Dyslexia is a common condition among both children and adults in the United States. Its prevalence is estimated to be between 5% and 17% of school-aged children. This chronic condition is characterized by a difficulty in reading unanticipated in children with otherwise average or above average intelligence, education and incentive. Although adults with a specific reading disability have demonstrated diminished diffusion anistropy, this relationship between reading ability and white matter connectivity is relatively unexplored in children. In this study, diffusion tensor magnetic resonance imaging (MRI) was utilized to compare the white matter connectivity in three groups of children with diverse reading ability- dysfluent and inaccurate (n= 42), dysfluent and accurate (n= 69) and nonimpaired (n= 23) children. ANOVA statistical analysis was performed detect any significant group differences in anisotropic indices between the three groups of readers. In contrast to past studies, our study did not reveal any statistically significant differences in fractional anisotropy, fiber coherence index and mean diffusivity between dysfluent and inaccurate, dysfluent but accurate, and non-impaired readers.

Functional magnetic resonance imaging (fMRI) is a non-invasive technique that allows for visualization of active brain regions. Although white matter (WM) constitutes approximately 50% of brain tissue, fMRI activation in WM has conventionally been dismissed. There are two main reasons WM fMRI remains controversial: 1) the blood oxygen level dependent (BOLD) fMRI signal depends on cerebral blood flow and volume, which are lower in WM than gray matter and 2) fMRI signal has been associated with post-synaptic potentials as opposed to action potentials. Despite these observations, there is no direct evidence against measuring fMRI activation in WM. This thesis is comprised of four manuscripts that investigate fMRI activation in WM at 4T. The first study evaluated whether it was possible to detect WM activation using an interhemispheric transfer task and examined whether certain MRI contrast mechanisms were more sensitive to activation in WM. Activation was detected in the anterior corpus callosum at the individual and group level and we discovered that T2 weighted imaging may provide increased sensitivity to activation in WM. The second study used two established interhemispheric transfer tasks to examine whether callosal activation could be experimentally manipulated using a within subjects design. The results replicated previous findings and demonstrated an ability to map functional activation in the corpus callosum that was task dependent. The third study examined WM fMRI activation in a different structure and focused on the posterior limb of the internal capsule using a motor task; activation was elicited at both individual and group levels. The fourth study linked advances in the ability to detect WM fMRI activation to current clinical approaches to the assessment of WM dysfunction. An adapted Symbol Digit Modalities Test was used to evaluate WM activation in healthy controls. The results revealed individual level activation in both the corpus callosum and internal capsule. Taken together this stream of research represents a major advance in the methods used to non-invasively study brain function. Future applications may include improved assessment methods for patients with WM dysfunction.

PURPOSE: To compare white matter neuron density across 3 regions, prefrontal, temporal, and posterior parietal (PFC, TE, PP) in macaque monkey, with further analysis of subdivisions within the gyral white matter. METHODS: Histological tissue from three adult macaque monkeys, previously prepared with the neuron-specific pan-neuronal marker Neuronal-N, was used for analysis. Tissue was digitized and processed electronically to investigate cross- and intra-regional differences in the distribution of white matter neurons. RESULTS: Statistical analysis showed significant differences across all regions sampled and across most intra-regional subdivisions, although the more conservative post-hoc tests failed to find significant differences between specific regions. CONCLUSIONS: The results of the current study support regional differences. Further studies using a larger sample size may help elucidate the relatively unknown properties of white matter neurons.

In this thesis, I have sought to address these issues by developing a toolbox called “Quantitative Tract Integrity Profiles (Q-TIPs)” that: is purely MATLAB-based (making it compatible with most operating systems), has a flexible and user-friendly interface, and will be distributed as a standalone toolbox for the popular Statistical Parametric Mapping (SPM) software package. This toolbox extracts the orientation of any ROI mask by calculating the medial axis using Voronoi and Delaunay algorithms, and then cross-sectional segments along the extracted medial axis are used to create profiles based on any quantitative white matter magnetic resonance imaging metric (e.g., diffusion tensor imaging, myelin water imaging, magnetization transfer imaging, etc.). As a result, this toolbox will enable future neuroimaging studies to more easily and flexibly quantify white matter imaging signals along individual tracts as a function of neural development, healthy aging, traumatic brain injury, or neurodegenerative disease. / February 2017

