Background and purpose: 5-Hydroxytryptamine (5-HT) has been shown to control and modulate many physiological and behavioural functions in insects. In this study, we report the cloning and pharmacological properties of a 5-HT1 receptor of an insect model for neurobiology, physiology and pharmacology. Experimental approach: A cDNA encoding for the Periplaneta americana 5-HT1 receptor was amplified from brain cDNA. The receptor was stably expressed in HEK 293 cells, and the functional and pharmacological properties were determined in cAMP assays. Receptor distribution was investigated by RT-PCR and by immunocytochemistry using an affinity-purified polyclonal antiserum. Key results: The P. americana 5-HT1 receptor (Pea5-HT1) shares pronounced sequence and functional similarity with mammalian 5-HT1 receptors. Activation with 5-HT reduced adenylyl cyclase activity in a dose-dependent manner. Pea5-HT1 was expressed as a constitutively active receptor with methiothepin acting as a neutral antagonist, and WAY 100635 as an inverse agonist. Receptor mRNA was present in various tissues including brain, salivary glands and midgut. Receptor-specific antibodies showed that the native protein was expressed in a glycosylated form in membrane samples of brain and salivary glands. Conclusions and implications: This study marks the first pharmacological identification of an inverse agonist and a neutral antagonist at an insect 5-HT1 receptor. The results presented here should facilitate further analyses of 5-HT1 receptors in mediating central and peripheral effects of 5-HT in insects.

Photosystem II is unique! It remains the only enzyme that can oxidize water using light as energy input. Water oxidation in photosystem II is catalyzed by the CaMn4 cluster. The electrons extracted from the CaMn4 cluster are transferred to P680+ via the tyrosine residue D1-Tyr161 (YZ). Favorable oxidation of YZ is coupled to a proton transfer along a hydrogen bond to the nearby D1-His190 residue, resulting in the neutral radical YZ•. By illuminating photosystem II at cryogenic temperatures, YZ• can be trapped in a stable state. Magnetic interaction between this radical and the CaMn4 cluster gives rise to a split electron paramagnetic resonance (EPR) signal with characteristics that depend on the oxidation state (S state) of the cluster. The mechanism by which the split EPR signals are formed is different depending on the S state. In the S0 and S1 states, split signal induction proceeds via a P680+-centered mechanism, whereas in the S2 and S3 states, our results show that split induction stems from a Mn-centered mechanism. This S state-dependent pattern of split EPR signal induction can be correlated to the charge of the CaMn4 cluster in the S state in question and has prompted us to propose a general model for the induction mechanism across the different S states. At the heart of this model is the stability or otherwise of the YZ•–(D1-His190)+ pair during cryogenic illumination. The model is closely related to the sequence of electron and proton transfers from the cluster during the S cycle. Furthermore, the important hydrogen bond between YZ and D1-His190 has been investigated by following the split EPR signal formation in the different S states as a function of pH. All split EPR signals investigated decrease in intensity with a pKa of ~4-5. This pKa can be correlated to a titration event that disrupts the essential hydrogen bond, possibly by a direct protonation of D1-His190.  This has important consequences for the function of the CaMn4 cluster as this critical YZ–D1-His190 hydrogen bond steers a multitude of reactions at the cluster.

