The influence of dolichols on fluidity of mouse synaptic plasma membranes

Dolichols are isoprenologues which constitute an important component of biological membranes. However, an understanding of the effects of dolichols on the organization and dynamics of biological membranes has not been forthcoming. The experiments reported here are aimed at understanding the effects of dolichols on the physical properties of mouse brain synaptic plasma membranes. The effect of dolichols incorporated into mouse brain synaptic plasma membranes on fluorescent and electron spin resonance probes sensing the hydrophobic core differed from that of probes reporting closer to the surface of membrane bilayers. Dolichols significantly (P less than 0.01) lowered the polarization, limiting anisotropy, and order parameter of diphenylhexatriene in synaptic plasma membranes and liposomes extracted from synaptic plasma membranes, without changing the rotational relaxation time. Similarly, dolichol increased the fluidity reported by 16-doxylstearic acid in synaptic plasma membranes or liposomes extracted from synaptic plasma membranes. In contrast, dolichols exerted no effect on those properties for trans-parinaric acid or 5-doxylstearic acid in synaptic plasma membranes or liposomes derived therefrom. Dolichols can dramatically alter the structure and dynamics of lipid motion in synaptic plasma membranes and these effects are dependent on the location of the probe in the membrane.     

A maturation-related differential phosphorylation of the plasma membrane proteins of the epididymal spermatozoa of the hamster by endogenous protein kinases

When the plasma membranes of caput and cauda epididymal spermatozoa of hamster were evaluated for their ability to undergo phosphorylation, a differential phosphorylation of the membrane proteins was observed. In the plasma membranes of the caput epididymal spermatozoa (immature), twelve proteins were phosphorylated (100, 76, 67, 65, 55, 52, 47, 42, 38, 32, 30, and 20 kD), whereas in the plasma membranes of cauda epididymal spermatozoa (mature), a differential phosphorylation pattern was observed with respect to the 94, 67, 52, and 47 kD proteins. The 94 kD protein was found to be phosphorylated and the 67 kD protein was found to be not phosphorylated in cauda spermatozoal plasma membrane (Cd SPM) in contrast to this protein in caput spermatozoal plasma membrane (Cpt SPM). The 52 and 47 kD proteins were also more intensely phosphorylated in Cd SPM than Cpt SPM. The 100 kilodalton protein, although present in both Cpt and Cd sperm plasma membranes, was found to be phosphorylated at the tyrosine residues only in the Cd SPM, as indicated by the Western blot using antiphosphotyrosine antibody. Further, a differential phosphorylation of the substrate proteins present in the Cpt and Cd SPM was seen when Mg²⁺ in the assay buffer was replaced by other divalent cations. For instance, Zn²⁺ stimulated the phosphorylation of an 85 kD protein in cauda SPM and not in the caput SPM and Ca²⁺ stimulated the phosphorylation of a 76 kD protein only in the cauda SPM. The phosphoproteins in both the plasma membranes were found to be phosphorylated predominantly at the tyrosine residue. The differential phosphorylation of a 100 kD protein at tyrosine in the Cd SPM (Western blot), which is absent in the immature Cpt SPM, also indicated that certain proteins in the hamster spermatozoa are phosphorylated in a maturation-specific manner. Mol. Reprod. Dev. 47:341–350, 1997. © 1997 Wiley-Liss, Inc.     

Characterization of the submitochondrial compartments: study of the site of synthesis of dolichol and dolichol-linked sugars

The fractionation of mitochondrial membranes on discontinuous sucrose gradient leads to the obtaining of free outer membranes, free inner membranes and two distinct membrane contact site populations characterized as follows. Only outer membrane contact sites and inner membrane contact sites bind hexokinase. Outer membranes and outer membrane contact sites are cholesterol-rich fractions. The endogenous dolichol content is twice fold higher in outer membranes and outer membrane contact sites than in inner membranes and inner membrane contact sites, only the biosynthesis of dolichol in inner membrane contact sites is not stimulated by addition of exogenous [14C]-IPP and FPP. The glycosylation of endogenous dolichol from labeled nucleotide-sugars (UDP-GlcNAc, GDP-Man and UDP-Glc) leads to the synthesis of dolichol-pyrophosphoryl-sugars and dolichol-monophosphoryl-sugars with the rate of synthesis proportional to the dolichol content of each submitochondrial fraction.     

