Nutrient restriction is a relatively common production insult to pregnancy in cattle. Recently, melatonin supplementation has been investigated as a possible therapeutic to rescue the negative effects of nutrient restriction. Neurotransmitters have been implicated as having negative programming effects in mouse and human pregnancies, manifested as metabolic and neurologic disorders. The role of neurotransmitters in fetal development has only begun to be understood in mice and humans. Neurotransmitters have not been analyzed in cattle, much less within the context of a compromised pregnancy. Brangus heifers were allotted to one of four treatments (ADQ-CON, RES-CON, ADQ-MEL, RES-MEL) in either Fall 2019 or Summer 2020. Cesarian sections, at day 240 of gestation, allowed for fetal and placental tissues to be collected for neurotransmitter and gene expression analysis. Alterations to neurotransmitter pathways were observed in a seasonally dependent manner. Future investigation is needed into the implications of altered neurotransmitters on post-natal life.

Cannabidiol (CBD), a non-psychotropic phytocannabinoid with structural similarity to Δ 9 -tetrahydrocannabinol (THC), is currently being investigated as a therapeutic for its immunosuppressive effects. One disease for which CBD is extensively researched is multiple sclerosis (MS), a demyelinating, autoimmune disorder, and its murine model counterpart, experimental autoimmune encephalomyelitis (EAE). The focus of this dissertation aimed to analyze the transcriptomic brain pathways in EAE and its comparison to MS in addition to CBD’s immunosuppressive mechanisms in the innate and adaptive immune systems. Evidence presented here showed that transcriptomic signaling pathways in the EAE brain of mice with clinical symptoms were similar to the transcriptome of active lesions from MS patients. The transcriptomic analysis also presented two differentially expressed genes that were increased in CBD-treated, asymptomatic EAE mouse brains: oxytocin and vasopressin. Expression of these genes was also increased in naïve, CBD-treated mouse brains, which may indicate potential as efficacy biomarkers. Subsequently, as disease progression requires input from the innate and adaptive immune systems, the mechanisms of CBD were analyzed under naïve and stimulatory conditions in macrophages and splenocytes. In macrophages, CBD exerted an anti-inflammatory effect by dampening the M1 polarization phenotype, decreasing pro-inflammatory cytokines and chemokines, reducing TNF-α through intracellular TACE retention, and diminishing the translocation of RelA to the nucleus. Notably, similar impact of CBD on TACE was evident in naïve macrophages, suggesting that CBD exerted an effect under naïve conditions. In splenocytes, CBD exhibited a long-term effect on the percentage of various immune populations during naïve and splenic T cell activation (with anti-CD3/anti-CD28) conditions but only provided temporary relief and short-term from TNF-α and IFN-γ cytokine secretion. CBD also increased early mRNA expression of Tnfa in CBD in stimulated splenocytes. In naïve splenocytes, CBD impacted key immune mediators discovered from a transcriptomic re-analysis of human neuroblastoma cells, including decreased early expression of Noxo1 but increased expression of Ctsb. In summary, this dissertation presented evidence that CBD impacts the immune system from the transcriptional level in the brain, the innate and adaptive immune systems at the cellular level, and the overall EAE disease phenotype.

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No description available.

Developing robust behavioral assays to study olfactory-driven behaviors allows for greater insight into the neural mechanisms behind them. Oftentimes, olfactory behavioral assays require a two-choice design, consistent variables, and controlled stimulus application. This can be challenging when working with marine dwelling organisms such as nudibranchs. Extensive work shows the importance of olfaction in both pre-metamorphic larval development and adult stage nudibranchs (Gastropoda, Mollusca). However, there is little research investigating how olfaction plays a role in rapidly developing, post-metamorphic juvenile nudibranchs such as Berghia stephanieae. To study olfactory-associated behaviors in juvenile Berghia, a novel behavioral microfluidic chip was designed which met the requirements for a reliable olfaction assay. Baseline motor behaviors such as general locomotion, turns, and contractions were observed. Despite individuals being raised in the same cohort, animal-to-animal variability was found in relation to their baseline behaviors. Xylene cyanol, a commonly used dye in microfluidics, was found to be aversive at a range of concentrations (0.075% - 0.025%) causing the animal to tightly contract and turn away from the stimulus. Juveniles showed no significant behavioral response to a 0.00125% dilution which was necessary to confirm a constant flow in the microfluidic chip. Juveniles showed a strong preference towards fluid that had been conditioned with their preferred food source odor, the sea anemone Exaiptasia diaphana. This was judged to be an innate preference because it occurred upon the first exposure of these animals to the odor. Serial dilution of Exaiptasia conditioned seawater (ECS) uncovered a threshold for behavioral preference of 60%. In summary, the development of this behavioral provides an opportunity to present controlled olfactory stimuli while observing the juvenile’s behaviors. This will allow future experiments to examine the neural mechanisms behind both aversive and attractive stimuli and provides a method for testing olfactory learning and memory in this species.

