The main goal of this research is to provide a novel bioanalytical approach to better understand α-synuclein (AS) aggregation linked to Parkinson’s disease (PD) and characterize the implications of contributing factors such as the presence of metal ions and potential therapeutics that would inhibit or reverse AS fibrillation. Current bioanalytical techniques have reported the fibrillation process of AS however, the detection of prefibrillar formation or the nucleation phase of AS has yet to be characterized. This research aimed to address this issue and monitor the primary stages of AS fibrillation from natively soluble monomer to fibrillar aggregates. The electrochemical measurement of these processes utilized the intrinsic electroactivity of 4 tyrosine (Tyr) residues in AS observed at ~0.6 V (vs. Ag/AgCl) to monitor its early fibrillation kinetics. The research presented here provided valuable evidence of the conformational changes attributed to prefibrillar forms of AS.

The main goal of this research is to provide a novel bioanalytical approach to better understand α-synuclein (AS) aggregation linked to Parkinson’s disease (PD) and characterize the implications of contributing factors such as the presence of metal ions and potential therapeutics that would inhibit or reverse AS fibrillation. Current bioanalytical techniques have reported the fibrillation process of AS however, the detection of prefibrillar formation or the nucleation phase of AS has yet to be characterized. This research aimed to address this issue and monitor the primary stages of AS fibrillation from natively soluble monomer to fibrillar aggregates. The electrochemical measurement of these processes utilized the intrinsic electroactivity of 4 tyrosine (Tyr) residues in AS observed at ~0.6 V (vs. Ag/AgCl) to monitor its early fibrillation kinetics. The research presented here provided valuable evidence of the conformational changes attributed to prefibrillar forms of AS.

The purpose of this study was to investigate whether locomotor training, in the form of treadmill training, could ameliorate neurochemical changes and behavioural deficits in the 6-hydroxydopamine (6-OHDA) rat model of Parkinsons disease. It has been recently demonstrated that rehabilitative forelimb motor training can attenuate dopamine loss and some deficits in forelimb movements in this animal model. In addition, brief locomotor treadmill training has been shown to attenuate forelimb deficits in 6-OHDA treated rats. However, it is not known whether locomotor training could result in an amelioration of locomotor deficits in these animals. Rats were lesioned with 6-OHDA injected intracerebrally. Lesioned rats were randomly assigned to one of 3 groups: early treadmill trained, late treadmill trained and untrained. Animals in the treadmill groups were trained to trot on a moving treadmill for 2 x 20 minute sessions daily for 30 days, beginning either 24 hours or 7 days after 6-OHDA injection. Untrained animals were exposed to a stationary treadmill for the same time periods. All animals were assessed on their abilities to perform several behavioural tasks designed to test locomotor and forelimb movement abilities prior to 6-OHDA injection and again at 3 and 6 weeks post-injection. These tests included measurement of ground reaction forces during overground locomotion, paw placements during a ladder crossing task, forelimb useage during exploratory behaviour and ability to initiate forelimb stepping movements. In addition, assessments of dopamine depletion in the striatum were carried out first in vivo, by measuring apomorphine-induced rotations at 2 weeks post 6-OHDA injection, and subsequently by post-mortem analysis of dopamine levels in the striatum using HPLC at the conclusion of the study. Treadmill training resulted in attenuation of dopamine depletion compared to non-treadmill trained animals, as measured by both apomorphine injection and HPLC. However, treadmill training produced no difference in behavioural deficits on a variety of tests compared to untrained animals. In some cases, early treadmill trained animals tended to display more severe behavioural deficits compared to untrained animals. Late treadmill training had a similar but smaller effect compared to early treadmill training. We conclude that treadmill training does not ameliorate locomotor deficits, in the 6-OHDA model of Parkinsons disease, even though this same training results in attenuation of dopamine loss in the striatum.