ACTH1–24 Releases a Protein from Synaptosomal Plasma Membranes

Abstract: Brain membranes contain several protein kinases, all of which appear to play a role in the regulation of neuronal functioning. These membranes also contain numerous (phospho) proteins. It has been proposed that the degree of phosphorylation of some of these proteins may affect neuronal membrane properties. In a series of previous reports we showed that ACTH_1-24 inhibits the endogenous phosphorylation of several synaptosomal plasmamembrane (SPM) proteins including the B-50 protein. Although we have speculated that the degree of phosphorylation of B-50 may be important in regulating the turnover of membrane (poly)-phosphoinositides, the exact nature of the interaction between ACTH_1-24 and B-50/B-50 protein kinase is unknown. The purpose of the present study was to determine whether treatment of SPM with ACTH_1-24 will lead to a specific release of proteins from SPM. We found that ACTH_1-24 specifically releases a 41,000 M_r protein from rat brain SPM. Although we are not certain about the biological significance of the release of this polypeptide, it is of sufficient interest for further research in view of the lack of success of finding binding of labeled ACTH to brain membranes.     

Hydrolysis of dolichyl esters by oviduct membranes and characterization of endogenous dolichyl esters

The chick oviduct system has been employed to study whether dolichol esters might serve as a storage form of dolichol to be converted to dolichyl phosphate (Dol-P) during periods when Dol-P levels increase. Chicken oviduct membranes catalyze the hydrolysis of dolichyl-[14C]oleate; the reaction is dependent on detergent (0.04% NP-40 is optimal), is unaffected by divalent cations and EDTA, and exhibits a pH optimum of 6.0. Oviduct membranes also hydrolyze cholesteryl-[14C]oleate, which exhibits similar properties except the pH optimum is 5.0-5.5. Neither Dol-[14C]palmitate nor Chol-[14C]palmitate is hydrolyzed by membranes. Chol-ester hydrolysis is more sensitive to heat-denaturation than is Dol-ester hydrolysis. Esterase activity was assayed in membranes prepared from immature chicks, chicks treated with diethylstilbestrol, chicks withdrawn from diethylstilbestrol, and mature hens. The highest esterase specific activity was observed in membranes obtained from chicks withdrawn from hormone. In order to characterize the fatty acid composition of Dol-esters they were purified from mature hen oviducts by chromatography on DEAE-cellulose and Fractogel ORPVA-6000, reverse-phase HPLC, and TLC. About 15-25% of oviduct dolichol is in the esterified form. Fatty acid analysis revealed that approximately 85% of the dolichol was esterified to oleic acid. The fact that the highest esterase activity is found in membranes from chicks withdrawn from hormone and that only 20% of the dolichol is esterified argues against a role for Dol-esters as a reservoir of dolichol for conversion to Dol-P.     

Influence of membrane physical state on lysosomal potassium ion permeability

Lysosomal permeability to potassium ions is an important property of the organelle. Influence of the membrane physical state on the potassium ion permeability of isolated lysosomes was assessed by measuring the membrane potential with bis(3-propyl-5-oxoisoxazol-4-yl)pentamethine oxonol and monitoring the lysosomal proton leakage with p-nitrophenol. The membrane fluidity of lysosomes was modulated by treatment with membrane fluidizer benzyl alcohol and rigidifier cholesteryl hemisuccinate. Changes in the membrane order were examined by steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene. The measurements of membrane potential and proton leakage demonstrated that the permeability of lysosomes to potassium ions increased with rigidification of their membranes by cholesteryl hemisuccinate treatment at 37 degrees C, and decreased with fluidization of their membranes by benzyl alcohol treatment at 2 degrees C. The changes in ion permeability could be recovered by fluidizing the rigidified membranes and rigidifying the fluidized membranes. The results suggest that the physical states of lysosomal membranes play an important role in the regulation of their K(+) permeability.     

