Two-microelectrode voltage clamp of Xenopus oocytes: Voltage errors and compensation for local current flow

Oocytes from Xenopus laevis are commonly used as an expression system for i on channel proteins. The most common method for their electrophysiological investigation is the two-microelectrode voltage clamp technique. The qualit y of voltage clamp recordings obtained with this technique is poor when mem brane currents are large and when rapid charging of the membrane is desired . Detailed mathematical modeling of the experimental setup shows that the r easons for this weak performance are the electrical properties of the oocyt es and the geometry of the setup. We measured the cytosolic conductivity to be similar to 5 times lower than that of the typical bath solution, and th e specific membrane capacitance to be similar to 6 times higher than that o f a simple lipid bilayer. The diameter of oocytes is typically similar to 1 mm, whereas the penetration depth of the microelectrodes is limited to sim ilar to 100 mu m. This eccentric current injection, in combination with the large time constants caused by the low conductivity and the high capacitan ce, yields large deviations from isopotentiality that decay slowly with tim e constants of up to 150 mu s. 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 a re optimized and initial experiments show that this method should allow for faster and more uniform control of membrane potential.