Deficiency of vitamin B6 due to mutations in key B6 metabolizing enzymes is suspected to contribute to several pathologies. Vitamin B6 in its active form, pyridoxal 5’-phosphate (PLP) is a cofactor for over 140 known B6 requiring (or PLP-dependent) enzymes, that serve vital roles in many biochemical reactions. There are three primary vitamin B6 forms, pyridoxine (PN), pyridoxamine (PM) and pyridoxal (PL) which are phosphorylated to pyridoxine 5’-phosphate (PNP), pyridoxamine 5’-phosphate (PMP) and PLP respectively. Pyridoxal kinase (PLK) and pyridoxine 5’-phosphate oxidase (PNPO) are the key enzymes involved in both salvage and de novo pathways of PLP biosynthesis. Mutations in these enzymes are one of the most important causes of PLP deficiency, apart from dietary insufficiency of vitamin B6 and drug inhibition of PLK and PNPO. One of our objectives is to understand the molecular basis of reduced catalytic activity of PNPO in case of the R95C homozygous missense natural mutant, which leads to the PLP deficiency and the debilitating disease, neonatal epilepsy encephalopathy. Using site-directed mutagenesis, circular dichroism, enzyme kinetics and fluorescence spectroscopy, we have shown that the reduced enzymatic activity exhibited by PNPO R95C mutant is due to reduced binding affinity of the oxidase cofactor, flavin mononucleotide (FMN), which is required by the enzyme for oxidizing the inactive B6 vitamers into the active PLP. High concentrations of B6 are linked to neurotoxic effects, which can be attributed to the highly reactive aldehyde group of PLP which reacts with many nucleophiles in the cell. This reactivity is most likely why the in vivo concentration of “free” PLP is about 1 μM, raising the intriguing question of how the cell supplies sufficient PLP to meet the requirements of the numerous B6 dependent enzymes. Our second objective is to determine how despite the low in vivo concentration of free PLP, enough of this co-factor is made available to activate PLP-dependent enzymes. We have used affinity pull down assays, fluorescence polarization and enzyme kinetics to show that PNPO forms specific interactions with B6 enzymes with dissociation constants less than 1 µM. We also show that transfer of PLP from PNPO possibly occurs by compartimentalization or channeling. Although, channeling is a controversial subject, it offers an efficient, exclusive, and protected means of delivery of the highly reactive PLP. High concentrations of B6 are linked to neurotoxic effects, which can be attributed to the highly reactive aldehyde group of PLP which reacts with many nucleophiles in the cell. This reactivity is most likely why the in vivo concentration of “free” PLP is about 1 ?M, raising the intriguing question of how the cell supplies sufficient PLP to meet the requirements of the numerous B6 dependent enzymes. Our second objective is to determine how despite the low in vivo concentration of free PLP, enough of this co-factor is made available to activate PLP-dependent enzymes. We have used affinity pull down assays, fluorescence polarization and enzyme kinetics to show that PNPO forms specific interactions with B6 enzymes with dissociation constants less than 1 µM. We also show that transfer of PLP from PNPO possibly occurs by compartimentalization or channeling. Although, channeling is a controversial subject, it offers an efficient, exclusive, and protected means of delivery of the highly reactive PLP.

Neonatal treatment of quinpirole in rats increases dopamine D2-like receptor sensitivity over the animal’s lifetime, a phenomenon referred to as D2 priming. Male and female Sprague-Dawley rats were given quinpirole (1mg/kg, i.p.) or saline on postnatal days (P)1-21. After habituation to a locomotor arena on P29-31, beginning P33, animals were administered nicotine (0.3 mg/kg, 0.5 mg/kg, or 0.7 mg/kg, i.p.) or saline and placed into a locomotor arena for behavioral testing every second day for a total of 9 treatments. The results showed that adolescents neonatally treated with quinpirole produced more enhanced sensitization to nicotine than controls. Brains tissues were analyzed for brain-derived neurotrophic factor (BDNF), a protein involved in neuron development and maintenance. The results showed that neonatal quinpirole treatment produced a significant increase in accumbal BDNF. Also, adolescent nicotine treatment produced a significant increase in BDNF in the nucleus accumbens and dorsal striatum. These findings help to broaden understanding of behavioral and chemical changes involved in schizophrenia and nicotine use and could have applications in aiding to alleviate this common comorbidity.

