Cell-cell signalling molecules play important roles in neural development. In response to extracellular signals, neuronal progenitor cells proliferate, differentiate, and form a neuronal network. In the vertebrate retina, retinal ganglion cells (RGCs) are the first neurons produced during development and are the only neurons that send projections to the brain. However, the molecular mechanisms for RGC development have not been fully understood. In this study, I have investigated the expression and functions of Nel (Neural Epidermal Growth Factor Like), an extracellular glycoprotein that contains chordin-like domains and epidermal growth factor-like domains, in the development of the chick RGCs and retinotectal projection. I found that on embryonic days (E) 2-3.5, Nel was expressed in the presumptive retinal pigment epithelium of the developing eye. Correspondingly, Nel-binding activity (Nel receptor activity) was detected in the retinal pigment epithelium and also the progenitor layer of the neural retina. At the early stages during RGC formation, Nel overexpression increased the total number of RGCs and accelerated the progression of RGC differentiation wave. Conversely, Nel expression knockdown decreased the total number of RGCs and slowed down the progression of RGC differentiation wave. At later stages (E3-E18), expression of Nel in the retina was in the retinal pigment epithelium and the RGC layer, whereas receptor activity for Nel was localized in the retinal pigment epithelium and the RGC axons. In vivo, Nel overexpression in the developing retina induced the inhibition of RGC axons and thus disrupting the intraretinal RGC axon projection. These results suggest that Nel can positively regulate the production of RGCs at the early stages during retinal development, and at the later stages, Nel can function as an inhibitory guidance cue in vivo for RGC axons.

The autonomic nervous system serves to maintain a constant inner environment, a process termed homeostasis. Thus, in response to the homeostatic challenge posed by infectious agents, the autonomic nervous system answers to signals from the immune system and elicits adaptive physiological and behavioral reactions. These so called sickness responses include fever, anorexia, hyperalgesia, social avoidance, and the release of stress hormones. Neuropeptides, used in the communication between neurons, are because of their release properties and sustained actions likely mediators of homeostatic responses. The enkephalinergic system constitutes one of the largest neuropeptidergic systems in the brain, but its involvement in inflammatory conditions has been little studied. We first examined the immune-induced activation of the parabrachial nucleus (paper I), an enkephalinergic autonomic relay center in the brain stem. We found that intravenous injection of bacterial endotoxin, lipopolysaccharide (LPS), activated the external lateral parabrachial subnucleus, as measured in terms of Fos expression, but that the enkephalinergic cell population in this subnucleus was largely separated from the LPS-activated neurons. Because Fos may not always be a reliable activity marker, we next examined by in situ hybridization the immune-induced expression of newly transcribed preproenkephalin (ppENK) heteronuclear RNA (hnRNA), which gives a direct indication of the utilization of enkephalin in a particular neuron (paper II). We detected induced expression of ppENK hnRNA in several autonomic structures in the brain, including the paraventricular hypothalamic nucleus (PVH) but not the parabrachial nucleus, indicating increased enkephalinergic signaling activity in the positively labeled structures during inflammatory condition. We then examined the projections of the immune-induced ppENK transcribing PVH neurons by injecting rats intraperitoneally with the retrograde tracer substance Fluoro-Gold, hence labeling neurons with axonal projections outside the blood-brain barrier, followed by systemic injection of LPS (paper III). Dual-labeling histochemical and hybridization techniques showed that the vast majority of the ppENK hnRNA expressing cells were hypophysiotropic cells, hence being involved in neuroendocrine regulation. These findings suggest that centrally produced enkephalin is involved in the coordination of the sickness responses during systemic immune challenge, including the modulation of the release of stress hormones or other hypothalamic hormones during inflammatory conditions. We next turned to the role of prostaglandins in the hypothalamic-pituitary-adrenal (HPA) axis response to inflammation. We injected mice deficient for the terminal prostaglandin (PG) E2 synthesizing enzyme mPGES-1 with LPS and studied their stress hormone release (paper IV). The genetically modified mice displayed attenuated plasma levels of adrenocorticotropic hormone (ACTH) and corticosterone during the later phases of the HPA-axis response compared with wild type mice, and this impairment did not depend on a changed activation pattern in the brain, but instead correlated to an early decrease in corticotropin-releasing hormone mRNA expression in the PVH, hence being the likely cause of the blunted ACTH and corticosterone responses at later time-points. Based on these findings we suggest that a neural, mPGES-1-independent pathway, and a humoral, mPGES-1-dependent pathway act in concert but in distinct temporal patterns to initiate and maintain the HPA-axis response during immune challenge. In addition to activating the central limb of the HPA-axis, inflammatory mediators have been suggested to act directly on the adrenal gland to induce the release of corticosterone, but little is known about the underlying mechanisms. We examined adrenal tissue isolated from rats injected with LPS or interleukin-1β (IL-1β) (paper V), and found that immune stimulation resulted in dynamic changes in the adrenal immune cell population, implying a rapid depletion of dendritic cells in the inner cortical layer and the recruitment of immature cells to the outer layers. These changes were accompanied by an induced production of IL-1β and IL-1 receptor type 1, as well as of cyclo-oxygenase-2 and mPGES-1 in these cells, implying local cytokine-mediated PGE2 production in the adrenals, which also displayed EP1 and EP3 receptors in the cortex and medulla. Additional mechanistic studies using an IL-1 receptor antagonist showed that IL-1β acts locally to affect its own synthesis, as well as that of cyclooxygenase-2. Taken together these data demonstrate a mechanism by which systemic inflammatory agents activate an intrinsically regulated local signaling circuit that may influence the adrenals’ response to immune stress and may help explain the dissociation between plasma levels of ACTH and corticosteroids during chronic immune perturbations.

