Occurrence of 28- and 27-Carbon ecdysteroids and sterols in developing worker pupae of the leaf-cutting ant acromyrmex octospinosus (reich) (hymenoptera,formicidae: Attini)

The major ecdysteroids in large worker pupae of the leaf-cutting ant Acromyrmex octospinosus were characterized at the peak ecdysteroid concentration by using high-performance liquid chromatography, enzyme immunoassay, and mass spectrometry. In decreasing amounts, they were determined to be makisterone A, an unidentified C₂₈ ecdysteroid bearing a molecular weight of 494, 20-hydroxyecdysone (ratio of 1 to 6 as compared to makisterone A), and putative but negligible ecdysone. The presence of both C₂₈ and C₂₇ ecdysteroids is discussed in relation to the content of 4-desmethylsterols determined by gas chromatography and mass spectrometry to be ergosta-5,7,24 (28)-trien-3β-ol, ergosterol, ergosta-5,7-dien-3β-ol and ergosta-7,24(28)-dien-3β-ol for the main sterols, and with a small amount of cholesterol.     

Human and rat chromosomal localization of two genes for 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase by analysis of somatic cell hybrids and in situ hybridization

Two genes encoding 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase were localized in human and rat chromosomes. PFKFB1 (previously PFRX), which encodes the liver and muscle isozymes, was assigned to Xq22-q31 in the rat and to Xq27–q28 in the human by in situ hybridization using probes generated by the polymerase chain reaction. PFKFB2, which encodes the heart isozyme of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase, was assigned to chromosome 13 in the rat and to chromosome 1 in the human by hybridization of DNA from somatic cell hybrids. By in situ hybridization, this gene was localized to the regions 13q24–25 in the rat and 1q31 in the human.     

Rhodopsin-stimulated activation-deactivation cycle of transducin: kinetics of the intrinsic fluorescence response of the alpha subunit

The intrinsic tryptophan fluorescence of the alpha subunit of transducin (alpha T) has been shown to be sensitive to the binding of guanine nucleotides, with the fluorescence being enhanced by as much as 2-fold upon the binding of GTP or nonhydrolyzable GTP analogues [cf. Phillips and Cerione (1988) J. Biol. Chem. 263, 15498-15505]. In this work, we have used these fluorescence changes to analyze the kinetics for the activation (GTP binding)-deactivation (GTPase) cycle of transducin in a well-defined reconstituted phospholipid vesicle system containing purified rhodopsin and the alpha T and beta gamma T subunits of the retinal GTP-binding protein. Both the rate and the extent of the GTP-induced fluorescence enhancement are dependent on [rhodopsin], while only the rate (and not the extent) of the GTP gamma S-induced enhancement is dependent on the levels of rhodopsin. Comparisons of the fluorescence enhancements elicited by GTP gamma S and GTP indicate that the GTP gamma S-induced enhancements directly reflect the GTP gamma S-binding event while the GTP-induced enhancements represent a composite of the GTP-binding and GTP hydrolysis events. At high [rhodopsin], the rates for GTP binding and GTPase are sufficiently different such that the GTP-induced enhancement essentially reflects GTP binding. A fluorescence decay, which always follows the GTP-induced enhancement, directly reflects the GTP hydrolytic event. The rate of the fluorescence decay matches the rate of [32P]Pi production due to [gamma-32P]GTP hydrolysis, and the decay is immediately reversed by rechallenging with GTP. The GTP-induced fluorescence changes (i.e., the enhancement and ensuing decay) could be fit to a simple model describing the activation-deactivation cycle of transducin. The results of this modeling suggest the following points: (1) the dependency of the activation-deactivation cycle on [rhodopsin] can be described by a simple dose response profile; (2) the rate of the rhodopsin-stimulated activation of multiple alpha T(GDP) molecules is dependent on [rhodopsin] and when [alpha T] greater than [rhodopsin], the activation of the total alpha T pool may be limited by the rate of dissociation of rhodopsin from the activated alpha T(GTP) species; and (3) under conditions of optimal rhodopsin-alpha T coupling (i.e., high [rhodopsin]), the cycle is limited by GTP hydrolysis with the rate of Pi release, or any ensuing conformational change, being at least as fast as the hydrolytic event.     

