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Description
Gene manipulation techniques, such as RNA interference (RNAi), offer a powerful method for elucidating gene function and discovery of novel therapeutic targets in a high-throughput fashion. In addition, RNAi is rapidly being adopted for treatment of neurological disorders, such as Alzheimer's disease (AD), Parkinson's disease, etc. However, a major challenge

Gene manipulation techniques, such as RNA interference (RNAi), offer a powerful method for elucidating gene function and discovery of novel therapeutic targets in a high-throughput fashion. In addition, RNAi is rapidly being adopted for treatment of neurological disorders, such as Alzheimer's disease (AD), Parkinson's disease, etc. However, a major challenge in both of the aforementioned applications is the efficient delivery of siRNA molecules, plasmids or transcription factors to primary cells such as neurons. A majority of the current non-viral techniques, including chemical transfection, bulk electroporation and sonoporation fail to deliver with adequate efficiencies and the required spatial and temporal control. In this study, a novel optically transparent biochip is presented that can (a) transfect populations of primary and secondary cells in 2D culture (b) readily scale to realize high-throughput transfections using microscale electroporation and (c) transfect targeted cells in culture with spatial and temporal control. In this study, delivery of genetic payloads of different sizes and molecular characteristics, such as GFP plasmids and siRNA molecules, to precisely targeted locations in primary hippocampal and HeLa cell cultures is demonstrated. In addition to spatio-temporally controlled transfection, the biochip also allowed simultaneous assessment of a) electrical activity of neurons, b) specific proteins using fluorescent immunohistochemistry, and c) sub-cellular structures. Functional silencing of GAPDH in HeLa cells using siRNA demonstrated a 52% reduction in the GAPDH levels. In situ assessment of actin filaments post electroporation indicated a sustained disruption in actin filaments in electroporated cells for up to two hours. Assessment of neural spike activity pre- and post-electroporation indicated a varying response to electroporation. The microarray based nature of the biochip enables multiple independent experiments on the same culture, thereby decreasing culture-to-culture variability, increasing experimental throughput and allowing cell-cell interaction studies. Further development of this technology will provide a cost-effective platform for performing high-throughput genetic screens.
ContributorsPatel, Chetan (Author) / Muthuswamy, Jitendran (Thesis advisor) / Helms Tillery, Stephen (Committee member) / Jain, Tilak (Committee member) / Caplan, Michael (Committee member) / Vernon, Brent (Committee member) / Arizona State University (Publisher)
Created2012
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Description
Advances in implantable MEMS technology has made possible adaptive micro-robotic implants that can track and record from single neurons in the brain. Development of autonomous neural interfaces opens up exciting possibilities of micro-robots performing standard electrophysiological techniques that would previously take researchers several hundred hours to train and achieve the

Advances in implantable MEMS technology has made possible adaptive micro-robotic implants that can track and record from single neurons in the brain. Development of autonomous neural interfaces opens up exciting possibilities of micro-robots performing standard electrophysiological techniques that would previously take researchers several hundred hours to train and achieve the desired skill level. It would result in more reliable and adaptive neural interfaces that could record optimal neural activity 24/7 with high fidelity signals, high yield and increased throughput. The main contribution here is validating adaptive strategies to overcome challenges in autonomous navigation of microelectrodes inside the brain. The following issues pose significant challenges as brain tissue is both functionally and structurally dynamic: a) time varying mechanical properties of the brain tissue-microelectrode interface due to the hyperelastic, viscoelastic nature of brain tissue b) non-stationarities in the neural signal caused by mechanical and physiological events in the interface and c) the lack of visual feedback of microelectrode position in brain tissue. A closed loop control algorithm is proposed here for autonomous navigation of microelectrodes in brain tissue while optimizing the signal-to-noise ratio of multi-unit neural recordings. The algorithm incorporates a quantitative understanding of constitutive mechanical properties of soft viscoelastic tissue like the brain and is guided by models that predict stresses developed in brain tissue during movement of the microelectrode. An optimal movement strategy is developed that achieves precise positioning of microelectrodes in the brain by minimizing the stresses developed in the surrounding tissue during navigation and maximizing the speed of movement. Results of testing the closed-loop control paradigm in short-term rodent experiments validated that it was possible to achieve a consistently high quality SNR throughout the duration of the experiment. At the systems level, new generation of MEMS actuators for movable microelectrode array are characterized and the MEMS device operation parameters are optimized for improved performance and reliability. Further, recommendations for packaging to minimize the form factor of the implant; design of device mounting and implantation techniques of MEMS microelectrode array to enhance the longevity of the implant are also included in a top-down approach to achieve a reliable brain interface.
ContributorsAnand, Sindhu (Author) / Muthuswamy, Jitendran (Thesis advisor) / Tillery, Stephen H (Committee member) / Buneo, Christopher (Committee member) / Abbas, James (Committee member) / Tsakalis, Konstantinos (Committee member) / Arizona State University (Publisher)
Created2013
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Description
The basal ganglia are four sub-cortical nuclei associated with motor control and reward learning. They are part of numerous larger mostly segregated loops where the basal ganglia receive inputs from specific regions of cortex. Converging on these inputs are dopaminergic neurons that alter their firing based on received and/or predicted

