Renal microvasculature in the adult pipid frog,Xenopus laevis: A scanning electron microscope study of vascular corrosion casts

We studied the opisthonephric (mesonephric) kidneys of adult male and female Xenopus laevis using scanning electron microscopy (SEM) of vascular corrosion casts and light microscopy of paraplast embedded tissue sections. Both techniques displayed glomeruli from ventral to mid-dorsal regions of the kidneys with single glomeruli located dorsally close beneath the renal capsule. Glomeruli in general were fed by a single afferent arteriole and drained via a single thinner efferent arteriole into peritubular vessels. Light microscopy and SEM of vascular corrosion casts revealed sphincters at the origins of afferent arterioles, which arose closely, spaced from their parent renal arteries. The second source of renal blood supply via renal portal veins varied interindividually in branching patterns with vessels showing up to five branching orders before they became peritubular vessels. Main trunks and their first- and second-order branches revealed clear longish endothelial cell nuclei imprint patterns oriented parallel to the vessels longitudinal axis, a pattern characteristic for arteries. Peritubular vessels had irregular contours and were never seen as clear cylindrical structures. They ran rather parallel, anastomosed with neighbors and changed into renal venules and veins, which finally emptied into the ventrally located posterior caval vein. A third source of blood supply of the peritubular vessels by straight terminal portions of renal arteries (vasa recta) was not found.     

Combined immunofluorescence and field emission scanning electron microscope study of plasma membrane-associated organelles in highly vacuolated suspensor cells of white spruce somatic embryos

Highly vacuolated suspensor cells of spruce somatic embryos were examined by immunofluorescence light microscopy using butyl-methyl-methacrylate (BMM) and polyethylene glycol (PEG) embedded sections, transmission electron microscopy (TEM) and field emission scanning electron microscopy (FESEM). The use of PEG embedded embryos provided a rapid method for light microscope detection of antigens before committing to FESEM analysis. BMM embedded specimens provided well preserved suspensor cells for immunofluorescence. FESEM permitted high resolution observation of large areas of the inner surface of the plasma membrane and associated cell organelles. Suspensor cells contained mostly transversely oriented cortical microtubules linked to the plasma membrane and adjacent microtubules by cross- bridges. Light and electron microscopy revealed numerous clathrin coated structures on the plasma membrane. These included flat patches of clathrin, coated pits and coated vesicles. Many coated vesicles were associated with microtubules. Both tubular and lamellar endoplasmic reticulum were observed on the plasma membrane by FESEM.     

A scanning electron microscope study of the development of free axonal sprouts at the cut ends of dorsal spinal nerve roots in the rat

Summary Rat dorsal spinal nerve roots were cut and the tip on the ganglionic side of the cut was examined by scanning electron microscopy at 0, 7, 20 and 48 h after operation. Seven hours after cutting, free axonal sprouts had started to protrude from the cut end of the nerve. After 20 h the free sprouts were more profuse than at 7 h but were smaller and had a rougher surface. At both 7 and 20 h many of the sprouts consisted of a stalk 2–7 m in diameter with a bulbous end 5–20 m in diameter. A few branching sprouts were seen. At 48 h the sprouts were shrunken with a deeply furrowed surface. The significance of the surface structure of the sprouts is discussed.I.R.D. is supported by the Medical Research Council; P.K. thanks the Mental Health Trust for a project grant     

Low-temperature electron microscopy for the study of polysaccharide ultrastructures in hydrogels. I. Theoretical and technical considerations

The high-pressure freezing (HPF) technique was applied to the cryo-immobilization of alginate gels and the quality of the freezing analyzed on a TEM by comparison of the segregation pattern of samples of decreasing thickness. Dynamic simulations of heat transfer within an idealized slab of pure water surrounded by two walls of aluminium were performed to illustrate the effect of the heat-transfer coefficient by convection on the cooling rate of the sample. Heat-transfer coefficients in liquid nitrogen and liquid propane at ambient pressure were measured using a carefully characterized thermocouple and the values incorporated as parameters in heat-transfer simulations to compare the efficiency of the plunge-freezing technique with the high-pressure freezing technique. Values of the heat-transfer coefficient in liquid nitrogen and liquid propane, calculated between 273 K and 173 K were 670 and 18420 W/m(2)/K, respectively. Based on TEM observations and the results of heat-transfer simulations, the HPF technique was adapted to the cryo-fixation of 50-microm-thick alginate gels. The occurrence of artifacts was rejected because no differences were observed in the pattern of cryo-fixed and freeze-substituted samples of various thickness, with and without ethanol as cryo-protectant. A sample thickness of 50 microm was found to ensure an adequate preservation of structures as small as a few nanometers, as verified by TEM and SEM observations. Finally, DSC measurements on alginate solutions and alginate beads revealed that under the experimental conditions (0-3%), alginate cannot be considered to be an efficient cryo-protectant.     

The functional morphology of the scolex of Diphyllobothrium Cobbold (Cestoda, Pseudophyllidea). A scanning electron and light microscopical study on scoleces of adult D. dendriticum (Nitzsch), D. latum (L). and D. ditremum (Creplin).

Scoleces of adult D. dendriticum, D. latum and D. ditremum were studied using scanning electron and light microscopy. In D. dendriticum and D. latum the transverse bar between the bothria at the scolex apex always possessed a frontal pit while in D. ditremum no such invagination was seen. The scoleces attach to the host intestine by taking a firm grip with each bothrium around one or two intestinal villi. In D. ditremum and D. latum a layer of secreted substance is present between the scolex apex and the host intestine and between the inner bothrium surface and the host intestinal villi. In D. dendriticum, however, such a secreted layer was only observed at the scolex apex. A general account of the musculature and nervous and excretory systems of a Diphyllobothrium scolex is given.     