Host fruit chemical stimuli eliciting distinct ovipositional responses from sibling species of Rhagoletis fruit flies

Laboratory experiments tested whether two economically-important sibling species of tephritid fruit flies have evolved distinct egg-laying responses to chemical stimuli on the fruits of their respective hostplants. The egg-laying preferences displayed by apple maggot flies, R. pomonella, and blueberry maggot flies, R. mendax, on artificial fruits treated with apple and blueberry extract paralleled their egg-laying responses to whole apples and blueberries. R. pomonella flies laid more eggs than R. mendax flies in artificial fruits treated with extract from ripe McIntosh apples, and vice versa for artificial fruits treated with extract from ripe Bluehaven blueberries. Furthermore, both species laid more eggs in artificial fruits treated with extract from their respective host fruits than control artificial fruits which were not treated with fruit extract. Prior electroantennogram recordings from R. mendax and R. pomonella flies exposed to volatiles from pentane extracts of apples and blueberries indicate that the antennal sensitivity of both species is selectively tuned to their respective host fruit odors. This differentiation in their olfactory responses to fruit odors could be important in mediating their distinct ovipositional responses to blueberry and apple fruits. Extract from unripe McIntosh apples also elicited egg laying by R. pomonella flies, however, artificial fruits treated with unripe apple extract received 1.9 times fewer eggs than those treated with ripe apple extract. Moreover, the numbers of R. pomonella ovipositor punctures and eggs placed in wax artificial fruits were increased when the artificial fruits were treated with a blend of 7 identified apple esters. Black coloration on these artificial fruits and the presence of apple esters had a synergistic effect on the egg-laying behavior of R. pomonella flies, which caused them to lay substantially more eggs per black fruit than white fruit treated with the same concentration of apple esters. In summary, our results indicate that the egg-laying responses of R. pomonella flies are mediated by the integration of information from fruit chemical and visual cues, and that R. mendax and R. pomonella flies have evolved divergent egg-laying responses to chemical stimuli on the fruits of their respective hostplants. These findings are discussed in the context of other studies on plant compounds which influence the ovipositional behavior of phytophagous Diptera.

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Stimuli chimiques des pommes et des myrtilles induisant la ponte des espèces jumelles, Rhagoletis pomonella et R. mendax

Résumé Des fruits artificiels en cire traités avec des extraits de fruits ont provoqué chez les espèces jumelles de R. mendax (Curran) et R. pomonella (Walsh) des réactions de ponte différentes suivant les stimulations chimiques par les fruits. Le comportement de ponte sur des fruits artificiels traités avec des extraits au pentane des myrtilles mûres (Vaccinium corymbosum L.) et de pommes mûres (Malus pumila Miller = Pyrus malus L.), est le même que sur des fruits naturels, ce qui montre que la réponse aux stimulations chimiques provenant du fruit constitue un aspect important de la reconnaissance de l'hôte. R. pomonella pond plus d'ufs que R. mendax sur les fruits artificiels traités à l'extrait de pommes mûres; c'est l'inverse pour les fruits traités aux extraits de myrtille. Les fruits artificiels traités avec des pommes ou des myrtilles provoquent la ponte de R. pomonella, tandis que les myrtilles mûres seules provoquent la ponte de R. mendax. Les extraits de pommes vertes stimulent la ponte de R. pomonella mais elle est alors 2 fois plus faible qu'avec des extraits de pommes mûres. Un mélange de 7 esters identifiés dans l'extrait de pomme induit aussi la ponte de R. pomonella. Le nombre de piqûres de tarièresfli dans les fruits artificiels en cire et le nombre d'ufs par fruit ont été augmentés par addition d'esters de pommes à des fruits blancs ou noirs. La couleur des fruits artificiels influence aussi la réaction de ponte de R. pomonella; la fréquence des piqûres de tarière contenant un uf et le nombre d'ufs par fruit étaient significativement plus élevés sur les fruits noirs que sur les fruits blancs traités avec la même concentration d'esters de pomme. Les fruits artificiels noirs traités avec la concentration la plus stimulante d'esters de pommes ont reçu 2, 3 fois plus d'ufs que les fruits blancs avec les mêmes concentrations en esters. Ces résultats montrent que les esters de pomme et la couleur noire stimulent synergiquement la ponte de R. pomonella sur des fruits artificiels.

     

Vanadium-containing tunicate blood cells are not highly acidic

The intracellular pH of intact blood cells of the tunicate Ascidia nigra was measured by transmembrane equilibration of [¹⁴C]methylamine. The pH of unfractionated blood cells is 7.39±0.10. The pH of vanadocytes, determined in a fractionation study, is 7.2. Previously used methods, in which pH values less than 3.0 are inferred from cell lysis or vital staining experiments, are shown to be unsuitable for intracellular pH determination due to the chemical composition of these vanadium-containing cells.     

