Phase Separation In Vesicles Research Articles

Membrane nanodomains homeostasis during propofol anesthesia as function of dosage and temperature

Some anesthetics bind and potentiate γ-aminobutyric-acid-type receptors, but no universal mechanism for general anesthesia is known. Furthermore, often encountered complications such as anesthesia induced amnesia are not understood. General anesthetics are hydrophobic molecules easily dissolving into lipid bilayers. Recently, it was shown that general anesthetics perturb phase separation in vesicles extracted from fixed cells. Unclear is whether under physiological conditions general anesthetics induce perturbation of the lipid bilayer, and whether this contributes to the transient loss of consciousness or anesthesia side effects. Here we show that propofol perturbs lipid nanodomains in the outer and inner leaflet of the plasma membrane in intact cells, affecting membrane nanodomains in a concentration dependent manner: 1 μM to 5 μM propofol destabilize nanodomains; however, propofol concentrations higher than 5 μM stabilize nanodomains with time. Stabilization occurs only at physiological temperature and in intact cells. This process requires ARP2/3 mediated actin nucleation and Myosin II activity. The rate of nanodomain stabilization is potentiated by GABAA receptor activity. Our results show that active nanodomain homeostasis counteracts the initial disruption causing large changes in cortical actin. Significance statementGeneral anesthesia is a routine medical procedure with few complications, yet a small number of patients experience side-effects that persist for weeks and months. Very young children are at risk for effects on brain development. Elderly patients often exhibit subsequent amnesia. Here, we show that the general anesthetic propofol perturbs the ultrastructure of the lipid bilayer of the cell membrane in intact cells. Initially propofol destabilized lipid nanodomains. However, with increasing incubation time and propofol concentration, the effect is reversed and nanodomains are further stabilized. We show that this stabilization is caused by the activation of the actin cortex under the membrane. These perturbations of membrane bilayer and cortical actin may explain how propofol affects neuronal plasticity at synapses.

Budding, fission and domain formation in mixed lipid vesicles induced by lateral phase separation and macromolecular condensation.

Phase separation in vesicles of lipid mixtures creates an entirely new variety of shapes. The process may be described in terms of a combination of the theory of spinodal decomposition and the membrane bending energy concept. One major aspect discussed is the stabilization of domain structures in intrinsically unstable (phase segregated) states by the coupling between local bending and phase separation. A second topic is the role of interfacial tension for the stability of the domains. We show that vesicle fission is a consequence (1) of the reduction of the interfacial chemical energy or (2) of local phase separation within the neck connecting bud and mother vesicle. Finally, the formation of coated buds in giant vesicles caused by the lateral condensation of membrane-bound macromolecular amphiphiles is described.

Combined electron-spin-resonance, X-ray-diffraction studies on phospholipid vesicles obtained from cold-hardened wheats

The contents of free sterols and phospholipids in leaves of wheat, Triticum aestivum L., cultivars of different frost resistances, as well as the physical state of isolated phospholipids in the presence and absence of sterols, were compared before and after hardening. There was an inverse relationship between the sterol/phospholipid ratio and frost tolerance as a consequence of both a decrease in the free sterol, and an increase in the total phospholipid content. Sterol-sterol interactions were investigated using wide angle X-ray diffraction, while the phase behaviour of phospholipid vesicles was studied using the electron-spin-resonance (ESR) technique. No sterol-sterol interactions at-10° C were detected in vesicles obtained from the hardened most cold-tolerant cultivar (Miranovskaja 808), containing sterols in a ratio (0.08) found in the original lipid extracts. In contrast, when the sterol-phospholipid ratio in the vesicles was set to the level (0.39) found in the extracts of the most sensitive cultivar, Penjamo 62, the appearance of sharp reflexion rings at 4.5·10(-1), 4.8·10(-1) and 5.0·10(-1) nm indicated strong sterol-sterol interactions. The temperatures for the onset of phase separation for vesicles of identical sterol/phospholipid ratios found in lipid extracts of hardened Miranovskaja 808 were almost the same as those measured in purified phospholipids (-15 vs.-16° C). In contrast, the temperature for the onset of phase separation of vesicles with a sterol/phospholipid ratio characteristic of hardened Penjamo 62 was shifted upwards (from-6 to-2° C). Phase separation was not completed in the vesicles of Miranovskaja 808 in the temperature range scanned (-30° C) but was shifted from-22 to-18° C in the presence of sterols in the case of Penjamo 62. The results are discussed in terms of the composition and physical state of membranes in relation to survival at freezing temperatures.

Micromolar concentrations of Al 3+ induce phase separation, aggregation and dye release in phosphatidylserine-containing lipid vesicles

The interaction of Al 3+, Cd 2+ and Mn 2+ with phosphatidylserine-containing lipid vesicles was studied. Phase separation of vesicles was investigated by monitoring fluorescence quenching of the phospholipid analogue 1-palmitoyl-2-(6-[ N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)]aminocaproyl)phosphatidylcholine (C 6-NBD-PC). Aggregation was determined by turbidimetry and leakage of vesicles content during fusion was monitored by the fluorescence of released 6-carboxyfluorescein. Al 3+ demonstrated quenching at less than 30 μmol/1 with a maximum effect at 100 μmol/1. Al 3+-induced aggregation and dye release from the lipid vesicles were observed in the same concentration range. The effect of Cd 2+ and Mn 2+ on quenching was much less pronounced and could only be demonstrated in the 0.1–1 mmol/1 range. Increasing amounts of phosphatidylcholine or phosphatidylethanolamine in the vesicles decreased both Al 3+-induced quenching and aggregation, wheras cholesterol only slightly increased aggregation without affecting quenching.