Attenuation of ethanol intoxication by alpha-2 adrenoceptor antagonists

The interaction of a highly potent and selective alpha-2 adrenoceptor antagonist, atipamezole with ethanol was investigated in tests assessing a number of ethanol's behavioral effects. Atipamezole antagonized ethanol's effects on directed exploration in a holeboard test, reduced observer-rated intoxication and also reduced the duration of loss of righting reflex caused by ethanol. Similar effects were produced by another alpha-2 adrenoceptor antagonist idazoxan. The magnitude of the effects was comparable to that produced in the same animal models by the imidazodiazepine Ro 15-4513, which antagonizes ethanol by an action at central benzodiazepine receptors. Whereas Ro 15-4513 possesses marked behavioral effects on its own, atipamezole is comparatively inactive in all paradigms so far tested. The data suggest that alpha-2 adrenoceptors can play an important role in modulating the intoxicating effects of ethanol.     

Detection of spread of malignant lymphoma to the liver by low field strength magnetic resonance imaging.

The accuracy of spin lattice relaxation time (T1) measurement obtained with a low field strength magnetic resonance imager for the detection of spread of malignant lymphoma to the liver was assessed. The results of histological examination obtained at open liver biopsy were compared with liver T1 values in 27 patients with lymphoma. The normal range for T1 was established by scanning 61 healthy volunteers. Magnetic resonance imaging was highly sensitive in detecting hepatic lymphoma, all seven patients with liver lymphoma proved by biopsy having considerably higher T1 values. Specificity was less good. Five out of 20 patients with no histological evidence of hepatic lymphoma had abnormal T1 values. this level of sensitivity is considerably better than that reported for other imaging methods and contrasts with the results of one previous study using a different magnetic resonance system. Low field strength magnetic resonance imaging may prove to be a useful screening test in patients with lymphoma. The presence of a normal liver T1 seems to be a reliable guide to the absence of hepatic disease.     

Short-term pigment changes in Norway spruce needles

The photosynthetic pigment contents and the chlorophyll fluorescence induction in Norway spruce [ Picea abies (L.) Karst.] needles were monitored at 3 h intervals over a 10 day period. This covered 6 days of high insolation, one day of low insolation, followed by three more days of high insolation. During the first six days only small changes in mean pigment contents were observed but we interpret them as suggesting a daily rhythm in total chlorophyll content with two maxima and minima. The day of low insolation was paralleled by larger pigment changes, mostly an increase in chlorophyll b along with a decrease in carotenoid content. With the resumption of high insolation, pigments gradually return to their previous levels. During the entire period chlorophyll a fluorescence induction remained relatively constant. These short-term responses may reflect features of thylakoid organization in relation to pigment content and suggest possible long-term adaptive mechanisms in non deciduous conifers.     

Solubilization, purification, and characterization of a membrane-bound phospholipase A2 from the P388D1 macrophage-like cell line

The release of free arachidonic acid from membrane phospholipids is believed to be the rate-controlling step in the production of the prostaglandins, leukotrienes, and related metabolites in inflammatory cells such as the macrophage. We have previously identified several different phospholipases in the macrophage-like cell line P388D1 potentially capable of controlling arachidonic acid release. Among them, a membrane-bound, alkaline pH optimum, Ca2+-dependent phospholipase A2 is of particular interest because of the likelihood that the regulatory enzyme has these properties. This phospholipase A2 has now been solubilized from the membrane fraction with octyl glucoside and partially purified. The first two steps in this purification are butanol extractions that yield a lyophilized, stable preparation of phospholipase A2 lacking other phospholipase activities. This phospholipase A2 shows considerably more activity when assayed in the presence of glycerol, regardless of whether the substrate, dipalmitoylphosphatidylcholine, is in the form of sonicated vesicles or mixed micelles with the nonionic surfactant Triton X-100. Glycerol (70%) increases both the Vmax and the Km with both substrate forms, giving a Vmax of about 15 nmol min-1 mg-1 and an apparent Km of about 60 microM for vesicles and a Vmax of about 100 nmol min-1 mg-1 and an apparent Km of about 1 mM for mixed micelles. Vmax/Km is slightly greater for vesicles than for mixed micelles. The lyophilized preparation of the enzyme is routinely purified about 60-fold and is suitable for evaluating phospholipase A2 inhibitors such as manoalide analogues. Subsequent steps in the purification are acetonitrile extraction followed by high performance liquid chromatography on an Aquapore BU-300 column and a Superose 12 column. This yields a 2500-fold purification of the membrane-bound phospholipase A2 with a 25% recovery and a specific activity of about 800 nmol min-1 mg-1 toward 100 microM dipalmitoylphosphatidylcholine in mixed micelles. When this material was subjected to analysis on a Superose 12 sizing column, the molecular mass of the active fraction was approximately 18,000 daltons.     

