Caffeine is the most consumed psychoactive substance in the world, and most caffeine consumption in coffee and energy drinks is intended to produce a psychoactive effect. However, caffeine is not a primary reinforcer in preclinical paradigms – non-human species do not reliably take the drug to produce a psychoactive effect. However, caffeine is a ‘reinforcement enhancer’ in preclinical models; the effects of caffeine increase the motivation to obtain other non-drug reinforcers. The overall goal of this project was to determine if these reinforcement enhancing effects of caffeine could promote caffeine self-administration and to subsequently investigate the behavioral and neurochemical underpinnings of this effect. We hypothesized reliable caffeine self-administration would occur by adventitious pairing of caffeine with saccharin, a primary reinforcer. Second, we hypothesized that caffeine enhances reinforcement by increasing the salience of incentive stimuli, which are stimuli that come to evoke approach behaviors through associative learning (e.g., Pavlovian conditioning). Finally, incentive salience is moderated by dopamine release in the nucleus accumbens (NAc), an area highly involved in reward-learning and substance dependence. Therefore, we hypothesized that if caffeine enhanced control of approach behavior by incentives, then it would increase the ability of incentive stimuli to evoke dopamine in the NAc. These studies show that intravenous delivery of caffeine with oral saccharin increases operant relative to control groups responding for intravenous caffeine or oral saccharin. The effect was also dose-dependent, confirming that the psychoactive effects of caffeine increased behavior. We also extended this effect to an oral model of caffeine self-administration, which included a simple sweetener (saccharin) or a complex oral vehicle (saccharin with decaffeinated coffee) to mask the bitter taste of caffeine. Presenting caffeine with oral saccharin promoted self-administration, relative to saccharin alone and did not depend on the nature of the complexity of the vehicle. Caffeine also dose-dependently increased approach to an incentive stimulus and this effect was associated with increased extracellular dopamine in the NAc. These findings suggest caffeine enhances incentive motivation and that this effect may result from increases in CS-evoked striatal dopamine.

The hippocampus is a brain region that can undergo tremendous plasticity in adulthood. The hippocampus is related to the formation of spatial memories in birds and mammals. In birds, plasticity in the hippocampus occurs when formation of such memories is directly relevant to survival or reproduction, such as for breeding or food caching. In reptiles, the homologues to the hippocampus are the dorsal and medial cortices (DC and MC). In several lizard, snake and turtle species, these structures have been related to spatial memory. Experimental investigations indicate that differences in DC volume are related to space use associated with differing foraging ecologies. Differences in MC volume have been associated with territory size-based mate acquisition strategies. Furthermore, territory size has previously been correlated with plasma testosterone (T) levels. Therefore, I hypothesized that neuroplasticity within the MC/DC is controlled by demands on spatial navigation and seasonal differences and that these changes may involve the action of T. During two experimental trials, male Western Fence Lizards (Sceloporus occidentalis) were placed into either large or small semi-natural enclosures and allowed to interact with a female and intruder males over the span of seven weeks. One trial was performed during the spring breeding season and the other during the summer non breeding season, to examine seasonal differences in plasticity. Blood samples were collected at initial time of capture and before sacrifice to measure plasma T. Immunostaining for doublecortin was used to determine the density of immature neurons in each region, and cresyl violet staining allowed for volume measurements of specific regions. MC cell layer neurogenesis was higher in lizards placed in large enclosures than those in small enclosures and higher in the summer than in the spring. DC volume was smaller in lizards held in large enclosures than those in small enclosures. The decreased DC volume seen lizards held in large enclosures may indicate a cost to the increased neurogenesis in the MC of lizards in the same enclosures. These results indicate a possible trade-off between DC volume and MC neurogenesis that allows for switching between the ability to solve novel spatial tasks using the DC while storing a cognitive map in the MC. During the spring, T had no relationship with MC volume, while during the summer this was negative, so effects of T on the MC may be seasonal.

<p>Temporal order judgment (TOJ) refers to one’s ability to successively report the temporal order of two tactile stimuli delivered to independent skin sites. The brain regions involved in processing TOJ remain unclear. Research has shown that TOJ performance can be impaired with a conditioning background stimuli and this phenomenon, known as TOJ synchronization (TOJ-S), is suggested to be mediated by inhibitory neural mechanisms within the primary somatosensory cortex (SI) that create perceptual binding across the two skin sites. Continuous theta-burst stimulation (cTBS) over SI impairs tactile spatial and temporal acuity. This dissertation examines the effects of cTBS on TOJ and TOJ-S performance on the hand. In Experiment 1, TOJ and TOJ-S were measured from the right hand before and for up to 34 minutes following 50 Hz cTBS over SI. In Experiment 2, same measurements were obtained bilaterally for up to 42 minutes following 30 Hz cTBS over SI. Compared to pre-cTBS values, TOJ was impaired for up to 42 minutes on the right hand following 30 Hz cTBS. TOJ-S performance was improved for up to 18 minutes on the right hand following 50 Hz cTBS. These experiments reveal two major findings. First, cTBS act upon different inhibitory circuits that are suggested to mediate TOJ and TOJ-S. Second, cTBS parameters may dictate cTBS effects over SI excitability. The findings of this work not only emphasize the significant contributions of SI on tactile temporal perception, it provides novel insight of the underlying neural mechanisms of cTBS effects on SI cortical excitability.</p> / Master of Science in Kinesiology

