Increase In Membrane Microviscosity Research Articles

Effect of colchicine on physiological and biochemical properties of Rhodococcus qingshengii.

The genus Rhodococcus includes polymorphic non-spore-forming gram-positive bacteria belonging to the class Actinobacteria. Together with Mycobacterium and Corynebacterium, Rhodococcus belongs to the Mycolata group. Due to their relatively high growth rate and ability to form biof ilms, Rhodococcus are a convenient model for studying the effect of biologically active compounds on pathogenic Mycolata. Colchicine was previously found to reduce biof ilm formation by P. carotovorum VKM B-1247 and R. qingshengii VKM Ac-2784D. To understand the mechanism of action of this alkaloid on the bacterial cell, we have studied the change in the fatty acid composition and microviscosity of the R. qingshengii VKM Ac-2784D membrane. Nystatin, which is known to reduce membrane microviscosity, is used as a positive control. It has been found that colchicine at concentrations of 0.01 and 0.03 g/l and nystatin (0.03 g/l) have no signif icant effect on the survival of R. qingshengii VKM Ac-2784D cultivated in a buffered saline solution with 0.5 % glucose (GBSS). However, colchicine (0.03 g/l) signif icantly inhibits biof ilm formation. Rhodococcus cells cultivated for 24 hours in GBSS with colchicine acquire a rounded shape. Colchicine at 0.01 g/l concentration increases C16:1(n-7), C17:0, C20:1(n-9) and C21:0 fatty acids. The microviscosity of the membrane of individual cells was distributed from the lowest to the highest values of the generalized laurdan f luorescence polarization index (GP), which indicates a variety of adaptive responses to this alkaloid. At a higher concentration of colchicine (0.03 g/l) in the membranes of R. qingshengii VKM Ac-2784D cells, the content of saturated fatty acids increases and the content of branched fatty acids decreases. This contributes to an increase in membrane microviscosity, which is conf irmed by the data on the GP f luorescence of laurdan. All of the above indicates that colchicine induces a rearrangement of the Rhodococcus cell membrane, probably in the direction of increasing its microviscosity. This may be one of the reasons for the negative effect of colchicine on the formation of R. qingshengii VKM Ac-2784D biof ilms.

Modulation of Membrane Microviscosity by Protein-Mediated Carotenoid Delivery as Revealed by Time-Resolved Fluorescence Anisotropy.

Carotenoids are potent antioxidants with a wide range of biomedical applications. However, their delivery into human cells is challenging and relatively inefficient. While the use of natural water-soluble carotenoproteins capable to reversibly bind carotenoids and transfer them into membranes is promising, the quantitative estimation of the delivery remains unclear. In the present work, we studied echinenone (ECN) delivery by cyanobacterial carotenoprotein AnaCTDH (C-terminal domain homolog of the Orange Carotenoid Protein from Anabaena), into liposome membranes labelled with BODIPY fluorescent probe. We observed that addition of AnaCTDH-ECN to liposomes led to the significant changes in the fast-kinetic component of the fluorescence decay curve, pointing on the dipole-dipole interactions between the probe and ECN within the membrane. It may serve as an indirect evidence of ECN delivery into membrane. To study the delivery in detail, we carried out molecular dynamics modeling of the localization of ECN within the lipid bilayer and calculate its orientation factor. Next, we exploited FRET to assess concentration of ECN delivered by AnaCTDH. Finally, we used time-resolved fluorescence anisotropy to assess changes in microviscosity of liposomal membranes. Incorporation of liposomes with β-carotene increased membrane microviscosity while the effect of astaxanthin and its mono- and diester forms was less pronounced. At temperatures below 30 °C addition of AnaCTDH-ECN increased membrane microviscosity in a concentration-dependent manner, supporting the protein-mediated carotenoid delivery mechanism. Combining all data, we propose FRET-based analysis and assessment of membrane microviscosity as potent approaches to characterize the efficiency of carotenoids delivery into membranes.

