This manuscript uses sound localization circuitry in the chick brainstem to examine the balance of inhibitory and excitatory neurotransmission maintained by neural circuits. This is a particular challenge in sensory systems, like the auditory system, in which this balance must be maintained despite variation in the acoustic environment. The studies described in this manuscript help clarify why and how inhibition is limited during two periods of changing auditory input: deafness and auditory development. In the cochlear nucleus, nucleus magnocellularis (NM), deprivation of excitatory input induced by deafness triggers neuronal death. While this neuronal death has previously been accredited to the loss of excitatory drive, the present experiments examine an alternative hypothesis: that inhibitory input to NM, which may also be affected by deafness, contributes to neuronal death in NM. Using an in vitro slice preparation in which excitatory input from the auditory nerve is absent, we pharmacologically altered GABA receptor activation in NM, and assayed an early marker of neuronal health, antigenicity for the ribosomal antibody Y10B (Y10B-ir). We found that GABA decreases Y10B-ir, and that GABAA activation is necessary for the GABA-induced effect. We further found that endogenous GABAA activation similarly decreases Y10B-ir and this decrease requires extracellular Ca2+ Our results suggest that, in the absence of excitatory input, endogenous activation of ionotropic GABAA receptors is detrimental to NM neurons. During auditory development, changes in membrane properties promote the ability of excitatory neurons in the brainstem to code aspects of sound, including the level and timing of a stimulus. Some of these changes coincide with the onset of hearing, suggesting that sound-driven activity in the nervous system produces developmental plasticity of ion channel expression. While it is known that the coding properties of excitatory neurons are modulated by inhibition in the mature system, it is unknown whether there are also developmental changes in the membrane properties of brainstem inhibitory neurons. We investigated the primary source of inhibition in the avian auditory brainstem, the superior olivary nucleus (SON), which displays sound-driven activity in mature animals. The present studies test the hypothesis that, as in excitatory neurons, the membrane properties of these inhibitory neurons also change following hearing onset. We examined SON neurons at different stages of auditory development: embryonic days 14-15 (E14-15), a time point at which cochlear ganglion neurons are just beginning to respond to sound, later stages of embryonic development (E18-19), and after hatching (P0-P1). We used in vitro whole-cell patch electrophysiology to explore physiological changes in SON. Age-related changes were observed at the level of a single spike and in multi-spiking behavior. In particular, tonic behavior, measured as a neuron's ability to sustain tonic firing over a range of current steps, became more common later in development. Voltage-clamp recordings and biophysical models were employed to examine how age-related increases in sodium and potassium currents enhance excitability in SON. Together the two sets of experiments suggest a new role for inhibition in the auditory brainstem. While it was previously known that inhibition complements excitation to improve auditory coding in a stable system, these experiments reveal that under less stable circumstances, inhibition actively contributes to plasticity in the brainstem circuit. / A Dissertation submitted to the Department of Psychology in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Spring Semester 2017. / April 6, 2017. / Includes bibliographical references. / Richard Hyson, Professor Directing Dissertation; Doug Schrock, University Representative; Richard Bertram, Committee Member; Frank Johnson, Committee Member; Chris Schatschneider, Committee Member.

