IMPROVED TECHNIQUE FOR STUDYING ION CHANNELS EXPRESSED IN XENOPUS-OOCYTES, INCLUDING FAST SUPERFUSION

The study of whole-cell currents from ion channels expressed in Xenopu s oocytes with conventional two-electrode Voltage clamp has two major limitations. First, the large diameter and spherical geometry of oocyt es prevent extremely fast solution changes. Second, the internal mediu m is not controlled, which limits the experimental versatility of the oocyte expression system. For example, because the internal medium is not controlled, endogenous calcium-activated chloride conductances can contaminate currents measured with channels that are permeable to cal cium. We describe a new technique that combines vaseline-gap voltage c lamp for oocytes with a fast superfusion system. The vaseline-gap proc edure is simplified by having the micropipette that monitors voltage s erve a dual role as a perfusion micropipette that controls the interna l solution. In addition, the technique provides fast external solution changes that are complete in 30-50 ms. We applied the approach to mea sure the calcium permeability of a muscle and a neuronal nicotinic ace tylchotine receptor. Very fast agonist induced currents were measured without contamination by the secondary activation of calcium-dependent chloride channels.