Dolichol induces membrane leakage of liposomes composed of phosphatidylethanolamine and phosphatidylcholine

Dolichol promotes the leakage of membranes in liposomes composed of phosphatidylethanolamine and phosphatidylcholine but not liposomes composed only of phosphatidylcholine. The membrane leakage was assayed by measuring the entrapment of TEMPOcholine, a cationic spin probe, in liposomes using ESR methods. The percent of membrane leakage induced by dolichol was found to be linearly proportional to the concentrations of dolichol. It is proposed that dolichol enhances the formation of non-bilayer configurations in liposomes containing phosphatidylethanolamine, thereby membrane leakage.     

Content of dolichol and retinol in isolated rat non-parenchymal liver cells

The liver sinusoids, that are considered as a functional unit, harbour four types of sinusoidal cells (Ito, Kupffer, endothelial and pit cells). Dolichol content has been determined in many tissues and subcellular compartments, alteration has been reported in many types of liver injury, but until now no data are available on its content in every type of sinusoidal non-parenchymal liver cells. Dolichol and retinol metabolism might intersect in their traffic in biological membranes. Intercellular as well as intracellular exchange of retinoids is an essential element of important processes occurring in liver cells. It has been suggested that the role of dolichol, besides being a carrier of oligosaccharides in the biosynthesis of N-linked glycoproteins, may be to modify membrane fluidity and permeability, and facilitate fusion of membranes. Dolichol in the membrane is intercalated between the two halves of the phospholipid bilayer, but its exact disposition is not known and the movement and distribution of retinoid in membranes may vary with the geometry of the membranes. Therefore the aim of this study is to obtain a global understanding of the sinusoidal system regarding dolichol and retinol content in each type of isolated rat liver sinusoidal cell, in normal conditions and after vitamin A administration. The information that can be drawn from the present results is that with normal vitamin A status of the animal, the dolichol content is almost uniform in all liver cells. After vitamin A supplementation, a great increase of dolichol, together with the known increase of retinol, can be measured only in a subpopulation of the Ito cells, the Ito-1 subfraction. Therefore in the cells that are present in the hepatic sinusoid, different pools of dolichol may have separate functions. Because retinol traffic among cells, membranes and plasma still remains to be fully understood, roles of dolichol in the exchange of vitamin A among sinusoidal liver cells are discussed. © 1998 John Wiley & Sons, Ltd.     

Influence of Membrane Physical State on the Lysosomal Proton Permeability

Influence of membrane physical state on the proton permeability of isolated lysosomes was assessed by measuring the membrane potential with 3,3′-dipropylthiadicarbocyanine iodide and monitoring their proton leakage with p-nitrophenol. Changes in the membrane order were examined by the steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene. Both the membrane potential and proton leakage increased with fluidizing the lysosomal membranes by benzyl alcohol and decreased with rigidifying the membranes by cholesteryl hemisuccinate. The proton permeability increased to the maximum of 42% by the benzyl alcohol treatment and decreased to the minimum of 38.1% by the cholesteryl hemisuccinate treatment. Treating the lysosomes with protonophore CCCP increased the proton permeability by 58%. The effects of the membrane fluidization and rigidification can be reversed by rigidifying the fluidized membranes and fluidizing the rigidified membranes, respectively. The results indicate that the proton permeability of lysosomes increased and decreased with increasing and decreasing their membrane fluidity, respectively. Moreover, the lysosomal proton permeability did not alter further if the changes, either an increase or a decrease, in the fluidity exceeded some amount. The results suggest that the proton permeability of lysosomes can be modulated finitely by the alterations in their membrane physical state. Received: 27 September 1999 / Revised: 27 December 1999     

Comparative studies on mannosylphosphoryl dolichol and glucosylphosphoryl dolichol synthases

