Friedhelm Achenbach

Geotactic behavior of the acellular myxomycete Physarum polycephalum

Abstract Geotaxis in Physarum polycephalum was tested using plasmodia migrating on vertical agar plates. 65.8% of all plasmodia showed positive geotaxis, 29.4% were indifferent, and 13% migrated opposite to the gravity vector. Single plasmodia on a vertical agar plate could repeatedly be forced to change the direction of migration by rotating the agar plate stepwise 90 ° on a horizontal axis under continuous vertical orientation.

Calcium accumulation in plasmodia of Physarum polycephalum

Abstract The total concentration of calcium in the endoplasm of plasmodia of Physarum polycephalum was measured using a calcein fluorescence-quenching technique. The calcium concentration of the endoplasm increases with the time of cultivation on different substrates under culture conditions frequently used for routine experiments. Calcium accumulation within endo- and ectoplasm as well as in microplasmodia is most pronounced when plasmodia are starved and illuminated. Starvation and illumination lead to calcium concentrations frequently exceeding 200 mM in the case of macroplasmodia and 20 mM in the case of microplasmodia.

Oscillating contractions in protoplasmic strands of physarum: Effects of externally applied ouabain, sodium-, potassium- and calcium-ions

1. Concentrations of 0.1-10 mM Ouabain do not affect oscillating contractions, when applied externally in physiological solutions. 2. Ouabain has no effect, when applied in solutions of increased sodium- (100 mM) and lowered potassium-concentration (0.1 or zero mM). 3. De novo generation of oscillating contractions in protoplasmic drops is not suppressed in Ouabain-solutions. 4. Biochemical studies of Na-K-ATPase did not show any Ouabain sensitive ATPase-activity. 5. Reaction of veins on high concentrations (300 mosm) is discussed as to be a physical effect. 6. It is concluded that no Na-K-ATPase is engaged in triggering oscillating contraction automaticity.

Lateral apertures as passage-ways between ectoplasm and endoplasm in plasmodial strands of Physarum.

Abstract Plasmodial strands of Physarum polycephalum are differentiated into an ectoplasmic tube and an endoplasmic core. The border of the ectoplasmic tube represents a circular arranged system of plasmalemma invaginations containing apertures which allow a continuous exchange of endoplasm and ectoplasm during the oscillating contraction-relaxation cycles. The apertures of thick plasmodial strands are described using phase-microscopy of semithin sections. Their physiological importance for signal transmission, endoplasmic mass transport and thereby plasmodial locomotion is discussed.

Calcium binding sites in plasmodia of Physarum polycephalum as revealed by the pyroantimonate technique.

Plasmodia of the acellular slime mold, Physarum polycephalum, were treated with an osmium tetroxide fixative containing potassium pyroantimonate to precipitate calcium and thereby localize calcium binding sites and sites of increased calcium concentration. Dense calcium pyroantimonate precipitates were detected within the nucleoli. The distribution of these precipitates during interphase and mitosis coincides with the distribution of the unique minichromosomes in Physarum, i.e., the numerous short pieces of extrachromosomal nucleolar chromatin containing segments of amplified DNA coding for ribosomal RNA. Calcium pyroantimonate precipitates were present as frequent dense granules in the mitochondrial matrix and as fine precipitates in the mitochondrial nucleoid. Large calcium-containing precipitates were seen within cytoplasmic vacuoles, confirming reports by others. In addition, we have identified calcium binding sites along the cytoplasmic surface of the plasma membrane. The distribution of calcium within the plasmodium is discussed in relation to the assembly of the mitotic spindle and the regulation of cell motility.

PROTEIN KINASE C AFFECTS REFORMATION OF ENDOTHELIAL JUNCTIONS IN XENOPUS XTH-2 CELLS

Endothelial cells can reversibly be forced to suppress the formation of endothelial junctions (EJ) by cultivation in a low calcium medium. The authors localized vinculin and cadherin as marker proteins of EJ and actin as a cytoskeletal component by fluorescence microscopy, and used this cell model to study the reformation of endothelial junctions under conditions of activation and inhibition of protein kinase C (PKC). Inhibition of PKC by H‐7 leads to an acceleration of EJ reformation, while constitutive activation by TPA inhibits the reformation process.

Progress in plasmodial differentiation improves regularity of oscillating contractions in Physarum polycephalum

Based on the knowledge about subcellular morphogenetic processes in the acellular slime mold Physarum polycephalum, we hypothesized that during differentiation of undifferentiated endoplasm to the highly differentiated complex structure of the contractile apparatus of this organism, the regularity of oscillating contractions must improve.

Quantification and preservation of lectin binding by isolated cardiomyocytes

During preparation of cells for experimentation a considerable amount of bound substance is lost. Our aim was to develop a protocol which retained lectin binding to an extent similar to living cells. This procedure would use fixation procedures suited for fluorescent lectin conjugates and gold-conjugates to be visualized by light- and electron microscopy, respectively. We tested glutaraldehyde and paraformaldehyde in different concentrations before and after lectin binding, different buffers and divalent cations, as additives, to determine the effects on preservation of lectin binding. Lectin binding was visualized and semiquantitatively evaluated by image analysis in the light microscope after silver enhancement of lectin-gold conjugates and by using tetramethyl rhodaminyl isothiocyanate (TRITC)-conjugated lectins. Preservation of lectin binding was best visualized with fluorescent lectin conjugates, whereas during silver enhancement procedures of gold-conjugated lectins, a considerable amount of bound lectins was lost. In general, lectin binding to living cells followed by fixation is superior to fixation before lectin binding. Unfavourable combinations of fixatives and buffers can cause a loss of more than 90% bound lectin. In our experiments with freshly isolated guinea pig cardiomyocytes, lectin binding was best when we used Na-cacodylate buffer with glutaraldehyde fixation (0.1%) after binding of lectins to the living cells.

Effect of magnesium on the contractile behavior of actomyosin aggregates isolated from Physarum plasmodia

The dependence on calcium concentration of the contractile behavior of actomyosin isolated from Physarum plasmodia according to Kohama & Kendrick-Jones (1986) was investigated under different magnesium conditions. The inhibitory calcium sensitivity is reduced at magnesium concentrations above or below 1 mM, i.e., contraction of actomyosin aggregates is most effectively inhibited in the presence of 1 mM calcium in combination with physiological magnesium concentrations. In the absence of calcium reactivation optimum is obtained at 8.5 mM Mg2+.