Nicotine use is very prevalent in the schizophrenic population, which is a 2.5 fold greater than the general population. In the present study, the drug quinpirole (dopamine D2/D3 agonist) or saline was given neonatally to 25 Sprague-Dawley male and female rats. Rats were randomly assigned to condition. Beginning postnatal day 33 animals were given twice daily administrations of nicotine (0.5 mg/kg free base). After the first of the daily injections they were placed in a locomotor arena every other day for behavioral testing. One day after behavioral testing, the dorsal striatum and nucleus accumbens were removed for brain-derived neurotrophic factor (BDNF) assay. BDNF is a neurotrophin that plays an important role in neuronal development, neuronal maintenance and plasticity, and synaptic activity. Results showed that nicotine produced locomotor sensitization but this was not enhanced by neonatal quinpirole, unlike past work. Regarding BDNF, there was a significant increase in the nucleus accumbens in rats treated with nicotine; neonatal quinpirole increased the BDNF response produced by nicotine. Nicotine produced an increase in dorsal striatum BDNF that was not affected by neonatal quinpirole treatment. Importantly, it appears that nicotine administrations, that occurred in two different contexts, may result in differential behavioral results relative to nicotine administrations given consistently in the same context.

Incremental improvements are continuously being made to P300-Speller BCI paradigms. Accurate classification depends on a high signal-to-noise ratio (SNR) between the target and nontarget items. Fixed presentation rates produce a large flash-evoked response that persists throughout the recording epoch, which can potentially undermine the classification of P300-responses. By introducing a random interstimulus interval (ISI) to a previously improved P300-Speller paradigm (i.e., Checkerboard Paradigm; CBP) we expect to reduce the deleterious flash-evoked responses and increase the P300 classification SNR. Data were recorded from 32 EEG locations (right mastoid referenced) from 13 subjects using the CBP with two conditions. In the Random ISI (RI) condition, ISI varied between 0 ms and 187.5 ms and averaged 93.75 ms. In the Fixed ISI (SI) condition, ISI remained static at 93.75 ms. In both conditions, participants were instructed to spell out 72 characters using an 8x9 matrix of alphanumeric characters by silently counting each target flash. The first 36 characters served as ‘calibration’ data for a stepwise linear discriminant analysis (SWLDA; 0 - 800 ms poststimulus epochs). This SWLDA classifier was then used to provide online feedback for an additional 36 character selections. Absolute amplitude of target and nontarget responses were summed across the recording epoch for each subject and averaged between Pz and Cz (maximum). Target averages were then divided by nontarget averages to create a SNR measure and compared between RI and FI conditions. The RI manipulation produced a significantly (p = .04) larger SNR (M = 5.85) than the FI condition (M =4.07).Further analysis of the averaged waveforms revealed a significantly (p = .05) greater positive peak at Cz (253 ms peak latency) for the RI condition. Classification performance measures for RI and FI conditions were high for accuracy (84 and 85%, respectively; NS) and bitrate (21 and 23 bits/min, respectively; NS). Together these results suggest that while randomizing ISI can yield higher SNR, response classification is not affected. It is possible that SWLDA is a useful classification method, in general; however, these data suggest that it does not capitalize on the additional information gained from the increase in SNR. Alternative classification techniques that can take advantage of specific subcomponents of the response may be able to utilize this additional information to improve BCI speed and accuracy.

No abstract.

The purpose of this study was to compare muscle activation in eight superficial lower limb muscles during execution of barbell back squats while in barefoot and athletic shoe conditions. It was hypothesized that greater muscle activity would be seen when squats were performed in barefoot conditions. Six participants were included in the study (means: 21.33±1.53 years, 170.45±11.33cm height, 69.85±12.46kg mass, 3.4±1.40 years training). Each met specific inclusion criteria. Participants came in three separate days for data collection (Day 1 – 1 repetition maximum [1RM] was determined, Day 2 – maximum voluntary contraction tests were held, Day 3 – squat tests performed with two footwear conditions). Squat tests were performed at 60, 70, and 80% of participants’ 1RM for each footwear condition and EMG data was recorded for these tests. Paired-sample T-tests were used to see if any differences were present between footwear conditions during eccentric and concentric phases of the squat, regardless of intensity. To test for differences between eccentric and concentric phases of the squat by intensity, 2x3 repeated measure ANOVAs were performed. Results showed some statistical difference between footwear conditions for two muscles in eccentric phase and no statistical significance for difference in concentric phase when compared without regard to intensity. When comparing footwear conditions at each intensity, main effects, as well as statistical significance, were found between footwear conditions in the eccentric phase. Main effects, but no statistical significance, were found in the concentric phase. The results indicate that EMG activity is greater for certain lower extremity muscles during the eccentric portion of a squat when under barefoot conditions.

