Cell-Free Ion-Channel Recording

When membrane patches are isolated from cells, one gains the ability to regulate precisely the composition of the solution bathing both surfaces of the membrane and to change the composition rapidly. However, in detaching the membrane from the underlying cytoskeleton, one irreversibly changes the integrated system of which the channel protein is a part, and begins an inexorable loss of channel function. In this chapter we will deal primarily with the techniques available for measuring the response of patch currents to changes in bathing solution and the practical problems to be minimized, or overcome, in implementing such techniques and analyzing the results obtained. Other chapters will deal with patch-clamping in general. Chapter 6 deals specifically with methods of changing the concentration of the solution bathing the intrapipet face of the membrane, and we will concentrate on consideration of the extrapipet solution changes, i.e., changes at the intracellular membrane surface in inside-out patches, or the external face in outside-out membrane patches.