Factors affecting the synthesis of mannosylphosphoryl dolichol and glucosylphosphoryl dolichol hen oviduct microsomes were compared in order to gain insight into the properties of their respective synthases. A stabilized form of mannosylphosphoryl dolichol synthase, but not glucosylphosphoryl dolichol synthase, was released from microsomes by freezing the membranes after exposure to the detergent CHAPSO. The activation energy for mannosylphosphoryl dolichol synthesis in membranes was 9.4 glucosylphosphoryl dolichol synthesis in membranes had a similar activation energy, 8.1 kcal/mol, but below 18 degrees C the value was 16.7 kcal/mol. Tryptic digestion of sealed microsomes preferentially inactivated mannosylphosphoryl dolichol synthase; however, both synthases were equally inactivated in detergent-permeabilized microsomes. Periodate-oxidized UDP-Glc was used to probe the topological orientation of glucosylphosphoryl dolichol synthase in rat liver microsomes. Sealed microsomes treated with oxidized UDP-Glc were inactive in synthesis of glucosylphosphoryl dolichol. However, when these treated microsomes were permeabilized, glucosylphosphoryl dolichol synthase activity was readily detected. From these studies we conclude that although mannosyl- and glucosylphosphoryl dolichol synthases catalyze chemically similar reactions in the endoplasmic reticulum, they differ in several respects. These differences were interpreted in terms of a topological model in which the active sites of the two enzymes reside on opposite faces of the endoplasmic reticulum, with that of the glucosyl lipid synthase facing the lumen and that of the mannosyl lipid synthase facing the cytosol.     

Interactive effects of dietary (n-3) polyunsaturated fatty acids and chronic ethanol intoxication on synaptic membrane lipid composition and fluidity in rats.

The influence of dietary polyunsaturated fatty acids on fatty acid composition, cholesterol and phospholipid content as well as 'fluidity' (assessed by fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) probes) of brain synaptic plasma membranes (SPM) and their interactions with chronic ethanol effects were studied in rats fed for two generations with diets either devoid of (n-3) fatty acids (sunflower oil diet), rich in alpha-linolenic acid (soya oil diet) or in long chain (n-3) fatty acids (sunflower + cod liver oil diet). Results were compared with rats fed standard lab chow. Sunflower oil led to an increase in the (n-6)/(n-3) ratio in the membranes with an increase of the 'fluidity' at membrane apolar level; sunflower + cod liver oil decreased the (n-6)/(n-3) ratio without affecting membrane 'fluidity' while no difference was seen between the SPM of rats fed soya oil and standard diet. After 3 weeks alcohol intoxication in rat fed the standard diet: oleic alpha-linoleic acids and cholesterol levels were increased, arachidonic acid and the double bond index/saturated fatty acids were decreased and there was a decrease of 'fluidity' in the lipid core of the SPM. Soya oil almost totally abolished these usually observed changes in the SPM fatty acids composition but increased oleic acid and cholesterol without any change in fluidity. Sunflower oil led to the same general alterations of fatty acid as seen with standard diet but to a greater extent, with decrease of the 'fluidity" at the apolar level and in the region probed by TMA-DPH. When sunflower oil was supplemented with cod liver oil, oleic and alpha-linoleic acids were increased while the 'fluidity' of the apolar core of SPM was decreased. So, the small changes in fatty acid pattern seem able to modulate neural properties i.e. the responses to a neurotoxic like ethanol. A structurally specific role of PUFA is demonstrated by the pernicious effects of the alpha-linolenic acid deficient diet which are not totally prevented by the supply of long chain (n-3) PUFA.     

