Sympathetic and adrenocorticoid influences on diet-induced thermogenesis and brown fat activity in the rat

Bilateral adrenalectomy markedly reduced body weight and energy gain and energetic efficiency of adult cafeteria-fed rats but enhanced the thermogenic response to food and stimulated brown fat activity. These changes were totally prevented by replacement of the animals with corticosterone (1 mg/rat/day). Unilateral denervation of the sympathetic nerves supplying the interscapular brown adipose tissue abolished the enhanced activity resulting from adrenalectomy and inhibited thermogenic activity in brown fat from cafeteria rats with intact adrenals, but had no effect in adrenalectomised animals treated with a high dose of corticosterone.     

Identification of a possible nucleotide binding site in CheW, a protein required for sensory transduction in bacterial chemotaxis

CheW is an essential component of the system which mediates chemotaxis in Salmonella typhimurium and Escherichia coli. Here we report the nucleotide sequence of the cheW gene as well as the purification and characterization of the CheW protein. The DNA sequence predicts a protein of 18,000 molecular weight. The pure protein exhibits an apparent molecular weight of 18,000 during sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Molecular sieve chromatography under nondenaturing conditions indicates a molecular weight of approximately 35,000, however. This result suggests that CheW is a homodimer. The predicted amino acid sequence between Thr-128 and Asp-160 fits a consensus exhibited by many proteins which bind purine nucleotides.     

Development of a single probe for documentation of chimerism following bone marrow transplantation.

Although numerous genetic markers are available for studying chimerism after bone marrow transplantation (BMT), there remains a need for a practical and highly informative method that is applicable in the early posttransplantation period. Using DNA restriction-fragment-length polymorphisms (RFLPs), we have evaluated the feasibility of developing a single synthetic oligonucleotide probe to study post-BMT chimerism. We have thus tested three candidate probes, termed O-3315-32, O-3315-80, and O-AY-29, that are homologous to tandemly repetitive sequences. Our results demonstrated donor-specific and recipient-specific fragments in 11 of 11 HLA-matched sibling pairs tested using probes O-3315-32 and O-3315-80. When probe O-AY-29 was used, 14 of 17 sibling pairs showed both donor and recipient markers, one had only a recipient marker, and two were identical. We showed that each of the three synthetic probes was effective in documenting donor marrow engraftment, mixed hematopoietic chimerism, the patient's pre-BMT phenotype (by using cultured skin fibroblasts obtained after BMT), and the origin of the malignant hematopoietic cells (i.e., of donor or recipient origin) in patients who developed recurrent hematologic malignancy following BMT. Compared with the use of cloned genomic probes, there are several important advantages to the use of synthetic oligonucleotide probes in studying post-BMT chimerism. Synthetic probes have absolute hybridization specificity and can be designed to suit the purposes of an individual study, since they have adjustable specificity that can be altered by changes in the length of the probe and by changes in the hybridization temperature. A single synthetic probe analogous to several highly polymorphic loci can have a polymorphism information content sufficiently high so that all but a small percentage of BMT patients could be followed easily; for example, if a probe were complementary to three highly polymorphic unlinked loci, it would discriminate approximately 98% of sibling donor/recipient pairs. This would be accomplished using only one restriction-endonuclease digestion and only one gel electrophoresis. Since other genetic markers, e.g., red blood cell antigens, immunoglobulin allotypes, and chromosome analysis, are not uniformly informative and, in some cases, cannot be used in the early posttransplantation period, the use of synthetic oligonucleotide probes for analysis of DNA RFLP is emerging as the method of choice for studies of post-BMT chimerism. This method will allow for the development of new knowledge that has not been possible with previous methods.     

Pharmacological profile of a novel carbacyclin derivative with high metabolic stability and oral activity in the rat