In preclinical research, females are often excluded because of the presumed variability associated with the cyclic fluctuations of the estrous cycle and pregnancy. But how much variability in biological data actually occurs as a result of these events? As biological events unique to females, the estrous cycle and pregnancy deserve to be examined not only for their unique attributes, but also to incorporate this data into the larger biological context of male and female reproductive experience. To address this, we analyzed the transcriptome of the C57BL/6J (B6) mouse hypothalamus, hippocampus, neocortex, and cerebellum to identify gene expression changes between the four brain regions as well as gene expression changes associated with the estrous cycle, pregnancy, and the postpartum period using RNA-sequencing. In naturally cycling virgin mice, we found 15,947 genes are expressed in one or more of the brain regions and ~92% of these genes are differentially expressed between the four brain regions. In contrast, the genes differentially expressed as a result of the estrous cycle only account for ~1.3% of the expressed genes and the 210 estrous cycle-specific differentially expressed genes (DEGs) are primarily expressed in only one brain region during one stage transition. In a separate study, we identified 3,737 DEGs as a result of pregnancy, parturition, and the postpartum period. Each brain region and time point shows a unique molecular signature, with only 49 DEGs in all four brain regions. Several genes previously implicated in postpartum depression also change expression during pregnancy and the postpartum period. Interestingly, the unique set of genes in each of the brain regions that change during pregnancy, parturition, and the postpartum period are distinct from the genes that change in response to the estrous cycle. Taken together, the work presented in this manuscript emphasizes the stability of the female mouse brain during the estrous cycle, especially in contrast to the dramatic changes that occur between the four brain regions and also as a result of pregnancy. To put these numbers into perspective, there are 18 times more gene expression changes associated with pregnancy, parturition, and the postpartum period in these four brain regions than as a result of the estrous cycle. Furthermore, in the hippocampus, the DEGs between the sexes (Vied et al., 2016) are 11 times greater than the DEGs as a result of the estrous cycle. These findings enhance our understanding of female-specific changes in gene expression and supports the inclusion of females in preclinical research. / A Dissertation submitted to the Department of Biomedical Sciences in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Spring Semester 2017. / March 3, 2017. / cerebellum, estrous cycle, hippocampus, hypothalamus, neocortex, pregnancy / Includes bibliographical references. / Richard S. Nowakowski, Professor Directing Dissertation; Brian Inouye, University Representative; Pradeep Bhide, Committee Member; Thomas Houpt, Committee Member; Yi Ren, Committee Member.

Oxytocin (OXT) is a peptide with important regulatory roles in both physiological and behavioral contexts. OXT binds to the OXT receptor (OXTR) in the central and peripheral nervous systems, with diverse patterns of expression dependent on many variables including but not limited to species, sex, and stage of development. OXT is an important hormone during adulthood for the transition to parenthood in many species, from mating behaviors through parturition all the way to parental care. OXT signaling in the parent is crucial during these stages, and deficits in parental OXT can have persisting developmental consequences for the offspring. However, less research has investigated how the infant processes OXT-mediated parental care. The aims of this research were to assess peripheral sites of OXTR in the infant that may inform how the infant perceives OXT from the environment. This cross-species analysis sought to identify regions that were conserved among species and identify differences in OXTR expression that may correlate with developmental behaviors. A novel site of OXTR-binding, the eye, was further assessed in neonates and adults for the presence of mRNA that could potentially inform synthesis and function. Finally, a battery of behavioral tasks to assess differences in pupillary responsiveness were performed in adult transgenic mice strains of Oxt and Oxtr wild-type and knockout. Results from OXTR autoradiography indicated several sites of specific binding in the mouse, prairie vole, and rat. There were species and strain differences in regions of interest including the periodontium and the ciliary bodies of the eye. OXTR autoradiography in adult mice demonstrated that OXTR in the ciliary bodies persist into adulthood. Oxtr mRNA was detected by in situ hybridization (ISH) in the neonatal mouse eye and by reverse-transcriptase polymerase chain reaction (RT-PCR) in the adult mouse eye. Regions relevant to pupillary modulation and processing sensory information from the eye displayed robust Oxtr signal by ISH, including the conjunctiva, ciliary bodies, ciliary ganglion, oculomotor nerve, and superior cervical ganglion. Behavioral assessments in adult mice demonstrated a significantly constricted baseline pupil diameter in Oxt knockout mice, which is rescued by the topical application of OXT. These data support a potential role for OXT mediating autonomic development in the visual system. / A Dissertation submitted to the Department of Psychology in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Spring Semester 2019. / April 5, 2019. / Cross-species, Knockout, Mouse, Oxytocin, Pupil / Includes bibliographical references. / Elizabeth A. D. Hammock, Professor Directing Dissertation; Xiuwen Liu, University Representative; Walter Richard Boot, Committee Member; James M. Fadool, Committee Member; Frank Johnson, Committee Member.