The basal ganglia are four sub-cortical nuclei associated with motor control and reward learning. They are part of numerous larger mostly segregated loops where the basal ganglia receive inputs from specific regions of cortex. Converging on these inputs are dopaminergic neurons that alter their firing based on received and/or predicted rewarding outcomes of a behavior. The basal ganglia's output feeds through the thalamus back to the areas of the cortex where the loop originated. Understanding the dynamic interactions between the various parts of these loops is critical to understanding the basal ganglia's role in motor control and reward based learning. This work developed several experimental techniques that can be applied to further study basal ganglia function. The first technique used micro-volume injections of low concentration muscimol to decrease the firing rates of recorded neurons in a limited area of cortex in rats. Afterwards, an artificial cerebrospinal fluid flush was injected to rapidly eliminate the muscimol's effects. This technique was able to contain the effects of muscimol to approximately a 1 mm radius volume and limited the duration of the drug effect to less than one hour. This technique could be used to temporarily perturb a small portion of the loops involving the basal ganglia and then observe how these effects propagate in other connected regions. The second part applied self-organizing maps (SOM) to find temporal patterns in neural firing rate that are independent of behavior. The distribution of detected patterns frequency on these maps can then be used to determine if changes in neural activity are occurring over time. The final technique focused on the role of the basal ganglia in reward learning. A new conditioning technique was created to increase the occurrence of selected patterns of neural activity without utilizing any external reward or behavior. A pattern of neural activity in the cortex of rats was selected using an SOM. The pattern was then reinforced by being paired with electrical stimulation of the medial forebrain bundle triggering dopamine release in the basal ganglia. Ultimately, this technique proved unsuccessful possibly due to poor selection of the patterns being reinforced.
ContributorsBaldwin, Nathan Aaron (Author) / Helms Tillery, Stephen I (Thesis advisor) / Castaneda, Edward (Committee member) / Buneo, Christopher A (Committee member) / Muthuswamy, Jitendran (Committee member) / Si, Jennie (Committee member) / Arizona State University (Publisher)
Created2014
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Description
Computed tomography (CT) is one of the essential imaging modalities for medical diagnosis. Since its introduction in 1972, CT technology has been improved dramatically, especially in terms of its acquisition speed. However, the main principle of CT which consists in acquiring only density information has not changed at all

