Negative inotropic effect of cyclic GMP in cardiac fiber fragments

Summary The force of spontaneously beating cardiac cellular fragments obtained from mice heart by homogenization was recorded in presence of cyclic guanosine –3.5-monophosphate (cGMP) and cyclic 8-bromguanosine –3.5-monophosphate in concentrations of 3×10^–6 M –33×10^–6 M. The nucleotide decreased the force and reduced the rate of spontaneity. Eventually the preparation became quiescent. It is thought that this nucleotide either reduces the capacity to sequester calcium or affects its release from the sarcotubular system.     

Activation of respiration of liver mitochondria in various metabolic states by cyclic 3′,5′-AMP

Cyclic 3,5-AMP (10^–6M) activates respiration of the liver mitochondria in all metabolic states and neither changes nor increases the rate of phosphorylation during oxidation of saturating concentrations of isocitrate and succinate. For the effect to be manifested, preincubation of the mitochondria or liver homogenate with cyclic AMP is necessary. The fifth fraction of serum albumin and EDTA do not abolish the effect. Noradrenalin (NA) increases mitochondrial respiration only on incubation with the homogenate. Effects of NA and cyclic AMP do not undergo summation, and the effect of the former is probably mediated by cyclic AMP. The results do not confirm the decisive role of uncoupling of respiration and phosphorylation or accumulation of the oxidation substrate, but instead they suggest activation of mitochondrial enzymes.Department of Biochemistry and Central Research Laboratory, Krasnoyarsk Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR S. S. Debov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 85, No. 3, pp. 291–294, March, 1978.     

Synchronizing action of cyclic AMP on the mitotic cycle of ehrlich's ascites carcinoma cells

The number of mitoses and of DNA-synthesizing cells in an Ehrlich's ascites carcinoma was studied during the 24 h after injection of dibutyryl cyclic 3,5-adenosine monophosphate. As a result of preprophase inhibition and, probably, of stimulation of entry of the cells into the synthetic period, a large number of cells commences mitosis simultaneously 8 h after the injection. The resulting synchronization of mitosis in the cell population of the tumor is evaluated.Department of General Biology, N. I. Pirogov Second Moscow Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR V. V. Kupriyanov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 1, pp. 66–68, January, 1976.     

Effect of benzodiazepines in 5′-nucleotidase activity of rat brain

Bulletin of Experimental Biology and Medicine -     

Inhibition by somatostatin of adenosine-3′,5′-monophosphate stimulated hepatic ribosomal protein synthesis

Summary Somatostatin which inhibits the secretion of various pituitary and intestinal hormones has been suspected to exert these effects by inhibiting adenosine-3,5-monophosphate accumulation in the respective endocrine gland. Our results, obtained by cell free protein synthesis and by sedimentation through sucrose gradients of ribosomes, prepared from rat liver after incubation with cyclic AMP and/or somatostatin also suggest this antagonism between somatostatin and cyclic AMP. In addition, they indicate that this antagonism is not restricted to endocrine tissues.Supported by a grant (M2-2777) of the Österreichischer Fonds zur Förderung der wissenschaftlichen Forschung     

Renal responsiveness to parathyroid hormone in a case of nonfamilial hypophosphatemic osteomalacia

Summary Plasma immunoreactive parathyroid hormone level, urinary excretion of adenosine cyclic 3,5-monophosphate (cyclic AMP) and the sensitivity of the renal tubule to calcium infusion and to parathyroid extract were investigated in a patient with nonfamilial hypophosphatemic osteomalacia. Plasma immunoreactive parathyroid hormone concentration was normal and basal urinary excretion of cyclic AMP was increased. Renal cortical adenylate cyclase, as measured by urinary cyclic AMP excretion, was certainly as sensitive to exogenous parathyroid extract as in normal subjects. After a previous calcium infusion, a greater parathyroid-hormone-sensitive component of phosphorus transport in the kidney was present than in two control subjects. Our results indicate that in nonfamilial hypophosphatemic osteomalacia the renal tubule could be hyperresponsive to parathyroid hormone.This work was supported by a grant (n^o 20,463) from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

The isolated frog skin epithelium: Presence of α and β adrenergic receptors regulating active sodium transport and water permeability

