Bradykinin dilates rat middle cerebral artery and its large branches via endothelial B2 receptors and release of nitric oxide

Ring segments of rat middle cerebral artery (MCA) were prepared for measurement of isometric force and precontracted with 10⁻⁴ M uridine triphosphate (UTP). Concentration-effect curves (CEC) were constructed for bradykinin (BK, 10⁻⁸–10⁻⁵ M) in segments with functionally intect (E+) or denuded (E−) endothelium. E− segments did not dilate to BK. The BK receptor was characterized by application of specific B₁ or B₂ antagonists [des-Arg⁹-Leu⁸] BK (10⁻⁵ M) and [ -Arg⁰-Hyp³-Thi⁵- -Tic⁷-Oic⁸] BK (HOE140,3 × 10⁻⁷ M), respectively, or B₁ agonist [des-Arg⁹] BK (10⁻⁸–10⁻⁴ M). Involvement of nitric oxide (NO) was tested with N^G-nitro- -arginine (LNNA, 10⁻⁴ M). BK induced concentration-dependent relaxation with a maximal effect (E_max) of 40.86 ± 1.50% at 10⁻⁶ M and a pD₂ (−log₁₀ EC₅₀) of 6.818 ± 0.044. This relaxation could be prevented with HOE140 or LNNA, but was not influenced by [des-Arg⁹-Leu⁸] BK. [des-Arg⁹] BK did not induce any effect. These results demonstrate that BK induced relaxation via endothelial B₂ receptors and release of NO in isolated rat MCA.     

A Simple Histochemical Reaction for Aldehydes

A study has been made on the possibility of replacing leucofuchsin by colored basic fuchsin for the histochemical demonstration of aldehydes. Several tissues from mammals and various pertinent fixatives were used. Aldehydes were freed from carbohydrates by oxidation and from thymonucleic acid by hydrolysis.

It was found that the colored form and not necessarily the leucoform of basic fuchsin can be used histochemically in demonstrating aldehydes. The technic used is as follows: (1) Treat with 1.0-0.5% H₅IO₆ (or in 1% KIO₄ in M/1 H₂SO₄) for 5 to 10 min. and wash thoroughly. For thymonucleic acid hydrolize with N HCl 5 min. at room temperature, 10 min. at 60°C. and 5 min. at room temperature. (2) Stain for 2-3 min. with 0.05% basic fuchsin in 5% ethanol, 3% phenol. (3). Transfer immediately to 1 or 2 changes of 1% sodium bisulphite or potassium metabisulphite in 0.1-0.2 N H₂SO₄ for a total of 5 min. (4) Rinse with water and treat with M H₂SO₄ in 95% ethanol for 3-5 min. 6. Wash thoroughly in water and dehydrate, clear, and mount. For glycogen and mucin the following counterstaining solution is recommended: orange G, 0.25 g.; light green SFY, 0.10 g.; phosphotungstic acid 0.50 g.; 50% ethanol, 100 ml.; glacial acetic acid, 0.25 ml.     

Characterization and Separation of Some of the Coordination Compounds of Chromium and Aluminon

Chromic nitrate and aluminon (ATA) react to form highly colored lakes. Examination of these lakes by the Vosburgh and Cooper method has led to the identification of a pigment [(ATA)Cr(H₂O)_x] which is insoluble in water and absolute alcohol, a deep red anionic component [(ATA)_BCr(H₂O)_x]^-y, and a purplish-red cationic component [(ATA)Cr₄(H₂O)_x]^+y. These components of the lake have been isolated and separated in the dry form. The anionic compound is apparently a simple coordination compound and can be used as a cytoplasmic stain while the cationic component is a chelate and can be used as a rather selective nuclear stain. In addition to these components, a number of other components were also found. Not only the ratio between chromium and aluminon but also the concentration of the reactants influences the formation of these different components. The amount of pigment formed is maximal with a molar ratio of 1 and a concentration of 10^-2 M or 10^-1 M. The anionic component is maximal with a molar ratio of 1 Cr to 6 aluminon at the 10^-1 M concentration, the cationic component is maximal with a molar ratio of 4 chromium to 1 aluminon at the 10^-1 M concentration. None of these are formed at the 10^-4 M concentration but only a deep redpurple soluble compound at the 1 : 1 ratio, which was not further investigated.     

