Fusion of native or reconstituted membranes to liposomes, optimized for single channel recording.
We here describe a protocol for fusing vesicles into large structures suitable for patch clamp recording. The method may be used with native membrane vesicles or with liposomes containing reconstituted/purified ion channels. The resulting unilamellar membranes exhibit high channel surface abundance, yielding multiple channels in the average excised patch. The procedure has been used to record voltage-sensitive Na channels from three native membrane preparations (eel electroplax, rat skeletal muscle, squid optic nerve), and from reconstituted protein purified from eel electroplax. Channels treated with batrachotoxin (BTX) displayed characteristic activation voltage dependence, conductances, selectivity, and sensitivity to saxitoxin (STX).
Correa
A M
AM
Department of Cellular and Molecular Physiology, Yale University, School of Medicine, New Haven, Connecticut 06510.
Agnew
W S
WS
D000818:Animals; D001496:Batrachotoxins / Q000494:pharmacology; D002462:Cell Membrane / Q000502:physiology; D008081:Liposomes; D008564:Membrane Potentials; D008954:Models, Biological; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010718:Phosphatidylserines; D012530:Saxitoxin / Q000494:pharmacology; D015222:Sodium Channels / Q000187:drug effects / Q000502:physiology*
D001496:Batrachotoxins; D008081:Liposomes; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010718:Phosphatidylserines; D015222:Sodium Channels; D012530:Saxitoxin