Distribution of histamine in the rat kidney during pregnancy and development

Summary An antiserum against conjugated histamine and two oligonucleotide probes that detect the mRNA encoding L-histidine decarboxylase (HDC) involved in histamine synthesis were used to study the appearance of histamine and its location in the kidneys of fetal, newborn and young postnatal rats and in the kidneys of pregnant rats. On embryonic days 16 and 18 (E16 and E18), some HA-immunoreactive (HA-ir) cells were found within the largest S-shaped bodies. Histamine was found to appear rapidly between the 18th and 20th embryonic days in the convoluted tubules of the kidneys. On postnatal day 0 (P0), the distal convoluted tubules and collecting ducts exhibited bright fluorescence, the intensity of which decreased quickly so that it was faint on day P4 and absent at later stages. In kidneys of pregnant rats HA-ir was found in the epithelium of both the Bowman's capsule, collecting ducts and in a few cells within the tubules. Nonuniform HA-ir was also detected within glomeruli. No evidence for the presence of L-histidine decarboxylase mRNA in kidneys of fetuses or pregnant rats was seen. It is concluded that distinct structures in the developing rat kidney contain histamine during a period around birth from day E20 to day P4. In the pregnant rat, the epithelium that is in direct contact with the urine flow is immunoreactive for histamine from day 16 to 20 of pregnancy. The results suggest that histamine is not synthesized locally in the kidneys but rather originates from other tissues.     

A Kdp-like,high-affinity,K+-translocating ATPase is expressed during growth of Rhodobacter sphaeroides in low potassium media

Cells of the purple non-sulphur bacterium Rhodobacter sphaeroides express a high-affinity K⁺ uptake system when grown in media with low K⁺ concentrations. Antibodies againts the catalytic KdpB protein or the whole KdpABC complex of Escherichia coli crossreact with a 70.0 kDa R. sphaeroides protein that was expressed only in cells grown in media with low K⁺ concentrations. In membranes derived from R. sphaeroides cells grown with low K⁺ concentrations (induced cells), a high ATPase activity could be detected when assayed in Tris-HCl pH 8.0 containing 1 mM MgSO₄. This ATPase activity increased upon addition of 1 mM KCl from 166 to 289 mol ATP hydrolysed x min^-1 x g protein^-1 (1.7-fold stimulation). The K⁺-stimulated ATPase activity was inhibited approximately 93% by 0.5 mM vanadate but hardly by N,N-dicyclohexylcarbo-diimide (DCCD). These results indicate that the inducible K⁺-ATPase in R. sphaeroides resembles the Kdp K⁺-translocating ATPase of Escherichia coli. This Kdp-like transport system is also expressed in R. capsulatus and Rhodospirillum rubrum during growth in media with low K⁺ concentrations suggesting a wide distribution of this transport system among phototrophic bacteria.Abbreviations electrical potential difference across the cytoplasmic membrane - pH pH difference across the cytoplasmic membrane - BSA bovine serum albumine - PAGE polyacrylamide gel electrophoresis - HEPES 4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid - PMSF phenyl-methyl-sulfonyl fluoride - DCCD N,N-dicyclohexylcarbodiimide - AIB 2--aminoisobutyric acid - TMG methyl--d-thiogalactopyranoside     

Chromosome aberrations (CA), sister-chromatid exchanges (SCE) and mitogen-induced blastogenesis in cultured peripheral lymphocytes from 48 control individuals sampled 8 times over 2 years

Confidence in the measurement of positive effects determined by monitoring of environmentally or occupationally exposed individuals can be enhanced by a knowledge of the normal variability in these endpoints in the general population. Confounding effects can be determined and study interpretation improved by correlation of this variability with various lifestyle factors such as sex and age of donor, smoking and drinking habits, viral infections, exposure to diagnostic X-rays, etc.

8 blood samples were taken from each of 24 male and 24 female volunteers over a period of 2 years. Questionnaires pertaining to lifestyle were completed at the time of each sampling. Whole blood was cultured and slides prepared for CA or SCE analysis. Separated mononuclear cells were cultured with a range of phytohaemagglutinin concentrations and the maximum level of mitogen-induced blastogenesis was determined by measurement of [³H]thymidine uptake.

There was a significant effect of both year and season of sampling for all 3 endpoints. No significant effects in any of the 3 endpoints were found with respect to sex or age of donor nor any of the other lifestyle factors, although SCE frequency and mitogen-induced blastogenesis were nearly always higher in females than males. These results point to the need for concurrent sampling of controls with exposed populations.     

