Some Effects of Calcium Ions on the Action Potential of Single Nodes of Ranvier

Action potentials of single frog nerve fibers were recorded with the air-gap method in "low Ca" (0.26 mM) and "high Ca" (4.2 mM) solutions and compared to spikes in normal Ringer''s (1.05 mM Ca). On increasing (Ca)_o the action potentials became shorter, the "knee" during the falling phase as well as the threshold for abolition moved to internal potentials more positive, and the spike recovery during the relative refractory period was faster. Outward current pulses applied during an action potential affected its configuration more in low Ca than in high Ca. The onset of the delayed rectification (in the absence of Na) was found faster in high Ga. After-potentials during anelectrotonus declined more rapidly in high Ca than in low Ca. The results are compared primarily with the voltage-clamp analysis of Ca effects on squid axons and satisfactory qualitative agreement is reached.     

STUDIES ON THE FIXATION OF ARTIFICIAL AND BACTERIAL DNA PLASMS FOR THE ELECTRON MICROSCOPY OF THIN SECTIONS

The process of fixation of DNA-containing plasms is investigated by macroscopical and electron microscopical observations on solutions of DNA, nucleohistones, as well as on bacterial nuclei. The following treatments were found to produce a gelation of a solution of DNA or nucleohistones: (a) OsO₄ fixation at pH 6 in the presence of amino acids (tryptone) and Ca⁺⁺. (b) Exposure to aqueous solutions of uranyl acetate. (c) Exposure to aqueous solutions of indium chloride. Observed in the electron microscope, these gels show a fine fibrillar material. From experiments in which solutions of DNA or nucleohistones are mixed with bacteria and treated together, it is concluded that the behavior of the bacterial nucleoplasm is similar to that of the DNA solutions. The appearance of birefringence indicates that uranyl acetate and indium chloride produce an orientation of the molecules of a DNA solution during gelation. Bacterial chromosomes fixed by these agents also show a certain order, while those fixed by the OsO₄-amino acid-Ca⁺⁺ formula do not. Whether or not the order can be considered to be artificial is discussed, and a tentative conclusion is presented: (a) Uranyl acetate may induce artificial order. (b) Fixatives which do not gel DNA probably result in the grossest artifacts. (c) OsO₄ fixation at pH 6 in the presence of amino acids (tryptone) and Ca⁺⁺ may give the most accurate preservation of the in vivo disposition of DNA (RK⁺ fixation).     

Stephanoscyphus Allman (Scyphozoa Coronatae), ein rezenter Vertreter der Conulata?

The scyphopolypStephanoscyphus Allman 1874 represents the polyp generation of the scyphomedusan order Coronatae. The benthonic polyp is known to occur with several species on the continental shelves and in greater depths of the oceans. Participating in the International Indian Ocean Expedition 1964 to 1965 on board of the German research vessel «Meteor», the author was able to collect living specimens. For the first time, it was possible to rear these polyps in laboratory cultures.Stephanoscyphus differs from all other living scyphopolyps by the possession of a firm periderm tube enclosing completely the soft body. It is this primitive feature whichStephanoscyphus has in common with the Conulata being the ancestors of the recent Scyphozoa. The characteristics found in a detailed investigation of the periderm tube ofStephanoscyphus conform well with those found in the tubes of the Conulata except for the closure of the aperture by triangular flaps which are absent inStephanoscyphus. The soft body contains primitive features as well. Hence from the existence, morphology and life history it must be concluded that the type of organization which the Conulata exhibited has survived inStephanoscyphus.     

Comparative Study of Cultured Burkitt Tumor Cells by Immunofluorescence, Autoradiography, and Electron Microscopy

Cultured Burkitt cells were examined by immunofluorescence, autoradiography, and electron microscopy in an effort to identify the stainable cells with those harboring herpes-type virus particles. Immediately after a 2-hr pulse of (3)H-thymidine, from 30 to 60% of the cells revealed heavy nuclear labeling. In most cases the grains were evenly dispersed, but in about 3 to 5% the grains showed a focal distribution and occasionally they extended into the cytoplasm. Such nuclear foci were rarely seen at 8 hr after the pulse. When the analysis was restricted to preselected immunofluorescent cells, up to 80% showed label at 8 hr and cytoplasmic grains were prominent. To reduce cellular deoxyribonucleic acid (DNA) synthesis, cells were X-irradiated with 3,000 to 6,000 R, and the isotope pulse was applied 1, 4, or 7 days later. Whereas the total number of labeled cells decreased in roughly twofold steps at the respective intervals (from 40 to 10%), the incorporation of (3)H-thymidine into fluorescent cells was not affected by X irradiation. In each series, about 70% of the fluorescent cells contained label when they were examined at 24 and 48 hr after the pulse, whereas at 8 and 72 hr fewer were positive. At the earlier intervals, unlabeled fluorescent cells most likely represented cells which had completed viral DNA synthesis prior to the pulse; at the later intervals, unlabeled fluorescent cells were probably cells which commenced viral replication after the pulse. These data support the conclusion that the immunofluorescent cells are the ones which harbor virus, and also confirm the expectation that the virus is a DNA virus from a member of the herpes group. This conclusion was firmly established by sectioning and electron microscopic examination of individual fluorescent cells, all of which contained numerous virus particles, whereas the nonstained cells prepared in a similar manner were free of them.     

