Dipalmitoylphosphatidylcholine membranes modified with carotenoid pigment lutein: Experiment versus Monte Carlo simulation study of the membrane organization

Organization of bilayer lipid membrane formed with dipalmitoylphosphatidylcholine and containing a xanthophyll pigment lutein was studied by both the Monte Carlo simulation and UV-Vis absorption spectroscopy. The simulations were based on ten-state Pink model. The proposed model consisted of two monolayers represented by a two-dimensional triangular lattice with vacancies. The orientation and aggregation state of lutein, obtained from the analysis of the spectroscopic measurements, were used to calibrate intermolecular interactions in the model. In accordance with the experimental data, the model allows two orientations of lutein molecules: one spanning the membrane and the other parallel to its plane. The influence of the intermolecular interactions on the main phase transition as well as on the aggregation of lutein molecules is discussed. The analysis of the model enables us to learn about molecular mechanisms that govern the effects of lutein on the membrane properties as well as the effects of the lipid matrix on lutein organization in the membrane. A concept is discussed according to which increasing domination of parallel lutein orientation, observed at high temperatures, can protect the membrane against penetration by water molecules and reactive oxygen species and against loss of the membrane compactness, especially in the regions of oxidized acyl chains.     

The visual cortex "on command": 2 modes of operation of neuronal mechanisms (a topical article)]

Orientation tuning (TO) of 152 single units in the 17th area of cat cerebral cortex was studied under different contrasts of the light bar flashed in RF (2.3, 10 and 100) and also under local and general anaesthesia (Sombrevin, 4 mg/kg or Nembutal, 35 mg/kg). An invariance of preferred orientation to contrast or anaesthesia was revealed in 40-50% of cells. Noninvariant cells shifted significantly their preferred orientation on 22-90 degrees. Invariant units more often have a simple RF, more sharp OT and preferred horizontal and vertical orientations. Under the low-contrast conditions or under general anaesthesia, noninvariant neurons shifted their preferred diagonal orientations. Different role of the two neuronal groups in detection of the reper (horizontal and vertical) and other orientations in a normal conditions (high-contrast, alert state) in comparison with their behavior under worse conditions (low-contrast, narcosis or sleep) is discussed.     

Asymmetric dipping of bacteriophage M13 coat protein with increasing lipid bilayer thickness

Knowledge about the vertical movement of a protein with respect to the lipid bilayer plane is important to understand protein functionality in the biological membrane. In this work, the vertical displacement of bacteriophage M13 major coat protein in a lipid bilayer is used as a model system to study the molecular details of its anchoring mechanism in a homologue series of lipids with the same polar head group but different hydrophobic chain length. The major coat proteins were reconstituted into 14:1PC, 16:1PC, 18:1PC, 20:1PC, and 22:1PC bilayers, and the fluorescence spectra were measured of the intrinsic tryptophan at position 26 and BADAN attached to an introduced cysteine at position 46, located at the opposite ends of the transmembrane helix. The fluorescence maximum of tryptophan shifted for 700 cm^-1 on going from 14:1PC to 22:1PC, the corresponding shift of the fluorescence maximum of BADAN at position 46 was approximately 10 times less (∼ 70 cm^-1). Quenching of fluorescence with the spin label CAT 1 indicates that the tryptophan is becoming progressively inaccessible for the quencher with increasing bilayer thickness, whereas quenching of BADAN attached to the T46C mutant remained approximately unchanged. This supports the idea that the BADAN probe at position 46 remains at the same depth in the bilayer irrespective of its thickness and clearly indicates an asymmetrical nature of the protein dipping in the lipid bilayer. The anchoring strength at the C-terminal domain of the protein (provided by two phenylalanine residues together with four lysine residues) was estimated to be roughly 5 times larger than the anchoring strength of the N-terminal domain.     

Effects of lutein and cholesterol on alkyl chain bending in lipid bilayers: a pulse electron spin resonance spin labeling study.