Systems for exposing mice to 2,450-MHz electromagnetic fields

Two systems for exposing mice to 2,450-MHz electromagnetic fields are described. In a waveguide system, four mice were placed in a Styrofoam cage and exposed dorsally to circularly polarized electromagnetic fields. The temperature and humidity in the mouse holder were kept constant by forced-air ventilation. For 1-W input power to the waveguide, the average specific absorption rate (SAR) was determined by twin-well calorimetry to be 3.60 ± 0.11 (SE) W/kg in 27-g mice. The maximum SAR at the skin surface determined thermographically was 8.36 W/kg in the head of the mouse. The second system was a miniature anechoic chamber. Six mice were irradiated dorsally to far field plane waves. Copper shielding and high-temperature absorbing material were lined inside the chamber to accommodate the high input power. The air ventilation at the location of the mice was separately controlled so that any heating in the absorber would not affect the animals. For 1-W input power, the average SAR was 0.17 ± 0.01 W/kg and the maximum SAR at the skin surface was 0.41 W/kg in the animal when irradiated with body axis parallel to the E field; the SARs were 0.11 ± 0.01 W/kg and 0.64 W/kg, respectively, when irradiated perpendicular to the E field.     

Role of glycerol 3-phosphate and glycerophosphate acyltransferase in the nutritional control of hepatic triacylglycerol synthesis

1. Glycerol 3-phosphate content of isolated hepatocytes from starved rats and of glycogen-depleted hepatocytes from fed rats was low and severely limited triacylglycerol synthesis. 2. Raising the glycerol 3-phosphate content by addition of precursors to the cells resulted in a hyperbolic-like relationship between triacylglycerol synthesis and cellular glycerol 3-phosphate content. Statistical analysis of the curves showed no significant differences between the nutritional states either at saturating or at subsaturating glycerol 3-phosphate content. 3. V_max. of glycerophosphate acyltransferase measured in homogenized hepatocytes was decreased by 30–40% in starvation. There was no change in apparent K_m for glycerol 3-phosphate. Since at saturating glycerol 3-phosphate content esterification rates in hepatocytes of both nutritional states were identical, the enzyme is not limiting esterification under this condition. 4. At subsaturating glycerol 3-phosphate content the flux through glycerophosphate acyltransferase necessarily limits esterification. Therefore one would expect a decrease in esterification in starvation under this condition. This was the case when triacylglycerol synthesis was plotted against intracellular glycerol 3-phosphate concentration, calculated from the cellular glycerol 3-phosphate content and the intracellular water space, which was smaller in hepatocytes from starved rats. 5. The data obtained in hepatocytes were extrapolated to the intact liver by using the number of parenchymal cells per g of liver as determined from marker-enzyme analysis and the liver weight per 100g body weight. The extrapolation suggested that glycerol 3-phosphate is limiting esterification in vivo for contents below 0.3–0.4 and 0.5–0.65μmol/g for livers from fed and starved animals respectively. Also for a given fatty acid load and a glycerol 3-phosphate content below 0.3μmol/g the liver may esterify less in the starved state. However, at the glycerol 3-phosphate contents measured in freeze-clamped livers (0.30 and 0.44μmol/g for the fed and starved state respectively), livers in both nutritional states seemed capable of esterifying similar amounts of fatty acids.     

Protonation and light synergistically convert plasmalemma sugar carrier system in mesophyll protoplasts to its fully activated form

The course of sugar fluxes into and out of protoplasts isolated from the mesophyll of Pisum sativum L. has been followed over brief time intervals (minutes). Light strongly stimulated net sugar influx at pH 8 as well as at pH 5.5. The proton conductor carbonyl cyanide m-chlorophenylhydrazone (CCCP) inhibited initial influx in the light, both at pH 8.0 and at pH 5.5. CCCP was without effect in the dark at either pH. All these results applied both to sucrose and to the nonmetabolizable glucose analog 3-O-methyl-d-glucose.When protoplasts at pH 5.5 were transferred from light to darkness, "stored" light driving force maintained uptake in the dark at the full light rate for the first 7 minutes. At pH 8, however, even 4 minutes after transfer to dark, uptake was well below the light rate. Initial uptake rates over a range of external concentrations were derived from progress curves obtained in the light and in the dark, both at pH 5.5 and at 7.7. When initial rate was plotted against concentration, simple Michaelis-Menten kinetics were observed only under the condition pH 5.5, light. In the dark at both pH values, and in the light at pH 7.7, complex curves with intermediate plateaus were obtained, strongly resembling curves reported for systems where mixed negative and positive cooperativity is operating.The same "K(m) for protons" was observed in the dark and in the light (10(-7) molar). Switching protoplasts in the dark from pH 8 to 5.5 failed to drive sugar transport by imposed protonmotive force, as judged by lack of sensitivity to CCCP. Switching protoplasts which had taken up sugar in the dark at pH 5.5 to pH 7 induced net efflux of sugar. Flux analysis showed that this effect was entirely due to the prompt fall in influx.It is concluded from the kinetic experiments that protonation alone is not sufficient to convert the sugar transport system to its fully activated high affinity form. A further light-dependent factor which acts synergistically with protonation is required.