Structural state of the cardiomyocyte sarcolemma in animals of different ages

Cardiomyocyte plasmalemma membranes of adult (12 months) and old (24-26 months) rats have been studied. Ageing is accompanied with an essential decrease of the content of the membrane protein SH-groups. We have found an increase in the fraction of strongly immobilized spin-labeled SH-groups, a decrease in that of unsaturated lipids, as well as an increase in that of lysophosphatidylcholine in old rats membranes. The pattern of the response of beta-adrenergic receptors to isoproterenol is changed with age. The mobility of spin probe 5NS in the membranes of old rats is lowered at 22.5 degrees C. However there is no statistically significant age difference in the spin-probe mobility at 37 degrees C. Possible role of lysophosphatidylcholine in the stabilization of membrane lipid "fluidity" at physiological temperatures is suggested.     

Fluidity of natural membranes and phosphatidylserine and ganglioside dispersions. Effect of local anesthetics, cholesterol and protein.

The microviscosity of artificial lipid membranes and natural membranes was measured by the fluorescence polarization technique employing perylene as the probe. Lipid dispersions composed of brain gangliosides exhibited greater microviscosity than phosphatidylserine (268 cP vs 173 cP, at 25 degrees C). Incorporation of cholesterol (30-50%) increased the microviscosity of lipid phases by 200-500 cP. Cholesterol's effect on membrane fluidity was completely reversed by digitonin but not by amphotericin B. Incorporation of membrane proteins into lipid vesicles gave varying results. Cytochrome b5 did not alter membrane fluidity. However, myelin proteolipid produced an apparent increase in microviscosity, but this effect might be due to partitioning of perylene between lipid and protein binding sites since tha latter have a higher fluorescence anisotropy than the lipid. The local anesthetics tetracain and butacaine increased the fluidity of lipid dispersions, natural membranes and intact ascites tumor cell membranes. The effect of anesthetics appears to be due to an increased disordering of lipid structure. The fluidity of natural membranes at 25 degrees C varied as follows: polymorphonuclear leukocytes, 335 cP; bovine brain myelin, 270 cP; human erythrocyte, 180 cP; rat liver microsomes, 95 cP; rat liver mitochondria, 90 cP. In most cases the microviscosity of natural membranes reflects their cholesterol: phospholipid ratio. The natural variations in fluidity of cellular membranes probably reflect important functional requirements. Similarly, the effects of some drugs which alter membrane permeability may be the result of their effects on membrane fluidity.     

Alterations in the physical state and composition of brush border membrane lipids of rat enterocytes during differentiation

The physical state of the membrane lipid of brush border membranes, prepared from rat small intestinal villus and crypt cells, was examined by steady-state fluorescence polarization using three lipid-soluble fluorophors. Membranes prepared from crypt cells were found to possess a higher lipid fluidity than those of villus cells with each probe. Analysis of the composition of these membranes revealed that those from crypt cells had lower ratios of cholesterol/phospholipid (mol/mol), protein/lipid (w/w), and saturated fatty acyl chains/unsaturated chains (w/w). Alterations in the levels of stearic (18:0) and oleic (18:1) acids were responsible for differences in the latter ratio. The results, therefore, demonstrate that alterations in the lipid composition and fluidity of brush border membranes of enterocytes occur during the process of differentiation.     

