A Golgi-electron-microscopical study of the structure and development of the lamina ganglionaris of the locust optic lobe

Summary The gross structure as well as the neuronal and non-neuronal components of the lamina ganglionaris of the locust Schistocerca gregaria are described on the basis of light- and electron-microscopical preparations of Golgj (selective silver) and ordinary histological preparations. The array of optic cartridges within the lamina neuropile — their order and arrangement — and the composition of the cartridges are described. There are six types of monopolar neurons: three whose branches reach to other cartridges and three whose branches are confined to their own cartridges. Retinula axons terminate either in the lamina or the medulla neuropiles. There are three types of centrifugal neurons, two types of horizontal neuron, as well as glia and trachea in the lamina neuropile. The development of the lamina neuropile is described in terms of developing monopolar and centrifugal axons, growing retinula fibres, and composition of the developing optic cartridges.MSN was supported in part by a Fulbrights-Hays Scholarsship. We are grateful to the Science Research Council for its grant to PMJS.     

Implication of growth hormone in experimental induction of vitellogenesis by estradiol-17 beta in female silver eel (Anguilla anguilla L.)]

The effect of different preparations of growth hormone (GH) was assayed with 17 beta-estradiol on vitellogenesis in hypophysectomized or normal female silver eels. Vitellogenin (Vg) plasma levels were taken as the index of hepatic vitellogenesis. The E2 doses were chosen to give the same pattern for the plasma Vg level as in the controls. They decreased or remained undetectable in hypophysectomized or normal animals. GH also failed to induce alone a significant modification. When E2 was injected together with a GH, plasma Vg levels were 5.13 +/- 1.30 times higher with salmon GH in hypophysectomized eels and 2.01 +/- 0.25 times higher with bovine GH in normal eels. GH is shown to enhance the effects of E2 on hepatic vitellogenesis induction in a teleost.     

STUDIES ON THE GOLGI APPARATUS BY ELECTRON MICROSCOPY WITH PARTICULAR REFERENCE TO AOYAMA'S TECHNIQUE

1. Aoyama's silver impregnation method for the Golgi apparatus has been used on exocrine cells of the pancreas of the mouse and studied by electron microscopy in order to determine as precisely as possible where the silver is deposited. Similar cells have also been fixed in buffered osmium tetroxide solution and compared with cells treated by the silver technique. 2. Examination of the Aoyama preparations usually revealed a light deposition of silver in the cytoplasm (hyaloplasm or matrix) and a heavy deposition of silver around a series of closely apposed vacuoles. The heavy deposition of silver was regarded as revealing the chromophilic region of the Golgi apparatus while the vacuoles were identified as the chromophobic component. 3. Comparison of the silver preparations with those fixed in buffered osmium tetroxide solution showed that the silver was primarily deposited in the region of the Golgi membranes.     

Studies on the Golgi apparatus by electron microscopy with particular reference to Aoyama's technique

1. Aoyama's silver impregnation method for the Golgi apparatus has been used on exocrine cells of the pancreas of the mouse and studied by electron microscopy in order to determine as precisely as possible where the silver is deposited. Similar cells have also been fixed in buffered osmium tetroxide solution and compared with cells treated by the silver technique. 2. Examination of the Aoyama preparations usually revealed a light deposition of silver in the cytoplasm (hyaloplasm or matrix) and a heavy deposition of silver around a series of closely apposed vacuoles. The heavy deposition of silver was regarded as revealing the chromophilic region of the Golgi apparatus while the vacuoles were identified as the chromophobic component. 3. Comparison of the silver preparations with those fixed in buffered osmium tetroxide solution showed that the silver was primarily deposited in the region of the Golgi membranes.     

Histochemical Differentiation of Chloride from Other Ions Precipitated by Silver Nitrate in Freeze-Substitution Fixation

The method is based on substitution fixation at —25° C of quickly frozen tissue with a 90% alcohol solution saturated with silver nitrate. The silver salts are photochemically reduced in the histological preparations. At this low temperature very little staining of the protein structure of the tissue takes place. Silver ions adsorbed by the tissue can be removed by treatment with a sodium nitrate solution. About 2/3 of the brown material in the histological preparations of cerebral cortex was due to the chloride in the tissue, 1/6 to the phosphate, 1/10 to an unidentified (probably organic) anion, and 1/20 to bicarbonate. When the alcoholic silver nitrate solution used for the fixation is acidified, or the sections are treated with nitric acid, the colored material consists of reduced silver chloride only. A comparison of the light absorption in histological preparations of cortex treated with neutral and with acid solutions supported the conclusion that about 2/3 of the colored material in the tissue is reduced silver chloride.     

