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- All Subjects: Neuroscience
- Creators: School of Life Sciences
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Description
Immediate early genes (IEGs) are rapidly activated in response to an environmental stimulus, and most code for transcription factors that mediate processes of synaptic plasticity, learning, and memory. EGR3, an immediate early gene transcription factor, is a mediator of biological processes that are disrupted in patients with schizophrenia (SCZ). A microarray experiment conducted by our lab revealed that Egr3 also regulates genes involved in DNA damage response. A recent study revealed that physiological neuronal activity results in the formation of DNA double-stranded breaks (DSBs) in the promoters of IEGs. Additionally, they showed that these DSBs are essential for inducing the expression of IEGs, and failure to repair these DSBs results in the persistent expression of IEGs. We hypothesize that Egr3 plays a role in repairing activity- induced DNA DSBs, and mice lacking Egr3 should have an abnormal accumulation of these DSBs. Before proceeding with that experiment, we conducted a preliminary investigation to determine if electroconvulsive stimulation (ECS) is a reliable method of inducing activity- dependent DNA damage, and to measure this DNA damage in three subregions of the hippocampus: CA1, CA3, and dentate gyrus (DG). We asked the question, are levels of DNA DSBs different between these hippocampal subregions in animals at baseline and following electroconvulsive stimulation (ECS)? To answer this question, we quantified γ-H2AX, a biomarker of DNA DSBs, in the hippocampal subregions of wildtype mice. Due to technical errors and small sample size, we were unable to substantiate our preliminary findings. Despite these shortcomings, our experimental design can be modified in future studies that investigate the role of Egr3 in activity-induced DNA damage repair.
ContributorsKhoshaba, Rami Samuel (Author) / Newbern, Jason (Thesis director) / Gallitano, Amelia (Committee member) / Marballi, Ketan (Committee member) / School of Molecular Sciences (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
Damage to DNA can affect the genes it encodes; if this damage is not repaired, abnormal proteins may be produced and cellular functions may be disturbed. DNA damage has been implicated in the initiation and progression of a variety of diseases. Conversely, DNA damage has also been discovered to contribute to beneficial biological processes. Madabhushi and colleagues (2015) determined that activity-dependent DNA double strand breaks (DSBs) in the promoter region of immediate early genes (IEGs) induced their expression. EGR3 is an IEG transcription factor which regulates the expression of growth factors and synaptic plasticity-associated genes. In a previously conducted microarray experiment, it was revealed that EGR3 regulates the expression of genes associated with DNA repair such as Cenpa and Nr4a2. These findings inspired us to investigate if EGR3 affects DNA repair in vivo. Before conducting this experiment, we sought to standardize and optimize a method of inducing DNA damage in the hippocampus. Electroconvulsive stimulation (ECS) is utilized to induce neuronal activity. Since neuronal activity leads to the formation of DNA DSBs, we theorized that ECS could be used to induce DNA DSBs in the hippocampus. We predicted that mice that receive ECS would have more DNA DSBs than those that receive the sham treatment. Gamma H2AX, a biomarker for DNA damage, was utilized to quantify DNA DSBs. Gamma H2AX expression in the dentate gyrus, CA1 and CA3 regions of the hippocampus was compared between mice that received the sham treatment and mice that received ECS. Mice that received ECS were sacrificed either 1 or 2 hours post-administration, constituting treatment conditions of 1 hr post-ECS and 2 hrs post-ECS. Our results suggest that ECS has a statistically significant effect exclusively in the CA1 region of the hippocampus. However, our analyses may have been limited due to sample size. A power analysis was conducted, and the results suggest that a sample size of n=4 mice will be sufficient to detect significant differences across treatments in all three regions of the hippocampus. Ultimately, future studies with an increased sample size will need to be conducted to conclusively assess the use of ECS to induce DNA damage within the hippocampus.
ContributorsAden, Aisha Abubakar (Author) / Newbern, Jason (Thesis director) / Gallitano, Amelia (Thesis director) / Marballi, Ketan (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
Chronic stress is a risk factor for many diseases that impact the brain, including Alzheimer’s Disease. Unlike acute stress, chronic stress reduces neuronal plasticity, which can lead to neuronal remodeling and suppression. This project investigates the effect of stress on the dendritic complexity of hippocampal neurons in rats, demonstrating a methodology for procuring and analyzing these neurons. The brains of the 160 rats from the Sustained Threat and Timing (STAT) experiment were frozen. The STAT experiment investigated the effect chronic variable stress had on prospective and retrospective timing in rodents. Using a cryostat, thin coronal slices of brain tissue were placed on microscopic slides. The tissue samples were then stained using the Golgi method of silver staining. Hippocampal neurons were assessed using Sholl Analysis; the dendritic complexity of these neurons was quantified. The method of using Sholl Analysis was found to be an effective process in measuring dendritic length of hippocampal neurons.
ContributorsMiller, Amara Delaney (Author) / Sanabria, Federico (Thesis director) / Gupta, Tanya (Committee member) / School of Life Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
Parkinson’s disease (PD) is a debilitating neurodegenerative disease characterized primarily by physical impairments such as tremors, poor balance, and bradykinesia; however, some individuals with PD will additionally experience numerous nonmotor symptoms such as dementia, depression, and sleep disturbances amongst various other life-altering ailments. Two of the key pathological hallmarks of PD include the death of melanated dopaminergic neurons in the nigrostriatal pathway and the accumulation of Lewy bodies, which are primarily composed of aggregates of the protein α-synuclein (α-syn). Interestingly, members of the chitinase protein family, namely chitinase-3-like protein-1 (L1), have heightened concentrations in a number of neurodegenerative diseases other than PD. To investigate the specific role L1 plays in PD etiology, we evaluated if astrocytic L1 expression was elevated in postmortem brain tissue of PD patients as well as in an α-syn overexpression rat model, and further tested if manipulating astrocytic-specific L1 expression correlated with neuroinflammation and nigral neuronal degeneration in the model. Preliminary histological analysis has shown increased levels of L1 expression in the α-syn model before neuronal loss occurs, and in human tissue, L1 was found to be significantly increased in the postmortem tissue of individuals with PD versus non-diseased controls. Investigations in identifying an astrocytic-specific virus capsid and manipulating L1 expression in the α-syn model are ongoing. This preliminary data thus far supports that increased astrocytic expression of L1 is associated with PD pathology.
ContributorsPettigrew, Tiffany (Author) / Manfredsson, Fredric (Thesis director) / Sandoval, Ivette (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor)
Created2023-12