Asymmetric pore distribution and loss of membrane lipid in electroporated DOPC vesicles.
An externally applied electric field across vesicles leads to transient perforation of the membrane. The distribution and lifetime of these pores was examined using 1,2-di-oleoyl-sn-glycero-3-phosphocholine (DOPC) phospholipid vesicles using a standard fluorescent microscope. The vesicle membrane was stained with a fluorescent membrane dye, and upon field application, a single membrane pore as large as approximately 7 microm in diameter was observed at the vesicle membrane facing the negative electrode. At the anode-facing hemisphere, large and visible pores are seldom found, but formation of many small pores is implicated by the data. Analysis of pre- and post-field fluorescent vesicle images, as well as images from negatively stained electron micrographs, indicate that pore formation is associated with a partial loss of the phospholipid bilayer from the vesicle membrane. Up to approximately 14% of the membrane surface could be lost due to pore formation. Interestingly, despite a clear difference in the size distribution of the pores observed, the effective porous areas at both hemispheres was approximately equal. Ca(2+) influx measurements into perforated vesicles further showed that pores are essentially resealed within approximately 165 ms after the pulse. The pore distribution found in this study is in line with an earlier hypothesis (E. Tekle, R. D. Astumian, and P. B. Chock, 1994, Proc. Natl. Acad. Sci. U.S.A. 91:11512--11516) of asymmetric pore distribution based on selective transport of various fluorescent markers across electroporated membranes.
Tekle
E
E
Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA. ephrem@helix.nih.gov
Astumian
R D
RD
Friauf
W A
WA
Chock
P B
PB
D002463:Cell Membrane Permeability; D018274:Electroporation; D005456:Fluorescent Dyes / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry* / Q000378:metabolism*; D008561:Membrane Fusion; D008563:Membrane Lipids / Q000378:metabolism*; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D010713:Phosphatidylcholines / Q000378:metabolism*
D005456:Fluorescent Dyes; D008081:Liposomes; D008563:Membrane Lipids; D010713:Phosphatidylcholines; C017251:1,2-oleoylphosphatidylcholine