Distribution and fluidizing action of soluble and aggregated amyloid beta-peptide in rat synaptic plasma membranes.

The effects of soluble and aggregated amyloid beta-peptide (Abeta) on cortical synaptic plasma membrane (SPM) structure were examined using small angle x-ray diffraction and fluorescence spectroscopy approaches. Electron density profiles generated from the x-ray diffraction data demonstrated that soluble and aggregated Abeta1-40 peptides associated with distinct regions of the SPM. The width of the SPM samples, including surface hydration, was 84 A at 10 degrees C. Following addition of soluble Abeta1-40, there was a broad increase in electron density in the SPM hydrocarbon core +/-0-15 A from the membrane center, and a reduction in hydrocarbon core width by 6 A. By contrast, aggregated Abeta1-40 contributed electron density to the phospholipid headgroup/hydrated surface of the SPM +/-24-37 A from the membrane center, concomitant with an increase in molecular volume in the hydrocarbon core. The SPM interactions observed for Abeta1-40 were reproduced in a brain lipid membrane system. In contrast to Abeta1-40, aggregated Abeta1-42 intercalated into the lipid bilayer hydrocarbon core +/-0-12 A from the membrane center. Fluorescence experiments showed that both soluble and aggregated Abeta1-40 significantly increased SPM bulk and protein annular fluidity. Physico-chemical interactions of Abeta with the neuronal membrane may contribute to mechanisms of neurotoxicity, independent of specific receptor binding.     

Amyloid β-Peptides Increase Annular and Bulk Fluidity and Induce Lipid Peroxidation in Brain Synaptic Plasma Membranes

Abstract: Amyloid β-peptides (Aβ) may alter the neuronal membrane lipid environment by changing fluidity and inducing free radical lipid peroxidation. The effects of Aβ_1–40 and Aβ_25–35 on the fluidity of lipids adjacent to proteins (annular fluidity), bulk lipid fluidity, and lipid peroxidation were determined in rat synaptic plasma membranes (SPM). A fluorescent method based on radiationless energy transfer from tryptophan of SPM proteins to pyrene and pyrene monomer-eximer formation was used to determine SPM annular fluidity and bulk fluidity, respectively. Lipid peroxidation was determined by the thiobarbituric acid assay. Annular fluidity and bulk fluidity of SPM were increased significantly ( p ≤ 0.02) by Aβ_1–40. Similar effects on fluidity were observed for Aβ_25–35 ( p ≤ 0.002). Increased fluidity was associated with lipid peroxidation. Both Aβ peptides significantly increased ( p ≤ 0.006) the amount of malondialdehyde in SPM. The addition of a water-soluble analogue of vitamin E (Trolox) inhibited effects of Aβ on lipid peroxidation and fluidity in SPM. The fluidizing action of Aβ peptides on SPM may be due to the induction of lipid peroxidation by those peptides. Aβ-induced changes in neuronal function, such as ion flux and enzyme activity, that have been reported previously may result from the combined effects of lipid peroxidation and increased membrane fluidity.     

Expression of alpha-synuclein in different brain parts of adult and aged rats.

The synucleins are a family of presynaptic proteins that are abundant in neurons and include alpha-, beta, and gamma-synuclein. Alpha-synuclein (ASN) is involved in several neurodegenerative age-related disorders but its relevance in physiological aging is unknown. In the present study we investigated the expression of ASN mRNA and protein in the different brain parts of the adult (4-month-old) and aged (24-month-old) rats by using RT-PCR technique and Western blot, respectively. Our results indicated that mRNA expression and immunoreactivity of ASN is similar in brain cortex, hippocampus and striatum but markedly lower in cerebellum comparing to the other brain parts. Aging lowers ASN mRNA expression in striatum and cerebellum by about 40%. The immunoreactivity of ASN in synaptic plasma membranes (SPM) from aged brain cortex, hippocampus and cerebellum is significantly lower comparing to adult by 39%, 24% and 65%, respectively. Beta-synuclein (BSN) was not changed in aged brain comparing to adult. Age-related alteration of ASN may affect the nerve terminals structure and function.     

P400 protein is one of the major substrates for Ca2+/calmodulin-dependent protein kinase II in the postsynaptic density-enriched fraction isolated from rat cerebral cortex, hippocampus and cerebellum.

