Pipet Perfusion Methods: Techniques and Applications

The patch-clamp method was introduced in 1976 (Hamill et al., 1981; Neher and Sakmann, 1976). It has been widely used to study whole-cell and single-channel currents. In many cases, the experimenter needs to test how changing the pipet solution composition can affect membrane-channel function (see Subheading 2. ) Applications of the Pipet Perfusion Technique). This can be accomplished by applying pipets containing different solutions to different cells or to different membrane patches. The effects are inferred by comparing the properties of currents recorded under these different conditions. Problems in data interpretation may arise because of cell-to-cell variations in cell membrane-channel properties. This is especially the case for heterogeneous systems such as the central nervous system (CNS). An alternative approach is to excise the membrane patch to form inside-out or outside-out patch recording configuration. In this way, the experimenter can perfuse the bath with various solutions, thus controlling the milieu facing the intracellular (inside-out patch) or extracellular (outside-out patch) surface of the cell membrane. Potential problems here are channel density (should be high enough to allow a reasonable chance of recording channel currents from a patch), and the stability of channel function in cell-free patches.