Summary The effects of stimulation of and adrenergic receptors on short circuit current (S.C.C.), Na⁺ and Cl^– fluxes and osmotic water permeability were studied on isolated frog skin epithelial layers separated from the dermis.Low norepinephrine doses (final concentrations in the incubation medium ranging from 5×10^–9 to 10^–8 M) produced increased water permeability and S.C.C. The latter was entirely accounted for by an increase in the active Na⁺ influx. Na⁺ outflux and Cl^– fluxes were not modified. Both these effects disappeared after treatment with the blocking agent, Propranolol. Higher norepinephrine doses (final concentrations: 10^–7 to 10^–6 M) produced: 1. an increase in water permeability lower than that produced by low doses, the highest doses failing to increase water permeability, and 2. a triphasic change in S.C.C.: after an initial increase, S.C.C. dropped to its resting value and then rose again to a sustained value. Na⁺ and Cl^– flux measurements showed that the variation in S.C.C. reflected variations in active Na⁺ transport. When the same high norepinephrine doses were applied after treatment with the blocking agent Phentolamine, the effects observed were identical to those obtained with low doses.On blocked preparations, large doses of norepinephrine inhibited the water permeability and sodium transport increases induced by theophylline or oxytocin but did not modify those induced by 35-cyclic AMP. The inhibition was suppressed after blocking receptors.From the foregoing, it was concluded that both and adrenergic receptors are present in frog skin epithelial cells and are involved in the regulation of water and sodium permeability.It is suggested that the inhibitory effect of stimulation resulted from the inhibition of cyclic-AMP generating system, the activity of which is under the positive control effect of oxytocin and stimulation.     

Aspects of the oxidative metabolism of nicotine

Conclusions The balance of the excreted sum of nicotine and its known metabolites from confirmed results reflects great progress in recent years and today accounts for about 40–60% of the estimated or known intake of the alkaloid, calculating on the basis of parts of well-defined and reliably determined species. The main part of this balance is accounted for by products of the -oxidation of nicotine in the 5-position. A further 40% has recently been attributed to those parts of metabolites which are additionally found after the treatment of urine samples with enzymes generally known to act on phase II metabolites. Conjugates of nicotine, cotinine, and trans-3-hydroxycotinine are held responsible for this. Recently, some of these products have been definitely characterized and determined by direct measurements.Quantitative data are missing on the 1,2-iminium ion, the very recently identified tautomer of 2-hydroxynicotine in urine, and the nature of its prevailing tautomeric form which both are biologically high importance, have there been reports so far on the mechanism for their metabolic formation. Further, the metabolic fate of nornicotine, the first metabolite of the intermediary methylene iminium species, is still awaiting elucidation. Nornicotine is excreted in only a very small ratio. This may be caused by demythylation or by oxidation. Further conclusions require knowledge of the actual structures of the nicotine metabolites under different analytical pH values or in organic solvents. The respective proportions of the tautomeric structures in the biological media in general and especially at the active sites of enzymes determine the metabolic pathways of nicotine and its subsequent metabolites and should be the aim of intense investigations.     

Comparison of the 5′ Leader Sequences of North American Isolates of Reference and Field Strains of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV)

The 5 leader is documented to be an important regulatory element in many (+) ssRNA virus genome. To understand the significance of the 5 leader RNA of PRRSV, we determined the complete leader sequences of fifteen different North American strains of PRRSV and predicted their secondary structures. Viruses analysed included three reference strains and nine field strains originating from different geographic locations. To further examine the leader region, one of the field strains was adapted to grow in tissue culture, and three clones were isolated. We also predicted the secondary structures of two European strains based on their published sequences. The predicted RNA secondary structures of the leader sequences suggested the existence of three conserved domains formed by the 5 region of the leader among the North American strains, two of which were conserved in the European strains. A variable structural domain was predicted from the 3 region of the leader sequences of the North American strains, where all tissue culture-adapted isolates were characterized by a stem-loop while field isolates were characterized by an internal bulge within the stem-loop.     

Synergistic effects of tumour necrosis factor-α and interferon-γ on feline haemopoietic progenitors

Pathogenic mechanisms that underlie feline leukaemia virus subgroup-C (FeLV-C) induced erythroid aplasia are unknown. FeLV-C infection is associated with higher serum levels of interferon- (IFN-) and tumour necrosis factor- (TNF-), which may act synergistically to cause haemopoietic suppression. In the present studies, the synergistic effects of TNF- and IFN- on feline bone marrow progenitors in vitro were evaluated. Bone marrow mononuclear cells from specific-pathogen-free cats were exposed to TNF- (100 and 200 pg/ml) and IFN- (100 or 200 units/ml), alone or in combination, for 2 h before plating for clonal assays of colony forming units. Our results show that TNF- and IFN- in combination caused marked suppression of feline colony forming units-erythroid (CFU-E), burst forming units-erythroid (BFU-E), and colony forming units-fibroblasts (CFU-F), whereas colony forming units-granulocyte/macrophage (CFU-GM) were minimally affected. The same concentrations of TNF- and IFN- alone had minimal effects on CFU-E, BFU-E and CFU-F. These results suggest that TNF- and IFN- may play a significant role in regulating haemopoiesis in cats and may be involved in the pathogenesis of erythroid aplasia in cats infected with feline leukaemia virus.     

