Measuring ion channels on solid supported membranes.
Application of solid supported membranes (SSMs) for the functional investigation of ion channels is presented. SSM-based electrophysiology, which has been introduced previously for the investigation of active transport systems, is expanded for the analysis of ion channels. Membranes or liposomes containing ion channels are adsorbed to an SSM and a concentration gradient of a permeant ion is applied. Transient currents representing ion channel transport activity are recorded via capacitive coupling. We demonstrate the application of the technique to liposomes reconstituted with the peptide cation channel gramicidin, vesicles from native tissue containing the nicotinic acetylcholine receptor, and membranes from a recombinant cell line expressing the ionotropic P2X2 receptor. It is shown that stable ion gradients, both inside as well as outside directed, can be applied and currents are recorded with an excellent signal/noise ratio. For the nicotinic acetylcholine receptor and the P2X2 receptor excellent assay quality factors of Z' = 0.55 and Z' = 0.67, respectively, are obtained. This technique opens up new possibilities in cases where conventional electrophysiology fails like the functional characterization of ion channels from intracellular compartments. It also allows for robust fully automatic assays for drug screening.
Schulz
Patrick
P
Max Planck Institut für Biophysik, D-60438 Frankfurt/Main, Germany.
Dueck
Benjamin
B
Mourot
Alexandre
A
Hatahet
Lina
L
Fendler
Klaus
K
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D004557:Electric Organ / Q000187:drug effects / Q000502:physiology; D004594:Electrophysiology / Q000379:methods*; D006096:Gramicidin / Q000737:chemistry; D006801:Humans; D007473:Ion Channels / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008566:Membranes / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D018733:Nicotinic Antagonists / Q000494:pharmacology; D058919:Purinergic P2 Receptor Antagonists; D051381:Rats; D011978:Receptors, Nicotinic / Q000737:chemistry / Q000378:metabolism; D018048:Receptors, Purinergic P2 / Q000737:chemistry / Q000378:metabolism; D058476:Receptors, Purinergic P2X2; D012964:Sodium / Q000378:metabolism; D014101:Torpedo
D007473:Ion Channels; D008081:Liposomes; D018733:Nicotinic Antagonists; C546401:P2RX2 protein, human; D058919:Purinergic P2 Receptor Antagonists; D011978:Receptors, Nicotinic; D018048:Receptors, Purinergic P2; D058476:Receptors, Purinergic P2X2; D006096:Gramicidin; D000255:Adenosine Triphosphate; D012964:Sodium