Melatonin reverses neurochemical alterations induced by 6-OHDA in rat striatum

The present work showed that the intrastriatal injection of 6-OHDA significantly decreases DA, DOPAC and HVA levels in that rat brain structure. Although there is also a decrease in 5-HT levels no changes were observed in 5-HIAA levels as compared to controls. On the other hand, melatonin (2, 5, 10 and 25 mg/kg. i.p., daily for 7 days) treatment starting 1 h after 6-OHDA lesions, partially reverses the decreases caused by 6-OHDA lesions on these neurotransmitter levels, and contents were brought to approximately 50% of that observed in the contralateral sides of controls or of melatonin treated group. Melatonin was more efficient at the doses of 5 and 10 mg/kg, i.p., and effects were similar between the lowest and highest doses characteristic of a bell-shaped type of response. The apomorphine-induced rotational behavior (3 mg/kg, i.p.) was blocked by 60, 89, 78 and 47% after the doses of 2, 5, 10 and 25 mg/kg, i.p., respectively. Similarly, in this case the doses of 5 and 10 mg/kg were also more efficient. Melatonin (5 mg/kg) produced an upregulation of D1 receptors associated with a decrease in Kd value. While no change was observed in maximum density of D2 receptors, the Kd value was also decreased.     

Rapid alterations in cellular morphology and plasma membrane structure induced in rat thymocytes by mitogenic stimuli.

Under conditions where a maximum stimulation of 3-O-methyl-glucose transport is observed, three thymocyte mitogens (concanavalin A, ionophore A23187 and hydrogen peroxide) cause cell rounding and a decrease in the density of intra-membrane particles on the plasma membrane. The early effects of mitogens on the thymocyte plasma membrane are similar to those of osmotic shock.     

Compartmentalization and ultrastructural alterations induced by chromium in aquatic macrophytes

The aim of the present study was to identify the sites of accumulation of Cr in the species of macrophytes that are abundant in the Cachoeira river, namely, Alternanthera philoxeroides, Borreria scabiosoides, Polygonum ferrugineum and Eichhornia crassipes. Plants were grown in nutritive solution supplemented with 0.25 and 50 mg l^?1 of CrCl₃·6H₂O. Samples of plant tissues were digested with HNO₃/HCl in a closed-vessel microwave system and the concentrations of Cr determined using inductively-coupled plasma mass spectrometry (ICP-MS). The ultrastructure of root, stem and leaf tissue was examined using transmission electron microscopy (TEM) and secondary ion mass spectrometry (SIMS) in order to determine the sites of accumulation of Cr and to detect possible alterations in cell organelles induced by the presence of the metal. Chromium accumulated principally in the roots of the four macrophytes (8.6?C30 mg kg^?1 dw), with much lower concentrations present in the stems and leaves (3.8?C8.6 and 0.01?C9.0 mg kg^?1 dw, respectively). Within root tissue, Cr was present mainly in the vacuoles of parenchyma cells and cell walls of xylem and parenchyma. Alterations in the shape of the chloroplasts and nuclei were detected in A. philoxeroides and B. scabiosoides, suggesting a possible application of these aquatic plants as biomarkers from Cr contamination.     

DNA-mediated transfer of multiple drug resistance and plasma membrane glycoprotein expression.

Colchicine-resistant Chinese hamster ovary (CHO) cell mutants whose resistance results from reduced drug permeability have been isolated previously in our laboratories. This reduced permeability affects a wide range of unrelated drugs, resulting in the mutants displaying a multiple drug resistance phenotype. A 170,000-dalton cell surface glycoprotein (P-glycoprotein) was identified, and its expression appears to correlate with the degree of resistance. In this study we were able to confer the multiple drug resistance phenotype on sensitive mouse L cells by DNA-mediated gene transfer of DNA obtained from the colchicine-resistant mutants. P-glycoprotein was detected in plasma membranes of these DNA transformants by staining with an antiserum raised against membranes of mutant CHO cells. These results are consistent with a causal relationship between P-glycoprotein expression and the multiple drug resistance phenotype.     