Genetic assay of misincorporation

A system to characterize mutations arising from in vitro nucleotide misincorporation, which avoids the effects of in vivo mismatch repair on recovery of mutants, was constructed and evaluated. The lacI gene of Escherichia coli was inserted into phage M13 and the M13-lacI recombinant was introduced into a strain of E. coli lacking a resident lacI gene. In this system the function of the M13-bearing lacI gene can be detected by plaque color. Mutants in the 5'-region of the lacI gene (encoding operator-binding domain) are seen as blue plaques when the host strain is grown in the presence of chromogenic substrate, X-gal, in the absence of inducer. The use of uracil-containing single stranded DNA from M13-lacI as template for DNA synthesis avoids the contribution of mismatch repair (in transfection recipients) on the recovery of mutants. To demonstrate the usefulness of the M13-lacI system we produced nucleotide misincorporations by in vitro DNA synthesis in the N-terminal region of the lacI template in the presence of only 3 deoxynucleoside triphosphates (dNTPs). Such mutagenic reactions were conducted in the absence of dATP with 4 different primers and in the absence of dGTP with 2 primers. The type of mutants produced by these reactions were identified through sequencing of DNA from progeny phage after screening for i- (blue plaque) phenotype. Mutations recovered in this system consisted of single and multiple base substitutions in the region of the template near the 3'-terminus of the primer. Nearly all of the mutants induced by '-A' conditions were T----C base substitutions, and those induced by '-G' conditions were C----T transitions. In general, the results were consistent with the spectrum of spontaneous mutants produced in strains deficient in mismatch repair, although some differences were noted. Several new base substitutions within the lacI gene (producing i- phenotype and unobserved by others) were isolated by the procedures described in this paper.     

Effects of spines and thorns on Australian arid zone herbivores of different body masses

Summary We investigated the effects of thorns and spines on the feeding of 5 herbivore species in arid Australia. The herbivores were the rabbit (Oryctolagus cuniculus), euro kangaroo (Macropus robustus), red kangaroo (Macropus rufus), sheep (Ovis aries), and cattle (Bos taurus). Five woody plants without spines or thorns and 6 woody plants with thorns were included in the study. The spines and thorns were not found to affect the herbivores' rates of feeding (items ingested/min), but they did reduce the herbivores' rates of biomass ingestion (g-dry/item). The reduction in biomass ingested occurred in two ways: at a given diameter, twigs with spines and thorns had less mass than undefended plants, and the herbivores consumed twigs with smaller diameters on plants with spines and thorns. The relative importance of the two ways that twigs with spines and thorns provided less biomass varied with herbivore body mass. Reduced twig mass was more important for small herbivores, while large herbivores selected smaller diameters. The effectiveness of spines and thorns as anti-herbivore defenses did not vary with the evolutionary history of the herbivores (i.e. native vs. introduced). Spines and thorns mainly affected the herbivores' selection of maximum twig sizes (reducing diameter and mass), but the minimum twig sizes selected were also reduced.     

Continuous shoot growth monitoring in hydroponics

A weighing apparatus for automatic recording of fresh weight of shoots of spinach plants ( Spinacia oleracea L., cv. Subito) growing in nutrient solution is described. The system was tested for 17 days in a controlled environment and enabled the determination of the relative growth rate (RGR) of the shoot fresh weight. Results from three consecutive growth experiments demonstrated diurnal fluctuations in the relative growth rate of the shoot fresh weight. In general, relative growth rates were between 0.32 and 0.36 day⁻¹ 16 days after sowing and decreased to between 0.11 and 0.18 day⁻¹ during the 12 following days. The variance between three replicate growth curves was compared with the variance of a growth function fitted through destructively obtained spinach shoot weight data.     

Distribution and immunolocalization of stachyose synthase in Cucumis melo L.

Indirect evidence for the site of stachyose biosynthesis has been provided by determining the occurrence and distribution of stachyose, raffinose and galactinol, the donor of the galactosyl moiety for stachyose synthesis, in Cucumis melo L. cv. Ranjadew. Studies of enzyme activities for the synthesis of these sugars and their distribution in different plant organs and isolates has led to the conclusion that stachyose is synthesized mainly in mature leaves and seeds. Nevertheless, stachyose-synthase activity varied with leaf age, the developmental stage of a plant, the growing season and the plant cultivar used. No stachyose or stachyose-synthase activity could be detected in isolated mesophyll protoplasts and chloroplasts, whereas both were found in a minor-vein-enriched fraction isolated from mature leaves. The conclusion that stachyose biosynthesis is associated with minor veins was confirmed by immunolocalization of the enzyme. Positive specific immunoreactivity of stachyose synthase with polyclonal anti-stachyose-synthase antibodies, labeled with protein A-gold, was detected in intermediary cells of leaf minor veins. The implication of this local synthesis of the main transport sugar for phloem loading in mature leaves of Cucumis melo is discussed.Abbreviation RUBPCase ribulose-1,5-bisphosphate carboxylase This work was supported by Deutsche Forschungsgemeinschaft. The excellent assistance of Ms. B. Müller in preparing the samples for electron microscopy is gratefully acknowledged. The authors thank Professor H.J. Schneider-Poetsch for anti-RuBPCase antibodies.