A new approach to analysis of protein-ligand interactions, termed proteochemometrics, has been developed. Contrary to traditional quantitative structure-activity relationship (QSAR) methods that aim to correlate a description of ligands to their interactions with one particular target protein, proteochemometrics considers many targets simultaneously. Proteochemometrics thus analyzes the experimentally determined protein-ligand interaction activity data by correlating the data to a complex description of all interaction partners and; in a more general case even to interaction environment and assaying conditions, as well. In this way, a proteochemometric model analyzes an “interaction space,” from which only one cross-section would be contemplated by any one QSAR model. Proteochemometric models reveal the physicochemical and structural properties that are essential for protein-ligand complementarity and determine specificity of molecular interactions. From a drug design perspective, models may find use in the design of drugs with improved selectivity and in the design of drugs for multiple targets, such as mutated proteins (e.g., drug resistant mutations of pathogens). In this thesis, a general concept for creating of proteochemometric models and approaches for validation and interpretation of models are presented. Different types of physicochemical and structural description of ligands and macromolecules are evaluated; mathematical algorithms for proteochemometric modeling, in particular for analysis of large-scale data sets, are developed. Artificial chimeric proteins constructed according to principles of statistical design are used to derive high-resolution models for small classes of proteins. The studies of this thesis use data sets comprising ligand interactions with several families of G protein-coupled receptors. The presented approach is, however, general and can be applied to study molecular recognition mechanisms of any class of drug targets.

In the subject of Computational Fluid Dynamics (CFD), there seems to be small number of important progress in the pressure-based methods for several decades. Recent studies on the implicit coupled algorithms for pressure-based methods have brought a new insight. This method seems to provide a huge reduction in the solution times over segregated methods. Fully implicit coupled algorithm for pressure-based methods is very new subject with only few papers in literature. One of the most important work in this area is referenced as [1] in this thesis. Another source of information about the method comes from a commercially available code FLUENT which includes the algorithm as an option for pressure-based solver. However the algorithm in FLUENT does not seem to be a fully implicit with a little information in its manual. In this thesis, a fully implicit coupled pressure-based solver is developed mainly based on the available literature. The developed code is succesfully tested against some test cases.

This study introduces a multiscale model for analyzing nonlinear thermo-viscoelastic responses of particulate composites. A simplified micromechanical model consisting of four sub-cells, i.e., one particle and three matrix sub-cells is formulated to obtain the effective thermal and mechanical properties and time-dependent response of the composites. The particle and matrix constituents are made of isotropic homogeneous viscoelastic bodies undergoing small deformation gradients. Perfect bonds are assumed along the sub-cell⁰́₉s interfaces. The coupling between the thermal and mechanical response is attributed to the dissipation of energy due to the viscoelastic deformation and temperature dependent material parameters in the viscoelastic constitutive model. The micromechanical relations are formulated in terms of incremental average field quantities, i.e., stress, strain, heat flux and temperature gradient, in the sub-cells. The effective mechanical properties and coefficient of thermal expansion are derived by satisfying displacement- and traction continuities at the interfaces during the thermo-viscoelastic deformations. The effective thermal conductivity is formulated by imposing heat flux- and temperature continuities at the subcells⁰́₉ interfaces. The expression of the effective specific heat at a constant stress is also established. A time integration algorithm for simultaneously solving the equations that govern heat conduction and thermoviscoelastic deformations of isotropic materials is developed. The algorithm is then incorporated within each sub-cell of the micromechanical model together with the macroscopic energy equation to determine the effective coupled thermoviscoelastic response of the particulate composite. The numerical formulation is implemented within the ABAQUS, general purpose displacement based FE software, allowing for analyzing coupled heat conduction and deformations of composite structures. Experimental data on the effective thermal properties and time dependent responses of particulate composites available in the literature are used to verify the micromechanical model formulation. The multiscale model capability is also examined by comparing the field variables, i.e., temperature, displacement, stresses and strains, obtained from heterogeneous and homogeneous composite structures, during the transient heat conduction and deformations. Examples of coupled thermoviscoelastic analyses of particulate composites and functionally graded structures are also presented. The present micromechanical modeling approach is found to be computationally efficient and shows good agreement with experiments in predicting the effective thermo-mechanical response of particulate composites and functionally graded materials. Our analyses forecast a better design for creep resistant and less dissipative structures using particulate composites and functionally graded materials.

