Potentiometric probes are dyes which, when bound to the membranes of neurons, cardiac and skeletal muscle, glands, and other cells, behave as molecular indicators of membrane potential. The optical properties of these molecules vary linearly with potential and may be used to monitor action potentials, synaptic potentials, or other changes in membrane voltage from a large number of sites at once, without the use of electrodes. For twenty five years our laboratory has pioneered the technology for using potentiometric probes, and developed new optical methods for use in cellular neurophysiology, including a high resolution system for Multiple Site Optical Recording of Transmembrane Voltage (MSORTV), capable of monitoring changes in membrane potential from as many as 464 loci at once. We will continue to apply these techniques to the study of a quasi-2- dimensional nervous system that is uniquely amenable to analysis by optical means, the submucous plexus of the guinea-pig small intestine. Our eventual goal is the complete analysis of the ensemble behavior of this intact mammalian neural network. First, we will use potentiometric dyes to monitor the electrical activity of all of the neurons in what are, perhaps, the closest approximations to mammalian simple nervous systems: rings of connected ganglia in the submucous plexus. Second, we will employ a novel analytical tool (the gravitational transformation) to elucidate with unprecedented completeness the ensemble behavior of an intact mammalian neural network. Third, we will use multiple site optical recording methods and the gravitational transformation, to study the dynamic response of this network to pharmacological and physical interventions that alter the functional connectivity of the circuit and modify its behavior. Fourth, we will examine the persistence and spatial coherence of neuronal assemblies by using fluorescent Ca-indicator dyes to monitor spatio-temporal patterns of electrical activity over time scales lasting tens of minutes.
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