A validated silver‐nanoparticle‐enhanced chemiluminescence method for the determination of citalopram in pharmaceutical preparations and human plasma

A simple and sensitive chemiluminescence (CL) method was developed for the determination of citalopram in pharmaceutical preparations and human plasma. The method is based on the enhancement of the weak CL signal of the luminol–H₂O₂ system. It was found that the CL signal arising from the reaction between alkaline luminol and H₂O₂ was greatly increased by the addition of silver nanoparticles in the presence of citalopram. Prepared silver nanoparticles (AgNPs) were characterized by UV–visible spectroscopy and transmission electron microscopy (TEM). Various experimental parameters affecting CL intensity were studied and optimized for the determination of citalopram. Under optimized experimental conditions, CL intensity was found to be proportional to the concentration of citalopram in the range 40–2500 ng/mL, with a correlation coefficient of 0.9997. The limit of detection (LOD) and limit of quantification (LOQ) of the devised method were 3.78 and 12.62 ng/mL, respectively. Furthermore, the developed method was found to have excellent reproducibility with a relative standard deviation (RSD) of 3.65% (n = 7). Potential interference by common excipients was also studied. The method was validated statistically using recovery studies and was successfully applied to the determination of citalopram in the pure form, in pharmaceutical preparations and in spiked human plasma samples. Percentage recoveries were found to range from 97.71 to 101.99% for the pure form, from 97.84 to 102.78% for pharmaceutical preparations and from 95.65 to 100.35% for spiked human plasma. Copyright © 2013 John Wiley & Sons, Ltd.     

Modified cobalt staining and silver intensification techniques for use with whole-mount gastropod ganglion preparations

While silver intensification of cobalt-filled cells is a common procedure for use with many arthropod preparations, it has not been routinely applied to gastropods. Several modifications in cobalt-staining techniques currently used with gastropods along with adaptations of silver intensification techniques used in insects are described. Cobalt was introduced into cells through axonal filling of cut nerve trunks or by either pressure injection or iontophoresis from intracellular, micropipette electrodes that had previously had their tips etched in dilute hydrofluoric acid. Etching produced consistent, sharp tips with large lumina. Further procedural modifications allowed complete, even intensification of neurons in large gastropod ganglia. These techniques have proved to be reliable and apparently broadly applicable, having been successfully used on three diverse gastropod species.     

Golgi potassium-dichromate silver-nitrate impregnation

Summary Golgi preparations were made by consecutive treatment of formalin-fixed brain and liver with potassium dichromate and silver nitrate. Impregnated tissue dissected from thin slices of the blocks were studied by X-ray powder diffraction methods, in a diffractometer and a Guinier camera. Such tissue proved to contain crystalline silver chromate, Ag₂CrO₄, both while still in the silver nitrate solution and after dehydration in ethanol and clearing in xylene and xylene-Dammar resin. No other compounds containing chromium or silver were detectable. Formalin-fixed tissue merely treated with silver nitrate contained silver chloride, but in impregnated tissue the amount was too scarce to be visible. Hence, silver chloride was no integral part of the Golgi precipitate.A number of mostly ethereal oils traditionally used for clearing histological sections, did not cause the appearance of metallic silver in detectable amount in the Golgi preparations. However, after treatment with clove oil and creosote metallic silver was detected in the tissue.This study was supported by U.S. P.H. S. Grant NS 07998. This aid is gratefully acknowledged.We are indebted to Miss I. Madsen and Mrs. K. Sörensen for technical assistance.     

Binding and movement of silver in the intestinal epithelium of a marine teleost fish,the European flounder (Platichthys flesus)

The intestine has been indicated as a site of waterborne silver toxicity in marine fish and chronic effects at the intestine have been observed at concentrations far below acutely toxic level. Thus, models of silver toxicity to marine fish need to consider the intestine as a biotic ligand. The present study characterises binding of silver to the intestine of the European flounder (Platichthys flesus). Everted intestinal sacks were prepared and submersed in a solution mimicking the intestinal fluid of the fish at the acclimation salinity (21 per thousand). Silver was added as (110m)AgNO(3) or (110m)AgNO(3)/AgNO(3) mixtures at concentrations ranging from 1.6 to 950 nM total silver. Appearance of (110m)Ag was analysed in mucosal scrapings, muscle layers, and in the plasma saline on the serosal side of the intestine. The latter represented uptake into blood and other extra-intestinal compartments. Mucosal scrapings consisted of both epithelial cells and mucus and, thus contained adsorbed as well as absorbed silver. Most of the silver in mucosal scrapings was bound to mucus. There was no difference in silver binding between the anterior, mid, and posterior regions of the intestine. Concentration-dependency of silver binding was sigmoidal and saturated above 100 nM total silver. The saturable appearance of (110m)Ag in the plasma saline suggest that silver passage across the intestine is transcellular and carrier mediated. Mucus likely influences uptake of silver by altering its speciation from that in the lumen and by serving as physical barrier for silver binding to the brushborder membrane. A biotic ligand model for marine fish to silver may have to consider these interactions.     

