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Description
In somatic cells, the mitotic spindle apparatus is centrosomal and several isoforms of Protein Kinase C (PKC) have been associated with the mitotic spindle, but their role in stabilizing the mitotic spindle is unclear. Other protein kinases such as, Glycogen Synthase Kinase 3â (GSK3â) also have been shown to be associated with the mitotic spindle. In the study in chapter 2, we show the enrichment of active (phosphorylated) PKCæ at the centrosomal region of the spindle apparatus in metaphase stage of 3T3 cells. In order to understand whether the two kinases, PKC and GSK3â are associated with the mitotic spindle, first, the co-localization and close molecular proximity of PKC isoforms with GSK3â was studied in metaphase cells. Second, the involvement of inactive GSK3â in maintaining an intact mitotic spindle was shown. Third, this study showed that addition of a phospho-PKCæ specific inhibitor to cells can disrupt the mitotic spindle microtubules. The mitotic spindle at metaphase in mouse fibroblasts appears to be maintained by PKCæ acting through GSK3â. The MAPK pathway has been implicated in various functions related to cell cycle regulation. MAPKK (MEK) is part of this pathway and the extracellular regulated kinase (ERK) is its known downstream target. GSK3â and PKCæ also have been implicated in cell cycle regulation. In the study in chapter 3, we tested the effects of inhibiting MEK on the activities of ERK, GSK3â, PKCæ, and á-tubulin. Results from this study indicate that inhibition of MEK did not inhibit GSK3â and PKCæ enrichment at the centrosomes. However, the mitotic spindle showed a reduction in the pixel intensity of microtubules and also a reduction in the number of cells in each of the M-phase stages. A peptide activation inhibitor of ERK was also used. Our results indicated a decrease in mitotic spindle microtubules and an absence of cells in most of the M-phase stages. GSK3â and PKCæ enrichment were however not inhibited at the centrosomes. Taken together, the kinases GSK3â and PKCæ may not function as a part of the MAPK pathway to regulate the mitotic spindle.
ContributorsChakravadhanula, Madhavi (Author) / Capco, David G. (Thesis advisor) / Chandler, Douglas (Committee member) / Clark-Curtiss, Josephine (Committee member) / Newfeld, Stuart (Committee member) / Arizona State University (Publisher)
Created2012
Description
Sensory gating is a process by which the nervous system preferentially admits stimuli that are important for the organism while filtering out those that may be meaningless. An optimal sensory gate cannot be static or inflexible, but rather plastic and informed by past experiences. Learning enables sensory gates to recognize stimuli that are emotionally salient and potentially predictive of positive or negative outcomes essential to survival. Olfaction is the only sensory modality in mammals where sensory inputs bypass conventional thalamic gating before entering higher emotional or cognitive brain regions. Thus, olfactory bulb circuits may have a heavier burden of sensory gating compared to other primary sensory circuits. How do the primary synapses in an olfactory system "learn"' in order to optimally gate or filter sensory stimuli? I hypothesize that centrifugal neuromodulator serotonin serves as a signaling mechanism by which primary olfactory circuits can experience learning informed sensory gating. To test my hypothesis, I conditioned genetically-modified mice using reward or fear olfactory-cued learning paradigms and used pharmacological, electrophysiological, immunohistochemical, and optical imaging approaches to assay changes in serotonin signaling or functional changes in primary olfactory circuits. My results indicate serotonin is a key mediator in the acquisition of olfactory fear memories through the activation of its type 2A receptors in the olfactory bulb. Functionally within the first synaptic relay of olfactory glomeruli, serotonin type 2A receptor activation decreases excitatory glutamatergic drive of olfactory sensory neurons through both presynaptic and postsynaptic mechanisms. I propose that serotonergic signaling decreases excitatory drive, thereby disconnecting olfactory sensory neurons from odor responses once information is learned and its behavioral significance is consolidated. I found that learning induced chronic changes in the density of serotonin fibers and receptors, which persisted in glomeruli encoding the conditioning odor. Such persistent changes could represent a sensory gate stabilized by memory. I hypothesize this ensures that the glomerulus encoding meaningful odors are much more sensitive to future serotonin signaling as such arousal cues arrive from centrifugal pathways originating in the dorsal raphe nucleus. The results advocate that a simple associative memory trace can be formed at primary sensory synapses to facilitate optimal sensory gating in mammalian olfaction.