Ethanol induced alterations in plasma membrane protein phosphorylation of neurons and astrocytes

The endogenous protein phosphorylation patterns in plasma membranes of bulk isolated neurons and astroglia from control and chronic ethanol treated rats have been investigated. Chronic ethanol treatment resulted in increased phosphorylation of specific proteins with molecular weights 116, 63 and 60 kDa in both neurons and astrocytes. These proteins were further resolved by 2-DE and the analysis suggested an increased phosphorylation of 10–15 proteins, of which 116 kDa protein is phosphorylated to a higher extent by ethanol. Further, decreased phosphorylation was noticed in D-95 and D-63 proteins in neurons and D-78 and D-54 proteins in astrocytes. Alkali stability experiments for identifying the phosphoamino acid involved in phosphorylation of 116, 63 and 60 kDa proteins suggested that tyrosine and threonine are not involved and probably serine is the likely site for phosphorylation during chronic ethanol treatment. The phosphorylation of specific membrane proteins during chronic ethanol treatment might contribute to ethanol evoked cellular dysfunction.     

Seasonal ultrastructural alterations in the plasma membrane produced by slow freezing in cortical tissues of mulberry (Morus bombyciz Koidz. cv. Goroji)

Seasonal alterations in the ultrastructure of the plasma membrane produced by slow freezing were examined in cortical parenchyma cells of mulberry twigs (Morus bombyciz Koidz. cv. Goroji) grown in northern Japan. In freezing-sensitive summer, freezing produced distinct aparticulate domains with accompanying inverted hexagonal_II (H_II) phase transitions in the plasma membrane. In autumn and spring, during cold acclimation and deacclimation, freezing produced aparticulate domains in the plasma membrane without accompanying H_ii phase transitions. In winter, when the twigs were freezing-tolerant, freezing did not produce ultrastructural alterations in the plasma membrane. A significant relationship was recognized between the percentages of cells with aparticulate domains in the plasma membrane, regardless of the presence or absence of H_II phase transitions, and the occurrence of freezing injury throughout all seasons and at all freezing temperatures tested in each season. The aparticulate domains in the plasma membranes were shown to be produced by the close apposition of membranes due to freezing-induced dehydration and deformation of cells. Although the precise mechanisms that cause injury as a result of the formation of aparticulate domains in the plasma membrane remain unclear, our results indicate that the development of cold acclimation paralleled the process whereby cells developed the ability to reduce and finally to prevent the formation of aparticulate domains in the plasma membrane that would otherwise result from freezing-induced cellular dehydration and deformation that brings membranes into close proximity with one another.     

Vincristine induced ultrastructural alterations of cardiac and smooth muscles in mice.

A morphological analysis of the effect of vincristine (VCR) on the cardiac muscle and smooth muscles of blood-vessels and alimentary canal of mice has been carried out. These animals show a high resistance to sublethal doses of VCR. The changes in the cardiac muscle involve mitochondria. The widening of the space between the intercalated discs is seen and accompanied by the occurrence of spheromembranous bodies (SMB) in it. The only alteration typical of the effect of VCR in the smooth muscles is the presence of SMB inside the mitochondria. The number and extent of changes observed increased in course of experiment.     

Heat shock induced ultrastructural alterations in Stylonychia mytilus cells

Fine structural observations on heat shocked cells of S. mytilus reveal that cell organelles undergo structural alterations. Mitochondria show distorted shapes with disorganized cristae and vacuolar spaces. Pulse heat shock results in dilated rough endoplasmic reticulum, abundant polysomes as well as smooth endoplasmic reticulum. Heat shocked cells show membrane bound bodies containing osmiophilic cores. In macronuclei, dense chromatin breaks up into discrete bodies accompanied by the appearance of bundles of fine filaments and clustering of nuclear pores. The most prominent changes are noticed in nucleoli. Within 15 min of heat shock, nucleoli show hypertrophy and fine fibrillar zone which gradually replaces the granular zone by 120 min giving the nucleoli ring shaped configuration. In S phase cells, macronuclei show the arrested replication band in which the diffused zone (the site of DNA replication) is absent.     