Hypertension has been associated with causing deleterious effects to the cerebrovasculature, which are thought to underlie the formation of white matter lesions (WML) and predispose individuals to age related cognitive decline. In humans hypertension frequently occurs concomitantly with other vascular risk factors making it difficult to ascertain the primary mechanisms of hypertension in isolation. Animal models of hypertension have been used in an aid to establish the mechanisms of hypertension in isolation. To date the knowledge gleaned from animal models has undoubtedly provided an insight as to the role of hypertension and cerebrovasculature remodelling but, these models have limitations such as lack of genetically matched controls and the inability to control the severity of hypertension, restricting the understanding of the underlying mechanisms. All studies within this thesis used the Cyp1a1 Ren2 inducible hypertensive rat model, induced by dietary addition of Indole-3-carbinol (I3C), allowing the severity and duration of hypertension to be tightly controlled and compared to genetically matched controls. This thesis set out to address the hypothesis that sustained hypertension will lead to alterations to the structural integrity of the cerebrovasculature and white matter, which will be exacerbated with age and that hypertension will be associated with alterations to gene expression and cognitive function. Initially this thesis sought to investigate the effect of hypertension on the structural integrity of the vasculature in the Cyp1a1 Ren2 rat model. Firstly, blood pressure in the Cyp1a1 Ren2 rat model was characterised and it was found that the dietary addition of I3C, caused a sustained level of increased blood pressure in all three cohorts. Cerebrovascular alterations were found to consist of increased eNOS expression in the young brain, which progressed with increased duration of hypertension to vascular morphological alterations of decreased vessel width and a redistribution of tight junction protein claudin-5. With age, hypertensive vascular alterations consisted of increased eNOS expression and vascular density. Additionally, there was evidence that hypertension caused a vascular inflammatory response in the young and aged brain. Secondly, this thesis investigated the effect of hypertension on gene expression. Overall it was found that hypertension altered genes related to collagen growth factors, ion channels, eNOS related Map-Kinase pathway and inflammatory genes. Thirdly, this thesis sought to investigate the impact of hypertension on the overall structural integrity of the brain and white matter examining neurons, myelin, oligodendrocytes, axons and microglia, in several regions of the young and aged brain. In general, this study found that hypertension did not cause overt structural or myelin alterations in the majority of regions analysed, with only evidence of myelin alterations occurring within the subcortex of hypertensive animals from each of the young cohorts analysed. However, an adverse subcortical inflammatory response was found in hypertensive animals of the young 6-month cohort and also in hypertensive animals from the aged 4-month cohort, where the inflammatory response was not exclusive to the subcortex of hypertensive animals but also occurred in multiple white matter tracts. Lastly this thesis chose to examine the effect of hypertension on cognitive function, specifically spatial reference and working memory using the Morris water maze and found no evidence of alterations in the cognitive functions examined. Conclusions The results presented within this thesis demonstrated that hypertension in isolation leads to modest alterations to the integrity of the cerebrovasculature and white matter, with no evidence of alterations to specific cognitive functions examined, demonstrating the importance of studying hypertension in isolation. Additionally, this study highlights the initial hypertensive induced alterations to the cerebrovasculature, such as endothelial signalling, vascular structure and inflammation, providing a window for therapeutic intervention at a time point when there are minimal alterations to the overall structural integrity of the brain. Future studies in this model should concentrate on examining different severities of hypertension and also hypertension concomitantly with other vascular risk factors to try and recapitulate pathological alterations found in humans.

While diffusion MRI promises an insight into white matter microstructure in vivo, the axonal pathways that connect different brain regions together can only partially be segmented using current methods. Here we present a novel method for estimating the tissue composition of each voxel in the brain from diffusion MRI data, thereby providing a foundation for computing the volume of different pathways in both health and disease. With the tissue dependent diffusion model described in this thesis, white matter is segmented by removing the ambiguity caused by the isotropic partial volumes: both grey matter and cerebrospinal fluid. Apart from the volume fractions of all three tissue types, we also obtain estimates of fibre orientations for tractography as well as diffusivity and anisotropy parameters which serve as proxy indices of pathway coherence. We assume Gaussian diffusion of water molecules for each tissue type. The resulting three-tensor model comprises one anisotropic (white matter) compartment modelled by a cylindrical tensor and two isotropic compartments (grey matter and cerebrospinal fluid). We model the measurement noise using a Rice distribution. Markov chain Monte Carlo sampling techniques are used to estimate posterior distributions over the model’s parameters. In particular, we employ a Metropolis Hastings sampler with a custom burn-in and proposal adaptation to ensure good mixing and efficient exploration of the high-probability region. This way we obtain not only point estimates of quantities of interest, but also a measure of their uncertainty (posterior variance). The model is evaluated on synthetic data and brain images: we observe that the volume maps produced with our method show plausible and well delineated structures for all three tissue types. Estimated white matter fibre orientations also agree with known anatomy and align well with those obtained using current methods. Importantly, we are able to disambiguate the volume and anisotropy information thus alleviating partial volume effects and providing measures superior to the currently ubiquitous fractional anisotropy. These improved measures are then applied to study brain differences in a cohort of healthy volunteers aged 25-65 years. Lastly, we explore the possibility of using prior knowledge of the spatial variability of our parameters in the brain to further improve the estimation by pooling information among neighbouring voxels.

