Alterations of human erythrocyte membrane fluidity by oxygen-derived free radicals and calcium

Two possible reasons for the structural alterations of cell membranes caused by free radicals are lipid peroxidation and an increase in the intracellular calcium ion concentration. To characterize the alterations in membrane molecular dynamics caused by oxygen-derived free radicals and calcium, human erythrocytes were spin-labeled with 5-doxyl stearic acid, and alterations in membrane fluidity were quantified by electron spin resonance oxidase (0.07 U/mL) decreased membrane fluidity, and the addition of superoxide dismutase and catalase inhibited the effect on membrane fluidity of the hypoxanthine-xanthine oxidase system. Hydrogen peroxide (0.1 and 1 nM) also decreased membrane fluidity and caused alterations to erythrocyte morphology. In addition, a decrease in membrane fluidity was observed in erythrocytes incubated with 2.8 mM CaCl₂. On the other hand, incubation of erythrocytes with calcium-free solution decreased the changes in membrane fluidity caused by hydrogen peroxide.

These results suggest that changes in membrane fluidity are directly due to lipid peroxidation and are indirectly the result of increased intracellular calcium concentration. We support the hypothesis that alterations of the biophysical properties of membranes caused by free radicals play an important role in cell injury, and that the accumulation of calcium amplifies the damge to membranes weakened by free radicals.     

Tissue water relations in elfin cloud forest tree species of Serrania de Macuira,Guajira, Colombia

Summary Diurnal courses of stomatal conductance, leaf water potential, and the components of tissue water potential were measured in six canopy species in an elfin cloud forest. High values of stomatal conductance were measured on cloudy days and during early morning and late afternoon of sunny days. Decreases in stomatal conductance with increases in vapour pressure deficit may have been a response to avoid further water deficits and suggested a stomatal response to changes in relative humidity. Daily transpiration varied between 470 and 1014 g m^-2 day^-1 during cloudy days and between 532 and 944 g m^-2 day^-1 during clear days. Stomatal conductance may have also responded to changes in leaf water potential, which was minimum at noon. The minimum tissue water potential measured in the field was -1.8 MPa in Myrcianthes fragrans, and the minimum turgor pressure was 0.49 MPa also in M. fragrans. There was a correlation between the osmotic potential and the minimum tissue water potential, suggesting that osmotic potential plays a major role in the maintenance of turgor in these species, in spite of the great variability in the elastic properties of leaf tissues. Turgor pressure decreased during the day following the course of water potential but never approached the turgor loss point, as it has been measured in some lowland rain forest trees. This is a strong indication that elfin cloud forest trees do not suffer severe water deficits, and that small tree stature is not directly related to water shortage.     

The arthropod gap junction and pseudo-gap junction: Isolation and preliminary biochemical analysis

Summary The hepatopancreas of the crayfish, Procambarus clarkii, contains an unusual abundance of gap junctions, suggesting that this tissue might provide an ideal source from which to isolate the arthropod-type of gap junction. A membrane fraction obtained by subcellular fractionation of this organ contained smooth septate junctions, zonulae adhaerentes, gap junctions and pentalaminar membrane structures (pseudo-gap junctions) as determined by electron microscopy. A further enrichment of plasma membranes and gap junctions was achieved by the use of linear sucrose gradients and extraction with 5 mM NaOH. The enrichment of gap junctions correlated with the enrichment of a 31 Kd protein band on polyacrylamide gels. Extraction with 20 mM NaOH or 0.5% (w/v) Sarkosyl NL97 resulted in the disruption and/or solubilization of gap junctions. Negative staining revealed a uniform population of 9.6 nm diameter subunits within the gap junctions with an apparent sixfold symmetry. Using antisera to the major gap junctional protein of rat liver (32 Kd) and to the lens membrane protein (MP 26), we failed to detect any homologous antigenic components in the arthropod material by immunoblotting-enriched gap junction fractions or by immunofluorescence on tissue sections. The enrichment of another membrane structure (pseudo-gap junctions), closely resembling a gap junction, correlated with the enrichment of two protein bands, 17 and 16Kd, on polyacrylamide gels. These structures appeared to have originated from intracellular myelin-like figures in phagolysosomal structures. They could be distinguished from gap junctions on the basis of their thickness, detergent-alkali insolubility, and lack of association with other plasma membrane structures, such as the septate junction. Pseudo-gap junctions may be related to a class of pentalaminar contacts among membranes involved in intracellular fusion in many eukaryotic cell types. We conclude that pseudo-gap junctions and gap junctions are different cellular structures, and that gap junctions from this arthropod tissue are uniquely different from mammalian gap junctions of rat liver in their detergentalkali solubility, equilibrium density on sucrose gradients, and protein content (antigenic properties).     