Fluidity of natural membranes and phosphatidylserine and ganglioside dispersions: Effects of local anesthetics,cholesterol and protein

The microviscosity of artificial lipid membranes and natural membranes was measured by the fluorescence polarization technique employing perylene as the probe. Lipid dispersions composed of brain gangliosides exhibited greater microviscosity than phosphatidylserine (268 cP vs 173 cP, at 25 °C). Incorporation of cholesterol (30–50%) increased the microviscosity of lipid phases by 200–500 cP. Cholesterol's effect on membrane fluidity was completely reversed by digitonin but not by amphotericin B. Incorporation of membrane proteins into lipid vesicles gave varying results. Cytochrome b₅ did not alter membrane fluidity. However, myelin proteolipid produced an apparent increase in microviscosity, but this effect might be due to partitioning of perylene between lipid and protein binding sites since the latter have a higher fluorescence anisotropy than the lipid. The local anesthetics tetracaine and butacaine increased the fluidity of lipid dispersions, natural membranes and intact ascites tumor cell membranes. The effect of the anesthetics appears to be due to an increased disordering of lipid structure. The fluidity of natural membranes at the 25 °C varied as follows:polymorphonuclear leukocytes, 335 cP; bovine brain myelin, 270 cP; human erytherocyte, 180 cP; rat liver microsomes, 95 cP; rat liver mitochondria, 90 cP. In most cases the microviscosity of natural membranes reflects their cholesterol : phospholipid ratio. The natural variations in fluidity of cellular membranes probably reflect important fuctional requirements. Similarly, the effects of some drugs which alter membrane permeability may be the result of their effects on membrane fluidity.     

Dexamethasone-induced alterations in lipid composition and fluidity of rat proximal-small-intestinal brush-border membranes.

A series of experiments were conducted to examine the possible effects of subcutaneous administration of the synthetic glucocorticoid dexamethasone (100 micrograms/day per 100 g body wt.) on the lipid fluidity and lipid composition of rat proximal-small-intestinal brush-border membranes. After 4 days of treatment, membranes and their liposomes prepared from treated animals possessed a greater fluidity than did their control (diluent, 0.9% NaCl) counterparts, as assessed by steady-state fluorescence-polarization techniques using several different fluorophores. Examination of the effects of temperature on the anisotropy values of 1,6-diphenylhexa-1,3,5-triene, using Arrhenius plots, moreover, revealed that the mean break-point temperatures of the treated preparations were approx. 3-4 degrees C lower than those of their control-preparation counterparts. Changes in the sphingomyelin/phosphatidylcholine (PC) molar ratio as well as in certain of the fatty acids of the PC fraction of treated membranes, secondary to alterations in membrane PC levels and in lysophosphatidylcholine acyltransferase activities respectively, were also noted after dexamethasone administration. These compositional alterations appeared to be responsible, at least in part, for the differences in fluidity noted between treated and control plasma membranes. These results therefore demonstrate that dexamethasone administration can modulate the lipid fluidity and lipid composition of rat proximal-small-intestinal brush-border membranes.     

Incorporation of sterol into chloroplast thylakoid membranes and its effect on fluidity and function

The sterol, cholesteryl hemisuccinate, has been incorporated into isolated thylakoid membranes of pea and lettuce chloroplasts in order to modify the fluidity of the lipid matrix. Changes in fluidity have been monitored using fluorescence polarization of the hydrophobic probe, 1,6-diphenyl-1,3,5-hexatriene and the electron-spin-resonance, spin-label probe, 5-doxyl stearate. Both methods indicate that incorporation of increasing levels of sterol reduces the fluidity of the thylakoid lipid matrix. At room temperature the thylakoid lipid matrix is relatively fluid and the effect of increasing the viscosity is to inhibit partially the maximum rate of steady-state electron flow and reduce the dark rate of reduction of flash-oxidised cytochrome f. The results are discussed in terms of lipid fluidity influencing the rate of lateral diffusion of reduced plastoquinone from photosystem II to photosystem I.     