Dopamine is a neurotransmitter found in central nerve system which has a vital role for human health. Dopamine oxidation in body is an important issue since it may form reactive metabolites which can be toxic to the cell. Surface-enhanced Raman scattering (SERS) is currently recognized as one of the most sensitive spectroscopic tools, which can be exploited for ultrasensitive chemical and biological detection, addition to providing structural information on the systems of interest. SERS of dopamine displays three strong bands at 1269, 1331 and 1479 cm-1. These bands are the signature of dopamine molecule. The most intense band at 1479 cm-1 is contributed mainly from stretching of the carbon-carbon bond to which the oxygens are attached. A bidentate silver-dopamine complex or in general bidentate metal-dopamine complex formation is required for the SERS detection of dopamine and other catecholamines. In other words, for acquiring the characteristic dopamine SERS signature, both of the catechol oxygens should take a part in the adsorption of dopamine to the silver metal surface which is used as a SERS substrate. Therefore, the reactivity of different oxidation forms of dopamine for the formation of bidentate silver-dopamine complex was investigated by obtaining their SERS spectra and following the characteristic C-C ring vibration at 1479 cm-1. Dopamine oxidation was carried out electrochemically, utilizing platinum and silver electrodes as working electrode. Oxidation products formed were identified with UV-vis Spectrometer. Also, silver metal ions were used to oxidize dopamine, leading to formation silver nanoparticles. Dopamine functionalized silver nanoparticles were characterized by Scanning Electron Microscope, Transmission Electron Microscope, UV-vis Spectrometer. Surface- enhanced Raman spectra of polydopamine on the surface of synthesized silver nanoparticles and the electrodeposited dopamine on the porous surface of silver electrode were also obtained.

Schizophrenia (SCZ) is a severe neuropsychiatric disorder with a genetic component. Most candidate gene association studies have given mixed results. We investigated the GABAA receptor gamma2 subunit gene GABRG2, the dopamine receptor gene DRD3, and the Brain-derived neurotrophic factor gene BDNF that is required for D3 expression by genotyping polymorphisms spanning and surrounding these genes for association with SCZ, as well as suicidal behaviour. We also examined the BDNF, DRD3, as well as the dopamine receptor gene DRD2 and Protein Kinase B gene AKT1 for association with Tardive Dyskinesia (TD), a potentially irreversible motor side effect of long-term antipsychotic medication. Our analysis included single-marker tests, haplotype tests, and gene-gene interactions. We found a haplotype in the 5’ region of GABRG2 to be associated with SCZ in both families and matched case-control samples. We also found two synonymous DRD2 polymorphisms, rs6275 (C939T) and rs6277 (C957T), and their haplotypes, as well as a polymorphism 5’ of DRD3, rs905568, to be associated with TD. Further, we reviewed two putative functional DRD2 polymorphisms, -141C Ins/Del and TaqIA, in TD and found TaqIA 3’ of the gene to be associated with TD in a meta-analysis. Lastly, we found a significant interaction between AKT1 rs3730358 and DRD2 C939T in TD. Though replication studies are required, these results contribute to the future development of genetic tests to assess for the risks of SCZ and TD, leading to better outcome for patients suffering from these debilitating conditions.

In addition to acting in synaptic transmission, neurotransmitters have been shown to play roles in the development of nervous system. Developing neurons extend neurites to connect to their target cells, and growth cones at the tip of growing neurites are critical for pathfinding. Although evidence for the regulation of axonal growth and growth cone guidance by neurotransmitters and neuromodulators is emerging, less is known about the mechanisms by which neurotransmitters affect developing neurons. Here, I focus on three neurotransmitters/ neuromodulators and describe their actions (a) at the level of growth cone, especially on filopodia, which serve as sensors that allow growth cones to probe the environment they are traversing, and (b) on how neurotransmitters modulate neuronal electrical properties, which, in itself, have been shown to affect neurite extension. The goals of this dissertation are to investigate 1) the cholinergic modulation of neuronal activity and its effects on growth cone motility; 2) the excitatory modulation of neuronal excitability by nitric oxide (NO); and 3) the inhibitory modulation of neuronal activity by dopamine (DA). The work uses a well-established model system to investigate growth cone motility and neuronal activity: identified neurons from the pond snail Helisoma trivolvis studied in cell culture or in the intact ganglion in situ. The study of B5 neurons demonstrates that acetylcholine (ACh) induces filopodial elongation, which is mediated by opening of nicotinic ACh receptors, membrane depolarization, and elevation of intracellular Ca level in growth cones. This dissertation also shows that NO inhibits two types of Ca-activated K channels to depolarize the membrane potential of B19 neurons. Additionally, the study reveals that DA serves as an inhibitory neurotransmitter to hyperpolarize and silence the electrical activity of firing B5 neurons via a D2-like receptor/PLC/K channel pathway. Taken together, this dissertation elucidates novel cellular mechanisms through which neurotransmitters can regulate growth cone motility and neuronal electrical properties, further supporting evidence for potential roles of neurotransmitters in axon pathfinding and synaptic transmission in vivo.