Transport of Ca2+ and Ca2+-dependent permeability transition in heart mitochondria in the early stages of Duchenne muscular dystrophy

Duchenne muscular dystrophy (DMD) is a progressive skeletal muscle disease that is associated with severe cardiac complications in the late stages. Significant mitochondrial dysfunction is reportedly responsible for the development of cardiomyopathy with age. At the same time, adaptive changes in mitochondrial metabolism in cardiomyocytes were identified in the early stages of DMD. In this work, we evaluate the functioning of calcium transport systems (MCU and NCLX), and MPT pore in the heart mitochondria of young dystrophin-deficient mice. As compared to wild-type animals, heart mitochondria of mdx mice have been found to be more efficient both in respect to Ca2+ uniport and Na+-dependent Ca2+ efflux. The data obtained indicate that the increased rate of Ca2+ uptake by heart mitochondria of mdx mice may be due to an increase in the ratio of MCU and MCUb subunits. In turn, an increase in the rate of Ca2+ efflux from organelles in DMD may be the result of a significant increase in the level of NCLX. Moreover, the heart mitochondria of mdx mice were more resistant to MPT pore opening, which may be due to an increase in the microviscosity of mitochondrial membranes of DMD mice. At the same time, the level of putative MPT pore proteins did not change. The paper discusses the effect of rearrangements of the mitochondrial proteome involved in the transport and accumulation of calcium on the adaptation of this organ to DMD.

Study of cytotoxicity performance of carbon nanohorns by method of spin probes

The effects of as-produced and treated by HNO3(3M) carbon nanohorns on the microviscosity of rat erythrocyte membranes and the viscosity of the water-containing plasma protein matrix were investigated by the method of spin probes. Addition of nanohorns at the concentration of 100 μg/ml to a suspension of erythrocytes led to an increase in membrane microviscosity during 4 h (about 60% effect). In addition, it was shown that nanohorns also induced an increased polarity of the microenvironment for lipophilic probes in the outer layer of membrane phospholipids, as well as disorders in erythrocytes membranes. Addition of nanohorns to plasma led to a little decrease in the viscosity of water and protein matrix, apparently, due to its partial destruction, impacting especially albumin. Pristine and treated by HNO3(3M) acid nanohorns was found more cytotoxic than nanoparticles of oxidized graphene, and significantly less than carbon nanotubes, which are known to dramatically increase the microviscosity of the membranes of erythrocytes and disrupt their integrity.

Changes in the spectrum of proteins and phospholipids in tissues of rats exposed to thiosulfonates

Esters of thiosulfoacids demonstrate a wide range of biological activity. One of their effects is the influence on the metabolism of proteins and lipids in the body. Therefore, the purpose of our experiment was to study the impact of synthesized thiosulfonates on the total content of proteins and phospholipids, as well as their spectrum in the blood, liver, and kidney of rats. For the experiment, allyl, ethyl, and methyl esters of thiosulfoacid were used. The protein profile of rat tissues was investigated by electrophoresis, and the ratio of different fractions of phospholipids by thin-layer chromatography. Our results have shown that short-term administration of thiosulfonates in a dose of 300 mg/kg of body weight did not cause significant changes in the content of total protein and its fractions in liver tissue, whereas the effect of allyl and ethyl esters of thiosulfoacid was accompanied by an increase in the total protein and albumin in the blood plasma. The decrease in total protein was found in the kidney tissue of rats injected with allyl- and methyl thiosulfonates. The newly synthesized compounds did not lead to significant changes in the total content of phospholipids in blood plasma and tissues of rats, except for methyl thiosulfonate, the effect of which was accompanied by an increase in the total phospholipids in the liver of rats. These data may indicate an adaptive reaction of the rat’s organism. Tissue-specific features of the phospholipid spectrum were detected in rats after short-term exposure to thiosulfonates. The most significant effect on the phospholipid profile in the blood was shown for allyl- and ethyl esters of thiosulfoacid. Their action was accompanied by a decrease in the phosphatidylserine and phosphatidylinositol fractions, while phosphatidylethanolamine and phosphatidylcholine increased, respectively. Esters of thiosulfoacid significantly influenced the ratio of different fractions of phospholipids in the liver and kidney tissues. The phospholipid composition of the liver was more influenced by the allyl and methyl esters of thiosulfoacid, whereas for the kidney tissue a greater effect was observed for ethyl and methyl esters. Thus, the action of allyl ester of thiosulfoacid caused a decrease in the asymmetry coefficient of hepatocyte membranes, indicating an elevation of the lipid bilayer saturation and the increase of membrane microviscosity. Similar changes were found in the kidney of rats treated with allyl- and ethyl thiosulfonates.