Regional differences in the lipid composition and fluidity of rat colonic brush-border membranes

The lipid composition and fluidity of brush-border membranes prepared from rat proximal and distal colonocytes were determined. Fluidity, as assessed by steady-state fluorescence polarization techniques using the fluorophores 1,6-diphenyl-1,3,5-hexatriene, DL-2(9-anthroyl)stearic acid and DL-12(9-anthroyl)stearic acid, was decreased in distal compared to proximal plasma membranes. This pattern was similar to that previously described for both antipodal plasma membranes in rat enterocytes of the small intestine. The decrease in fluidity of the distal as compared to the proximal membranes resulted from an increase in cholesterol content, cholesterol/phospholipid molar ratio and degree of saturation of the fatty acid residues in the distal membranes. The specific activities of total alkaline phosphatase and cysteine-sensitive alkaline phosphatase, enzymes previously shown to be functionally dependent on the physical state of the colonic brush-border membrane's lipid, were also significantly lower in distal as compared to proximal clonic plasma membranes. These studies, therefore, demonstrate that differences in the lipid fluidity, lipid composition and certain enzymatic activities exist in brush-border membranes prepared from rat proximal and distal colonocytes. The regional variation in rat colonic luminal membrane lipid fluidity and composition may, at least partially, be responsible for differences in these enzymatic activities as well as in sodium and water absorption along the length of this organ.     

Spin label studies on rat liver and heart plasma membranes: Effects of temperature,calcium, and lanthanum on membrane fluidity

The structures of rat liver and heart plasma membranes were studied with the 5-nitroxide stearic acid spin probe, I(1 2,3). The polarity-corrected order parameters (S) of liver and heart plasma membranes were independent of probe concentration only if experimentally determined low I(1 2,3)/lipid ratios were employed. At higher probe/lipid ratios, the order parameters of both membrane systems decreased with increasing probe concentration, and these effects were attributed to enhanced nitroxide radical interactions. Examination of the temperature dependence of approximate and polarity-corrected order parameters indicated that lipid phase separations occur in liver (between 19° and 28°C) and heart (between 21° and 32°C) plasma membranes. The possibility that a wide variety of membrane-associated functions may be influenced by these thermotropic phase separations is considered. Addition of 3.9 mM CaCl₂ to I(1 2,3)-labeled liver plasma membrane decreased the fluidity as indicated by a 5% increase in S at 37°C. Similarly, titrating I(1 2,3)-labeled heart plasma membranes with either CaCl₂ or LaCl₃ decreased the lipid fluidity at 37°C, although the magnitude of the La³⁺ effect was larger and occurred at lower concentrations than that induced by Ca²⁺; addition of 0.2 mM La³⁺ or 3.2 mM Ca²⁺ increased S by approximately 7% and 5%, respectively. The above cation effects reflected only alterations in the membrane fluidity and were not due to changes in probe–probe interactions. Ca²⁺ and La³⁺ at these concentrations decrease the activities of such plasma membrane enzymes as Na⁺, K⁺-ATPase and adenylyl cyclase, and it is suggested that the inhibition of these enzymes may be due in part to cation-mediated decreases in the lipid fluidity.     

Dimethylnitrosamine inhibits the glucagon-stimulated adenylate cyclase activity of rat liver plasma membranes and decreases plasma membrane fluidity

The effect of the hepatocarcinogen dimethylnitrosamine on rat liver plasma membrane adenylate cyclase activity and lipid fluidity was assessed. Glucagon-stimulated adenylate cyclase activity exhibited a complex response to increasing concentrations of dimethylnitrosamine, whereas fluoride-stimulated adenylate cyclase activity was progressively inhibited. Maximal inhibitory effects were observed at a concentration of 15 mM in both cases. The activity of detergent-solubilized adenylate cyclase was unaffected by dimethylnitrosamine. ESR analysis using a fatty acid spin probe showed that dimethylnitrosamine produced a marked, dose-dependent reduction in the fluidity of the plasma membrane with a maximal effect occurring at 20 mM. Dimethylnitrosamine also elevated the temperature at which the lipid phase separation occurred in rat liver plasma membranes, from 28 degrees C to 31 degrees C. The non-carcinogenic but structurally similar compound, dimethylamine hydrochloride neither inhibited adenylate cyclase nor decreased plasma membrane fluidity. It is suggested that the decrease in membrane fluidity, induced by dimethylnitrosamine, via its effects on membrane fluidity, could influence plasma membrane function and cellular regulation.     

