18] Cut-open recording techniques

Abstract

Publisher Summary This chapter describes a new technique for voltage-clamp recording of Xenopus oocytes that enables fast and stable monitoring of transmembrane current, as well as intracellular perfusion by cutting a part of the plasma membrane to open the cytoplasm to the recording chamber, which is filled with an artificial intracellular solution. The original method was developed and described by E. Stefani and colleagues, and this chapter has made a modification by exchanging intracellular solution using a push-pull cannula. This technique has certain characteristic features. High-frequency response and relatively low current noise is the first feature. Fast activation and deactivation of ionic currents can be precisely evaluated. Stable recording conditions that lasts for several hours is a marked advantage over the conventional and patch-clamp recording. Control of the ionic composition of both the internal and external media is another feature. Although it is possible to study the action of compounds on the internal surface of oocytes using inside-out patches or by injecting drugs under a whole-cell clamp, the cut-open configuration enables the desired manipulation of the internal milieu of oocytes easily. These features allow reliable measurements of gating currents and fast ionic currents of Shaker K + channels in single and unsubtracted traces. Moreover, the easy access to the intracellular milieu is suitable for the study of channel modulation by second messengers and drugs. This method has been applied for the studies of transient K + channels, rapid delayed rectifier K + channels, voltage-dependent Ca 2+ channels, Na + -glucose co-transporters, and Na + -independent amino acid transporters.