Immunofluorescence staining of thin-filament sections not participating in actomyosin crossbridges: Studies by use of a monoclonal antibody specific to actin

Summary Monoclonal antibodies (mcab) were produced in vitro by fusing mouse X63-Ag8.653 plasmacytoma cells with spleen cells from a Balb/c mouse immunized with primary cultures of chick skeletal muscle (pmcc). After cloning on agar, stable clones were obtained, the antibodies of which stain specifically the I-band of myofibrils in the immunofluorescence (IF) procedure. For further characterization of these mcab their affinities to muscle proteins were tested by immunoblotting and by enzyme-linked immunosorbent assay (ELISA). Mcab specific for actin were revealed by these criteria. One of the anti-actin antibodies, mcab 647, reveals a variety of IF-staining patterns on myofibrils. On rest-length myofibrils the I-band is labeled only. However, at sarcomere lengths below 2 m, where the thin filaments meet in the middle of the A-band and form a region of double overlap, an additional fluorescent band appears in this position. The fluorescence intensity of this band is increased significantly in shorter sarcomeres. Finally, when the I-band has disappeared at a sarcomere length of 1.5 m, fluorescence is located exclusively in the middle of the A-band. These IF-staining patterns suggest that only those sections of the thin filament are stained that do not participate in actomyosin crossbridges.     

Chronic administration of an organophosphorus insecticide to rats alters cholinergic muscarinic receptors in the pancreas

Male rats were treated for 10 days with the organophosphorus insecticide, acetylcholinesterase inhibitor, O,O-diethyl S-[2-(ethylthio)ethyl]phosphorodithioate (disulfoton, 2 mg/kg/day by gavage). At the end of the treatment, binding of [³H]quinuclidinyl benzilate ([³H]QNB) to cholinergic muscarinic receptors and cholinesterase (ChE) activity were assayed in the pancreas. Functional activity of pancreatic muscarinic receptor was investigated by determining carbachol-stimulated secretion of α-amylase in vitro. ChE activity and [³H]QNB binding were significantly decreased in the pancreas from disulfoton-treated rats. The alteration of [³H]QNB binding was due to a decrease in muscarinic receptor density with no change in the affinity. Basal secretion of amylase from pancreas in vitro was not altered, but carbachol-stimulated secretion was decreased. The effect appeared to be specific since pancreozymin was able to induce the same amylase release from pancreases of control and treated rats. The results suggest that repeated exposures to sublethal doses of an organophosphorus insecticide lead to a biochemical and functional alteration of cholinergic muscarinic receptors in the pancreas.     

Temporal Distribution of [3H]-Imipramine Binding in Rat Brain Regions is Not Changed by Chronic Methamphetamine

Specific binding of [³H]-imipramine in the rat suprachiasmatic nuclei, occipital cortex and caudate putamen underwent significant and replicable changes throughout 24 hr under a light-dark cycle or under constant conditions. Daily variations were also found in the medial and dorsal raphe nuclei and the lateral hypothalamus. Methamphetamine, a psychoactive drug with marked effect on circadian rhythms in physiological and hormonal parameters and adrenergic receptors, did not have any significant effect on imipramine binding rhythms in eight discrete brain regions. Thus a drug known to reduce serotoninergic neurotransmission did not change characteristics of the modulatory binding site related to serotonin uptake.     

The effects of adrenal and gonadal steroids and K+-canrenoate on the metabolism of aldosterone by rat liver microsomes

The synthesis of polar aldosterone metabolites by rat liver microsomes at physiological concentrations of aldosterone (21.5 nM), was markedly inhibited by progesterone, testosterone, corticosterone, K+-canrenoate and estradiol-17 beta. In contrast, corticosterone and estradiol-17 beta significantly increased the synthesis of reduced aldosterone metabolites by 8- and 15-fold respectively, the majority of which were 5 alpha-reduced products of aldosterone. In experiments at higher substrate (aldosterone) concentrations (20-200 microM) the synthesis of ring A-reduced aldosterone metabolites by liver microsomes followed Michaelis-Menten kinetics with a Km[app] for aldosterone of 160 microM and Vmax[app] of 12.2 nmoles/mg protein/5 min. In these experiments progesterone, testosterone and K+-canrenoate all competitively inhibited the synthesis of reduced metabolites with inhibition constants (Ki [app]) of 70, 85 and 55 microM respectively; however, corticosterone did not. In contrast, estradiol-17 beta increased the rate of synthesis of reduced products by 40%, lowering the Km[app] to 83 microM.     

Preparative Electrophoresis of Isotopically Labeled L-Cell Interferons

The preparative method of polyacrylamide gel electrophoresis was adapted for purification and characterization of isotopically labeled L-cell interferons. Re-covery of interferon activity was quantitative, and purification and resolution were comparable to those obtained by analytical polyacrylamide gel electrophoresis. Ultimate specific activities attainable ranged from 2 x 10(6) to 3 x 10(6) international units per mg of protein.     

Der Einfluß von Elektrolyten auf Chloroplasten beim Gefrieren und Trocknen

Summary The effect of freezing, desiccation and various electrolytes on photophosphorylation, electron transport and some enzyme reactions of isolated spinach chloroplasts has been investigated. Freezing of broken chloroplasts took place at-25°C for 3 hrs; desiccation was performed at +2°C in vacuo over CaCl₂ for 3 hrs. The influence of various electrolytes during freezing or drying or during incubation of thylakoids or stroma enzymes for 3 hrs at +2°C in electrolyte solutions was determined. After treatment, the activities of a number of enzymes and enzyme systems were measured under normal conditions, e. g. in the absence of elevated electrolyte levels in a reaction medium which contained only the substrates and cofactors which are necessary for the respective enzyme reactions.Only photophosphorylation and electron transport were affected by freezing, desiccation and high concentrations of electrolytes; various soluble enzymes investigated here were not inactivated under the same conditions. In general, mild dehydration and lower concentrations of electrolytes resulted in an irreversible inactivation of ATP synthesis but did not impair ferricyanide reduction. With increasing dehydration or at higher concentrations of electrolytes the Hill reaction was also inhibited. In a certain range of dehydration and electrolyte concentration uncoupling of photophosphorylation from electron transport took place. Sugar protects the sensitive structures against the deleterious effect of both dehydration and high concentration of electrolytes.Various electrolytes affected thylakoid membranes differently. Inactivation of the membranes increased with increasing ion radius and decreasing hydration envelope of univalent or divalent cations. Divalent cations were more destructive than univalent cations. Anions did not follow these rules. Within a group of similar anions (halides or organic anions) effectivity decreased with increasing hydration envelope. On a molar basis, polyvalent anions were less effective than univalent anions. Inactivation by anions followed Hofmeister's series in seversed order. However, exceptions were observed and it appears that various ions affect the membrane in a specific manner.Inactivation of photophosphorylation and electron transport due to freezing or desiccation is identical to that due to high concentrations of electrolytes. This suggests that during dehydration due to freezing or drying the concentration of electrolytes in the remaining solution is responsible for the inactivation of the sensitive membranes.     

Flavonglykoside in Blüten vonPaeonia arborea undPaeonia suffruticosa

Summary Three glycosides have been isolated fromPaeonia arborea: kaempferol-3--glucoside-7--glucoside (Paeonoside), apigenin-7--glucoside, and apigenin-7-rhamnoglucoside (Rhoifolin).Paeonia suffruticosa also contains these three compounds but its main glycoside is kaempferol-3--glucoside (astragalin), which is present inPaeonia arborea only in traces.     

Über die genetischen Ursachen der Samenbildung

Zusammenfassung Innerhalb von Primula malacoides Franchet wurden zwischen verschiedenen Formen Valenzkreuzungen durchgeführt. Es traten 2x-, 3x-, 4x- und 5x-Pflanzen auf. An der Entstehung dieser Valenztypen sind sowohl reduzierte als auch unreduzierte Gameten beteiligt. Auf Grund der erzielten Kreuzungsergebnisse wird versucht, Rückschlüsse auf die genetischen Ursachen der Samenbildung zu ziehen.Es wird angenommen, daß der jeweiligen Konstitution des Endosperms eine vorrangige Bedeutung für die Entstehung von lebensfähigen Samen zukommt.

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On the genetic causes of seed formation

Summary New crossings were made between different diploid and polyploid forms of Primula malacoides Franchet. 2x, 3x, 4x and 5x plants appeared. Both reduced and unreduced gametes participated in the formation of these types. Based on the results conclusions are drawn about the genetic causes of seed development. It is suggested that the endosperm constitution has prime importance for the development of viable seeds.

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Herrn Prof. Dr. Hans Stubbe zum 65. Geburtstag gewidmet.

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Quedlinburger Beiträge zur Züchtungsforschung Nr. 78.     

Zytologische Beobachtungen an der hyperplastischen Nebennierenrinde des Menschen

Summary Clinical observations and the light and electron microscopic appearance of the adrenal cortex are described in a case of Cushing's disease associated with adrenocortical hyperplasia and a basophilic adenoma of the pituitary gland. The electron microscopic investigation concentrates upon the occurrence of osmiophil material in the different cytoplasmic compartments of the cell type rich in organelles and relatively poor in paraplasmic inclusions. It seems that the observation of osmiophil material in the perinuclear space, the mitochondrial matrix, Golgi vacuoles, and cytoplasmic vesicles has been made possible by a rather long fixation in Wohlfarth-Bottermann's chromium-osmium fixative. Mitochondria and related particles (named X-Einschlüsse) may contain a light center with a few dense filaments and masses resembling those described by other authors as being of deoxyribonucleic acid nature. Relatively large Golgi complexes are found, sometimes in close topographic relation with a centriole. A large population of osmiophil bodies (lipid droplets, pigment granules, microbodies) is described. Apart from these findings the cytoplasmic organization in the tissue studied is similar to that of the adrenal cortex as described in other mammals. The observations are discussed with respect to hypotheses about lipid droplet formation and to the possibility of artifacts due to preparation procedures.

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Herrn Prof. Dr. med. W. Bargmann zum 60. Geburtstag gewidmet. — Herrn Prof. Dr. E. Uehlinger, Direktor des Pathologischen Instituts der Universität Zürich, danken wir für die Überlassung des Materials und für die freundlicherweise zur Verfügung gestellte pathologisch-anatomische Diagnose, dem Schweizerischen Nationalfonds für die Unterstützung und Frau Jäger, Mlle. Challamel und Herrn Krieger für die Ausführung der histologischen und photographischen Arbeiten.