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Biophys J, October 1999, p. 1980-1991, Vol. 77, No. 4
*Institute for Biophysics, University of Linz, A-4040 Linz, Austria; #Institute of Anatomy, University of Würzburg, D-97070 Würzburg, Germany; and §Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06520 USA
Oocytes from Xenopus laevis are commonly used
as an expression system for ion channel proteins. The most common
method for their electrophysiological investigation is the
two-microelectrode voltage clamp technique. The quality of
voltage clamp recordings obtained with this technique is poor when
membrane currents are large and when rapid charging of the membrane is
desired. Detailed mathematical modeling of the experimental setup shows
that the reasons for this weak performance are the electrical
properties of the oocytes and the geometry of the setup. We measured
the cytosolic conductivity to be ~5 times lower than that of the
typical bath solution, and the specific membrane capacitance to be ~6 times higher than that of a simple lipid bilayer. The diameter of
oocytes is typically ~1 mm, whereas the penetration depth of the
microelectrodes is limited to ~100 µm. This eccentric current injection, in combination with the large time constants caused by the
low conductivity and the high capacitance, yields large deviations from
isopotentiality that decay slowly with time constants of up to 150 µs. The inhomogeneity of the membrane potential can be greatly
reduced by introducing an additional, extracellular current-passing
electrode. The geometrical and electrical parameters of the setup are
optimized and initial experiments show that this method should allow
for faster and more uniform control of membrane potential.
Biophys J, October 1999, p. 1980-1991, Vol. 77, No. 4
© 1999 by the Biophysical Society 0006-3495/99/10/1980/12 $2.00
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