Expression of Ion Channels in Xenopus Oocytes

Xenopus oocytes have been widely used for studying ion channels in a controlled in vivo environment since the system was initially developed for this purpose by Miledi and coworkers [1, 2]. There have been at least five major types of studies using oocytes to examine ion channel expression. The earliest use was to examine the properties of specific ion channels in a living cell free from other responses. The oocytes were injected with RNA isolated from whole brains, and the responses were analyzed using the two-microelectrode whole cell voltageclamp [2, 3], the patch clamp [4], or a variety of biochemical techniques [5, 6]. Once the responses were isolated, Xenopus oocytes were then used in a second type of study as an assay system to isolate cDNA clones encoding the proteins involved. For example, cDNA clones encoding the 5-HT1C receptor were isolated using electrophysiological assays, both by hybrid depletion [7] and by directly transcribing RNA from a cDNA library and injecting the transcripts into oocytes [8]. These types of studies are much less commonly used now because of the large number of available heterologous expression systems and cDNA clones encoding ion channels. The third major type of study for which the Xenopus oocyte expression system has been, and continues to be, particularly useful is the correlation of molecular structure with electrophysiological function of a specific channel. The two basic approaches have been to construct defined mutations whose effects are determined by expression in oocytes, and to construct chimeric molecules between two closely related channels or receptors followed by expression in oocytes and electrophysiological analysis. These types of approach are still commonly used but with more sophisticated structural alterations and functional analyses. The fourth general approach utilizing expression in oocytes is to determine the functional effects of mutations that cause human diseases. These types of studies are also performed using expression in other heterologous systems such as mammalian cells, and the advantages and disadvantages of each approach will be discussed in Section 1.5.3. 1