<p>Immune-brain communication has important influences on stress circuitry and stress-related behaviours. Adaptive immune deficiency through loss of lymphocytes or an absence of gut bacteria has been linked to anxiety behaviours and stress responsiveness. In these models, there is a common deficit of T lymphocytes leading to the central hypothesis that T lymphocytes influence stress responsiveness and stress-related behaviours. This project considers the effects of T lymphocyte deficiency on anxiety and fear related behaviours as well as stress responsiveness in the hypothalamic pituitary adrenal (HPA) axis. Mice lacking T lymphocytes through knockout of the T cell receptor (TCR) β and δ chains, and B lymphocytes through knockout of the immunoglobulin M μ chain, were obtained and compared to C57BL/6 control mice. Activity, exploration, anxiety, fear and spatial learning tests were employed. Separately, gene expression was assessed for genes related to stress circuitry following chronic restraint stress. Additionally, lipopolysaccharide was used to determine the stress response to an innate immune challenge that was previously shown to elicit an exaggerated stress response in mice lacking Class I Major Histocompatibility Complex (MHC) and CD8+ T lymphocytes. It was found that mice lacking T lymphocytes, but not B lymphocytes, have reduced anxiety-like behaviour but an increased fear response. TCRβ-/-δ-/- mice also had altered expression of components of the HPA axis, serotonergic receptors and NMDA receptor subunits indicating an altered response to chronic stress. Finally, TCRβ-/-δ-/- mice do not display an exaggerated stress response to an innate immune challenge suggesting a central role for Class I MHC in the stress response that is not due to the CD8+ T lymphocyte deficiency that accompanies the functional loss of Class I MHC. These studies reflect an important role for T lymphocytes specifically in the development of the stress system and stress-related behaviours and enables a deeper understanding of neuroimmune influences on stress.</p> / Doctor of Philosophy (PhD)

<p>Microinjection is an established and reliable method to deliver transgenic constructs and other reagents to specific locations in the animal. Specifically, microinjection of a desired DNA construct into the distal gonad is the most widely used method to generate germ-line transformation of <em>C. elegans</em>. Although, current <em>C. elegans</em> microinjection method is an effective manner for creating transgenic worms, it requirements such as expensive multi DOF micromanipulator, detailed injection alignment procedure and skilled operator which makes the microinjection process slow and not suitable for scale to high throughput. Although many microfabricated microinjectors exist, none of them are capable of immobilizing a freely mobile animal such as <em>C.elegans</em> worm. In this research, a microfluidic microinjector was developed to simultaneously immobilize a freely mobile animal such as <em>C.elegans</em> and perform microinjection by using a simple and fast mechanism for needle actuation. The entire process of the microinjection takes ~30 seconds which includes 10s for worm loading and aligning, 5s needle penetration, 5s reagent injection and 5s worm unloading. The capability of the microinjector chip for creating transgenic <em>C. elegans</em> was illustrated (with success rate between 4% to 20%)</p> / Master of Science (MSc)

Injury causes increased pain sensation in humans and animals but the mechanisms underlying the emergence of persistent pathological pain states, which arise in the absence of on-going physical damage, are unclear. Therefore, elucidating the physiological regulation of such intractable pain is of exceptional biomedical importance. It is well known that endogenous activation of µ-opioid receptors (MORs) provides relief from acute pain but the consequences of prolonged endogenous opioidergic signaling have not been considered. Here we test the hypothesis that the intrinsic mechanisms of MOR signaling promote pathological sensitization of pain circuits in the spinal cord. We found that tissue inflammation produces agonist-independent MOR signaling in the dorsal horn of the spinal cord, which tonically represses hyperalgesia for months, even after complete recovery from injury and re-established normal pain thresholds. Disruption of this constitutive activity with MOR inverse agonists reinstated pain and precipitated cellular, somatic and aversive signs of physical withdrawal. This phenomenon required N-methyl-D-aspartate receptor activation of calcium-sensitive adenylyl cyclase type 1. Thus, we present a novel mechanism of long-lasting opioid analgesia that regulates the transition from acute to chronic pain while, in parallel, generates physical dependence. In conclusion we propose that the prevalence of chronic pain syndromes may result from a failure in constitutive signaling of spinal MORs and a loss of endogenous analgesic control.

