Avinoam Livne

Differential inhibition of mitochondrial monoamine oxidase from brain by hashish components

Abstract Monoamine oxidase (MAO) of porcine brain mitochondria was differentially affected by hashish components: with benzylamine as a substrate, Δ 1 -tetrahydrocannabinol (Δ 1 -THC) inhibited MAO activity markedly, while cannabidiol (CBD) was essentially innocuous at the same concentrations. When added concomitantly, CBD obviated the inhibitory effect of Δ 1 -THC. An extract of hashish was over 10-fold more inhibitory toward MAO than Δ 1 -THC on weight basis. A prior incubation of the mitochondrial preparation with the cannabis compounds was required to express the inhibitory effect. Liver mitochondrial MAO was not affected by either Δ 1 -THC, CBD or hashish extract, despite a prolonged preincubation period, thus demonstrating tissue selectivity with respect to the cannabinoid effect.

Differential effects of lipids on the osmotic fragility of erythrocytes

Abstract Stearic, oleic, linoleic and linolenic acids, the methyl esters of these acids, as well as their hydroxy analogs were tested for their potency to stabilize human erythrocytes against hypotonic hemolysis. The erythrocyte stabilization was affected by the polarity and the degree of unsaturation of the added compound and by the level of hemolysis employed. The stabilization afforded by the unsaturated fatty acids decreased with increasing number of double bonds while the opposite was true for the alcohol series. The interaction of both the polar and the hydrophobic portions of a stabilizing compound with the erythrocyte membrane contributes to the overall stabilization effect. To account for the differential effects of the lipids. is it proposed that some modifications of the membrane are being revealed on excessive swelling of the erythrocytes in a hypotonic medium.

Water permeability of bilayer lipid membranes: Sterol-lipid interaction.

SummaryBilayer lipid membranes were generated in an aqueous medium from synthetic, egg or plant phosphatidyl choline (PC) or from plant monogalactosyl diglyceride (MG). The water permeability of the black membranes was determined by measuring the net volume flux produced by a NaCl gradient. The osmotic permeability coefficient,Pos, was markedly affected by the number of double bonds in the fatty acid conjugates of the lipids: the greater the degree of unsaturation, the higher the value ofPos. The temperature dependence ofPos of the lipid membranes was studied over a range of 29 to 40°C. The experimental activation energy,Ea, estimated from the linear plots of log (Pos)versus 1/T, was significantly higher for MG membranes (17 kcal/mole) than for the various PC membranes (11 to 13 kcal/mole), probably owing to hydrogen bonding between MG and water molecules. In comparison with PC membranes, the membranes generated from PC and cholesterol (1∶1 molar ratio) had lowerPos but similarEa values. Likewise, either stigmasterol or β-sitosterol decreasedPos of MG membranes, whileEa was not affected by the sterols. MG-cholesterol membranes were specifically characterized by a unique value ofEa (−36 kcal/mole) thus indicating temperature dependent structural changes.

Cytoplasmic acidification and activation of Na+/H+ exchange during regulatory volume decrease in Ehrlich ascites tumor cells

SummaryEhrlich ascites tumor cells undergoing regulatory volume decrease (RVD) exhibit cytoplasmic acidification as measured by an intracellular fluorescent pH indicator. The acidification results in an activation of the Na+/H+ exchanger. The intracellular pH ‘set point’ for the activation is estimated to be around 7.0. The activation of the Na+/H+ exchanger leads to an incomplete RVD. In support of this conclusion, amiloride and Na+-free medium, known to limit the Na+/H+ exchange, indeed enhance the RVD response. Intracellular acidification and activation of Na+/H+ exchange may be a general response of cells undergoing RVD.

Initiation of RVD response in human platelets: mechanical-biochemical transduction involves pertussis-toxin-sensitive G protein and phospholipase A2.

