Properties of human brain glycine receptors expressed in Xenopus oocytes

Glycine and gamma-aminobutyric acid (GABA) receptors from the foetal human brain were 'transplanted' into the Xenopus oocyte membrane by injecting the oocytes with poly(A)+-mRNA extracted from the cerebral cortex. Activation of both glycine and GABA receptors induced membrane currents carried largely by chloride ions. However, unlike the GABA-activated current, the glycine current was blocked by strychnine, and was not potentiated by barbiturate. At low doses, the glycine current increased with concentration following a 2.7th power relation, suggesting that binding of three molecules of glycine may be required to open a single membrane channel. The current induced by steady application of glycine decreased with hyperpolarization beyond about -60 mV.     

Increased organization of cytoskeleton accompanying the aging of human fibroblasts in vitro

Fibroblastic cells of human origin have a limited lifespan in culture. One of the senescence-associated phenotypic changes is an increase in the abundance of cytoplasmic filaments. Human skin fibroblasts (strain 0011), derived from an 8-week-old male fetus, were passaged according to a predetermined schedule and examined at successive population doubling levels. In young rapidly growing cultures, fluorescence microscopy with NBD-Phallacidin shows a normal organization of the actin-containing fibers, microtubules and intermediate filaments, as has been described previously. At stages close to the end of the in vitro lifespan of the cell strain, large flat fibroblasts are the predominant cell type in culture. These large senescent fibroblasts contain numerous prominent actin fibers traversing the entire long axis of the cytoplasm. The fibers are often located adjacent to each other and appear to form a sheet on the ventral side of the cytoplasm. Staining of senescent cells with anti-tubulin antibody reveals an increase in the abundance of microtubules per cell and the distribution pattern is altered through the increase in the number of organization centers. Intermediate filaments are also more abundant and display tightly packed fibrillar sheets or bundles. Electron microscopic studies have confirmed the increased organization of microfilaments into bundles in senescent cells. These results suggest that during in vitro senescence, the increase in cell size is correlated with increased organization of the cytoskeleton. The presence of a rigid cytoskeletal structure may contribute in part to the inability of the senescent cell to replicate.     

Further studies of dopamine metabolism and function in Tetrahymena

The large amounts of dopamine accumulated by cells of Tetrahymena pyriformis strain NT-1 and secreted into their growth medium were found to depend primarily upon an extracellular, non-enzymatic conversion of tyrosine to L-dihydroxyphenylalanine (L-DOPA); L-DOPA was then rapidly taken into the cells and transformed into dopamine enzymatically. Efforts to find physiologically significant dopamine binding sites on the cell surface or dopamine-sensitive adenylate cyclase activity were unsuccessful, suggesting that the catecholamine does not function in Tetrahymena as it does in higher animals.     

Deoxyribonucleotide Pools in Plant Tissue Cultures

Two dimensional thin-layer chromatography on anion-exchange cellulose enables the separation of the normally occurring ribo- and deoxyribonucleoside triphosphates. This technique was applied to perchloric acid extracts of callus tissue of sycamore and tobacco and of pine pollen grown in ³²P-orthophosphate labelled media to quantitate the nucleoside triphosphate pools under different growth conditions. The results showed that the ratio of the deoxyribonucleo-side triphosphates to their corresponding ribonucleoside triphosphates is low in plant cells, similar to the ratio previously found for animal cells. During the period of most rapid DNA synthesis in the callus tissue, the deoxyribonucleoside triphosphate pools reach their highest values. This effect is also demonstrated with cells of Escbericbia coli.     

Influence of sewage discharge on nitrogen fixation and nitrogen flux from coral reefs in Kaneohe Bay, Hawaii.

Nitrogen fixation was investigated in Kaneohe Bay, Oahu, Hawaii, a subtropical eutrophic estuary, by using the acetylene reduction technique on algal samples. No active, planktonic, N2-fixing blue-green algae or bacteria were observed. However, Calothrix and Nostoc capable of fixing N2 were cultured from navigational buoys and dead coral heads. Nitrogen fixation associated with these structures was greater in the middle sector than in the south and north sectors of the estuary. Experiments demonstrated that the fixation was photosynthetically dependent. Examination of the data showed that there was no significant correlation between rates of nitrogen fixation and concentration of combined nitrogen compounds in the Bay water. Fixation was significantly correlated to the inorganic N/P (atomic) ratio in the south and middle sectors but not in the north sector. The nutrient data indicate there was a flux of combined nitrogen, but not phosphate, from the reef flats.     