Lipid-protein interactions in rat renal subcellular membranes: a biophysical and biochemical study

The phase behavior of plasma membrane (PM), endoplasmic reticulum (ER), and nuclear membranes (NM) isolated from adult rat papillary cells was studied using the molecular probe Laurdan. The steady-state fluorescence data analysis was correlated with the lipid composition obtained by biochemical assays. The comparison between intact membranes and protein-free reconstituted vesicles using the whole lipid extract shows the essential role of proteins on the temperature response of natural membranes. The phospholipid (PL) and cholesterol (Cho) content was measured in the three membrane fractions, the PL/Cho molar ratio being between 1.5 and 1.9. However, Laurdan's parameters in NM show a fluid phase state pattern even at low temperature (5 degrees C), with a restricted dipole relaxation in comparison with that displayed in liquid crystalline phase state lipid model membranes. PM and ER are in a gel-like state at temperatures below 20 degrees C, showing increasing dipole relaxation with temperature. The curved fits obtained are characteristic of cholesterol-enriched membranes. The distinctive phase behavior of nuclear membranes vanishes when proteins are extracted. However, relaxation is still faster in this fraction, which correlates with the native lipid composition. NM has the lowest percentage of phosphatidylinositol and sphingomyelin-the latter being a highly saturated phospholipid- and the highest percentage of phosphatidylcholine and phosphatidylethanolamine (PE), nuclear PE being enriched in arachidonic acid. All these changes agree with the higher fluidity of NM compared with ER or PM in the conditions assayed.     

Effect of dehydroepiandrosterone and its sulfate and fatty acid ester derivatives on rat brain membranes

The effects of dehydroepiandrosterone (DHEA) as well as its sulfate and fatty acid ester derivatives on rat brain membrane fluidity was investigated by fluorescence depolarization of a lipid probe 1,6-diphenyl-1,3,5-hexatriene and compared to its effect on phospholipid conformation investigated by Fourier transform infrared spectroscopy. In rat brain, membrane fluidity varied rostro-caudally, the frontal cortex showing the highest fluidity compared to the hypothalamus, hippocampus, striatum, thalamus, and hindbrain. As previously reported, it was observed that cholesteryl hemisuccinate and stearic acid rigidify striatal membrane whereas linoleic acid and L-alpha-phosphatidylcholine increase the membrane fluidity. Striatal fluidity was increased in vitro with increasing concentrations of DHEA, this effect was greater with the DHEA fatty acid ester derivatives (DHEA-L), DHEA-undecanoate, and DHEA-stearate, whereas no effect was observed with DHEA-sulfate (DHEA-S). In the frontal cortex only the two DHEA-L derivatives increased membrane fluidity, whereas DHEA and DHEA-S were without effect. The effect of DHEA-L on synthetic dimyristoylphosphatidylcholine-d54 phospholipid membranes indicates a disordering effect of DHEA-undecanoate and DHEA-stearate as reflected by increased trans-gauche isomerization of the acyl chains of the lipid. Hence, DHEA-L increase the disorder and/or fluidity of brain membranes; interestingly, these compounds are abundant in the brain where they are generally considered as storage compounds that slowly release the active unconjugated steroid hormone.     

Dexamethasone influences the lipid fluidity, lipid composition and glycosphingolipid glycosyltransferase activities of rat proximal-small-intestinal Golgi membranes.