High-resolution two-dimensional electrophoresis of nuclear proteins: a comparison of HeLa nuclei prepared by three different methods.

A comparative analysis of HeLa cell nuclear proteins is presented using Iso-Dalt methods of protein resolution in two dimensions. The nuclear proteins were prepared by (1) spin through glycerol cushion, (2) spin through sucrose cushion, or (3) Triton wash. Improved resolution of total nuclear proteins in the range of pH 4.5-6.0 was achieved by substituting longer isotubes in combination with broad-range ampholines during the isoelectric focusing step. An attempt to indicate silver stainable protein spots common to total cellular extracts and nuclear preparations has been made. Also, proteins that appear to be well represented in all three nuclear preparations and remain undetectable in the total cellular protein pattern have been marked as probably being enriched nuclear proteins. Such a comparative analysis of whole nuclear protein preparations made it possible to document that the different preparations preserved the same set of proteins. The Triton-wash method of obtaining nuclei was identified as the preferred choice. Coomassie-stained gels and blots of these nuclear proteins could serve as a guide for accessing relevant protein spots for further biochemical analysis such as immunoblotting.     

Ultrastructural analysis of preparations impregnated with silver salts]

The work was devoted to the ultrastructural analysis of the neurohistological preparations. Sections of the tissue from the precardial parts of the pulmonary and caval dog veins were impregnated with silver salts after Campos and embedded in the araldite by a special method. Electronmicroscopi studies showed reduced silver adsorbed by the tissue of the impregnated preparations to exhibit a granular structure (the granules were 30-400 A in size). The largest of them were revealed in the axoplasm of the myelinated and unmyelinated nerve conductors, whereas the smaller ones found in various cellular and fibrous formations of the tissue substrate; silver granules were as a rule absent within the thickness of the myelin sheath. The noted regularities of adsorption and distribution of the silver granules in the impregnated preparations caused a morphologically expressed phenomenon of argentophilia.     

A quantitative electron microscope autoradiographic study on 125I-human choriogonadotropin internalization in bovine luteal slices

Very few silver grains were seen on the cell surface and none intracellularly after incubation for 2 h at 4 degrees C. However, numerous grains were seen in various subcellular organelles when the tissues were incubated for 2 h at 22 degrees or 38 degrees C. The grain distribution was qualitatively similar, but quantitatively, there were fewer grains at 22 degrees than at 38 degrees C. Co-incubation of 125I-hCG with excess unlabelled hCG resulted in the virtual disappearance of silver grains from all the subcellular organelles. Excess unlabelled human luteinizing hormone (but not follicle-stimulating hormone or prolactin) inhibited the appearance of silver grains in luteal tissue. There were no silver grains in bovine liver slices incubated with 125I-hCG. The plasma membrane-associated grains progressively decreased, while intracellular organelle-associated grains increased with time at 38 degrees C. There were no grains in nuclei at 5 min, but they appeared at 10 min and increased until 120 min. After correction for radiation spread by three-step mask analysis, smooth endoplasmic reticulum and mitochondria did not contain any grains. The grain density was the highest in Golgi, followed by lysosomes, rough endoplasmic reticulum, nuclei, and plasma membranes after incubation for 2 h at 38 degrees C. Thus, the electron microscope autoradiography approach confirmed our biochemical data in the preceding paper (Chegini et al., Exp cell res 151 (1984) 466 [5]) on time, temperature dependency and specificity of 125I-hCG internalization, association of internalized hormone with a variety of intracellular organelles, and the highest uptake in Golgi.     

Pharmacokinetics of omega-3-fatty acids during ingestion of fish oil preparations.

