Frequency selectivity and dopamine-dependence of plasticity at glutamatergic synapses in the subthalamic nucleus

In Parkinson's disease, subthalamic nucleus (STN) neurons burst fire with increased periodicity and synchrony. This may entail abnormal release of glutamate, the major source of which in STN is cortical afferents. Indeed, the cortico-subthalamic pathway is implicated in the emergence of excessive oscillations, which are reduced, as are symptoms, by dopamine-replacement therapy or deep brain stimulation (DBS) targeted to STN. Here we hypothesize that glutamatergic synapses in the STN may be differentially modulated by low-frequency stimulation (LFS) and high-frequency stimulation (HFS), the latter mimicking deep brain stimulation. Recordings of evoked and spontaneous excitatory post synaptic currents (EPSCs) were made from STN neurons in brain slices obtained from dopamine-intact and chronically dopamine-depleted adult rats. HFS had no significant effect on evoked (e) EPSC amplitude in dopamine-intact slices (104.4±8.0%) but depressed eEPSCs in dopamine-depleted slices (67.8±6.2%). Conversely, LFS potentiated eEPSCs in dopamine-intact slices (126.4±8.1%) but not in dopamine-depleted slices (106.7±10.0%). Analyses of paired-pulse ratio, coefficient of variation, and spontaneous EPSCs suggest that the depression and potentiation have a presynaptic locus of expression. These results indicate that the synaptic efficacy in dopamine-intact tissue is enhanced by LFS. Furthermore, the synaptic efficacy in dopamine-depleted tissue is depressed by HFS. Therefore the therapeutic effects of DBS in Parkinson's disease appear mediated, in part, by glutamatergic cortico-subthalamic synaptic depression and implicate dopamine-dependent increases in the weight of glutamate synapses, which would facilitate the transfer of pathological oscillations from the cortex.

Studies of recognition and selectivity by fluorescent polymers imprinted with N1-benzylidene pyridine-2-carboxamidrazones

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Ionic and molecular diffusion in cementitious materials

The work described in this thesis is an attempt to provide improved understanding of the effects of several factors affecting diffusion in hydrated cement pastes and to aid the prediction of ionic diffusion processes in cement-based materials. Effect of pore structure on diffusion was examined by means of comparative diffusion studies of quaternary ammonium ions with different ionic radii. Diffusivities of these ions in hydrated pastes of ordinary portland cement with or without addition of fly ash were determined by a quasi-steady state technique. The restriction of the pore geometry on diffusion was evaluated from the change of diffusivity in response to the change of ionic radius. The pastes were prepared at three water-cement ratios, 0.35, 0.50 and 0.65. Attempts were made to study the effect of surface charge or the electrochemical double layer at the pore/solution interface on ionic diffusion. An approach was to evaluate the zeta potentials of hydrated cement pastes through streaming potential measurements. Another approach was the comparative studies of the diffusion kinetics of chloride and dissolved oxygen in hydrated pastes of ordinary portland cement with addition of 0 and 20% fly ash. An electrochemical technique for the determination of oxygen diffusivity was also developed. Non-steady state diffusion of sodium potassium, chloride and hydroxyl ions in hydrated ordinary portland cement paste of water-cement ratio 0.5 was studied with the aid of computer-modelling. The kinetics of both diffusion and ionic binding were considered for the characterization of the concentration profiles by Fick's first and second laws. The effect of the electrostatic interactions between ions on the overall diffusion rates was also considered. A general model concerning the prediction of ionic diffusion processes in cement-based materials has been proposed.

