Acs. Costa et al., IMPROVED TECHNIQUE FOR STUDYING ION CHANNELS EXPRESSED IN XENOPUS-OOCYTES, INCLUDING FAST SUPERFUSION, Biophysical journal, 67(1), 1994, pp. 395-401
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.