Concanavalin A-binding glycoprotein with 250 K M(r) found in the postsynaptic density (PSD)-enriched preparation (or synaptic cytoskeleton) from rat cerebellum was identified with P400 protein from the physicochemical properties and enrichment in the cerebellum. Proteins homologous to the cerebellar 250 K M(r) protein occurred in the PSD-enriched preparations from rat cerebral cortex and from hippocampus, although the contents in the preparations were very low. The 250 K M(r) proteins in the PSD-enriched preparations from cerebellum and from cerebrum were highly phosphorylated by Ca2+/calmodulin (CaM)-dependent protein kinase II. The protein of synaptic plasma membrane (SPM) and PSD-enriched fractions prepared from cerebral cortex were not phosphorylated by the cAMP-dependent protein kinase endogenous to the fractions, whereas the protein from cerebellum was done in SPM and PSD-enriched fractions. The facts suggest that P400 or P400-like protein is closely associated with Ca2+/CaM-dependent protein kinase II in the PSD-enriched preparations, especially in the preparation from cerebral cortex. Phosphorylation of the protein by Ca2+/CaM-dependent protein kinase II may play an important role in the postsynaptic function in both cerebellum and at least in some areas of cerebrum.     

The effect of bupivacaine·HCl on the physical properties of neuronal membranes

Fluorescent probe techniques were used to evaluate the effect of bupivacaine.HCl on the physical properties (transbilayer asymmetric lateral and rotational mobilities, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMVs) isolated from bovine cerebral cortex. An experimental procedure was used based on selective quenching of both 1,3-di(1-pyrenyl)propane (Py-3-Py) and 1,6-diphenyl-1,3,5-hexatriene (DPH) by trinitrophenyl groups, and radiationless energy transfer (RET) from the tryptophans of membrane proteins to Py-3-Py. Bupivacaine.HCl increased the bulk lateral and rotational mobilities, and annular lipid fluidity in SPMVs lipid bilayers, and had a greater fluidizing effect on the inner monolayer than that of the outer monolayer. The magnitude of increasing effect on annular lipid fluidity in SPMVs lipid bilayer induced by bupivacaine.HCl was significantly far greater than magnitude of increasing effect of the drug on the lateral and rotational mobilities of bulk SPMVs lipid bilayer. It also caused membrane proteins to cluster. These effects of bupivacaine.HCl on neuronal membranes may be responsible for some, though not all, of the local anesthetic actions of bupivacaine.HCl.     

Cerebral microvascular and parenchymal phospholipid composition in the mouse

Cerebral microvessels consisting predominantly of capillaries and small arterioles (<30 m dia.) were isolated from the cerebral cortex and cerebellum of 3-month-old mice. Lipids were extracted from both microvascular and brain parenchymal fractions and the major phospholipid classes (choline phosphoglyceride, ethanolamine phosphoglyceride, inositol phosphoglyceride, serine phosphoglyceride, and sphingomyelin) separated by 2-dimensional TLC. Comparison of mol % determined by phosphate analysis of each phospholipid revealed significant differences in membrane composition of ethanolamine phosphoglyceride, inositol phosphoglyceride, and sphingomyelin between microvascular and parenchymal components of the central nervous system. Moreover, the choline phosphoglyceride/sphingomyelin mol ratio, one of three determinants of membrane fluidity, is significantly lower for microvessel membrane than for membranes of the brain parenchyma.     

Corticosterone administration and lipid metabolism in brain regions during development.

Effect of corticosterone on lipid contents of different brain regions and the effect of age on the sensitivity of these regions to corticosterone have been studied. Corticosterone administration (40 mg/kg body wt, sc) to 17-day-old rat for 3 days led to significant decrease in phospholipid content of cerebellum and increase in cholesterol contents of hippocampus and striatum. However, there was no effect on cerebral cortex and brain stem lipids. This alteration in lipids was associated with decrease in [U-14C] glucose incorporation into cholesterol and phospholipids, decrease in plasma beta-hydroxy butyrate levels and increase in beta-hydroxy butyrate dehydrogenase activity in hippocampus and striatum, thereby suggesting that suppression of glucose utilization by corticosterone was compensated by higher utilization of ketone bodies for lipid synthesis in these regions. The sensitivity to corticosterone appears to be age-specific as, at 20-day, cerebellum, hippocampus and striatum were susceptible, at 10-day only hippocampus and at 40- and 90-day none of these regions responded to the treatment.     