Verbal memory deficits are among the most prominent cognitive sequelae in individuals with left temporal lobe epilepsy (LTLE). However, relationships between verbal memory function and white matter integrity (WMI) in the left temporal lobe remain unclear. Current study aims included determining fractional anisotropy (FA) and mean diffusivity (MD) differences as an index of WMI between participants with left temporal lobe epilepsy (LTLE), participants with right TLE (RTLE), and controls, establishing group differences based on verbal memory function between TLE groups, and describing relationships between WMI and verbal memory function within TLE groups. Probabilistic tractography defined the left fornix (FRX), left uncinate fasciculus (UF), left parahippocampal cingulum (PHC), and a control region, the left corticospinal tract (CST), in 26 LTLE, 29 RTLE, and 20 control participants. The LTLE group demonstrated significantly lower fractional anisotropy (FA) along the PHC compared with controls. LTLE and RTLE groups did not differ significantly on measures of verbal memory until analyses were restricted to participants with left-lateralized language functioning. PHC FA was negatively correlated with semantic memory function in LTLE, but positively associated with episodic memory functioning in RTLE. Overall, findings highlight the PHC as vulnerable in LTLE, and differentially related to verbal memory functioning based on TLE group. Both findings are likely secondary to left-lateralized white matter disruption in LTLE. The current study also highlighted the importance of identifying homogenous groups to more clearly identify brain-behavior relationships. Current findings further define left-lateralized white matter alternations and related verbal memory deficits in TLE. Implications for these findings are presented in context with previous TLE literature, and future directions for further study are discussed.