哺乳动物不活动化肌纤维的动作电位

用河豚毒素(TTX)慢性阻断大鼠坐骨神经的冲动传导,使后肢不活动,经过不同时间(最长7d)后离体观察了快肌伸趾长肌(EDL)和慢肌比目鱼肌(SOL)肌纤维终板区的诱发动作电位。我们发现在不活动期间动作电位超射和上升速率逐步下降,并从第4天起部分肌纤维能在含有1×10~(-7)g/ml TTX的溶液中被诱发产生动作电位(称抗TTX动作电位),待至第7天时全部SOL肌纤维和90％的EDL肌纤维都能被诱发出抗TTX动作电位。与去神经肌纤维相比,不仅抗TTX动作电位出现较晚,并且其超射和上升速率较低。在去掉TTX阻断使肌肉恢复活动后,动作电位超射和上升速率渐趋恢复,抗TTX动作电位逐渐消失。无论是动作电位的恢复还是抗TTX动作电位的消失,EDL肌纤维均快于SOL肌纤维。本文还讨论了不活动化使肌纤维动作电位变化以及快、慢肌差别的可能原因。     

Cell age-related differences in the interaction of a potential-sensitive fluorescent dye with nuclear envelopes of Acetabularia mediterranea

Abstract. To study whether an electrical potential difference exists across the nuclear envelope or inner nuclear membrane of plant cells, the authors have used an optical probe of membrane potential, the cationic fluorescent dye, DiOC₆(3) (MW = 572.5). This dye was microinjected into the nucleoplasm of isolated Acetabularia nuclei (which are still surrounded by a thin layer of cytoplasm) and its subnuclear localization visualized by fluorescence microscopy. Striking differences, which seemed to be correlated with the developmental stage of the isolated nucleus, were observed. In nuclei isolated from cells at the stage of early cap stage formation, the dye was restricted to the nuclear envelope. In nuclei isolated from cells with intermediate or fully developed caps, there was increased nucleoplasmic staining, and the staining of the envelope was frequently diminished or abolished. In all nuclei, the dye remained within the nucleus after injection. Cytoplasmic staining was only observed when nuclei isolated from cells at the stage of early cap formation were incubated in a hyper- or hypo-tonic medium. Various ionophores, injected before the dye into the nucleoplasm, had no effect on the subsequent nuclear localization of DiOC₆(3), although they did rapidly induce nucleolar condensation in nuclei isolated from cells at the stage of early cap formation. The results suggested that the electrical properties of Acetabularia nuclear envelopes or inner nuclear membranes change during cell maturation. Furthermore, the retention of the dye in the nucleoplasm under isotonic conditions indicated that the nuclear pores were not open channels for molecules of this size.     

The efficiency of water use in water stressed plants is increased due to ABA induced stomatal closure

Gas exchange and abscisic acid content of Digitalis lanata EHRH. have been examined at different levels of plant water stress. Net photosynthesis, transpiration and conductance of attached leaves declined rapidly at first, then more slowly following the withholding of irrigation. The intercellular partial pressure of CO₂ decreased slightly. The concentration of 2-cis(S)ABA increased about eight-fold in the leaves of non-irrigated plants as compared with well-watered controls. A close linear correlation was found between the ABA content of the leaves and their conductance on a leaf area basis. In contrast, the plot of net assimilation versus ABA concentration was curvilinear, leading to an increased efficiency of water use during stress. After rewatering, photosynthesis reached control values earlier than transpiration, leaf conductance and ABA content. From these data it is concluded that transpiration through the stomata is directly controlled by the ABA content, whereas net photosynthesis is influenced additionally by other factors.Possible reasons for the responses of photosynthesis and water use efficiency to different stress and ABA levels are discussed.Abbreviations A net CO₂ assimilation - ABA abscisic acid - C_i intercellular CO₂ concentration - g stomatal conductance - T transpiration - WUE water use efficiency     

Effect of Maternal Alcohol Consumption on the Lipid Composition of CNS in the Offspring

Maternal alcohol consumption at a level that does not affect calorie intake increases cholesterol concentration and content as well as incorporation of labeled glucose into cholesterol in the brain and spinal cord of newborn rat pups. Continued consumption of alcohol during lactation also affects the galactolipid concentration in the brain and spinal cord of pups at 21 days of age, and this increase seems mainly to be due to an increase in content of myelin lipids. Analysis of myelin shows that the concentration of phospholipids also increases in this fraction. The increase in incorporation of labeled glucose into these membrane lipids suggests an increase in the synthesis of these lipids, which prevents fluidization of the membrane by alcohol. That in the brainstem the increase in levels of cholesterol and galactolipids is higher than in other regions and that there is also an increase in content of sphingomyelin, phosphatidylcholine, and phosphatidylethanolamine suggest that the brainstem needs better protection against fluidization.     

