Rapid Activation of Tyrosine Hydroxylase in Response to Nerve Growth Factor

Abstract: Nerve growth factor protein (NGF) was found to rapidly promote the activation of tyrosine hydroxylase in cultured rat PC 12 pheochromocytoma cells. PC 12 cultures were exposed to NGF for periods of less than 1 h and the soluble contents of homogenates prepared from the cells were assayed for tyrosine hydroxylase activity. Under these conditions, the specific enzymatic activity was increased by 60 ± 10% (n = 13) in comparison with that in untreated sister cultures. The increase was half maximal by 2–5 min of exposure and at NGF concentrations of about 10 ng/ml (0.36 n M ). Antiserum against NGF blocked the effect. Tyrosine hydroxylase activity could also be rapidly increased by NGF in cultures of PC12 cells that had been treated with the factor for several weeks in order to produce a neuron-like phenotype. This was achieved by withdrawing NGF for about 4 h and then readding it for 30 min. The NGF-induced increase of tyrosine hydroxylase activity in PC12 cultures was not affected by inhibition of protein synthesis and therefore appeared to be due to activation of the enzyme. Kinetic experiments revealed that NGF brought about no change in the apparent K_m of the enzyme for tyrosine or for co-factor (6-methyltetrahydropteridine), but that it did significantly increase the apparent maximum specific activity of the enzyme. These observations suggest that NGF (perhaps released by target organs) could promote a rapid and local enhancement of noradrenergic transmission in the sympathetic nervous system.     

Defective mononuclear phagocyte function in systemic lupus erythematosus: dissociation of Fc receptor-ligand binding and internalization

Fc receptor-mediated mononuclear phagocyte system (MPS) clearance is impaired in systemic lupus erythematosus (SLE) and may contribute to the pathogenesis of the immune complex disease. To investigate the basis of MPS dysfunction, we have examined concurrent in vivo and in vitro Fc receptor function in 22 patients with SLE and 23 disease-free adults. Blood monocyte Fc receptor binding was increased rather than decreased as predicted by the saturation hypothesis of MPS blockade. Rosette formation of IgG-sensitized bovine erythrocytes (EA) with monocytes demonstrated increased Fc receptor-ligand binding in SLE (percent rosettes: 40 +/- 12 vs 27 +/- 8, p less than 0.001). Scatchard analysis of the binding of radiolabeled IgG oligomers to SLE monocytes indicated a mean receptor number 30% higher than control, although this did not reach statistical significance. Despite enhanced Fc receptor-ligand (EA) binding, Fc-mediated phagocytosis of EA was decreased in SLE (1.7 +/- 0.7 erythrocytes/monocytes/hour vs 2.6 +/- 1.0, p less than 0.004). This decrease in phagocytosis by blood monocytes from SLE patients was significantly greater than that attributable to the predominance in SLE of individuals with certain HLA B cell alloantigens and intrinsically lower phagocytic rates (p less than 0.05 for all groups). This decrease therefore represents a disease-acquired characteristic. Furthermore, the phagocytic rate of the four SLE patients with marked prolongation in MPS clearance was significantly lower than that of the eight patients with near normal clearance values (p less than 0.01). Saturation of Fc receptors by immune complexes does not explain impaired immune clearance in SLE. Our results indicate that despite increased binding of the EA ligand, Fc receptor-mediated phagocytosis is markedly impaired in SLE monocytes. This impairment cannot be explained on the basis of HLA-related differences in phagocytosis among lupus patients. The defect in phagocytosis of EA is most profound in those patients with the most significantly impaired MPS clearance. Thus, the dissociation of receptor-ligand binding and receptor-mediated internalization may contribute significantly to the in vivo clearance defect in SLE.     

A cortical granule-specific enzyme,B-1,3-glucanase,in sea urchin eggs

The ultrastructural localization of B-1,3-glucanase in three species of sea urchin eggs was determined using a monospecific antibody in an electronmicroscopic immunogold procedure. In all three species, Lytechinus variegatus, Strongylocentrotus purpuratus, and Arbacia punctulata, B-1,3-glucanase was localized specifically to the cortical granules. No other organelle within the egg contained significant label. During the fertilization reaction, B-1,3-glucanase was released from cortical granules into the perivitelline space and became associated with the hyaline layer. No significant label was found in association with the fertilization envelope.     