ContributorsLi, Monica (Author) / Tyler, William J (Thesis advisor) / Smith, Brian H. (Thesis advisor) / Duch, Carsten (Committee member) / Neisewander, Janet (Committee member) / Vu, Eric (Committee member) / Arizona State University (Publisher)
Created2012
Description
Research suggests that the more positive the first drug experience, the more likely addiction will develop. Since smoking is initiated in a social setting, it is surprising how little is known about social context effects on acquisition of nicotine self-administration. We investigated this issue in rats during late adolescence using conjoined self-administration chambers that had a removable shared wall. Rats were assigned to training conditions with either a solid black plexiglass or wire mesh partition in place throughout 22 subsequent 2-hour daily training sessions. Initially, 58 day-old (late-adolescent) male and female rats received 2, 30-min habituation sessions/day over 2 consecutive days, with only an inactive lever present. Sessions began with presentation of a retractable lever and thereafter each response on that lever resulted in simultaneous delivery of saline or 1 of 2 doses of nicotine (0.015 or 0.030 mg/kg, IV) and lever retraction for a 20-second time out. The findings indicate that the social context inhibits nicotine self-administration in female rats during the development of addiction, but has little effect on the initial stages of drug acquisition. Furthermore, the data suggest that in male rats the social context enhances responding independent of nicotine, but has few effects on nicotine self-administration during the development of addiction. The findings have important implications for substance use disorders.
ContributorsDufwenberg, Martin (Author) / Neisewander, Janet (Thesis director) / Deviche, Pierre (Committee member) / Peartree, Natalie (Committee member) / Barrett, The Honors College (Contributor) / School of Mathematical and Statistical Sciences (Contributor) / School of Politics and Global Studies (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
Consequences of drug abuse and addiction affect both men and women, but women tend to rapidly progress through drug addiction phases, have higher drug dependency, and have higher relapse rates. Ovarian hormones fluctuate with female reproductive cycles and are thought to cause increased sensitivity to psychostimulants. Additionally, intermittent social defeat stress induces social avoidance, weight loss, and long-lasting cross-sensitization to psychostimulants, which is associated with increased FosB/ΔFosB expression in the nucleus accumbens (NAc) shell. In this study, we examined the estrous cycle in female rats on social defeat stress-induced amphetamine cross-sensitization through FosB/ΔFosB expression in the NAc shell. Every third day for ten days, we induced social defeat stress in rats through short confrontations with a lactating female resident rat and her pups. In parallel, a group of rats were handled for control. Vaginal swabs were taken daily to assess estrous stage. Ten days after the last stress exposure, rats were administered a low dose of amphetamine (0.5 mg/kg, i.p.), which induced cross-sensitization in stressed rats, evidenced by enhanced locomotor activity. Approximately 3-10 days after amphetamine challenge, brain tissue was collected for immunohistochemistry analyses. Stressed female rats had lower body weight gain, higher social avoidance, and increased FosB/ΔFosB expression in the NAc shell. Differences in FosB/ΔFosB expression in the NAc shell was also observed in handled animals in different estrous stages. Furthermore, rats in proestrous/estrous stages displayed enhanced social defeat stress-induced amphetamine cross-sensitization in comparison to rats in metestrous/diestrous stages. Elucidating the effects of the female reproductive cycle on drug use may provide a novel approach to treatments or therapies in preventing women’s stress-induced vulnerability to substance abuse.
