Adaptations of Goldner's Masson Trichrome Stain for the Study of Undecalcified Plastic Embedded Bone

Specialized adaptations for application of Goldner's Masson trichrome stain to plastic embedded undecalcified bone specimens are presented. This stain can be used successfully on methyl-glycol methacrylate, glycol methacrylate and Spurr embedded bones. The stain affords the advantage of good cellular staining due to the hematoxylin component with concomitant sharp discrimination of mature bone matrix which stains green, immature new bone matrix which stains red, and calcified cartilage which stains very pale green. Use of red filters during photomicrography aids in bone-osteoid discrimination in black and white photographs.     

Technique-dependent variations in cerebral microvessel blood volumes and hematocrits in the rat.

To quantitate small parenchymal microvessel blood volumes in the brain, the distribution spaces of radiolabeled red blood cells (RBC) and serum albumin (RISA) were assessed in rats by different methods of tissue sampling and radioassay. Three minutes after intravenous administration of 55Fe-RBCs and/or 125I-RISA, the rats were decapitated. The brain was either immediately frozen within the skull and later removed (head-frozen group) or rapidly removed from the skull and then frozen (brain-frozen group). Radioactivity was measured either by liquid scintillation counting of tissue pieces, which contained pial plus large and small parenchymal microvessels, or by quantitative autoradiography (QAR) of tissue sections, which indicated small parenchymal microvessel blood only. In 12 of 15 areas, the RISA, RBC, and blood volumes determined by liquid scintillation counting of head-frozen tissue pieces were equal to or greater than those of brain-frozen tissue; this indicated less than or equal to 25% greater blood retention in pial and parenchymal microvessels with head freezing. At the parenchymal microvessel level (QAR assay), the distribution volumes of RBCs, RISA, and blood were similar with the two freezing techniques; hence with QAR either freezing procedure can be used to assess small parenchymal microvessel blood volumes.     

Increases in arterial blood oxygen during exercise in the lemon shark (Negaprion brevirostris)

Summary Polyethylene cannulae were implanted in pre- and post-branchial blood vessels allowing nonstressful blood sampling over a variety of activity ranges in an active tropical elasmobranch, the lemon shark (Negaprion brevirostris). TheP ₅₀ was found to be 11.8 Torr at 24°C and pH of 7.7. A Bohr shift of –0.36 was also found. BloodP _{o 2} and oxygen content were measured during rest, routine swimming, and exercise in unanesthetized, free swimming juveniles. Under all conditions venous oxygen levels were low with venousP _{o 2} of 7.1±2.7 Torr, and venous oxygen content ( ) of 0.56±0.4 vol%. However, a large variability was found in arterial blood measurements. ArterialP _{o 2} ranged from 7 to 80 Torr, while arterial oxygen content (Ca_{o 2}) varied from 1.6 vol% to 6.8 vol% among ten experimental animals. A significant increase in arterialP _{o 2}, oxygen content, and hematocrit was noted during increased activity. Since the venous system provides little or no oxygen reserve, increased oxygen extraction from the blood ( ) appears to be met by an increase inCa_{o 2} rather than a decrease in . Mechanisms to accomplish this may include increasing hematocrit and vacular gill shunts.     

Endocytosis of lactate dehydrogenase isoenzyme M4 in rats in vivo. Experiments with enzyme labelled with O-(4-diazo-3,5-di[125I]iodobenzoyl)sucrose.

1. Pig lactate dehydrogenase isoenzyme M4 was labelled with O-(4-diazo-3,5-di[125I]iodobenzoyl)sucrose and injected intravenously into rats. Previous work has shown that this label does not influence the clearance of the enzyme (half-life about 26 min) and that it is retained within the lysosomes for several hours after endocytosis and breakdown of the protein [De Jong, Bouma & Gruber (1981) Biochem. J. 198, 45--51]. 2. The distribution of the radioactivity over a large number of tissues was determined 2 h after injection. A high percentage of the injected dose was found in liver (41%), spleen (10%) and bone including marrow (21%). 3. Autoradiography indicated uptake of the enzyme mainly by Kupffer cells of the liver, by spleen macrophages and by bone marrow macrophages. 4. Liver cells were isolated 1 h after injection of the enzyme. Kupffer cells, endothelial cells and parenchymal cells were found to endocytose the enzyme at rates corresponding to 4230, 35 and 25 ml of plasma/day per g of cell protein, respectively. 5. Previous injection of carbon particles greatly reduced the uptake of the enzyme by liver and spleen, but the uptake by bone marrow was not significantly changed.     

Identification of Strychnine Binding Sites in the Rat Retina

Abstract: [³H]Strychnine specifically binds to membrane fractions isolated from rat retinae. The binding is saturable, with an apparent dissociation constant, K _D, of 14.3 × 10⁻⁹ M and 205 fmol bound/mg protein. Specific binding is time-dependent and proportional to protein concentration. Glycine and taurine are equally potent inhibitors of [³H]strychnine binding ( K _i= 4 × 10⁻⁵ M); no other amino acids endogenously present in the retina inhibited [³H]strychnine binding.     

Nucleoside uptake by rat retina cells

Adenosine and guanosine uptake have been studied in the rat retina. Both nucleosides are taken up in a time- and temperature-dependent manner by dispersed rat retinal cells. The uptake of both nucleosides is Na⁺-dependent and Ca⁺⁺-independent. Initial rate studies of guanosine and adenosine uptake demonstrate a single uptake process for each nucleoside with K_D values of 2.1 and 2.9 uM, and maximal rates of 24 and 17 pmol/mg protein/min, respectively. Guanosine uptake was inhibited by adenosine with a K_I of 12.1 uM whereas guanosine inhibited adenosine uptake with a K_I value greater than 10^?3 M. LN⁶-phenylisopropyladenosine, a nucleoside analog, was the most potent inhibitor of adenosine and guanosine uptake with K_I values of 25 and 8 uM, respectively. Phosphodiesterase inhibitors (isobutylmethylxanthine and theophylline) and biogenic amines (dopamine, norepinephrine, and histamine) had no significant effect on the uptake of guanosine or adenosine at concentrations up to 100 uM.     

An ultrastructural search for lectin-binding sites on surfaces of spinach leaf organelles

Organelles isolated from leaves of spinach (Spinacia oleracea L.) were prefixed in glutaraldehyde and then incubated with ferritin conjugates of four lectins — Concanavalin A (Con A), Ricinus communis L. agglutinin, MW 120,000 (RCA), soybean agglutinin (SBA), and wheat germ agglutinin (WGA) — in order to probe their cytoplasmic surfaces for saccharide residues. In each case the major leaf organelles, including microbodies, mitochondria and chloroplast derivatives, failed to exhibit labeling when examined with the electron microscope. Tobacco (Nicotiana tabacum L.) leaf protoplasts, incubated simultaneously with and under identical conditions to the spinach organelles, showed specific labeling of their plasma membranes with all four lectin conjugates, thus establishing the efficacy of the procedure for demonstrating the presence of binding sites when they exist. Further attempts to show binding of one of the lectins, Con A, by labeling with fluorescein-Con A and by organelle agglutination, yielded results consistent with the absence of ultrastructural labeling. It is concluded that no saccharide residues recognized by the four lectins are present on the cytoplasmic surfaces of organelles and that those residues reported to be constituents of intracellular membranes, therefore, are most likely exposed on the luminal (extracytoplasmic) surfaces.Abbreviations Con A Concanavalin A - RCA Ricinus communis agglutinin, MW 120,000 - SBA soybean agglutinin - WGA wheat germ agglutinin