In neuroscience, zebrafish can be used as a model organism to study cognitive deficits and human physiology. Previous methods to study behavior in animal models have relied on manual observations, which can lack repeatability, until recently when automatic operant boxes have been developed. Due to the automated testing, automatic operant boxes are a robust method with high throughput and minimal handling which reduce stress in the animal. The aim of this study was to establish and optimize a method and a protocol for studying reversal learning in Zantiks AD units (an automatic operant box). The Zantiks AD unit was controlled by predefined scripts and the purpose of the script was supposed to make the fish associate a visual stimulus with a positive reinforcement, in this study, dry food. Three pilot studies were made, each with one parameter in focus (habituation days, intertrial time and motivation) to see if there was any impact on learning, between two groups. Neither the habituation days (pilot 1) nor the intertrial time (pilot 2) seemed to have a pronounced effect but the impact of motivation (pilot 3) did. The impact of motivation needs to be analyzed further, where different types of food could be compared.

Alzheimer’s disease (AD) is a progressive neurodegenerative disease, which is characterized by formation of amyloid plaques in the brain. The major constituent of these plaques is the hydrophobic peptide Aβ. Aβ accumulation is considered to be the main cause of the pathology seen in AD brains. Aβ is produced through sequential cleavage of the amyloid precursor protein (APP). APP can be processed by two different enzymatic pathways. Formation of Aβ requires cleavage of APP by β- and γ-secretase. However, most proteolytic processing of APP does not result in Aβ formation. Instead, APP is mainly cleaved by α-secretase, which not only precludes formation of the toxic Aβ peptide but also generates the neuroprotective sAPPα fragment. Increasing the α-secretase processing of APP is thereby a potential therapeutic strategy for AD. APP is a member of a conserved gene family, also including the APP-like proteins-1 and -2 (APLP1 and APLP2). The APP family members have essential and overlapping functions and have been reported to be processed in a similar way by the same enzymes. The processing of all APP family members is increased in response to several stimuli, including retinoic acid (RA) and insulin-like growth factor-1 (IGF-1), which also induce a shift towards α-secretase processing. The aim of this thesis was to investigate the mechanisms and signaling involved in induced α-secretase processing of the APP family. The main α-secretase candidates are ADAM10 and TACE. In this thesis we wanted to study the effects on expression levels of ADAM10 and TACE during RA treatment. We also wanted to investigate the mechanism behind IGF-1-induced processing of APP and APLP2. We found that both ADAM10 and TACE are up-regulated in response to RA, but that the signaling pathways involved differed between the two enzymes. Similarly, we showed that IGF-1-induced processing of APLP2, but not of APP, is dependent on PKC. Furthermore, we showed that ADAM10 is the main α-secretase for APP, whereas TACE cleaves APLP2 in response to IGF-1. We conclude that although APP and APLP2 proteolytic processing are induced by the same stimuli, the processing is dependent on different signaling pathways and processing enzymes, which in turn are differentially regulated.

Sleep has become less important in western society during modern times, where many have the habit of prioritizing productive activities instead of sufficient sleep. However, recent studies have indicated the importance of sleep for emotional processing. A crucial finding in literature regarding sleep and emotions has been the enhancement of negative emotions after sleep deprivation. The aim of this systematic literature review was to investigate the neural basis of the effects of sleep deprivation on emotional reactivity. In order to conduct this review, three databases were used to obtain relevant articles. Out of the total 1041 articles, 11 fulfilled the inclusion criteria and were included in the review. The selected articles exclusively contained results regarding reactivity to visual emotional stimuli. Results showed that total and partial sleep deprivation result in enhanced amygdala activity in response to negative stimuli. Enhanced amygdala activity was also found in response to positive and neutral stimuli after sleep deprivation. The insula was another brain region that displayed enhanced activity toward all types of valenced stimuli after sleep deprivation. Moreover, weaker connectivity between the amygdala and prefrontal areas (specifically the medial prefrontal cortex) was found after total and partial sleep loss. Together, these results suggest that sleep deprivation induces hyperreactivity toward emotional stimuli and disrupts top-down regulation of emotional reactivity.