A Gelatin Film Method for Improved Histochemical Localization of Dehydrogenases in Plant Cells

Glucose-6-phosphate and 6-phosphogluconate dehydrogenases diffused from frozen sections of Vicia faba embryos during histochemical incubation. In the liquid incubation medium, the dehydrogenases catalysed the oxidation of substrate and reduction of NADP. NADPH₂ thus formed could lead to artifactual deposition of formazan in frozen sections. The addition of 20% polyvinyl alcohol to the incubation medium was found unsatisfactory in preventing this loss which appeared to be overcome by incorporating the reaction mixture into a gelatin film. Equal volumes of 10% gelatin solution in 0.05 M phosphate buffer at pH 7.8, and the enzyme reaction medium containing twice the normal concentration of substrate (0.014 M), of 0.007 M pyridine nucleotide, of 0.02 M KCN and of 0.0024 M NBT in the buffer, were mixed and layered onto polyethylene, and allowed to set in the dark at room temperature for 30-60 min. The solidified medium and its support were cut into strips and layed onto unfixed, frozen sections of plant tissues which were incubated at 20 C. Evidence is presented to support the supposition that the enzymes are retained in the sections during the reaction.     

Hydrodynamic properties of the fractions of mannan formed by Rhodotorula rubra yeast

Aqueous solutions of fractions of an extracellular linear mannan formed by Rhodotorula rubra yeast have been investigated by hydrodynamic methods (high-speed sedimentation, translation isothermic diffusion and viscometry). The molecular weight was determined according to Svedberg ( ) and the polydispersity parameters of the initial sample were also determined (M_w/M_n = 1·20 and M_z/M_w = 1·21). Relationships between the molecular weight (M) and s_o, D_o and [η] in the range were: [η] = 2·33 × 10⁻² M^0.75, D_o = 1·65 × 10⁻⁴ M^0·58, s_o = 2·24 × 10⁻¹⁵ M^0·43. The equilibrium rigidity and hydrodynamic diameter of chains representing mannan molecules were evaluated.     

Removal of Ribonucleic Acid from Bacterial Cells by Dilute Salt Solutions

Bacterial cells were impressed upon a clean glass slide, fixed in ethyl alcohol and immersed at 37°C in either of the following two salt solutions: (A) NaCl, 7.8 gm; KCl, 0.7 gm; distilled water, 1000 ml; adjusted to pH 7.0; or (B) 0.1M NaH₂PO₄, 400 ml; 0.1M Na₂HPO₄, 600 ml; KCl, 0.7 gm. After 1-5 hr soaking to remove ribonucleic acid, the slide was stained by Giemsa's method as usual. The staining revealed slender chromatinic bodies with reasonable clarity extending the whole diameter of the moderately swollen cell. The results of this method seemed to be much like those obtained after ribonuclease digestion.     

Fine-Structure Locations of α-Glucan Phosphorylase, As Shown by Lead Precipitation and Electron Microscopy

Sites of glucan phosphorylase activity in fine structures, as shown by the lead precipitation method (Hori, Stain Techn., 39: 275, 1964) were studied by electron microscopy. Rat livers were fixed 2 hr at 0 C in buffered 2.5% glutaraldehyde, frozen-sections cut and incubated in the medium containing glucose-1-phosphate, 2.7 mM; NaF, 20 mM; acetate buffer, pH 5.8, 80 mM; Pb(NO₃)₂, 4.2 mM; and sucrose, 0.44 M; refixed in buffered 1% OsO₄, dehydrated and embedded in Epon 812 as usual. The reaction product was found in close association with endoplasmic reticulum, but not in mitochondria, nuclear membrane and the cisternae of endoplasmic reticulum. The possibility of demonstrating by the present method the indirect hydrolysis of glucose-1-phosphate through the phosphoglucomutase-glucose-6-phosphatase system was ruled out by inhibiting glucose-6-phosphatase with fluoride and ethanol.     

