Thesis (Ph. D.)--Ohio State University, 2006. / Title from first page of PDF file. Includes bibliographical references (p. 213-226).

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In recent years, the determination of several high-resolution structures of transient receptor potential (TRP) channels has led to significant progress within this field. The primary focus of this dissertation is to elucidate the structural characterization of TRPML1 and TRPP2. Mutations in TRPML1 cause mucolipidosis type IV (MLIV), a rare neurodegenerative lysosomal storage disorder. We determined the first high-resolution crystal structures of the human TRPML1 I-II linker domain using X-ray crystallography at pH 4.5, pH 6.0, and pH 7.5. These structures revealed a tetramer with a highly electronegative central pore which plays a role in the dual Ca2+/pH regulation of TRPML1. Notably, these physiologically relevant structures of the I-II linker domain harbor three MLIV-causing mutations. Our findings suggest that these pathogenic mutations destabilize not only the tetrameric structure of the I-II linker, but also the overall architecture of full-length TRPML1. In addition, TRPML1 proteins containing MLIV-causing mutations mislocalized in the cell when imaged by confocal fluorescence microscopy. Mutations in TRPP2 cause autosomal dominant polycystic kidney disease (ADPKD). Since novel technological advances in single-particle cryo-electron microscopy have now enabled the determination of high-resolution membrane protein structures, we set out to solve the structure of TRPP2 using this technique. Our investigations offer valuable insight into the optimization of TRPP2 protein purification and sample preparation procedures necessary for structural analysis.

Wong, Ching On. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 150-167). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

The transient receptor potential (TRP) channel superfamily is a non-selective Ca2+-permeable cation channels involved in sensory physiology. Here we show that 2-aminoethoxydiphenyl borate (2-APB), a compound commonly used as TRP channel inhibitor, dose-dependently induce a significant facilitation on the frequency of spontaneous neurotransmitter release at developing Xenopus neuromuscular junction through, surprisingly, TRP channel activation. Bath application of universal TRP channel inhibitors either SKF96365, flufenamic acid or RuR cease the 2-APB-induced synaptic facilitation. Exclusion of Ca2+ from culture medium or bath application of the pharmacological Ca2+ channel inhibitor cadmium, membrane-permeable Ca2+ chelator BAPTA-AM, effectively hampered the facilitation of neurotransmitter release induced by 2-APB, suggesting Ca2+ influx is requisite for 2-APB-induced synaptic facilitation. Blockade of the voltage-dependent Ca2+ channel with either nifedipine, verapamil or £s-CTX failed to abolish the SSC facilitating effect of 2-APB. Electrophysiological recording of 2-APB induced single channel currents by using cell-attached patch-clamp technique reveals 2-APB evoked a robust single channel activity recorded at different pipette voltages. Furthermore, the 2-APB-evoked single-channel events are significantly abolished in the presence of SKF96365. Either pretreatment of the cultures with inhibitor of phospholipase C (U73122) or tyrosine kinase (Genistein) abolishes 2-APB induced potentiation of synaptic transmission. The structure of PMA is analogous to diacylglycerol (DAG), which abolishes 2-APB induced synaptic facilitation. 2-APB no longer elicited any changes in SSC frequency when serum is eliminated from culture medium. Overall, results from our current study provide evidences that 2-APB induces the opening of TRP channels and Ca2+ influx which resulting in facilitation of spontaneous neurotransmitter release at developing Xenopus neuromuscular synapse. Serum may activate tyrosine kinase to turn on PI3K and phospholipase C. Then phospholipase C cleavage PIP2 to IP3 and diacylglycerol, and diacylglycerol induced TRP channel opening. 2-APB potentiates and sensitizes the TRP channel, increasing Ca2+ inffux. Elevated [Ca2+]i resulted in enhancement of neurotransmitter release from presynaptic nerve terminal.

Auditory receptor cells rely on force-gated channels to transform sound stimuli into neural activity. These primary auditory neurons form the first stage of the neural circuits that support a host of higher-order functions, such as the localization of sound or the comprehension of speech. The mechanisms of sound transduction, as well as higher-order processes such as acoustic communication during courtship, can be studied in the fruit fly Drosophila melangogaster, a model organism with a suite of powerful genetic tools. However, this work is hampered by incomplete knowledge of the components of the Drosophila auditory system and a lack of high resolution techniques for investigating their function. We used several approaches to identify candidate Drosophila central auditory neurons and developed techniques for measuring the activity of identified neurons in vivo. As an outgrowth of this work, we also developed a non-invasive method for measuring generator currents in the primary auditory neurons. Chapter 4 describes this technique and provides a basic characterization of the sensitivity of the Drosophila auditory system to sound. Determining the sensitivity of the Drosophila auditory system is necessary for understanding the neural basis of acoustic communication and has implications for the mechanism of transduction. The force-gated ion channel that transforms sound into an electrical signal has not been identified in any species. Several TRP channels have been implicated in Drosophila auditory transduction, but mechanistic studies have been hampered by the inability to record subthreshold signals from auditory receptor neurons. We recorded generator currents from primary auditory neurons to assess the roles of several TRP family members in transduction. We found that the TRPN family member NompC is not required for transduction, despite the fact that it is required for the active amplification of motion by the auditory organ. Instead, NompC is required for a process that sensitizes the transduction complex to movement and regulates the resting forces on the complex. In contrast, the TRPV channels Nanchung and Inactive are required for responses to sound, suggesting they are components of the transduction complex. Thus, transduction and active amplification are genetically separable processes in the Drosophila auditory system.

Mode of access: World Wide Web. / Thesis (Ph. D.)--University of Hawaii at Manoa, 2005. / Includes bibliographical references (leaves 102-113). / Electronic reproduction. / Also available by subscription via World Wide Web / xvi, 113, pp leaves, bound ill. (some col.) 29 cm

Thesis (Ph. D.)--Ohio State University, 2005. / Title from first page of PDF file. Document formatted into pages; contains xviii, 178 p.; also includes graphics. Includes bibliographical references (p. 163-178). Available online via OhioLINK's ETD Center

Thesis (Ph.D.)--University of California, San Francisco, 2005. / Includes bibliographical references. Also available online.

Thesis (Ph.D.)--University of California, San Francisco, 2004. / Includes bibliographical references. Also available online.