The brain renin-angiotensin system (RAS), of which angiotensin II (AngII) is the primary effector peptide, plays a critical role in the neurohumoral regulation of cardiovascular and body fluid homeostasis by modulating blood pressure, secretion of hypothalamic and pituitary hormones, and water intake. AngII produced locally in the brain or in the systemic circulation can act on brain regions called circumventricular organs (CVO), which lack the blood-brain-barrier. Dysregulation of central AngII signaling is implicated in the pathogenesis of hypertension; therefore, understanding the mechanisms of AngII in the CNS is an important area of investigation. Recently, a novel signaling mechanism for AngII in the periphery has been shown to involve NAD(P)H oxidase-derived reactive oxygen species (ROS). Although ROS are now known to be involved in numerous AngII-regulated processes in peripheral tissues, and are increasingly implicated in CNS neurodegenerative diseases, the role of ROS in central regulation of AngII-induced cardiovascular function remains unexplored. The hypothesis that ROS are critically involved in central AngII signaling and in AngII-dependent blood pressure and drinking behavior was tested by harnessing the power of an array of selective genetic tools, in combination with novel technologies for analysis of cardiovascular function in conscious mice. More specifically, central injections of adenoviruses encoding ROS-modulating molecules were used to examine the redox mechanisms in central AngII-mediated cardiovascular responses in vivo. Neuronal cell cultures were also used to investigate the involvement of NAD(P)H oxidase-derived ROS in AngII signaling, as well as to examine a link between calcium and ROS in intra-neuronal AngII signaling. Finally, in order to better understand the potential role of ROS in the brain in the pathogenesis of AngII-dependent hypertension, a mouse model that recapitulates the characteristics of human hypertension was employed in conjunction with genetic modulation of the redox state of the brain. These studies provide new evidence that ROS are involved in the intracellular signaling mechanism of AngII in the brain under normal and pathological conditions and offer new insight to how dysregulation of redox mechanisms in the brain may lead to the pathogenesis of AngII-dependent hypertension.

