Yu. S. Chentsov

Mitochondrial framework (reticulum mitochondriale) in rat diaphragm muscle.

Reconstitution of rat diaphragm mitochondria has been carried out with the use of the serial section technique. It is shown that mitochondrial material is organized as networks transpiercing the I band regions of the muscle near the Z-discs. Each network forms tubules, oriented perpendicular to its plane, and branches, connecting the network with mitochondrial clusters in the fiber periphery. Such a system, defined as mitochondrial reticulum, is found to be characteristic of the diaphragm of adult animals. It is absent in the diaphragm of rat embryos and newborn rats. The junctions of the branches of mitochondrial reticulum are described. In the junction site, the outer membranes of two mitochondrial branches are in contact, and spaces between outer and inner membranes are filled with an osmiophilic substance. No junctions were found in the embryos and in newborn animals whose diaphragm contains single, elliptical or worm-like mitochondria. The hypothesis is put forward that the mitochondrial reticulum serves as a system for transport of energy, oxygen and fatty acid residues along mitochondrial membranes over distances commensurable with the muscle fiber diameter.

Intermitochondrial contacts in myocardiocytes

Intermitochondrial contacts are described in rat myocardiocytes. They are characterized by a maximal drawing together (or maybe fusion) of two outer mitochondrial membranes of neighbouring mitochondria, accompanied by a strong local increase in the electron density of both of the outer and inner mitochondrial membranes as well as of the intermembrane spaces. Mitochondrial contacts were shown to connect all the single mitochondria of myocardiocyte. Contacts of the mitochondrial membranes with the plasma membrane in the nexus zone are also found. Mitochondrial contacts are absent from myocardiocytes of 3-day-old rats.

Chromonema and chromomere

A study of ultrathin sections of normal Chinese hamster cells and cells treated with decreasing concentrations of bivalent cations (Ca2+ and Mg2+) in situ revealed several discrete levels of compaction of DNA-nucleoprotein (DNP) fibrils in mitotic chromosomes and the chromatin of interphase nuclei. At concentrations ranging from 3 mM CaCl2 and 1 mM MgCl2 to ten times less, the chromosomes are found to contain fibrous elements (chromonemata) about 100 nm in diameter. As Ca2+ concentration is gradually decreased to 0.2–0.1 mM, the chromosomes decondense into a number of discrete chromatin structures, the chromomeres. As decondensation proceeds, these chromomeres acquire a rosettelike structure with DNP fibrils radiating from an electron-dense core. Upon complete decondensation of chromosomes, individual chromomeres persist only in the centromeric regions. The following levels of DNP compaction in mitotic chromosomes are suggested: a 10-nm nucleosomal fibril, a 25-nm nucleomeric fibril, and the chromonema, a fibrous structure, about 100 nm in diameter, composed of chromomeres. Interphase nuclei also contain structures which are morphologically similar to the chromomeres of mitotic chromosomes.

The ultrastructure of centriole in mammalian tissue culture cells

Structural polarity of centriole has been shown by analyzing serial sections of centrioles in the tissue culture cells of mouse, man, pig and Chinese hamster. Its major component is nine microtubule triplets. The inclination of the triplets towards the radius at the proximal end of the centriole is smaller than at the distal one. The internal tubule of the triplet has a smaller diameter than the middle and external ones; The triplets are bound by links of various nature all over their length. In the middle part, in the centriole lumen there is an amorphous hub; in the distal part, a thin fibre that is helically wound. In the proximal part, there are bases along the triplets, and handles stretch from the internal tubules. In the middle and distal parts, there are accumulations of an electron dense substance along the middle tubules. At the distal end, the centriole lumen is filled with an amorphous substance, whereas the proximal end is free from it. From outside, appendages are attached to the triplets at the distal end. The centriole structure is identical in all the cell types studied, except for those of Chinese hamster.

The effect of oncotic pressure on heart muscle mitochondria

Abstract The role of oncotic pressure ( i.e. pressure created by non-penetrants of high molecular weight) in structural responses of mitochondria has been studied. It has been found that treatment of beef of rabbit heart mitochondria by a synthetic non-penetrant of high molecular weight, polyvinyl pyrrolidone, induces a decrease in the intermembrane (intracristal) space and an increase in the matrix space of mitochondria. As a result, the appearance of the in vitro mitochondria proves to be similar to that of the in situ ones. If a Waring blender is used to homogenize the tissue, only a portion of the mitochondria respond to polyvinyl pyrrolidone. If a glass-Teflon homogenizer is used instead all the mitochondria prove responsive. The addition of 0.5 mM polyvinyl pyrrolidone is found to be sufficient for the effect to be observable. In the presence of polyvinyl pyrrolidone, energy-dependent changes in mitochondrial structure can be demonstrated. The increase in matrix space by polyvinyl pyrrolidone treatment enlarges even more when an energy source, a penetrating weak acid and a penetrating cation are added. The size of the matrix increases in the following order: (1) de-energized mitochondria without polyvinyl pyrrolidone, (2) de-energized + polyvinyl pyrrolidone, (3) energized + polyvinyl pyrrolidone, (4) as (3) + phosphate (“twisted” configuration of cristae), (5) as (3) + phosphate + Ca 2+ . Structural changes resembling those indicated in points (2)–(5) are shown for mitochondria in the tissue, when pieces of rat diaphragm muscle treated with an uncoupler, phosphate, and Ca 2+ were studied in conditions excluding anaerobiosis. The effect of polyvinyl pyrrolidone is suggested to be due to it balancing the oncotic pressure created by high molecular weight compounds dissolved in the intermembrane water, which are incapable of penetrating the outer mitochondrial membrane. A concept is discussed considering mitochondrial structure changes as a function of the osmotic gradient across the inner membrane and the oncotic gradient across the outer membrane of mitochondria.

