Use of Xenopus oocytes to measure ionic selectivity of pore-forming peptides and ion channels.
The Xenopus laevis oocyte is a widely used system for heterologous expression of exogenous ion channel proteins (1, 2). Among other advantages, these easy to obtain, mechanically and electrically stable, large-sized cells enable multiple types of electrophysiological recordings: two-electrode voltage-clamp, single-cell attached or cell-free patch-clamp, and macropatch recordings. The size of an oocyte (1 mm in diameter) also allows the use of additional electrodes (1-3) for injection of diverse materials (Ca2+ chelators, peptides, chemicals, antibodies, proteic-partners, and so on) before or during the course of the electrophysiological experiment. We have successfully used this system to analyze the biophysical properties of pore-forming peptides. Simple perfusion of these peptides induced the formation of channels in the oocyte plasma membrane; these channels can then be studied and characterized in diverse ionic conditions. The ease of the perfusion and the stability of the voltage-clamped oocyte make it a powerful tool for such analyses. Compared with artificial bilayers, oocytes offer a real animal plasma membrane where biophysical properties and toxicity can be studied in the same environment.
Cens
Thierry
T
CRBM, CNRS, France.
Charnet
Pierre
P
D000818:Animals; D015220:Calcium Channels / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives; D007267:Injections; D015640:Ion Channel Gating; D007473:Ion Channels / Q000378:metabolism*; D008564:Membrane Potentials; D009865:Oocytes / Q000502:physiology*; D018408:Patch-Clamp Techniques / Q000379:methods*; D010455:Peptides / Q000378:metabolism*; D012996:Solutions; D013816:Thermodynamics; D014982:Xenopus laevis / Q000502:physiology*
D015220:Calcium Channels; D007473:Ion Channels; D010455:Peptides; D012996:Solutions; D004533:Egtazic Acid; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid