Pyruvic-acid-containing polysaccharide in the cell wall of Bacillus polymyxa AHU 1385

Three acidic polymer fractions with molecular masses of about 16 kDa, 35 kDa and 70 kDa were isolated from lysozyme digests of N-acetylated cell walls of Bacillus polymyxa AHU 1385 by ion-exchange chromatography and gel chromatography. These fractions, containing mannosamine, glucosamine and pyruvic acid in a molar ratio of about 1:1:1 together with glycopeptide components, were characterized as polysaccharide-linked glycopeptides with one, two and more polysaccharide chains. On the other hand, treatment of the cell walls with glycine/HC1 buffer, pH 2.5, at 100 degrees C for 10 min followed by separation of water-soluble products on ion-exchange chromatography gave three polysaccharide fractions, PS-I-III, which contained different amounts of pyruvic acid (0,0.6 and 0.9 residue/mannosamine residue) along with equimolar amounts of mannosamine and glucosamine. Pyruvate-free polysaccharides similar to PS-I were also obtained from PS-II, PS-III and polysaccharide-linked glycopeptides by treatment with 10 mM HC1 at 100 degrees C for 1 h. Results of analyses of these polysaccharide preparations by 1H-NMR and 13C-NMR measurement and methylation, together with data from characterization of fragments obtained by hydrogen fluoride hydrolysis, lead to the most likely structure, ----3)[4,6-O-(1-carboxyethylidene)]ManNAc(beta 1----4)GlcNac(beta 1----, for the acidic polysaccharide of this strain.     

Potassium channels in synaptosomal membrane examined using patch-clamp techniques and reconstituted giant proteoliposomes

Synaptosomes isolated from the rat cerebral cortex were mixed with sonicated phospholipid vesicles and subjected to freezing-thawing to acquire giant proteoliposomes. Membranes of these giant proteoliposome could thus be studied using patch-clamp techniques. Single-channel currents were measured with the inside-out patch of the membrane, in KCl solutions. Three different potassium channels were detected and unit conductances were 15.1, 28.6 and 91.0 pS, respectively, in a symmetrical 150 mM KCl solution. All these channels are more permeable to potassium than to sodium ions, the permeability ratio being about 2:1. Tetraethylammonium ions blocked these channels. The gating of these potassium channels is independent of the membrane potential, Presumably, these channels play a role in the resting membrane potential of presynaptic nerve terminals.     

A novel action of activin A: stimulation of insulin secretion in rat pancreatic islets

The present study was conducted to examine an action of activin A on insulin secretion from rat pancreatic islets. In a batch incubation system, activin A stimulated insulin secretion in a dose-dependent manner at concentrations higher than 1 nM. Furthermore, activin A greatly potentiated glucose-induced insulin release. When islets were perifused with 1 nM activin A, insulin secretion was barely affected in this system. However, the insulin response to 16.7 mM glucose was greatly enhanced. Both the first and second phases of insulin response were enhanced by 1 nM activin A. These results indicate that, in addition to its known actions on pituitary-gonadal and hematopoietic systems, activin A modulates the function of pancreatic islets and stimulates insulin secretion.     

Establishment of a new hereditary-cataract mouse (CSM) strain derived from SENCAR mouse

A mouse representing a new hereditary cataract strain was found in a mouse colony and a new line was established strain CSM. These mice were investigated genetically, histologically and biochemically. The results suggested that this cataract was apparently inherited through two recessive autosomal genes. Histologically the denucleation process of lens fibers was abnormal and small vacuoles appeared in the equatorial region of the lens cortex at 12 days. Biochemically, insoluble protein and sodium increased in the lens with age.     

Increased erythrocyte catalase activity in patients with hyperthyroidism

Levels of human erythrocyte catalase activity were determined in 38 patients with thyroidal dysfunction. In patients with hyperthyroidism, erythrocyte catalase activities were found to be higher than the levels of normal subjects (P less than 0.001). In hypothyroidism, erythrocyte catalase activities were of the same order as those of normal subjects. Significantly high positive correlation was found between erythrocytes catalase activity and the levels of thyroxine (r = 0.5794, n = 36, P less than 0.001), and slight positive correlation was detected between catalase activity and the levels of triiodothyronine (r = 0.3978, n = 33, P less than 0.05). A decreased erythrocyte catalase activity was observed when erythrocytes lysate was incubated with thyroid hormones. It was suggested that erythrocyte catalase activity had close relationship with thyroid state, however, direct effect of thyroid hormones were not observed on erythrocyte catalase assay system in vitro.     

Stimulation of Amino Acid Uptake and Na+, K+-ATPase Activity by Norepinephrine in Superior Cervical Sympathetic Ganglia Excised from Adult Rats

Active uptake of a labelled nonmetabolizable amino acid, alpha-aminoisobutyric acid (AIB), into isolated superior cervical sympathetic ganglia (SCG) excised from adult rats was considerably stimulated by the addition of either norepinephrine (NE, 50 microM) or 3,4-dihydroxyphenylethylamine (dopamine, DA, 100 microM) to the medium during aerobic incubation for 2 h at 37 degrees C. The NE-induced increase in AIB uptake was significantly antagonized by the addition of an alpha 1-adrenoceptor antagonist (prazosin, 10 microM) in SCG axotomized 1 week prior to the examination, in which most of the ganglionic neurons had degenerated and reactive proliferation of the satellite glial components was in progress. The addition of neither acetylcholine (ACh, 1 mM) plus eserine (0.1 mM) nor cyclic nucleotides (1 mM) changed the AIB uptake by the SCG. In the axotomized SCG, the NE-evoked increase in AIB uptake was much more pronounced than that of intact or denervated SCG. A kinetic study of the active AIB uptake in the SCG showed that NE produced a decrease of the Km value and an increase in the Vmax, especially in the axotomized SCG. Ganglionic Na+, K+-ATPase activity was greatly stimulated in the presence of NE, but not by ACh. These results strongly suggest that the NE-induced enhancement of active AIB uptake in the isolated SCG is occurring in glial cells rather than in neuronal cells, with a possible alteration of membrane properties for amino acid uptake and with an apparent regulation by the stimulated transport enzyme Na+, K+-ATPase.     