Transbilayer effects of ethanol on fluidity of brain membrane leaflets

Previous work on membrane effects of ethanol focused on fluidization of the bulk membrane lipid bilayer. That work was extended in the present study to an examination of ethanol's effect on lipid domains. Two independent methods were developed to examine the effects of ethanol on the inner and outer leaflets of synaptic plasma membranes (SPM). First, differential polarized phase and modulation fluorometry and selective quenching of diphenyl-1,3,5-hexatriene (DPH) were used to examine individual leaflets. Both limiting anisotropy and rotational relaxation time of DPH in SPM indicated that the outer leaflet was more fluid than the inner leaflet. Second, plasma membrane sidedness selective fluorescent DPH derivatives, cationic 1-[4-(trimethylammonio)phenyl]-6-phenylhexa-1,3,5-triene (TMA-DPH) and anionic 3-[p-6-phenyl)-1,3,5-hexatrienyl]phenylpropionic acid (PRO-DPH), confirmed this transmembrane fluidity difference. TMA-DPH and PRO-DPH preferentially localized in the inner and outer leaflets of SPM, respectively. Ethanol in vitro had a greater fluidizing effect in the outer leaflet as compared to the inner leaflet. Thus, ethanol exhibits a specific rather than nonspecific fluidizing action within transbilayer SPM domains. This preferential fluidization of the SPM outer leaflet may have a role in ethanol affecting transmembrane signaling in the nervous system.     

Phosphatidylcholine and sphingomyelin containing an elaidoyl fatty acid can form cholesterol-rich lateral domains in bilayer membranes

Elaidic acid is a trans-fatty acid found in many food products and implicated for having potentially health hazardous effects in humans. Elaidic acid is readily incorporated into membrane lipids in vivo and therefore affects processes regulating membrane physical properties. In this study the membrane properties of sphingomyelin and phosphatidylcholine containing elaidic acid (N-E-SM and PEPC) were determined in bilayer membranes with special emphasis on their interaction with cholesterol and participation in ordered domain formation. In agreement with previous studies the melting temperatures were found to be about 20 degrees C lower for the elaidoyl than for the corresponding saturated lipids. The trans-unsaturation increased the polarity at the membrane-water interface as reported by Laurdan fluorescence. Fluorescence quenching experiments using cholestatrienol as a probe showed that both N-E-SM and PEPC were incorporated in lateral membrane domains with sterol and saturated lipids. At low temperatures the elaidoyl lipids were even able to form sterol-rich domains without any saturated lipids present in the bilayer. We conclude from this study that the ability of N-E-SM and PEPC to form ordered domains together with cholesterol and saturated phospho- and sphingolipids in model membranes indicates that they might have an influence on raft formation in biological membranes.     

The relationship between plasma membrane lipid composition and physical-chemical properties. II. Effect of phospholipid fatty acid modulation on plasma membrane physical properties and enzymatic activities

The fatty acid composition of plasma membrane phospholipids of the murine T lymphocyte tumor EL4 were systematically modified in an attempt to understand the relationship between lipid bilayer composition and plasma membrane physical and biological properties. Two plasma membrane enzyme activities, adenylate cyclase and ouabain-sensitive (Na+ + K+)-ATPase, were measured in normal and fatty acid-substituted EL4 plasma membrane fractions. The fatty acid effect on enzyme activities was similar to previously reported effects of fatty acids on cytotoxic T cell function. The activity of both enzymes was inhibited by saturated fatty acids, while unsaturated fatty acids had a moderate enhancing effect on both enzyme activities. Using two different nitroxide derivatives of stearic acid, the order parameter and approximate rotational correlation times were calculated from ESR spectra of normal and fatty acid-modified plasma membranes. No significant differences was found in either parameter in these membranes. These results, in conjunction with earlier data from our laboratory and others, suggest that caution should be exercised in inferring changes in membrane 'fluidity' based on lipid modulation of biological membranes.     

