Independent activation of subfornical organ and hypothalamo-neurohypophysial system during administration of angiotensin II

The autoradiographic deoxyglucose method was employed to investigate: 1) whether the increased glucose utilization in the subfornical organ (SFO) during administration of angiotensin II (AII) depends on the neural inputs to the SFO; and 2) to investigate whether the activation of the hypothalamo-neurohypophysial system during administration of AII depends on inputs from the SFO. The ventral stalk of the SFO, which contains the majority of efferent and afferent projections of this circumventricular structure, was interrupted with knife cuts three days before the deoxyglucose experiments. Intravenous infusion of AII (2.5 micrograms/min) for 45 min increased glucose utilization in the SFO and neural lobe in the lesioned animals to the same extent as in the sham-operated animals. Drinking, however, was significantly reduced in lesioned animals. These experiments disclose independent parallel mechanisms responsible for activation of the SFO and the hypothalamo-neurohypophysial system by AII.     

Purification and partial characterization of 1-aminocyclopropane-1-carboxylate synthase from tomato pericarp

1-Aminocyclopropane-1-carboxylate synthase was purified 5000-fold from LiCl-induced tomato fruit slices by conventional and high-performance liquid chromatography. The final preparation was estimated to be between 25% and 50% pure. Two-dimensional gel electrophoresis indicates that 1-aminocyclopropane-1-carboxylate synthase activity is associated with a 45-kDa polypeptide, with a pI of 5.8 +/- 0.2. The enzyme is inactivated both by its substrate, S-adenosyl-L-methionine (AdoMet) and by one of its products, 1-aminocyclopropane-1-carboxylate. Due to the extremely low abundance of the protein it was necessary to scale up the extraction in order to obtain reasonable amounts for sequence analysis. Therefore, 200 kg tomatoes were extracted on semi-industrial scale and 1-aminocyclopropane-1-carboxylate synthase purified. This yielded approximately 150 micrograms enzyme.     

Abundant alkali-sensitive sites in DNA of human and mouse sperm

The DNA of human and mouse sperm cells was analyzed by single-cell microgel electrophoresis, by agarose gel electrophoresis, and by alkaline elution--three techniques that can detect single-strand DNA breaks and/or labile sites. Under these conditions a surprisingly large number of single-strand DNA breaks, approximately 10(6) to 10(7) per genome, were detected in human and mouse sperm but not in human lymphocytes or in mouse bone marrow cells. These breaks were also present in chicken erythrocyte DNA, which is also highly condensed. These breaks were not observed under neutral pH conditions nor under denaturing conditions not involving alkali, suggesting that these sites are alkali-sensitive and do not represent preexisting single-strand breaks. The high frequency of such sites in sperm from healthy mouse and human donors suggests that they represent a functional characteristic of condensed chromatin rather than DNA damage.     

Genetic and Molecular Characterization of Suppressors of Sir4 Mutations in Saccharomyces Cerevisiae

In order to learn more about other proteins that may be involved in repression of HML and HMR in Saccharomyces cerevisiae, extragenic suppressor mutations were identified that could restore repression in cells defective in SIR4, a gene required for function of the silencer elements flanking HML and HMR. These suppressor mutations, which define at least three new genes, SAN1, SAN2 and SAN3, arose at the frequency expected for loss-of-function mutations following mutagenesis. All san mutations were recessive. Suppression by san1 was allele-nonspecific, since san1 could suppress two very different alleles of SIR4, and was locus-specific since san1 was unable to suppress a SIR3 mutation or a variety of mutations conferring auxotrophies. The SAN1 gene was cloned, sequenced, and used to construct a null allele. The null allele had the same phenotype as the EMS-induced mutations and exhibited no pleiotropies of its own. Thus, the SAN1 gene was not essential. SAN1-mediated suppression was neither due to compensatory mutations in interacting proteins, nor to translational missense suppression. SAN1 may act posttranslationally to control the stability or activity of the SIR4 protein.     