Problems involving coupled thermo-poro-chemo-mechanical processes are of great importance in geothermal and petroleum reservoir systems. In particular, economic power production from enhanced geothermal systems, effective water-flooding of petroleum reservoirs, and stimulation of gas shale reservoirs are significantly influenced by coupled processes. During such procedures, stress state in the reservoir is changed due to variation in pore fluid pressure and temperature. This can cause deformation and failure of weak planes of the formation with creation of new fractures, which impacts reservoir response. Incorporation of geomechanical factor into engineering analyses using fully coupled geomechanics-reservoir flow modeling exhibits computational challenges and numerical difficulties. In this study, we develop and apply efficient numerical models to solve 3D injection/extraction geomechanics problems formulated within the framework of thermo-poro-mechanical theory with reactive flow. The models rely on combining Displacement Discontinuity (DD) Boundary Element Method (BEM) and Finite Element Method (FEM) to solve the governing equations of thermo-poro-mechanical processes involving fracture/reservoir matrix. The integration of BEM and FEM is accomplished through direct and iterative procedures. In each case, the numerical algorithms are tested against a series of analytical solutions. 3D study of fluid injection and extraction into the geothermal reservoir illustrates that thermo-poro-mechanical processes change fracture aperture (fracture conductivity) significantly and influence the fluid flow. Simulations that consider joint stiffness heterogeneity show development of non-uniform flow paths within the crack. Undersaturated fluid injection causes large silica mass dissolution and increases fracture aperture while supersaturated fluid causes mineral precipitation and closes fracture aperture. Results show that for common reservoir and injection conditions, the impact of fully developed thermoelastic effect on fracture aperture tend to be greater compare to that of poroelastic effect. Poroelastic study of hydraulic fracturing demonstrates that large pore pressure increase especially during multiple hydraulic fracture creation causes effective tensile stress at the fracture surface and shear failure around the main fracture. Finally, a hybrid BEFEM model is developed to analyze stress redistribution in the overburden and within the reservoir during fluid injection and production. Numerical results show that fluid injection leads to reservoir dilation and induces vertical deformation, particularly near the injection well. However, fluid withdrawal causes reservoir to compact. The Mandel-Cryer effect is also successfully captured in numerical simulations, i.e., pore pressure increase/decrease is non-monotonic with a short time values that are above/below the background pore pressure.

Magnetic components, such as inductors and transformers, have important effects on the efficiency and performance of switching power supplies; their parasitic properties directly impact the high frequency properties which can cause lot-to-lot variation or unanticipated and non-ideal operation. They are also amongst the most problematic components to design, often requiring numerous design-prototype-test interactions. The electrostatic and electromagnetic analysis of wound components has become more important recently to predict their performance and frequency behavior. Accurate prediction and design of winding parasitic parameters of leakage inductance and winding capacitance for high frequency inductors and transformers in switching power supplies is fundamental to improve performance, lower cost, and speed time to market. This thesis presents a methodology and process to obtain accurate prediction of the inter- and intra-winding capacitances of high frequency magnetic components. Application examples considered are a single-winding choke, a coupled inductor filter, and a multi-winding transformer. Analytical approach for determination of parasitic capacitances in high frequency magnetic components will be covered also. Comparison of the FEA results using JMAG with experimental and empirical formula results show good agreement, supporting the method as a model-based design tool with the potential to significantly reduce the design-prototype-test cycle commonly needed with sophisticated magnetic designs.