Computed tomography (CT) is one of the essential imaging modalities for medical diagnosis. Since its introduction in 1972, CT technology has been improved dramatically, especially in terms of its acquisition speed. However, the main principle of CT which consists in acquiring only density information has not changed at all until recently. Different materials may have the same CT number, which may lead to uncertainty or misdiagnosis. Dual-energy CT (DECT) was reintroduced recently to solve this problem by using the additional spectral information of X-ray attenuation and aims for accurate density measurement and material differentiation. However, the spectral information lies in the difference between two low and high energy images or measurements, so that it is difficult to acquire the accurate spectral information due to amplification of high pixel noise in the resulting difference image. In this work, a new model and an image enhancement technique for DECT are proposed, based on the fact that the attenuation of a high density material decreases more rapidly as X-ray energy increases. This fact has been previously ignored in most of DECT image enhancement techniques. The proposed technique consists of offset correction, spectral error correction, and adaptive noise suppression. It reduced noise, improved contrast effectively and showed better material differentiation in real patient images as well as phantom studies.
ContributorsPark, Kyung Kook (Author) / Metin, Akay (Thesis advisor) / Pavlicek, William (Committee member) / Akay, Yasemin (Committee member) / Towe, Bruce (Committee member) / Muthuswamy, Jitendran (Committee member) / Arizona State University (Publisher)
Created2011
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Description
Biosensors offer excellent diagnostic methods through precise quantification of bodily fluid biomarkers and could fill an important niche in diagnostic screening. The long term goal of this research is the development of an impedance immunosensor for easy-to-use, rapid, sensitive and selective simultaneously multiplexed quantification of bodily fluid disease biomarkers. To

Biosensors offer excellent diagnostic methods through precise quantification of bodily fluid biomarkers and could fill an important niche in diagnostic screening. The long term goal of this research is the development of an impedance immunosensor for easy-to-use, rapid, sensitive and selective simultaneously multiplexed quantification of bodily fluid disease biomarkers. To test the hypothesis that various cytokines induce empirically determinable response frequencies when captured by printed circuit board (PCB) impedance immunosensor surface, cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) methods were used to test PCB biosensors versus multiple cytokine biomarkers to determine limits of detection, background interaction and response at all sweep frequencies. Results indicated that sensors for cytokine Interleukin-12 (IL-12) detected their target over three decades of concentration and were tolerant to high levels of background protein. Further, the hypothesis that cytokine analytes may be rapidly detected via constant frequency impedance immunosensing without sacrificing undue sensitivity, CV, EIS, impedance-time (Zt) methods and modeling were used to test CHITM gold electrodes versus IL-12 over different lengths of time to determine limits of detection, detection time, frequency of response and consistent cross-platform sensor performance. Modeling and Zt studies indicate interrogation of the electrode with optimum frequency could be used for detection of different target concentrations within 90 seconds of sensor exposure and that interrogating the immunosensor with fixed, optimum frequency could be used for sensing target antigen. This informs usability of fixed-frequency impedance methods for biosensor research and particularly for clinical biosensor use. Finally, a multiplexing impedance immunosensor prototype for quantification of biomarkers in various body fluids was designed for increased automation of sample handling and testing. This enables variability due to exogenous factors and increased rapidity of assay with eased sensor fabrication. Methods were provided for simultaneous multiplexing through multisine perturbation of a sensor, and subsequent data processing. This demonstrated ways to observe multiple types of antibody-antigen affinity binding events in real time, reducing the number of sensors and target sample used in the detection and quantification of multiple biomarkers. These features would also improve the suitability of the sensor for clinical multiplex detection of disease biomarkers.
ContributorsFairchild, Aaron (Author) / La Belle, Jeffrey T (Thesis advisor) / Muthuswamy, Jitendran (Committee member) / Nagaraj, Vinay (Committee member) / Pizziconi, Vince (Committee member) / Vernon, Brent (Committee member) / Arizona State University (Publisher)
Created2012
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Description
Noninvasive neuromodulation could help treat many neurological disorders, but existing techniques have low resolution and weak penetration. Ultrasound (US) shows promise for stimulation of smaller areas and subcortical structures. However, the mechanism and parameter design are not understood. US can stimulate tail and hindlimb movements in rats, but not forelimb,

Noninvasive neuromodulation could help treat many neurological disorders, but existing techniques have low resolution and weak penetration. Ultrasound (US) shows promise for stimulation of smaller areas and subcortical structures. However, the mechanism and parameter design are not understood. US can stimulate tail and hindlimb movements in rats, but not forelimb, for unknown reasons. Potentially, US could also stimulate peripheral or enteric neurons for control of blood glucose.