An in vivo comparison of three dosages (3 g, 6 g, 12 g) of two different fish oil preparations in terms of plasma concentrations of their major active components eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) was performed. The plasma accumulation was measured during 28 days of ingestion and an equally long wash out period. Data were scrutinized for bioavailability in order to distinguish between the efficiency of the two preparations. Rapid increases in EPA and DHA plasma concentrations can be demonstrated at all dosages during a 28-day ingestion period. EPA accumulated more during ingestion of high than of low dosages of fish oil. DHA revealed almost identical increases and peak values in plasma concentrations in all subgroups. The present data demonstrate dose dependent increases of EPA concentrations whereas DHA plasma concentrations are comparable in all dosages investigated. Measurable EPA and DHA plasma concentration levels are inappropriate means to explain clinical effectiveness. These results were found in both commercially available fish oil preparations. Direct comparison of both preparations revealed no differences in bioavailability.     

Water pH and urinary excretion in silver catfish Rhamdia quelen

The effect of acute exposure to different water pH levels on urinary excretion and plasma ion levels in silver catfish Rhamdia quelen was analysed. Fish were exposed to pH 4·0, 5·0, 7·5, 8·0, and 9·0 for 4 days and urine was collected. Other specimens were also exposed to the experimental pH for 24 h and blood was sampled. Urine flow rate, urine and plasma pH showed a significant trend to increase with the increase of water pH. Urinary Na⁺ excretion rate also increased and ammonia urinary excretion rate decreased with the increase of water pH. There was a significant trend to decrease volume, ammonia, Cl⁻ and Na⁺ urinary excretion rate with increasing mass in fish exposed to all pH levels studied. Plasma ammonia levels showed a slight decrease in fish exposed to water pH from 4·0 to 8·0, but those exposed to water pH 9·0 presented the highest ammonia levels. Most plasma ions and urinary excretion changes observed in silver catfish exposed to acidic or alkaline water were similar to those already detected in rainbow trout Oncorhynchus mykiss . In addition, the kidney and urinary bladder might participate on acid–base balance in silver catfish, since urine pH changed according to plasma pH.     

Advances in plasmatic fractions used in blood transfusion.

I tried to give a bird's eye view of some of the intriguing recent advances in the production and clinical use of albumin, immunoglobulins, coagulation factors, protease inhibitors and plasma enzyme preparations. I aimed to put into the limelight of my lecture the basic differences between the reaction of plasma proteins in solutions in the test tube or on model surfaces in vitro as opposed to the well-balanced intricate protein-protein and cell membrane-plasma protein interactions in vivo. In the light of these recent advances we have to reassess the biological properties, structural and functional integrity of our plasma protein preparations. In spite of technical and regulatory difficulties new fractionation techniques have to break through. Especially gentle techniques which do not denature proteins are badly needed. Purified albumin with an adequate fatty acid content seems to be the safest and therapeutically most useful preparation among the albumin containing fractions. PPF and even albumin are, however, not so safe as we have thought. There is an urgent need to fight more intensively against their unjustified use. There is a growing need for specific immunoglobulins, coagulation factors, protease inhibitors and therapeutically useful enzyme preparations.     

硝酸银硅胶柱层析分离血浆不饱和脂肪酸

目的:利用硝酸银硅胶填料有效分离出血浆混合脂肪酸中的亚油酸。方法:通过改进的FOLCH法提取血浆总脂后,再采用皂化、酸化水解的方法将总脂转化为混合脂肪酸。用ghosh法将硅胶改性为硝酸银硅胶后,以亚油酸为对象,通过静态吸附试验了解硝酸银硅胶对不饱和脂肪酸的吸附特性,采用柱层析的方法分离血浆混合脂肪酸中的亚油酸。结果:血浆与有机溶剂在1∶5时既能有效萃取血浆总脂,用正己烷∶二氯甲烷∶乙醚=89∶10∶1作为洗脱剂,将洗脱液甲酯化后进行GC和GC-MS检测,硝酸银硅胶柱的洗脱液中亚油酸的纯度60.74%,硅胶柱的为23.65%,不饱和脂肪酸得到了较好的纯化。     

Comparative effectiveness of antimicrobial action of antiseptics against pathogens of chronic purulent otitis media]

Comparable antimicrobial and disinfecting action of decamethoxine and silver preparations on pathogens of chronic purulent otitis media (CPOM) was studied. The clinical isolates of staphylococci proved to be most sensitive to decamethoxine whose MBcC conformed to 16.5 micrograms/ml. The antimicrobial action on Proteus spp. and Pseudomonas aeruginosa was less pronounced. The required concentrations for bactericidal action on these pathogens were 69 and 93.5 micrograms/ml, respectively. The antimicrobial activity of the silver preparations such as poviargol, collargol and protargol was low. Depending on the microbial species, the bactericidal effect of the silver preparations was 12-235 times lower than that of decamethoxin. It was also shown that decamethoxin had a high disinfecting action on CPOM pathogens. It was noted that decamethoxin had a marked ability to increase the bactericidal action of poviargol (by 2-14 times) and its disinfecting action (by 2 times) on Proteus spp., E. coli and Ps. aeruginosa.     

