Biology and Biomedical Sciences: Neurosciences
Date of Award
Doctor of Philosophy (PhD)
Chair and Committee
There are an estimated 1.6-3.8 million concussions in the United States annually. Individuals who experience a single concussion are at low risk for long-term consequences. However, there is mounting evidence that experiencing multiple concussions can lead to persistent symptoms, cognitive impairment, and increased risk for neurodegenerative disease. The underlying pathophysiology of concussions is not well understood.
To study the mechanisms that lead to these long-term consequences, a mouse model of repetitive concussive traumatic brain injury (rcTBI) was developed. Initial studies sought to characterize the histological and functional changes that occur after two closed-skull impacts in mouse. Similar to human traumatic brain injury, rcTBI produced axonal injury evident by amyloid precursor protein, neurofilament, and silver staining abnormalities in the absence of gross structural changes or cell loss. Microglia and astrocytes both became activated and were prominent in injured white matter by 7 days. Behaviorally, injury resulted in acute Morris Water Maze deficits that were not completely recovered by 7 weeks post-injury. Functionally, the velocity of axonal compound action potentials was slowed in both myelinated and unmyelinated axons. These alterations were accompanied by changes in white matter that were detectable by in vivo diffusion tensor magnetic resonance imaging (MRI). At 7 days post-injury, mean diffusivity and the diffusion of water parallel to axons in the corpus callosum and external capsule was reduced. However, these parameters did not correlate with increases in silver staining or microglial activation and indicate a need to develop better histological methods for assessing axonal injury after mild trauma. Future experiments will be conducted to quantify axonal injury by array tomography, a method outlined here briefly.
The second goal of this work was to determine what factors might contribute to axonal injury in concussion. Specifically, the hypothesis that microglia may increase axonal injury acutely following rcTBI was tested. The CD11b-TK mouse line, a valganciclovir-inducible model of microglial depletion, was used to reduce microglia within the corpus callosum and external capsule by 35%. Quantification of silver staining determined that this had no effect on axonal injury at 7 or 21 days after rcTBI. Further reduction by 56% also did not alter axonal injury detectable by silver staining, APP or neurofilament labeling, or by electron microscopy. We additionally tested several pharmacological compounds to determine whether they could reduce the microglial response. None of the compounds tested--including minocycline, (RS)-2-Chloro-5-hydroxyphenylglycine (CHPG), brilliant blue g (BBG), and microRNA-124 (miRNA-124)--were able to reduce the number of iba-1-positive microglia present in the corpus callosum after injury, which were quantified by stereology. Silver staining was unaffected. Use of the targeted toxin, Mac-1-Saporin, was found to dramatically reduce microglial number but also result in non-specific neuronal loss days 7 after rcTBI. Collectively, these experiments indicate that microglia appear to play a neutral role in regards to axonal injury acutely after repeat concussion. To test the role of microglia in the long-term, additional tools to manipulate the microglial response will need to be developed.
Last, the contribution of Apolipoprotein E to axonal injury after moderate-severe traumatic brain injury was assessed in a transgenic mouse model carrying three human familial AD mutations (PS1M146V, tauP301L, and APPSWE). The Apolipoprotein E4 (APOE4) genotype is a risk factor for poor outcome following traumatic brain injury, especially in young patients. By analogy to APOE4's effects on the risk of Alzheimer's disease, one hypothesis is that APOE genotype influences amyloid-beta (Aβ) and tau deposition following injury. Surprisingly, the amount of amyloid-beta and tau as measured by stereology was similar between mice possessing the APOE2, 3, or 4 allele. However, APOE4 mice had significantly greater numbers of APP-positive axons. These results suggest that the APOE4 genotype may have a primary effect on the severity of axonal injury in the setting of acute traumatic brain injury.
Altogether, this work presents the characterization of a mouse model of repetitive concussive traumatic brain injury that can be utilized to determine what factors contribute to pathophysiological changes and to aid in the design of future therapeutics.
Bennett, Rachel, "Axonal Damage in Repetitive Concussive Traumatic Brain Injury: Characterization and Contributing Factors" (2014). All Theses and Dissertations (ETDs). 1217.