Effects of Prolonged Depolarization on the Nicotinic Acetylcholine Receptors of PC12 Cells

To determine whether prolonged depolarization and/or changes in intracellular Ca2+ concentrations stimulate adaptive responses of neuronal nicotinic acetylcholine receptors, PC12 pheochromocytoma cells were grown in medium containing various concentrations of K+. Nicotinic receptor function was determined as carbachol-stimulated uptake of 86Rb+. Cells were exposed to 50 mM K+ for up to 4 days and then allowed to repolarize for 60 min. Under these conditions, no changes in basal or carbachol-stimulated uptake of 86Rb+ were observed. Furthermore, neither the time course of carbachol-stimulated uptake or the carbachol concentration dependence of 86Rb+ uptake was altered. Finally, concurrent depolarization did not affect the functional down-regulation produced by chronic exposure of the cells to carbachol. Thus, neuronal nicotinic acetylcholine receptors on PC12 cells do not appear to be regulated by depolarization or prolonged elevation of the intracellular Ca2+ level.     

Effects of drought on host and endophyte development in mycorrhizal soybeans in relation to water use and phosphate uptake

Bethlenfalvay, G. J., Brown, M. S., Ames, R. N. and Thomas, R. S. 1988. Effects of drought on host and endophyte development in mycorrhizal soybeans in relation to water use and phosphate uptake. - Physiol. Plant. 72: 565–571.
Soybean [ Glycine max (L.) Merr.] plants were grown in pot cultures and inoculated with the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus mosseae (Nicol. & Gerd.) Gerd. and Trappe or provided with P fertilizer (non-VAM plants). After an initial growth period (21 days), plants were exposed to cycles of severe, moderate or no drought stress over a subsequent 28-day period by rewatering at soil water potentials of -1.0, -0.3 or -0.05 MPa. Dry weights of VAM plants were greater at severe stress and smaller at no stress than those of non-VAM plants. Phosphorus fertilization was applied to produce VAM and non-VAM plants of the same size at moderate stress. Root and leaf P concentrations were higher in non-VAM plants at all stress levels. All plants were stressed to permanent wilting prior to harvest. VAM plants had lower soil moisture content at harvest than non-VAM plants. Colonization of roots by G. mosseae did not vary with stress, but the biomass and length of the extraradical mycelium was greater in severely stressed than in non-stressed plants. Growth enhancement of VAM plants relative to P-fertilized non-VAM plants under severe stress was attributed to increased uptake of water as well as to more efficient P uptake. The ability of VAM plants to deplete soil water to a greater extent than non-VAM plants suggests lower permanent wilting potentials for the former.     

Effects of UV-C radiation on extension growth, H+-extrusion and transmembrane electric potential in maize coleoptile segments

The effect of 253.7 nm ultraviolet radiation on elongation growth, medium acidification and changes in electric potential difference between vacuole and external medium in cells of maize ( Zea mays L.) coleoptile segments was investigated. It was found that irradiation with 390, 1170, 3900 and 5 850 J m⁻² UV-C (ultraviolet radiation 253.7 nm) inhibited elongation growth, whereas at 195 J m⁻² stimulation of growth was observed. The administration of IAA (10⁻⁵ M ) to the incubation medium of coleoptile segments partially abolished the inhibitory effect of UV-C. The pH of the incubation medium, measured simultaneously with growth, showed that the exposure of the segments to UV-C caused inhibition of H⁺-extrusion (or stimulation of H⁺ uptake). The presence of IAA (10⁻⁵ M ) in the incubation medium promoted (except after 5850 J m⁻² irradiation) H⁺-extrusion to a level comparable with that produced by IAA in non-irradiated segments. In UV-C irradiated segments the potential difference underwent significant alterations. Irradiation of coleoptile segments with 390 J m⁻² caused a transient depolarization, which was fully reversible within 30 min, while at higher doses depolarization was irreversible. The hyperpolarization of the membrane potential (MP) in cells of maize coleoptile induced by IAA was completely nullified by subsequent irradiation with UV-C. It is suggested that UV-C inhibited IAA-induced growth by a mechanism independent of cell wall acidification.