Corticotropin-releasing factor (CRF) is the primary hormone of the hypothalamo-pituitary adrenal axis (HPA-axis). In addition to its endocrine function, it has been proposed that CRF acts as a neurotransmitter. The widespread distribution of CRF immunoreactivity and CRF receptors in the rat central nervous system (CNS) supports this theory. Immunohistochemical studies have demonstrated high levels of CRF immunoreactivity the rat hypothalamus, a brain region involved in the regulation and integration of a variety of endocrine and autonomic homeostatic mechanisms. CRF has been shown to be involved in a number of these activities such as blood pressure control, food and water intake, behaviour and emotional integration. Many of these activities demonstrate progressive dysfunction as ageing proceeds. The aim of this thesis was to investigate the regulation of CRF in the rat CNS, particularly over the period of maturation and ageing. Tissue extraction and peptide radioimmunoassay (RIA) techniques were developed in order to measure regional CRF concentrations as a function of age in the rat CNS. Seven brain regions were examined including the hypothalamus, pituitary, medulla oblongata, pons, cerebral cortex, cerebellum and midbrain. Three age ranges were investigated: 3 – 4 weeks, 4 – 5 months and 14 – 18 months, representing young, mature and old age groups. Data for the tissues of individual rats from each age group were analysed using one-way analysis of variance (ANOVA) with post-hoc Scheffé tests (SPSS Release 6 for Windows, 1989 – 1993). CRF were detected in measurable quantities in all brain regions examined. Different age-related patterns of change were observed in each brain region. CRF concentrations (ng/g tissue) were highest in the pituitaries of young rats and were significantly reduced over the period of maturation (P< 0.05). However, the high CRF concentration of the young rat pituitary was likely to be a factor of the smaller tissue mass. Although the absolute CRF content (ng/tissue) of this tissue appeared to decline with maturation and ageing, the reduction was not significant (P>0.05). Therefore the pituitary of the young rat was relatively enriched with CRF per gram tissue. The highest CRF concentration in mature and aged rats was measured in the hypothalamus, in accordance with previous immunohistochemical studies. Hypothalamic CRF concentrations (ng/g tissue) demonstrated no significant alterations with maturation and ageing. The absolute CRF content (ng/tissue) of the hypothalamus was significantly less in the young rat compared to mature and aged animals, however this was accompanied by a smaller tissue mass (P<0.05). The CRF concentrations (ng/g tissue) of the rat cerebral cortex and medulla oblongata demonstrated significant reduction with advancing age (P<0.05), however in both cases this appeared to be due to significant increases in mean tissue mass. The absolute CRF content of these tissues (ng/tissue) were not significantly different over the period of maturation and ageing (P>0.05). CRF concentration (ng/g tissue) and absolute content (ng/tissue) of the pons demonstrated a trend to increase with advanced age in the rat, however this was not significant in both cases (P>0.05). Of interest were the significant increases observed in the CRF concentrations of the cerebellum and midbrain (ng/g tissue with advanced ageing (P<0.05). Significant increases were also observed in the mean tissue mass and absolute CRF content (ng/tissue) of these regions in aged rats (P<0.05). These findings perhaps indicate increased CRF synthesis and or decreased CRF turnover in these tissues with advancing age. The second stage of these studies examined age-related alterations in basal and potassium-stimulated hypothalamic CRF and overflow over the period of maturation and ageing in the rat, and required the preliminary development of an in vitro tissue superfusion system. The concomitant release of the co-modulatory compound, neuropeptide Y (NPY) was also measured. NPY has been shown to positively regulate CRF release and gene expression in the hypothalamus. In addition, NPY has been demonstrated to be involved in a number of hypothalamic activities, including blood pressure control and food intake regulation. Hypothalamic superfusion data were analysed using one factor repeated measures ANOVA (SPSS Release 6 for Windows, 1989-1993) followed by least significant difference tests ( Snedecor and Cochran, 1967) to enable both time and age comparisons. Basal hypothalamic CRF overflow was unaltered with maturation and ageing in the rat. Potassium stimulation (56 mM) elicted a significant 2 – 3 fold increase in hypothalamic CRF overflow across age groups (P<0.05). Stimulated hypothalamic CRF overflow was significantly greater in the young rat compared to the mature and aged animals (P<0.05). The enhanced response to depolarizing stimulus was observed at an age when the absolute CRF content of the hypothalamus was significantly less that of other age groups. It is possible that the enhanced responsiveness of the young rat may be of survival advantage in life threatening situations. Basal hypothalamic NPY overflow was much less than that of CRF, and potassium stimulation resulted in a very different age-related profile. The hypothalamic NPY response to depolarization was significantly reduced in the young rat and declined significantly with advanced ageing (P<0.05). The contrasting profiles of stimulated CRF and NPY overflow may indicate the activity of alternative regulatory factors present in the hypothalamus, whose activity may also be affected in an age-related manner. The final stage of these studies examined the nature of NPY modulation of hypothalamic CRF overflow in the mature rat. The facilitatory effect of NPY on hypothalamic CRF overflow was confirmed. The application of NPY (0.1 µM) significantly increased CRF overflow approximately 4 fold of basal (P<0.05). In addition, the role of the NPY-Y1 receptor was investigated by the prior application of Y1 receptor antagonists, GW1229 (0.05 µM). At this concentration GW1229 significantly reduced hypothalamic CRF overflow induced by perfusion with NPY (0.1 µm), P<0.05. It was concluded the Y1 receptor does have a role in the regulation of hypothalamic CRF overflow by NPY.

Thesis (Ph. D.)--University of Missouri-Columbia, 2007. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file (viewed on December 18, 2007) Vita. Includes bibliographical references.

