No description available.

No description available.

No description available.

The peripheral nervous system (PNS) is composed of a diverse array of neurons that mediate sensation. This includes sensory circuits that encode external stimuli, as well as circuits that provide information flow from our internal organs. My PhD training has focused on addressing two questions: 1) what molecular mechanisms underlie this functional diversity, and 2) can we engineer non-invasive therapeutics to modulate PNS activity? To study the molecular mechanisms of sensory function, I employed the Merkel-cell neurite complex as a model system. Merkel cells are mechanosensory epidermal cells that have long been proposed to activate neuronal afferents through chemical synaptic transmission. RNA sequencing of adult mouse Merkel cells demonstrated that they express presynaptic molecules and biosynthetic machinery for adrenergic transmission. Moreover, live-cell imaging showed that Merkel cells mediate activity- and VMAT- dependent release of fluorescent catecholamine neurotransmitter analogues. Touch-evoked firing in Merkel-cell afferents was inhibited either by silencing of SNARE-mediated vesicle fusion from Merkel cells or by neuronal deletion of b2-adrenergic receptors. Next, to develop non-invasive technologies for peripheral nerve modulation, I employed targeted focused ultrasound (FUS) stimulation and electrophysiology to record activity of individual mechanosensory neurons. Parameter space exploration showed that stimulating neuronal receptive fields with high-intensity, millisecond FUS sonication reliably and repeatedly evoked action potentials in peripheral neurons. FUS elicited action potentials with latencies comparable to electrical stimulation, demonstrating both speed and reliability of the technique. Lastly, I show that peripheral neurons can be both excited by FUS stimulation targeted to either skin receptive fields or peripheral nerve trunks, a key finding that increases the therapeutic range of FUS-based peripheral neuromodulation.

Thesis (Ph.D.)--Georgetown University, 2009. / Includes bibliographical references.

Glycinergic neurotransmission is a major inhibitory influence in the CNS and defects are associated with paroxysmal neuromotor disorder, hyperekplexia with mutations in subunits of the inhibitory glycine receptor which facilitates postsynaptic ligand-binding, ion-channels. This study investigates the human glycinergic system by; 1) Mutation analysis of glycinergic candidate genes in hyperekplexia: the DNA sequencing of GLRAl in 88 hyperekplexia patients revealed 30 sequence variants; 21 were inherited in recessive mode or part of compound heterozygosity, indicating that recessive hyperekplexia is more common than previously expected. Further screening of the glycine transporter-2 gene (SLC6A5) as a candidate gene, 12 SLC6A5 mutations were found in 7 human hyperekplexia cases inherited predominantly by compound heterozygosity. 2) Biophysical analysis and molecular modelling of GLRAl mutations: which demonstrated that subcellular localisation defects were the major mechanism underlying recessive mutations. Other mutants typically show alterations in the dose-response curve for glycine suggestive of disrupted signal transduction. This study reports the first hyperekplexia mutation associated with leaky current suggesting tonic channel opening as a new receptor mechanism and fully-supported by molecular modelling. 3) Molecular and immunoreactive analysis of gephyrin heterogeneity in human brain: gephyrin encodes a multifunctional cytoplasmic protein important for organizing glycine and GABAa receptors at the postsynaptic membrane. Gephyrin has many different transcript isoforms and the study describes the population / distribution of gephyrin isoforms in neuronal tissues using molecular and immunohistochemical techniques. The heterogeneity of gephyrin cassettes indicates that each cassette is temporally and spatially regulated with unique patterns of glycine receptors co-localisation and we hypothesise that different gephyrin isoforms exhibit differential binding specificity affecting protein-protein interactions. This thesis describes that hyperekplexia is definitively a glycinergic disorder with several mechanism of molecular pathogenicity. Moreover, the underlying complexity of proteins, such as gephyrin, reveals further challenges in interpretating the functional significance of the neuronal heterogeneity.

No description available.

Mammalian neurons exhibit extreme sensitivity to oxygen deprivation and undergo rapid and irreversible degeneration when oxygen supply is curtailed. Though several neuroprotective pathways are activated during oxygen deprivation, their analyses are masked by the complex series of pathological events which are triggered simultaneously. Such events can be analyzed in the anoxia tolerant fresh water turtle, which can inherently survive the conditions of oxygen deprivation and post-anoxic reoxygenation without brain damage. It is likely in such a model that modulation of a particular molecular pathway is adaptive rather than pathological. The major objective behind this study was to analyze the intracellular signaling pathways mediating the protective effects of adenosine, a potential neuromodulator, and its effect on cell survival by influencing the key prosurvival proteins that prevent apoptosis. In vivo and in vitro studies have shown that adenosine acts as a neuroprotective metabolite and its action can be duplicated or abrogated using specific agonist and antagonists. Stimulating the adenosine receptors using selective A1 receptor agonist N6-cyclopentyladenosine (CPA) activated the presumed prosurvival ERK and P13-K/AKT cascade promoting cell survival, and suppression of the receptor using the selective antagonist DPCPX (8- cyclopentyl-1,3-dipropylxanthine) activated the prodeath JNK and P38 pathways. The complex regulation of the MAPK's/AKT signaling cascades was also analyzed using their specific inhibitors. The inhibiton of the ERK and AKT pathway increased cell death, indicating a prosurvival role, whereas inhibiton of the JNK and p38 pathway increased cell survival in this model. In vitro studies have also shown a high Bcl-2/BAX ratio during anoxia and reoxygenation, indicating a strong resistance to cell death via apoptosis. / Silencing of the anti-apoptotic Bcl-2 gene using specific siRNA upregulated levels of prodeath BAX, thus altering the Bcl-2/BAX ratio and elevating cleaved Caspase-3 levels leading to increased cell death. Another promising neuroprotective target which we analyzed was Neuroglobin, which was induced during oxygen crisis and silencing this gene indicated that its plays a major role in modulation of ROS. This study strongly emphasizes the advantages of an alternate animal model in elucidating neuroprotective mechanisms and revealing novel therapeutic targets which could eventually help clinicians to design new stroke therapies based on naturally tolerant organisms. / by Gauri Nayak. / Thesis (Ph.D.)--Florida Atlantic University, 2009. / Includes bibliography. / Electronic reproduction. Boca Raton, Fla., 2009. Mode of access: World Wide Web.

