Store-operated Ca2+ entry in astrocytes: different spatial arrangement of endoplasmic reticulum explains functional diversity in vitro and in situ

Ca(2+) signaling is the astrocyte form of excitability and the endoplasmic reticulum (ER) plays an important role as an intracellular Ca(2+) store. Since the subcellular distribution of the ER influences Ca(2+) signaling, we compared the arrangement of ER in astrocytes of hippocampus tissue and astrocytes in cell culture by electron microscopy. While the ER was usually located in close apposition to the plasma membrane in astrocytes in situ, the ER in cultured astrocytes was close to the nuclear membrane. Activation of metabotropic receptors linked to release of Ca(2+) from ER stores triggered distinct responses in cultured and in situ astrocytes. In culture, Ca(2+) signals were commonly first recorded close to the nucleus and with a delay at peripheral regions of the cells. Store-operated Ca(2+) entry (SOC) as a route to refill the Ca(2+) stores could be easily identified in cultured astrocytes as the Zn(2+)-sensitive component of the Ca(2+) signal. In contrast, such a Zn(2+)-sensitive component was not recorded in astrocytes from hippocampal slices despite of evidence for SOC. Our data indicate that both, astrocytes in situ and in vitro express SOC necessary to refill stores, but that a SOC-related signal is not recorded in the cytoplasm of astrocytes in situ since the stores are close to the plasma membrane and the refill does not affect cytoplasmic Ca(2+) levels.     

Evidence for changes in the conformational status of rat liver microsomal glucose-6-phosphate:phosphohydrolase during detergent-dependent membrane modification. Effect of p-mercuribenzoate and organomercurial agarose gel on glucose-6-phosphatase of native and detergent-modified microsomes

Comparative studies investigating influences of temperature and time of preincubation on the interactions of an organomercurial agarose gel and p-mercuribenzoate with glucose-6-phosphatase of native and Triton X-114-modified rat liver microsomes were carried out. The effect of p-mercuribenzoate on glucose 6-phosphate hydrolysis is a result of two processes, a moderate membrane perturbation connected with release of some latency and temperature- and time-dependent inhibition of the catalytic activity. Short-term preincubation with both organic mercurials at 37 degrees C is a necessary condition for the entire inhibition of the enzyme activity of native as well as of Triton X-114-modified microsomes. A binding site of the phosphohydrolase itself is accessible to p-mercuribenzoate and the phenyl mercury residue of the affinity gel from the cytoplasmic surface even in native microsomes. Kinetic analyses reveal a formally competitive mechanism of inhibition using native microsomes, but the kinetic picture changes to a noncompetitive pattern of Lineweaver-Burk plots when the inhibitor-loaded microsomes are modified optimally by Triton X-114. This behavior can be evaluated as the first convincing evidence for drastic changes of the conformational status of the phosphohydrolase during the membrane modification process. A combined conformational flexibility-substrate transport model characterizing the microsomal glucose-6-phosphatase as an integral channel-protein embedded within the hydrophobic interior of the membrane is proposed.     

Untersuchungen über Ernährung und Fruchtkörperbildung von Myxobakterien

Ohne ZusammenfassungDissertation der Mathem.-Naturwiss. Fakultät der Universität Göttingen.     

Distinct colonization waves underlie the diversification of the freshwater sculpin (Cottus gobio) in the Central European Alpine region

Ecological speciation and adaptive radiation are key processes shaping northern temperate freshwater fish diversity. Both often involve parapatric differentiation between stream and lake populations and less often, sympatric intralacustrine diversification into habitat‐ and resource‐associated ecotypes. However, few taxa have been studied, calling for studies of others to investigate the generality of these processes. Here, we test for diversification within catchments in freshwater sculpins in a network of peri‐Alpine lakes and streams. Using 8047 and 13 182 restriction site‐associated (RADseq) SNPs, respectively, we identify three deeply divergent phylogeographic lineages associated with different major European drainages. Within the Aare catchment, we observe populations from geographically distant lakes to be genetically more similar to each other than to populations from nearby streams. This pattern is consistent with two distinct colonization waves, rather than by parapatric ecological speciation after a single colonization wave. We further find two distinct depth distribution modes in three lakes of the Aare catchment, one in very shallow and one in very deep water, and significant genomewide differentiation between these in one lake. Sculpins in the Aare catchment appear to represent an early‐stage adaptive radiation involving the evolution of a lacustrine lineage distinct from parapatric stream sculpins and the repeated onset of depth‐related intralacustrine differentiation.     