The synthesis and application of room temperature ionic liquids

This research project is concerned with the development and use of eco-friendly reaction media for a variety of organic transformations in the preparation of organic chemicals with potential pharmaceutical applications. These chemicals will then be investigated for their anti-cancer, anti-bacterial and anti-inflammation properties. In this project, different methods were used to synthesize various kinds of ionic liquids. Some new ionic liquids were prepared. In addition, Knoevenagel condensation reactions were investigated in RTILs. For the first time, some neutral ionic liquids such as [BMIM]+[BF4]-, [MeOEtMIM]+[CF3COO]- acted as both catalysts and solvents to promote Knoevenagel reactions. All these experiments indicated that RTILs have a great potential as alternative solvents in synthetic chemistry. Furthermore, nucleoside chemistry is an important research area in drug discovery. Various chemical modified nucleosides have therapeutic activities. However, these compounds usually have poor solubility in common organic solvents. RTILs such as [MeOEtMIM]+[CH3SO3]- have good dissolving capability for these chemicals. A range of thio-substituted nucleobases and nucleosides with potential pharmaceutical applications have been synthesized in several RTILs. These chemicals will then be investigated for their anti-cancer properties.

Ionic and neutral hydrogels for dermal and ophthalmic applications

This thesis is concerned with the use of ionic and neutral hydrogels in dermal and ocular applications with particular reference to controlled release applications. The work consists of three interconnected themes.The first area of study is the use of skin adhesive bioelectrode hydrogels as ground plate electrodes for ophthalmic iontophoresis applications. The work provides a basis of understanding the relative contributions made by ionic monomers (such as sodium s-(acrylamide)-2-methyl propane sulphonate and acrylic acid-bis-(3-sulfopropyl-ester, potassium salt) and neutral monomers (such as acryloymorpholine, N,N-dimethylacrylamide and N-vinyl pyrrolidone) to adhesion, rheology and impedance of bioelectrode gels. The general advantage of neutral monomers, which have been used to successfully replace ionic monomers, is that they enable more effective control of independent anion and cation species (for example potassium chloride and sodium chloride) unlike ionic monomers where polymerisation produces an immobile polyanion thus limiting cation mobility. Secondly, release from a completely neutral hydrogel under the influence of mechanical shaking was studied for the case of crosslinked polyvinyl alcohol (PVA) containing low concentration of linear soluble PVA in a contact lens application. The soluble PVA was observed to be eluting by reptation from the lens matrix due to the mechanical action of the eyelid. This process was studied in an in vitro model, which in this research was used as a basis for developing a lens made with enhanced release polymer. The third area of work is related to the factors that control drug release (in particular non-steroidal anti-inflammatory drugs) from a hydrogel matrix. This links both electrotherapy applications, such as transcutaneous electrical nerve stimulation, in which the passive diffusion from the gel could be used in conjunction with enhanced transmission across the dermal surface with passive diffusion from a contact lens matrix and the development of therapeutic contact lenses.

Protein-protein interactions in ovalbumin solutions studied by small angle scattering: effect of ionic strength, and the chemical nature of cations

The influence of ionic strength and of the chemical nature of cations on the protein-protein interactions in ovalbumin solution was studied using small-angle X-ray and neutron scattering (SAXS/SANS). The globular protein ovalbumin is found in dimeric form in solutions as suggested by SANS/SAXS experiments. Due to the negative charge of the proteins at neutral pH, the protein-protein interactions without any salt addition are dominated by electrostatic repulsion. A structure factor related to screened Coulombic interactions together with an ellipsoid form factor was used to fit the scattering intensity. A monovalent salt (NaCl) and a trivalent salt (YCl3) were used to study the effect of the chemical nature of cations on the interaction in protein solutions. Upon addition of NaCl, with ionic strength below that of physiological conditions (150 mM), the effective interactions are still dominated by the surface charge of the proteins and the scattering data can be understood using the same model. When yttrium chloride was used, a reentrant condensation behavior, i.e., aggregation and subsequent redissolution of proteins with increasing salt concentration, was observed. SAXS measurements reveal a transition from effective repulsion to attraction with increasing salt concentration. The solutions in the reentrant regime become unstable after long times (several days). The results are discussed and compared with those from bovine serum albumin (BSA) in solutions.