Synaptic plasma membrane-bound acetylcholinesterase activity is not affected by docosahexaenoic acid-induced decrease in membrane order

We investigated the effect of administration of docosahexaenoic acid (C22:6, n-3; 300 mg/kg.day, for 12 weeks) on the degree of membrane order and membrane-bound acetylcholinesterase activity of the cerebral cortex synaptic plasma membrane in male Wistar rats. Docosahexaenoic acid levels in the synaptic plasma membrane increased significantly by 16% over levels in control rats concomitant with an increase in the molar ratio of docosahexaenoic acid to arachidonic acid. Synaptic plasma membrane order, assessed by 1,6-diphenyl-1,3,5-hexatriene, which measures order of the bulk internal hydrophobic lipid core, decreased significantly in the docosahexaenoic acid-fed rats. Lateral mobility of both global and annular lipids measured by pyrene also increased. Acetylcholinesterase activity of the synaptic plasma membrane was unaffected, and synaptic plasma membrane phospholipid contents increased in the docosahexaenoic acid-fed rats, with a concomitant decrease in the cholesterol/phospholipid molar ratio. Lipid peroxide and reactive oxygen species, indicators of tissue oxidative stress, decreased in both the cerebral cortex synaptosome and homogenate of the docosahexaenoic acid-fed rats. Arrhenius plot showed a break point in acetylcholinesterase activity at 22 degrees C and 24 degrees C in plasma membranes from docosahexaenoic acid-fed and control rats, respectively. The present experiment indicates that chronic administration of docosahexaenoic acid does not affect synaptic acetylcholinesterase activity and evoke oxidative stress, although it increases the disorder of the global and annular lipids of rat synaptic plasma membranes.     

The effect of propoxycaine.HCl on the physical properties of neuronal membranes

Fluorescent probe techniques were used to evaluate the effect of propoxycaine.HCl on the physical properties (transbilayer asymmetric lateral and rotational mobilities, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMVs) isolated from bovine cerebral cortex. An experimental procedure was used based on selective quenching of both 1,3-di(1-pyrenyl)propane (Py-3-Py) and 1,6-diphenyl-1,3,5-hexatriene (DPH) by trinitrophenyl groups, and radiationless energy transfer (RET) from the tryptophans of membrane proteins to Py-3-Py. Propoxycaine.HCl increased the bulk lateral and rotational mobilities, and annular lipid fluidity in SPMVs lipid bilayers, and had a greater fluidizing effect on the inner monolayer than that of the outer monolayer. The magnitude of increasing effect on annular lipid fluidity in SPMVs lipid bilayer induced by propoxycaine.HCl was significantly far greater than magnitude of increasing effect of the drug on the lateral and rotational mobilities of SPMVs lipid bilayer. It also caused membrane proteins to cluster. These effects of propoxycaine.HCl on neuronal membranes may be responsible for some, though not all, of the local anesthetic actions of propoxycaine.HCl.     

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.     

The effect of cholesterol and monosialoganglioside (GM1) on the release and aggregation of amyloid beta-peptide from liposomes prepared from brain membrane-like lipids

In order to investigate the influence of cholesterol (Ch) and monosialoganglioside (GM1) on the release and subsequent deposition/aggregation of amyloid beta peptide (Abeta)-(1-40) and Abeta-(1-42), we have examined Abeta peptide model membrane interactions by circular dichroism, turbidity measurements, and transmission electron microscopy (TEM). Model liposomes containing Abeta peptide and a lipid mixture composition similar to that found in the cerebral cortex membranes (CCM-lipid) have been prepared. In all, four Abeta-containing liposomes were investigated: CCM-lipid; liposomes with no GM1 (GM1-free lipid); those with no cholesterol (Ch-free lipid); liposomes with neither cholesterol nor GM1 (Ch-GM1-free lipid). In CCM liposomes, Abeta was rapidly released from membranes to form a well defined fibril structure. However, for the GM1-free lipid, Abeta was first released to yield a fibril structure about the membrane surface, then the membrane became disrupted resulting in the formation of small vesicles. In Ch-free lipid, a fibril structure with a phospholipid membrane-like shadow formed, but this differed from the well defined fibril structure seen for CCM-lipid. In Ch-GM1-free lipid, no fibril structure formed, possibly because of membrane solubilization by Abeta. The absence of fibril structure was noted at physiological extracellular pH (7.4) and also at liposomal/endosomal pH (5.5). Our results suggest a possible role for both Ch and GM1 in the membrane release of Abeta from brain lipid bilayers.     