The cell membrane as a biosensor of oxidative stress induced by radiation exposure: a multiparameter investigation

The role of biological membranes as a target in biological radiation damage remains unclear. The present study investigates how the biochemical and biophysical properties of a simple biological model, i.e. human erythrocyte membranes, are altered after exposure to relatively low doses of (60)Co gamma rays. Lipid peroxidation increased in the hours after radiation exposure, based on measurements of MDA and on the lipid peroxidation index after parinaric acid incorporation. Protein carbonyl content also increased rapidly after radiation exposure. An imbalance between the radiation-mediated oxidative damages and the antioxidant capacity of the erythrocytes was observed in the hours after radiation exposure. Antioxidant enzyme activities, mainly catalase and glutathione peroxidase, were found to decrease after irradiation. The development of a radiation-induced oxidative stress probably explains the reorganization of the fatty acid pattern 72 h after radiation exposure. The phosphatidylethanolamine (PE) fatty acids of the (n-3) and (n-6) series decreased, while the PE saturated fatty acid content increased. All these modifications may be involved in the variation of the biophysical properties of the membranes that we noted after radiation exposure. Specifically, we observed that the lipid compartment of the membrane became more fluid while the lipid-protein membrane interface became more rigid. Taken together, these findings reinforce our understanding that the cell membrane is a significant biological target of radiation. Thus the role of the biological membrane in the expression and course of cell damage after radiation exposure must be considered.     

Uptake of dolichol by Vero cells.

Characterization and kinetics of dolichol uptake by a Vero cell line are reported. Vero cells incorporate dolichol in a time- and dose-dependent manner. Optimal uptake is found at 37 degrees C and at a pH of 7.4. In contrast to cholesterol, an inhibitory effect on the dolichol incorporation is found for farnesol, geraniol, and retinol. Long chain polyprenols were slightly stimulatory. The translocation seems not to be highly energy dependent. The lack of substantial inhibition by chloroquine does not plead for a receptor-mediated endocytosis. Incorporated dolichol was distributed over both membranes and supernatant fractions, paralleling the distribution of the lysosomal marker beta-N-acetylhexosaminidase. The incorporated dolichol is subject to a fast efflux process, which is potentiated by the presence of lipid acceptors in the extracellular medium.     

Cholesterol sulfate inhibits the fusion of Sendai virus to biological and model membranes

Cholesterol sulfate is a component of several biological membranes. In erythrocytes, cholesterol sulfate inhibits hypotonic hemolysis, while in sperm, it can decrease fertilization efficiency. We have found cholesterol sulfate to be a potent inhibitor of Sendai virus fusion to both human erythrocyte and liposomal membranes. Cholesterol sulfate also raises the bilayer to hexagonal phase transition temperature of dielaidoyl phosphatidylethanolamine as demonstrated by differential scanning calorimetry and 31P nuclear magnetic resonance spectrometry. Although hexagonal phase structures are not readily found in biological membranes, there is a correlation between the effects of membrane additives on bilayer/non-bilayer equilibria and membrane stabilization. It is proposed that the ability of cholesterol sulfate to alter the physical properties of membranes contributes to its stabilization of biological membranes and the inhibition of membrane fusion.     

Formation of dolichol-linked sugar intermediates during the postnatal development of skeletal muscle

