A novel voltage clamp technique for mapping ionic currents from cultured skeletal myotubes.
The biophysical properties and cellular distribution of ion channels largely determine the input/output relationships of electrically excitable cells. A variety of patch pipette voltage clamp techniques are available to characterize ionic currents. However, when used by themselves, such techniques are not well suited to the task of mapping low-density channel distributions. We describe here a new voltage clamp method (the whole cell loose patch (WCLP) method) that combines whole-cell recording through a tight-seal pipette with focal extracellular stimulation through a loose-seal pipette. By moving the stimulation pipette across the cell surface and using a stationary whole-cell pipette to record the evoked patch currents, this method should be suitable for mapping channel distributions, even on large cells possessing low channel densities. When we applied this method to the study of currents in cultured chick myotubes, we found that the cell cable properties and the series resistance of the recording pipette caused significant filtering of the membrane currents, and that the filter characteristics depended in part upon the distance between the stimulating and recording pipettes. We describe here how we determined the filter impulse response for each loose-seal pipette placement and subsequently recovered accurate estimates of patch membrane current through deconvolution.
Anson
B D
BD
Institute of Neuroscience, University of Oregon, Eugene 97403-1254, USA. bdanson@facstaff.wisc.edu
Roberts
W M
WM
D000818:Animals; D016477:Artifacts; D002478:Cells, Cultured; D002642:Chick Embryo; D004558:Electric Stimulation; D004867:Equipment Design; D008564:Membrane Potentials; D018485:Muscle Fibers, Skeletal / Q000502:physiology*; D018482:Muscle, Skeletal / Q000502:physiology*; D018408:Patch-Clamp Techniques / Q000295:instrumentation; D015222:Sodium Channels / Q000502:physiology*
D015222:Sodium Channels