Electron microscopic observations on isolated mitochondrial membranes,prepared by various histological methods

Summary The pictures of isolated mitochondrial membranes, as seen on the electron-microscope, depend very much on the method of specimen preparation. Subunits of linear dimensions of about 25 m, (electron transport particles) are observed in carbon-replicas of the membranes and in specimens treated with trypsin or pepsin (0.02% for 30 mins) and shadowed with platinum. A three-layered structure of the unit membrane is seen in sections of specimens fixed with osmium tetroxide or formalin followed by post-fixation with osmium tetroxide. But fixation with potassium permanganate or with formalin, followed by post-fixation with potassium permanganate reveals an electron-dense globular structural element in the unit membrane. An electron-transparent ultrastructural element of the unit membrane is observed after treatment with trypsin (0.2% for 5 mins) and fixation with osmium tetroxide. Unsectioned specimens treated with 0.02% trypsin for 30 mins show a honeycomb-like structure of the membrane. Thus, part of the results appear to support the concept of a mosaic-like structure of the unit membrane, whereas other results are in agreement with the classical concept of a three-layered structure.The authors wish to express their gratitude to Dr. Sina Rosenthal, Department of Physiological Chemistry, Humboldt University, Berlin, who prepared the isolated membranes, to Mr. E. Fischer, Head Technician of the Department of Electron Microscopy, Greifswald University, who took most of the electron micrographs, to Mr. G. Bartsch, Department of Electron Microscopy, Greifswald University, and especially to Prof. W. Bargmann and to Doz. E. Lindner, Department of Anatomy, Kiel University, for many valuable suggestions.     

Stephanoscyphus (Scyphozoa,Coronatae) und seine direkte Abstammung von den fossilen Conulata

The scyphopolypStephanoscyphus Allman 1874 represents the polyp generation of the scyphomedusan order Coronatae. Though this polyp has been known for more than a hundred years its general morphology, systematics, and evolution have been inadequately described. Participating in the International Indian Ocean Expedition, 1964 to 1965, on board of the German research vessel “Meteor”, the author was able to collect a sufficient supply of livingStephanoscyphus off the coasts of South Arabia and East Africa. For the first time, it was possible to rear these polyps in laboratory cultures. A thorough investigation of morphology, developmental history and behaviour based on longterm observations of the living polyps gave clear indications thatStephanoscyphus directly descended from the fossil group of Conulata, the scyphozoan nature of which has been affirmed byKiderlen (1937) andKnight (1937). The main feature whichStephanoscyphus has in common with the Conulata is the possession of a periderm tube. The characteristics found in a detailed investigation of the periderm tube conform well with those found in the periderm of the Conulata except for the closure of the aperture by triangular flaps which are absent inStephanoscyphus. The soft body contains primitive features as well. Hence it must be concluded finally that the type of organization which the fossil ancestors exhibited has survived inStephanoscyphus and that the Coronatae represent the most basic group of all living Scyphozoa. On the other hand, the results give strong support for the scyphozoan nature of the Conulata, the organization and life history of which have been elucidated by the observations of the living representatives ofStephanoscyphus.     

Fatty Acid Composition of the Peribacteroid Membrane and the ER in Nodules of Glycine max Varies after Infection by Different Strains of the Microsymbiont Bradyrhizobiumjaponicum

The fatty acid composition of ER, Golgi and peribacteroid membrane (PBM) from root nodules formed on Glycine max after infection with different strains of Bradyrhizobium japonicum has been analysed by gas chromatography. In each plant-microsymbiont combination the fatty acid composition (FAC) of the PBM is distinct from ER and Golgi. The similarity between ER and PBM fatty acid composition is significantly stronger than between Golgi and PBM. In addition the fatty acid composition of all membrane systems in nodules is affected by the microsymbiont strain. A comparison of four strains of Bradyrhizobium japonicum grown in agar surface culture and isolated as the symbiotic bacteroids reveals a decrease in oleic acid during bacteroid differentiation.     

Luciferin derivatives in bioluminescence-enhanced enzyme immunoassays

Ultrasensitive bioluminescence immunoassays for the determination of peptides and proteins (illustrated with human urinary kallikrein, bradykinin and the determination of human urinary kallikrein antibody titres) have been developed. The usable ranges of the standard curves are from 5 pg to 5000 pg per litre. The relative intra-assay coefficients of variation of the tests were between 2% and 6%, and the inter-assay coefficients of variation between 4% and 12%.