Tübingen, Univ., Diss., 2007.

Thesis (Ph. D.)--University of Washington, 2006. / Vita. Includes bibliographical references (leaves 105-120).

Frankfurt (Main), Univ., Diss., 2001.

Universiẗat, Diss., 2003--Kiel.

abstract: Patients with Alzheimer's disease (AD) exhibit a significantly higher incidence of unprovoked seizures compared to age-matched non-AD controls, and animal models of AD (i.e., transgenic human amyloid precursor protein, hAPP mice) display neural hyper-excitation and epileptic seizures. Hyperexcitation is particularly important because it contributes to the high incidence of epilepsy in AD patients as well as AD-related synaptic deficits and neurodegeneration. Given that there is significant amyloid-β (Aβ) accumulation and deposition in AD brain, Aβ exposure ultimately may be responsible for neural hyper-excitation in both AD patients and animal models. Emerging evidence indicates that α7 nicotinic acetylcholine receptors (α7-nAChR) are involved in AD pathology, because synaptic impairment and learning and memory deficits in a hAPPα7-/- mouse model are decreased by nAChR α7 subunit gene deletion. Given that Aβ potently modulates α7-nAChR function, that α7-nAChR expression is significantly enhanced in both AD patients and animal models, and that α7-nAChR play an important role in regulating neuronal excitability, it is reasonable that α7-nAChRs may contribute to Aβ-induced neural hyperexcitation. We hypothesize that increased α7-nAChR expression and function as a consequence of Aβ exposure is important in Aβ-induced neural hyperexcitation. In this project, we found that exposure of Aβ aggregates at a nanomolar range induces neuronal hyperexcitation and toxicity via an upregulation of α7-nAChR in cultured hippocampus pyramidal neurons. Aβ up-regulates α7-nAChRs function and expression through a post translational mechanism. α7-nAChR up-regulation occurs prior to Aβ-induced neuronal hyperexcitation and toxicity. Moreover, inhibition of α7-nAChR or deletion of α7-nAChR prevented Aβ induced neuronal hyperexcitation and toxicity, which suggests that α7-nAChRs are required for Aβ induced neuronal hyperexcitation and toxicity. These results reveal a profound role for α7-nAChR in mediating Aβ-induced neuronal hyperexcitation and toxicity and predict that Aβ-induced up-regulation of α7-nAChR could be an early and critical event in AD etiopathogenesis. Drugs targeting α7-nAChR or seizure activity could be viable therapies for AD treatment. / Dissertation/Thesis / Ph.D. Neuroscience 2011

Introduction: Beat-to-beat variability in cycle length exists in spontaneously beating cardiac preparations of varying complexities from the level of the isolated whole heart to the single sinoatrial nodal cell (SANC). The nature of this variability is poorly characterised as are its fundamental physiological origins. Methods: Recordings of spontaneous electrical activity were made from hearts in vivo, during Langendorff-perfusion, and from single SANC. Heart rate variability (HRV) was calculated in the time- and frequency-domains at baseline and in response to pharmacological mediators that interfered with critical processes involved in automaticity (catecholamines, carbachol, ivabradine, zatebradine, ryanodine and thapsigargin). In addition, a novel 2D technique for imaging Ca2+ fluorescence in spontaneously beating, fluo4-AM loaded, patched single sinoatrial nodal cells was developed to investigate the biophysical behaviour of Ca2+ during pacemaking to see if variability in this was responsible for SANC HRV. Results: Under baseline, temperature-stable conditions, levels of HRV were greatest in vivo (human > rat). SANC exhibited slightly lower levels of HRV, whereas HRV levels expressed by Langendorff-perfused hearts were the least (rabbit > rat), although still comprised a significant proportion of the variability witnessed in vivo. Anaesthetising in vivo rabbits decreased HRV to levels similar to those seen in the Langendorff-perfused heart. HRV was decreased by catecholamines and by ryanodine/thapsigargin in the Langendorff heart. Conversely, HRV was increased by carbachol, ivabradine, zatebradine and ryanodine in SANC. Heart rate changes had a marked effect on levels of HRV. 2D Ca2+ imaging of SANC showed that diastolic local Ca2+ releases (LCRs) occurred earlier than previously thought, with early LCRs having characteristics that were distinct from later LCRs. Mean time of occurrence of all the LCRs within a given diastole closely predicted the duration of the cycle. The rate of restitution of the whole cell Ca2+ transient (used as a surrogate for the pumping function of SERCA) in turn closely predicted the mean time of occurrence of LCRs. Tight synchronisation of the electrical activity of the cell with the biophysical behaviour of Ca2+ appeared to predict shorter cycle lengths. Isoprenaline increased LCR amplitude, though did not increase LCR number, size or duration. Isoprenaline caused LCRs to occur earlier, and synchronised their occurrence and the rate of pumping of Ca2+ back into the sarcoplasmic reticulum. Finally, LCRs were found to preferentially recur in certain regions of the cell, dubbed hotspots. Isoprenaline favoured hotspot production. Conclusion: Whilst greatest in vivo, significant HRV exists in spontaneously beating cardiac preparations devoid of a functioning autonomic nervous system. Studies in SANC indicate that the origin of this is likely to be variability in release of LCRs from the SR via ryanodine receptors. This in turn is controlled by SR refilling kinetics via SR Ca2+ pumping. The coupled system of membrane- and Ca2+-pacemaker clocks are so heavily intertwined that myriad factors will come to bear on generating such variability, including the amount of Ca2+ available for pumping and the phosphorylation state of key proteins, to the extent that variability in no one process can take the credit for generating such HRV.

