Membrane lipid domains and dynamics as detected by Laurdan fluorescence

Abstract

(Laurdan) is a membrane probe of recent characterization, which shows high sensitivity to the polarity of its environment. Steady-state Laurdan excitation and emission spectra have different maxima and shape in the two phospholipid phases, due to differences in the polarity and in the amount of dipolar relaxation, ha bilayers composed of a mixture of gel and liquid-crystalline phases, the properties of Laurdan excitation and emission spectra are intermediate between those obtained in the pure phases. These spectral properties are analyzed using the generalized polarization (GP). The GP value can be used for the quantitation of each phase. The wavelength dependence of the GP value is used to ascertain the coexistence of different phase domains in the bilayer. Moreover, by following the evolution of Laurdan emission vs. time after excitation, the kinetics of phase fluctuation in phospholipid vesicles composed of coexisting gel and liquid-crystalline phases was determined. GP measurements performed in several cell lines did not give indications of coexistence of phase domains in their membranes. In natural membranes, Laurdan parameters indicate a homogeneously fluid environment, with restricted molecular motion in comparison with the phospholipid liquid-crystalline phase. The influence of cholesterol on the phase properties of the two phospholipid phases is proposed to be the cause of the phase behavior observed in na~u'al membranes. In bilayers composed of different phospholipids and various cholesterol concentrations, Laurdan response is very similar to that arising from cell membranes. In the absence of cholesterol, from the steady-state and time-resolved measurements of Laurdan in phospholipid vesicles, the condition for the occurrence of separate coexisting domains in the bilayer has been determined: the molecular ratio between the two phases must be in the range between 30% and 70%. Below and above this range, a single homogeneous phase is observed, with the properties of the more concentrated phase, slightly modified by the presence of the other. Moreover, in this concentration range, the calculated dimension of the domains is very small, between 20 and 50 A.

Order of lipid phases in model and plasma membranes

“…Moreover, GP s values were not discrete for the phase states but changed with lipid composition, decreasing in value from Lo phases to coexisting Lo-Ld phases and Ld phases (Fig. 1B and SI Text) (38). Notably, high amounts of the LW TM peptide affected membrane order only marginally in the composition tested (Fig.…”

Use Liposome as a Designable Platform for Molecular Recognition ~ from “Statistical Separation” to “Recognitive Separation” ~

“…: hexagonal periodical; ripple-gel phase [17], ESR [18], molecular dynamics (Monte Carlo method) [23], and fluorescent probes [24], as summarized in Table 2. "Micro-"domains are known to be observed by using FRET, although there have been few reports about "nano-"sized domains (<5 nm) formed in the lipid membranes.…”

Deoxycholic acid modulates cell death signaling through changes in mitochondrial membrane properties

“…Both di-4-ANEPPDHQ ( 31,32 ) and Laurdan ( 33,34 ) have proved to be extremely sensitive to changes in membrane order through changes in their fl uorescence emission spectra or lifetimes. Both molecules sense the reorientation of solvent dipoles, whose strength is directly dependent on membrane fl uidity, this being related to water penetration within membranes ( 31,33 ). The measurement of shifts in Laurdan emission spectra through ratiometric imaging ( 25 ) and changes of di-4-ANEPPDHQ fl uorescence lifetime through FLIM ( 32 ) are widely used methodologies that allow for a quantitative assessment of membrane order in living cells.…”