Demethylation of the Protein Phosphatase PP2A Promotes Demethylation of Histones to Enable Their Function as a Methyl Group Sink

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Isolation and patch clamp measurements of xylem contact cells for the study of their role in the exchange between apoplast and symplast of leaves

Because of their importance for nutrition, a method was developed to patch xylem contact cells in leaves of Vicia faba and maize. Since the lignification of older cells was a major obstacle for isolating protoplasts which could be patched, only young leaves (fourth fully developed leaf) were used. An important step in the isolation of these cells was the infiltration of the leaves and their exposure to enzymes for several hours, allowing mesophyll cells to be removed whilst having most of the xylem contact cells attached to the xylem. Channel activity in cell-attached mode or in excised patches could only be observed if an internal coating of sigmacote was used to block diffusion ions out of the pipette glass. Two different types of K+ channels were identified by measuring the reversal potential at different concentrations of KCl. One channel (SC) had a symmetric IV curve with a high probability of remaining open, irrespective of membrane potential; the other channel was an inward rectifier. The symmetical channel could be blocked weakly by Na+ but it was permeable to .     

Characterization of entomopox virions of the army cutworm,Euxoa auxiliaris (Lepidoptera: Noctuidae)

Virions were released from virus-containing inclusions (VCI) of an entomopoxvirus of the army cutworm, Euxoa auxiliaris, with carbonate-thioglycolate solution. Knoblike projections present on the surface of the viral envelope were removed by digestion with trypsin. Trypsin-treated virions were homogeneous in both sucrose and CsCl gradients. The virions were similar to vertebrate poxviruses in morphology, contained 1.13 ± 0.3% DNA and had a buoyant density of 1.261 ± 0.003 gm/cm³ in CsCl. The virion preparations were infective and possessed RNA polymerase activity. Of eight species of Lepidoptera tested, only the species from which the virus was originally isolated proved susceptible to infection.     

Position-independent expression of a human nerve growth factor-luciferase reporter gene cloned on a yeast artificial chromosome vector.

Two yeast artificial chromosomes containing the entire human nerve growth factor gene were isolated and mapped. By homologous recombination a luciferase gene was precisely engineered into the coding portion of the NGF gene and a neomycin selection marker was placed adjacent to one of the YAC telomeres. Expression of the YAC-based NGF reporter gene and a plasmid-based NGF reporter gene were compared with the regulation of endogenous mouse NGF protein in mouse L929 fibroblasts. In contrast to the plasmid-based reporter gene, expression and regulation of the YAC-based reporter gene was independent of the site of integration of the transgene. Basic fibroblast growth factor and okadaic acid stimulated expression of the YAC transgene, whereas transforming growth factor-beta and dexamethasone inhibited it. Although cyclic AMP strongly stimulated production of the endogenous mouse NGF, no effect was seen on the human NGF reporter genes. Downregulation of the secretion of endogenous mouse NGF already occurred at an EC50 of 1-2 nM dexamethasone, but downregulation of the expression of NGF reporter genes occurred only at EC50 of 10 nM. This higher concentration was also required for upregulation of luciferase genes driven by the dexamethasone-inducible promoter of the mouse mammary tumor virus in L929 fibroblasts.     

Mechanisms of the antinociceptive action of (?) Epicatechin obtained from the hydroalcoholic fraction of Combretum leprosum Mart & Eic in rodents

ABSTRACT: BACKGROUND: The mechanisms of the antinociceptive activity of () epicatechin (EPI), a compound isolated from the hydroalcoholic fraction of Combreum leprosum Mart & Eicher. METHODS: were assessed in the model of chemical nociception induced by glutamate (20 mumol/paw). To evaluate the mechanisms involved, the animals , male Swiss mice (25-30 g), received EPI (50 mg/kg p.o.) after pretreatment with naloxone (2 mg/kg s.c. opioid antagonist), glibenclamide (2 mg/kg s.c. antagonist K + channels sensitive to ATP), ketanserin (0.3 mg/kg s.c. antagonist of receptor 5-HT2A), yoimbine (0.15 mg/kg s.c. alpha2 adrenergic receptor antagonist), pindolol (1 mg/kg s.c. 5-HT1a/1b receptor antagonist), atropine (0.1 mg/kg s.c. muscarinic antagonist) and caffeine (3 mg/kg s.c. adenosine receptor antagonist), ondansetron (0.5 mg/kg s.c. for 5-HT3 receptor) and L-arginine (600 mg/kg i.p.). RESULTS: The antinociceptive effect of EPI was reversed by pretreatment with naloxone and glibenclamide, ketanserin, yoimbine, atropine and pindolol, which demonstrates the involvement of opioid receptors and potassium channels sensitive to ATP, the serotoninergic (receptor 5HT1A and 5HT2A), adrenergic (receptor alpha 2) and cholinergic (muscarinic receptor) systems in the activities that were observed. The effects of EPI, however, were not reversed by pretreatment with caffeine, L-arginine or ondansetron, which shows that there is no involvement of 5HT3 receptors or the purinergic and nitrergic systems in the antinociceptive effect of EPI. In the Open Field and Rotarod test, EPI had no significant effect, which shows that there was no central nervous system depressant or muscle relaxant effect on the results. CONCLUSIONS: This study demonstrates that the antinociceptive activity of EPI in the glutamate model involves the participation of the opioid system, serotonin, adrenergic and cholinergic.