A short pulse saturation recovery electron spin resonance technique has been used to study the effects of polar carotenoid-lutein and cholesterol on interactions of 14N:15N stearic acid spin-label pairs in fluid-phase phosphatidylcholine (PC) membranes. Bimolecular collisions for pairs consisting of various combinations of [14N]-16-, [14N]-10-, [14N]-7-, or [14N]-5-doxylstearate and [15N]-16-doxylstearate in dimyristoyl-PC (DMPC) or egg yolk PC (EYPC) membranes were measured at 27 degrees C. In the absence and presence of lutein or cholesterol for both lipid systems, the collision rates were ordered as 16:5 < 16:7 < 16:10 < 16:16. For all spin-label pairs studied, interaction frequencies were greater in DMPC than in EYPC. Polar carotenoid-lutein reduces the collision frequency for all spin-label pairs, whereas cholesterol reduces the collision frequency for 16:5 and 16:7 pairs and increases the collision frequency in the membrane center for 16:10 and 16:16 pairs. The presence of unsaturated alkyl chains greatly reduces the effect of lutein but magnifies the effect of cholesterol in the membrane center. The observed differences in the effects of these modifiers on alkyl chain bending result from differences in the structure of cholesterol and polar carotenoid and from their different localization within the lipid bilayer membrane. These studies further confirm the occurrence of vertical fluctuations of alkyl chain ends toward the bilayer surface.     

Orientation and vertical fluctuations of spin-labeled analogues of cholesterol and androstanol in phospholipid bilayers

We have used ESR and NMR linewidth broadening by spin-labels to determine the overall orientation of spin-labeled analogues of cholesterol and androstanol in egg lecithin bilayers. While the cholesterol analogues were found to have a single orientation in each monolayer, with the acyl chain pointing towards the center of the bilayer, the androstanol analogue appeared, at least in sonicated vesicles, to experience two opposite orientations in the same monolayer, very likely with a rapid reorientation. The possibility of rapid vertical fluctuations of the sterol molecules within the phospholipid bilayer is also discussed.     

Enhanced lutein bioavailability by lyso-phosphatidylcholine in rats

The bioavailability of lutein solubilized in mixed micelles containing either phosphatidylcholine (PC) or lysophosphatidylcholine (lysoPC) was evaluated in male rats. Mixed micelles contained 2.5 mM monooleoylglycerol, 7.5 mM oleic acid, 12 mM sodium taurocholate and 200 μM lutein either with 3 mM PC or lysoPC. To study lutein bioavailability, single and repeated dose experiments were conducted. For single dose study, group of rats (n = 30/group) were fed single dose of lutein solubilized in lysoPC (LPC group), PC (PC group) and no phospholipids (NoPL group) in micellar form. Each group was further divided in to five sub-groups (n = 6/sub group) to measure lutein bioavailability over time up to 9 h. For repeated dose study, group of rats (n = 6/group) were fed daily for 10 days a dose of lutein in mixed micelles with NoPL, PC and LPC. A separate group (n = 6) not fed mixed micelles was considered as zero-time control. In both the experiments, mixed micelles (0.2 ml/rat) were fed to the rat by direct intubation to the stomach. Results of single dose studies showed that the mean lutein levels in the plasma and liver of the PC group was significantly lower (p < 0.05) than those of the other two groups. Moreover, the average lutein level in the plasma and liver was significantly (p < 0.05) different among the groups in the order LPC > NoPL > PC. But, repeated dose experiment followed the order LPC > PC > NoPL. The level of lutein excreted through urine and feces of PC group was significantly higher (p < 0.05) than those of the other two groups. Thus, the results indicate that the PC in the mixed micelles suppressed the intestinal uptake of lutein after single dose but not after repeated dose and that lysoPC enhanced the absorption. In both the experiments, plasma and liver level of lutein was higher in LPC compared with PC group. Results also suggest that the luminal hydrolysis of PC to lysoPC is necessary for intestinal uptake of lutein solubilized in mixed micelles.     

Phase II enzyme induction by a carotenoid,lutein, in a PC12D neuronal cell line

The mechanism by which lutein, a carotenoid, acts as an antioxidant in retinal cells is still not fully understood. Here, lutein treatment of a neuronal cell line (PC12D) immediately resulted in reduced intracellular ROS levels, implying that it has a direct role in ROS scavenging. Significantly, lutein treatment also induced phase II antioxidative enzyme expression, probably via a nuclear factor-like 2 (Nrf2) independent pathway. This latter mechanism could explain why lutein acts diversely to protect against oxidative/cytotoxic stress, and why it is physiologically involved in the human neural tissue, such as the retina.     