To better understand the inconsistent effects across rat motor cortex, US modulation of electrically-evoked movements was tested. A stimulation array was implanted on the cortical surface and US (200 kHz, 30-60 W/cm2 peak) was applied while measuring changes in the evoked forelimb and hindlimb movements. Direct US stimulation of the hindlimb was also studied. To test peripheral effects, rat blood glucose levels were measured while applying US near the liver.

No short-term motor modulation was visible (95% confidence interval: -3.5% to +5.1% forelimb, -3.8% to +5.5% hindlimb). There was significant long-term (minutes-order) suppression (95% confidence interval: -3.7% to -10.8% forelimb, -3.8% to -11.9% hindlimb). This suppression may be due to the considerable heating (+1.8°C between US
on-US conditions); effects of heat and US were not separable in this experiment. US directly evoked hindlimb and scrotum movements in some sessions. This required a long interval, at least 3 seconds between US bursts. Movement could be evoked with much shorter pulses than used in literature (3 ms). The EMG latency (10 ms) was compatible with activation of corticospinal neurons. The glucose modulation test showed a strong increase in a few trials, but across all trials found no significant effect.

The single motor response and the long refractory period together suggest that only the beginning of the US burst had a stimulatory effect. This would explain the lack of short-term modulation, and suggests future work with shorter pulses could better explore the missing forelimb response. During the refractory period there was no change in the electrically-evoked response, which suggests the US stimulation mechanism is independent of normal brain activity. These results challenge the literature-standard protocols and provide new insights on the unknown mechanism.
ContributorsGulick, Daniel Withers (Author) / Kleim, Jeffrey (Thesis advisor) / Towe, Bruce (Thesis advisor) / Muthuswamy, Jitendran (Committee member) / Herman, Richard (Committee member) / Helms Tillery, Steven (Committee member) / Arizona State University (Publisher)
Created2015
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Description
A direct Magnetic Resonance (MR)-based neural activity mapping technique with high spatial and temporal resolution may accelerate studies of brain functional organization.

The most widely used technique for brain functional imaging is functional Magnetic Resonance Image (fMRI). The spatial resolution of fMRI is high. However, fMRI signals are highly influenced

A direct Magnetic Resonance (MR)-based neural activity mapping technique with high spatial and temporal resolution may accelerate studies of brain functional organization.

The most widely used technique for brain functional imaging is functional Magnetic Resonance Image (fMRI). The spatial resolution of fMRI is high. However, fMRI signals are highly influenced by the vasculature in each voxel and can be affected by capillary orientation and vessel size. Functional MRI analysis may, therefore, produce misleading results when voxels are nearby large vessels. Another problem in fMRI is that hemodynamic responses are slower than the neuronal activity. Therefore, temporal resolution is limited in fMRI. Furthermore, the correlation between neural activity and the hemodynamic response is not fully understood. fMRI can only be considered an indirect method of functional brain imaging.

Another MR-based method of functional brain mapping is neuronal current magnetic resonance imaging (ncMRI), which has been studied over several years. However, the amplitude of these neuronal current signals is an order of magnitude smaller than the physiological noise. Works on ncMRI include simulation, phantom experiments, and studies in tissue including isolated ganglia, optic nerves, and human brains. However, ncMRI development has been hampered due to the extremely small signal amplitude, as well as the presence of confounding signals from hemodynamic changes and other physiological noise.