Effect of imidazole compounds on 3′,5′-AMP phosphodiestyerase activity

The effect of 24 imidazole compounds on activity on the phosphodiesterase (3, 5-AMP phosphohydrolase; EC 3.1.4.1) from rat brain and skeletal muscles ofRana temporaria was investigated. Imidazole compounds were shown to have both an activating and an inhibitory action on the enzyme. Imidazole itself and seven of its alkyl substitution products activated phosphodiesterase. Of the inhibitors, tetrachloro-2-trifluoromethyl-benzimidazole had the strongest action on the enzyme.Group for Biophysics of Synaptic Processes, I. M. Sechenov Institute of Evolutionary Physiology and Biochemistry, Academy of Sciences of the USSR, Leningrad. (Presented by Academician S. E. Severin.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 9, pp. 1055–1059, September, 1976.     

Cyclische Nucleotide als intracelluläre Überträgerstoffe bei Hormonwirkungen

Zusammenfassung Hormone dienen als extracelluläre Informationsüberträger zwischen ihrem Bildungsort, einer endokrinen Drüse, und den Zellen, deren Funktion sie regulieren. Durch die Reaktion des Hormons mit den an der Zellmembran gelegenen Receptoren wird die Aktivität der mit diesen eng verknüpften Adenyl-Cyclase beeinflußt. Die meisten Hormone erhöhen in ihrem Zielorgan die Aktivität dieses Enzyms und führen hierdurch zu einem raschen Anstieg der intracellulären Konzentration von Adenosin-3:5-monophosphat (Ado-3:5-P). Dieses cyclische Nucleotid wird durch eine spezifische Phosphodiesterase zu Adenosin-5-monophosphat abgebaut. Auch die Aktivität dieses Enzyms bestimmt die intracelluläre Ado-3:5-P-Konzentration, die im Vergleich zu der anderer Nucleotide sehr gering ist.Ado-3:5-P beeinflußt als zweiter, intracellulärer Überträgerstoff die Aktivität zahlreicher Schlüsselenzyme. Die Ado-3:5-P-Konzentration bestimmt hierdurch das Gleichgewicht verschiedener Stoffwechselwege zueinander und damit die Reaktion einer Zelle auf eine hormonale Stimulierung. An einer Reihe von Enzymen wird die durch Ado-3:5-P bedingte Aktivitäts-Änderung durch einen gleichartigen Mechanismus bewirkt. Das cyclische Nucleotid stimuliert Proteinkinasen, die eine Phosphatgruppe des ATP auf verschiedene Proteine übertragen und hierdurch deren Eigenschaften verändern können. So steigt bei Phosphorylierung durch eine Ado-3:5-P-stimulierbare Proteinkinase die Aktivität der Triglyceridlipase und der Glykogen-Phosphorylase-b-kinase an, dagegen nimmt die Aktivität der Glykogen-Synthetase ab; durch Phosphorylierung von Histonen kann deren Repressorcigenschaft vermindert und die Synthese von Enzymen gesteigert werden.In manchen tierischen Geweben wurde auch eine spezifisch durch Guanosin-3:5-monophosphat (Guo-3:5-P) stimulierbare Proteinkinase nachgewiesen. Dieses cyclische Nucleotid kommt wie Ado-3:5-P in allen Säugerorganen vor. Die Bildung von Guo-3:5-P aus GTP wird durch die Guanyl-Cyclase katalysiert, ein Ferment, das im Gegensatz zur Adenyl-Cyclase zum großen Teil nicht an die Zellmembranen gebunden ist. Die Konzentration von Guo-3:5-P in verschiedenen Geweben, im Blutplasma und im Urin wird durch Hormone beeinflußt. Es ist noch nicht bekannt, welche hormonalen Regulationen durch Guo-3:5-P vermittelt werden; dagegen ist bei vielen, rasch einsetzenden Hormonwirkungen die Beteiligung von Ado-3:5-P nachgewiesen worden.