High resolution scanning electron-microscopic study on the three-dimensional structure of the sarcoplasmic reticulum in the slow (tonic) muscle fibers of the frog,Rana nigromaculata

Summary The three-dimensional structure of the sarcoplasmic reticulum (SR) in the slow (tonic) fibers of the reclus abdominis muscle of the Japanese meadow frog (Rana nigromaculata nigromaculata Hallowell) was examined by high resolution scanning electron microscopy, after removal of the cytoplasmic matrices by the osmium-DMSO-osmium procedure. The SR forms a repetitive network throughout these fibers. At the level of the Z-line, a slender transverse tubule (T-tubule) runs transversely to the longitudinal axis of the myofibril. Small, spherical or ovoid terminal cisternae couple laterally with the T-tubule at intervals of 0.4–1.0 m, and form a terminal cisterna-T-tubule complex on whose surface tiny indentations are occasionally seen. Each terminal cisterna gives rise to a few sarcotubules that run in various directions, divide frequently and form circular or oval meshes of diverse sizes in front of the A- and I-bands. The sarcotubules usually form small meshes in the middle of the A-band, but occasionally fuse and form a poorly developed H-band (fenestrated) collar.     

A study of the relative importance of the peroxiredoxin-, catalase-, and glutathione-dependent systems in neural peroxide metabolism

Cells are endowed with several overlapping peroxide-degrading systems whose relative importance is a matter of debate. In this study, three different sources of neural cells (rat hippocampal slices, rat C6 glioma cells, and mouse N2a neuroblastoma cells) were used as models to understand the relative contributions of individual peroxide-degrading systems. After a pretreatment (30 min) with specific inhibitors, each system was challenged with either H₂O₂ or cumene hydroperoxide (CuOOH), both at 100 μM. Hippocampal slices, C6 cells, and N2a cells showed a decrease in the H₂O₂ decomposition rate (23-28%) by a pretreatment with the catalase inhibitor aminotriazole. The inhibition of glutathione reductase (GR) by BCNU (1,3-bis(2-chloroethyl)-1-nitrosourea) significantly decreased H₂O₂ and CuOOH decomposition rates (31-77%). Inhibition of catalase was not as effective as BCNU at decreasing cell viability (MTT assay) and cell permeability or at increasing DNA damage (comet test). Impairing the thioredoxin (Trx)-dependent peroxiredoxin (Prx) recycling by thioredoxin reductase (TrxR) inhibition with auranofin neither potentiated peroxide toxicity nor decreased the peroxide-decomposition rate. The results indicate that neural peroxidatic systems depending on Trx/TrxR for recycling are not as important as those depending on GSH/GR. Dimer formation, which leads to Prx2 inactivation, was observed in hippocampal slices and N2a cells treated with H₂O₂, but not in C6 cells. However, Prx-SO₃ formation, another form of Prx inactivation, was observed in all neural cell types tested, indicating that redox-mediated signaling pathways can be modulated in neural cells. These differences in Prx2 dimerization suggest specific redox regulation mechanisms in glia-derived (C6) compared to neuron-derived (N2a) cells and hippocampal slices.     

Hydration, structure, and molecular interactions in the headgroup region of dioleoylphosphatidylcholine bilayers: an electron spin resonance study

The relationship between bilayer hydration and the dynamic structure of headgroups and interbilayer water in multilamellar vesicles is investigated by electron spin resonance methods. Temperature variations of the order parameter of a headgroup spin label DPP-Tempo in DOPC in excess water and partially dehydrated (10 wt % water) show a cusp-like pattern around the main phase transition, Tc. This pattern is similar to those of temperature variations of the quadrupolar splitting of interbilayer D2O in PC and PE bilayers previously measured by 2H NMR, indicating that the ordering of the headgroup and the interbilayer water are correlated. The cusp-like pattern of these and other physical properties around Tc are suggestive of quasicritical fluctuations. Also, an increase (a decrease) in ordering of DPP-Tempo is correlated with water moving out of (into) interbilayer region into (from) the bulk water phase near the freezing point, Tf. Addition of cholesterol lowers Tf, which remains the point of increasing headgroup ordering. Using the small water-soluble spin probe 4-PT, it is shown that the ordering of interbilayer water increases with bilayer dehydration. It is suggested that increased ordering in the interbilayer region, implying a lowering of entropy, will itself lead to further dehydration of the interbilayer region until its lowered pressure resists further flow, i.e., an osmotic phenomenon.     

Biogenic silver nanoparticles reduce Toxoplasma gondii infection and proliferation in RAW 264.7 macrophages by inducing tumor necrosis factor-alpha and reactive oxygen species production in the cells

Owing to the serious adverse effects caused by pyrimethamine and sulfadiazine, the drugs commonly used to treat toxoplasmosis, there is a need for treatment alternatives for this disease. Nanotechnology has enabled significant advances toward this goal. This study was conducted to evaluate the activity of biogenic silver nanoparticles (AgNp-Bio) in RAW 264.7 murine macrophages infected with the RH strain of Toxoplasma gondii. The macrophages were infected with T. gondii tachyzoites and then treated with various concentrations of AgNp-Bio. The cells were evaluated by microscopy, and culture supernatants were collected for ELISA determination of their cytokine concentration. Treatment with 6 μM AgNp-Bio reduced the infection and parasite load in infected RAW 264.7 macrophages without being toxic to the cells. The treatment also induced the synthesis of reactive oxygen species and tumor necrosis factor-alpha (both pro-inflammatory mediators), which resulted in ultrastructural changes in the tachyzoites and their intramacrophagic destruction. Our findings suggest that AgNp-Bio affect T. gondii tachyzoites by activating microbicidal and pro-inflammatory mechanisms and may be a potential alternative treatment for toxoplasmosis.