Magnetic Resonance Electrical Impedance Tomography (MREIT) methods could have the potential for the detection of neuronal activity. In this technique, small external currents are applied to a body during MR scans. This current flow produces a magnetic field as well as an electric field. The altered magnetic flux density along the main magnetic field direction caused by this current flow can be obtained from phase images. When there is neural activity, the conductivity of the neural cell membrane changes and the current paths around the neurons change consequently. Neural spiking activity during external current injection, therefore, causes differential phase accumulation in MR data. Statistical analysis methods can be used to identify neuronal-current-induced magnetic field changes.
ContributorsFu, Fanrui (Author) / Sadleir, Rosalind (Thesis advisor) / Kodibagkar, Vikram (Committee member) / Kleim, Jeffrey (Committee member) / Muthuswamy, Jitendran (Committee member) / Helms Tillery, Stephen (Committee member) / Arizona State University (Publisher)
Created2019
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Description
From time immemorial, epilepsy has persisted to be one of the greatest impediments to human life for those stricken by it. As the fourth most common neurological disorder, epilepsy causes paroxysmal electrical discharges in the brain that manifest as seizures. Seizures have the effect of debilitating patients on a physical

From time immemorial, epilepsy has persisted to be one of the greatest impediments to human life for those stricken by it. As the fourth most common neurological disorder, epilepsy causes paroxysmal electrical discharges in the brain that manifest as seizures. Seizures have the effect of debilitating patients on a physical and psychological level. Although not lethal by themselves, they can bring about total disruption in consciousness which can, in hazardous conditions, lead to fatality. Roughly 1\% of the world population suffer from epilepsy and another 30 to 50 new cases per 100,000 increase the number of affected annually. Controlling seizures in epileptic patients has therefore become a great medical and, in recent years, engineering challenge.



In this study, the conditions of human seizures are recreated in an animal model of temporal lobe epilepsy. The rodents used in this study are chemically induced to become chronically epileptic. Their Electroencephalogram (EEG) data is then recorded and analyzed to detect and predict seizures; with the ultimate goal being the control and complete suppression of seizures.



Two methods, the maximum Lyapunov exponent and the Generalized Partial Directed Coherence (GPDC), are applied on EEG data to extract meaningful information. Their effectiveness have been reported in the literature for the purpose of prediction of seizures and seizure focus localization. This study integrates these measures, through some modifications, to robustly detect seizures and separately find precursors to them and in consequence provide stimulation to the epileptic brain of rats in order to suppress seizures. Additionally open-loop stimulation with biphasic currents of various pairs of sites in differing lengths of time have helped us create control efficacy maps. While GPDC tells us about the possible location of the focus, control efficacy maps tells us how effective stimulating a certain pair of sites will be.



The results from computations performed on the data are presented and the feasibility of the control problem is discussed. The results show a new reliable means of seizure detection even in the presence of artifacts in the data. The seizure precursors provide a means of prediction, in the order of tens of minutes, prior to seizures. Closed loop stimulation experiments based on these precursors and control efficacy maps on the epileptic animals show a maximum reduction of seizure frequency by 24.26\% in one animal and reduction of length of seizures by 51.77\% in another. Thus, through this study it was shown that the implementation of the methods can ameliorate seizures in an epileptic patient. It is expected that the new knowledge and experimental techniques will provide a guide for future research in an effort to ultimately eliminate seizures in epileptic patients.
ContributorsShafique, Md Ashfaque Bin (Author) / Tsakalis, Konstantinos (Thesis advisor) / Rodriguez, Armando (Committee member) / Muthuswamy, Jitendran (Committee member) / Spanias, Andreas (Committee member) / Arizona State University (Publisher)
Created2016
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Description
Over the past fifty years, the development of sensors for biological applications has increased dramatically. This rapid growth can be attributed in part to the reduction in feature size, which the electronics industry has pioneered over the same period. The decrease in feature size has led to the production of

