No description available.

No description available.

Parkinson’s disease (PD) is the most common movement neurodegenerative disease affecting approximately 1% of the population over 60. Though originally thought to be sporadic in nature, a genetic component is increasingly being linked to the disease. Of these genes, mutations in DJ-1 (PARK7) cause early onset autosomal recessive PD. Initial workup of the DJ-1 protein has suggested that it may act in the cell by combatting oxidative stress though the mechanism by which it does so is unclear. Thus, though much work has attempted to elucidate a function at the biochemical, cellular and organismal level, the overt physiological role of DJ-1 remains elusive. In this dissertation, we explore the mechanisms through which DJ-1 confers neuroprotection, particularly in the case of oxidative stress insult. We demonstrate that DJ-1 acts through the pro-survival protein AKT to accomplish its neuroprotective function. Moreover, we note that DJ-1 likely serves its role as an antioxidant through the NRF2 master antioxidant regulator pathway a pathway that is, itself, likely to be regulated by AKT. Together, our results demonstrate that neuroprotection by DJ-1 is done through a signaling pathway involving both AKT and NRF2 and that disruption of the former in PD likely results in abolishing this signaling pathway. Finally, to generate a better animal model of PD, we demonstrate that backcrossing DJ-1 null mice - which originally did not demonstrate any gross histopathological or behavioral phenotypes – display unilateral dopaminergic degeneration that progresses to bilateral degeneration with aging, a feature reminiscent of classical PD progression. Collectively, this thesis takes a two-sided approach to address the biochemical and physiological functions of DJ-1 within the cell and the mouse in hopes of elucidating mechanisms of neuronal death to devise better translational therapies.

The primary role of acetylcholinesterase (AChE) is hydrolysis of acetylcholine (ACh). However, observations by numerous groups have suggested that AChE may have non-cholinergic functions. Furthermore, developmental roles for AChE and its related enzyme, butyrylcholinesterase (BuChE), which is also capable of ACh hydrolysis, have been postulated. One line of evidence to support a non-cholinergic role for AChE is the apparent disparity in several brain areas between the distribution of AChE and the cholinergic marker choline acetyltransferase. The substantia nigra (SN), an area of the ventral mesencephalon (VM), which contains the dopaminergic cells that degenerate in Parkinson's disease (PD), is an area that displays such a disparity. One approach to treating PD involves implantation of embryonic dopaminergic VM cells into the parkinsonian brain. This procedure, known as foetal transplantation, has met with limited success, in part due to degeneration of dopaminergic cells within the donor preparation. It is known that incorporation of trophic factors into the preparation for grafting improves dopaminergic cell survival. It has previously been shown that AChE enhances survival and neurite outgrowth of postnatal dopaminergic cells in organotypic cultures of the VM. The aim of the studies in this thesis was to establish the effects of BuChE and AChE on embryonic dopaminergic neurons in a preparation analogous to that used in the animal model of foetal transplantation as a treatment for PD. Addition of BuChE and monomeric (G_1-) and tetrameric (G_4-) forms of AChE enhanced dopaminergic neurite outgrowth. Inhibition of the active site of BuChE and AChE by echothiophate had no effect upon neurite outgrowth or cell survival, demonstrating that the trophic effects of BuChE and AChE on neurite outgrowth were not dependent upon ACh hydrolysis. In contrast, inhibition of the peripheral anionic site (PAS) of AChE by BW284c51 markedly decreased cell survival and neurite outgrowth. The mechanism of action of BW284c51 toxicity was subsequently investigated using a mixture of nicotinic ACh receptor antagonists in order to demonstrate that the chronic toxic effects of BW284c51 were not a consequence of elevated ACh resulting from inhibition of AChE. Finally, the technique of whole-cell patch-clamp electrophysiology revealed a novel inhibitory effect of BuChE and G_1- and G_4-AChE on voltage-dependent calcium currents. It was postulated that these actions underlie the trophic effects of BuChE and AChE on embryonic dopaminergic neurons, a suggestion that was supported by the findings that established inhibitors of voltage-dependent calcium currents enhanced dopaminergic neurite outgrowth. The findings of this thesis are discussed in the context of other studies and are related to both physiological and pathological functions of the central nervous system.

No description available.