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Abkürzungen Ado-3:5-P Adenosin-3:5-monophosphat - dAdo-3:5-P Desoxy-adenosin-3:5-monophosphat - Guo-3:5-P Guanosin-3 : 5-monophosphate - Nuc-3:5-P Nucleosid-3:5-monophosphat - NTP Nucleosidtriphosphat - NMP Nuclcosid-5-monophosphat - dATP Desoxyadenosintriphosphat - P_i anorganisches Phosphat - PP_i anorganisches Pyrophosphat - DNS Desoxyribonucleinsäure - RNS Ribonucleinsäure - r-RNS ribosomale RNS - m-RNS Boten-RNS - Glykogen-Synthetase UDP-Glucose--1,4-glucan--4-glucosyltransferase - ICSH interstitial-cell-stimulating hormone     

Effect of the dephosphorylated core of 2′,5′-oligoadenylate (2′,5′-ApApA) on electrical activity of the snail neuron

Bulletin of Experimental Biology and Medicine -     

Control of metastatic properties of BL6 melanoma cells by H-2Kb gene: immunological and nonimmunological mechanisms

The effect of class I H-2 antigen expression on the metastatic properties of BL6 melanoma cells was investigated. The BL6-8 clone isolated from the highly metastatic BL6 melanoma did not express H-2K b gene. Following transfection with the H-2K^b gene, BL6-8 cells displayed a low metastatic potential in the immunocompetent as well as immunosuppressed (X-irradiated) or triple-immunodeficient mice with impaired T, B and natural killer (NK) cells function. The expression of H-2K^b gene and the low metastatic ability of transfected BL6 melanoma cells were associated with appearance of cell membrane soybean agglutinin (SBA) and Griffonia simplicifolia 1B₄ (GS1B₄) lectin-binding carbohydrataes. These alterations in cell surface carbohydrates were found to be a result of reduction in sialylation of SBA binding sites and upregulation of the 1.3 galactosyltransferase (1.3GT) gene. To assess the importance of H-2K^b-induced alterations in cell surface carbohydrates for metastasis formation, BL6-8 melanoma cells were transfected with H-2K^b gene without neo^r gene cotransfection and selected for adherence to SBA-lectin-conjugated agarose beads. The transfected clones that expressed SBA and GS1B₄ lectin-binding carbohydrates were low metastatic. Further analysis of these clones showed that presence of SBA and GS1B₄ lectin-binding carbohydrates rather than expression of H-2K^b molecules per se might be responsible for low metastatic potentials of H-2K^b-transfected cells in the immunocompromized mice. Studies of the possible mechanisms responsible for low metastatic ability of H-2K^b-transfected melanoma cells revealed that these cells displayed a reduced ability to adhere to murine pulmonary endothelial cells as well as to laminin and collagen IV. We hypothesized that the observed nonimmunological effects of H-2K^b gene in BL6 melanoma cells is a result of an interaction between the H-2K^b gene and B16 melanoma-specific ecotropic retrovirus. It results in inhibition of this retrovirus production with consecutive alteration in the expression of cellular genes controlling cell surface glycosylation and adhesion properties essential for the metastatic phenotype of BL6 melanoma.     

Activity of enzymes of adenosine metabolism in human natural killer (NK) cells during activation and inhibition of their cytotoxic activity

R. E. Kavetskii Institute for Problems in Oncology, Academy of Sciences of the Ukrainian SSSR, Kiev. Research Institute of Carcinogenesis, All-Union Oncologic Scientific Center, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR N. N. Trapeznikov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 107, No. 4, pp. 438–440, April, 1989.     

Nitric oxide mediated modulation of free radical generation response in the rat polymorphonuclear leukocytes: A flowcytometric study

Nitric oxide (NO) synthesis and free radical generation from polymorphonuclear leukocytes (PMNs) play an important role in several pathological conditions. It is therefore important to understand the regulatory mechanisms of free radical generation from PMNs. Flowcytometry can be used to assess generation of reactive oxygen and nitrogen species from PMNs by using fluorescent probes. In the present study regulation of NO synthesis in the control and lipopolysaccharide (LPS) treated rat PMNs has been investigated. Free radical generation was assessed by flow cytometry using a dye, 27-dichlorodihydrofluorescein diacetate (DCFDA), dihydrorhodamine-123 (DHR) and 4,5-diaminofluorescein diacetate (DAF). Superoxide dismutase (SOD), and catalase significantly attenuated the arachidonic acid (AA, 1 × 10^–6 M) induced free radical generation, while 4-aminobenzoicacid hydrazide (ABH), myeloperoxidase (MPO) inhibitor had no significant effect. Intracellular and extracellular calcium levels also modulated FR generation. AA induced free radical generation from PMNs was also enhanced significantly after LPS treatment. NO synthase (NOS) inhibitors, aminoguanidine (AG) and 7-nitroindazole (NI) inhibited arachidonic acid induced free radical generation from LPS treated PMNs, while in control PMNs NOS inhibition had no effect. Augmentation of free radical generation from rat PMNs following LPS treatment seems to be regulated by NO.