Studies of the blood antioxidant system and oxygen-transporting properties of human erythrocytes during 105-day isolation

Effects of strict 105-day isolation on the blood antioxidant status, processes in erythrocyte membranes, and oxygen-binding properties of hemoglobin were studied in six male volunteers (25 to 40 years old) in ground-based simulation of a mission to Mars (Mars-105 experiment). The study was performed with venous blood samples and red blood cells isolated from them, which were collected during the baseline data collection period, on days 35, 70, and 105 of the experiment, and on days 7 and 14–15 after its completion. Biochemical (determination of enzyme activity and thin-layer chromatography) and biophysical (laser interference microscopy and Raman spectroscopy) methods showed changes in the relative content of lipid and phospholipid fractions, suggesting an increase in the membrane microviscosity and the content of TBA-RP (active lipid peroxidation products interacting with thiobarbituric acid). A significant increase in the activity of glucose-6-phosphate dehydrogenase and superoxide dismutase and a reduction in the catalase activity was found, which indicates both reparative processes in red blood cells and imbalance between the amount of generated reactive oxygen species and antioxidant protection mechanisms in cells. The hemoglobin affinity for oxygen and the blood level of oxyhemoglobin also increased. It is assumed that the adaptation of the body to stresses experienced during and after the experiment can disturb the balance between the antioxidant defense systems. The latter, in turn, leads to peroxidation of membrane phospholipids, alteration in their content, increase in membrane microviscosity, and eventual disturbance of the gas-exchange function of red blood cells.

Toxic Mechanisms Induced by Fumonisin B1 Mycotoxin on Human Intestinal Cell Line

The gastrointestinal tract is the main target of exposure to mycotoxin fumonisin B1 (FB1), common natural contaminant in food. Previous studies reported that proliferating cells are more sensitive than confluent cells to the toxic effect of FB1. This study aims to investigate, by dose- and time-dependent experiments on human colon proliferating intestinal cell line (HT-29), the modifications induced by FB1 at concentrations ranging from 0.25 to 69μM. The choice of highest FB1 concentration considered the low toxicity previously reported on intestinal cell lines, whereas the lowest one corresponded to the lower FBs levels permitted by European Commission Regulation. Different functional parameters were tested such as cell proliferation, oxidative status, immunomodulatory effect and changes in membrane microviscosity. In addition FB1-FITC localization in this cell line was assessed by using confocal laser scanning microscopy. Lipid peroxidation induction was the main and early (12h) effect induced by FB1 at concentrations ranging from 0.5 to 69μM, followed by inhibition of cell proliferation (up to 8.6μM), the immunomodulatory effect (up to 17.2μM), by assessing IL-8 secretion, and increase in membrane microviscosity (up to 34.5μM). The toxic effects observed in different functional parameters were not dose-dependent and could be the consequence of the FB1 intracytoplasmatic localization as confirmed by confocal microscopy results. The different timescales and concentrations active of different functional parameters could suggest different cellular targets of FB1.