In addition to acting in synaptic transmission, neurotransmitters have been shown to play roles in the development of nervous system. Developing neurons extend neurites to connect to their target cells, and growth cones at the tip of growing neurites are critical for pathfinding. Although evidence for the regulation of axonal growth and growth cone guidance by neurotransmitters and neuromodulators is emerging, less is known about the mechanisms by which neurotransmitters affect developing neurons. Here, I focus on three neurotransmitters/ neuromodulators and describe their actions (a) at the level of growth cone, especially on filopodia, which serve as sensors that allow growth cones to probe the environment they are traversing, and (b) on how neurotransmitters modulate neuronal electrical properties, which, in itself, have been shown to affect neurite extension. The goals of this dissertation are to investigate 1) the cholinergic modulation of neuronal activity and its effects on growth cone motility; 2) the excitatory modulation of neuronal excitability by nitric oxide (NO); and 3) the inhibitory modulation of neuronal activity by dopamine (DA). The work uses a well-established model system to investigate growth cone motility and neuronal activity: identified neurons from the pond snail Helisoma trivolvis studied in cell culture or in the intact ganglion in situ. The study of B5 neurons demonstrates that acetylcholine (ACh) induces filopodial elongation, which is mediated by opening of nicotinic ACh receptors, membrane depolarization, and elevation of intracellular Ca level in growth cones. This dissertation also shows that NO inhibits two types of Ca-activated K channels to depolarize the membrane potential of B19 neurons. Additionally, the study reveals that DA serves as an inhibitory neurotransmitter to hyperpolarize and silence the electrical activity of firing B5 neurons via a D2-like receptor/PLC/K channel pathway. Taken together, this dissertation elucidates novel cellular mechanisms through which neurotransmitters can regulate growth cone motility and neuronal electrical properties, further supporting evidence for potential roles of neurotransmitters in axon pathfinding and synaptic transmission in vivo.

The purpose of this study was to investigate whether locomotor training, in the form of treadmill training, could ameliorate neurochemical changes and behavioural deficits in the 6-hydroxydopamine (6-OHDA) rat model of Parkinsons disease. It has been recently demonstrated that rehabilitative forelimb motor training can attenuate dopamine loss and some deficits in forelimb movements in this animal model. In addition, brief locomotor treadmill training has been shown to attenuate forelimb deficits in 6-OHDA treated rats. However, it is not known whether locomotor training could result in an amelioration of locomotor deficits in these animals. Rats were lesioned with 6-OHDA injected intracerebrally. Lesioned rats were randomly assigned to one of 3 groups: early treadmill trained, late treadmill trained and untrained. Animals in the treadmill groups were trained to trot on a moving treadmill for 2 x 20 minute sessions daily for 30 days, beginning either 24 hours or 7 days after 6-OHDA injection. Untrained animals were exposed to a stationary treadmill for the same time periods. All animals were assessed on their abilities to perform several behavioural tasks designed to test locomotor and forelimb movement abilities prior to 6-OHDA injection and again at 3 and 6 weeks post-injection. These tests included measurement of ground reaction forces during overground locomotion, paw placements during a ladder crossing task, forelimb useage during exploratory behaviour and ability to initiate forelimb stepping movements. In addition, assessments of dopamine depletion in the striatum were carried out first in vivo, by measuring apomorphine-induced rotations at 2 weeks post 6-OHDA injection, and subsequently by post-mortem analysis of dopamine levels in the striatum using HPLC at the conclusion of the study. Treadmill training resulted in attenuation of dopamine depletion compared to non-treadmill trained animals, as measured by both apomorphine injection and HPLC. However, treadmill training produced no difference in behavioural deficits on a variety of tests compared to untrained animals. In some cases, early treadmill trained animals tended to display more severe behavioural deficits compared to untrained animals. Late treadmill training had a similar but smaller effect compared to early treadmill training. We conclude that treadmill training does not ameliorate locomotor deficits, in the 6-OHDA model of Parkinsons disease, even though this same training results in attenuation of dopamine loss in the striatum.