The aim of this thesis is to examine the mechanisms of Methylphenidate (MPH) on Conditioned Place Preference (CPP), a behavioral test of reward. The psychostimulant MPH is therapeutically used in the treatment of ADHD, but has been implicated in many pharmacological actions related to drug addiction and is considered to have abuse potential. Past work in our lab and others have shown substantial sex-differences in the neuropharmacological profile of MPH. Here a discussion of the relevant mechanisms of action of MPH and its relationship to neurotrophins and CPP are reviewed. Furthermore, previous work is reviewed and a rationale for two experiments are presented. The study resulted in two experiments conducted earlier this year. Each experiment is presented, examining sex differences in CPP in two different methodological paradigms as well as the effects of MPH on levels of the dopamine transporter (DAT) in striatal and accumbal neurons.

This study examined nutritional status and fecundity of women and was approved for patient enrollment by the ETSU Institutional Review Board (IRB). Once enrolled, participants filled out two food frequency questionnaires (FFQ, fruit/vegetable intake, fat intake), self-reported anthropometric data, and had one 5 mL vacutainer of whole blood (EDTA) collected for measures of oxidative stress (OS), coenzyme Q10 level, and fatty acid profile of the red blood cell (RBC) membrane. Participants were either designated to the case group (13 women experiencing fertility problems and an anti-mullerian hormone level of 1 ng/ml or less (AMH, <1 ng/mL)) or the control group (13 women experiencing normal ovulation cycles and no evidence of polycystic disease). This study hypothesized that the case group would have poorer plasma antioxidant status, consume more dietary saturated fat and have lower levels of omega-3 and higher levels of omega-6 fatty acids in the RBC membrane. Results showed that case participants had significantly higher body mass index (BMI) when graduating from high school, BMI upon enrollment in the study, and trended to gain more weight since high school. Antioxidant status of the plasma tended to be higher for the case group. This result was surprising as case participants consumed significantly fewer servings of fruits and vegetables (sources of antioxidants) than control subjects. FFQ results indicated that case participants consumed significantly higher amounts of total fat (g), saturated fat (g), unsaturated fat (g), and their diets had a higher dietary fat percent. No significant differences were noted in the individual fatty acids isolated from the RBC membrane or in calculated ratios of fatty acid between case and control groups. The fatty acid profile from RBC membranes are in agreement with self-reported fat intake from FFQ data and may provide a rationale for no differences observed between the two groups.

Behavioral problems exhibited in early childhood can predict continued behavioral difficulties into adolescence and adulthood and can result in poor social functioning and health outcomes. Early identification of these concerns is necessary in order to connect families with appropriate interventions that thwart a negative trajectory. In pediatric offices, developmental screeners and pediatric surveillance are efficient and reliable methods of assessing behavioral concerns, which can help to quickly connect families with services. This study examines two methods of screening for behavioral concerns and the impact on on-site behavioral health referrals for children 4 and 5 years of age at a local pediatric clinic. In 2014, children were screened using the PEDS Developmental Questionnaire (PEDS) and referred to the on-site Behavioral Health Consultant (BHC). In 2015, children were screened using the Pediatric Symptom Checklist-17 (PSC-17) and referred to either the BHC or another new, on-site evidenced-based intervention called the Family Check Up (FCU). This study evaluates and compares the reach, effectiveness, and adoption of the two methods across both years. The results suggest that both screeners had the same rate of identification of behavioral concerns; however, the PSC-17 appears to improve rates of referrals to the BHC and the FCU.

No description available.