Binding and biologic activity of neurotensin in guinea pig ileum

Experiments were performed to relate receptor binding to biologic activity for the contractile effect of neurotensin (NT) in guinea pig ileum. The contractile response was examined on pieces of ileum under 1 g tension in a 5 ml bath of oxygenated Tyrode's at 38°C. NT contracted the longitudinal muscle (ED₅₀, 0.3 nM), the 2–3 g response peaking at 1 min and fading rapidly. In the presence of atropine (1 μM), ≥50% of the response was blocked and the residual effect gave an ED₅₀ of 1.4 nM. In the presence of atropine and CP-96,345, a substance P receptor antagonist (0.2 μM), no contraction was observed at 20 nM NT. Thus, there were two components to the response, one involving acetylcholine (ED₅₀, 0.3 nM) and one substance P (ED₅₀, 1.4 nM). Using membrane preparations and ¹²⁵I-labeled NT, specific, high affinty receptors for NT were demonstrated in the muscle and myenteric plexus. Scatchard analyses indicated the presence of two binding sites (K_ds: 0.1 nM and 2 nM). Sodiu ion and GTP analogs inhibited binding. Binding and biologic activity were similar in regard to dependence on specific groups within NT and sensitivity to metal ions. The high potency of Hg⁺⁺ was consistent with an involvement of free sulfhydryl group(s) in the binding reaction; this was supported by work with SH-directed agents. The results suggest that two receptor types or configurations may mediate the two components of the contractile effect of NT on guinea pig ileum.     

Simultaneous Formalin Fixation and Edta Decalcification, With Carbowax Embedding for Preservation of Acid Phosphatase

Carbowax serial sections from pubic symphyses of female mice, fixed and decalcified in a 10% formalin-5% Versenate solution for 18 hr at 4 C, pH 5.2, were incubated for 30 min with Burstone's simultaneous coupling reagent (pH 5.2); substrate: naphthol AS-TR and the diazonium salt, fast red violet L.B. All sections were counterstained with 1% methyl green at pH 4.0 in a phospho-citrate buffer. Inhibition by 0.01 M NaF, 0.0002 M CuCl₂, 10% tartaric acid and 0.01 M NaCN, as well as substrate-deficient and heat-inactivated controls, demonstrated conclusively that acid phosphatase was functionally preserved. Strong enzymatic activity was exhibited by osteoclasts, chondroclasts and free multinucleated giant cells. In addition, megakaryocytes, histiocytes, plasma cells, and monocytes exhibited moderate activity. The results demonstrated the technique to be consistently reproducible.     

Amino-functionalized zirconocene (C5H4CH(Me)NMe2)2ZrCl2, a catalyst for the dehydropolymerization of PhSiH3: Probing of peripheral substituent effects on catalyst dehydropolymerization activity

The amino-functionalized metallocene (C₅H₄CH(Me)NMe₂)₂ZrCl₂, [(Cp^N)₂ZrCl₂] was synthesized by salt metathesis of ZrCl₄ and 2 equiv. of C₅H₄CH(Me)NMe₂Li. The metallocene was obtained in good yield as a mixture of rac and meso diastereomers as established by NMR spectroscopy. The addition of 2 equiv. of n-BuLi to the metallocene (Cp^N)₂ZrCl₂ produced a co-catalyst system which was active, at a 1.0 mol% loading, in the dehydropolymerization of PhSiH₃ to poly(phenylsilane), PPSi. The PPSi was obtained as a 9:1 linear–cyclic mixture (M_w=3850, M_n=2300) as established by GPC analysis; ²⁹Si{¹H} NMR spectroscopy revealed an atactic polymer microstructure.     

The Destruction of Cytoplasmic Basophilia with Mineral Acids

Cytoplasmic basophilia may be selectively destroyed by the mineral acids, HNO₃, HCl and H₂SO₄. Their specificity is similar to that of ribonuclease. The optimal conditions for their use are 3°C. for 16 hours at 2M concentrations. Removal of cytoplasmic basophilia as with ribonuclease, malt diastase and perchloric acid is most effective on sections prepared from tissues fixed in solutions containing no chromates. Under the conditions herein reported the mineral acids appear to be a satisfactory and economical substitute for ribonuclease or perchloric acid.     

Identical properties of transmembrane synaptic vesicle protein Mr 100,000 in Torpedo and Mr 86,000 in bovine brain

Synaptic vesicles derived from the Torpedo electric organ and bovine cerebral cortex contain concanavalin A binding transmembrane glycoproteins of M_r 100,000 and 86,000, respectively. Their isolelectric points range from 5.5 to 6.0. On deglycosilation both glycoproteins yield identical products of M_r 62,000. The fully glycosilated and the deglycosilated proteins from both Torpedo and bovine brain are recognized by the monoclonal anti-SV2 antibody (Buckley and Kelly, J. Cell Biol. 100, 1284–1294, 1985) as well as by a monospecific IgG fraction raised against Torpedo vesicles and immunopurified against the bovine brain M_r 86,000 glycoprotein. This is shown by Western blotting as well as by immunoaffinity isolation with one antibody and immunodetection with the other antibody. Furthermore on immunohistochemical analysis of the Torpedo electric organ both antibodies recognize exactly the same nerve terminal ramifications. It is concluded that the glycoproteins of M_r 100,000 in Torpedo and of M_r 86,000 in bovine brain are corresponding proteins with different degrees of glycosilation.     