Molecular Analysis of a Transposon-Induced Deletion of the Nivea Locus in Antirrhinum Majus

The transposable element Tam3 of Antirrhinum majus is capable of causing large-scale chromosomal restructuring. It induced a large deletion at the nivea locus, to produce the allele niv-:529. The deletion removed the entire nivea coding region while the element remains intact with the potential to induce further rearrangements. Genetic experiments showed that the endpoint of the deletion (called x) is closely linked to nivea. The DNA sequences of niv-:529, a genomic excision of Tam3 from niv-:529, and the original genomic position of x have been determined. These data suggest that the deletion could have resulted from an abortive transposition or through breakage and religation.     

Transgene excision in zebrafish using the phiC31 integrase

Genome engineering strategies employing site‐specific recombinases (SSRs) have become invaluable to the study of gene function in model organisms. One such SSR, the integrase encoded by the Streptomyces bacteriophage phiC31, promotes recombination between heterotypic attP and attB sites. In the present study I have examined the feasibility of the use of phiC31 integrase for intramolecular recombination strategies in zebrafish embryos. I report here that (1) phiC31 integrase is functional in zebrafish cells, (2) phiC31 integrase can excise a transgene cassette flanked by an attB and an attP site, analogous to a common use of the Cre/lox SSR system, (3) phiC31 integrase functions in the zebrafish germline, and (4) a phiC31 integrase‐estrogen receptor hormone‐binding domain variant fusion protein catalyzes attB‐attP recombination in zebrafish embryos in a 4‐hydroxytamoxifen‐dependent manner, albeit less efficiently than phiC31 alone. These features should make this a useful approach for genome manipulations in the zebrafish. genesis 48:137–143, 2010. © 2010 Wiley‐Liss, Inc.     

Resetting and regulation of FLOWERING LOCUS C expression during Arabidopsis reproductive development

The epigenetic regulation of the floral repressor FLOWERING LOCUS C ( FLC ) is one of the critical factors that determine flowering time in Arabidopsis thaliana . Although many FLC regulators, and their effects on FLC chromatin, have been extensively studied, the epigenetic resetting of FLC has not yet been thoroughly characterized. Here, we investigate the FLC expression during gametogenesis and embryogenesis using FLC::GUS transgenic plants and RNA analysis. Regardless of the epigenetic state in adult plants, FLC expression disappeared in gametophytes. Subsequently, FLC expression was reactivated after fertilization in embryos, but not in the endosperm. Both parental alleles contributed equally to the expression of FLC in embryos. Surprisingly, the reactivation of FLC in early embryos was independent of FRIGIDA (FRI) and SUPPRESSOR OF FRIGIDA 4 (SUF4) activities. Instead, FRI , SUF4 and autonomous-pathway genes determined the level of FLC expression only in late embryogenesis. Many FLC regulators exhibited expression patterns similar to that of FLC , indicating potential roles in FLC reprogramming. An FVE mutation caused ectopic expression of FLC in the endosperm. A mutation in PHOTOPERIOD-INDEPENDENT EARLY FLOWERING 1 caused defects in FLC reactivation in early embryogenesis, and maintenance of full FLC expression in late embryogenesis. We also show that the polycomb group complex components, Fertilization-Independent endosperm and MEDEA, which mediate epigenetic regulation in seeds, are not relevant for FLC reprogramming. Based on our results, we propose that FLC reprogramming is composed of three phases: (i) repression in gametogenesis, (ii) reactivation in early embryogenesis and (iii) maintenance in late embryogenesis.