Increasing Age Alters Transbilayer Fluidity and Cholesterol Asymmetry in Synaptic Plasma Membranes of Mice

Abstract: Previous studies examining age differences in membrane fluidity and cholesterol content have reported on the average or total change in membrane structure, respectively. However, a membrane consists of an exofacial leaflet and a cytofacial leaflet that differ in fluidity and cholesterol distribution. The purpose of the present experiments was to determine fluidity and cholesterol distribution of the exofacial and cytofacial leaflets of brain synaptic plasma membranes (SPMs) from 3–4-, 14–15-, and 24–25-month-old C57BL/6NNIA mice by using trinitrobenzenesulfonic acid (TNBS)-quenching techniques and fluorescent probes. The exofacial leaflet of SPMs from young mice was significantly more fluid compared with the cytofacial leaflet. The large difference in fluidity between the two leaflets was abolished in SPMs of the oldest age group. Total SPM cholesterol and the cholesterol-to-phospholipid molar ratio did not differ among the three different age groups of mice. However, considerable differences were observed in the distribution of cholesterol in the two SPM leaflets. The exofacial leaflet contained substantially less cholesterol than did the cytofacial leaflet (13 vs. 87%, respectively) in SPMs of young mice. This asymmetric distribution of cholesterol was significantly modified with increasing age. There was an approximately twofold increase in exofacial leaflet cholesterol in the oldest group compared with the youngest age group. Transbilayer fluidity and cholesterol asymmetry were altered in SPMs of older mice. This approach is a new and different way of viewing how aging modifies membrane structure. Age differences in SPM leaflet structure may be an important factor regulating activity of certain membrane proteins.     

Effects of dimyristoylphosphatidylethanol on the structural parameters of neuronal membranes

Fluorescent probes located in different membrane regions were used to evaluate the effects of dimyristoylphosphatidylethanol (DMPEt) on the structural parameters (transbilayer rotational and lateral mobility, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMV) from the bovine cerebral cortex. An experimental procedure was used based on selective quenching of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1,3-di(1-pyrenyl)propane (Py-3-Py) by trinitrophenyl groups, and radiationless energy transfer from the tryptophans of membrane proteins to Py-3-Py. DMPEt increased the bulk lateral and rotational mobility, and annular lipid fluidity of SPMV lipid bilayers, and had a greater fluidizing effect on the outer monolayer than the inner monolayer. It also caused membrane proteins to cluster. These effects of DMPEt on neuronal membranes may be responsible for some, though not all, of the general anesthetic actions of ethanol.     

Regional Heterogeneity of Cerebral Microvessels and Brain Susceptibility to Oxidative Stress

The hippocampus is one of the earliest and most affected regions in Alzheimer’s disease (AD), followed by the cortex while the cerebellum is largely spared. Importantly, endothelial dysfunction is a common feature of cerebral blood vessels in AD. In this study, we sought to determine if regional heterogeneity of cerebral microvessels might help explain the susceptibility of the hippocampus and cortex as compared to the cerebellum. We isolated microvessels from wild type mice from the cerebellum, cortex, and hippocampus to characterize their vascular phenotype. Superoxide anion was significantly higher in microvessels isolated from the cortex and hippocampus as compared to the cerebellum. Importantly, protein levels of NADPH oxidase (NOX)-2 and NOX-4 were significantly higher in the cortical and hippocampal microvessels as compared to microvessels from the cerebellum. In addition, expression of manganese superoxide dismutase protein was significantly lower in microvessels from the cortex and hippocampus as compared to cerebellum while other antioxidant enzymes were unchanged. There was no difference in eNOS protein expression between the microvessels of the three brain regions; however, bioavailability of tetrahydrobiopterin (BH₄), an essential cofactor for eNOS activity, was significantly reduced in microvessels from the hippocampus and cortex as compared to the cerebellum. Higher levels of superoxide and reduced tetrahydrobiopterin bioavailability may help explain the vulnerability of the hippocampus and cortical microvessels to oxidative stress and development of endothelial dysfunction.     

