A Real-Time Device for Constructing Virtual Ion Channels in Living Cells

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0085177WhiteIntroduction. Electrical activity in nerve and muscle cells is generated by populations of ion channels that "gate" (open or close) in response to changes in transmembrane voltage and/or the concentrations of crucial chemicals. In studies of the biophysical processes underlying electrical activity, scientists and engineers rely upon two basic recording configurations. In the first recording configuration, commonly referred to as current clamping, the researcher controls the amount of net transmembrane current (i.e., current across the cell membrane) and measures transmembrane voltage. In the second recording configuration, called voltage clamping, the researcher uses an electrical feedback circuit to control transmembrane voltage and measures transmembrane current. Current clamping is useful for characterizing the patterns of electrical activity generated by a given nerve or muscle cell; voltage clamping is useful for studying the biophysical mechanisms underlying a particular pattern of electrical activity.More recently, a third very useful recording configuration has been developed, in which neither transmembrane current nor transmembrane voltage is the controlled variable. Instead, the researcher uses a sophisticated recording system to mimic in real time the electrical conductance associated with a given population of virtual ion channels. This recording mode, called dynamic clamping, allows the researcher to block native ion channels, and replace them with virtual analogs, the properties of which can be controlled precisely. Dynamic clamping and other real-time-computing-based protocols enable entirely new classes of experiments, in which (for example) the applied stimulus can mimic the behavior of a (blocked) dynamic component of the system, and thus be used to determine unequivocally the effects of the mimicked component of overall behavior.Dynamic clamping and other real-time experimental techniques show enormous promise as important research tools. Dynamic clamping could be used, for example, to study the electrical effects of computer-designed pharmaceutical agents even before the agents are developed in the laboratory. So far, however, the impact of this technique has been educed by three interrelated difficulties. First, the technical complexities of its design are beyond the skills of most end-users, and no one has yet provided a flexible, powerful, turn-key system. Second, existing dynamic clamp systems do not account for the seemingly stochastic (i.e., probabilistic) nature of voltage-gated ion channels. Adding this capability would allow researchers to attack entirely new sets of exciting problems that are as yet unapproachable. Third, existing dynamic clamp systems cannot account for measured or assumed spatial distributions of ion channels.Specific Aims. The specific aims of this proposal are (1) to complete construction of a stochastic dynamic clamp (SDC) system that can be used to study the actions of noisy virtual voltage- and ligand-gated ion channels in living cells; (2) to create a web-based system of support to help end-users adopt and use the SDC system for dynamic clamping and other real-time experimental applications; (3) to develop methods for representing virtual ion channels that are remote from the recording site in the cell body, and (4) to use the new SDC system to test specific hypotheses regarding the relative importance of noise from synaptic sources and noise from voltage-gated ion channels in limiting neuronal reliability.The project will have educational impact at both the graduate and undergraduate levels, in the context of the classroom and research projects. Innovations from Specific Aims 1 and 3 will extend the dynamic clamp method to account for stochastic and spatially distributed channels. The investigators' support (Aim 2) will be a crucial step in developing a turn-key system. The system will be easily adaptable to apply techniques of real-time computation in many biomedical engineering applications. Field tests of the device (aim 4) will push the field of neurobiology in a more quantitative, information-oriented direction. In particular, they will advance the study of the biophysical underpinnings of neuronal reliability, which play a fundamental role in the understanding of coding strategies used by the nervous system. Such advances are not practically achievable without real-time computing technology.