Experiments were performed to examine the effects of subcutaneous administration of the synthetic glucocorticoid dexamethasone (100 micrograms/day per 100 g body wt.) on the lipid fluidity, lipid composition and glycosphingolipid glycosyltransferase activities of rat proximal-small-intestinal Golgi membranes. After 4 days of treatment, Golgi membranes and liposomes prepared from treated rats were found to possess a greater fluidity than their control (diluent or 0.9% NaCl) counterpart, as assessed by steady-state fluorescence-polarization techniques using three different fluorophores. Moreover, analysis of the effects of temperature on the anisotropy values of 1,6-diphenylhexa-1,3,5-triene, using Arrhenius plots, demonstrated that the mean break-point temperatures of treated preparations were 4-5 degrees C lower than those of control preparations. Changes in the fatty acyl saturation index and double-bond index of treated membranes, secondary to alterations in stearic acid, linoleic acid and arachidonic acid, at least in part, appeared to be responsible for the differences in fluidity noted between treated and control Golgi membranes. Concomitant with these fluidity and lipid-compositional alterations, treated membranes possessed higher specific activities of UDP-galactosyl-lactosylceramide galactosyltransferase and CMP-N-acetylneuraminic acid:lactosylceramide sialyltransferase than their control counterparts. Experiments utilizing benzyl alcohol, a known fluidizer, furthermore suggested that the fluidity alteration induced by dexamethasone may be responsible for the increased activity of the former, but not the latter, glycosphingolipid glycosyltransferase.     

Alcohols inhibit adipocyte basal and insulin-stimulated glucose uptake and increase the membrane lipid fluidity

Benzyl alcohol and ethanol, at aqueous concentrations that cause local anesthesia of rat sciatic nerve, affect structural and functional properties of rat adipocytes. The data strongly suggest that structurally-intact membrane lipids are required for the proper cellular uptake of glucose and for the physiologic response of adipocytes to insulin. The structure of adipocyte membrane lipids was examined with the spin label method. Isolated adipocyte ‘ghost’ membranes were labeled with the 5-nitroxide stearate spin probe I(12,3). Order parameters that are sensitive to the fluidity of the lipid environment of the incorporated probe were calculated from ESR spectra of labeled membranes. Benzyl alcohol and ethanol dramatically increased the fluidity of the adipocyte ghost membrane, as indicated by decreases in the polarity-corrected order parameter S. This concentration-dependent fluidization commenced at approx. 10 mM benzyl alcohol and progressively increased at all higher concentrations tested (up to 107 mM). S decreased approx. 5.7% at 40 mM benzyl alcohol, a change in S comparable in magnitude to that induced by a 6°C increase in the incubation temperature. Benzyl alcohol and ethanol inhibited basal glucose uptake in adipocytes and uptake maximally stimulated by insulin. Temperature-induced increases in membrane fluidity, detected with 1(12,3), that closely paralleled the fluidity effects of alcohols were associated only with increases in basal and insulin-stimulated glucose uptake. The contention that the membrane lipid fluidity plays a role in insulin action needs further study.     

Variations in dietary triacylglycerol saturation alter the lipid composition and fluidity of rat intestinal plasma membranes

Rats were maintained on nutritionally complete diets enriched in unsaturated (corn oil) or saturated (butter fat) triacylglycerols. After 6 weeks, significant differences in the lipid composition and fluidity of a number of intestinal membranes were observed. The corn oil diet (enriched mainly in linoleic acid) increased the overall unsaturation of the acyl chains and enhanced the lipid fluidity, as assessed by the fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene, of enterocyte microvillus and basolateral membranes and of colonocyte basolateral membranes. Concomitantly, the cholesterol content and the cholesterol/phospholipid molar ratio were increased in the microvillus but not in the basolateral membranes. The increased cholesterol in ileal microvillus membranes can result from enhanced cellular biosynthesis, since ileal slices from rats fed the unsaturated diet incorporated [14C]octanoate more rapidly into digitonin-precipitable sterol. Increased fluidity of the enterocyte microvillus and basolateral membranes, respectively, enhanced the enzyme specific activities of p-nitrophenylphosphatase and (Na+ + K+)-dependent adenosine triphosphatase. The results indicate that the lipid composition, fluidity and enzyme activities of intestinal plasma membranes can be altered by dietary means. Moreover, rat enterocytes possess regulatory mechanisms which modulate the cholesterol content of the microvillus membranes so as to mitigate changes in lipid fluidity.     