Reaction-driven surface restructuring and selectivity control in allylic alcohol catalytic aerobic oxidation over Pd

Synchronous, time-resolved DRIFTS/MS/XAS cycling studies of the vapor-phase selective aerobic oxidation of crotyl alcohol over nanoparticulate Pd have revealed surface oxide as the desired catalytically active phase, with dynamic, reaction-induced Pd redox processes controlling selective versus combustion pathways.

Selectivity and timing performance of UK investment trusts

This study examines the selectivity and timing performance of 218 UK investment trusts over the period July 1981 to June 2009. We estimate the Treynor and Mazuy (1966) and Henriksson and Merton (1981) models augmented with the size, value, and momentum factors, either under the OLS method adjusted with the Newey-West procedure or under the GARCH(1,1)-in-mean method following the specification of Glosten et al. (1993; hereafter GJR-GARCH-M). We find that the OLS method provides little evidence in favour of the selectivity and timing ability, consistent with previous studies. Interestingly, the GJR-GARCH-M method reverses this result, showing some relatively strong evidence on favourable selectivity ability, particularly for international funds, as well as favourable timing ability, particularly for domestic funds. We conclude that the GJR-GARCH-M method performs better in evaluating fund performance compared with the OLS method and the non-parametric approach, as it essentially accounts for the time-varying characteristics of factor loadings and hence obtains more reliable results, in particular, when the high frequency data, such as the daily returns, are used in the analysis. Our results are robust to various in-sample and out-of-sample tests and have valuable implications for practitioners in making their asset allocation decisions across different fund styles. © 2012 Elsevier B.V.

Investigating the role of cholesterol in the formation of non-ionic surfactant based bilayer vesicles:thermal analysis and molecular dynamics

The aim of this research was to investigate the molecular interactions occurring in the formulation of non-ionic surfactant based vesicles composed monopalmitoyl glycerol (MPG), cholesterol (Chol) and dicetyl phosphate (DCP). In the formulation of these vesicles, the thermodynamic attributes and surfactant interactions based on molecular dynamics, Langmuir monolayer studies, differential scanning calorimetry (DSC), hot stage microscopy and thermogravimetric analysis (TGA) were investigated. Initially the melting points of the components individually, and combined at a 5:4:1 MPG:Chol:DCP weight ratio, were investigated; the results show that lower (90 C) than previously reported (120-140 C) temperatures could be adopted to produce molten surfactants for the production of niosomes. This was advantageous for surfactant stability; whilst TGA studies show that the individual components were stable to above 200 C, the 5:4:1 MPG:Chol:DCP mixture show ∼2% surfactant degradation at 140 C, compared to 0.01% was measured at 90 C. Niosomes formed at this lower temperature offered comparable characteristics to vesicles prepared using higher temperatures commonly reported in literature. In the formation of niosome vesicles, cholesterol also played a key role. Langmuir monolayer studies demonstrated that intercalation of cholesterol in the monolayer did not occur in the MPG:Chol:DCP (5:4:1 weight ratio) mixture. This suggests cholesterol may support bilayer assembly, with molecular simulation studies also demonstrating that vesicles cannot be built without the addition of cholesterol, with higher concentrations of cholesterol (5:4:1 vs 5:2:1, MPG:Chol:DCP) decreasing the time required for niosome assembly. © 2013 Elsevier B.V.

