Ulf-Rüdiger Heinrich

Ultrastructural and electron spectroscopic analyses of cyanobacteria and bacteria

The extracellular sheath material and some intracellular cell components of cyanobacteria and phosphate‐accumulating sewage bacteria were analysed by electron spectroscopic imaging (ESI) and electron energy‐loss spectroscopy (EELS). The specimens were embedded in water‐soluble Nanoplast resin without any previous fixation and ultrathin sections were examined in a Zeiss CEM 902 microscope. A high sulphur content was detected in the inner sheath of the cyanobacterium Gloeothece. The elemental composition of some cell components and inclusion bodies, such as carboxysomes and cyanophycin, was determined by ESI and EELS. In addition, the phosphate content in specific granules of phosphate‐accumulating sewage bacteria was estimated by EELS and nuclear magnetic resonance spectroscopy.

Elemental composition of pyroantimonate precipitates analysed by electron spectroscopic imaging (ESI) and electron energy-loss spectroscopy (EELS) in vitellogenic ovarian follicles of Drosophila

Ca2+ was precipitated with potassium antimonate in vitellogenic follicles of the fruit fly Drosophila melanogaster and the distribution of the precipitates formed was studied by electron microscopy. The microvilli of the oolemma in mid‐ and late vitellogenic follicles were lined with precipitates. The chemical composition of the precipitates was analysed by electron spectroscopic imaging (ESI). The images produced by inelastically scattered electrons at specific ionization edges were compared, and the nonspecific background signals were subtracted by an image processing system. The presence of Ca2+, antimony and oxygen in the precipitates formed could be demonstrated. The elemental composition of the precipitates and of yolk spheres was also analysed by electron energy‐loss spectroscopy (EELS). With respect to the precipitates, signals at the calcium L2,3‐edge, the oxygen K‐edge and the antimony M4,5‐edge were recorded without deconvolution and background subtraction. The yolk spheres, which were free of precipitates, gave the characteristic signal of the nitrogen K‐edge. The applied techniques combine good ultrastructural resolution with the possibility of analysing the elemental composition of histochemical reaction products and cellular structures.

Progress in electron microscopic diagnostics: semi-quantitative determination of precipitable calcium in different cell types of the organ of Corti in the guinea-pig

Potassium antimonate was used to precipitate calcium in the cochlea of the guinea‐pig. The distribution of the calcium antimonate precipitates was analysed by electron microscopy. The precipitate density was determined in different cell types in the organ of Corti by counting the number of calcium binding sites in a 10‐μm2 area. The size of the precipitates varied considerably, and thus the relative amount of the precipitable calcium was estimated only semi‐quantitatively. As the prominent carbon signal is superimposed over the nearby small Ca2+ ‐edge signals, the combined signal of the antimony M4,5‐edge and the oxygen K‐edge of the calcium antimonate salt formed was chosen for the semi‐quantitative estimation. Images of the inelastically scattered electrons of the precipitates at ΔE = 570 eV were recorded by electron spectroscopic imaging. The area covered by the calcium precipitates within a given cell type was determined in different ultrathin sections of the same organ of Corti by an image processing system.

Calcium-binding sites in the inner ear after pure-tone stimulation

SummaryFive guinea pigs were exposed to an interrupted 90 dB SPL pure tone of 3.2 kHz for a total application time of 5 min. Following sound application all animals were decapitated and the cochleae were removed. After that, calcium-binding sites were located by the potassium pyroantimonate precipitation method. Another three animals served as control animals and did not receive the sound treatment. Findings confirmed previous studies showing the spatial arrangements of precipitate rich regions in the inner ears two acellular structures (the basilar membrane and tectorial membrane) and the two cellular structures (the inner hair cells and Huschkes teeth). By using semiquantitative evaluation we found a relative diminution of precipitable calcium in inner hair cells and in Huschkes teeth of sound-exposed animals when compared to untreated animals.

Localisation of potassium pumps in Drosophila ovarian follicles

It has been shown previously that, in Drosophila oogenesis, potassium ions are important for bioelectric phenomena as well as for other physiological and developmental processes. In the present study we determined the spatial distribution and activity of the Na+,K+)-pump and of ouabain-insensitive K+ pumps in plasma membranes of vitellogenic ovarian follicles (stage 10). We used the light microscopic anthroylouabain method as well as the cytochemical lead and cerium precipitation methods in combination with electron spectroscopic imaging (ESI) and electron energy-loss spectroscopy (EELS). (Na+,K+)-ATPase activity was predominantly observed on the oolemma as well as on the membranes of the columnar follicle cells covering the oocyte, whereas on the membranes of the nurse cells and of the squamous follicle cells covering the nurse cells the activity was very low. The highest activity of the (Na+,K+)-pump was found at the anterior and posterior ends of the oocyte, and this on the oolemma as well as on the membranes of the follicle cells located here. Strong activity of the ouabain-insensitive K+-pumps was observed on most of the oolemma (except at the anterior of the oocyte) and on the membranes of some nurse cells located next to the oocyte, whereas less activity was found on the other nurse cell membranes and on the membranes of all follicle cells. The suitability of the different methods used for determining the localisation as well as the activity of K+-pumps is discussed. We further discuss the nature of the ouabain-insensitive K+ pumps and the relevance of the observed distribution of K+-pumps for K+ uptake, extrafollicular ionic current flow, intercellular signalling and other developmental processes in Drosophila oogenesis.

Localization of Ca2+ ‐stores and tissue compartments with a Ca2+ ‐binding capacity in the organ of Corti of the guinea‐pig by electron energy‐loss spectroscopy

The addition of 10 mM CaCl2 to glutaraldehyde fixative leads to the formation of small electron‐dense deposits in the organ of Corti of the guinea‐pig. These precipitates are mainly attached to cell membranes in contact with different extracellular lymphatic fluids. A higher number of precipitates is localized in the acellular parts of tectorial and basilar membrane. Electron energy‐loss spectroscopy (EELS) was used to determine the elemental composition of the deposits formed. The spectra showed a prominent signal at the Ca2+ L2,3 ionization edge. Oxygen could also be detected in all the precipitates analysed. EELS analysis of mitochondria of the inner and outer hair cells after conventional fixation (glutaraldehyde followed by post‐fixation in OsO4) revealed a small but significant calcium signal.