Platelets revert hypotonic-induced swelling by the process of regulatory volume decrease (RVD). We have recently shown that this process is under the control of endogenous hepoxilin A3. In this work, we investigated the mechanical-biochemical transduction that leads to hepoxilin A3 formation. We demonstrate that this process is mediated by pertussis-toxin-sensitive G protein, which activates Ca2+-insensitive phospholipase A2, and the sequential release of arachidonic acid. This conclusion is supported by the following observations: (i) RVD response is blocked selectively by the phospholipase A2 inhibitors manoalide and bromophenacyl-bromide (0.2 and 5 μm, respectively) but not by phospholipase C inhibitors. The addition of arachidonic acid overcame this inhibition; (ii) extracellular Ca2+ depletion by EGTA (up to 10 mm) does not affect RVD; (iii) intracellular Ca2+ depletion by BAPTAAM (100 μm) inhibits RVD but not hepoxilin A3 formation, as tested by the RVD reconstitution assay; (iv) RVD is inhibited by the G-protein inhibitors, GDPβS (1 μm) and pertussis toxin (1 ng/ml). This inhibition is overcome by addition of arachidonic acid or hypotonic cell-free eluate that contains hepoxilin A3; (v) NaF, 1 mm, induces hepoxilin A3 formation, tested by the RVD reconstitution assay; and (vii) GDPβS inhibits hepoxilin A3 formation associated with flow. Therefore, it seems that G proteins are involved in the initial step of the mechanical-biochemical transduction leading to hepoxilin A3 formation in human platelets.

The erythrocyte membrane site for the effect of temperature on osmotic fragility

The osmotic fragility of human erythrocytes is well known to decrease as the temperature is elevated. The cellular site for the temperature effect was studied by assessing possible roles of hemoglobin and of membrane lipids and by taking advantage of the unique response of camel erythrocytes to temperature. It is concluded that the erythrocyte membrane is the site for the temperature effect on osmotic fragility. The human erythrocyte is likely to rupture in protein--lipid boundary regions in the membrane, from which cholesterol is apparently excluded.

Acetylcholine esterase as a probe for erythrocyte-membrane intactness

Abstract Erythrocyte acetylcholine esterase can be assayed in intact cells and was tested as a probe for membrane changes. Acetylcholine esterase activity correlated with the erythrocyte relative volume. Antihemolytic acyl sorbitols, fatty acids and phenothiazines inhibit to varying extents the activity of acetylcholine esterase. The inhibition of acetylcholine esterase by linolenoyl sorbitol was further characterized and found to be non-competitive and critically dependent on cell intactness over a wide temperature range. Neither solubilized nor ghost acetylcholine esterase was affected by the acyl sorbitol while under conditions optimal for ghost resealing, the enzyme resumed the sensitivity to the acyl sorbitol. Acetylcholine esterase sensitivity thus appears to be a promising tool to follow the dynamics of membrane integrity.

Unique properties of the camel erythrocyte membrane

Abstract In view of the exceptionally high osmotic stability of camel erythrocytes, several aspects of the membranes were examined. 1. 1. Added linolenoyl sorbitol increased the osmotic stability of camel erythrocytes to only one-third of the increase of human erythrocytes similarly treated. Human, hamster, rat and sheep erythrocytes exhibited similar time courses of lysis by sonic irradiation, while camel erythrocytes were distinctly more stable. Unlike its effect on human erythrocytes, vinblastin did not impair the osmotic stability of camel erythrocytes. 2. 2. Lipid composition of camel erythorcytes was analyzed. Lipid content per packed cell volume, phospholipid:cholesterol molar ratio, phospholipid composition as well as composition of the phospholipid fatty acids were all close to values reported for other species. 3. 3. Ghosts prepared from camel erythrocytes had a protein:lipid ratio of 3.0, compared with a ratio of 1.25 in human erythrocyte ghosts, and showed a higher proportion of proline and arginine but lower proportion of glutamic acid. 4. 4. It is proposed that the unique lipoprotein structure of the camel erythrocyte membrane may account for the unusual features of these erythrocytes.

The interaction of hashish components with human erythrocytes

Abstract At 10 −5 M the hashish components, Δ 1 -tetrahydrocannabinol and cannabidiol, stabilize human erythrocytes reversibly against hypotonic hemolysis. Δ 1 -Tetrahydrocannabinol expanded the erythrocyte membrane in a hypotonic medium more effectively than did cannabidiol.

Inositol-1-phosphatase in red blood cells of manic-depressive patients before and during treatment with lithium

The present study was designed to answer whether or not Li-free manic-depressive patients show similar enzyme activities as normal controls and whether or not the enzyme in RBC of manic depressive patients is inhibitable after in vivo treatment to the extent expected from in vitro studies