Ion channels play key roles in the physiology of neuronal and non-neuronal cells, in information processing in the nervous system, in brain development, and in a broad spectrum of neurological and non-neurological disorders, such as cardiovascular disease and infertility conditions. To this day, the gold standard for studying ion channels is the patch clamp technique, a laborious technique based on carefully sealing the aperture of a pipette against the cell membrane (the gigaohm seal ). The technique is not easily amenable to automation, so the low throughput of pipette-based recordings is a serious bottleneck for pharmacological screening of ion channel-targeting compounds. Furthermore, recording from >2-3 cells simultaneously is not possible with pipettes;as a result, investigations of communication in complex mature and developing neural networks are limited to extracellular recordings or optical imaging of intracellular calcium activity, both of which are limited in their ability to provide detailed information about intracellular electrical activity. Several groups have recently reported the successful operation of various designs of patch clamp chips, all based on positioning the cell against a microfabricated aperture. We have recently developed a microfluidic patch clamp chip that allows for obtaining gigaohm seals with yields comparable to or surpassing those achievable with a pipette;the performance was evaluated on single rat basophilic leukemia cells during whole-cell recordings of the inward- rectifying potassium ion channel. Here we propose to extend our recent work to recordings from cultured embryonic cortical slices using a novel slice preparation that has a clean layer of cells on the surface of the slice. With the multi-unit patch clamp chip we will investigate the propagation of neuronal signals across developing cortical networks.
The successful completion of this project could reveal neuronal communication mechanisms that underlie the propagation of activity waves seen in development (as part of the normal developmental program) as well as in epilepsy. Furthermore, the same technology would enable low-cost screening of ion channel-targeting compounds (which constitute ~25% of drugs) for their effects on single ion channel currents (approximately 50% of safety-related withdrawals of drugs from the market are due to undesired side effects on ion channels, so the FDA now recommends that all drug candidates usually pools of >10,000 compounds be patch clamp-tested for their effects on the hERG channel).Ion channels play key roles in all known brain functions. Using patch clamp chips, it is now possible to monitor the ion channel activity of large numbers of single dissociate cells (but not of brain slices). We propose to develop a patch clamp chip design for monitoring multiple cells on the surface of brain slices. We will use the device to investigate the propagation of neuronal signals across developing cortical networks.
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