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At least 145 records · Page 8Linked to original sources
PubMed1991

Cytochrome oxidase activity as a measure of synaptogenesis by multifunctional neuron L7 of Aplysia.

Neuron L7 of the marine mollusc, Aplysia californica, is unique in that it innervates five different target tissues in the animal. We show that when L7 is grown in vitro with two of these targets, that is, muscle cells isolated from the auricle or the gill vein, newly formed L7 neurites contact the muscle cells. Chemical synapses are formed since intracellular stimulation of L7 elicits contraction of individual muscle cells. Interestingly, auricle muscles are also innervated by neuron RBhe and co-cultures of RBhe and auricle muscle cells also exhibit synapse formation. To explore the molecular basis for synaptogenesis between L7 and its targets, it would be useful to quantify the extent of synapse formation in vitro, that is, to determine how many muscle cells can be innervated by a single L7. We show that this can be attained by staining for cytochrome oxidase activity. Cultures of auricle and gill vein muscles were exposed to the appropriate neurotransmitter in order to elicit contraction. The cells were then fixed and stained. In both cases, only cells that contracted were stained and electron microscopy showed reaction product associated with the cristae of mitochondria. When this procedure was applied to cultures of L7 and muscle cells, 38 +/- 2.8% (S.E.M.; n = 7) of the cells on the neurites were stained and therefore responded to L7 stimulation. Thus, part of the L7-RBhe circuit can be assembled in vitro and the extent of synaptogenesis can be accurately quantitated.

Animals↗
PubMed1990

The mitochondrial genome of an exsymbiotic Chlorella-like green alga.

Mitochondrial (mt) DNA from the unicellular, exsymbiotic Chlorella-like green alga, strain Nla was isolated and cloned. The mtDNA has a buoyant density of 1.692 g/ml in CsCl and an apparent G/C base composition of 32.5%. The genome contains approximately 76 kbp of DNA based on restriction fragment summation and electron microscopic measurements. A map of restriction endonuclease sites using Sst I, Bam I, Sal I and Xho I was generated. The genome maps as a circular molecule and appears as such under the electron microscope. Eight genes were assigned to the map by hybridization to specific restriction fragments using heterologous mt-encoded specific probes. These include the genes for subunits 6, 9, and alpha of the F0-F1 ATPase complex, the large and small subunit rRNAs, cytochrome oxidase subunits I and II, and apocytochrome b.

Apoproteins↗
PubMed1991

Effects of unilateral climbing fibre deafferentation on cytochrome oxidase activity in the developing rat cerebellum.

In a previous study, we found a relationship between climbing fibre synaptogenesis and cytochrome oxidase activity in Purkinje cells during normal development of the rat cerebellum. To determine whether removal of a major depolarizing afferent would alter the level of cytochrome oxidase activity in a post-synaptic neuron, climbing fibre input to Purkinje cells in the right hemicerebellum was interrupted by unilateral pedunculotomy in postnatal day 1 rat pups. After survival to postnatal day 5 (P5) or postnatal day 10 (P10), the cytochrome oxidase reactivity of mitochondria, packing density of mitochondria and perikaryal area of Purkinje cell somata were quantified at the electron microscopic level and compared with the same parameters in both sham-operated animals and normal controls. We found that the areal and numerical densities of darkly reactive mitochondria were lower in deafferented cells than those in the sham-operated animals. Cells of sham-operated animals, however, had higher densities of darkly reactive mitochondria than those in normal animals of the same age group, indicating that cell shrinkage or retarded growth had an effect on the levels of cytochrome oxidase activity in the operated animals. In addition, both operated groups had higher numerical densities of mitochondria than cells of normal animals, reflecting the decreased cell size of the sham and deafferented groups. From these data, we concluded that neonatal destruction of climbing fibres leads to lower levels of cytochrome oxidase activity in Purkinje cell somata that survived to both P5 and P10. The data from the P5 animals was more striking than that from P10, perhaps reflecting the increased number of synaptic interactions of Purkinje cells at P10. We also concluded that destruction of excitatory input did not lead to changes in the total area or number of mitochondria in a post-synaptic neuron, indicating that there was a conversion from darkly to lightly reactive mitochondria in the partially deafferented neurons; however, this may also reflect the smaller cell size of the deafferented group. Thus, our results further substantiate the close relationship between the levels of cytochrome oxidase activity in Purkinje cell somata and the type of input that they receive or fail to receive.

Afferent Pathways↗
PubMed1986

Preparation and characterization of submitochondrial fractions from adrenal cells.