Evoked effects of cholesterol binding on integral proteins and lipid fluidity of dog brain synaptosomal plasma membranes

Binding of cholesterol into dog brain synaptosomal plasma membranes (SPM) within the limits of concentration used (0.5-5 microM) follows an exponential curve described by the general formula y = a.ebx. This curve, which represents the total binding (specific and nonspecific), acquires sigmoid character in the presence of 100 microM cholesterol glucoside, with a Hill coefficient of h = 2.98 +/- 0.18. The specific activity of the Na+/K+-transporting ATPase and Ca2+-transporting ATPase rose after a 2-h preincubation of SPM with cholesterol (up to 5 microM) or its glucoside (up to 50 microM) to at least 50% above their original values. Fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) increased with cholesterol glucoside (50 microM) incorporation. Cholesterol (5 microM) had no effect on the DPH fluorescence polarization. Arrhenius plots of Na+/K+-transporting ATPase activity exhibited a break point at 23.2 +/- 1.1 degrees C in control SPM, which was elevated to 29.5 +/- 1.4 degrees C in SPM treated with cholesterol glucoside (50 microM) and abolished in SPM treated with cholesterol (5 microM). The allosteric properties of SPM-bound Na+/K+-transporting ATPase inhibited by F- and Ca2+-transporting ATPase inhibited by Na+ (as reflected by changes in the Hill coefficient) were modulated by cholesterol. It could be stated that cholesterol glucoside (50 microM) produced an increased packing of the bulk lipids, while cholesterol (5 microM) increased the fluidity of the lipid microenvironment of both Na+/K+-transporting ATPase and Ca2+-transporting ATPase.     

Neural membrane phospholipids in alzheimer disease

Phospholipids form the backbone of neural membranes, providing fluidity and permeability. Two plasma membrane fractions, one from synaptosomes (SPM), the other glial and neuronal cell bodies (PM), were prepared from different regions of autopsied Alzheimer disease (AD) brains. Corresponding fractions were prepared from age-matched control brains. All fractions from AD brains showed significantly lower levels of ethanolamine glycerophospholipids and significantly higher levels of serine glycerophospholipids than the control brain. No differences were observed in phosphatidylcholine levels among these membranes. These results suggest that altered phospholipid composition of plasma membranes may be involved in the abnormal signal transduction and neurodegeneration in AD.Special issue dedicated to Dr. Leon S. Wolfe.     

Interaction between Alzheimer's Abeta(25-35) peptide and phospholipid bilayers: the role of cholesterol

There is mounting evidence that the lipid matrix of neuronal cell membranes plays an important role in the accumulation of beta-amyloid peptides into senile plaques, one of the hallmarks of Alzheimer's disease (AD). With the aim to clarify the molecular basis of the interaction between amyloid peptides and cellular membranes, we investigated the interaction between a cytotoxic fragment of Abeta(1-42), i.e., Abeta(25-35), and phospholipid bilayer membranes. These systems were studied by Electron Paramagnetic Resonance (EPR) spectroscopy, using phospholipids spin-labeled on the acyl chain. The effect of inclusion of charged phospholipids or/and cholesterol in the bilayer composition was considered in relation to the peptide/membrane interaction. The results show that Abeta(25-35) inserts in bilayers formed by the zwitterionic phospholipid dilauroyl phosphatidylcholine (DLPC), positioning between the outer part of the hydrophobic core and the external hydrophilic layer. This process is not significantly influenced by the inclusion of the anionic phospholipid phosphatidylglycerol (DLPG) in the bilayer, indicating the peptide insertion to be driven by hydrophobic rather than electrostatic interactions. Cholesterol plays a fundamental role in regulating the peptide/membrane association, inducing a membrane transition from a fluid-disordered to a fluid-ordered phase. At low cholesterol content, in the fluid-disordered phase, the insertion of the peptide in the membrane causes a displacement of cholesterol towards the more external part of the membrane. The crowding of cholesterol enhances its rigidifying effect on this region of the bilayer. Finally, the cholesterol-rich fluid-ordered membrane looses the ability to include Abeta(25-35).     