A novel carbacyclin derivative (16S)-13,14-dehydro-16,20-dimethyl-3-oxa-18,18,19,19-tetradehydro- 6a- carbaprostaglandin-I2 (3-oxa-analogue) has been synthesized in order to find chemically and metabolically stable prostacyclin-mimetics with a potency equal or even superior to PGI2. The 3-oxa-analogue was found to be stabilized against beta-oxidation, a main metabolic degradation step also for chemically stable PGI2-analogues. The compound is orally available and displays a long duration of 4.5-48 h of antiaggregatory and hypotensive action. The 3-oxa-analogue inhibits ADP-induced platelet aggregation with an IC50 of 3.0 nM. Following intravenous application the 3-oxa-analogue lowers diastolic blood pressure in a dose dependent manner, the ED20 being 0.1-0.2 micrograms/kg after injection and less than or equal to 0.05 micrograms/kg/min after infusion respectively. In vivo platelet aggregation is inhibited after i.v. infusion of the 3-oxa-analogue with an IC50 of 0.037 micrograms/kg/min. As compared to Iloprost, the 3-oxa-analogue is 5-12 fold more potent with respect to in vivo hypotensive and anti-aggregatory effects. The results of the present studies indicate that the 3-oxa-analogue has a pharmacological profile comparable to prostacyclin (PGI2) and Iloprost. Due to the fact that the 3-oxa-analogue is chemically and metabolically stable, long term oral treatment can be achieved in clinical conditions in which PGI2 and Iloprost have already been shown to be therapeutically useful principles.     

Oxidative metabolism of dermatophytes

A method for preparing young, actively respiring dermatophyte mycelia was obtained through the use of concentrated spore inocula and short growth periods in static culture. These hyphal elements were uniform in appearance, and vacuoles were absent. Concentrated mycelial suspensions were obtained which could be transferred easily and accurately. Glucose stimulated oxygen uptake in young mycelia which had been grown in a medium with low carbohydrate content. The level of endogenous respiration was affected by exogenous glucose only when this substrate stimulated oxygen uptake by less than 14%. Low nicotinamide adenine dinucleotide phosphate (NADP) dehydrogenase activity was noted in microconidia which have a low endogenous Q_o2 value, whereas the activity of this enzyme was greater in macroconidia and mycelia which possess higher endogenous Q_o2 values. Microsporum gypseum oxidizes 50% of exogenous glucose and assimilates the remainder. A large percentage of this substrate was assimilated into nitrogenous substances.     

Identification of the site of phosphorylation of the chemotaxis response regulator protein, CheY

The protein (Escherichia coli CheY) that controls the direction of flagellar rotation during bacterial chemotaxis has been shown to be phosphorylated on the aspartate 57 residue. The residue phosphorylated is present within a conserved sequence in every member of a family of bacterial regulatory proteins. The phosphorylation is transient, with a much shorter half-life than that expected of a simple acyl phosphate intermediate, indicating that the sequence and conformation of the protein is designed to achieve a rapid hydrolysis. The CheY-phosphate linkage can be reductively cleaved by sodium borohydride. High-performance tandem mass-spectrometric analysis of proteolytic peptides derived from [3H]borohydride-reduced phosphorylated CheY protein was used to identify the position of phosphorylation. Mutants with altered aspartate 57 exhibited no chemotaxis. When aspartate 13, another conserved residue, was changed, greatly reduced chemotaxis was observed, suggesting an important role for aspartate 13. The rate-determining step of chemotactic signaling is governed by the kinetics of formation and hydrolysis of the CheY protein phosphoaspartate bond. The CheY protein apparently functions as a protein phosphatase that possesses a transient covalent intermediate. Transient phosphorylation of an aspartate residue is an effective mechanism for producing a biochemical signal with a short concentration-independent half-life. The duration of the signal can be controlled by small structural elements within the phosphorylated protein.     

Grenzen der Kompensation: Energiebudgets von Ringelg?nsen (Branta b. bernicla) — die Wirkung von St?rreizen