Over the past fifty years, the development of sensors for biological applications has increased dramatically. This rapid growth can be attributed in part to the reduction in feature size, which the electronics industry has pioneered over the same period. The decrease in feature size has led to the production of microscale sensors that are used for sensing applications, ranging from whole-body monitoring down to molecular sensing. Unfortunately, sensors are often developed without regard to how they will be integrated into biological systems. The complexities of integration are underappreciated. Integration involves more than simply making electrical connections. Interfacing microscale sensors with biological environments requires numerous considerations with respect to the creation of compatible packaging, the management of biological reagents, and the act of combining technologies with different dimensions and material properties. Recent advances in microfluidics, especially the proliferation of soft lithography manufacturing methods, have established the groundwork for creating systems that may solve many of the problems inherent to sensor-fluidic interaction. The adaptation of microelectronics manufacturing methods, such as Complementary Metal-Oxide-Semiconductor (CMOS) and Microelectromechanical Systems (MEMS) processes, allows the creation of a complete biological sensing system with integrated sensors and readout circuits. Combining these technologies is an obstacle to forming complete sensor systems. This dissertation presents new approaches for the design, fabrication, and integration of microscale sensors and microelectronics with microfluidics. The work addresses specific challenges, such as combining commercial manufacturing processes into biological systems and developing microscale sensors in these processes. This work is exemplified through a feedback-controlled microfluidic pH system to demonstrate the integration capabilities of microscale sensors for autonomous microenvironment control.
ContributorsWelch, David (Author) / Blain Christen, Jennifer (Thesis advisor) / Muthuswamy, Jitendran (Committee member) / Frakes, David (Committee member) / LaBelle, Jeffrey (Committee member) / Goryll, Michael (Committee member) / Arizona State University (Publisher)
Created2012
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Description
Safety and efficacy of neuromodulation are influenced by abiotic factors like failure of implants, biotic factors like tissue damage, and molecular and cellular mechanisms of neuromodulation. Accelerated lifetime test (ALT) predict lifetime of implants by accelerating failure modes in controlled bench-top conditions. Current ALT models do not capture failure modes

Safety and efficacy of neuromodulation are influenced by abiotic factors like failure of implants, biotic factors like tissue damage, and molecular and cellular mechanisms of neuromodulation. Accelerated lifetime test (ALT) predict lifetime of implants by accelerating failure modes in controlled bench-top conditions. Current ALT models do not capture failure modes involving biological mechanisms. First part of this dissertation is focused on developing ALTs for predicting failure of chronically implanted tungsten stimulation electrodes. Three factors used in ALT are temperature, H2O2 concentration, and amount of charge delivered through electrode to develop a predictive model of lifetime for stimulation electrodes. Second part of this dissertation is focused on developing a novel method for evaluating tissue response to implants and electrical stimulation. Current methods to evaluate tissue damage in the brain require invasive and terminal procedures that have poor clinical translation. I report a novel non-invasive method that sampled peripheral blood monocytes (PBMCs) and used enzyme-linked immunoassay (ELISA) to assess level of glial fibrillary acidic protein (GFAP) expression and fluorescence-activated cell sorting (FACS) to quantify number of GFAP expressing PBMCs. Using this method, I was able to detect and quantify GFAP expression in PBMCs. However, there was no statistically significant difference in GFAP expression between stimulatory and non-stimulatory implants. Final part of this dissertation assessed molecular and cellular mechanisms of non-invasive ultrasound neuromodulation approach. Unlike electrical stimulation, cellular mechanisms of ultrasound-based neuromodulation are not fully known. Final part of this dissertation assessed role of mechanosensitive ion channels and neuronal nitric oxide production in cell cultures under ultrasound excitation. I used fluorescent imaging to quantify expression of nitric oxide in neuronal cell cultures in response to ultrasound stimulation. Results from these experiments indicate that neuronal nitric oxide production increased in response to ultrasound stimulation compared to control and decreased when mechanosensitive ion channels were suppressed. Two novel methods developed in this dissertation enable assessment of lifetime and safety of neuromodulation techniques that use electrical stimulation through implants. The final part of this dissertation concludes that non-invasive ultrasound neuromodulation may be mediated through neuronal nitric oxide even in absence of activation of mechanosensitive ion channels.
ContributorsVoziyanov, Vladislav (Author) / Muthuswamy, Jitendran (Thesis advisor) / Smith, Barbara (Committee member) / Greger, Bradley (Committee member) / Abbas, James (Committee member) / Okandan, Murat (Committee member) / Arizona State University (Publisher)
Created2022