Comparison of polymerase insertion and extension kinetics of a series of O2-alkyldeoxythymidine triphosphates and O4-methyldeoxythymidine triphosphate

The effect of alkyl group size on ability to act as deoxythymidine triphosphate (dTTP) has been studied for the carcinogen products O2-methyl-, O2-ethyl-, and O2-isopropyl-dTTP by using three types of nucleic acids as template and DNA polymerase I (Pol I) or Klenow fragment as the polymerizing enzymes. Apparent Km and relative Vmax values were determined in primer extension on M13 DNA at a single defined site, in poly[d(A-T)], and in nicked DNA. These data are the basis for calculation of the relative rate of insertion opposite A, relative to dTTP. The insertion rate for any O2-alkyl-dTTP is much higher than for a mismatch between unmodified dNTPs. Unexpectedly, O2-isopropyl-dTTP is more efficiently utilized than O2-methyl-dTTP or O2-ethyl-dTTP on each of the templates. O2-isopropyl-dTTP also substitutes for dTTP over extended times of DNA synthesis at a rate only slightly lower than that of dTTP. Parallel experiments using O4-methyl-dTTP under the same conditions show that it is incorporated opposite A more frequently than is O2-methyl-dTTP. Therefore, both the ring position and the size of the alkyl group influence polymerase recognition. Once formed, all O2-alkyl-T.A termini permit elongation, as does O4-methyl-T.A. In contrast to the relative difficulty of incorporating the O-alkyl-dTTPs, formation of the following normal base pair (C.G) occurs rapidly when dGTP is present. This indicates that a single O-alkyl-T.A pair does not confer significant structural distortion recognized by Pol I.     

The immunocytochemical localization of superoxide dismutase in the enterocytes of the avian intestine: The effect of vitamin D3

Summary Both light microscopical and electron microscopical immunocytochemical techniques were utilized to localize CuZnsuperoxide dismutase (SOD) in the duodenum of normal, rachitic and vitamin-D₃-replete chicks. This enzyme catalyses the dismutation of the superoxide anion, a toxic free radical generated during the normal aerobic metabolism of most respiring cells. Light microscopy showed no SOD activity associated with the duodenal enterocytes of normal and rachitic chicks. However, in rachitic animals subsequently treated with vitamin D, i.e. vitamin-D-replete chicks, intense immunoreactivity for the enzyme was seen in association with the apical border of the duodenal absorptive cells. Immunostaining for SOD was not seen in goblet cells. With electron microscopy, immunostaining for SOD activity was identified in association with the apical microvilli and, to a lesser degree, with the terminal web, a well as in association with both lysosomes and peroxisomes. From this report it appears that there is a physiological relationship between vitamin D, SOD and the intestinal absorptive cell. However, the precise relationship must await further clarification.     

Geographical distribution,morphology and water quality of caldera lakes: a review

The geographical distribution, morphometry and water quality of lakes within large calderas (> 2 km in diameter) were evaluated through a review of the literature and maps. Eighty-eight lakes in 75 calderas were located in 31 volcanic subregions. As a group, the lakes varied greatly in elevation, surface area, maximum depth, and shoreline development. The average surface area was 16.9 km², surface elevation 873 m, depth 151.1 m, and shoreline development 1.35. Water quality ranged from ultraoligotrophic to highly eutrophic. None of the lakes had an inlet that originated outside the calderas. Most lakes did not have a surface outlet, were circular or subcircular in shape, and covered only parts of the caldera basins. Water clarity in some lakes was among the highest recorded for freshwater systems, but there are indications of possible declining clarity in some cases. Secondary volcanic activity, such as primary (hydrothermal) water and eruptions, has been associated with deteriorated water quality conditions in some lakes.     

Role of cell-cycle in regulating neuroepithelial cell shape during bending of the chick neural plate

Summary Neuroepithelial cells transform from spindle-shaped to wedge-shaped within the median and paired dorsolateral hinge points of the bending neural plate, but the mechanisms underlying these localized changes are unclear. This study was designed to evaluate further the hypothesis that localized wedging of neuroepithelial cells during bending involves basal cellular expansion resulting from alteration of the cell-cycle. Neurulating chick embryos were treated with tritiated thymidine, and transverse sections through the midbrain were examined autoradiographically. Parameters of the cell-cycle as well as nuclear position and size were assessed in the median hinge point, which contains predominantly wedge-shaped cells, and in adjacent lateral areas of the neural plate, which contain predominantly spindle-shaped cells. Both the DNA-synthetic phase and non-DNA synthetic portion of the cell-cycle were significantly longer in the median hinge point than in lateral neuroepithelial areas, some nuclei in both regions were located basally during these phases, and virtually all basal nuclei in the median hinge point were large. Additionally, the mitotic phase was significantly shorter in the median hinge point than in lateral areas. We present a model to explain how alteration of the cell-cycle in the median hinge point could generate wedging of cells in this region.