<p>Deep brain stimulation (DBS) is an effective therapy for motor symptoms in Parkinson's disease (PD). DBS efficacy depends on the stimulation parameters, and the current gold standard therapy is high-frequency stimulation (>100 Hz) with constant interpulse intervals and short pulse widths (<210 &#956;s). However, the temporal pattern of stimulation is a novel parameter dimension that has not been thoroughly explored. We used non-regular temporal patterns of DBS to pursue two goals: to better understand the mechanisms of DBS, and to increase the efficacy and efficiency of DBS for PD.</p><p>First, we designed high frequency patterns of non-regular stimulation with distinct features proposed to be important for efficacy and evaluated these patterns in human subjects with PD. Unexpectedly, some non-regular patterns of stimulation improved performance of an alternating finger-tapping task-a proxy for bradykinesia-compared to high frequency regular stimulation. Performance in the motor task was correlated with suppression of beta band power in a computational model of the basal ganglia suggesting a possible mechanism for effective stimulation patterns.</p><p>Inspired by the increased clinical efficacy of non-regular patterns of stimulation with high average frequencies, we developed a non-regular pattern of stimulation that reduced motor symptoms in PD using a low average stimulation frequency. Since the number of potential combinations of interpulse intervals is exceedingly large and it is unclear how such timing should be selected, we applied computational evolution to design an optimal temporal pattern of deep brain stimulation to treat the symptoms of PD. Next, we demonstrated the efficacy of the resulting pattern of stimulation in hemi-parkinsonian rats and humans with PD. Both the optimized stimulation pattern and high frequency stimulation suppressed abnormal oscillatory activity in the basal ganglia in the rat and human, providing a shared mechanism of action for effective stimulation patterns. This innovation could allow patients to achieve battery life savings compared to traditional high frequency stimulation, which will reduce the costs and risks of frequent battery replacement procedures. Further, our approach can be used to design novel temporal patterns of stimulation in other applications of neural stimulation.</p><p>Finally, we explored evoked field potentials in the subthalamic nucleus (STN) in response to DBS. These potentials were evoked by stimulation through one of the contacts on the DBS lead and recorded from the two surrounding contacts. Subthalamic DBS local evoked potentials (DLEPs) have never before been recorded. We characterized the DLEPs, differences across DBS frequencies and time, their relationship to beta frequency oscillations and phase-amplitude coupling, and their dependence on electrode contact location.</p><p>A 3-dimensional biophysical model of DBS in the subthalamic nucleus-globus pallidus externus (GPe) subcircuit was built to explore the neural origin of the DLEPs. The computational model could reproduce the DLEP signal, and it revealed that the quasi-periodic DLEP oscillations are caused by excitatory synaptic currents in STN interrupted periodically by inhibition from GPe.</p><p>DLEP power was correlated with beta band oscillation power in the recordings without DBS, and significant phase-amplitude coupling was observed in a subset of subjects with robust DLEP responses. Together, all available evidence suggested the contact location was an important determinant for the presence and characteristics of DLEP signals. Predictions were made concerning contact location relative to the boundaries of the STN based on the DLEP recordings and insights gained using the computational model, and the predictions were in agreement with blinded post hoc imaging based contact localization for ~70% of contacts predicted to be within STN.</p><p>DLEPs are an exciting new signal with several useful applications. DLEPs could help neurosurgeons verify accurate DBS lead placement or optimal stimulation parameters, probe the pathological basal ganglia, and elucidate the mechanisms of DBS.</p> / Dissertation

No description available.

Among all symptoms of Parkinson’s Disease (PD), depression and sleep dysfunction have the highest impact on quality of life, yet sleep disturbances and depression symptoms are often left unrecognized and untreated. With the rising annual cost of pharmacologic treatments for PD and the increasing prevalence of PD in the United States, there is a need to implement effective non-pharmacologic regimens, such as physical exercise. Adherence to exercise regimens can often be challenging, especially for elderly patients with progressive neurological impairment. However, enjoyable exercises, such as dancing, involve socialization and musical stimuli that are associated with increased motivation in patients. Dance exercise has been shown to have significant improvement in motor symptoms, functional mobility, mood, and quality of life in PD patients compared to no intervention or traditional exercises. Previous studies have shown that Tango style dancing has additional benefits for PD patients as it may selectively activate areas of the brain associated with motor and non-motor symptoms (the basal ganglia and anterior putamen, respectively) during backward walking and metered rhythmic movement. However, no studies have yet investigated the effect of dance intervention on sleep quality in patients with PD. The proposed study is a randomized control trial that will compare the sleep quality improvement of 90 elderly PD patients in two treatment arms, a Tango dance intervention with walking and walking alone (control group). The Tango group will participate in a 6-month Adapted Tango class designed for PD patients. Sleep quality will be measured as the primary outcome using the Parkinson’s Disease Sleep Scale (PDSS-2). Depression will be measured as a secondary outcome using the BDI-II. This will be the first study to investigate the effects of dance intervention on sleep quality in patients with PD, applying an adapted Tango program similar to those used in previous studies. If the results of this study reveal positive effects of Tango on sleep quality, clinicians may be able to recommend Tango-style dance exercise as a therapeutic intervention to target sleep disturbances and improve quality of life for patients with PD.