Safety of human blood products: inactivation of retroviruses by heat treatment at 60 degrees C

Acquired immune deficiency syndrome (AIDS) can be transferred to patients by blood transfusions or human blood preparations, such as cryoprecipitates or factor VIII concentrates. Retroviruses have been discussed as infectious AIDS agents and more recently human T-lymphotropic retroviruses designated as HTLV type III and LAV (lymphadenopathy-associated virus) have been isolated from AIDS patients. Whether heat treatment at 60 degrees C (pasteurization) of liquid human plasma protein preparations inactivates retroviruses was therefore investigated. Pasteurization had already been included in the routine manufacturing process of human plasma protein preparations in order to guarantee safety with regard to hepatitis B. Since high titer preparations of human retroviruses were not available, heat inactivation was studied using Rous sarcoma virus added to the various plasma protein preparations tested. This retrovirus which was obtained in preparations of 6.0 log10 FFU/ml was shown to be at least as heat stable as two mammalian retroviruses studied, i.e., feline and simian sarcoma virus. In all of eight different plasma protein preparations tested, Rous sarcoma virus was completely inactivated after a heat treatment lasting no longer than 4 hr. It is thus concluded that pasteurization of liquid plasma protein preparations at 60 degrees C over a period of 10 hr must confer safety to these products with respect to AIDS, provided that the AIDS agents are retroviruses of comparable heat stability as Rous sarcoma virus and the mammalian retroviruses tested.     

Comparative study of antifungal activity of two preparations of green silver nanoparticles from Portulaca oleracea extract

The green silver nanoparticles (green AgNPs) exhibit an exceptional antimicrobial property against different microbes, including bacteria and fungi. The current study aimed to compare the antifungal activities of both the crude aqueous extract of Portulaca oleracea or different preparations of green AgNPs biosynthesized by mixing that aqueous extract with silver nitrate (AgNO₃). Two preparations of the green AgNPs were synthesized either by mixing the aqueous extract of P. oleracea with silver nitrate (AgNO₃) (normal AgNPs) or either irradiation of the AgNPs, previously prepared, under ⁶⁰Co γ-ray using chitosan (gamma-irradiated AgNPs). Characterization of different AgNPs were tested by Zeta potential analyzer, Ultraviolet (UV) Visible Spectroscopy, and Fourier-Transform Infrared (FTIR) spectrometry. Three different plant pathogenic fungi were tested, Curvularia spicifera, Macrophomina phaseolina, and Bipolaris sp. The antifungal activities were evaluated by Transmission Electron Microscope (TEM) for either the crude aqueous extract of P. oleracea at three doses (25%, 50%, and 100%) or the newly biosynthesized AgNPs, normal or gamma-irradiated. With a few exceptions, the comparative analysis revealed that the irradiated green AgNPs at all three concentrations showed a relatively stronger antifungal effect than the normal AgNPs against all the three selected fungal strains. UV–visible spectroscopy of both preparations showed surface plasmon resonance at 421 nm. TEM results showed that both AgNPs were aggregated and characterized by a unique spherical shape, however, the gamma-irradiated AgNPs were smaller than the non-irradiated AgNPs (0.007–0.026 µM vs. 0.009–0.086 µM). TEM photographs of the fungal strains treated with the two AgNPs preparations showed flaccid structures, condensed hyphae, and shrunken surface compared with control cells. The data suggested that the biosynthesized P. oleracea AgNPs have antifungal properties against C. spicifera, M. phaseolina, and Bipolaris sp. These AgNPs may be considered a fungicide to protect different plants against phytopathogenic fungi.     

Effects of Potassium Cyanide on Silver Stainability of Specific Cell Structures

Treatment with potassium cyanide prevents the occurrence of diffuse silver precipitates on chromosome preparations. It therefore allows greatly increased times of incubation with silver solutions on both mitotic and meiotic preparations. By this method selective silver staining of active nucleolar organizers, centromeric regions and kinetochores of mitotic chromosomes and of specifically staining structures of meiotic chromosomes has been obtained.