We create a novel framework to enable the exploration of spatial calcium dynamics in astrocytes via compartmentalization of the cell domain. The cell is split up into spatially partitioned compartments representing sub-cellular domains with varied physiology. These compartments are modeled with systems of ordinary differential equations with diffusive transfer between compartments as in previous work. Stochasticity is leveraged to investigate the effect of mGluR-mediated inositol-triphosphate production which models the main form input due to neuronal activity. The compartmentalized astrocyte is able to represent several hierarchies of calcium dynamics across the cell in both spatial and temporal scales. / A Dissertation submitted to the Department of Scientific Computing in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / 2019 / November 14, 2019. / Astrocytes, Calcium Modeling, Compartmental, glia, neurodegenerative / Includes bibliographical references. / Gordon Erlebacher, Professor Directing Dissertation; Richard Bertram, University Representative; Anke Meyer-Baese, Committee Member; Michael Mascagni, Committee Member; Brian Quaife, Committee Member.

As dementia grows at an increasingly faster rate around the world, it has become imperative to identify its symptoms at an early stage. Dementia of the Alzheimer’s type (DAT) mainly disrupts cognitive functioning, but behavioral changes such as slow walking also occur. In an aging world with a growing dementia population, early identification has come to play an important role in the management and treatment of the disease. A simultaneous assessment of the behavioral and cognitive changes is recommended to be used to assist with early identification of dementia. A dual task of walking and talking is especially practical to identify early signs of dementia since individuals usually walk while doing something else such as talking on the phone or with another person. This study was an investigation into longitudinal changes in gait under different levels of cognitive load conditions: 1) the baseline condition was defined as normal walking with no simultaneous cognitive load, 2) the low cognitive load condition was defined as walking and counting numbers from a random number assigned, and finally 3) the high cognitive load condition was defined as walking and simultaneously completing a categorical naming task (generating as many words in a specific category as possible). Fourteen individuals with a mean age of 78 years who in the early (mild or moderate) stage of DAT (Mean DRS-2 score = 88.14, SD = 12.6) participated. For the walking task, the GAITRite© Portable Walkway system incorporating a 580-cm mat with embedded sensors to measure spatiotemporal parameters of gait and the accompanying software was employed. Once a month over a one-year period, participants walked across the GAITRite© Walkway mat. The gait analyses included Functional Ambulation Profile (FAP), stride length (SL), velocity, and double support time (DST). To more precisely examine the effects of concurrent cognitive load, participants completed the low and high cognitive load tasks while seated. Values in FAP, SL, and velocity decreased as tasks became more complex and as the disease progressed, while DST increased at the same rate. The comparison of walking and seated conditions indicated that participants’ performance on both number-counting and categorical naming tasks was poorer when they were walking rather than when they were seated. The results imply effectiveness of gait assessment during multitasking conditions for early diagnosis of DAT. Identifying DAT at an earlier stage is beneficial for the person with the disease, family members and caregivers, as well as health professionals. In this regard, the findings of this current investigation may establish a bridgehead for advances in the early identification of DAT. / A Dissertation submitted to the School of Communication Science and Disorders in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Summer Semester 2018. / May 15, 2018. / Cognitive Load, Dementia, Early Identification, Gait / Includes bibliographical references. / Richard J. Morris, Professor Co-Directing Dissertation; Leonard L. LaPointe, Professor Co-Directing Dissertation; Colleen M. Kelley, University Representative; Katlin L. Lansford, Committee Member; Charles G. Maitland, Committee Member.