Experimental and computational investigations addressing how various neural functions are achieved in the brain converged in recent years to a unified idea that the neural activity underlying most of the cognitive functions is distributed over large scale networks comprising various cortical and subcortical areas. Modeling approaches represent these areas and their connections using diverse models of neurocomputational units engaged in graph-like or neural field-like structures. Regardless of the manner of network implementation, simulations of large scale networks have encountered significant difficulties mainly due to the time delay introduced by the long range connections. To decrease the computational effort, it is common to assume severe approximations to simplify the descriptions of the neural dynamics associated with the system's units. In this dissertation we propose an alternative framework allowing the prevention of such strong assumptions while efficiently representing th e dynamics of a complex neural network. First, we consider the dynamics of small scale networks of globally coupled non-identical excitatory and inhibitory neurons, which could realistically instantiate a neurocomputational unit. We identify the most significant dynamical features the neural population exhibits in different parametric configuration, including multi-cluster dynamics, multi-scale synchronization and oscillator death. Then, using mode decomposition techniques, we construct analytically low dimensional representations of the network dynamics and show that these reduced systems capture the dynamical features of the entire neural population. The cases of linear and synaptic coupling are discussed in detail. In chapter 5, we extend this approach for spatially extended neural networks. / We consider the dynamical behavior of a neural field-like network, which incorporates many biologically realistic characteristics such as heterogeneous local and global connectivity as well as dispersion in the neural membrane excitability. We show that in this case as well, we can construct a reduced representation, which may capture well the dynamical features of the full system. The method outlined in this dissertation provides a consistent way to represent complex dynamical features of various neural networks in a computationally efficient manner. / by Roxana A. Stefanescu. / Thesis (Ph.D.)--Florida Atlantic University, 2009. / Includes bibliography. / Electronic reproduction. Boca Raton, Fla., 2009. Mode of access: World Wide Web.

Neuronal cell adhesion molecules of L1 family play a critical role in proper nervous system development. Various mutations on human L1-CAM that lead to severe neurodevelopmental disorders like retardation, spasticity etc. termed under L1 syndrome. The vertebrr their roles in axon pathfinding, neurite extension and cell migration, howeverate L1CAM and its homolog in Drosophila, neuroglian (nrg) have been well studied fo, much less is known about the mechanisms by which they fine tune synaptic connectivity to control the development and maintenance of synaptic connections within neuronal circuits. Here we characterized the essential role of nrg in regulating synaptic structure and function in vivo in a well characterized Drosophila central synapse model neuron, the Giant Fiber (GF) system. Previous studies from our lab revealed that the phosphorylation status of the tyrosine in the Ankyrin binding FIGQY motif in the intracellular domain of Nrg iscrucial for synapse formation of the GF to Tergo-Trochanteral Motor neuron (TTMn) synapse in the GF circuit. The present work provided us with novel insights into the role of Nrg-Ank interaction in regulating Nrg function during synapse formation and maintenance. By utilizing a sophisticated Pacman based genomic rescue strategy we have shown that dynamic regulation of the Neuroglian–Ankyrin interaction is required to coordinate transsynaptic development in the GF–TTMn synapse. In contrast, the strength of Ankyrin binding directly controls the balance between synapse formation and maintenance at the NMJ. Human L1 pathological mutations affect different biological processes distinctively and thus their proper characterization in vivo is essential to understand L1CAM function. By utilizing nrg14;P[nrg180ΔFIGQY] mutants that have exclusive synaptic defects and the previously characterized nrg849 allele that affected both GF guidance and synaptic function, we were able to analyze pathological L1CAM missense mutations with respect to their effects on guidance and synapse formation in vivo. We found that the human pathological H210Q, R184Q and Y1070C, but not the E309K and L120V L1CAM mutations affect outside-in signaling via the FIGQY Ankyrin binding domain which is required for synapse formation and not for axon guidance while L1CAM homophilic binding and signaling via the ERM motif is essential for axon guidance in Drosophila. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2014. / FAU Electronic Theses and Dissertations Collection