This study presents a simple process for producing resistance-based humidity sensors utilizing dopamine (DA) coated gold nano-particles (AuNPs) as the sensing material. The sensing material for typical humidity sensors are solid state metal oxides, graft-polymers or salt-doped polymers. However, these humidity sensors may suffer from low sensing response or slow time response since water molecules have to diffuse into the sensing materials to induce the electrical property changes. Alternatively, AuNPs have large surface area for water molecule absorption and can be potentially for high performance humidity sensing. Nevertheless, the surface property of AuNPs is hydrophobic and needs to be modified. In this regards, this work uses a highly hydrophilic molecule of dopamine to modify the surface of AuNP into hydrophilic to enhance the humidity sensing performance. Highly hydrophilic bio-molecule of dopamine is physically bonded onto 4-6 nm AuNPs to enhance the humidity sensing performance. Results show that the DA coated AuNPs have nice humidity sensing responses in the measuring range of 20-90%RH. The measured resistance response shows >1500 times greater than the sensor using the same AuNPs without DA coating. The developed humidity sensor shows rapid time responses for water absorption (13 s) and desorption (30 s), respectively. Moreover, a 3-day long-term measurement at low, medium and high humidity ranges also shows the good stability of the developed sensor. The method developed in this study provides a simple and low-cost method to produce high-performance humidity sensors with DA-coated AuNPs.

Excessive risk-taking is a characteristic of several psychopathological disorders. In order to alleviate maladaptive risky behavior, a thorough understanding of the neurobiological and pharmacological substrates of risky choice must be developed. In this dissertation, the “risky decision-making task” was utilized to explore the mechanisms by which dopamine mediates risky choice. In experiment 1, we characterized rats in risky decision-making as well as a variety of other behavioral traits. This was performed to determine if the behavioral patterns obtained in the risky decision-making task represent an independent cognitive construct rather than a function of a separate behavioral trait. Risky decision-making performance was not correlated with measures of motivation, anxiety, pain tolerance, or other types of decision-making. In contrast, risky choice was correlated with impulsive action as assessed by the Differential Rates of Low Responding Task, suggesting that risky choice may be mechanistically similar to impulsive action. In experiment 2, the effects of various dopaminergic drugs on risky decision-making was investigated. Amphetamine administration attenuated risky choice, while the dopamine antagonist α-flupenthixol had no effect on risky choice. Agonists and antagonists specific to D1 dopamine receptors had no effects on risky choice; however, the D2 dopamine receptor agonist bromocriptine reduced risky choice in a manner similar to amphetamine. Furthermore, coadministration of amphetamine with a D2 antagonist abolished amphetamine’s effects on risky choice, and amphetamine’s effects were unaffected by coadministration of a D1 antagonist. These data suggest that D2 signaling at the receptor is particularly critical to risky decision-making behavior. In experiment 3, D2 dopamine receptor mRNA abundance was assessed in rats that had been previously characterized in risky decision-making using in situ hybridization. Levels of D2 cRNA hybridization in both orbitofrontal cortex (OFC) and medial prefrontal cortex (mPFC) predicted risky decision-making behavior as assessed by nonlinear curve estimation analyses. Interestingly, opposite relationships between D2 mRNA abundance and risky choice were observed in these two cortical areas, with OFC D2 mRNA abundance showing a U-shaped relationship with risky choice, and mPFC D2 mRNA resembling an inverted U-curve. Additionally, increased levels of D2 mRNA in dorsal striatum were observed in risk-averse rats in comparison to risk-taking rats. In conclusion, these data suggest that signaling via D2 dopamine receptors is an important mediator of risky decision-making behavior, and that D2 signaling in frontostriatal circuitry may be particularly relevant toward these behaviors.