Shedding light on nicotinic acetylcholine receptors: Creation and use of fluorescently tagged receptors

Neuroscience research advances forward each time a new tool is created. Hodgkin and Huxley ushered in the age of neurocomputation, before most labs had computers, by providing a useful electrochemical model of voltage changes across the plasma membrane. Magnetic resonance imaging and positron emission tomography have contributed greatly to our understanding of how brain regions work in concert during specific tasks and may one day reveal our innermost feelings in a way that would astound Freud. Additionally, Roger Tsien's group has quite literally shed new light on protein dynamics in neurons with their rainbow of fluorescent proteins.;One aim of this work was to exploit advances in molecular biology and bionanotechnology to create the next generation of photonic probes that will shed even more light on neural proteins for basic research, and then to demonstrate the utility of these tools for commercial uses, such as drug discovery and lead compound selection. To this end, a neurotransmitter receptor subunit was genetically labeled with a fluorescent protein and used to confirm the existence of a putative receptor subtype, which then provided clues as to the role of one of its subunits. Additionally, the fluorescent subunits were employed to identify small peptides from a random library for use as future probes. They may be used in a heterologous expression system to demonstrate their suitability for cell-based high throughput screening assays for lead drug compound selection. Another aim of this work was to expand the use of computational modeling from studying small areas of neurons in the brain to predicting inter-regional behavior. This was done with the hope that future imaging technologies will be able to test this model using fluorescent voltage sensitive dyes and possibly some of the probes which could be developed as a result of this work.