Horizontal-vertical filters in early vision predict anomalous line-orientation identification frequencies.

A characteristic of early visual processing is a reduction in the effective number of filter mechanisms acting in parallel over the visual field. In the detection of a line target differing in orientation from a background of lines, performance with brief displays appears to be determined by just two classes of orientation-sensitive filter, with preferred orientations close to the vertical and horizontal. An orientation signal represented as a linear combination of responses from such filters is shown to provide a quantitative prediction of the probability density function for identifying the perceived orientation of a target line. This prediction was confirmed in an orientation-matching experiment, which showed that the precision of orientation estimates was worst near the vertical and horizontal and best at about 30 degrees each side of the vertical, a result that contrasts with the classical oblique effect in vision, when scrutiny of the image is allowed. A comparison of predicted and observed frequency distributions showed that the hypothesized orientation signal was formed as an opponent combination and horizontal and vertical filter responses.     

Molecular dynamics simulation studies of the effect of phosphocitrate on crystal-induced membranolysis

In this study, following our earlier work on calcium pyrophosphate dihydrate (CPPD) crystal-induced membranolysis, we demonstrate, using the CHARMM method of molecular dynamics simulation, the protective role of phosphocitrate (PC) against solvated dimyristoyl phosphatidylcholine phospholipid bilayer disintegration on contact with the CPPD crystal. Our molecular dynamics simulations studies show that coverage of the CPPD crystal with a layer of phosphocitrate molecules results in the conservation of phospholipid bilayer integrity. We show that the rupture of the lipid bilayer in presence of CPPD and the protective effect of PC are primarily due to electrostatic interactions. The protective role of PC, which may also play an important and potentially therapeutic function against crystal-induced membranolysis is also discussed.     

Two-channel brain-stem frequency-following responses to pure tone and missing fundamental stimuli

In 2 separate experiments the brain-stem frequency-following response (FFR)was recorded to a pure tone (200 Hz) and complex “missing fundamental” (MF) stimuli differing in temporal fine structure and envelope modulation depth. FFRs were simultaneously recorded in 2 channels with horizontal and vertical dipole orientations. Horizontal electrodes were identical in both experiments (right-left ear), but the vertical configuration was varied (vertex-left ear; vertex-linked mastoids). The horizontal channel yielded a well defined FFR to tone stimulation at a latency consistent with an origin along the auditory nerve. However, there was no horizontal response to MF stimulation. This latter finding provides electrophysiological support for the conclusion that MFs are not directly coded in the peripheral neural response. Vertical recordings, however, showed equally well defined FFRs to tone and MF stimuli. Thus, a representation of the missing fundamental frequency is registered in the brain-stem. Vertical latencies were consistent with a source at the level of the lateral lemniscus. The FFR is well suited to elucidate certain brain-stem mechanisms of auditory information processing. Important additional information results when responses are compared in horizontal and vertical dipole orientations. Thus, the present results provide the first evoked response demonstration of a peripheral-brain-stem dichotomy of MF coding.     

Binary mixtures of asymmetric phosphatidylcholines with one acyl chain twice as long as the other

High-resolution differential scanning calorimetry and 31P NMR spectroscopy have been used to study aqueous phosphatidylcholine (PC) dispersions prepared from colyophilized mixtures of C(10):C(22)PC/C(22):C(12)PC of various molar ratios. These two lipid species are highly asymmetric but have a common structural feature; namely, one acyl chain in the fully extended conformation is about twice as long as the other. Our experimental results support two conclusions: (1) These two component lipids are miscible in all proportions in both gel and liquid-crystalline states. This type of system behaves as a nearly ideal mixture. Its calorimetric parameters are those expected on the basis of the mole fraction weighted average of the corresponding parameters for the pure components. (2) The component lipids appear to self-assemble, at T less than Tm, into a mixed interdigitated bilayer in excess water. In a mixed interdigitated bilayer, the short acyl chain of one asymmetric phosphatidylcholine on one side of the bilayer leaflet is apposed with the short acyl chain of another lipid molecule on the other side of the bilayer leaflet, while the longer acyl chain from each of the two leaflets crosses the entire hydrocarbon width of the bilayer. The fundamental packing unit, as well as the dynamic unit describing the axial rotator motion about the bilayer normal for this mixed interdigitated bilayer, is thus a dimer, whereas the packing unit assigned for the noninterdigitated bilayer such as C(16):C(16)PC lamellae is a monomer.     