abstract: As residential photovoltaic (PV) systems become more and more common and widespread, their system architectures are being developed to maximize power extraction while keeping the cost of associated electronics to a minimum. An architecture that has become popular in recent years is the "DC optimizer" architecture, wherein one DC-DC converter is connected to the output of each PV module. The DC optimizer architecture has the advantage of performing maximum power-point tracking (MPPT) at the module level, without the high cost of using an inverter on each module (the "microinverter" architecture). This work details the design of a proposed DC optimizer. The design incorporates a series-input parallel-output topology to implement MPPT at the sub-module level. This topology has some advantages over the more common series-output DC optimizer, including relaxed requirements for the system's inverter. An autonomous control scheme is proposed for the series-connected converters, so that no external control signals are needed for the system to operate, other than sunlight. The DC optimizer in this work is designed with an emphasis on efficiency, and to that end it uses GaN FETs and an active clamp technique to reduce switching and conduction losses. As with any parallel-output converter, phase interleaving is essential to minimize output RMS current losses. This work proposes a novel phase-locked loop (PLL) technique to achieve interleaving among the series-input converters. / Dissertation/Thesis / Masters Thesis Electrical Engineering 2014

abstract: Solution conformations and dynamics of proteins and protein-DNA complexes are often difficult to predict from their crystal structures. The crystal structure only shows a snapshot of the different conformations these biological molecules can have in solution. Multiple different conformations can exist in solution and potentially have more importance in the biological activity. DNA sliding clamps are a family of proteins with known crystal structures. These clamps encircle the DNA and enable other proteins to interact more efficiently with the DNA. Eukaryotic PCNA and prokaryotic β clamp are two of these clamps, some of the most stable homo-oligomers known. However, their solution stability and conformational equilibrium have not been investigated in depth before. Presented here are the studies involving two sliding clamps: yeast PCNA and bacterial β clamp. These studies show that the β clamp has a very different solution stability than PCNA. These conclusions were reached through various different fluorescence-based experiments, including fluorescence correlation spectroscopy (FCS), Förster resonance energy transfer (FRET), single molecule fluorescence, and various time resolved fluorescence techniques. Interpretations of these, and all other, fluorescence-based experiments are often affected by the properties of the fluorophores employed. Often the fluorescence properties of these fluorophores are influenced by their microenvironments. Fluorophores are known to sometimes interact with biological molecules, and this can have pronounced effects on the rotational mobility and photophysical properties of the dye. Misunderstanding the effect of these photophysical and rotational properties can lead to a misinterpretation of the obtained data. In this thesis, photophysical behaviors of various organic dyes were studied in the presence of deoxymononucleotides to examine more closely how interactions between fluorophores and DNA bases can affect fluorescent properties. Furthermore, the properties of cyanine dyes when bound to DNA and the effect of restricted rotation on FRET are presented in this thesis. This thesis involves studying fluorophore photophysics in various microenvironments and then expanding into the solution stability and dynamics of the DNA sliding clamps. / Dissertation/Thesis / Ph.D. Chemistry 2013