Enhanced formation of mouse hybridomas without hat treatment in a serum-free medium

Summary A newly developed, serum-free medium (NYSF-404) selects for antibody-producing hybridomas after fusion of antigen-sensitized mouse spleen cells with myeloma cell lines P3-X63-Ag8-U1 (P3-U1), P3-X63-Ag8-6.5.3 (Ag8.653), or P3-NSI/1-Ag4-1 (NS-1). Without the need for hypoxanthine-aminopterinthymidine (HAT) selection of hybrid cells, frequency of hybridoma formation in medium NYSF-404 is higher (twice) than that in serum- and HAT-containing medium. Colonies developed upon limiting dilution in the presence of the mortal parent myeloma cells in medium NYSF-404 and pure culture of antibody-secreting cells could be subsequently established. The results suggest that fusions can be done in serum-free medium and that the clonal growth of hybridomas is dependent on factors produced by parent myeloma cells under serum-free culture conditions. Such factors seem deficient in serum- and HAT-containing medium or are masked by serum.     

Release of endogenous prostaglandins by mild hyperosmotic saline inhibits tetragastrin-stimulated gastric acid secretion in rats

Filling of the gastric lumen of rats with 1.0 M NaCl solution (5 ml) for 10 min under urethane anesthesia caused an increase in the gastric fluid concentrations of prostaglandin (PG) E₂, 13, 14-dihydro-15-keto-PGE₂ and 6-keto-PGF_1α as determined by radioimmunoassay. PGE₂ was the major PG generated. The levels of PGE₂ in the gastric fluid were increased dose-dependently after filling the lumen with 0.3, 0.5, 0.7 or 1.0 M NaCl solutions. The pH of the gastric fluid increased similarly after 0.5 to 1.0 M NaCl solutions. Indomethacin (10 mg/kg, i.p.) suppressed the PGE₂ increase caused by 1.0 M NaCl solution, but did not prevent the increase of the pH of the gastric fluid induced by intragastric 1.0 M NaCl. Infusion of tetragastrin (62.5 μg/kg/hr, i.v., for 10 min) caused a marked increase of acid secretion without modifying intragastic concentration of PGE₂. The acid secretion due to tetragastrin was completely inhibited after intragastric administration of 1.0 M NaCl solution, while indomethacin restored the tetragastrin-induced acid secretion, with prevention of a rise of intragastric PGE₂ levels. These observations suggest that 1.0 M NaCl solutions suppress basal intragastric acid through a mechanism which is independent of prostaglandins. In contrast, the suppression of tetragastrin-induced acid secretion by intragastric 1.0 M NaCl solution appears to be mediated through a release of prostaglandins     

Chromosomal localization of the Epstein-Barr virus (EBV) genome in Bloom's syndrome B-lymphoblastoid cell lines transformed with EBV

The localization of the Epstein-Barr virus (EBV) genome in chromosomes of human B-lymphoblastoid cell lines (LCLs) transformed with EBV, and the effect of EBV DNA on the level of sister chromatid exchange (SCE) in Bloom's syndrome (BS) B-LCLs, were examined with chromosomal in situ hybridization techniques using a ³H-EBV DNA probe. EBV DNA was detected in chromosomes 1–5 and 13–15 at specific G band regions in BS as well as in normal B-LCLs, regardless of SCE. Several chromosomal sites (1p31, 1q31, 4q22–24, 5q21, 13q21, 14q21) carrying EBV DNA seemed to be very characteristic in normal as well as in BS B-LCLs. There was no statistically significant difference in silver grain counts due to EBV DNA and their distribution in different chromosomes or groups among normal and BS B-LCLs with normal and high SCE. These findings strongly indicate that EBV infection did not introduce a correcting factor for BS SCE.     

Immunohistochemical studies of the serotonergic supraependymal plexus in the mammalian ventricular system, with special reference to the characteristic reticular ramification

Distributional and morphological features, especially characteristics of the ramification of serotonin-containing supraependymal fibers (SEF), were studied in the ventricular systems of mammals (mouse, rat, guinea pig, rabbit, cat, dog, monkey) by means of a modified peroxidase antiperoxidase technique, using antiserotonin antiserum prepared in our laboratory. SEF were present in all ventricular systems, except on the third ventricle floor and in the choroid plexus. The density of SEF was higher in the smaller species. In the rat, light- and scanning electron microscopical SEF were almost completely abolished 1 week after intraventricular administration of 5,6-dihydroxytryptamine. Ramification of SEF was complicated; the SEF formed a true network with frequent anastomosing. In the ventricular system of rats rendered hydrocephalic by kaolin administration, the mode of axonal branching in the supraependymal plexus could best be analyzed by the scanning electron microscope because the meshes of the plexus were spread out.