Changes in islet plasma membrane calcium-ATPase activity and isoform expression induced by insulin resistance

We studied the effect of insulin resistance (IR) induced by administration of a fructose-rich diet (FRD) to normal Wistar rats for 21 days, upon islet plasma membrane calcium ATPases (PMCAs) and insulin secretion. FRD rats showed significantly higher triglyceride and insulin levels, insulin:glucose ratio and HOMA-IR index than controls. FRD islets released significantly more insulin in response to glucose and showed (a) marked changes in PMCA isoform protein content (decreased PMCA 2 and increased PMCA 3), (b) a decrease in total PMCAs activity, and (c) higher levels of cytosolic calcium [Ca²⁺]_i. The lower PMCAs activity with the resultant increase in [Ca²⁺]_i would favor the compensatory greater release of insulin necessary to cope with the IR state present in FRD rats and to maintain normal glucose homeostasis. Thus, changes in PMCAs activity and isoform expression play a modulatory role upon insulin secretion during long-term adaptation to an increased hormone demand.     

Modulation of P-glycoprotein-mediated multidrug resistance by acceleration of passive drug permeation across the plasma membrane

The drug concentration inside multidrug-resistant cells is the outcome of competition between the active export of drugs by drug efflux pumps, such as P-glycoprotein (Pgp), and the passive permeation of drugs across the plasma membrane. Thus, reversal of multidrug resistance (MDR) can occur either by inhibition of the efflux pumps or by acceleration of the drug permeation. Among the hundreds of established modulators of Pgp-mediated MDR, there are numerous surface-active agents potentially capable of accelerating drug transbilayer movement. The aim of the present study was to determine whether these agents modulate MDR by interfering with the active efflux of drugs or by allowing for accelerated passive permeation across the plasma membrane. Whereas Pluronic P85, Tween-20, Triton X-100 and Cremophor EL modulated MDR by inhibition of Pgp-mediated efflux, with no appreciable effect on transbilayer movement of drugs, the anesthetics chloroform, benzyl alcohol, diethyl ether and propofol modulated MDR by accelerating transbilayer movement of drugs, with no concomitant inhibition of Pgp-mediated efflux. At higher concentrations than those required for modulation, the anesthetics accelerated the passive permeation to such an extent that it was not possible to estimate Pgp activity. The capacity of the surface-active agents to accelerate passive drug transbilayer movement was not correlated with their fluidizing characteristics, measured as fluorescence anisotropy of 1-(4-trimethylammonium)-6-phenyl-1,3,5-hexatriene. This compound is located among the headgroups of the phospholipids and does not reflect the fluidity in the lipid core of the membranes where the limiting step of drug permeation, namely drug flip-flop, occurs.     

Ultrastructural alterations induced by Amphotericin B in the plasma membrane of Epidermophyton floccossum as revealed by freeze-etch electron microscopy

Freeze-etch electron microscopic observations demonstrated that Amphotericin B, a potential antifungal polyene induced profound ultrastructural alterations in the plasma membrane of cells of a human pathogenic fungus, Epidermophyton floccosum; aggregation of membrane-associated particles (85-Å) and formation of depressions (or craters) on the inner fracture face.Thin-sectioning electron microscopy showed “vesiculation” of plasma membranes of Amphotericin B-treated Epidemophyton cells.It can be suggested from the preliminary results that the fungal cells which contain ergosterol as the major sterol are highly sensitive to Amphotericin B.     

Plasma membrane lipid alterations induced by NaCl in winter wheat roots

A highly enriched plasma membrane traction was isolated by two phase partitioning from wheat roots ( Triticum aestivum L. cv. Vivant) grown with and without 100 m M NaCl. The lipids of the plasma membrane fraction were extracted and characterised. Phosphatidylcholine and phosphatidylethanolamine were the major phospholipids with lesser amounts of phosphandylinositol, phosphatidylglycerol, diphosphalidylglycerol, phosphatidic acid and phosphatidylseriae. NaCl decreased the total phospholipids and the phosphatidylcholine portion of the plasma membranes. Salt treatment had no effect on total sterols and glycolipids. but the relative abundance of the tree sterols was altered: cholesterol, stigma sterol and brassicasterol were significantly increased. Salt treatment resulted in an increase of the more planar/less planar ratio of the free sterols and in introduction of a double bond in the C₂₂ position in the side chain of stigma sterol and brassicasterol. The degree of fatty acid saturation of total phospholipids, phospha-tidylcholine and phosphatidylethanolamine was increased after salt treatment. These lipid changes are discussed in relation to the salt tolerance mechanism.     

