Localization of receptors for atrial natriuretic polypeptide (ANP) in the glomerulus: in vitro electron microscopic autoradiographical investigation using 125I-labeled ANP

The localization of receptors for atrial natriuretic polypeptide (ANP) in the glomerulus of the rat was studied by electron microscopic autoradiography using 125I-labeled ANP. A total of 1,134 silver grains counted in 30 glomeruli were distributed as follows: processes of the podocytes (36.4%), cell bodies of the podocytes (14.8%), basement membrane (12.3%), endothelial cells (6.3%), mesangial cells (7.7%) and others, for example the vascular spaces (22.5%). This finding indicates that ANP binding sites are mainly localized on the foot processes of the podocytes in the glomerulus.     

Time-resolved x-ray study of effect of sinusoidal length change on tetanized frog muscle.

Time-resolved x-ray diffraction studies were done on frog skeletal muscles with synchrotron radiation by applying sinusoidal length changes of frequency 10 Hz and amplitude approximately 1% to isometrically contracting muscles at approximately 17 degrees C. Distinct periodic intensity changes were observed in the 14.3-nm myosin meridional reflection and the equatorial 1,0 and 1,1 reflections. Response of the 14.3-nm reflection to the sinusoidal length change was nonlinear, as evidenced by a large second harmonic in its oscillatory intensity change, whereas the response of the equatorial 1,1 reflection was closely linear, as evidenced by almost sinusoidal intensity change. Intensity change of the 1,0 reflection was nearly antiphase to that of the 1,1 reflection. Integral widths of the 14.3-nm meridional reflection measured along the meridian and of the equatorial 1,1 reflection remained almost constant during tension development, while that of the 1,0 reflection tended to decrease. The widths of the 14.3-nm meridional reflection perpendicular to the meridian and of the equatorial 1,0 reflection appeared to undergo oscillatory changes in response to the sinusoidal length changes.     

Multiple Neurite Formation in Neuroblastoma Cell Lines by Griseolic Acid, a Potent Inhibitor of Cyclic Nucleotide Phosphodiesterases

The morphological change of several neuroblastoma cell lines induced by griseolic acid, a novel and potent inhibitor of cyclic nucleotide phosphodiesterase (PDE), was examined. In the cell lines tested, Neuro-2a (a murine neuroblastoma cell line) showed dose-dependent (1 microM-1 mM) neurite extension. Griseolic acid markedly increased the intracellular cyclic AMP level of Neuro-2a cells, suppressed DNA synthesis (82% at 1 mM), and induced multipolar (multiple-neurite-bearing)-type neuritogenesis. A similar type of neurite outgrowth was induced by 8-bromo-cyclic AMP, which also elevated the intracellular cyclic AMP concentration. In contrast, when Neuro-2a cells were treated with retinoic acid, neurite formation was of the monopolar (single-neurite-bearing) type. Papaverine and theophylline, which have been frequently used as PDE inhibitors, failed to induce these morphological changes up to 1 mM, probably owing to the lesser potency of these compounds as compared with griseolic acid on the inhibition of PDE. Retinoic acid, theophylline, and papaverine were ineffective at elevating the intracellular cyclic AMP level. These results suggest that multipolar-type cell shape change in Neuro-2a cells is correlated with the accumulation of intracellular cyclic AMP and that griseolic acid is a useful compound to induce neuroblastoma cells into terminal differentiation.     

Alternative and cyclic appearance of H2 and O2 photoproduction activities under non-growing conditions in an aerobic nitrogen-fixing unicellular cyanobacterium Synechococcus sp.

With synchronously grown cells of an aerobic unicellular marine cyanobacterium Synechococcus sp. Miami BG 043511, changes in the activities of anoxygenic, nitrogenase-dependent hydrogen photoproduction and oxygen photoevolution were measured under non-growing hydrogen production conditions. Interestingly, synchronously grown cells, incubated in light under an argon atmosphere, exhibited cyclic changes in the activity of nitrogenase-catalyzed hydrogen production for approximately 20- to 25-h intervals. Cyclic photosynthetic oxygen production activity also appeared in approximately the same intervals. However, changes in hydrogen production and oxygen production activities were inversely correlated and temporally separated. Stepwise accumulation of hydrogen in closed vessels was also observed in approximately 24-h cycles in these non-growing cells. These observations in non-growing cells suggest that this unicellular aerobic, nitrogen-fixing cyanobacterium may have an endogenous system to control the exhibition of cyclic rhythms, in addition to the previously studied cell cycle-oriented system. Expression and switching between these two systems may be related to the sufficiency or insufficiency of nitrogen growth nutrients. The possibility of the existence of a common control factor of the two systems involving glycogen is also discussed.     