Tumours of the central nervous system (CNS) are the second most common cancers diagnosed in children, yet the cause of this disease remains largely unknown. This thesis examines whether polymorphisms in folate-metabolising and glutathione S transferase (GST) genes influence the risk and disease outcome of childhood CNS tumours. 204 children aged ≤18 years diagnosed with a CNS tumour at the Sydney Children??s Hospital between 1989 and 2004 were included in the study. DNA samples were isolated from archival frozen and formalin-fixed paraffin-embedded tumour tissue. Polymorphisms in GST and folate pathway genes were examined using real-time PCR. Genotype distributions in children with CNS tumours were compared to those observed in a control panel of cord blood samples from 363 healthy newborns. Children carrying at least one variant allele for each of MTHFR 677 C>T, MTHFR 1298 A>C, MTR 2756 A>G, MTRR 66 A>G, and RFC 80 G>A were found to have a 2.8-fold greater risk of developing a CNS tumour than non-carriers (OR=2.80; 95%CI: 1.08-7.56, P=0.022), an association which was even more apparent in those children with an embryonal tumour (OR=4.54; 95%CI: 1.13-15.85, P=0.016). Results also showed that children with the GSTP1 105 Val/Val genotype were three times more likely to develop a CNS tumour of embryonal cell origin than children with the GSTP1 105 Ile/Ile or Ile/Val genotypes (OR=3.02; 95%CI: 1.34-6.46, P=0.005). No such association was observed for CNS tumours of glial cell origin. The GSTM1, GSTT1, and GSTP1 Ala114Val polymorphisms did not appear to be associated with the development of a childhood CNS tumour. In addition, children with the MTHFR 677 TT or RFC 80 AA genotypes were found to have a higher risk of death within 5 years of diagnosis compared to children with one or more MTHFR 677 C or RFC 80 G alleles, respectively (HR=5.52, 95%CI: 1.00-30.37, P=0.049 and HR=5.69, 95%CI: 1.38-23.51, P=0.016, respectively), after adjusting for other prognostic factors such as sex, age at diagnosis, period of diagnosis, and tumour grade. Conversely, children with the MTR 2756 AG or GG genotypes, or MTRR 66 AG or GG genotypes, were more likely to survive compared to those with the MTR 2756 AA or MTRR 66 AA genotypes, respectively (HR=0.21, 95%CI: 0.05-0.93, P=0.040 and HR=0.11, 95%CI: 0.02-0.53, P=0.006). Results presented in this thesis indicate that polymorphisms in folate-metabolising and GST genes may play a role in the aetiology and survival of childhood CNS tumours, and that this may vary depending on the histological sub-type of tumour.

<p>Persons suffering from Attention deficit/hyperactivity disorder (ADHD) struggle with complications within the functions that regulate and control the brain activities, due to deficiencies in these functions within the affected nerve-paths. ADHD is a cognitive function impairment characterised by inattention, impulsiveness and over activity. According to Diagnostic and Statistic Manual of Mental Disorders, American Psychiatric Association (DSM-IV), certain diagnostic criteria of ADHD must be fulfilled in order for a person to be diagnosed with ADHD. The everyday problems caused by ADHD are individual and medication can have positive effects relieving the person’s impairing behaviour. The study is based on scientific literature, three quantitative scientific articles and preview material from the last study by Johnson, Fransson, Kadesjö & Gillberg, presently being scrutinised. Swedish as well as English literature has been used. The purpose of this study is to shed some light upon whether medication facilitates the child’s school situation. The result deals with the ADHD diagnosis and pharmacological therapy involving drugs that stimulate the central nervous system, as well as naturopathic medicine like Omega-3/6. The pedagogical aspect for children with ADHD in school has been observed and evaluated. In this matter it is important for the pedagogue to encourage the child by letting it find out that it can manage more than it thinks.</p>