Handling Germinated Pollen on Millipore Membrane During the Feulgen and Autoradiographic Procedures

To prevent loss of pollen during the Feulgen's procedure, the pollen was grown on an autoclaved membrane filter (Millipore AA WP 025 00) in contact with a sterilized medium containing agar 0.5-1%, sucrose according to the genus (Malus 0.3-0.5 M; Persica and Tulipa 0.4 M), and H₃BO₃, 0.01%. To fix the germinated pollen of most species, the membrane was placed for 2 hr to overnight at 2-4 C on filter paper wet with the following mixture: OsO₄, 1 gm; CrO₃, 1.66 gm; and distilled water, 233 ml. To fix Persica pollen, 10% of glacial acetic acid had to be added to the fixative. Washing with distilled water and bleaching with a mixture of 3% H₂O₂ and sat. aq. ammonium oxalate, 1:1, were performed also on filter paper. Similarly, the preparation was processed for Feulgen staining by use of pieces of filter paper wet with the required fluids. Hydrolysis preceding the Schiff's reagent was performed at room temperature with 5 N HCl for 18 min. The differentiation after the Schiff's action was with 2% K₂S₂O₅ buffered to pH 2.3 with 9 ml of phosphate buffer (KH₂PO₄, 1.4 gm; conc. HCl, 0.35 ml and distilled water to make 100 ml). The stained pollen was floated off the membrane with a drop of glacial acetic acid to a gelatinized or an albumenized slide, and squashed. When the coverslip is removed the preparation may be either dehydrated and mounted or coated with autoradiographic film.     

林分密度对马尾松林下植物与土壤种子库多样性的影响

林分密度是影响林下植物多样性的重要因子。本研究以马尾松人工林为对象,设置低密度(1575株·hm^-2,D₁)、中密度(2474株·hm^-2,D₂)和高密度(3550株·hm^-2,D₃)3个林分密度,分析林下植物和土壤种子库多样性及二者的关系,为马尾松人工林实现多目标可持续发展提供科学依据。结果表明: 3种林分密度林下草本与灌木植物共有42科62属70种,D₁密度以喜光植物种类分布较多,而D₂、D₃密度以耐阴植物为主;3种林分密度间的草本、灌木的Margalef (M)、Shannon (H)、Simpson (D)、Pielou (J_sw)、Alatalo (Al)指数均随林分密度增加而呈下降趋势,且在草本与灌木层中对密度的响应不同。在草本层中,D₁与D₃的H、D、J_sw、Al指数均存在显著差异;在灌木层中,不同林分密度J_sw和Al指数存在显著差异,H和D指数差异不显著。土壤种子库H、D、J_sw和Al指数均随林分密度增加呈先下降后升高趋势,D₁密度的物种丰富度及多样性最高;不同林分密度下土壤种子库Jaccard与Sorensen相似性系数均较低。草本层M与J_sw指数呈显著正相关;灌木层林分密度与H、D、J_sw、Al指数的相关性大于草本层,而草本、灌木层的林分密度与J_sw指数均呈显著负相关。林分密度1575株·hm^-2是马尾松林下植被生长发育较为合适的密度,能够维持林下植物多样性,有利于马尾松人工林的可持续经营。     

Fluorescence characteristics of lyophilized maize chloroplasts suspended in buffer

Fluorescence transients were measured in lyophilized maize chloroplasts (suspended in Tris-maleate buffer (pH 6.6)) after extraction with heptane. (The fluorescence characteristics before extraction were qualitatively similar to those in the fresh chloroplasts.) The initial fluorescence level (m) in the (dry) heptane-extracted sample remained the same as in the unextracted material, but the variable fluorescence (Δm) was drastically diminished. A portion of variable fluorescence, however, could be restored by adding Na₂S₂O₄. If the heptane extraction was made in the presence of water (wet), the m level was almost as high as (or higher than) the final level (M) of the unextracted sample, and Δm was reduced. The “jet” of O₂ (that measures the pool size of the intersystem intermediate A) and the “microjet” (that measures the pool size of the reaction center complex E), present in the unextracted samples, were absent in both types of extracted samples. Some of the above data may be interpreted in a hypothesis in which two quenchers (Q₁ and Q₂) control the fluorescence (O → P) of chloroplasts — the reduction of Q₁ being responsible for the rapid and that of Q₂ for the slow fluorescence rise.     