University evaluations often reflect an individual’s ability to memorize and recall lecture material during exams. Consequently, the ability to effectively encode, store, and later retrieve information is an integral part of learning and academic success. Notably, students who are more physically active tend to have better academic performance. The neurobiology of stress is a strong candidate for the mechanism underlying this exercise-cognition interaction. Given that exercise is a physical stressor, it is hypothesized that exercise-induced adrenocortical activations increase cortisol levels. Critically, cortisol increases memory consolidation for newly learned information. One hundred twenty-eight young adults (36 males; age: M±SD =19.47±1.55 years) viewed a video lecture before exercise (n = 41), after exercise (n = 42), or after rest (n = 45). The exercise was high intensity interval training on a cycle ergometer and memory for the lecture material was assessed using a multiple-choice quiz conducted 14 minutes and 48 hours after the lecture. There was a significant positive correlation between aerobic fitness and grade point average [r(95) = 0.22, p < .05], immediate recall [r(100) = 0.39, p < .001], and delayed recall [r(98) = 0.28, p < .01]. A mixed model ANOVA found a significant main effect of group on comprehension of the lecture material, F(2, 96) = 3.34, p < .05, revealing greater memory benefits at both 14 minutes and 48 hour delays for those who exercised compared to those who did not exercise; however, pairwise comparisons found this effect specific to the exercise post group. There was also a main effect of group on cortisol levels, F(2, 107) = 3.97, p < .05; however, only the exercise prior group exhibited significantly greater levels than the control group. Thus cortisol levels collected during the experimental session did not clearly differentiate the exercise conditions or reflect the observed memory benefits for the exercise post group. This may have resulted from the gradual increase in cortisol following exercise that had time to increase when exercise was completed at the beginning of the exercise session (exercise prior) rather than at the end (exercise post). Overall, this study suggests that both physical fitness and an acute bout of aerobic exercise are associated with academic and memory performance. More research is needed to understand the mechanism. / Thesis / Master of Science (MSc)

Protein disulfide isomerase (PDI) is an endoplasmic reticulum (ER)-resident chaperone with oxidoreductase and isomerase activity. Unique to its normal function, PDI also appears to disassemble the A1 subunits of cholera toxin (CT) and heat-labile enterotoxin (LT). It does so using an unfolding mechanism that knocks the catalytic A1 subunit away from the rest of the holotoxin. Release of the A1 subunit is linked to the diarrheal diseases caused by V. cholerae and enterotoxicogenic E. coli (ETEC). Due to the previously established difference in disease potency between CT and LT, we investigated and established a distinction between the two toxins in their efficacy of disassembly by PDI. We further identified four amino acid differences between the CTA2 and LTA2 linkers, which connect the A1 and cell-binding B subunits of both toxins, as the basis for this difference. We believe these four amino acids result in changes to holotoxin architecture that lead to antiparallel binding of PDI to LT as opposed to CT, which translates to a loss of momentum for the physical disassembly of LT. We have shown this through algorithmic simulations of the binding event between PDI and either CT or LT. We hypothesized the unfolding mechanism of PDI, which dislodges the A1 subunit of both CT and LT, can also break down neurotoxic aggregates of β-Amyloid (AB) and α-Synuclein (AS). PDI is known to inhibit the aggregation of the amyloid proteins. We demonstrated here that PDI could also reverse oligomeric and post-oligomeric aggregates of AB and AS, respectively. Our work sheds light on the specifics of PDI's novel physical mechanism as well as introduce it as a possible therapeutic for both Alzheimer's and Parkinson's disease due to its unique ability to disaggregate early fibrillar structures of AS and AB proteins.

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As the most widely consumed drug around the globe, there is a vast array of contradicting research available on caffeine. One of the most debated and researched topics on caffeine is its effect on the brain. Meanwhile, the data on the neurological condition of migraine has information scattered throughout countless research articles and experiments. Although neither migraine or caffeine are completely understood by the medical world, this analysis attempts to give a more coherent understanding of the relationship between the two. This is done by first understanding the known and theorized mechanisms of caffeine as well as the pathologies of migraine. Discussions on channelopathies, current migraine medications, and case studies will be presented. After much background research, we hypothesized that caffeine could excite neurons at physiological concentrations to the point of activation. This was tested by targeting the transcription factor cFos using immunocytochemistry in vitro. The protein cFos was identified due to its rapid translation—just 15 minutes after stimuli—to indicate activation. In addition to a control culture, three different caffeine concentrations were tested on the neurons: 50 micromoles— average plasma level after 1-2 cups of coffee consumption, 100 micromoles—average plasma level after 5-6 cups of coffee also believed to be the therapeutic amount to defend against neurological diseases such as Alzheimers Disease, and 250 micromoles—the average plasma level considered to be toxic in humans. Indeed, we saw a 53.8% increase in cFos expression in the neurons as 100 micromolar of caffeine was added and exposed to the cell cultures for 24 hours. In order to ensure the results obtained in this study were physiologically relevant in vivo, known toxic levels were tested for in vitro neurotoxicity. It was found in vitro that at the non toxic plasma concentrations of 50 micromolar and 100 micromolar of caffeine did not display cellular death as tested by Trypan Blue viability testing, Crystal Violet morphologies, and fleurojade immunochemistry that tests for degeneration. Each of these experiments identified a significant death increase as the toxic level of 250 micromoles of caffeine were utilized. This allowed us to theorize that the activation of neurons found in these experiments due to caffeine exposure would apply the same effect in vivo.

The purpose of this study was to use retrograde tracing techniques to examine hepatic neuroanatomy in the rat model, with special emphasis upon the identification of previously undiscovered intrahepatic parasympathetic ganglia. Retrograde analysis was performed using Fluoro-Gold (FG) tracer injections of both male and female Sprague-Dawley rats. To accurately examine the neural connectivity of both the vasculature and the parenchyma, the FG-labelled livers were divided into two groups. In the first, vessel trees were extracted via dissection and whole-mounted for bright field and confocal visualization. Left lateral lobes taken from the male and female liver that constituted the second group were sectioned, and slices from various layers of tissue were fixed to slides and visualized. The results indicated the presence of several large, fluorescent structures bearing a strong resemblance to parasympathetic ganglia. However, the images were not detailed enough to properly differentiate true ganglia from similar paraganglia. Regardless, the importance of this experiment lies in its attempt to revisit an understudied field in neuroscience, and the findings of this study could potentially provide a starting point for further inquiry.