A likely role for the PH-domain containing protein, PEPP2/ PLEKHA5, at the membrane-microtubule cytoskeleton interface

PH (pleckstrin homology) domains are well known to bind membrane phosphoinositides with different specificities and direct PH domain-containing proteins to discrete subcellular compartments with as - sistances of alternative binding partners. PH domain-containing proteins have been found to be involved in a wide range of cellular events, including signalling, cytoskeleton rearrangement and vesicular trafficking. Here we showed that a novel PH domain-containing protein, PEPP2 (also known as PLEKHA5), displays mode- rate phosphoinositide binding specificity. Full length PEPP2 was observed to variably associate with both the plasma membrane and microtubules. The membrane-associated PEPP2 nucleated at cell-cell contacts and the leading edge of migrating cells. Overexpression of PEPP2 increased membrane microviscosity, in- dicating a potential role for PEPP2 in regulating function of microtubule-dependent membrane functions.

RETRACTED: A likely role for a novel PH-domain containing protein, PEPP2, in connecting membrane and cytoskeleton

PH domains (pleckstrin homology) are well known to bind membrane phosphoinositides with different specificities and direct PH domain-containing proteins to discrete subcellular apartments with assistances of alternative binding partners. PH domain-containing proteins are found to be involved in a wide range of cellular events, including signalling, cytoskeleton rearrangement and vesicular trafficking. Here we showed that a novel PH domain-containing protein, PEPP2, displayed moderate phosphoinositide binding specificity. Full length PEPP2 associated with both plasma membrane and microtubules. The membrane-associated PEPP2 nucleated at cell-cell contacts and the leading edge of migrating cells. Overexpression of PEPP2 increased membrane microviscosity, indicating a potential role of PEPP2 in regulating function of membrane and microtubules.

Mechanism of erythrocyte death in human population exposed to arsenic through drinking water

Arsenic contamination in drinking water is one of the biggest natural calamities, which has become an imperative threat to human health throughout the world. Abbreviation of erythrocyte lifespan leading to the development of anemia is a common sequel in arsenic exposed population. This study was undertaken to explore the mechanism of cell death in human erythrocytes during chronic arsenic exposure. Results revealed transformation of smooth discoid red cells into evaginated echinocytic form in the exposed individuals. Further distortion converted reversible echinocytes to irreversible spheroechinocytes. Arsenic toxicity increased membrane microviscosity along with an elevation of cholesterol/phospholipid ratio, which hampered the flexibility of red cell membrane and made them less deformable. Significant increase in the binding of merocyanine 540 with erythrocyte membrane due to arsenic exposure indicated disruption of lipid packing in the outer leaflet of the cell membrane resulting from altered transbilayer phospholipid asymmetry. Arsenic induced eryptosis was characterized by cell shrinkage and exposure of phosphatidylserine at the cell surface. Furthermore, metabolic starvation with depletion of cellular ATP triggered apoptotic removal of erythrocytes from circulation. Significant decrease in reduced glutathione content indicating defective antioxidant capacity was coupled with enhancement of malondialdehyde and protein carbonyl levels, which pointed to oxidative damage to erythrocyte membrane. Arsenic toxicity intervened into red cell membrane integrity eventually leading to membrane destabilization and hemoglobin release. The study depicted the involvement of both erythrophagocytosis and hemolysis in the destruction of human erythrocytes during chronic arsenic exposure.

Polarization of Plasma Membrane Microviscosity during Endothelial Cell Migration

Cell movement is characterized by anterior-posterior polarization of multiple cell structures. We show here that the plasma membrane is polarized in moving endothelial cells (EC); in particular, plasma membrane microviscosity (PMM) is increased at the cell leading edge. Our studies indicate that cholesterol has an important role in generation of this microviscosity gradient. In vitro studies using synthetic lipid vesicles show that membrane microviscosity has a substantial and biphasic influence on actin dynamics; a small amount of cholesterol increases actin-mediated vesicle deformation, whereas a large amount completely inhibits deformation. Experiments in migrating ECs confirm the important role of PMM on actin dynamics. Angiogenic growth factor-stimulated cells exhibit substantially increased membrane microviscosity at the cell front but, unexpectedly, show decreased rates of actin polymerization. Our results suggest that increased PMM in lamellipodia may permit more productive actin filament and meshwork formation, resulting in enhanced rates of cell movement.

Membrane microviscosity regulates endothelial cell motility.

