Dynamic fluorescence measurements on the main phase transition of dipalmytoylphosphatidylcholine vesicles

Genz, A.; Holzwarth, Josef F.

doi:10.1007/bf00265668

Cited by 46 publications

(40 citation statements)

References 22 publications

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“…It is apparent that this equation applies equally well to diffusive water permeability in bilayers made asymmetric with CS and to osmotic water permeability in . Previous studies had provided evidence that individual leaflets of membrane bilayers could alter their physical properties independent of the other leaflet (23,24). Our results demonstrate for the first time that individual leaflets of the membrane bilayer can independently regulate permeation.…”

Section: And Discussionsupporting

confidence: 60%

“…To determine whether, in our experiments, Pr 3ϩ was increasing the phase transition temperature of the outer leaflet, we monitored fluorescence anisotropy as an estimate of membrane fluidity. To examine the fluidity of the outer leaflet, we introduced a phospholipid-bound probe of anisotropy, 2-(3-(diphenylhexatrienyl)propanyl)-1-hexadecanoyl-snglycero-3-phosphocholine (DPH HPC) into the outer leaflet, and monitored fluorescence anisotropy as a function of temperature (24,25). Because phospholipids added to the outside of the liposomes enter the outer leaflet rapidly but cross to the inner leaflet slowly, this probe monitored exclusively the outer leaflet of the bilayer.…”

Section: And Discussionmentioning

confidence: 99%

“…A, fluidity of the outer leaflet was monitored using DPH HPC (excitation 360 nm, emission 430 nm) at a probe:lipid ratio of 1:400 at varying temperatures in the absence (filled circles) and presence (filled squares) of 10 mM extravesicular Pr 3ϩ on a SPEX spectrofluorimeter using standard methods (24,25). B, measurements of water flux at 44°C in the absence (DPPC) and presence (DPPC/Pr 3ϩ ) of 10 mM extravesicular Pr 3ϩ .…”

Section: Fig 1 Effect Of External Prmentioning

confidence: 99%

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Individual Leaflets of a Membrane Bilayer Can Independently Regulate Permeability

Negrete

Rivers

Gough

et al. 1996

Journal of Biological Chemistry

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Water rapidly crosses most membranes, but only slowly crosses apical membranes of barrier epithelia such as bladder and kidney collecting duct, a feature essential to barrier function. How apical membrane structure reduces permeabilities remains unclear. Cell plasma membranes contain two leaflets of distinct lipid composition; the role of this bilayer asymmetry in membrane permeability is unclear. To determine how asymmetry of leaflet composition affects membrane permeability, effects on bilayer permeation of reducing single leaflet permeability were determined using two approaches: formation of asymmetric bilayers in an Ussing chamber, with only one of two leaflets containing cholesterol sulfate, and stabilization of the external leaflet of unilamellar vesicles with praeseodymium (Pr 3؉ ). In both systems, permeability measurements showed that each leaflet acts as an independent resistor of water permeation. These results show that a single bilayer leaflet can act as the barrier to permeation and provide direct evidence that segregation of lipids to create a low permeability exofacial leaflet may act to reduce the permeability of barrier epithelial apical membranes.Epithelial cells generate and maintain apical membrane bilayers made up of leaflets of distinct composition by mechanisms involving asymmetric biosynthesis in the Golgi, oriented insertion into the plasma membrane, and the activity of ATPdriven phospholipid flippases (1-6). In several epithelia, such as those of the stomach, kidney collecting duct and thick ascending limb, and mammalian bladder, these apical membranes exhibit exceptionally low permeabilities to water, small nonelectrolytes, protons, and ammonia. These low permeabilities are critical to the barrier function of these epithelia (7-11). The structural features responsible for the low permeabilities of these apical membranes remain poorly defined as is the physiological role of the bilayer asymmetry observed in these membranes. The present studies examine the role of bilayer asymmetry in governing the permeability of membranes to water by measuring permeabilities across artificial symmetric and asymmetric membranes. The results show that a single leaflet of a membrane bilayer can act as a barrier to water flux and provide strong evidence that each leaflet acts as an independent resistor to permeation.