Inflammation and oxidative stress are increasingly recognised as important independent mediators of preterm brain injury and have been implicated in the pathogenesis of cerebral palsy and cognitive impairment. Such exposures are common for the premature infant in whom infection and inflammatory morbidities occur in around 60%. Furthermore, many preterm infants require oxygen therapy and respiratory support due to lung immaturity. Epidemiological and experimental studies indicate that in addition to the independent effects of inflammation and extreme hyperoxia on the developing brain, inflammation preconditions the developing brain resulting in variable injury when exposed to subsequent hypoxia-ischaemia. However experimental studies employing exposure to more modest oxygen fluctuations are lacking. This thesis characterises a clinically relevant model of prematurity where the developing brain is exposed to low grade inflammation and oxygen fluctuation around a hyperoxic mean. We hypothesise that antenatal inflammation and postnatal oxygen fluctuation, both alone and in combination, have detrimental effects on developing white matter. Pregnant dams received intraperitoneal lipopolysaccharide (LPS) or saline on G18 and G19. Dams and their pups were then reared in room air or fluctuating hyperoxia (circa 10kPa) for seven days. We measured longitudinal brain and body growth in different experimental groups to 12 weeks. Whole brains were examined for mRNA expression of inflammatory cytokines (TNFα, IL-1β, IL-6 and IL-10) and markers of oxidative injury (iNOS, SOD2). To determine the effect of perinatal insults on developing white matter, we analysed the expression of myelin basic protein (MBP) and glial fibrillary acidic protein (GFAP) in the internal and external capsule. We also examined white matter tracts for differences in microglia (CD68), oligodendrocyte progenitor cells (NG2), oligodendroglial cells (Olig2) and cell death (cleaved caspase3). Behavioural studies (Morris Watermaze Test, Elevated Plus Test and Open Field Test) were undertaken at 12 weeks of age to detect any long-term functional difference between the groups. Antenatal inflammation reduces both brain and body growth at P7. This normalises by P14 unless this inflammatory insult has been followed by postnatal oxygen fluctuation, where brain and body growth restriction persists until P14. We defined our inflammatory response at P1 following antenatal inflammation and did not observe elevation of mRNA at P1. We demonstrated increased SOD2 at this time point, indicating a reparative process. At P7 we observed a significant reduction in the oxidative response following combined exposure to antenatal inflammation and postnatal oxygen fluctuation, indicating a potential limit to, or suppression of, the reparative process. In terms of white matter injury, antenatal inflammation reduces myelination at P7. There is no synergistic effect of inflammation and oxygen fluctuation on MBP immunohistochemistry at P7. However, MBP mRNA expression is increased in pups exposed to both insults compared to those exposed to inflammation alone suggesting that the oxygen fluctuation may stimulate MBP production in response to oxidative injury. MBP mRNA levels and protein expression have all normalised by P14. We observed a reduction in total cell number in the external capsule and corpus callosum in the dual insult group, without an increase in caspase. In keeping with other studies we detected no effect of our perinatal insults on NG2+ve oligodendrocytes. Olig2+ve cell numbers were also consistent between experimental groups. In further characterisation of the cellular response, antenatal inflammation followed by postnatal oxygen fluctuation resulted in a decrease in GFAP mRNA at P7, an effect which was reversed and significantly increased by P14 suggesting delayed activation of the innate immune system. No difference was observed in microglial numbers between experimental groups. There was however, increased microglial cell death (CD68 + caspase) in the group exposed to antenatal inflammation. When this insult was combined with postnatal oxygen fluctuation there was a comparative decrease in microglial cell death, which may reflect an earlier peak of microglial cell death, due to an increased and sustained inflammatory stimulus. Morris Watermaze testing demonstrated that pups exposed to both insults took longer than controls to locate the hidden platform on day 1, which is a measure of spatial learning. The Elevated Plus Test and Open Field Test demonstrated that pups exposed to both insults were less anxious and took more risks than pups exposed to single insults. In conclusion, within a clinically relevant preterm model, antenatal inflammation transiently disrupts both brain and body growth and myelination of the motor tracts of the developing brain. Moreover, when combined with postnatal oxygen fluctuation, detrimental effects on growth are amplified and sustained. Decreased cell numbers are also observed within white matter tracts. In terms of long term functionality, these pups display disinhibition of behaviour as young adults. Collectively, this thesis demonstrates that synergistic actions of common low-grade perinatal insults may alter normal neurodevelopment, and that this may carry a risk of neurodevelopmental sequelae for preterm infants.

Cerebral ischemia and intracerebral hemorrhage (ICH) are both characterized by a prolonged inflammatory response and secondary injury phase, yet the spatial/temporal relationships between inflammation and white matter (WM) damage were largely unknown. Thus, I quantified the development of WM damage and inflammation over 7 days after ischemia, and 14 days after ICH. Following ischemia, myelin and axons were progressively damaged, and myelin damage coincided with neutrophil infiltration. Activated microglia/macrophages increased dramatically in the lesion core and edge, and selectively infiltrated damaged WM tracts while surrounding undamaged ones. To investigate the involvement of neutrophils in WM damage and inflammation after ICH, rats were rendered neutropenic before performing ICH. Neutrophil depletion reduced peri-hematomal axonal damage, BBB breakdown, and MMP-9 production at early times, and lessened microglia/macrophage and astrocyte responses at later times. Activated microglia/macrophages infiltrated peri-hematomal WM tracts, correlating with myelin fragmentation and axonal loss, and this was reduced with neutrophil depletion.

Cerebral ischemia and intracerebral hemorrhage (ICH) are both characterized by a prolonged inflammatory response and secondary injury phase, yet the spatial/temporal relationships between inflammation and white matter (WM) damage were largely unknown. Thus, I quantified the development of WM damage and inflammation over 7 days after ischemia, and 14 days after ICH. Following ischemia, myelin and axons were progressively damaged, and myelin damage coincided with neutrophil infiltration. Activated microglia/macrophages increased dramatically in the lesion core and edge, and selectively infiltrated damaged WM tracts while surrounding undamaged ones. To investigate the involvement of neutrophils in WM damage and inflammation after ICH, rats were rendered neutropenic before performing ICH. Neutrophil depletion reduced peri-hematomal axonal damage, BBB breakdown, and MMP-9 production at early times, and lessened microglia/macrophage and astrocyte responses at later times. Activated microglia/macrophages infiltrated peri-hematomal WM tracts, correlating with myelin fragmentation and axonal loss, and this was reduced with neutrophil depletion.