Endothelial cell (EC) movement is an initiating and rate-limiting event in the neogenesis and repair of blood vessels. Here, we explore the hypothesis that microviscosity of the plasma membrane (PM) is a key physiological regulator of cell movement. Aortic ECs treated with membrane-active agents, such as alpha-tocopherol, cholesterol and lysophospholipids, exhibited a biphasic dependency on membrane microviscosity, in which moderate increases enhanced EC migration, but increases beyond a threshold markedly inhibited migration. Surprisingly, angiogenic growth factors, that is, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), also increased membrane microviscosity, as measured in live cells by fluorescence recovery after photobleaching (FRAP). The localization of Rac to the PM was modified in cells treated with membrane-active agents or growth factors, suggesting a molecular mechanism for how membrane microviscosity influences cell movement. Our data show that angiogenic growth factors, as well as certain lipophilic molecules, regulate cell motility through alterations in membrane properties and the consequent relocalization of critical signalling molecules to membranes.

Synthetic Miniprion PrP106

Elucidation of structure and biological properties of the prion protein scrapie (PrP(Sc)) is fundamental to an understanding of the mechanism of conformational transition of cellular (PrP(C)) into disease-specific isoforms and the pathogenesis of prion diseases. Unfortunately, the insolubility and heterogeneity of PrP(Sc) have limited these studies. The observation that a construct of 106 amino acids (termed PrP106 or miniprion), derived from mouse PrP and containing two deletions (Delta 23-88, Delta 141-176), becomes protease-resistant when expressed in scrapie-infected neuroblastoma cells and sustains prion replication when expressed in PrP(0/0) mice prompted us to generate a corresponding synthetic peptide (sPrP106) to be used for biochemical and cell culture studies. sPrP106 was obtained successfully with a straightforward procedure, which combines classical stepwise solid phase synthesis with a purification strategy based on transient labeling with a lipophilic chromatographic probe. sPrP106 readily adopted a beta-sheet structure, aggregated into branched filamentous structures without ultrastructural and tinctorial properties of amyloid, exhibited a proteinase K-resistant domain spanning residues 134-217, was highly toxic to primary neuronal cultures, and induced a remarkable increase in membrane microviscosity. These features are central properties of PrP(Sc) and make sPrP106 an excellent tool for investigating the molecular basis of the conformational conversion of PrP(C) into PrP(Sc) and prion disease pathogenesis.

Altered membrane fluidity and signal transduction in the platelets from patients of thrombotic stroke.

Several earlier studies have implicated platelet activation with the pathogenesis of thrombotic stroke. In this report we have studied the changes in membrane physical microenvironment and signal transduction in the platelets obtained from the patients with thrombotic stroke. Aggregation induced by the synthetic agonist thrombin receptor-activating peptide was significantly enhanced (p < 0.001) in the platelets obtained from the patients. Steady-state fluorescence anisotropy measurements using diphenylhexatriene reflected a significant increase in membrane microviscosity from 3.315 (+/- 0.103) in the control to 4.600 (+/- 0.119) in the stroke. Proteins of relative mobilities of 131, 100, 47 and 38 kDa were found to remain phosphorylated on tyrosine in the resting platelets obtained from thrombotic stroke patients while they were not phosphorylated in the control counter-parts. Besides, calpain, a calcium dependent thiol protease present in the platelets, was found to remain active in this disease as reflected from the proteolysis of calpain substrates. Taken together, these data indicated abnormal circulating platelets in the patients ofthrombotic stroke. which could contribute to the etiopathogenesis of this disease.

Protective effect of erythropoietin on the oxidative damage of erythrocyte membrane by hydroxyl radical

Treatment of red blood cells with the copper (II) ascorbate system causes increased lipid peroxidation, increased membrane microviscosity, and phospholipid translocation with a concurrent decrease in cytosolic catalase and glutathione peroxidase activities. All these changes are prevented if the cells are treated with erythropoietin prior to the exposure to copper (II) ascorbate. The present investigation further indicates that the oxidative damage brought about by copper (II) ascorbate is due to generation of hydroxyl radical and that erythropoietin plays a unique role in protecting the membrane from oxidative damage.