Consideration of the efficacy of non-ionic vesicles in the targeted delivery of oral vaccines

The fundamentals of this research were to exploit non-ionic surfactant technology for delivery and administration of vaccine antigens across the oral route and to gain a better understanding of vaccine trafficking. Using a newly developed method for manufacture of non-ionic surfactant vesicles (niosomes and bilosomes) lower process temperatures were adopted thus reducing antigen exposure to potentially damaging conditions. Vesicles prepared by this method offered high protection to enzymatic degradation, with only ~10 % antigen loss measured when vesicles incorporating antigen were exposed to enzyme digestion. Interestingly, when formulated using this new production method, the addition of bile salt to the vesicles offered no advantage in terms of stability within simulated gastro-intestinal conditions. Considering their ability to deliver antigen to their target site, results demonstrated that incorporation of antigen within vesicles enhanced delivery and targeting of the antigen to the Peyer's Patch, again with niosomes and bilosomes offering similar efficiency. Delivery to both the Peyer's patches and mesentery lymphatics was shown to be dose dependent at lower concentrations, with saturation kinetics applying at higher concentrations. This demonstrates that in the formulation of vaccine delivery systems, the lipid/antigen dose ratio is not only a key factor in production cost, but is equally a key factor in the kinetics of delivery and targeting of a vaccine system. © 2013 Controlled Release Society.

Protein interactions studied by SAXS:effect of ionic strength and protein concentration for BSA in aqueous solutions

We have studied a series of samples of bovine serum albumin (BSA) solutions with protein concentration, c, ranging from 2 to 500 mg/mL and ionic strength, I, from 0 to 2 M by small-angle X-ray scattering (SAXS). The scattering intensity distribution was compared to simulations using an oblate ellipsoid form factor with radii of 17 x 42 x 42 A, combined with either a screened Coulomb, repulsive structure factor, S-SC(q), or an attractive square-well structure factor, S-SW(q). At pH = 7, BSA is negatively charged. At low ionic strength, I <0.3 M, the total interaction exhibits a decrease of the repulsive interaction when compared to the salt-free solution, as the net surface charge is screened, and the data can be fitted by assuming an ellipsoid form factor and screened Coulomb interaction. At moderate ionic strength (0.3-0.5 M), the interaction is rather weak, and a hard-sphere structure factor has been used to simulate the data with a higher volume fraction. Upon further increase of the ionic strength (I >= 1.0 M), the overall interaction potential was dominated by an additional attractive potential, and the data could be successfully fitted by an ellipsoid form factor and a square-well potential model. The fit parameters, well depth and well width, indicate that the attractive potential caused by a high salt concentration is weak and long-ranged. Although the long-range, attractive potential dominated the protein interaction, no gelation or precipitation was observed in any of the samples. This is explained by the increase of a short-range, repulsive interaction between protein molecules by forming a hydration layer with increasing salt concentration. The competition between long-range, attractive and short-range, repulsive interactions accounted for the stability of concentrated BSA solution at high ionic strength.

Consideration of the efficacy of non-ionic vesicles in the targeted delivery of oral vaccines

The fundamentals of this research were to exploit non-ionic surfactant technology for delivery and administration of vaccine antigens across the oral route and to gain a better understanding of vaccine trafficking. Using a newly developed method for manufacture of non-ionic surfactant vesicles (niosomes and bilosomes) lower process temperatures were adopted thus reducing antigen exposure to potentially damaging conditions. Vesicles prepared by this method offered high protection to enzymatic degradation, with only ~10 % antigen loss measured when vesicles incorporating antigen were exposed to enzyme digestion. Interestingly, when formulated using this new production method, the addition of bile salt to the vesicles offered no advantage in terms of stability within simulated gastro-intestinal conditions. Considering their ability to deliver antigen to their target site, results demonstrated that incorporation of antigen within vesicles enhanced delivery and targeting of the antigen to the Peyer's Patch, again with niosomes and bilosomes offering similar efficiency. Delivery to both the Peyer's patches and mesentery lymphatics was shown to be dose dependent at lower concentrations, with saturation kinetics applying at higher concentrations. This demonstrates that in the formulation of vaccine delivery systems, the lipid/antigen dose ratio is not only a key factor in production cost, but is equally a key factor in the kinetics of delivery and targeting of a vaccine system. © 2013 Controlled Release Society.

