Hypoxia-induced fibre type transformation in rat hindlimb muscles Histochemical and electro-mechanical changes

Twelve male Sprague-Dawley rats (21 days old) were randomly assigned into two experimental groups: sea level control (CONT) and hypobaric hypoxia (HYPO). The HYPO rats were kept in an hypobaric chamber maintaining a simulated altitude of 4000 m (61.1 kPa). After 10 weeks of treatment, the rat hindlimb muscles [soleus (SOL) and extensor digitorum longus (EDL)] were subjected to histochemical and electro-mechanical analyses. Results indicated that compared to CONT the HYPO SOL muscle had a significantly greater relative distribution of fast-twitch-oxidative-glycolytic (FOG) fibres (28.9% SEM 2.0 vs 18.3% SEM 1.8, P less than 0.01) with a significant decrease in slow twitch oxidative fibre distribution (69.5% SEM 2.4 vs 82.9% SEM 3.1, P less than 0.01). Compared to CONT the HYPO EDL muscle also manifested a significant increase in FOG fibre distribution (51.6% SEM 0.8 vs 46.6% SEM 1.1, P less than 0.01), but this was accompanied by a significant decrease in fast twitch glucolytic fibres (44.3% SEM 0.9 vs 49.2% SEM 1.7, P less than 0.05). These histochemical fibre type transformations accompanied significant and expected changes in the electro-mechanical parameters tested in situ, e.g. maximal twitch force, maximal rate of force development, contraction time, half relaxation time, force: frequency curve, and fatigability. It was concluded that chronic hypobaric hypoxia could have a potent influence upon the phenotype expression of muscle fibres.     

Immunohistochemical colocalization of glucagon,serotonin, and aromatic L-amino acid decarboxylase in islet A cells of chicken pancreas

Summary The identity of monoamine-emitted, formaldehyde-induced fluorescence in some pancreatic islet cells was studied in pancreatic tissue of male chickens by fluorescence and immunohistochemistry either on the same tissue section or on serial tissue sections. Pancreatic islet cells emitting intense formaldehyde-induced fluorescence also react immunohistochemically with antisera directed against glucagon, serotonin and aromatic L-amino acid decarboxylase. These results show that chicken pancreatic islet A cells contain glucagon, serotonin, and aromatic L-amino acid decarboxylase, an enzyme involved in the synthesis of serotonin. The islet B cells identified with anti-insulin immunoreactivity, which displayed a very weak formaldehyde-induced fluorescence, did not react with anti-serotonin serum.     

Rhodamine 123 as a vital stain for mitochondria of plant cells

Abstract. Rhodamine 123 (Rh 123), a relatively new mitochondrial marker, little used in the study of plant cells, was tested on excized leaves of Elodea canadensis Michx. and on suspension-cultured cells of Ranunculus serbicus Vis. In both preparations, the dye accumulated rapidly and selectively in the mitochondria whose number, morphology and cell distribution could be easily observed. In the presence of Rh 123, cytoplasmic movements could also be perceived and the spatial arrangement of the mitochondria with respect to that of the auto-fluorescent chloroplasts was studied in connection with a normal or altered cytoskeletal framework. The specific uptake of Rh 123 by the organdies seemed to be potential-dependent since it was influenced by cations, ionophores and inhibitors of electron transport. Short exposures to the stain were practically non-toxic, whereas prolonged treatments (6–20 h) provoked specific alterations in structure of the mitochondria. The data reported here indicate that Rh 123 may be an excellent vital stain to study the morphology, function and dynamics of the mitochondria in living plant cells.     

Pigment organization and energy transfer in the green photosynthetic bacterium Chloroflexus aurantiacus. III. Energy transfer in whole cells

The transfer of excitation energy in intact cells of the thermophilic green photosynthetic bacterium Chloroflexus aurantiacus was studied both at low temperature and under more physiological conditions. Analysis of excitation spectra measured at 4K indicates that the minor fraction of bacteriochlorophyll a present in the chlorosome functions as an intermediate in energy transfer between the main light-harvesting pigment BChl c and the membrane-bound B808-866 antenna complex. This supports the hypothesis that BChl a is associated with the base plate which connects the chlorosome with the membrane. The overall efficiency for energy transfer from the chlorosome to the membrane is only 15% at 4K. High efficiencies of close to 100% are observed above 40°C near the temperature where the cultures are grown. Cooling to 20°C resulted in a sudden drop of the transfer efficiency which appeared to originate in the chlorosome. This decrease may be related to a lipid phase transition. Further cooling mainly affected the efficiency of transfer between the chlorosome and the membrane. This effect can only partially be explained by a decreased Förster overlap between the chlorosomal BChl a and BChl a 808 associated with the membrane-bound antenna system. The temperature dependence of the fluorescence yield of BChl a 866 also appeared to be affected by lipid phase transitions, suggesting that this fluorescence can be used as a native probe of the physical state of the membrane.     

Effect of chloramphenicol and lincomycin on chloroplast DNA amplification in greening pea leaves

The light-induced increase in chloroplast DNA was not inhibited by two inhibitors of protein synthesis on 70S polysomes, chloramphenicol and lincomycin, in greening pea leaves. The changes in chloroplast DNA were observed by fluorescence microscopy and measured by hybridization to specific cloned probes. The results suggest that the light-induced increase in chloroplast DNA proceeds without de novo protein synthesis in the chloroplast, in agreement with those with mutants and cultured leaf tissue.     