Membrane microviscosity modulates mu-opioid receptor conformational transitions and agonist efficacy.

The influence of membrane microviscosity on mu-opioid agonist and antagonist binding, as well as agonist efficacy, was examined in membranes prepared from SH-SY5Y cells and from a C6 glioma cell line stably expressing the rat mu-opioid receptor (C6mu). Addition of cholesteryl hemisuccinate (CHS) to cell membranes increased membrane microviscosity and reduced the inhibitory effect of sodium and guanine nucleotides on the affinity of the full agonists sufentanil and [D-Ala2,N-MePhe4,Gly-ol5]enkephalin (DAMGO) for the mu-opioid receptor. Binding of the antagonists [3H]naltrexone and [3H]diprenorphine and the partial agonist nalbuphine was unaffected by CHS. The effect of CHS on agonist binding was reversed by subsequent addition of cis-vaccenic acid, suggesting that the effect of CHS is the result of increased membrane microviscosity and not a specific sterol-receptor interaction. CHS addition increased the potency of DAMGO to stimulate guanosine-5'-O-(3-[35S]thio)triphosphate binding by fourfold, whereas the potency of nalbuphine was unaffected. However, nalbuphine efficacy relative to that of the full agonist DAMGO was strongly increased in CHS-treated membranes compared with that in control membranes. Membrane rigidification also resulted in an increased efficacy for the partial agonists meperidine, profadol, and butorphanol relative to that of DAMGO as measured by agonist-stimulated GTPase activity in control and CHS-modified membranes. These findings support a regulatory role for membrane microviscosity in receptor-mediated G protein activation.

A Neurotoxic and Gliotrophic Fragment of the Prion Protein Increases Plasma Membrane Microviscosity

Prion-related encephalopathies are characterized by astrogliosis and nerve cell degeneration and loss. These lesions might be the consequence of an interaction between the abnormal isoform of the cellular prion protein that accumulates in nervous tissue and the plasma membranes. Previously we found that a synthetic peptide, homologous to residues 106–126 of the human prion protein, is fibrillogenic and toxic to neurons and trophic to astrocytes in vitro.This study dealt with the ability of the peptide to interact with membranes. Accordingly, we compared PrP 106–126 with different synthetic PrP peptides (PrP 89–106, PrP 127–147, a peptide with a scrambled sequence of 106–126, and PrP 106–126 amidated at the C-terminus) as to the ability to increase the microviscosity of artificial and natural membranes. The first three had no effect on nerve and glial cells in vitro,whereas the amidated peptide caused neuronal death. Using a fluorescent probe that becomes incorporated into the hydrocarbon core of the lipid bilayer and records the lipid fluidity, we found PrP 106–126 able to increase significantly the membrane microviscosity of liposomes and of all cell lines investigated. This phenomenon was associated with the distribution of the peptide over the cell surface, but not with changes in the membrane lipid or protein content, or with membrane lipid phase transitions. Accordingly, we deduced that increased membrane microviscosity was unrelated to changes in the membrane native components and was the result of increased lipid density following PrP 106–126 embedding into the lipid bilayer. No control peptides had comparable effects on the membrane microviscosity, except PrP 106–126 amidated at the C-terminus. Since the latter was as neurotoxic, but not as fibrillogenic, as PrP 106–126, we argued that the ability of PrP 106–126 to increase membrane microviscosity was unrelated to the propensity of the peptide to raise fibrils. Rather, it could be connected with the primary structure of PrP 106–126, characterized by two opposing regions, one hydrophilic and the other hydrophobic, that enabled the peptide to interact with the lipid bilayer. Based on these findings, we speculated that the glial and nerve cell involvement occurring in prion-related encephalopathies might be caused by the interaction with the plasma membrane of a PrP 106–126-like fragment or of the sequence spanning residues 106–126 of the abnormal isoform of the prion protein.

