Ion trapping with fast-response ion-selective microelectrodes enhances detection of extracellular ion channel gradients.
Previously, functional mapping of channels has been achieved by measuring the passage of net charge and of specific ions with electrophysiological and intracellular fluorescence imaging techniques. However, functional mapping of ion channels using extracellular ion-selective microelectrodes has distinct advantages over the former methods. We have developed this method through measurement of extracellular K+ gradients caused by efflux through Ca2+-activated K+ channels expressed in Chinese hamster ovary cells. We report that electrodes constructed with short columns of a mechanically stable K+-selective liquid membrane respond quickly and measure changes in local [K+] consistent with a diffusion model. When used in close proximity to the plasma membrane (<4 microm), the ISMs pose a barrier to simple diffusion, creating an ion trap. The ion trap amplifies the local change in [K+] without dramatically changing the rise or fall time of the [K+] profile. Measurement of extracellular K+ gradients from activated rSlo channels shows that rapid events, 10-55 ms, can be characterized. This method provides a noninvasive means for functional mapping of channel location and density as well as for characterizing the properties of ion channels in the plasma membrane.
Messerli
Mark A
MA
BioCurrents Research Center, Cell Dynamics Program, Marine Biological Laboratory, Woods Hole, Massachusetts 02543, USA. mmesserli@mbl.edu
Collis
Leon P
LP
Smith
Peter J S
PJ
D000818:Animals; D016466:CHO Cells; D002462:Cell Membrane / Q000502:physiology; D003198:Computer Simulation; D006224:Cricetinae; D003412:Cricetulus; D004058:Diffusion; D005110:Extracellular Space / Q000737:chemistry; D015640:Ion Channel Gating / Q000502:physiology*; D008839:Microelectrodes; D009010:Monte Carlo Method; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000032:analysis / Q000378:metabolism*; D024681:Potassium Channels, Calcium-Activated / Q000502:physiology*
D024681:Potassium Channels, Calcium-Activated; D011188:Potassium