The postnatal development of skeletal muscle is characterized by changes in membrane function associated with N-linked glycoproteins. In the present study, early reactions involved in the synthesis of the dolichol-linked core oligosaccharide were examined in neonatal and adult rabbit skeletal muscle sarcoplasmic reticulum membranes. The initial rate of N-acetylglucosamine incorporation in the presence of exogenous dolichol phosphate was similar between neonate and adult (3.5-4.1 pmol of GlcNAc/min/mg). The Km values for UDP-GlcNAc and exogenous dolichol phosphate were similar. Tunicamycin (0.04-0.08 micrograms/ml) inhibited N-acetylglucosamine incorporation by 50%. UDP-GlcNAc pyrophosphatase activity was greater in neonatal membranes than adult (840 versus 350 pmol of GlcNAc-1-P/min/mg), explaining, in part, the greater enhancement of neonatal GlcNAc incorporation by pyrophosphatase inhibitors. Nucleotide-sugar pyrophosphatase inhibitors (alpha, beta-methylene ATP and dimercaptopropanol) increased the capacity of neonatal activity 4-fold and adult enzyme 2-fold. Analysis of dolichol-linked products by mild acid hydrolysis however, revealed that neonate had higher capacity for N,N'-diacetylchitobiosyl(pyro)phosphoryldolichol synthesis than adult. Mannosyltransferase and glucosyltransferase were elevated 6- and 5-fold in neonate compared to adult membranes. Neonate exhibited 4-fold greater GDP-Man pyrophosphatase activity than adult (500 versus 125 pmol of Man-1-P/min/mg). The Km for GDP-Man increased in the presence of exogenous dolichol phosphate. Increasing concentrations of exogenous dolichol phosphate did not equalize neonate and adult mannosyltransferase activity, indicating that the decline in activity during development was not due to a decrease in a pool of dolichol phosphate accessible to mannosyltransferase. Glucosyltransferase for the synthesis of glucosylphosphoryldolichol was also elevated 5-fold in neonatal compared to adult sarcoplasmic reticulum (7 versus 1.4 pmol of Glc/min/mg). In a previous study, it was reported that glycoprotein sialyltransferase activity decreased by a factor of 6.5 during the postnatal maturation and that total membrane hexose content of sarcoplasmic reticulum decreased by a factor of 8. Together, these results suggest that the postnatal development of skeletal muscle is characterized by coordinated changes in the expression of enzymes involved in both the "early" and "late" reactions of N-linked oligosaccharide biosynthesis.     

Acute and Chronic Effects of Ethanol on Transbilayer Membrane Domains

Alcohols, including ethanol, have a specific effect on transbilayer and lateral membrane domains. Recent evidence has shown that alcohols in vitro have a greater effect on fluidity of one leaflet as compared to the other. The present study examined effects of chronic ethanol consumption on fluidity of synaptic plasma membrane (SPM) exofacial and cytofacial leaflets using trinitrobenzenesulfonic acid (TNBS) labeling and differential polarized fluorometry of 1,6-diphenyl-1,3,5-hexatriene (DPH). Mice were administered ethanol or a control liquid diet for 3 weeks. Animals were killed and SPM prepared. The exofacial leaflet of SPM was significantly more fluid than the cytofacial leaflet in both groups, as indicated by limiting anisotropy of DPH. However, differences between the two leaflets were much smaller in the ethanol-treated group. Ethanol at concentrations seen clinically had a greater effect in vitro on the more fluid exofacial leaflet. This asymmetric effect of ethanol was significantly diminished in the exofacial leaflet of the ethanol-treated mice. Chronic ethanol consumption has a specific effect on membranes. Membrane functions that may be regulated by asymmetry of fluidity and lipid distribution may be altered by chronic ethanol consumption.     

The influence of dolichol, dolichol esters, and dolichyl phosphate on phospholipid polymorphism and fluidity in model membranes

The effect of dolichols, polyprenols, dolichol esterified with fatty acids, and dolichyl phosphate on the structure and fluidity of model membranes was studied using 31P NMR, small-angle x-ray scattering, differential scanning calorimetry, and freeze-fracture electron microscopy. These studies suggest that dolichol and dolichol derivatives destabilize unsaturated phosphatidylethanolamine containing bilayer structures and promote hexagonal II phase formation; high concentrations of dolichol induce lipid structures characterized by "isotropic" 31P NMR and particulate fracture faces; dolichol, contrary to cholesterol, has no effect on the thermotropic behavior of membranes consisting of phosphatidylcholine, while dolichyl-P incorporation abolishes the transition from the gel to liquid crystalline phase in 1,2-dimyristoyl-sn-glycero-3-phosphocholine; both dolichol and dolichyl-P increase the fatty acid fluidity in phosphatidylethanolamine mixtures; the effect of dolichol on bilayer structure and fluidity is more pronounced with increasing number of isoprene residues; dolichol esters are only soluble to a limited extent in the bilayer and segregates into domains at low concentrations; the results are consistent with a localization of dolichyl-P in which the phosphate group is oriented to the water interphase. The induction of hexagonal II phase by dolichyl-P may elicit the transmembrane movement of glycosylated lipid intermediate.