Nodulation ofTrifolium subterraneum byRhizobium leguminosarum

Summary Four cultivars ofTrifolium subterraneum were nodulated by five strains ofRhizobium leguminosarum; all combinations except one gave 100% nodulation. Rates of nodule formation and total nodule numbers were similar to those with an effectiveR. trifolii strain. The nodules were more commonly associated with lateral roots and were ineffective in nitrogen fixation.     

Response of chloride efflux from skeletal muscle ofRana pipiens to changes of temperature and membrane potential and diethylpyrocarbonate treatment

Summary Efflux of³⁶Cl^– from frog sartorius muscles equilibrated in two depolarizing solutions was measured. Cl^– efflux consists of a component present at low pH and a pH-dependent component which increases as external pH increases.For temperatures between 0 and 20°C, the measured activation energy is 7.5 kcal/mol for Cl^– efflux at pH 5 and 12.6 kcal/mol for the pH-dependent Cl^– efflux. The pH-dependent Cl^– efflux can be described by the relationu=1/(1+10n(pK _a -pH)), whereu is the Cl^– efflux increment obtained on stepping from pH 5 to the test pH, normalized with respect to the increment obtained on stepping from pH 5 to 8.5 or 9.0. For muscles equilibrated in solutions containing 150mm KCl plus 120mm NaCl (internal potential about –15 mV), the apparent pK _a is 6.5 at both 0 and 20°C, andn=2.5 for 0°C and 1.5 for 20°C. For muscles equilibrated in solutions containing 7.5mm KCl plus 120mm NaCl (internal potential about –65 mV), the apparent pK _a at 0°C is 6.9 andn is 1.5. The voltage dependence of the apparent pK _a suggests that the critical pH-sensitive moiety producing the pH-dependent Cl^– efflux is sensitive to the membrane electric field, while the insensitivity to temperature suggests that the apparent heat of ionization of this moiety is zero. The fact thatn is greater than 1 suggests that cooperativity between pH-sensitive moieties is involved in determining the Cl^– efflux increment on raising external pH.The histidine-modifying reagent diethylpyrocarbonate (DEPC) applied at pH 6 reduces the pH-dependent Cl^– efflux according to the relation, efflux=exp(–k·[DEPC]·t), wheret is the exposure time (min) to DEPC at a prepared initial concentration of [DEPC] (mm). At 17°C,k ^–1=188mm·min. For temperatures between 10 and 23°C,k has an apparent Q₁₀ of 2.5. The Cl^– efflux inhibitor SCN^– at a concentration of 20mm substantially retards the reduction of the pH-dependent Cl^– efflux by DEPC. The findings that the apparent pK _a is 6.5 in depolarized muscles, that DEPC eliminates the pH-dependent Cl^– efflux, and that this action is retarded by SCN^– supports the notion that protonation of histidine groups associated with Cl^– channels is the controlling reaction for the pH-dependent Cl^– efflux.     

Detection and quantification of triazine herbicides using algal cell fluorescence

Summary A continuous-flow system is described which, by measuring fluorescence of the unicellular alga Chlorella, is capable of measuring concentrations of the triazine herbicide, simazine, as low as 60nM (approx 12g l^-1) within 5 minutes. Further developments are suggested to achieve the desired detection limit of 0.5nM. The use of such an instrument in environmental analysis is discussed.     

Long-term culture of human aortas

Summary Segments of human thoracic aorta were maintained in long-term explant culture for 18 weeks in serum-supplemented medium. The aortas were grossly normal in appearance, and random samples fixed for light microscopy prior to culture revealed a normal morphology. The intima contained no more than five layers of smooth muscle cells. After 7 days in culture, the intima was noticeably thicker than the uncultured segments. The increased thickness was due to proliferating smooth muscle cells and production of extracellular material. After several months in culture, extracellular material consisting of collagen and flocculent material was present in areas resembling atherosclerotic fibrous plaques. A peripheral growth, which formed around the explant, was composed of fibroblastlike cells and added to the overall thickness of the intima. However, aortic segment maintained for up to 2 months in serum-free culture medium showed no cellular proliferation. This study demonstrates that changes resembling early stages of atherosclerosis occur in human aortas maintained in explant culture using routine culture procedures. Supported in part by the Pangborn Fund and the Graduate School of the University of Maryland. This is publication 443 from the Cellular Pathobiology Laboratory.     

Cyclin D1 and cyclin D3 show divergent responses to distinct mitogenic stimulation

D‐type cyclins predominantly regulate progression through the cell cycle by their interactions with cyclin‐dependent kinases (cdks). Here, we show that stimulating mitogenesis of Swiss 3T3 cells with phorbol esters or forskolin can induce divergent responses in the expression levels, localization and activation state of cyclin D1 and cyclin D3. Phorbol ester‐mediated protein kinase C stimulation induces S phase entry which is dependent on MAPK activation and increases the levels and activation of cyclin D1, whereas forskolin‐mediated cAMP‐dependent protein kinase A stimulation induces mitogenesis that is independent of MAPK, but dependent upon mTor and specifically increases the level and activation of cyclin D3. These findings uncover additional levels of complexity in the regulation of the cell cycle at the level of the D‐type cyclins and thus may have important therapeutic implications in cancers where specific D‐cyclins are overexpressed. J. Cell. Physiol. 225: 638–645, 2010. © 2010 Wiley‐Liss, Inc.