Today's product development process should be rapid andcost-efficient, and should result in innovative and reliableproducts. A crucial factor is the dynamic behaviour of theproduct. This thesis focuses on theoretical, numerical andexperimental approaches to achieve a comprehensiveunderstanding of dynamical phenomena occurring in nonlinearproducts, especially in products with parts that includeimpacts. The aim is to show the usefulness of nonlineartheories to better understand and optimise the dynamicbehaviour of products and thereby account for nonlinearphenomena already in the product development process. This is achieved through an investigation of researchefforts in the field of nonlinear dynamics; identification ofimportant research directions; a study on the effect ofcouplings between nonlinear parts; a detailed study on thedynamic behaviour of a product component; investigations oflow-cost strategies for controlling the dynamics of a nonlinearsystem; and the design and implementation of experimentalset-ups of two studied products. The investigation of research efforts shows that nonlinearparts are frequently included in products. Most common areparts that are nonlinear due to impacts and friction. Twoimportant areas are identified; to study coupling effectsbetween nonlinear subsystems and to study how nonlinearanalysis can be used to improve existing designs. Considering the studied products; a pantograph on a trainand a Braille printer, it can be concluded that thecharacteristics of a part can largely affect the dynamicbehaviour of the product. Typical nonlinear behaviour, such ascoexisting solutions and irregular motions, do occur. Theanalysis of the pan- tograph motion shows important aspects toconsider in the modelling process; coupling effects. In thecase of the Braille printer it is shown possible to create alow-cost control, by taking advantage of an existingdiscontinuity, to achieve a desired motion. Altogether, this work contributes to improved understandingof the be- haviour of nonlinear parts in products, especiallythose including impacts, pro- viding greater knowledge aboutaspects to consider in the design process. Keywords:Nonlinear Dynamics, Impacts, Discontinuities,Subsystems, Chaos, Irregular Behaviour, Printer Dynamics,Suspensions, Coupled Systems, Control. / QC 20100621

The neuropeptides orexin-A and orexin-B were discovered in 1998 and were first described as regulators of feeding behaviour. Later research has shown that they have an important role in the regulation of sleep. Two G protein-coupled receptors, OX1 and OX2 orexin receptors, mediate the cellular responses to orexins. The overall aim of this thesis was to investigate the OX1 orexin receptors signalling to phospholipases. Previous investigations have determined that orexin receptors induce Ca2+ elevations through both receptor-operated Ca2+ channels (ROCs) and store-operated Ca2+ channels (SOCs). In this thesis we investigated the importance of these influxpathways on orexin-mediated phospholipase (PLC) activation. The results demonstrate that ROC influx is enough to fully support orexin-stimulated PLC activation but that SOC influx has a further amplifying role. We also investigated the metabolites generated after PLC activation, inositolphosphates and diacylglycerol (DAG). The results indicate involvement of two different PLC activities with different substrate specificities one of them leading to DAG production without co-occurring IP3 production at low orexin receptor stimulation. The results also suggest that at even lower orexin receptor stimulation DAG is produced via the activation of phospholipase D. In this thesis we also investigated if the ubiquitous phospholipase A2 (PLA2) signalling system is involved in orexin receptor signalling. The results demonstrate that stimulation of the OX1 orexin receptors leads to arachidonic acid (AA) release. This release is fully dependent on Ca2+ influx, probably through ROC, and at the same time the studies demonstrate that ROC influx is partly dependent on PLA2 activation. At low orexin receptor activation the AA release seemed to in part rely on extracellular signal-regulated kinase. We also devised two methods to aid in these investigations. The first method enabled studies of the receptor-operated Ca2+ influx without interference of the co-occurring store-operated Ca2+ influx. This was done by the expression of IP3-metabolising enzymes IP3-3-kinase-A and IP3-5-phosphatase-I. The second method enables quantification of DAG and IP3 signalling in fixed cells using GFP-fused indicators, leading to a semi-quantitative but easily applicable pharmacological assay.

The increasing number of biological databases today requires that users are able to search more efficiently among as well as in individual databases. One of the most widespread problems is redundancy, i.e. the problem of duplicated information in sets of data. This thesis aims at implementing an algorithm that distinguishes from other related attempts by using the genomic positions of sequences, instead of similarity based sequence comparisons, when making a sequence data set non-redundant. In an automatic updating procedure the algorithm drastically increases the possibility to update and to maintain the topicality of a non-redundant database. The procedure creates a biologically sound non-redundant data set with accuracy comparable to other algorithms focusing on making data sets non-redundant