Chlorpromazine and other psychoactive drug induced alterations of a membrane bound enzyme in rat brain

Psychoactive drugs like chlorpromazine (CPZ), imipramine, lithium and amphetamine in one way or another affect behaviour. The drug responses are presumably mediated by inducing a change in the activity of membrane bound enzymes. CPZ is very potent in inhibiting the alkaline phosphatase activity in rat brain. The combined effect of CPZ with other drugs shows that CPZ and imipramine together inhibit the enzyme activity significantly greater than the individual inhibition either by CPZ or by imipramine alone. Effective inhibition of the alkaline phosphatase activity with a single drug or combined drugs may lead to a change in neuronal permeability through glucocorticoids thereby affecting mood.     

Cell volume alterations induced by platinum-thymine in sarcoma-180 ascites cells: ultrastructural study

Ultrastructural studies of the effects of the chemotherapeutic agent platinum-thymine, on the morphology of sarcoma-180 ascites cells were studied to elucidate the cancer cells immediate response to therapy and the possible mode of cancer cell regression. Sarcoma-180 ascites cells treated with platinum-thymine in vitro at concentrations of 60 micrograms/ml at varying time intervals demonstrated that the drug is capable of killing cancer cells. The cells exhibited drastic nuclear and cytoplasmic alterations with few lipid spherules in the cytoplasm. The cell volume increased tremendously, with the nucleus relatively larger than the cytoplasm as time of treatment increased. Degradative features of the nucleus, and especially the nucleolus, inhibited metabolic processes. Nucleolar and cytoplasmic necrosis with increased cell volume and vacuolation are potential signals for cancer cells response to the drug and a possible mode of causing the eventual demise of the cancer cells.     

Specific 125I-iodination of cell surface lipids: plasma membrane alterations induced during humoral immune attack.

A method whereby lactoperoxidase-catalyzed 125I-iodination of plasma membrane lipids can be achieved is described. The reaction results in a uniform and stable labeling of neutral lipids, phospholipids, lysophosphatides, free fatty acids, and triacylglycerides. By the use of this method, the action of antibody plus complement (C) on the specific release of lipid from the plasma membrane of line-10 tumor cells was studied. Within 15 min after the addition of C to antibody-sensitized cells, the enhanced release of specific lipid classes from the cell surface was observed; these lipids included sphingomyelin, phosphatidylserine, and phosphatidylcholine. The release of phosphatidylethanolamine and, in some instances, triglycerides, was reduced after antibody-C attack. Neither the specificity of the antibody used to sensitize the cells nor the ability of the antibody plus C to be cytotoxic to the cells appeared to affect the identity or amounts of lipids released from the cells.     

Closure of the plasma membrane around microneedle in Amoeba proteus. An ultrastructural study

Microneedle perforations of the plasma membrane of Amoeba proteus were studied on the ultrastructural level. In each individual cell one hole was produced, which subsequently was marked with an eyelash left in place. Cells were quickly fixed, and sections cut parallel to the longitudinal axis of the eyelash. It was clear that the eyelash penetrated the plasma membrane, and that its free tip was located in the interior of the cell. A gap remained between the plasma membrane and the eyelash which may correspond to the electrical leak sometimes found by microelectrode punctures. The edges of the broken plasma membrane curled back into the cytoplasm. Here, a great redundancy of the plasma membrane was observed. Within these membrane accumulations, a large quantity of dense droplets was apposed at the inner leaflet of the plasma membrane. Their involvement in the formation and expansion of the plasma membrane in Amoeba proteus and Xenopus laevis has been suggested previously [1, 18]. Present studies offer more supportive evidence to that effect. Therefore, the interpretation seems to be plausible, that these membrane accumulations are the result of membrane expansion to minimize the hole produced during injury. This is in agreement with Holtfreter's [8] and Bluemink's [1] concept that the wound closure may occur by proliferation of the plasma membrane.     

