The essential role of sodium bioenergetics and ATP homeostasis in the developmental transitions of a cyanobacterium

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An improved method for electron microscopic observation of cerebrospinal fluid cells

Human cerebrospinal fluid (CSF) cells were entrapped between two bovine serum albumin cylinders linked by glutaraldehyde using microhematocrit tubes. The "sandwiched" specimen could be processed in further preparatory steps like tissue for routine electron microscopy. This rapid procedure resulted in adequate cell preservation for transmission electron microscopy without substantial loss of CSF cells.     

Comparison of two prepill cortisol concentrations in dogs with hypercortisolism treated with trilostane

Background

The ideal method for monitoring trilostane therapy in dogs with hypercortisolism is still open to debate. Recently, determination of the pre-trilostane (prepill) cortisol concentration has been proposed to be more repeatable than either post-trilostane or post-ACTH cortisol. The aim of this study was to compare two prepill cortisol concentrations in dogs with hypercortisolism during trilostane therapy. Sixteen client-owned dogs with naturally occurring hypercortisolism were prospectively included and cortisol concentrations were measured twice, 1?h apart, before the morning trilostane dose (prepill 1 and 2 cortisol).

Results

A total of 47 prepill cortisol measurement pairs were included. Compared to prepill 1, prepill 2 cortisol was higher in 15, equal in 8 and lower in 24 pairs. Group agreement between prepill 1 and 2 cortisol was 70% (moderate agreement - weighted kappa 0.55). In 30% of the pairs, group assignment was discrepant, implying a different therapeutic decision. In some dogs certain circumstances (e.g. excessive barking, difficulties during blood collection, excitement at arrival) were identified as potential factors explaining the discrepancy between prepill 1 and 2 cortisol measurements.

Conclusions

In a substantial number of dogs treated with trilostane, the two prepill cortisol concentrations differed. Part of this difference might be ascribable to stressful events during test performance. When using prepill cortisol measurements to monitor trilostane therapy, recording of any incident during handling that might affect cortisol release might be helpful to make a reliable decision about a trilostane dose adaptation.

     

Molecular Requirements for Bi-directional Movement of Phagosomes Along Microtubules

Microtubules facilitate the maturation of phagosomes by favoring their interactions with endocytic compartments. Here, we show that phagosomes move within cells along tracks of several microns centrifugally and centripetally in a pH- and microtubuledependent manner. Phagosome movement was reconstituted in vitro and required energy, cytosol and membrane proteins of this organelle. The activity or presence of these phagosome proteins was regulated as the organelle matured, with “late” phagosomes moving threefold more frequently than “early” ones. The majority of moving phagosomes were minus-end directed; the remainder moved towards microtubule plus-ends and a small subset moved bi-directionally. Minus-end movement showed pharmacological characteristics expected for dyneins, was inhibited by immunodepletion of cytoplasmic dynein and could be restored by addition of cytoplasmic dynein. Plus-end movement displayed pharmacological properties of kinesin, was inhibited partially by immunodepletion of kinesin and fully by addition of an anti-kinesin IgG. Immunodepletion of dynactin, a dynein-activating complex, inhibited only minus-end directed motility. Evidence is provided for a dynactin-associated kinase required for dyneinmediated vesicle transport. Movement in both directions was inhibited by peptide fragments from kinectin (a putative kinesin membrane receptor), derived from the region to which a motility-blocking antibody binds. Polypeptide subunits from these microtubule-based motility factors were detected on phagosomes by immunoblotting or immunoelectron microscopy. This is the first study using a single in vitro system that describes the roles played by kinesin, kinectin, cytoplasmic dynein, and dynactin in the microtubule-mediated movement of a purified membrane organelle.     

Overexpression of the Dynamitin (p50) Subunit of the Dynactin Complex Disrupts Dynein-dependent Maintenance of Membrane Organelle Distribution