Oscillation frequencies of plant stems with apical loads

The frequency of free oscillations of plant stems with apical loads, as found in some cereals, is different depending on whether the stems are oriented vertically or horizontally. Neglecting the stem's own weight the differential equations describing the oscillation can be solved for both cases, although in the vertical orientation only for a limited set of conditions including constant bending stiffness along the stem. Comparison with experimental data shows that the difference between the oscillation frequencies in vertical and horizontal orientations can be attributed to the fact that in the vertical orientation the top load due to gravity induces a bending moment varying with the oscillation, while in the horizontal case this bending moment is nearly constant.     

Temporal and topographic characteristics of evoked potentials in the conflict of two consecutive visual stimuli in a working memory task

Behavioral reactions and brain mechanisms involved in processing two matching or mismatching (conflicting) visual stimuli were studied in healthy subjects (mean age 22.57 ± 0.46 years). Line orientations (vertical, horizontal, or 45°) were used as stimuli and were presented with an interval of 1500–1800 ms. The reaction time was shown to increase in the case of a conflict of two orientations as compared with matching orientations. The reaction time depended on the orientation of the reference stimulus and was minimal when a vertical line was used as a reference. An increase in N2 negativity (time window 200–280 ms) in the frontal and parietal cortical areas was identified as an informative indicator of a conflict between the current orientation and the orientation stored in working memory. The dipole sources of N2 were localized to the prefrontal cortex (middle frontal gyrus, frontal pole, and pars orbitalis). The N2 amplitude was found to depend on the orientation of the first stimulus in a pair, being higher in the case of a 45° orientation. The visual areas were shown to play a role in detecting a conflict of two consecutive signals because the early sensory components increased in amplitude. The results implicate cortical structures, including the sensory-specific visual, parietal, and prefrontal areas, in comparing consecutive visual signals and detecting their conflict.     

A computational model of visual anisotropy

Visual anisotropy has been demonstrated in multiple tasks where performance differs between vertical, horizontal, and oblique orientations of the stimuli. We explain some principles of visual anisotropy by anisotropic smoothing, which is based on a variation on Koenderink's approach in [1]. We tested the theory by presenting gaussian elongated luminance profiles and measuring the perceived orientations by means of an adjustment task. Our framework is based on the smoothing of the image with elliptical gaussian kernels and it correctly predicted an illusory orientation bias towards the vertical axis. We discuss the scope of the theory in the context of other anisotropies in perception.     

Atomic force microscopy of supported lipid membranes and their complexes with polycations

The method of atomic force microscopy has been used to investigate the morphology of mica-supported bilayer lipid membranes and stability of their complexes with a cationic polymer, poly-(N-ethyl-4-vinylpyridinium bromide). Lipid bilayers with a minimum of defects were obtained by the fusion of monolamellar neutral or mixed anionic bilayer vesicles (liposomes) on the mica surface, followed by excessive solvent removal by means of rapid rotation of a plate in horizontal plane (spin-coating). It has been shown that the cationic polymer does not interact with the bilayers, where the outer leaflet (i.e., the monolayer exposed to the surrounding aqueous solution) is made of an electroneutral phosphatidylcholine (PC). At the same time, the polymer irreversibly binds to the bilayer containing an anionic lipid.     

Anisotropy in the chromatic channel: a horizontal-vertical effect

We compared the chromatic contrast thresholds of drifting (2Hz) red-green sine-wave gratings of horizontal, vertical, and two oblique orientations at three spatial frequencies (2, 4, 8 cpd). Luminance contrast thresholds for yellow-black gratings were also obtained. The classic oblique effect was found for high spatial frequency luminance and chromatic stimuli. For chromatic thresholds, a significant difference was found between the horizontal and vertical thresholds of all observers. One observer was retested with her head tilted 45 deg and demonstrated that the anisotropy was specific to retinal coordinates. These results give evidence for orientation selectivity in the chromatic channel which is at least partially independent of that in the luminance channel. We estimated the degree of lateral chromatic aberration in our observers' eyes and discuss the possible contribution of this aberration to the horizontal-vertical difference in the chromatic channel.     

