Single-channel dose-response studies in single, cell-attached patches

Abstract

A method for carrying out dose-response studies of ion channel currents in cell-attached patches has been devised. Patch pipettes are filled at the tip with a solution containing one concentration of ligand and then backfilled with another. The concentration of ligand at the membrane is described as a function of time by the equation for diffusion in a cone, allowing response vs. time data to be transformed into a dose-response curve. For Xenopus myocyte cholinergic receptors, examples of the use of this method are given for several concentration-dependent reactions including blockade by the local anesthetic QX-222, activation by acetylcholine, and modulation of current amplitude by sodium ions. Several methods of analyzing the nonstationary channel kinetics are presented, including a pseudo-stationary approach that uses interval likelihood maximization.