Comparative Amperometric Study of Uptake Hydrogenase and Hydrogen Photoproduction Activities between Heterocystous Cyanobacterium Anabaena cylindrica B629 and Nonheterocystous Cyanobacterium Oscillatoria sp. Strain Miami BG7

Heterocystous filamentous cyanobacterium Anabaena cylindrica B629 and nonheterocystous filamentous cyanobacterium Oscillatoria sp. strain Miami BG7 were cultured in media with N₂ as the sole nitrogen source; and activities of oxygen-dependent hydrogen uptake, photohydrogen production, photooxygen evolution, and respiration were compared amperometrically under the same or similar experimental conditions for both strains. Distinct differences in these activities were observed in both strains. The rates of hydrogen photoproduction and hydrogen accumulation were significantly higher in Oscillatoria sp. strain BG7 than in A. cylindrica B629 at every light intensity tested. The major reason for the difference was attributable to the fact that the heterocystous cyanobacterium had a high rate of oxygen-dependent hydrogen consumption activity and the nonheterocystous cyanobacterium did not. The activity of oxygen photoevolution and respiration also contributed to the difference. Oscillatoria sp. strain BG7 had lower O₂ evolution and higher respiration than did A. cylindrica B629. Thus, the effect of O₂ on hydrogen photoproduction was minimized in Oscillatoria sp. strain BG7.     

Examination of the role of serine phosphorylation in phospholipase C-gamma and its related P47 in cAMP-mediated depression of epidermal growth factor signal transduction.

Forskolin-pretreatment of A431 cells reduced both intrinsic and epidermal growth factor (EGF)-induced EGF receptor phosphorylation, however, phosphorylation of phospholipase C-gamma (PLC-gamma) was stimulated under the same conditions. No significant difference was detected in the amount of phosphotyrosine of PLC-gamma between two cultures with or without forskolin treatment followed by EGF. On the other hand, phosphorylation of a 47 kDa protein (P47) which cross-reacted with an anti-PLC-gamma monoclonal antibody, was stimulated by both forskolin and EGF. Phosphorylation was exclusively on serine residues in this case. These results indicate that both PLC-gamma and P47 are phosphorylated by a cAMP-dependent protein kinase and the EGF-stimulated serine kinase, and suggest that serine phosphorylation of PLC-gamma has no effect on ligand-dependent coupling with the EGF receptor.     

Effects of multiple dosing of phenacetin in the micronucleus test

As a part of the international cooperative study to identify the most sensitive regimen in the micronucleus test, phenacetin was given i.p. to male CD-1 mice at doses of 37.5, 75, 150, 300, 400, and 600 mg/kg once, twice, thrice or four times and the bone marrow cells were harvested 24 h after the final dosing. Positive responses were seen at 600 mg/kg after single and triple dosing and at 400 and 600 mg/kg after double dosing. No dose level gave a positive response after quadruple dosing. A repeated-dosing effect was detected at double and triple dosing. Although triple dosing gave the highest magnitude of micronuclei at 600 mg/kg, double dosing showed a sufficient sensitivity and was more convenient from the viewpoint of selecting a suitable test dose and carrying out the micronucleus test.     

Time-resolved x-ray diffraction studies on the intensity changes of the 5.9 and 5.1 nm actin layer lines from frog skeletal muscle during an isometric tetanus using synchrotron radiation.

Time-resolved x-ray diffraction studies have been made on the 5.9- and 5.1-nm actin layer lines from frog skeletal muscles during an isometric tetanus at 6 degrees C, using synchrotron radiation. The integrated intensities of these actin layer lines were found to increase during a tetanus by 30-50% for the 5.9-nm reflection and approximately 70% for the 5.1-nm reflection of the resting values. The intensity increase of both reflections was greater than that taking place in the transition from rest to rigor state. The intensity change of the 5.9-nm reflection preceded those of the myosin 42.9-nm off-meridional reflection and of the equatorial reflections, as well as the isometric tension development. The intensity profile of the 5.9-nm layer line during contraction was found to be different from that observed in the rigor state.     

Crystal and molecular structure of RNase Rh, a new class of microbial ribonuclease from Rhizopus niveus.

The crystal structure of RNase Rh, a new class of microbial ribonuclease from Rhizopus niveus, has been determined at 2.5 A resolution by the multiple isomorphous replacement method. The crystal structure was refined by simulated annealing with molecular dynamics. The current crystallographic R-factor is 0.200 in the 10-2.5 A resolution range. The molecular structure which is completely different from the known structures of RNase A and RNase T1 consists of six alpha-helices and seven beta-strands, belonging to the alpha+beta type structure. Two histidine and one glutamic acid residues which were predicted as the most probably functional residues by chemical modification studies are found to be clustered. The steric nature of the active site taken together with the relevant site-directed mutagenesis experiments (Irie et al.) indicates that: (i) the two histidine residues are the general acid and base; and (ii) an aspartic acid residue plays a role of recognizing adenine moiety of the substrate.