abstract: Following a traumatic brain injury (TBI) 5-50% of patients will develop post traumatic epilepsy (PTE). Pediatric patients are most susceptible with the highest incidence of PTE. Currently, we cannot prevent the development of PTE and knowledge of basic mechanisms are unknown. This has led to several shortcomings to the treatment of PTE, one of which is the use of anticonvulsant medication to the population of TBI patients that are not likely to develop PTE. The complication of identifying the two populations has been hindered by the ability to find a marker to the pathogenesis of PTE. The central hypothesis of this dissertation is that following TBI, the cortex undergoes distinct cellular and synaptic reorganization that facilitates cortical excitability and promotes seizure development. Chapter 2 of this dissertation details excitatory and inhibitory changes in the rat cortex after severe TBI. This dissertation aims to identify cortical changes to a single cell level after severe TBI using whole cell patch clamp and electroencephalogram electrophysiology. The work of this dissertation concluded that excitatory and inhibitory synaptic activity in cortical controlled impact (CCI) animals showed the development of distinct burst discharges that were not present in control animals. The results suggest that CCI induces early "silent" seizures that are detectable on EEG and correlate with changes to the synaptic excitability in the cortex. The synaptic changes and development of burst discharges may play an important role in synchronizing the network and promoting the development of PTE. / Dissertation/Thesis / Masters Thesis Biology 2014

This thesis presents work to help enable the transition of sensitive nanoscale instruments from research laboratory demonstration to societal use. It focuses on nanomechanical resonators made by field-directed assembly, with contributions to understanding effects of materials, clamp geometries and laser measurement of motion, towards their use as commercial scientific instruments. Nanomechanical resonators in their simplest form are cantilevered or doubly- clamped nanowires or nanotubes made to vibrate near one of their resonant frequencies. Their small mass and high frequency enable extraordinary mass sensitivity, as shown in published laboratory-scale demonstrations of their use for detection of a few molecules of prostate cancer biomarker and of their response to mass equal to that of a single proton. However such sensitive devices have been prohibitively expensive for societal use, since the fabrication process cost scales with number of devices and the chip area covered, when they are made using standard electron beam lithography. Our laboratory has published new results for the method of field-directed assembly, in which the nanofabrication process cost is independent of the number of devices. While drastically lowering the cost, this method also broadens the range of device materials and properties that can be used in instrument applications for sensitive mass and force detection. Unanswered questions affecting the performance of devices made by this method are studied in this thesis. Clamping variability can cause uncertainties in the device resonant frequency (effective stiffness), raising manufacturing metrology costs to track reduced homogeneity in performance. Using a numerical model, we quantify how compliant clamp material and insufficient clamp depth reduce the effective stiffness and resonance frequency. Obliquely clamped nanowires and defects at the clamp-nanowire interface break the symmetry and split the resonance frequency into fast and slow modes. The difference of resonance frequency between the fast and slow modes corresponds to the degree of asymmetry and must be controlled in fabrication to keep device error bounded. Optical transduction has been used for measuring the nanoresonator frequency spectrum; however, the influence of the laser in the measurement process is only recently receiving attention and is not well understood. We found that the measured spectrum is significantly influenced by laser-nanowire interaction. Variation of input laser power could result in resonance peak shifts in the kHz range for a resonance frequency in the MHz range, which could reduce device mass resolution by a factor of 100 or greater. As the laser power is increased, the resonance frequency decreases. The heating effect of the laser on temperature-dependent Young’s modulus could explain this phenomenon. To our surprise, we also found that the amplitude and frequency of the resonance peak signal vary significantly with the angle made by the plane of laser polarization with the nanowire axis. Our measurements established that the maximum signal amplitude is seen when the plane of the linearly polarized laser is parallel to SiNW or perpendicular to RhNW. Maximum resonance frequency was found when laser is polarized perpendicular to SiNW or parallel to RhNW. / Graduate / 0537 / 0548 / 0752