Bio-synthesis of PGD2 and TXB2 by rabbit blood monocytes

A method for the preparation of a highly purified sample of rabbit blood monocytes is described. The metabolism of arachidonic acid (AA) in these cells was studied. Mononuclear cells were prepared by centrifugation on Ficoll-Paque gradients and the monocytes were obtained by further centrifugation and adherence onto plastic culture dishes. These procedures provided a preparation which contained 95% monocytes (non-specific esterase positive). Incubation of [1-14C]-AA with these cells produced four major metabolites which were separated by TLC; these corresponded to prostaglandin (PG) D2, thromboxane (TX) B2, 12-hydroxyheptadecatrienoic acid (HHT) and 12-/15-hydroxyeicosatetraenoic acid (HETE). A minor product which co-migrated with PGE2 was also detected but neither 6-keto-PGF1 alpha nor PGF2 alpha were detected. Also, there was no evidence of the formation of 5-lipoxygenase products (5-HETE and LTB4) by rabbit monocytes with or without calcium-ionophore A23187-stimulation. The production of PGD2, TXB2 and PGE2 was further confirmed by analyzing [3H]-AA metabolites using high-performance liquid chromatography (HPLC) with tritiated standards as references. The biosynthesis of these compounds from endogenous substrate in A23187-stimulated monocytes was confirmed by specific radioimmunoassays with or without prior HPLC separation. The synthesis of immunoreactive LTB4 and LTC4 by A23187-stimulated cells was also monitored and found to be relatively low. The synthesis of PGD2, TXB2 and PGE2 from both exogenous and endogenous substrate was suppressed by treatment of the monocytes with indomethacin (10(-6) M).     

Selenium modifies carcinogen metabolism by inhibiting enzyme induction

Since selenium has been found to exert a protective action against carcinogenesis in various systems, the mechanism where-by sodium selenite inhibits DNA binding of the carcinogen, 7,12-dimethylbenz[a]anthracene, was investigated. It was found that selenite preferentially reduced DNA binding occurring through ananti-dihydrodiol epoxide metabolite of this carcinogen by inhibiting the induction of an enzyme system that generates this specific reactive metabolite.     

Social stimulation and the resumption of copulation in Rhesus (Macaca mulatto) and stumptail (Macaca arctoides) Macaques

Copulatory data derived from observations of social groups of rhesus and stumptail macaques were analyzed to test the hypothesis that pairs of animals would resume copulation significantly sooner if a second male copulated with the female shortly after the first male’s ejaculation. Data from both groups supported the hypothesis. These results, extending previous studies in Macaca nemestrina,suggest that the shortening of copulatory intervals by social stimuli occurs in several species, both in social groups and in experimentally created triads. These findings also are consistent with the hypothesis that socially mediated resumption of mating is related to intrasexual competition among males.     

An ultrastructural study of embryonic chick retinal neurons in culture

Summary The differentiation of cells and synapses in explants of 9-day-old chick embryo retina has been studied by light and electron microscopy over a period of 35 days in vitro, and samples of retina from the 9-day chick foetus were directly fixed and prepared for study.At the time of explantation the retinae were poorly differentiated and no lamination was apparent. From day 14 onwards, (i) outer and inner nuclear layers (ONL, INL) separated by a layer of neuropil corresponding to the outer plexiform layer (OPL) and (ii) a layer of scattered large ganglion cells separated from the INL by a zone of neuropil resembling the inner plexiform layer (IPL) were apparent, and (iii) a well-differentiated outer limiting membrane was established close to the surface of the explants. In the oldest cultures some development of photoreceptor outer segments occurred but a distinct optic nerve fibre layer did not form.Although cell identification presented problems even in the oldest cultures, the major retinal cell types described in vivo could be identified. Photoreceptor cells developed pedicles in the OPL which became filled with synaptic vesicles and synaptic ribbons and established ribbon synapses (including triads) with and were commonly invaginated by processes from horizontal and bipolar cells. Processes of bipolar cells in the IPL formed simple and dyad synapses. At least two types of presynaptic amacrine cells were also identified in the INL, one of which contained large numbers of dense-core vesicles. The ganglion cells, though sparse, were large and well differentiated.These findings show that all the major neuronal types of the retina are capable of developing and differentiating in vitro, lagging behind the time-table of development and differentiation in vivo by approximately 7 days, but resulting in a histotypically organised retina with synaptic neuropil showing many similarities to the corresponding neuropil in vivo.     

Host receptor sites involved in the attachment of Bdellovibrio bacteriovorus and Bdellovibrio stolpii

Abstract The location of lipoteichoic acid and 3 cell wall-associated protein antigens in fresh isolates of Streptococcus mutans (serotype c ) and in variants derived from these parental strains by repeated subculture in vitro has been examined by electrophoretic and dot-immunobinding techniques. Following subculture there was a marked shift in the distribution of all four antigens from the cell wall to the extracellular culture supernatants. It is therefore apparent that that the decreased hydrophobicity and the impaired ability of the subcultured variants to colonise in vivo, as observed by others, cannot be correlated with changes in a single surface molecule.     

Relationships among New World species of Hibiscus section furcaria (Malvaceae)

Seventeen species ofHibiscus sectionFurcaria are native to the New World, of which 12 (one diploid, nine tetraploids, one octoploid and one decaploid) have been studied cytotaxonomically. New chromosome counts (2n=4x=72) are reported forH. cucurbitaceus, H. flagelliformis, H. kitaibelifolius, andH. laxiflorus. Seventeen types of tetraploid interspecific hybrids (seven new to this study) all showed complete meiotic chromosome homology (genome formulaGGPP) and normal floral development. That all hybrids were nevertheless almost completely sterile suggests a cytoplasmic component to the genetic differentiation of the species. The diversification of the tetraploid species in habits, habitats and geographical ranges is considerable, despite their similar genome constitutions. A key to 17 native and four introduced African species is presented.Hibiscus cerradoensis sp. nov. is described.