Dynactin is a multisubunit complex that plays an accessory role in cytoplasmic dynein function. Overexpression in mammalian cells of one dynactin subunit, dynamitin, disrupts the complex, resulting in dissociation of cytoplasmic dynein from prometaphase kinetochores, with consequent perturbation of mitosis (Echeverri, C.J., B.M. Paschal, K.T. Vaughan, and R.B. Vallee. 1996. J. Cell Biol. 132:617–634). Based on these results, dynactin was proposed to play a role in linking cytoplasmic dynein to kinetochores and, potentially, to membrane organelles. The current study reports on the dynamitin interphase phenotype. In dynamitin-overexpressing cells, early endosomes (labeled with antitransferrin receptor), as well as late endosomes and lysosomes (labeled with anti–lysosome-associated membrane protein-1 [LAMP-1]), were redistributed to the cell periphery. This redistribution was disrupted by nocodazole, implicating an underlying plus end–directed microtubule motor activity. The Golgi stack, monitored using sialyltransferase, galactosyltransferase, and N-acetylglucosaminyltransferase I, was dramatically disrupted into scattered structures that colocalized with components of the intermediate compartment (ERGIC-53 and ERD-2). The disrupted Golgi elements were revealed by EM to represent short stacks similar to those formed by microtubule-depolymerizing agents. Golgi-to-ER traffic of stack markers induced by brefeldin A was not inhibited by dynamitin overexpression. Time-lapse observations of dynamitin-overexpressing cells recovering from brefeldin A treatment revealed that the scattered Golgi elements do not undergo microtubule-based transport as seen in control cells, but rather, remain stationary at or near their ER exit sites. These results indicate that dynactin is specifically required for ongoing centripetal movement of endocytic organelles and components of the intermediate compartment. Results similar to those of dynamitin overexpression were obtained by microinjection with antidynein intermediate chain antibody, consistent with a role for dynactin in mediating interactions of cytoplasmic dynein with specific membrane organelles. These results suggest that dynamitin plays a pivotal role in regulating organelle movement at the level of motor–cargo binding.     

Synthesis and characterization of 8-methoxy-2'- deoxyadenosine-containing oligonucleotides to probe the syn glycosidic conformation of 2'-deoxyadenosine within DNA.

The synthesis of 8-methoxy-2'-deoxyadenosine (moA) protected at N6 as an N,N-dimethylformamidine derivative and incorporation of the modified nucleoside into oligodeoxynucleotides via the phosphoramidite method are described. UV thermal denaturation studies were conducted on duplexes containing moA:G, moA:C and moA:T base pairs to determine the thermodynamic stability of duplexes containing moA relative to their adenosine (A)-containing counterparts. In the case of moA:G base pairs the effect of moA substitution is sequence dependent. In A:G mismatch-containing sequences, which have been shown by structural characterization to have a syn conformational preference at the glycosidic bond of A, moA substitution results in stabilization of the duplex. In contrast, in sequences where the A in the A:G mismatch has been shown to prefer the anti conformation moA substitution is destabilizing to the duplex. Thus moA may be a useful probe for investigating the conformational preferences of the N-glycosidic bond of adenosine within DNA. In addition, moA nucleoside is more resistant to acid-catalyzed depurination than previously described 8-bromo-2'-deoxyadenosine, allowing for facile incorporation into oligonucleotides via automated solid phase DNA synthesis.     

Bau und optische Leistungen des Komplexauges der StielaugenfliegeCyrtodiopsis dalmanni Wiedemann

Zusammenfassung Die großen Sehfelder der Komplexaugen der malayischen StielaugenfliegeCyrtodiopsis dalmanni überschneiden sich 3–4 mm vor den Augen. Eine Entfernungswahrnehmung von Objekten, die sich in den Sehfeldern beider Facettenaugen befinden, ist deshalb sehr wahrscheinlich.Interferometrische Messungen amCyrtodiopsis-Auge führten zu gleichen Brechungsquotienten für die Ommatidienstrukturen wie beiCalliphora erythrocephala. Die geometrischoptischen Berechnungen ergaben für den dioptrischen Apparat des Ommatidiums imCyrtodiopsis-Auge eine objektseitige Brennweite von 29 m. Hieraus und aus den Durchmessern der Rhabdomeren-Kappen — sie liegen in der Brennebene — resultieren die Öffnungswinkel von2,5° für die Rhabdomere 1–6 und von1,6° für das 7. Rhabdomer. Diese Öffnungswinkel sind kleiner als die der Ommen, die bei8,8° liegen. Im einzelnen Ommatidium kann eine höhere Auflösung erzielt werden, da die Beugungsscheibe jeweils nur eine Kappe bedeckt.

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Morphology and physiology of the compound eye of the stalked-eye flyCyrtodiopsis dalmanni Wiedemann

Summary The large visual fields of the compound eyes of the Malayan flyCyrtodiopsis dalmanni overlap 3–4 mm in front of the eyes. Depth perception of objects placed within the visual fields of both eyes is therefore very probable. Interferometrical studies on theCyrtodiopsis eye indicated that refractive indices of the ommatidial structures are the same as in the blowfly. The focal length of the dioptric apparatus of the stalked-eye fly ommatidium was found to be 29 m on the object side. The angles of aperture calculated from this focal length and from the diameters of the corresponding rhabdomeric prolongations—located in the focal plane-are 2.5° for the rhabdomeres 1–6, and 1.6° for the rhabdomere 7. These opening angles are smaller than those of the ommatidia, which are about 8.8°. Calculations based on the diffraction theory lead to the conclusion that higher resolution is obtained in the single ommatidium because the airy disc covers only one rhabdomeric prolongation.

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Mit Unterstützung der Deutschen Forschungsgemeinschaft.