Rosette-like structures from nuclei with condensed (chromomeric) chromatin but not from nuclei with diffuse (nucleomeric or nucleosomic) chromatin

The structure of chromatin of rat hepatocyte nuclei has been studied. At low ionic strength (20-50) chromatin in isolated nuclei, depending on the concentration of MgCl2 in the solution (0-2 and 4-5 mM), may be present in two states, respectively, diffuse and condensed. The major structural component of the nuclei with condensed chromatin is globular structures 100 nm in diameter, i.e. chromomers. By treating chromomer-containing nuclei with heparin and dextransulfate (polyanions/DNA equal 1), one can isolate rosette-like structures having an electron-dense core and numerous loops (the number loops in the rosette, 15-30, total length of all the loops, 15-20 micrometers, core diameter, 30-60 nm). The action of endogeneous nuclease on the nuclei and DNase I (but not RNase) on the rosette results in the break-down of the loops. Pronase or higher concentrations of polyanions (polyanions/DNA equal 4) induces partial or total decondensation of the rosette core and unfolding of the loops into a continuous linear structure. Rosette structures are not isolated from nuclei with diffuse chromatin. Rosette structures are discussed in terms of the known levels of the organization of chromatin.

The silver-stained NOR and argentophilic nuclear proteins in early mouse embryogenesis: a cytological study

Silver staining (Howell and Black, 1980) was used in light and electron microscopic studies for detecting the localization of argentophilic nuclear proteins in fertilized ova and cleaving mouse embryos. No silver-stained nucleolus organizing regions (NORs) (Ag-NORs) were visualized in the metaphase chromosomes of the first cleavage mitosis. From the 2-cell stage on, metaphase chromosomes contained Ag-NORs. Argentophilic proteins were detected in the pronuclei of the 1-cell embryos, i.e. before transcription of the ribosomal genes started. After fertilization these proteins accumulated on the decondensing sperm chromatin and telophase maternal chromosomes, then migrated into the pronuclei to be stored in pronucleoli, and, during mitosis, were transferred into the cytoplasm. In the metaphase chromosomes of the cleaving embryos Ag-NORs adequately reflected the transcriptional activity of the ribosomal genes, whereas in pronuclei of the 1-cell stage embryos argentophilic proteins were not involved in this process, but are likely to play a part in the formation and maturation of pronucleoli, and in the cell cycle regulation.

Reversible differential decondensation of unfixed Chinese hamster chromosomes induced by change in calcium ion concentration of the medium

Differential decondensation of isolated unfixed Chinese hamster metaphase chromosomes was obtained by decreasing the calcium ion concentration in the surrounding medium. A banded appearance of the swollen chromosomes could be observed either directly by phase contrast microscopy or after glutaraldehyde fixation and staining. There was a gradual transition from homogeneously dense to banded and finally to extensively decondensed chromosomes. The patterns induced at different stages were similar to those observed on fixed chromosomes after standard banding procedures (i.e., G-, C-, Cd−, Ag-NOR-staining). Chromosome decondensation could be reversed by the addition of calcium ions to the medium. Ca++-dependent reversible differential chromosome decondensation was not observed if the chromosomes were previously treated with 0.35 M NaCl. Chromosome regions which had incorporated BrdU into their DNA were more resistant to a decrease in calcium ion concentration than BrdU non-substituted regions.

Mouse Centromeric Heterochromatin: Isolation and Some Characteristics

A method is suggested for isolation of highly purified mouse centromeric heterochromatin. Treatment of mouse liver nuclei with decreasing concentrations of Ca2+ resulted in the gradual unraveling of chromatin in the nucleus and at 0.1 mM Ca2+ electron microscopy revealed several dense particles per nucleus, surrounded by decondensed chromatin. These particles, assumed to represent centromere regions of interphase chromosomes by in situ hybridization with radioactive mouse satellite DNA and by differential staining for centromere heterochromatin, were isolated in preparative amounts and their DNA and protein composition was analyzed. The preparation represented practically pure mouse centromere heterochromatin, since more than 90% of its DNA was satellite DNA.

Electron microscopic studies on the polytene chromosomes of Chironomus thummi salivary glands

The method of ultrathin sections of unsquashed salivary gland polytene chromosomes of Ch. thummi was applied to their ultrastructural mapping. There was a good agreement between electron micrographs and Hägeles light microscopic map (1970) with respect to the pattern and number of bands. 94% of bands were identified in larval and prepupal chromosomes. In Ch. thummi, band thickness varied from 0.05–0.5 μm. Most characteristic were 0.2–0.3 μm bands. Morphologically, bands were classified as: continuous (frequently with holes and gaps), discrete, dotted and continuous-discrete, discrete-dotted.Band morphology is related to band size, such that smaller bands, as a rule, were also dotted. Bands beginning to puff likewise became dotted. Interbands in unsquashed chromosome sections were from 0.05–0.15 μm. The smallest interbands contained only fibrils, in the larger interbands few granules could be observed. This makes interbands distinguishable from a typical puff with many such granules.