One-channel Cell-attached Patch-clamp Recording |
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| Authors: | Bruce A Maki Kirstie A Cummings Meaghan A Paganelli Swetha E Murthy Gabriela K Popescu |
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| Institution: | 1.Graduate Program in Neuroscience, SMBS, University at Buffalo, SUNY;2.Department of Biochemistry, SMBS, University at Buffalo, SUNY;3.Molecular and Cellular Neuroscience Department, The Scripps Research Institute;4.Department of Biochemistry and Graduate Program in Neuroscience, SMBS, University at Buffalo, SUNY |
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| Abstract: | Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate depends on properties intrinsic to each channel protein as well as the mechanism by which it is generated and controlled and represents an important area of current research. Information about the operational dynamics of ion channel proteins can be obtained by observing long stretches of current produced by a single molecule. Described here is a protocol for obtaining one-channel cell-attached patch-clamp current recordings for a ligand gated ion channel, the NMDA receptor, expressed heterologously in HEK293 cells or natively in cortical neurons. Also provided are instructions on how to adapt the method to other ion channels of interest by presenting the example of the mechano-sensitive channel PIEZO1. This method can provide data regarding the channel’s conductance properties and the temporal sequence of open-closed conformations that make up the channel’s activation mechanism, thus helping to understand their functions in health and disease. |
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| Keywords: | Neuroscience Issue 88 biophysics ion channels single-channel recording NMDA receptors gating electrophysiology patch-clamp kinetic analysis |
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