Giant membrane vesicles as a model to study cellular substrate uptake dissected from metabolism.
In order to use giant vesicles for substrate uptake studies in metabolically important tissues, we characterized giant vesicles isolated from heart, liver, skeletal muscle and adipose tissue. We investigated which cell types and which plasma membrane regions are involved in giant vesicle formation and we examined the presence of transporters for metabolic substrates. Analysis of giant vesicles with markers specific for distinct cell types and distinct domains of the plasma membrane reveals that the plasma membrane of parenchymal cells, but not endothelial cells, are the source of the vesicle membranes. In addition, plasma membrane regions enriched in caveolae and involved in docking of recycling vesicles from the endosomal compartment are retained in giant vesicles, indicating that KCl-induced alterations in recycling processes are involved in giant vesicle formation. Giant vesicles contain vesicular lumen consisting of the soluble constituents of the cytoplasm including, fatty-acid binding proteins. Furthermore, giant vesicles isolated from heart, liver, skeletal muscle and adipose tissue are similar in size (10-15 microm) and shape and do not contain subcellular organelles, providing the advantage that substrate fluxes in the different organs can be studied independently of the surface/volume ratio but most importantly in the absence of intracellular metabolism.
Koonen
D P Y
DP
Department of Physiology, Cardiovascular Research Institute Maastricht, Maastricht University, Maastricht, the Netherlands. d.koonen@fys.unimaas.nl
Coumans
W A
WA
Arumugam
Y
Y
Bonen
A
A
Glatz
J F C
JF
Luiken
J J F P
JJ
D000273:Adipose Tissue / Q000378:metabolism / Q000648:ultrastructure; D000818:Animals; D018955:CD36 Antigens; D002352:Carrier Proteins / Q000378:metabolism; D022461:Caveolins / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D051275:Glucose Transporter Type 4; D008099:Liver / Q000378:metabolism / Q000648:ultrastructure; D008562:Membrane Glycoproteins / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D009124:Muscle Proteins; D018482:Muscle, Skeletal / Q000648:ultrastructure; D009206:Myocardium / Q000378:metabolism / Q000648:ultrastructure; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D027361:Organic Anion Transporters / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D022161:Transport Vesicles / Q000737:chemistry / Q000378:metabolism*
D018955:CD36 Antigens; D002352:Carrier Proteins; D022461:Caveolins; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids; D051275:Glucose Transporter Type 4; D008562:Membrane Glycoproteins; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D027361:Organic Anion Transporters; C495694:Slc2a4 protein, rat; D005947:Glucose