Glutaraldehyde induced alterations of membrane anionic sites

Prefixation of the plasma membranes of cultured baby hamster kidney cells and human fibroblasts with glutaraldehyde can induce an anomalous distribution of surface anionic sites. However, such glutaraldehyde induced changes do not appear to occur in isolated inner mitochondrial membranes.     

Changes induced in the permeability barrier of the yeast plasma membrane by cupric ion.

A specific effect of Cu2+ eliciting selective changes in the permeability of intact Saccharomyces cerevisiae cells is described. When 100 microM CuCl2 was added to a cell suspension in a buffer of low ionic strength, the permeability barrier of the plasma membranes of the cells was lost within 2 min at 25 degrees C. The release of amino acids was partial, and the composition of the amino acids released was different from that of those retained in the cells. Mostly glutamate was released, but arginine was mainly retained in the cells. Cellular K+ was released rapidly after CuCl2 addition, but 30% of the total K+ was retained in the cells. These and other observations suggested that Cu2+ caused selective lesions of the permeability barrier of the plasma membrane but did not affect the permeability of the vacuolar membrane. These selective changes were not induced by the other divalent cations tested. A novel and simple method for differential extraction of vacuolar and cytosolic amino acid pools by Cu2+ treatment was established. When Ca2+ was added to Cu2+-treated cells, a large amount of Ca2+ was sequestered into vacuoles, with formation of an inclusion of a Ca2+-polyphosphate complex in the vacuoles. Cu2+-treated cells also showed enhanced uptake of basic amino acids and S-adenosylmethionine. The transport of these substrates showed saturable kinetics with low affinities, reflecting the vacuolar transport process in situ. With Cu2+ treatment, selective leakage of K+ from the cytosolic compartment appears to create a large concentration gradient of K+ across the vacuolar membrane and generates an inside-negative membrane potential, which may provide a driving force of uptake of positively charged substances into vacuoles. Cu2+ treatment provides a useful in situ method for investigating the mechanisms of differential solute pool formation and specific transport phenomena across the vacuolar membrane.     

Phospholipid supplementation reverses behavioral and biochemical alterations induced by n-3 polyunsaturated fatty acid deficiency in mice

This study investigated the effects of a diet deficient in alpha-linolenic acid followed or not by supplementation with phospholipids rich in n-3 polyunsaturated fatty acids (PUFA) on behavior and phospholipid fatty acid composition in selected brain regions. Three weeks before mating, two groups of mice were fed a semisynthetic diet containing both linoleic and alpha-linolenic acid or a diet deficient in alpha-linolenic acid. Pups were fed the same diet as their dams. At the age of 7 weeks, a part of the deficient group was supplemented with n-3 PUFA from either egg yolk or pig brain phospholipids for 2 months. In the open field, rearing activity was significantly reduced in the deficient group. In the elevated plus maze (anxiety protocol), the time spent on open arms was significantly smaller in deficient mice than in controls. Using the learning protocol with the same task, the alpha-linolenic acid deficiency induced a learning deficit. Rearing activity and learning deficits were completely restored by supplementation with egg yolk or cerebral phospholipids, though the level of anxiety remained significantly higher than that of controls. There were no differences among the 4 diet groups for either the Morris water maze or passive avoidance. In control mice, the level of 22:6 n-3 was significantly higher in the frontal cortex compared to all other regions analysed. The frontal cortex and the striatum were the most markedly affected by the deficiency. Supplementation with phospholipids restored normal fatty acid composition in brain regions except for frontal cortex. Egg yolk or cerebral phospholipids are an effective source of n-3 PUFA for reversing behavioral changes and altered fatty acid composition induced by a diet deficient in n-3 PUFA.