Humans spend a third of their lives sleeping but very little is known about the physiological and genetic mechanisms controlling sleep. Increased data from sleep phenotyping studies in mouse and other species, genetic crosses, and gene expression databases can all help improve our understanding of the process. Here, we present analysis of our own sleep data from the large-scale phenotyping program at The Jackson Laboratory (JAX), to identify the best gene candidates and phenotype predictors for influencing sleep traits. The original knockout mouse project (KOMP) was a worldwide collaborative effort to produce embryonic stem (ES) cell lines with one of mouse’s 21,000 protein coding genes knocked out. The objective of KOMP2 is to phenotype as many as of these lines as feasible, with each mouse studied over a ten-week period (www.mousephenotype.org). The phenotyping for sleep behavior is done using our non-invasive Piezo system for mouse activity monitoring. Thus far, sleep behavior has been recorded in more than 6000 mice representing 343 knockout lines and nearly 2000 control mice. Control and KO mice have been compared using multivariate statistical approaches to identify genes that exhibit significant effects on sleep variables from Piezo data. Using these statistical approaches, significant genes affecting sleep have been identified. Genes affecting sleep in a specific sex and that specifically affect sleep during daytime and/or night have also been identified and reported. The KOMP2 consists of a broad-based phenotyping pipeline that consists of collection of physiological and biochemical parameters through a variety of assays. Mice enter the pipeline at 4 weeks of age and leave at 18 weeks. Currently, the IMPC (International Mouse Phenotyping Consortium) database consists of more than 33 million observations. Our final dataset prepared by extracting biological sample data for whom sleep recordings are available consists of nearly 1.5 million observations from multitude of phenotyping assays. Through big data analytics and sophisticated machine learning approaches, we have been able to identify predictor phenotypes that affect sleep in mice. The phenotypes thus identified can play a key role in developing our understanding of mechanism of sleep regulation.

Feeding behavior and its associated neural circuitry is complex and intricate in mammalian systems, however, a simple model organism, such as C. elegans provides a more basic approach to understand factors and molecules involved. The fruit-dwelling nematode provides a unique set of resources; it only consists of 959 cells, 302 of which are neurons. In addition, each neuron’s connectivity and position within the worm is known and consistent between animals. Conservation of neurotransmitters and biochemical processes add to this impressive list. These resources provide an excellent background to address feeding behavior and the neural structures governing it. Feeding behavior in worms mimics feeding behavior in more complex organisms. They decide when to eat based on recent feeding behavior, current nutritional status, availability of food, and familiarity with the food available. Following starvation and refeeding worms enter a behavioral state similar to post-prandial sleep. The worms will stop eating and stop moving, in a state referred to as satiety quiescence. The ability to enter this state and maintain it is dependent on a pair of neurons in the head of C. elegans called ASI. Using calcium imaging and an automated satiety quiescence assay, our lab has found that this neuron pair is important for entering satiety quiescence and senses food. Feeding behavior, such as satiety quiescence, is regulated by numerous factors internal and external to the worm. Another pair of head neurons, ASH are capable of suppressing ASI’s activity in the presence of noxious stimuli and the presence of nutrients (potentially acting via ASI) can suppress ASH’s activation to noxious stimuli under starvation conditions. The interaction between these two neuron pairs can be regulated by other signals from the rest of the worm. We identified an opioid signal that can modulate the response of ASI to noxious stimulus signaling from ASH under starvation conditions. Other signals were identified to influence satiety behavior and this circuit including serotonin, octopamine, glutamate, and adenosine. In addition to these signals, a group of transcription factors were identified that may play a role in conveying the status of fat storage within the worm to its nervous system. Nuclear hormone receptors were found to increase their expression during starvation then decrease their expression upon refeeding. Upon completion of this work, we have a reached a greater understanding of the internal and external conditions governing feeding and avoidance behaviors.

No description available.

Using matched pairs of schools having a diagnosis of Sick Building Syndrome (SBS) and those not identified, and not having the potential of being identified as having the diagnosis, this study examined the relationship between factors assumed or determined to have an impact on a school's ability to carry out its mission as they related to the SBS diagnosis. The factors studied were student average daily attendance (ADA); students' mean standardized test scores in math and reading on the Iowa Test of Basic Skills and the Tests of Achievement and Proficiency; the need for students to take prophylactic medications; the rate of staff turnover; the percentages of short term suspensions (less than 10 days), long term suspensions (more than 10 days), and the percentage of recommendations for expulsion; as well as the impact of the public identification of a school as having a diagnosis of SBS on student ADA, mean standardized test scores in math and reading and staff turnover. A factorial ANOVA model (2 by 4) for repeated measures revealed no statistical significance for main or interaction effects on any of the six hypotheses addressed.