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Section: And Discussionsupporting

confidence: 60%

Section: And Discussionmentioning

confidence: 99%

Section: Fig 1 Effect Of External Prmentioning

confidence: 99%

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Individual Leaflets of a Membrane Bilayer Can Independently Regulate Permeability

Negrete

Rivers

Gough

et al. 1996

Journal of Biological Chemistry

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“…In other studies relaxation times faster than the ones we measured and up to five relaxation processes were reported [24,25,26,28,29,62,41,38,39]. Different spectroscopic detection methods were used.…”

Section: Discussionmentioning

confidence: 88%

How Anesthetics, Neurotransmitters, and Antibiotics Influence the Relaxation Processes in Lipid Membranes

2007

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We used pressure perturbation calorimetry to investigate the relaxation time scale after a jump into the melting transition regime of artificial lipid membranes. This time is equivalent to the characteristic rate of domain growth. The studies were performed on single-component large unilamellar and multilamellar vesicle systems with and without the addition of small molecules such as general anesthetics, neurotransmitters, and antibiotics. These drugs interact with membranes and affect melting points and profiles. In all systems, we found that heat capacity and relaxation times are linearly related to each other in a simple manner, and we outline the theoretical origin of this finding. Thus, the influence of a drug on the time scale of domain formation processes can be understood on the basis of their influence on the heat capacity profile. This allows estimations of the characteristic relaxation time scales in biological membranes.

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“…The increase in fluorescence lifetime seems to be slightly less pronounced for DPPC containing~-sitosterol. It is well known from a variety of measurements employing different physical techniques, that complex formation and phase segregation of cholesterol occurs at high concentrations of cholesterol, and that at a concentration of about 50 mol-OJo cholesterol, the gel to liquid-crystalline phase transition is suppressed [5,6,[40][41][42][43][44][45][46].…”

Section: Resultsmentioning

confidence: 99%

Influence of Cholesterol and β‐Sitosterol on the Dynamic Behaviour of DPPC as Detected by TMA‐DPH and PyrPC Fluorescence: A Fluorescence Lifetime Distribution and Time‐Resolved Anisotropy Study

Bernsdorff

Winter

Hazlett

et al. 1995

Ber Bunsenges Phys Chem

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Fluorescence lifetime and time‐resolved anisotropy measurements on 1‐(4‐trimethylammonium‐phenyl)‐6‐phenyl‐1,3,5‐hexatriene (TMA‐DPH) and 1‐palmitoyl‐2‐[10‐(1‐pyrenyl)decanoyl]‐phosphatidylcholine (PyrPC) in lipid vesicles of 1,2‐dipalmitoyl‐sn‐glycero‐3‐phosphatidylcholine (DPPC) with and without cholesterol and β‐sitosterol, using multifrequency cross‐correlation phase fluorometry, are presented. We used TMA‐DPH and PyrPC as fluorescence probe for detecting rotational dynamics. Contrary to TMA‐DPH, which is tethered to the bilayer interface and therefore detects wobbling rotational dynamics in the interfacial and headgroup region, PyrPC senses the deeper acyl chain dynamics of the lipid bilayer. We have investigated the influence of sterol structure on the lipid dynamics. The experiments were carried out at two temperatures, one below and one above the main phase transition temperature of DPPC. Data were analyzed in terms of Lorentzian distribution functions for the fluorescence lifetime results and in terms of hindered rotation for the time‐resolved anisotropy results. We noticed, that addition of cholesterol and β‐sitosterol results in a distinct increase in average fluorescence lifetime of TMA‐DPH in both lipid phases. The rotational rates of TMA‐DPH depolarizing motions are only modestly increased in going from the gel to the liquid‐crystalline phase of DPPC. However, a marked increase in the contribution of fast rotational motions of TMA‐DPH is found as the temperature is increased through the main transition. No significant differences in rotational rates of TMA‐DPH exist between samples with embedded cholesterol or β‐sitosterol at T = 55°C. The rotational rate of TMA‐DPH in the gel state of the lipid bilayer at T = 35°C is slightly larger in the case of β‐sitosterol, however. Cholesterol and β‐sitosterol, then, have a similar effect on the contribution of fast rotational dynamics in the interfacial region of the bilayer. It decreases about 40% upon addition of 50 mol‐% sterol. The incorporation of either sterol into DPPC vesicles leads to an increase in fluorescence lifetime of PyrPC in both lipid phases. The rotational rates of PyrPC in the bilayer are an order of magnitude smaller than those of TMA‐DPH. Also the contributions of fast rotational motions significantly differ for the two fluorophores. The motional freedom in the deeper acyl chain region of DPPC in its fluid‐like state at T = 55°C is scarcely affected by addition of either sterol. However, differences in the rotational rates of the two sterols are observed for high sterol concentrations.

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Dynamic fluorescence measurements on the main phase transition of dipalmytoylphosphatidylcholine vesicles

Cited by 46 publications

References 22 publications

Individual Leaflets of a Membrane Bilayer Can Independently Regulate Permeability

Individual Leaflets of a Membrane Bilayer Can Independently Regulate Permeability

How Anesthetics, Neurotransmitters, and Antibiotics Influence the Relaxation Processes in Lipid Membranes

Influence of Cholesterol and β‐Sitosterol on the Dynamic Behaviour of DPPC as Detected by TMA‐DPH and PyrPC Fluorescence: A Fluorescence Lifetime Distribution and Time‐Resolved Anisotropy Study

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