Dynamic field testing of coating chemistry candidates by a rotating disk system

Quick and reliable testing is crucial for the development of new fouling release (FR) coatings. Exposure of these coatings to natural multispecies communities is essential in evaluating their efficacy. To this end, we present a rotating disk setup for dynamic field exposure. To achieve a well-defined flow on the surface of the disk, an easy to use sample mounting system was developed that provides a smooth and even surface. We related the angular velocity of the disk to the wall shear stress on the surface with a hydrodynamic model. The wall shear stress was adjusted to values previously found to be suitable to discriminate dynamic diatom attachment on different coating chemistries in the lab. The effect of the dynamic conditions was shown by comparing polystyrene slides under static and dynamic exposure. Using a set of self-assembled monolayers, the discrimination potential of the assay in a multispecies environment was demonstrated.     

Effects of fluorescent and nonfluorescent tracing methods on lymphocyte migration in vivo.

BACKGROUND: The use of fluorescent dyes to monitor in vivo cellular migration and proliferation has greatly expanded, but little is known about their potential influence on cell migration. METHODS: Adoptive transfer studies of lymphocytes labeled with various dyes were performed, and their in vivo homing was compared with that of coinjected unlabeled control cells. In addition, in vitro migration and binding studies were performed to analyze the various steps of transmigration separately. RESULTS: These data showed that the intracellular fluorescent dyes calcein acetoxymethyl ester, 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein acetoxymethyl ester, 5-chloromethylfluorescein diacetate, 5-(and-6)-carboxyfluorescein diacetate, succinimidyl ester, and fluorescein isothiocyanate affect in vivo homing of especially B lymphocytes to lymphoid organs, without any direct effect on in vitro chemotactic or adhesive activity. The only label that did not affect migration was the extracellular and nonfluorescent molecule biotin, provided that the labeling was performed at room temperature. Interestingly, by using the highly versatile congenic Ly5.1-Ly5.2 system, we also demonstrated intrinsic differences in lymphocyte migration based on allelic differences. CONCLUSIONS: Our data showed that fluorescent labeling of lymphocytes has a severe effect on their homing capacity in vivo. Labeling of cells with biotin appeared to be a good alternative for this purpose; however, if direct fluorescence is required, the negative effects on cell migration should be considered.     

Aufnahme von Trypanblau und Ferritin in die Blasenzellen des Bindegewebes von Helix pomatia und Cepaea nemoralis (Stylommatophora,Pulmonata)

Zusammenfassung Untersuchungen des Bindegewebes der stylommatophoren Pulmonaten Helix pomatia and Cepaea nemoralis zeigen, daß die Blasenzellen des Bindegewebes befähigt sind, Substanzen aus der Hämolymphe aufzunehmen. Licht- und elektronenmikroskopische Untersuchungen ausgewählter Bindegewebsbereiche, die in bestimmten Zeitintervallen nach der Injektion von Trypanblau oder Ferritin in die Körperhöhle präpariert wurden (Stufenuntersuchungen), ergaben, daß — mit Ausnahme einiger Blutzellen — ausschließlich die Blasenzellen die injizierten Substanzen aufgenommen hatten. Weder andere Bindegewebszellen noch die Zellen von Organen (u.a. Gonaden, Schlundringganglien), die von Bindegewebe umgeben werden, enthielten diese Substanzen oder hatten sie akkumuliert. Die elektronenmikroskopischen Aufnahmen zeigen, daß die Aufnahme von Ferritin in die Blasenzellen wahrscheinlich durch Pinocytose erfolgt.

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Uptake of trypan blue and ferritin into the globular cells of the connective tissue of Helix pomatia and Cepaea nemoralis (Stylommatophora, Pulmonata)

Summary Investigations of the connective tissue of the stylommatophoran pulmonates Helix pomatia and Cepaea nemoralis have demonstrated, that so-called globular cells have the capability for the uptake of substances out of the hemolymph. Light- and electron-microscopic investigations of pieces of connective tissues fixed at different intervals after the injection of Trypane Blue or Ferritin into the cavity of the body give striking evidence for the uptake of these substances exclusively into the globular cells (with the exception of some blood cells). Neither other connective tissue cells nor cells of the organs (e.g. gonads, central nervous ganglions) surrounded by connective tissue incorporate or accumulate the injected substances. The uptake of Ferritin into the cytoplasm of the globular cells, normally filled with a high amount of glycogen, takes place by pinocytosis.