Orientation and motion of spin-labels in rabbit small intestinal brush border vesicle membranes

The temperature dependence of the packing (order) and fluidity (microviscosity) of rabbit small, intestinal brush border vesicle membranes and of liposomes made from their extracted lipids has been investigated by using a variety of lipid spin probes. The lipids in the brush border membrane are present essentially as a bilayer. Compared to other mammalian membranes, the brush border membrane appears to be characterized by a relatively high packing order as well as microviscosity. At body temperature, the lipid molecules undergo rapid, anisotropic motion, which is essentially a fast rotation about an axis approximately perpendicular to the bilayer normal. Both the order (motional anisotropy) and the microviscosity increase with decreasing temperature and with increasing distance from the center of the bilayer. Qualitatively similar motional or fluidity gradients have been reported for other mammalian and bacterial membranes. The liposomes made from the extracted lipids have a somewhat lower packing order and a slightly higher fluidity than brush border vesicle membranes. The differences are, however, small indicating that the packing and the fluidity (microviscosity) of the membrane are primarily determined by the lipid composition. Membrane-associated proteins and cytoskeleton cannot play a dominant role in determining the order and fluidity of the lipid bilayer. Discontinuities are observed in the temperature dependence of various spectral parameters, the order parameter S, the rotational correlation time tau, and 2,2,6,6-tetramethylpiperidinyloxy partitioning. They are assigned to phase transitions and/or phase separations of the membrane lipids. These discontinuities occur at about 30, 20, and 13 degrees C for 5-doxyl-, 12-doxyl-, and 16-doxylstearic acid, respectively. The apparent transition temperature depends on the location of the spin probe along the bilayer normal, being higher the closer the probe is to the membrane surface. This indicates the possibility that chain melting is progressive and spreads with increasing temperature from the center of the membrane outward.     

Modulation of Na+-H+ exchange by ethinyl estradiol in rat colonic brush-border membrane vesicles

Prior studies by our laboratory have suggested that a relationship may exist between rat colonic brush-border membrane vesicular fluidity and Na+-H+ exchange. To further explore this possible relationship, in the present studies the effects of ethinyl estradiol (17 alpha-ethinyl-1,3,5-estratriene-3,17-beta-diol) administration subcutaneously (5 mg/kg body wt. per day) for 5 days, on rat colonic brush-border membrane fluidity and Na+-H+ exchange were examined. This treatment regimen has previously been shown to decrease the lipid fluidity of rat hepatic and rabbit small intestinal plasma membranes. In agreement with these prior studies, the present results demonstrate that this agent decreases the lipid fluidity of treated-rat colonic brush-border membranes compared to control membranes, as assessed by steady-state fluorescence polarization techniques using three different fluorophores. An increase in the cholesterol content and cholesterol/phospholipid molar ratio of treated-membranes appear to, at least partially, be responsible for the fluidity differences. Furthermore, examination of the kinetic parameters for amiloride-sensitive sodium-stimulated proton efflux in treated and control membrane vesicles, utilizing the pH-sensitive fluorescent dye, Acridine orange, revealed that ethinyl estradiol administration decreased the Vmax for this exchange mechanism, expressed in arbitrary fluorescence units, by approx. 25% but did not influence its Km for sodium. These data, therefore, lend further support to the contention that alterations in fluidity may modulate Na+-H+ exchange in rat colonic brush-border membrane vesicles.     