Electrical signaling in the olfactory bulb (OB) is modulated by changes in metabolic state. The voltage-gated potassium channel, Kv1.3, makes up 60 - 80% of the outward current flow in mitral cells (MCs), the primary projection neurons of the OB. The metabolic molecules GLP-1, insulin, and glucose are present in the OB and modulate MC signaling by reducing the activity of Kv1.3. In obesity and diabetes, modulation of MC signaling and Kv1.3 current flow by these molecules is absent. Gene-targeted deletion of Kv1.3 (Kv1.3 -/-) produces a phenotype that encompasses changes in olfactory ability and metabolism. Kv1.3 -/- mice are “supersmellers”, with an enhanced ability to detect and discriminate odors, are leaner than their wild-type counterparts and resistant to diet-induced obesity. These observations suggest metabolism, the OB and Kv1.3 are intimately linked, providing opportunity for therapeutic intervention at the level of the voltage-gated potassium channel. Strategies towards targeting Kv1.3 in the OB, but not other regions of the brain or periphery, are desired. There are several natural modulators of Kv1.3 that can be utilized for therapeutic targeting of the channel. Nedd4-2 is an ubiquitin ligase that mediates ubiquination and degradation of target proteins and can act to regulate Kv1.3 channel density, while the adaptor protein Grb10 can mediate Nedd4-2 activity. Patch-clamp electrophysiology in HEK293 cells, SDS-PAGE, immunoprecipitation, and mutagenesis strategies demonstrated a channel/adaptor/ ligase signalplex. Mutation of the C-terminal, SH3-recognition or ubiquitination sites on Kv1.3 retained the observed co-immunoprecipitation between Nedd4-2/Kv1.3, while the latter prevented a reduction in channel density. A model based on these data is presented for which an atypical interaction may permit Nedd4-2/Kv1.3 interactions that lead to protein degradation and reduced current density, and can be disrupted by Nedd4-2/Grb10 interactions. Venom-derived ion channel inhibitors are a strong alternative to natively expressed Kv1.3 modulators. These inhibitors have strong channel selectivity, potency, and stability; however, tracking delivery to their target can be challenging. Margatoxin (MgTx) is a potent Kv1.3 inhibitor and conjugation to luminescent quantum dots (QDs) can provide a means to track its delivery. Towards this, two approaches were taken. Covalent conjugation of MgTx to QDs produced QD-MgTx, which exhibited a retention of known biophysical properties associated with block of the vestibule of Kv1.3. Towards a more efficient and controlled conjugation, a polyhistidine tagged MgTx (HisMgTxFSU) was produced and conjugated to QDs via polyhistidine-mediated self assembly (QDHisMgTxFSU). Similar to QD-MgTx, QDHisMgTxFSU had a strong ability to inhibit Kv1.3 in HEK293 cells, excite mitral cells of the OB, and label Kv1.3 expressing HEK293 cells. When delivered to the OB via cannula guided delivery, QDHisMgTxFSU failed to label Kv1.3 expressing mitral cells. Therapeutic effects due to delivery of QDHisMgTxFSU, however, were observed. To better understand the role of Kv1.3 in the OB and metabolism, HisMgTxFSU and QDHisMgTxFSU were delivered to the OB of obese mice via cannula-guided delivery and changes in metabolism were measured. Compared to control animals, targeted inhibition of Kv1.3 was found to prevent weight gain and reduce the respiratory quotient (RER). These data provide evidence for the first time that the OB plays a key role in energy regulation and the pathways involved in this regulation are proposed. / A Dissertation submitted to the Institute of Molecular Biophysics in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Summer Semester 2018. / July 16, 2018. / Electrophysiology, Ion Channel, Metabolism, Olfactory Bulb, Pharmacology, Quantum Dots / Includes bibliographical references. / Debra Ann Fadool, Professor Directing Dissertation; Thomas Houpt, University Representative; Timothy Cross, Committee Member; Richard Bertram, Committee Member; Yi Zhou, Committee Member.

The error-related negativity (ERN) is a widely-used electrophysiological measure of error processing in the brain. Whether the ERN supports conscious awareness of erring has yet to be determined. Some researchers report no interaction between ERN amplitude and error awareness, while others report a strict dependence of ERN amplitude on error awareness. Methodologically, it is unclear whether differences in reporting error awareness and/or different task demands across studies can account for the extant discrepancies. For example, it is unclear whether differences in stimulus perceptibility can account for conflicting results alone. Theoretically, discoveries from within error-awareness research as well as the larger current paradigm for studying the neural correlates of consciousness (NCC) have burgeoning implications for the phenomenology of erring. In order to address both methodological and theoretical problems with the literature on the electrophysiology of error awareness, this experiment used a novel visual search task. Results suggest that the ERN’s amplitude covaries with error awareness, and may be sensitive to the timing of subjective error sensations. However, due to a small sample size (n = 10) and in light of theoretical advancements of NCC research, the results obtained here are tentative. Suggestions for future research of the ERN and error awareness are discussed.