Synchrotron-excited time-resolved fluorescence spectroscopy of adenosine,protonated adenosine and 6N, 6N-dimethyladenosine in aqueous solution at room temperature

The fluorescence behavior of adenosine in neutral solution has been studied by time-resolved spectroscopy using synchrotron excitation and timecorrelated single photon counting, and by decay time measurements. Three emissions have been identified and correlated with three excitation spectra. The assignment of these transitions has been made by comparison with similar measurements on ⁶N, ⁶N-dimethyladenosine (6 DMA), and on adenosine in acid solution (ADO H⁺). It is proposed that two of the transitions of adenosine which correlate with 6DMA originate from coplanar and orthogonal rotational conformers of the amino group. The other transition, correlating with ADO H⁺ may originate either from the ³H-imino tautomer, or from a differently solvated rotational conformer.A partial presentation of this work has been made at the Second Congress of the European Society for Photobiology Padova, Italy, 6–10 September 1987     

Circumnutation in Phaseolus vulgaris. I. Growth, osmotic potential and cell ultrastructure in the free-moving part of the shoot

Circumnutation of Phaseolus vulgaris L. cv. Blanc de Juillet twining shoots was examined at the cellular level. Cell growth was monitored inside the free-moving part of the shoot as a function of time and position with respect to the terminal bud. Complete elongation of epidermal cells required >7 days. A 60% increase in length of the cells in the bending zone was observed. Scanning and transmission electron microscopy studies showed the elasticity of cell walls, the extent of plasmodesmata and endoplasmic reticulum of epidermal cells. The osmotic potential in epidermal cells of the bending zone displayed significant differences only between the concave and convex sides. The pattern of cell growth in twining shoots is discussed. Circumnutation could be seen as a consequence of periodic turgor changes inducing periodic growth.     

Photoinhibition of photosynthesis in intact kiwifruit (Actinidia deliciosa) leaves: Effect of temperature

Photoinhibition of photosynthesis was induced in attached leaves of kiwifruit grown in natural light not exceeding a photon flux density (PFD) of 300 mol·m^-2·s^-1, by exposing them to a PFD of 1500 mol·m^-2·s^-1. The temperature was held constant, between 5 and 35° C, during the exposure to high light. The kinetics of photoinhibition were measured by chlorophyll fluorescence at 77K and the photon yield of photosynthetic O₂ evolution. Photoinhibition occurred at all temperatures but was greatest at low temperatures. Photoinhibition followed pseudo first-order kinetics, as determined by the variable fluorescence (F _v) and photon yield, with the long-term steady-state of photoinhibition strongly dependent on temperature wheareas the observed rate constant was only weakly temperature-dependent. Temperature had little effect on the decrease in the maximum fluorescence (F _m) but the increase in the instantaneous fluorescence (F _o) was significantly affected by low temperatures in particular. These changes in fluorescence indicate that kiwifruit leaves have some capacity to dissipate excessive excitation energy by increasing the rate constant for non-radiative (thermal) energy dissipation although temperature apparently had little effect on this. Direct photoinhibitory damage to the photosystem II reaction centres was evident by the increases in F _o and extreme, irreversible damage occurred at the lower temperatures. This indicates that kiwifruit leaves were most susceptible to photoinhibition at low temperatures because direct damage to the reaction centres was greatest at these temperatures. The results also imply that mechanisms to dissipate excess energy were inadequate to afford any protection from photoinhibition over a wide temperature range in these shade-grown leaves.Abbreviations and symbols fluorescence yield correction coefficient - F _o, F _m, F _v instantaneous, maximum, variable fluorescence - K _D, K _F, K _P, K _T rate constants for non-radiative energy dissipation, fluorescence, photochemistry, energy transfer to photosystem I - PFD photon flux density - PSI, II photosystem I, II - _i photon yield of photosynthesis (incident light)     

Metal ion stabilization of the conformation of a recombinant 19-kDa catalytic fragment of human fibroblast collagenase.

A recombinant 19-kDa human fibroblast collagenase catalytic fragment modeled on a naturally occurring proteolytic product was purified from E. coli inclusion bodies. Following renaturation in the presence of zinc and calcium, the fragment demonstrated catalytic activity with the same primary sequence specificity against small synthetic substrates as the full-length collagenase. Unlike the parent enzyme, it rapidly cleaved casein and gelatin but not native type I collagen. Intrinsic fluorescence of the three tryptophan residues was used to monitor the conformational state of the enzyme, which underwent a 24-nm red shift in emission upon denaturation accompanied by quenching of the fluorescence and loss of catalytic activity. Low concentrations of denaturant unfolded the fragment while the full-length enzyme displayed a shallow extended denaturation curve. Calcium remarkably stabilized the 19-kDa fragment, zinc less so, while together they were synergistically stabilizing. Among divalent cations, calcium was the most effective stabilizer, EC50 approximately 60 microM, and similar amounts were required for substrate hydrolysis. Catalytic activity was more sensitive to denaturation than was tryptophan fluorescence. Least sensitive was the polypeptide backbone secondary structure assessed by CD. These observations suggest that the folding of the 19-kDa collagenase fragment is a multistep process stabilized by calcium.