Zusammenfassung Der Einfluß von Störreizen auf das Energiebudget von Ringelgänsen wurde im Frühjahr 1991 in zwei Vorland-Salzwiesen des Nationalparks Schleswig-Holsteinisches Wattenmeer vergleichend untersucht. Die Häufigkeit störreizbedingter Reaktionen der Gänse (Störreizhäufigkeit) wurde als Index für die anthropogene Beeinflussung der Gebiete herangezogen. Die Beeinflussung war in Westerhever mit 1,5±0,7 Reaktionen/h signifikant größer als vor dem Norderheverkoog (1,0±0,6 Reaktionen/h). Mit zunehmender Aktivitätszeit wurde in beiden Gebieten im Verlauf des Frühjahres mehr umsetzbare Energie aufgenommen. In Westerhever haben die Gänse im Vergleich zu denen vor dem Norderheverkoog mit Ausnahme des Mai jeweils mehr umsetzbare Energie aufgenommen. Die energetischen Kosten (DEE) der Gänse wurden anhand der Zeit-Energie-Budget Methode und über die gemessene Körpermasseentwicklung von Fänglingen berechnet. Beide Methoden erzielten vergleichbare Ergebnisse; die Unterschiede betrugen je nach Gebiet 0,4 bis 4,5 %. In Westerhever verzeichneten die Gänse im Vergleich zu denen vor dem Norderheverkoog im Monatsmittel höhere energetische Kosten. Aus dem budgetierten Energieüberschuß (E) wurde die theoretisch mögliche Reservestoffanlagerung (Änderung der Körpermasse) der Gänse berechnet. In Westerhever konnten die Gänse im Verlauf des Frühjahres 357 g Körperreserven anlagern. Bei den Gänsen vor dem Norderheverkoog waren es 399 g. Der Unterschied betrug 11,7 %, war aber nicht signifikant. Anhand von Fangdaten ist bekannt, daß Ringelgänse im langjährigen Mittel Körperreserven von 380 bis 400 Gramm anlagern. Die Störreizhäufigkeit des Tages bewirkte eine signifikante Veränderung der Budgetparameter DME und DEE und E auf stündlicher Basis. In Westerhever stieg die Aufnahme an umsetzbarer Energie signifikant mit steigender Störreizhäufigkeit an. Vor dem Norderheverkoog verringerte sich diese jedoch signifikant. Die Energieaufwendungen stiegen in beiden Gebieten signifikant mit steigender Reizhäufigkeit an. Der Überschuß in der Energiebilanz der Gänse in Westerhever war an Tagen mit einer großen Störreizhäufigkeit um 8,7 % gegenüber den Tagen mit einer geringen Reizhäufigkeit verringert. Aufgrund fehlender kompensatorischer Nahrungsaufnahme war der Überschuß bei den Gänsen vor dem Norderheverkoog um maximal 27,5 % vermindert. Die Gänse in Westerhever haben störreizbedingte Zeitverluste bei der Nahrungsaufnahme und erhöhte energetische Kosten durch Verhaltensänderung und eine erhöhte Nahrungsaufnahme pro Zeit kompensiert. Aufgrund physiologischer Zwänge sind sie jedoch in dem stark vom Menschen beeinflußten Gebiet an die Grenzen ihrer Kompensationsmöglichkeit angelangt. Die errechnete Reservestoffanlagerung und auch der Jungvogelanteil im nachfolgenden Herbst waren geringer als bei den Gänsen vor dem Norderheverkoog.

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Compensatory limits: energy budgets of Brent Geese,Branta b. bernicla, the influence of human disturbance

The impact of human disturbances on the energy budget of Brent Geese,Branta b. bernicla, during spring migration was investigated in two different salt marsh areas within the national park Schleswig-Holsteinisches Wattenmeer, Germany. We used the frequency of disturbance-related reactions of the birds as an indicator of the anthropogenic influencing of the sites. The disturbance frequency was significantly higher in the Westerhever salt marsh (1,5±0,7 reactions/h) than in the Norderheverkoog area (1,0±0,6 reactions/h). Monthly activity and energy budgets of the birds were calculated from March to May, based on several simultaneous day-round field observations per month. The amount of daily metabolizable energy (DME) was calculated via dropping rate and digestability of the food. The daily energy expenditure (DEE) was calculated by means of the time-energy-budget methods and by calculations, using the bodymass changes of caught birds during spring. With both methods very similar results were obtained; they varied between 0,4 to 4,5 % per areas and month. The daily energy budget (Æ E) was calculated from DEE — DME. Due to an increasing daily activity time from March to Mai the DME increased in both areas. Compared to the Norderheverkoog area the DME of the birds in Westerhever was higher in March and April, and similar in Mai. DEE increased in both areas too and was in all months higher in Westerhever than in Norderheverkoog. The calculated bodymass changes of the birds during spring was 357 g for the birds in Westerhever and 399 g for the birds in Norderheverkoog. Bodymass measures from caught birds revealed a mean mass gain of 380 to 400 g during spring. The daily disturbance frequency caused a significant change in the budget parameters DME, DDE and E on a hourly base. In Westerhever DME/h increased, while in Norderheverkoog DME/h decreased significantly with increasing disturbance frequency. At the same time DEE/h increased in both areas significantly. As a consequence E/h was reduced at days with a high disturbance frequency; in Westerhever by 8,7 % and in Norderheverkoog by 27,5%. In the disturbed area the birds compensated the time-loss during feeding and the higher energetic costs during disturbance-related flights by a change in activity pattern and by an increased food consumption per time unit. Based on physiological constraints the birds reached their compensatory abilities. The calculated bodymass gain and the proportion of young birds in the flocks was reduced in autumn.