No description available.

This manuscript-based thesis explores the relationship between learning and development. The first manuscript reviews the important empirical regularities identified in human discrimination shift learning, including a qualitative age-related change in performance observed in childhood. Leading theoretical accounts of the empirical data are discussed, suggesting that none provides a comprehensive interpretation. The manuscript presents the novel, spontaneous overtraining interpretation. It hypothesizes that age-related changes in human shift learning stem from differences in amount of processing. Successful neural network simulations of the reversal and nonreversal shift tasks and of the optional shift task are reported as tests of the hypothesis. / The second manuscript reports simulations of additional discrimination shift tasks. These are the intradimensional and extradimensional shift tasks, in which novel stimuli are introduced in the relearning phase. Preschoolers and adults exhibit the same pattern of behavior in this variant of shift learning. Simulation results show that the spontaneous overtraining hypothesis captures this effect. / The third chapter reports an empirical validation of the shift learning model. If the shift learning performance of adults is a consequence of more extensive processing, it follows that adults in whom such processing is prevented should perform as preschoolers. Sixty adults took part in a shift learning experiment with a Brown-Peterson task as a cognitive load. Results mirror those observed with preschoolers. As a control, 40 adults performed the shift learning experiment without the cognitive load. These results replicate the typical adult performance. Overall, these experiments lend additional support to the model developed in Manuscript 1. / The final manuscript is a theoretical discussion of the relationship between learning and development. Two classes of neural networks are discussed, and their underlying assumptions about learning and development are highlighted. These are static architecture and generative architecture networks. It is argued that only generative algorithms, such as used in the shift learning simulations, qualify as developmental models. Both classes of networks are further contrasted with respect to innateness. The comparison suggests that only generative networks can acquire genuinely new representations. The manuscript proposes a novel formulation of Piaget's constructivism from the generative neural network perspective.

This thesis is concerned with the function and implementation of synchronization in networks of oscillators. Evidence for the existence of synchronization in cortex is reviewed and a suitable architecture for exhibiting synchronization is defined. A number of factors which affect the performance of synchronization in networks of laterally coupled oscillators are investigated. It is shown that altering the strength of the lateral connections between nodes and altering the connective scope of a network can be used to improve synchronization performance. It is also shown that complete connective scope is not required for global synchrony to occur. The effects of noise on synchronization performance are also investigated and it is shown that where an oscillator network is able to synchronize effectively, it will also be robust to a moderate level of noise in the lateral connections. Where a particular oscillator model shows poor synchronization performance, it is shown that noise in the lateral connections is capable of improving synchronization performance. A number of applications of synchronizing oscillator networks are investigated. The use of synchronized oscillations to encode global binding information is investigated and the relationship between the form of grouping obtained and connective scope is discussed. The potential for using learning in synchronizing oscillator networks is illustrated and an investigation is made into the possibility of maintaining multiple phases in a network of synchronizing oscillators. It is concluded from these investigations that it is difficult to maintain multiple phases in the network architecture used throughout this thesis and a modified architecture capable of producing the required behaviour is demonstrated.

Traumatic brain injury (TBI) is a major cause of death and disability throughout the world with few pharmacological treatments available for individuals who suffer from neurological morbidities associated with TBI. Cellular and molecular pathological processes initiated at the time of injury develop into neurological impairments, with chronic sleep disorders (insomnia, hypersomnolence) being among the somatic, cognitive and emotional neurological impairments. Immediately post-injury, TBI patients report excessive daytime sleepiness, however, discordant opinions suggest that individuals should not be allowed to sleep or should be frequently awoken following brain injury. To provide adequate medical care, it is imperative to understand the role of acute post-traumatic sleep on the recovery of neurological function after TBI. The aim of this thesis was to examine post-traumatic sleep after experimental TBI, defined as an increase in sleep during the first hours post-injury. In these studies, we non-invasively measured sleep activity following diffuse brain injury induced by midline fluid percussion injury to examine the architecture of post-traumatic sleep in mice. We detected significant injury-induced increases in acute sleep for six hours regardless of injury severity or time of day injury occurred. We found concurrent increases in cortical levels of the sleep promoting inflammatory cytokine interleukin 1-beta. We extended the timeline of post-injury sleep recording and found increases in post-traumatic sleep are distinctly acute with no changes in chronic sleep following diffuse TBI. Further, we investigated if post-traumatic sleep was beneficial to neurological outcome after brain-injury by disrupting post-traumatic sleep. Disruption of post-traumatic sleep did not worsen functional outcome (neuromotor, sensorimotor, cognition) at one week after diffuse TBI. With sufferers of TBI not always seeking medical attention, our final studies investigated over-the-counter analgesics and their effect on post-traumatic sleep and functional outcome. Acute administration of analgesics with varying anti-inflammatory properties had little effect on post-traumatic sleep and functional outcome. Overall, these studies demonstrated translational potential and suggest sleep after a concussion is part of the natural recovery from injury. While disrupting sleep does not worsen outcome, it is in no way beneficial to recovery. Additionally, a single analgesic dose for pain management following concussion plays little role in short term outcome.