Effect of L-α-glycerylphosphorylcholine on muscarinic receptors and membrane microviscosity of aged rat brain

1. 1. Old rats showed a significant decrease in the number of muscarinic M 1 receptors and a significant increase in membrane microviscosity in the striatum and hippocampus as compared to young animals. In contrast, no significant changes in the density of muscarinic M 2 receptors were observed with aging. 2. 2. Chronic treatment of aged rats with L-α-glycerylphosphorylcholine (L-α-GPC) restored the number of M 1 receptors to levels found in the striatum and hippocampus from young animals. The same treatment to aged rats partially restored membrane microviscosity in both regions studied and hence increased membrane fluidity. 3. 3. None of the major metabolites of L-α-GPC (choline, glycerophosphate or phosphorylcholine) was able to restore the number of striatal and hippocampal M 1 sites and membrane microviscosity of aged rats, neither did any of these treatments (including treatment with L-α-GPC) modify the level of m 1 receptors and microviscosity values in young rats.

Age-specific alterations in muscarinic stimulation of K +-evoked dopamine release from striatal slices by cholesterol and S-adenosyl- l-methionine

The present experiments were carried out in order to test the hypothesis that age-related signal transduction (ST) deficits may occur as a result of structural changes in the membrane that are reflected partially as increased membrane microviscosity. Oxotremorine (oxo) enhancement of K +-evoked release of dopamine (K +-ERDA) was examined in superfused striatal slices from mature (6 months) and old (24 months) Wistar rats incubated (1 or 4 h, 37°C) with graded concentrations of S-adenosyl- l-methionine (SAM) or cholesterol hemisuccinate (CHO) in a modified Krebs medium. Tissue was then assessed for one of the following: (a) the degree of oxo-enhanced K +-ERDA, (b) carbachol stimulated low K m GTPase activity, or (c) alterations in membrane microviscosity. In other experiments the tissue was incubated in CHO followed by SAM (or the reverse), and oxo-enhanced K +-ERDA examined. Results indicated that SAM treatment increased all the parameters in the striatal tissue from old animals, while CHO had selective, opposite effects in the striatal tissue obtained from young animals. CHO-SAM, or the reverse, produced the same pattern of results. These results suggest that ST deficits may involve age-related structural alterations in membranes that interfere with receptor-G protein coupling/uncoupling.

Altered membrane microviscosity in essential hypertension

To investigate the alterations in erythrocyte ghost membrane microviscosity in essential hypertensive patients and to determine the relationship between these changes and the sodium-lithium countertransport activity as a sensitive marker of membrane function. Forty-three normolipidaemic essential hypertensive patients (23 treated, 20 untreated) and 27 normotensive controls were studied. Patients were attending the hospital hypertension clinic or a local general practitioner's surgery. Erythrocyte sodium-lithium countertransport activity was measured. The Michaelis constant (Km) for extracellular sodium and maximal reaction velocity for sodium-lithium countertransport were measured in a subgroup consisting of 22 essential hypertensive patients and 11 normotensive controls. Erythrocyte membrane microviscosity was measured using fluorescence polarization anisotropy of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-[4-trimethylammoniumphenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH). There was no significant difference in the fluorescence polarization anisotropy of DPH or TMA-DPH between normotensive and essential hypertensive patients. However, the fluorescence polarization anisotropy of TMA-DPH was increased significantly (reflecting increased membrane microviscosity) in hypertensive patients with a family history of hypertension compared with in patients without a family history of hypertension. The standard sodium-lithium countertransport activity was elevated in essential hypertensive patients compared with normotensive controls, and the Km for sodium was significantly lower in patients with a family history of hypertension than in patients without a family history of hypertension. Patients with a family history of hypertension were clustered, with significantly lower Km for sodium and higher TMA-DPH anisotropies than either hypertensive patients without a family history of hypertension or normotensive controls. These findings suggest that a high membrane microviscosity affecting the outer region of the lipid bilayer is associated with altered sodium-lithium countertransport kinetics in a subgroup of essential hypertensive patients consisting of those with a family history of hypertension.