Sequence analysis of mouse cDNAs encoding ribosomal proteins L12 and L18

Mouse cDNAs encoding ribosomal proteins (r-proteins), L12 and L18, were isolated and their sequences determined. The L12 cDNA was found to contain 639 bp, including a coding sequence of 498 nucleotides (nt), 5' (78 nt) and 3' (45 nt) untranslated regions (UTRs), and a poly(A) tail of 18 nt. The L18 cDNA was shown to consist of 648 bp, including a coding sequence of 567 nt, 5' (26 nt) and 3' (39 nt) UTRs, and a poly(A) tail of 16 nt. The nt sequences of the protein-coding region from the mouse L12 and L18 cDNAs were found to exhibit 96% and 92% identity, respectively, with those of the rat. With the use of mouse L12 and L18 cDNA probes, multiple (at least 10) copies of the L12 and L18 gene families were shown to be present in the mouse and rat genomes. However, there was no sequence heterogeneity detected among seven L18 cDNA clones, indicating that only one copy of the L18 gene-related sequences is functional, and the other copies are presumably nonfunctional pseudogenes. The complete amino acid (aa) sequences of the mouse r-proteins, L12 and L18, were deduced from the nt sequences of their cDNA clones. L12 has 165 aa and a M_r, of 17 790, while L18 has 188 aa and a M_r of 21 570. The aa sequences of the mouse r-proteins, L12 and L18, exhibit 98% and 94% identity, respectively, to those of rat.     

A Rhodocyan Technic for Staining the Anterior Pituitary

A reproducible, one-step, differential staining technic which uses routine formalin-fixed tissue and gives brilliantly contrasting results is produced by incubating sections for 1 hr in a 60° C oven in the following dye mixture: 1% eosin B (CI#771), 8 ml; 1% anilin blue (CI#707), 2 ml; and buffer solution (0.1M citric acid, 1.1 ml; 0.2M Na₂HPO₄, 0.9 ml; distilled water, 28.0 ml) at pH 4.5. No differentiation is necessary. The method can be modified for duodenal enterochromaffin cells and alpha cells of pancreatic islets by adjusting the buffer to pH 3.6 and staining for only 3 min at 60° C.     

Circadian change of heat loss in response to change in core temperature in rats

1. 1.|Circadian changes in heat production (M), heat loss (H), core temperature (T_c) and feeding activity (FA) of ad-lib fed rats were observed by direct and indirect calorimetry.

2. 2.|M, H and T_c showed a clear nocturnal increase associated with several discrete increases.

3. 3.|Whereas the slope of M vs T_c relation did not change appreciably within a day, the slope of H vs T_c or thermal conductance vs T_c relation tended to decrease at night, implying that the correlation between heat loss and body temperature is also a function of time of day in rats.

猴头菌高产麦角甾醇液体发酵工艺优化

以菌丝体中麦角甾醇产量为考察指标,在单因素实验的基础上用响应面分析法考察碳源、氮源和无机盐3个因素对麦角甾醇发酵产量的影响,以获得猴头菌液体发酵产麦角甾醇最优工艺,建立高产、稳定的猴头菌液体发酵产麦角甾醇生产工艺.经响应面分析,各因素按照对响应值的影响顺序为:氮源＞碳源＞无机盐,且氮源对麦角甾醇产量的影响极显著,碳源和...     

Day-night variation of thermoregulatory responses to intraperitoneal electric heating in rats

1. 1.Hypothalamic temperature (T_hy), nonevaporative heat loss (R + C + K), evaporative heat loss (E), thermal conductance (k), metabolic heat production (M) and heat storage (S) of rats were simultaneously measured by direct and indirect calorimetry during internal heat loading (2 W per rat) with an intraperitoneal electric heater.

2. 2.The tests were made twice a day; once during the day (1000–1200 h) and once at night (2200–2400 h) at an ambient temperature of 24°C.

3. 3.The resting values of T_hy, colonic temperature, (R + C + K), E, M and heart rate, and the T_hy threshold for tail skin vasodilation (T_th) during internal heat load were significantly higher at night than during the day.

4. 4.The slopes showing the relationshiop between (R + C + K), k or M and T_hy were significantly steeper during the day than at night after T_hy exceeded T_th.

5. 5.The slopes showing the relationship between E or S and T_hy were not different during the day and at night.

6. 6.These results indicate that the responses of nonevaporative heat loss and heat production to internal heat load vary with the time of day in rats.