University of Macau / Institute of Chinese Medical Sciences

α-synuclein (αS) is an intrinsically disordered protein that is strongly connected with Parkinson’s disease (PD) and a number of other neurodegenerative disorders, including Parkinson’s disease with dementia, dementia with Lewy bodies and multiple system amyotrophy. Fibrillar aggregates of αS have been identified as the major constituents of proteinaceous inclusions known as Lewy bodies that form inside the neurons of patients suffering from these conditions. A number of missense mutations, as well as duplications and triplications of the gene encoding αS have also been associated with familial forms of early onset PD. Despite the association between αS aggregation and neurodegeneration is now established, the specific function of αS is still currently unclear, however, a general consensus is forming on its key role in regulating the process of neurotransmitter release, which is associated with the ability of αS to bind a variety of biological membranes. Indeed, in dopaminergic neurons, αS exists in a tightly regulated equilibrium between water-soluble disordered state and membrane-associated forms that are rich in α-helix. Characterising the nature of this binding as well as the structural and functional properties of αS at the surface of biological membranes is currently a top challenge. In particular the intrinsic limitation of current analytical techniques in studying highly heterogeneous protein states in rapid equilibrium between different physical phases demands for novel approaches to be formulated. This PhD thesis describes major achievements in developing and applying a multidisciplinary approach based on solution and solid-state NMR and extending to a number of other biophysical techniques, including cryo electron microscopy, super resolution microscopy, FRET and cellular biophysics, which enabled us to elucidate in detail the balance between structural order and disorder associated with the membrane interaction of αS in view of its physiological and pathological roles. Using this approach, we identified the key elements that govern the binding of αS to synaptic vesicles (Chapter III). In particular, three regions of αS were shown to possess distinct structural and dynamical properties at the surface of synaptic vesicles, including an N-terminal helical segment having a role of membrane anchor, an unstructured C-terminal region that is weakly associated with the membrane and a central region acting as a sensor of the lipid properties and determining the affinity of αS membrane binding. We refined the structural ensemble of the N-terminal membrane anchor at the surface of synaptic membranes, showing that the partial insertion of this region in the membrane core promotes strong but reversible binding with biological membranes in such a way to enable a fast equilibrium between membrane-bound and cytosolic forms of the protein (Chapter III). Further studies of two mutational variants of αS that are associated to early onset PD, namely A30P and E46K, revealed that two key regions of the protein, namely the N-terminal membrane-anchor (residues 1 to 25) and the central segment of the sequence (residues 65–97, having significant overlap with the non-amyloid β component - NAC - region), have independent membrane-binding properties and therefore are not only able to interact with a single SV, but can also simultaneously bind to two different vesicles thereby promoting their clustering (Chapter IV). The resulting “double-anchor” mechanism explains the biological property of αS to promote clusters of synaptic vesicles within the processes of formation of distal pools to the active zone. The double-anchor mechanism reconciles literature data showing that the deletion of the segment 71–82 in the NAC region of αS or the impairment of the membrane affinity of the N-terminal anchor region of the protein severely affect vesicle clustering in vivo. Thus our data revealed that the NAC region is not only involved in the aggregation of αS, as extensive literature evidence has previously indicated, but also has a specific role in a key molecular mechanism associated with the normal function of αS. The structural characterisation also showed that the active conformations of αS to initiate the double-anchor mechanism are particularly vulnerable to self-association leading to αS aggregation at membrane surfaces, thereby providing a new mechanistic link between functional and pathological roles of αS. In addition to studying the physiological membrane interactions by αS, we characterised the fundamental mechanism of membrane disruption by αS oligomers resulting in the generation of neuronal toxicity in PD (Chapter V). Indeed, while fibrillar aggregates of αS represent the major histopathological hallmarks of PD, small oligomeric assemblies of this protein are believed to play a crucial role in neuronal impairment. We obtained a detailed structural characterisation of toxic αS oligomers and compared these results to the study of non-toxic oligomeric species. The results reveal the fundamental structural characteristics driving the toxicity of αS oligomers, including a highly lipophilic element that promotes strong interactions with biological membranes and a structured region that inserts into lipid bilayers and disrupts their integrity. We obtained additional support for these conclusions by showing that mutations targeting the region of αS promoting such interactions with the membrane dramatically suppress the toxicity of αS aggregates in neuroblastoma cells and primary cortical neurons. Taken together our studies enabled the characterisation of a series of structural properties of the membrane-bound states of αS in both its monomeric and oligomeric forms. The results revealed the nature of the fine balance between functional and pathological membrane interactions of αS and delineated how subtle perturbations of this equilibrium can lead to the rapid evolution of processes that trigger pathological mechanisms. Understanding this balance is a top challenge for advancing the research in PD and requires innovation across different disciplines to overcome current limitations in probing the conformational transitions of this disordered and metamorphic neuronal protein.