Selectivity control in Pt-catalyzed cinnamaldehyde hydrogenation

Chemoselectivity is a cornerstone of catalysis, permitting the targeted modification of specific functional groups within complex starting materials. Here we elucidate key structural and electronic factors controlling the liquid phase hydrogenation of cinnamaldehyde and related benzylic aldehydes over Pt nanoparticles. Mechanistic insight from kinetic mapping reveals cinnamaldehyde hydrogenation is structure-insensitive over metallic platinum, proceeding with a common Turnover Frequency independent of precursor, particle size or support architecture. In contrast, selectivity to the desired cinnamyl alcohol product is highly structure sensitive, with large nanoparticles and high hydrogen pressures favoring C=O over C=C hydrogenation, attributed to molecular surface crowding and suppression of sterically-demanding adsorption modes. In situ vibrational spectroscopies highlight the role of support polarity in enhancing C=O hydrogenation (through cinnamaldehyde reorientation), a general phenomenon extending to alkyl-substituted benzaldehydes. Tuning nanoparticle size and support polarity affords a flexible means to control the chemoselective hydrogenation of aromatic aldehydes.

A simple and efficient procedure for Knoevenagel reaction promoted by imidazolium-based ionic liquids

Various room temperature ionic liquids (RTILs), notably, 1-methoxyethyl-3-methylimidazolium trifluoroacetate [MeOEtMIM]+[CF3COO]ˉ , have been used to promote the Knoevenagel condensation to afford substituted olefins. All reactions proceeded effectively in the absence of any other catalysts or co-solvents with good to excellent yields. This method is simple and applicable to reactions involving a wide range of aldehydes and ketones with methylene compounds. The ionic liquid can be recycled without noticeable reduction of its catalytic activity. A plausible reaction mechanism is proposed.

Tailoring heme -thiolate proteins into efficient biocatalysts with high specificity and selectivity

Cytochrome P450 monooxygenases, one of the most important classes of heme-thiolate proteins, have attracted considerable interest in the biochemical community because of its catalytic versatility, substrate diversity and great number in the superfamily. Although P450s are capable of catalyzing numerous difficult oxidation reactions, the relatively low stability, low turnover rates and the need of electron-donating cofactors have limited their practical biotechnological and pharmaceutical applications as isolated enzymes. The goal of this study is to tailor such heme-thiolate proteins into efficient biocatalysts with high specificity and selectivity by protein engineering and to better understand the structure-function relationship in cytochromes P450. In the effort to engineer P450cam, the prototype member of the P450 superfamily, into an efficient peroxygenase that utilizes hydrogen peroxide via the “peroxide-shunt” pathway, site-directed mutagenesis has been used to elucidate the critical roles of hydrophobic residues in the active site. Various biophysical, biochemical and spectroscopic techniques have been utilized to investigate the wild-type and mutant proteins. Three important P450cam variants were obtained showing distinct structural and functional features. In P450camV247H mutant, which exhibited almost identical spectral properties with the wild-type, it is demonstrated that a single amino acid switch turned the monooxygenase into an efficient preoxidase by increasing the peroxidase activity nearly one thousand folds. In order to tune the distal pocket of P450cam with polar residues, Leu 246 was replaced with a basic residue, lysine, resulting in a mutant with spectral features identical to P420, the inactive species of P450. But this inactive-species-like mutant showed catalytic activities without the facilitation of any cofactors. By substituting Gly 248 with a histidine, a novel Cys-Fe-His ligation set was obtained in P450cam which represented the very rare case of His ligation in heme-thiolate proteins. In addition to serving as a convenient model for hemoprotein structural studies, the G248H mutant also provided evidence about the nature of the axial ligand in cytochrome P420 and other engineered hemoproteins with thiolate ligations. Furthermore, attempts have been made to replace the proximal ligand in sperm whale myoglobin to construct a heme-thiolate protein model by mimicking the protein environment of cytochrome P450cam and chloroperoxidase.