ContributorsAzuma, Alyssa (Author) / Neisewander, Janet (Thesis director) / Nikulina, Ella (Thesis director) / Hammer, Ronald (Committee member) / School of Mathematical and Natural Sciences (Contributor) / Watts College of Public Service & Community Solut (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
MicroRNAs are small, non-coding transcripts that control gene expression by preventing mRNA from translating into proteins. They have been implicated to play a role in many drug addictions. We previously found that miR-495 targets several addiction-related genes (ARGs) and is highly expressed in the nucleus accumbens (NAc). We also found miR-495 is downregulated in the NAc following acute cocaine administration, and cocaine motivation measured by breakpoint on a progressive ratio schedule of cocaine reinforcement is decreased when miR-495 is overexpressed. In this study, we manipulated the endogenous levels of miR-495 by using a viral vector. Using an animal model, rats were first trained for self-administration on a fixed ratio (FR) schedule of reinforcement. After they were infused with a lentivirus to overexpress (LV-miR-495) or decrease (LV-Sponge) miR-495, in the NAc shell. The rats were then tested for extinction and reinstatement of cocaine-seeking behavior, which are measures of motivation for cocaine. We measured the relative levels of miR-495 in the NAc shell using qRT-PCR. Our results show that overexpression of miR-495 decreased cocaine-seeking behavior during extinction and cocaine reinstatement, as we hypothesized. Surprisingly, miR-495 LV-sponge also decreased cocaine-seeking behavior in extinction, not as we hypothesized. However, we found that LV-Sponge failed to significantly decrease levels of miR-495 as intended. In conclusion, understanding why LV-Sponge decreased, rather than increased, miR-495 will need further study, however, the results with LV-miR-495 extend previous findings that miR-495 plays a vital role in the molecular mechanism that influences motivation to seek cocaine.
ContributorsChaudhury, Trisha (Author) / Neisewander, Janet (Thesis director) / Newbern, Jason (Committee member) / Powell, Gregory (Committee member) / Department of Psychology (Contributor) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2017-05
Description
Systems biology studies complex biological systems. It is an interdisciplinary field, with biologists working with non-biologists such as computer scientists, engineers, chemists, and mathematicians to address research problems applying systems’ perspectives. How these different researchers and their disciplines differently contributed to the advancement of this field over time is a question worth examining. Did systems biology become a systems-oriented science or a biology-oriented science from 1992 to 2013?
This project utilized computational tools to analyze large data sets and interpreted the results from historical and philosophical perspectives. Tools deployed were derived from scientometrics, corpus linguistics, text-based analysis, network analysis, and GIS analysis to analyze more than 9000 articles (metadata and text) on systems biology. The application of these tools to a HPS project represents a novel approach.
The dissertation shows that systems biology has transitioned from a more mathematical, computational, and engineering-oriented discipline focusing on modeling to a more biology-oriented discipline that uses modeling as a means to address real biological problems. Also, the results show that bioengineering and medical research has increased within systems biology. This is reflected in the increase of the centrality of biology-related concepts such as cancer, over time. The dissertation also compares the development of systems biology in China with some other parts of the world, and reveals regional differences, such as a unique trajectory of systems biology in China related to a focus on traditional Chinese medicine.
This dissertation adds to the historiography of modern biology where few studies have focused on systems biology compared with the history of molecular biology and evolutionary biology.
This project utilized computational tools to analyze large data sets and interpreted the results from historical and philosophical perspectives. Tools deployed were derived from scientometrics, corpus linguistics, text-based analysis, network analysis, and GIS analysis to analyze more than 9000 articles (metadata and text) on systems biology. The application of these tools to a HPS project represents a novel approach.
The dissertation shows that systems biology has transitioned from a more mathematical, computational, and engineering-oriented discipline focusing on modeling to a more biology-oriented discipline that uses modeling as a means to address real biological problems. Also, the results show that bioengineering and medical research has increased within systems biology. This is reflected in the increase of the centrality of biology-related concepts such as cancer, over time. The dissertation also compares the development of systems biology in China with some other parts of the world, and reveals regional differences, such as a unique trajectory of systems biology in China related to a focus on traditional Chinese medicine.
This dissertation adds to the historiography of modern biology where few studies have focused on systems biology compared with the history of molecular biology and evolutionary biology.