Chronic intermittent ethanol exposure (CIE) is associated with degeneration of hippocampal neurons. The present study used hippocampal cultures to examine the loss of NeuN immunoreactivity, a relaible marker or neuronal density, after 1, 2, or 3 cycles of 5 days EtOH exposure (50 mM), followed by a 24-hour period of EWD or continuous EtOH exposure. NeuN immunoreactivity was decreased by 13%, 19%, and 16% in the CA1, CA3, and dentate gyrus after 3 cycles of CIE respectively; thionine staining confirmed significant cellular losses within each hippocampal subregion. Two cycles of CIE in aged tissue cultures resulted in significant decreases in NeuN immunoreactivity in all hippocampal subregions; however continuous ethanol exposure or exposure to one cycle of CIE did not. Further, exposure to the N-Methyl-D-aspartate receptor (NMDAR) antagonist 2-amino-7-phosphonvaleric acid (APV) (30 uM) during periods of EWD attenuated the loss of NeuN in all hippocampal subregions, while exposure to APV (40 uM) prevented the loss of NeuN in the CA1 and dentate gyrus. These results suggest that the loss of mature neurons after CIE is associated with the overactivation on the NMDAR.

Severe injury may result in disconnection between the peripheral and central nervous system. Regeneration of the central portion of sensory neurons into the spinal cord is notoriously poor in adult mammals, with low regenerative drive and an unpermissive central environment, most likely resulting in persistent loss of sensory function. A variety of strategies have been addressedto augment regeneration, including application of growth promoting factors, counteraction of inhibitory molecules, and provision of growth permissive substrates. Stem cells have been investigated in these contexts, as well as for the possibility of providing new neurons to act as a relay between the periphery and spinal cord. Here we have investigated different sources of neural stem cells for their ability to form neurons and glia after transplantation to the periphery; to project axons into the spinal cord; and to assist regeneration of surviving sensory neurons. These have been performed at two locations: the "dorsal root ganglion cavity", and the transitional zone following dorsal root avulsion. Neurons and glia were generated form mouse boundary cap neural crest stem cells and embryonic stem cell derived ventral spinal cord progenitors, and in addition to this, regeneration of sensory fibers was observed after transplantation of human fetal spinal cord derived progenitors and human embryonic stem cell derived ventral spinal cord progenitors. Further, delivery of neurotrophic factor mimetics via mesoporous silica nanoparticles proved a valuable tool for stem cell survival and differentiation. While technological advances make in vivo differentiation a realistic goal, our findings indicate that so far assisting regeneration of host sensory fibers to reconnect with the spinal cord by transplantation of stem cells is a more reliable strategy.

No description available.

During neuronal development, the axonal and dendritic projections are polarized and oriented along specific body axis. To further explore the molecular basis of neuritogenesis in vivo, we used the nematode Caenorhabditis elegans as a developmental model and performed a forward genetic screen to identify genes that specify the polarity of neurite outgrowth. We examined the VC4 and VC5 neurons, members of the six VC motor neurons using the Pcat-1::gfp transgene cyIs4. The VC motor neurons are ventrally located neurons that extend two processes. VC1, VC2, VC3, and VC6 extend axons along the anterior-posterior (A/P) axis; VC4 and VC5 extend axons around the vulva along a mediolateral left-right (L/R) axis perpendicular to the A/P axis. We identified and showed that vang-1/Van Gogh, a core component of planar cell polarity (PCP) signalling pathway, acts cell-autonomously in VC4 and VC5 neurons and non-autonomously from the epithelial cells to restrict neurite formation along the A/P axis. vang-1 mutant animals display ectopic neurites along the A/P axis. Using a candidate gene approach, we further identified and revealed two additional core members of PCP signalling, Prickle (PRKL-1) and Dishevelled (DSH-1), to play a role in A/P-directed neurite suppression. We also showed prkl-1 and dsh-1 genetically interact with vang-1 and VANG-1 is required to suppress A/P-directed neurite outgrowth from larval stage 4 to adulthood. Overexpression of VANG-1 results in a loss-of-function (lof) phenotype, suggesting that an appropriate level of VANG-1 activity is important. Additionally, vang-1/prkl-1, and dsh-1 may interact in parallel pathways. Our findings implicate PCP genes to play a previously unidentified role in maintaining polarized neuronal morphology by inhibiting neuronal outgrowth responses to environmental cues.