Supratherapeutic doses of anabolic androgenic steroids (AASs) are administered, not only as performance-enhancing drugs in the world of sports, but also in order to modify behaviour. AAS abusers are at risk of developing serious physical and psychological side effects such as dependence and aggressive behaviour. The aim of this thesis was to investigate the impact of supratherapeutic doses of nandrolone decanoate after subchronic administration on dopamine and serotonin pathways involved in drug dependence and aggression, in the male rat brain. Adult male Sprague-Dawley rats received intramuscular injections of nandrolone decanoate (3 or 15 mg/kg) or vehicle once daily for 14 days. Nandrolone decanoate pre-exposure abolished the effect of amphetamine on the 3,4-dihydroxyphenylacetic acid (DOPAC) tissue level in the hypothalamus and on the DOPAC/dopamine ratio in the hypothalamus and the hippocampus. A significant decrease of the basal extracellular DOPAC and homovanillic acid (HVA) levels could be detected in the nucleus accumbens, which remained low during the first hour following the amphetamine challenge. Nandrolone decanoate significantly reduced the activity of both monoamine oxidase A and B (MAO-A and -B) in the caudate putamen and amygdala. The gene transcript levels of MAO-B, and the dopamine D1 and D4 receptors were altered in limbic regions. No changes in transcriptional levels could be detected among the serotonin receptor genes examined. However, the density of the serotonin transporter protein was elevated in a range of aggression-related brain regions. Taken together, subchronic administration of nandrolone decanoate causes dopaminergic and serotonergic dysregulations in distinct brain regions. These areas of the brain are involved in the development of drug dependence and expression of impulsive and aggressive behaviours. These results may contribute to explain some of the behavioural changes often reported in AAS abusers, such as polydrug use and impaired impulse control.

Persons suffering from Attention deficit/hyperactivity disorder (ADHD) struggle with complications within the functions that regulate and control the brain activities, due to deficiencies in these functions within the affected nerve-paths. ADHD is a cognitive function impairment characterised by inattention, impulsiveness and over activity. According to Diagnostic and Statistic Manual of Mental Disorders, American Psychiatric Association (DSM-IV), certain diagnostic criteria of ADHD must be fulfilled in order for a person to be diagnosed with ADHD. The everyday problems caused by ADHD are individual and medication can have positive effects relieving the person’s impairing behaviour. The study is based on scientific literature, three quantitative scientific articles and preview material from the last study by Johnson, Fransson, Kadesjö &amp; Gillberg, presently being scrutinised. Swedish as well as English literature has been used. The purpose of this study is to shed some light upon whether medication facilitates the child’s school situation. The result deals with the ADHD diagnosis and pharmacological therapy involving drugs that stimulate the central nervous system, as well as naturopathic medicine like Omega-3/6. The pedagogical aspect for children with ADHD in school has been observed and evaluated. In this matter it is important for the pedagogue to encourage the child by letting it find out that it can manage more than it thinks.

The regenerative capacity of mature mammalian CNS neurons after axonal injury is severely limited by a variety of mechanisms. Retrograde repression is the continuous inhibition of the expression of growth phenotypes by tonic signals produced by target tissues and transmitted to the neuron cell body via retrograde axonal transport. Loss of target contact through axonal injury is thought to interrupt this retrograde signal and allow the up-regulation of growth-associated proteins. Most CNS neurons, however, possess many widespread axon collaterals, such that retrograde repression is maintained by intact sustaining collaterals even if some axons are injured.<p>In this project we investigated whether or not retrograde repression plays a role in limiting the expression of GAP-43 in transcallosal neurons. Because TCNs possess local axon collaterals to nearby cortex and project distal axons to homologous areas of contralateral cortex, we hypothesized that the simultaneous interruption of retrograde repressive signals from both ipsilateral and contralateral cortex would result in an up-regulation of GAP-43 expression in at least some TCNs.<p>We found that a bilateral infusion of a function blocking antibody to FGF-2 into the parietal cortex of rats using implanted osmotic mini-pumps resulted in a significant increase in the level of expression of GAP-43 mRNA in TCNs identified by retrograde fluorescent labeling. In contrast, neither ipsilateral or contralateral antibody infusions alone increased GAP-43 expression significantly compared to controls. The level of expression of GAP-43 in TCNs did not significantly increase after stereotactic callosotomy alone, but callosotomized animals treated with an ipsilateral infusion of anti-FGF-2 had levels of increased GAP-43 expression equivalent to those seen in animals that had received bilateral antibody infusions.<p>We conclude that FGF-2 provides a retrograde repressive signal for at least some mature mammalian TCNs, and that the expression of growth-associated proteins can be up-regulated in CNS neurons by simultaneously blocking retrograde repressive signals from all existing axon collaterals. The ability to alter the gene expression of mature CNS neurons in both normal and injured states through the targeted infusion of a pharmacological agent may have potential clinical implications in the future.

