediate. Subsequent studies including mass spectrometry, 13C NMR spectroscopy, infrared spectroscopy, X-ray crystallography and molecular dynamics simulations, further confirmed the viability of the intermediate. This acyl-enzyme intermediate of OAT2 was found to be exceptionally stable at physiological pH, as compared to the acyl-enzyme intermediates involved in catalysis by hydrolytic enzymes including proteases, Ntn hydrolases and β-lactamses. The X-ray studies revealed possible reason for this unusual stability. The infrared studies revealed two conformations for the acyl-enzyme. Modeling (MDS) studies assigned one of these to the structure observed by X-ray and proposed the other one to result from a hydroxyl hydrogen 'flip' involving the oxyanion hole component Thr-111 resulting in a singly hydrogen bonded acyl-enzyme intermediate. α, β Subunit co-expression studies with OAT2 were used to investigate the autocatalytic cleavage step. In one case an interesting N-acyl enzyme species was observed.
Part II of this thesis describes efforts carried out to characterise the ORF10 and ORF15 proteins of clavulanic acid biosynthesis. ORF10 was characterised to be an 'active' cytochrome P450 and ORF10 crystals were obtained in the presence spinach ferredoxin, highlighting the role of the ferredoxin interaction in assisting ORF10 crystallisation. ORF15 was shown to be a probable peptide transporter, which binds bradykinin as observed in the crystal structure.
Solitary waves and wave groups at the shore

A significant proportion of the world's population and physical assets are located in low lying coastal zones. Accurate prediction of wave induced run-up and overtopping of sea defences are important in defining the extent and severity of wave action, and in assessing risk to people and property from severe storms and tsunamis.
This thesis describes a one-dimensional numerical model based on the Boussinesq equations of Madsen and Sorensen (1992) and the non-linear shallow water equations. The model is suitable for simulating propagation of weakly non-linear and weakly dispersive waves from intermediate to zero depth, such that any inundation and/or overtopping caused by the incoming waves is also calculated as part of the simulation. Wave breaking is approximated by locally switching to the non-linear shallow water equations, which can model broken waves as bores. A piston paddle wavemaker is incorporated into the model for complete reproduction of laboratory experiments. A domain mapping technique is used in the vicinity of the paddle to transform a time-varying domain into a fixed domain, so that the governing equations can be more readily solved.
First, various aspects of the numerical model are verified against known analytical and newly derived semi-analytical solutions. The complete model is then validated with laboratory measurements of run-up and overtopping involving solitary waves. NewWave focused wave groups, which give the expected shape of extreme wave events in a linear random sea, are used for further validation. Simulations of experiments of wave group run-up on a plane beach yield very good agreement with the measured run-up distances and free surface time series. Wave-by-wave overtopping induced by focused wave groups is also successfully simulated with the model, with satisfactory agreement between the experimental and the predicted overtopping volumes. Repeated simulations, now driven by second order paddle displacement signals, give insight into second order error waves spuriously generated by using paddle signals derived from linear theory. Separation of harmonics reveals that the long error wave is significantly affecting the wave group shape and leading to enhanced runu-up distances and overtopping volumes. An extensive parameter study is carried out using the numerical model investigating the influence on wave group run-up of linear wave amplitude at focus, linear focus location, and wave group phase at focus. For a given amplitude, both the phase and the focus location significantly affect the wave group run-up. It is also found that the peak optimised run-up increases with the wave amplitude, but wave breaking becomes an inhibiting factor for larger waves. This methodology is proposed for extreme storm wave induced run-up analysis.
Biochemical and biophysical studies of the prokaryotic proton dependent oligopeptide transporters

The proton dependent oligopeptide transporters (POT family) are members of the Major Facilitator Superfamily of secondary active transporter proteins. They use the transmembrane proton gradient to drive the uptake of di- and tripeptides into the cytoplasm. Members of the family are highly conserved in pro- and eukaryotic genomes, and in humans they are responsible for the oral absorption of many drug families, including β-lactam antibiotics. Recently, the crystal structures of PepTSo and PepTSt, two prokaryotic homologues of the human proteins PepT1 and PepT2, captured the proteins in two distinct conformations, providing insight into the structural aspects of the transport mechanism. A protocol was designed for functional liposome reconstitution of POT proteins, and transport assays were conducted to characterise their substrate specificity, pH dependence and kinetic properties. Using site-directed mutagenesis, we identified binding site residues involved in peptide recognition and proton translocation, and distinguished between the two roles by comparing protein activity in proton- and peptide-driven conditions. We also investigated the roles of key residues in the conformational transitions that accompany the transport cycle, using data from biochemical assays, molecular dynamics simulations and modeling, as well as electron paramagnetic resonance measurements. In addition, several bacterial POT members were screened for crystallisation, in order to assess their stability and crystal diffraction quality in different detergents. Further work was performed with bacterial POT homologues YdgR and GkPOT, including binding studies using NMR spectroscopy and assaying drug transport in vivo and in vitro. Together, the data establish bacterial POTs as model systems for studying the mammalian oligopeptide transporters, and a mechanistic model for peptide transport is proposed.
Measurement of charmed hadron production in e±p deep inelastic scattering with the ZEUS detector

Charmed hadron production has been studied in 225 pb-1 of data collected with the ZEUS detector in the HERA-II runnning period. Studies were made in the D* → D0π → Kππ decay channel to determine the potential for the microvertex detector to improve charm signals. Integrated cross sections were measured for the decays D0 → K0sπ⁺ π⁻, D± → K0s π±, D±_s → K0s K± and Lambda±_c → Lambda0π± in the kinematic region 5< Q² < 1000 GeV², 0.02 < y < 0.7, 1.5 < pT(D,Lambda_c) < 20.0 GeV, -1.6 < pseudorapidity(D, Lambda_c) < 1.6. The D0 cross section was additionally measured down to pT(D0) > 1.0 GeV. Differential cross sections as a function of Q², x, pT(D) and pseudorapidity(D) were measured for the D0, D±, and D_s± channels. The measurements were compared to next-to-leading order (NLO) QCD calculations, and good agreement found for all variables in all channels over the measured kinematic region. The strangeness-suppression factor, gamma_s = D_s±/(D± + D0), was measured and found to be in good agreement with values previously measured at HERA and elsewhere, and with the DJANGOH and RAPGAP Monte Carlo. The ratio of charmed baryons to mesons, Lambda_c±/(D± + D0), was measured and also found to be in good agreement with Monte Carlo.