ContributorsZou, Yawen (Author) / Laubichler, Manfred (Thesis advisor) / Maienschein, Jane (Thesis advisor) / Creath, Richard (Committee member) / Ellison, Karin (Committee member) / Newfeld, Stuart (Committee member) / Arizona State University (Publisher)
Created2016
Description
MicroRNAs are small, non-coding transcripts that post-transcriptionally regulate expression of multiple genes. Recently microRNAs have been linked to the etiology of neuropsychiatric disorders, including drug addiction. Following genome-wide sequence analyses, microRNA-495 (miR-495) was found to target several genes within the Knowledgebase of Addiction-Related Genes (KARG) database and to be highly expressed in the nucleus accumbens (NAc), a pivotal brain region involved in reward and motivation. The central hypothesis of this dissertation is that NAc miR-495 regulates drug abuse-related behavior by targeting several addiction-related genes (ARGs). I tested this hypothesis in two ways: 1) by examining the effects of viral-mediated miR-495 overexpression or inhibition in the NAc of rats on cocaine abuse-related behaviors and gene expression, and 2) by examining changes in NAc miR-495 and ARG expression as a result of brief (i.e., 1 day) or prolonged (i.e., 22 days) cocaine self-administration. I found that behavioral measures known to be sensitive to motivation for cocaine were attenuated by NAc miR-495 overexpression, including resistance to extinction of cocaine conditioned place preference (CPP), cocaine self-administration on a high effort progressive ratio schedule of reinforcement, and cocaine-seeking behavior during both extinction and cocaine-primed reinstatement. These effects appeared specific to cocaine, as there was no effect of NAc miR-495 overexpression on a progressive ratio schedule of food reinforcement. In contrast, behavioral measures known to be sensitive to cocaine reward were not altered, including expression of cocaine CPP and cocaine self-administration under a low effort FR5 schedule of reinforcement. Importantly, the effects were accompanied by decreases in NAc ARG expression, consistent with my hypothesis. In further support, I found that NAc miR-495 levels were reduced and ARG levels were increased in rats following prolonged, but not brief, cocaine self-administration experience. Surprisingly, inhibition of NAc miR-495 expression also decreased both cocaine-seeking behavior during extinction and NAc ARG expression, which may reflect compensatory changes or unexplained complexities in miR-495 regulatory effects. Collectively, the findings suggest that NAc miR-495 regulates ARG expression involved in motivation for cocaine. Therefore, using microRNAs as tools to target several ARGs simultaneously may be useful for future development of addiction therapeutics.
ContributorsBastle, Ryan (Author) / Neisewander, Janet (Thesis advisor) / Newbern, Jason (Committee member) / Nikulina, Ella (Committee member) / Perrone-Bizzozero, Nora (Committee member) / Sanabria, Federico (Committee member) / Arizona State University (Publisher)
Created2016
Description
Intervertebral Disc Degeneration (IVDD) is a complex phenomenon characterizing the desiccation and structural compromise of the primary joint in the human spine. The intervertebral disc (IVD) serves to connect vertebral bodies, cushion shock, and allow for flexion and extension of the vertebral column. Often presenting in the 4th or 5th decades of life as low back pain, this disease was originally believed to be the result of natural “wear and tear” coupled with repetitive mechanical insult, and as such most studies focus on patients between 40 and 50 years of age. Research over the past two decades, however, has demonstrated that environmental factors have only a modest effect on disc degeneration, with genetic influences playing a much more substantial role. Extensive research has focused on this process, though definitive risk factors and a clear pathophysiology have proven elusive. The aim of this study was to assemble a cohort of patients exhibiting definitive signs of degeneration who were well below the average age of presentation, with minimal or no exposure to suspected environmental risk factors and to conduct a targeted genome analysis in an attempt to elucidate a common genetic component. Through whole genome sequencing and analysis, the results corroborated findings in a previous study, as well as demonstrated a potential connection and influence between mutations found in IVD structural or functional genes, and the provocation of IVDD. Though the sample size was limited in scale and age, these findings suggest that further IVDD research into the association of variants in collagen, aggrecan and the insulin-like growth factor receptor genes of young patients with an early presentation of disc degeneration and minimal exposure to suspected risk factors is merited.
ContributorsFulton, Travis (Author) / Liebig, Juergen (Thesis advisor) / Neisewander, Janet (Committee member) / Theodore, Nicholas (Committee member) / Arizona State University (Publisher)
Created2016