A method for preparing submitochondrial fractions from adrenocortical cells was developed by adapting a procedure that has been successful with yeast mitochondria. The method is based upon osmotic swelling, sonication and centrifugation in sucrose. The preparation yields highly purified fractions of outer membrane, intermembrane space, inner membrane and a less purified fraction of matrix. Recoveries are good so that 10(7) cells yield approximately 170 micrograms of inner membrane protein and 12 micrograms of outer membrane protein. Electron microscopy shows that the outer membrane fraction consists of vesicles (0.2-0.6 micron diameter) while inner membrane appears as densely packed sheets of membranous material. Two-dimensional polyacrylamide gels (isoelectric focusing followed by electrophoresis) of all the fractions give highly reproducible patterns of protein spots with Coomassie staining. Steroidogenic proteins were found only in inner membrane fractions which were shown to contain cytochrome P-450 C27 side-chain cleavage and P-450 11 beta-hydroxylase together with adrenodoxin and adrenodoxin reductase. Inner membrane catalyzes side-chain cleavage of cholesterol (conversions to pregnenolone) and 11 beta-hydroxylation (DOC----corticosterone) when substrate and NADPH are added. The preparation yields highly purified submitochondrial fractions from Y-1 mouse adrenal tumor cells and from porcine and bovine adrenocortical mitochondria. The method has the virtue of yielding highly purified intermembrane fluid which is not true of other methods for fractionation of adrenal mitochondria. The procedure also yields cleaner preparations of the two membranes than two other published methods currently used to prepare submitochondrial fractions from adrenocortical cells.

Adrenal Glands↗
PubMed2004

Cytochrome oxidase I gene sequence of Hypoderma sinense infecting yaks in the Qinghai-Tibet high plateau of China.

The previous scanning electron microscopy study of the Hypoderma species suggested that Hypoderma sinense Pleske (H. sinense) was different from Hypoderma bovis (H. bovis) and Hypoderma lineatum (H. lineatum). In this study, the mitochondrial cytochrome oxidase I (COI) gene sequence of H. sinense was compared with those of the other two species. The H. sinense sequence showed only 88.3% homology to H. bovis and 88.5% to H. lineatum. The results of the sequencing analysis confirm that H. sinense is a different species from H. bovis and H. lineatum.

Animals↗
PubMed1982

Preparation of two-dimensional arrays from purified beef heart cytochrome c oxidase.

A method for preparing two-dimensional crystals from highly purified beef heart cytochrome c oxidase is described. This involves mixing the enzyme with phosphatidylcholine and then extracting excess lipid with deoxycholate. The reconstituted crystals show PI symmetry. Alternating rows of monomers are more closely packed (along the b axis) than in the previously described P121 crystals. However, the monomer structure in projection is the same in the two crystal forms. The P121 crystal form has been reacted with trypsin. This treatment did not alter the crystals but removed most of the impurities present in these cytochrome c oxidase preparation of low purity.

Animals↗
PubMed1962

The isolation of cell nuclei from rat brain.

A method for preparing highly pure cell nuclei from adult rat brain, using both differential and isopycnic centrifugation in sucrose media, is described. The morphology of these preparations was examined by both phase contrast and electron microscopy. The isolated nuclei retained many aspects of their in situ morphology; in particular, the nuclear envelope was double layered and interrupted by pore-like discontinuities, and the nucleoli consisted of irregular masses of densely packed granules. Analyses of these nuclear preparations for cytochrome oxidase and cholinesterase activity, as well as RNA/DNA ratio, indicated minimal contamination with mitochondria and microsomes. Problems involving the homogenization technique, choice of ionic conditions in the homogenization medium, and choice of optimal density of the sucrose solution used for the final purification of nuclei are discussed. Results of application of the technique to isolation of adult rat liver nuclei are also reported.

Animals↗
PubMed1977

Anaerobiosis and symbiosis with bacteria in free-living ciliates.

Marine, sediment-dwelling ciliates were examined for cytochrome oxidase activity by a cytochemical method and for fine structural details. Species of Plagiopylidae (Trichostomatida), i.e. Plagiopyla frontata, Sonderia vorax and Sonderia sp., and of Heterotrichda, i.e., Parablepharisma pellitum, Parablepharisma sp., Metopus contortus, Metopus vestitus and Caenomorpha capucina; previously considered to be obligate anaerobes because of their sulfide-containing habitat, do not have cytochrome oxidase activity or mitochondria with cristae or tubuli. The evolutionary origin and significance of anaerobic ciliates is discussed. Most of the anaerobic ciliates harbor a flora of ecto- and endosymbiotic bacteria as demonstrated by transmission and scanning electron micrographs. It is speculated that the bacteria may utilize the metabolic end products of the protozoa for growth and energy yielding processes. These associations are also compared with other, previously described cases of symbiosis involving prokaryotes and protozoa.

Anaerobiosis↗