Rapid transmembrane diffusion of ceramide and dihydroceramide spin-labelled analogues in the liquid ordered phase

In order to study the basic physical phenomena underlying complex lipid transbilayer movement in biological membranes, we have measured the transmembrane diffusion of spin-labelled analogues of sphingolipids in phosphatidylcholine (PC) large unilamellar vesicles in the absence or presence of cholesterol, going from a fluid ( liquid disordered) l(d), phase to a more viscous, liquid ordered (l(o)), phase. We have found cholesterol to reduce the transverse diffusion of glucosylceramide (GlcCer) and galactosylceramide (GalCer) in a concentration-dependent manner. However, surprisingly, we could neither detect any influence of cholesterol on the rapid flip-flop of ceramide nor on the flip-flop of dihydroceramide, for which the tau(1/2) of flip-flop remains in the order of 1 minute at 20 degrees C in the presence of cholesterol. As a consequence of rapid flip-flop of ceramide in both the l(o) and the l(d) phase, ceramide is likely to distribute between the two monolayers of a membrane, and could in principle partition into segregated domains in each side of the plasma membrane of eukaryotic cells.