The Quantitative Yield in Purification of Cytokinins. Model-experiments with Kinetin, 6-Furfuryl-amino-purine

Model-experiments with kinetin, 6-furfuryl-amino-purine, to determine the quantitative yield in purification of cytokinin extracts have been performed. If kinetin in acidic water solution is partitioned three times with equal volumes of ethyl ether, about 50 per cent of the kinetin passes into the ether phases. In similar experiments with ethyl acetate more than 90 per cent of the kinetin goes over to the ethyl acetate phases. Accordingly, use of these two solvents in purification of cytokinin extracts leads to very large losses. Use of petroleum ether or n-hexane on the other hand leads to none or very small losses. Partitioning of alkaline kinetin solutions three times with equal volumes of 1-butanol results in an almost quantitatitve extraction of the kinetin from the water solution. About 7 per cent of the original amount of kinetin follows the acid auxin, when a saturated ether solution of kinetin is extracted according to a common method for acid auxins. Kinetin may thus interfere in later analyses for auxins. The dangers involved when cytokinins are “purified” according to normal practice are pointed out.     

Larval aestivation and direct development as alternative strategies in the speckled wood butterfly, Pararge aegeria, in Sweden

ABSTRACT.

• 1 Sweden has two disjunct populations of the speckled wood butterfly, Pararge aegeria L. The southern population has two generations per year but the central Swedish population is univoltine. When rearing larvae from central Sweden under normal photoperiodic conditions but at temperatures slightly above the ambient, 42% of the larvae developed directly and produced a second generation of adults the same summer. The egg—larval development time of the directly developing individuals was about 40 days, whereas that of the individuals developing along the univoltine pathway was about 100 days.
• 2 Larvae of the central Swedish population normally aestivate during part of the summer even though abundant food is available. In the closely related Lasiommata petropolitana F., which is the only Swedish satyrid that overwinters in the pupal stage besides P.aegeria, larvae do not aestivate, indicating that there does not seem to be any obligatory association between pupal hibernation and larval aestivation.
• 3 Development rates of aestivating and directly developing P.aegeria are equal up to the third larval instar. During the third and fourth instars, however, the development rate of aestivating individuals is retarded and females also have an additional fifth instar.
• 4 Since the central Swedish P.aegeria have the capacity to develop directly, and the southern Swedish ones have the capacity to aestivate, the evidence indicates that the outcome of the cost/benefit balance of univoltine versus bivoltine development differs between the two areas.

     

Summer mastitis experimentally induced by Hydrotaea irritans exposed to bacteria

Abstract. Summer mastitis is an acute suppurative bacterial infection of the udder in heifers and dry cows. To ascertain the possible role of flies in the transmission of the disease, experimental exposures of recipient heifers to Hydrotaea irritans previously exposed to bacteria were carried out. Flies were allowed to feed on secretions from clinical cases of summer mastitis. The pathogens present were the bacteria Staphylococcus aureus, Streptococcus dysgalactiae, Actinomyces pyogenes , Stuart-Schwan cocci, Peptococcus indolicus, Fusobacterium necrophorum and Bacterioides species. The teats of eight heifers were exposed to flies with verified pathogen content. Two teats of each animal were deliberately damaged before fly exposure. One teat was cut, another pricked with insect needles to mimic insect bites. Two of the heifers developed summer mastitis in the quarters where teats had been cut. The bacterial species isolated from these quarters corresponded to those that had previously been fed to the flies. For the first time, it is now demonstrated that H.irritans is capable of transmitting summer mastitis pathogens and so causing summer mastitis in recipient heifers. Lesions on the teat orifice may be a predisposing factor in the development of the disease.     

Nested PCR for ultrasensitive detection of the potato ring rot bacterium, Clavibacter michiganensis subsp. sepedonicus.

Oligonucleotide primers derived from sequences of the 16S rRNA gene (CMR16F1, CMR16R1, CMR16F2, and CMR16R2) and insertion element IS1121 of Clavibacter michiganensis subsp. sepedonicus (CMSIF1, CMSIR1, CMSIF2, and CMISR2) were used in nested PCR to detect the potato ring rot bacterium C. michiganensis subsp. sepedonicus. Nested PCR with primer pair CMSIF1-CMSIR1 followed by primer pair CMSIF2-CMSIR2 specifically detected C. michiganensis subsp. sepedonicus, while nested PCR with CMR16F1-CMR16R1 followed by CMR16F2-CMR16R2 detected C. michiganensis subsp. sepedonicus and the other C. michiganensis subspecies. In the latter case, C. michiganensis subsp. sepedonicus can be differentiated from the other subspecies by restriction fragment length polymorphism (RFLP) analyses of the nested PCR products (16S rDNA sequences). The nested PCR assays developed in this work allow ultrasensitive detection of very low titers of C. michiganensis subsp. sepedonicus which may be present in symptomiess potato plants or tubers and which cannot be readily detected by direct PCR (single PCR amplification). RFLP analysis of PCR products provides for an unambiguous confirmation of the identify of C. michiganensis subsp. sepedonicus.