Amyloid beta-protein interactions with membranes and cholesterol: causes or casualties of Alzheimer's disease

Amyloid beta-protein (Abeta) is thought to be one of the primary factors causing neurodegeneration in Alzheimer's disease (AD). This protein is an amphipathic molecule that perturbs membranes, binds lipids and alters cell function. Several studies have reported that Abeta alters membrane fluidity but the direction of this effect has not been consistently observed and explanations for this lack of consistency are proposed. Cholesterol is a key component of membranes and cholesterol interacts with Abeta in a reciprocal manner. Abeta impacts on cholesterol homeostasis and modification of cholesterol levels alters Abeta expression. In addition, certain cholesterol lowering drugs (statins) appear to reduce the risk of AD in human subjects. However, the role of changes in the total amount of brain cholesterol in AD and the mechanisms of action of statins in lowering the risk of AD are unclear. Here we discuss data on membranes, cholesterol, Abeta and AD, and propose that modification of the transbilayer distribution of cholesterol in contrast to a change in the total amount of cholesterol provides a cooperative environment for Abeta synthesis and accumulation in membranes leading to cell dysfunction including disruption in cholesterol homeostasis.     

Transbilayer Distribution of Cholesterol Is Modified in Brain Synaptic Plasma Membranes of Knockout Mice Deficient in the Low-Density Lipoprotein Receptor, Apolipoprotein E, or Both Proteins

Abstract: Both apolipoprotein E (apoE) and the low-density lipoprotein (LDL) receptor are present in brain; however, little is known regarding the function of these proteins in brain, in particular with respect to brain cholesterol. The role of apoE and the LDL receptor in modulating the transbilayer or asymmetric distribution of cholesterol in the exofacial and cytofacial leaflets of synaptic plasma membranes (SPMs) was examined in mutant mice deficient in apoE, the LDL receptor, or both proteins by using the fluorescent sterol dehydroergosterol and fluorescent quenching procedures. Fluidity of the exofacial and cytofacial leaflets was also measured. Cholesterol asymmetry of SPMs was altered in the mutant mice, with the largest effect observed in the LDL receptor-deficient mice. There was an approximately twofold increase in the percent distribution of cholesterol in the exofacial leaflet of the LDL receptor-deficient mice (32%) compared with C57BL/6J mice (15%). Mice deficient in apoE or both proteins also showed a significantly higher percent distribution of cholesterol (23 and 26%, respectively) in the exofacial leaflet compared with the C57BL/6J mice. Although the percent distribution of cholesterol was highest in the exofacial leaflet of the LDL receptor-deficient mice, fluidity of the exofacial leaflet of that group was significantly lower. However, the cholesterol-to-phospholipid ratio of SPMs of the LDL receptor-deficient mice was significantly lower, and this difference was largely the result of a significant increase in the total amount of SPM phospholipid. This study demonstrates for the first time that SPM lipid structure is altered in mice deficient in apoE or the LDL receptor. Although the mechanism that maintains the asymmetric distribution of cholesterol in plasma membranes is not well understood, data of the present experiments indicate that both apoE and the LDL receptor are involved in maintaining the transbilayer distribution of cholesterol.     

Changes in cerebral membrane lipid composition and fluidity during thioacetamide-induced hepatic encephalopathy

Lipids are an essential structural and functional component of cellular membranes. Changes in membrane lipid composition are known to affect the activities of many membrane-associated enzymes, endocytosis, exocytosis, membrane fusion and neurotransmitter uptake, and have been implicated in the pathophysiology of many neurodegenerative disorders. In the present study, we investigated changes in the lipid composition of membranes isolated from the cerebral cortex of rats treated with thioacetamide (TAA), a hepatotoxin that induces fulminant hepatic failure (FHF) and thereon hepatic encephalopathy (HE). HE refers to acute neuropsychiatric changes accompanying FHF. The estimation of membrane phospholipids, cholesterol and fatty acid content in cerebral cortex membranes from TAA-treated rats revealed a decrease in cholesterol, phosphatidylserine, sphingomyelin, a monounsaturated fatty acid, namely oleic acid, and the polyunsaturated fatty acids gamma-linolenic acid, decosa hexanoic acid and arachidonic acid compared with controls. Assessment of membrane fluidity with pyrene, 1,6-diphenyl-1,3,5-hexatriene and 1-[4-(trimethylammonio)phenyl]-6-phenyl-1,3,5-hexatriene revealed a decrease in the annular membrane fluidity, whereas the global fluidity was unaffected. The level of the thiobarbituric acid reactive species marker for lipid peroxidation also increased in membranes from TAA-treated rats, thereby indicating the prevalence of oxidative stress. Results from the present study demonstrate gross alterations in cerebral cortical membrane lipid composition and fluidity during TAA-induced HE, and their possible implications in the pathogenesis of this condition are also discussed.