An acute demyelinating syndrome (ADS) in a child may be a monophasic illness or may represent the incident attack of multiple sclerosis (MS) – an inflammatory demyelinating neurodegenerative disorder affecting the brain, spinal cord and optic nerves. The central objective of this dissertation was to identify MRI parameters present at ADS that predict MS diagnosis. A scoring tool was first created containing 14 parameters identified from the literature and demonstrating substantial inter-rater agreement (Cohen’s kappa values ≥0.6). Children aged <16 years were enrolled at incident ADS and are currently followed for five years at 23 Canadian centers. Standardized MRI scans were acquired at onset and serially. MS was defined based on the occurrence of a second demyelinating attack or MRI evidence of new lesions in accordance with McDonald criteria for dissemination in time. Multivariable Cox proportional hazards regression models were used to identify MRI parameters that predicted MS diagnosis. Over 1100 MRI scans in 284 children with ADS were evaluated. To date, 57(20%) children have been diagnosed with MS. For those that developed MS, the median (IQR) time from incident attack to diagnosis was 6.2 (4.7-11.1) months. The presence of ≥1 T1-hypointense lesion (HR 20.6, 95% CI 5.5-78.0) and ≥1 T2 periventricular lesion (3.3, 1.3-8.8) were associated with an increased likelihood for MS diagnosis (sensitivity 84%, specificity 93%, PPV 76%, NPV 96%). The predictive parameters were validated in an independent Dutch cohort of 45 children with ADS (n=15, 33% MS): sensitivity 93%, specificity 87%, PPV 78%, NPV 96%. Finally, it was determined that the 2010 McDonald criteria are applicable for diagnosis of pediatric-onset MS diagnosis in older children with non-ADEM presentations. The work embodied herein emphasizes the value of MRI in predicting MS diagnosis in children with incident ADS. Early identification of children with MS is important for planning clinical care and will be valuable in future pediatric MS treatment trials.

The regenerative capacity of mature mammalian CNS neurons after axonal injury is severely limited by a variety of mechanisms. Retrograde repression is the continuous inhibition of the expression of growth phenotypes by tonic signals produced by target tissues and transmitted to the neuron cell body via retrograde axonal transport. Loss of target contact through axonal injury is thought to interrupt this retrograde signal and allow the up-regulation of growth-associated proteins. Most CNS neurons, however, possess many widespread axon collaterals, such that retrograde repression is maintained by intact sustaining collaterals even if some axons are injured.<p>In this project we investigated whether or not retrograde repression plays a role in limiting the expression of GAP-43 in transcallosal neurons. Because TCNs possess local axon collaterals to nearby cortex and project distal axons to homologous areas of contralateral cortex, we hypothesized that the simultaneous interruption of retrograde repressive signals from both ipsilateral and contralateral cortex would result in an up-regulation of GAP-43 expression in at least some TCNs.<p>We found that a bilateral infusion of a function blocking antibody to FGF-2 into the parietal cortex of rats using implanted osmotic mini-pumps resulted in a significant increase in the level of expression of GAP-43 mRNA in TCNs identified by retrograde fluorescent labeling. In contrast, neither ipsilateral or contralateral antibody infusions alone increased GAP-43 expression significantly compared to controls. The level of expression of GAP-43 in TCNs did not significantly increase after stereotactic callosotomy alone, but callosotomized animals treated with an ipsilateral infusion of anti-FGF-2 had levels of increased GAP-43 expression equivalent to those seen in animals that had received bilateral antibody infusions.<p>We conclude that FGF-2 provides a retrograde repressive signal for at least some mature mammalian TCNs, and that the expression of growth-associated proteins can be up-regulated in CNS neurons by simultaneously blocking retrograde repressive signals from all existing axon collaterals. The ability to alter the gene expression of mature CNS neurons in both normal and injured states through the targeted infusion of a pharmacological agent may have potential clinical implications in the future.