Modelling of the UV Index on vertical and 40° tilted planes for different orientations

In this study, estimated data of the UV Index on vertical planes are presented for the latitude of Valencia, Spain. For that purpose, the UVER values have been generated on vertical planes by means of four different geometrical models a) isotropic, b) Perez, c) Gueymard, d) Muneer, based on values of the global horizontal UVER and the diffuse horizontal UVER, measured experimentally. The UVER values, obtained by any model, overestimate the experimental values for all orientations, with the exception of the Perez model for the East plane. The results show statistical values of the MAD parameter (Mean Absolute Deviation) between 10% and 25%, the Perez model being the one that obtained a lower MAD for all levels. As for the statistic RMSD parameter (Root Mean Square Deviation), the results show values between 17% and 32%, and again the Perez model provides the best results in all vertical planes. The difference between the estimated UV Index and the experimental UV Index, for vertical and 40° tilted planes, was also calculated. 40° is an angle close to the latitude of Burjassot, Valencia, (39.5°), which, according to various studies, is the optimum angle to capture maximum radiation on tilted planes. We conclude that the models provide a good estimate of the UV Index, as they coincide or differ in one unit compared to the experimental values in 99% of cases, and this is valid for all orientations. Finally, we examined the relation between the UV Index on vertical and 40° tilted planes, both the experimental and estimated by the Perez model, and the experimental UV Index on a horizontal plane at 12 GMT. Based on the results, we can conclude that it is possible to estimate with a good approximation the UV Index on vertical and 40° tilted planes in different directions on the basis of the experimental horizontal UVI value, thus justifying the interest of this study.     

A mechanism underlying stimulation and inhibition of protein kinase C by lyso-PC: A role of membrane physical state

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Melittin-induced bilayer leakage depends on lipid material properties: evidence for toroidal pores

The membrane-lytic peptide melittin has previously been shown to form pores in lipid bilayers that have been described in terms of two different structural models. In the "barrel stave" model the bilayer remains more or less flat, with the peptides penetrating across the bilayer hydrocarbon region and aggregating to form a pore, whereas in the "toroidal pore" melittin induces defects in the bilayer such that the bilayer bends sharply inward to form a pore lined by both peptides and lipid headgroups. Here we test these models by measuring both the free energy of melittin transfer (DeltaG degrees ) and melittin-induced leakage as a function of bilayer elastic (material) properties that determine the energetics of bilayer bending, including the area compressibility modulus (K(a)), bilayer bending modulus (k(c)), and monolayer spontaneous curvature (R(o)). The addition of cholesterol to phosphatidylcholine (PC) bilayers, which increases K(a) and k(c), decreases both DeltaG degrees and the melittin-induced vesicle leakage. In contrast, the addition to PC bilayers of molecules with either positive R(o), such as lysoPC, or negative R(o), such as dioleoylglycerol, has little effect on DeltaG degrees , but produces large changes in melittin-induced leakage, from 86% for 8:2 PC/lysoPC to 18% for 8:2 PC/dioleoylglycerol. We observe linear relationships between melittin-induced leakage and both K(a) and 1/R(o)(2). However, in contrast to what would be expected for a barrel stave model, there is no correlation between observed leakage and bilayer hydrocarbon thickness. All of these results demonstrate the importance of bilayer material properties on melittin-induced leakage and indicate that the melittin-induced pores are defects in the bilayer lined in part by lipid molecules.     

Anisotropies in peripheral vernier acuity

Vernier acuity for short horizontal, vertical and oblique target lines was measured in many locations in the periphery of the visual field in normal human observers. In the 10 deg periphery, the average alignment threshold with oblique vernier lines in eight locations for three observers was 2.29 times higher than that with vertical and horizontal target lines. This oblique effect was found everywhere in the visual field. Similar conclusions are drawn for configurations in which the lines were replaced by just their distal endpoints, but here, additionally, performance was distinctly better when the dot pair was collinear with the fixation point, i.e. oriented radially, than when it was oriented tangentially. Both for vernier lines and for dot pairs, in all observers, horizontal configurations showed somewhat better thresholds than vertical ones. These results suggest an inherent pattern of connectivity throughout the visual field favoring processing in the cardinal orientations over the obliques, the radial over the tangential and, to a limited extent, the horizontal over the vertical.