Localization of the lipid probe 1,6-diphenyl-1,3,5 hexatriene (DPH) in intact cells by fluorescence microscopy

The lipophilic fluorescent probe DPH, generally used to determine the microviscosity of membrane lipids, has been visualized in intact cells by fluorescence microscopy. All lipid material of the cells, including cytoplasmic lipid droplets, was found to be labelled with DPH. The fluorescent signal from inside the cells contributes to a large extent to the total cell fluorescence. The results indicate that fluorescence polarization data obtained from intact cells, using DPH as probe, give information on the total lipid material of the cells rather than exclusive information on microviscosity and fluidity of plasma membranes of these cells, as has been repeatedly suggested.     

Effect of lipid composition on rat liver nuclear membrane fluidity

Nuclear membrane fluidity is measured in rat liver by use of the fluorescence anisotropy of two probes: diphenylhexatriene and its cationic derivative trimethylammonium-diphenylhexatriene. It has been shown that, in 2-month-old rat liver cells, the bilayer surface is less fluid than the hydrophobic core. The fluidity was higher in 6-day-old rat liver nuclei, in which both the amount of cholesterol and the cholesterol/phospholipid ratio decreased. The influence of the single phospholipids, and in particular of phosphatidylcholine, has been studied by increasing the phosphatidylcholine with a choline base exchange reaction in isolated nuclear membranes. After this reaction, the fluorescence anisotropy of the bilayer surface increased, whereas at the hydrophobic core it decreased. Analysis of fatty acid composition shows an increase of phosphatidylcholine unsaturated fatty acids. The results show that the fluidity of nuclear membranes changes in relation to the lipid content and to the fatty acid composition. The role of nuclear membrane fluidity in cell function is discussed. © 1997 John Wiley & Sons, Ltd.     

The effects of cell proliferation on the lipid composition and fluidity of hepatocyte plasma membranes.

The fluorescence probe, 1,6-diphenyl-1,3,5-hexatriene, has been used to investigate the effects of controlled and uncontrolled growth on the dynamic properties of the lipid regions of hepatocyte plasma membranes. DPH was incubated with plasma membranes derived from quiescent and regenerating liver and Morris hepatoma 7777, and the resulting systems were studied by fluorescence polarization spectroscopy. Membranes from the rapidly growing hepatoma exhibited a significantly lower fluorescence polarization than observed in quiescent liver, suggesting the presence of a more fluid membrane lipid domain. Membranes from regenerating liver exhibited a time-dependent increase in membrane fluidity, reaching a maximum 12 h after growth stimulation. A close correspondence between membrane fluidity and the cholesterol-phospholipid ratio was also observed where a decrease in this ratio resulted in a more fluid lipid matrix. These results suggest that cell cycling, as observed in regenerating liver and Morris hepatoma 7777, results in significant increases in membrane fluidity, a property which may play an important regulatory role in various cell functions.     

Time-dependent decay and anisotropy of fluorescence from diphenylhexatriene embedded in the chloroplast thylakoid membrane

The hydrophobic fluorescent probe 1,6-diphenyl-1,3,5-hexatriene has been incorporated into the membranes of isolated thylakoids, separated granal and stromal lamellae and aqueous dispersions of extracted thylakoid galactolipids. Time-resolved fluorescence decays have been recorded on a nanosecond scale using single-photon counting in order to assess the motional properties of the probe. All the experimental systems used showed biphasic decay kinetics and the anisotropies of the decays have been interpreted in terms of a model for wobbling diffusion confined to a cone. The analysis has given information about dynamic and structural restraints of the lipid acyl chains. In the intact thylakoid membrane the degree of order of the fatty acid acyl chains is higher and their rate of motion slower than for isolated lipids. Even so, the dynamic and structural parameters indicate that the thylakoids can be considered as a relatively fluid membrane system when compared with many other biological membranes, a property which is probably required to facilitate efficient long-range diffusion of lipophilic mobile electron-transport components. It is suggested that the optimization of thylakoid fluidity is linked to regulation of the membrane protein/lipid ratio which is also likely to be responsible for the higher fluidity of stromal membranes relative to those of the grana.