Outer retinal anoxia during dark adaptation is not a general property of mammalian retinas

The purpose of this study was to measure the intraretinal oxygen distribution across the retina under conditions, which maximise outer retinal oxygen consumption. In particular, we looked for evidence of increased oxygen delivery from the choroid and the deep retinal capillary layer, and whether or not this was sufficient to avoid the development of intraretinal anoxia. Under dark-adapted conditions the photoreceptors need additional energy, at least part of which is derived from increased oxidative metabolism. In earlier studies in the cat retina it was revealed that dark adaptation could render some regions of the outer retina anoxic. The present study of the in vivo oxygen distribution across the rat retina in light and dark found no evidence of outer retinal anoxia in the dark. This was despite a mean increase of 52.6+/-11.4% (n=7) in outer retinal oxygen consumption in the dark. The mean value for the minimum outer retinal PO(2) in the dark was 5.2+1.2 mmHg. Oxygen delivery from both the choroid and the deep retinal capillary layer increased in the dark (P<0.01, and P<0.001, respectively). It is argued that the ability of the deep capillary layer to compensate for changes in oxygen demand in the outer retina is an important element in the maintenance of homeostasis in the retina. This is in addition to the role of the deep capillary layer in supplying oxygen to the highly consuming plexiform layers within the inner retina. These findings in the rat retina also demonstrate that intraretinal anoxia in the dark, is not, as implied by earlier work in the cat, a general feature of mammalian retinas.     

Simultaneous Measurement of Oxygen Evolution and Endogenous Adenosine-Triphosphate Levels in Isolated 'Intact' Chloroplast Suspensions

Simultaneous continuous polarographic estimation of oxygen evolutionand measurement of ATP levels by the firefly luciferase methodwere made in suspensions containing predominantly intact spinachchloroplasts. Measurements were made during light to dark anddark to light transitions in the presence and absence of bicarbonate,3-phosphoglycerate and ribose-5-phosphate and following theaddition of these compounds during steady state conditions. Absolute levels of ATP were usually in the range 3.7 µmolATP per g chlorophyll and the results of kinetic experimentsare interpreted within the established concepts of photophosphorylationand the carbon reduction cycle. In experiments involving dark to light transitions in the absenceof added bicarbonate, light to dark transitions in the presenceof 3-phosphoglycerate and during photosynthesis in the presenceof bicarbonate, 3-phosphoglycerate and ribose-5-phosphate rapidtransient changes in ATP levels were observed.     

Growth of Ectothiorhodospira mobilis in the dark

The purple photosynthetic bacterium Ectothiorhodospira mobilis, like E. shaposhnikovii, can grow in the dark in the presence of oxygen on organic media, in particular, containing acetate or malate. The source of sulfur may be sulfate or thiosulfate. The two bacteria grown in the light and in the dark display the activity of all the enzymes of the citric acid cycle, with the exception of alpha-ketoglutarate dehydrogenase, and possess the enzymes of the glyoxylate shunt (isocitrate lyase and malate synthase). Irrespective of the conditions of the cultural growth, active fixation of carbon dioxide by the cells of E. mobilis was found only in the light.     

A comparison of the photosynthesis — light intensity relationship in phylogenetically different marine microalgae

Measured under equivalent physiological conditions, the photosynthesis-light intensity relationship based on oxygen production/mg chlorophyll a was found to be the same in five species representing the different chlorophyll c containing divisions of marine phytoplankton. Other non-photochemical metabolic processes related to photosynthesis such as diurnal variations, maximal photosynthesis rates, and dark oxygen uptake were quite different, and so these are the more significant factors in production and ecological distribution of diatoms, dinoflagellates, and coccolithophores. In contrast, the green algae tested showed a significantly different photosynthesis-light intensity curve from the chlorophyll c group.     

Some effects of light on algal respiration and the validity of the light and dark hottle technique for measuring primary productivity

SUMMARY. Measurements of the rate of oxygen uptake in a number of blue-green algae and diatoms were carried out under both field and laboratory conditions to determine the effects of light on such rates. The light history of algal cells was an important controlling factor of oxygen uptake. When measured in the light, with dichlorophenyl-dimethylurea (DCMU), oxygen uptake was sometimes different from uptake measured in the dark. The results cast some doubt on the validity of the light and dark bottle method for determining primary productivity. It is suggested that oxygen uptake measurements should be made in the presence of DCMU.     

The effect of dissolved oxygen concentration on the growth physiology of Saccharomyces cerevisiae whi2 mutants

Isogenic whi2 and WHI2+ strains of Saccharomyces cerevisiae were grown in a 2-litre bioreactor as batch cultures on a medium containing yeast extract and peptone with either glucose or ethanol as carbon and energy source. The concentration of dissolved oxygen within the medium was varied over the range of 0 to 100% saturation. Expression of the whi2 phenotype only occurred above 40% oxygen saturation with either glucose or ethanol as carbon and energy source. Under these conditions the whi2 cells could be distinguished from WHI2+ cells in that they were phase dark, highly budded and very small during the stationary growth phase, and reached final cell densities four to six times higher than WHI2+ cells. The results clearly show that the WHI2 gene of S. cerevisiae plays an important role in cell proliferation and that the availability of oxygen, or some product of oxidative metabolism, is involved in regulating the phenotypic expression of mutations within this gene.     

Effect of lectins on auxin-induced elongation and wall loosening in oat coleoptile and azuki bean epicotyl segments

A marine unicellular aerobic nitrogen-fixing cyanobacterium Synechococcus sp. strain Miarni BG 043511 was pretreated with different light and dark regimes in order to induce higher growth synchrony. A pretreatment of two dark and light cycles of 16 h each yielded good synchrony for 3 cell division cycles. Longer dark treatments decreased the degree of synchrony and shorter dark treatments caused irregular cell division. Once synchronous culture was established, distinct phases of cellular carbohydrate accumulation and cellular carbohydrate degradation were observed even under continuous illumination. Changes in carbohydrate content were repeated in a cyclic manner with approximately 20 h intervals, the same as the cell division cycle. This change in carbohydrate metabolism provided a good index of growth synchrony under nitrogen-fixing conditions.
Photosynthetic oxygen evolution and nitrogen fixation capabilities and their activities in near, in situ, culture conditions were measured in well synchronized cultures of this strain under continuous illumination. Distinct oscillations of both photosynthetic oxygen evolution and nitrogen fixation capabilities with ca 20-h intervals, similar to the interval of the cell division cycle, were observed for three cycles. However, the activities of photosynthetic oxygen evolution were inversely correlated with those of nitrogen fixation. During the nitrogen fixation period, net oxygen consumption was observed even in the light under conditions approximating in situ culture conditions. The phase of temporal appearance of nitrogenase activity during the cell division cycle coincided with the phase of carbohydrate net degradation. These data indicate that this unicellular cyanobacterium can grow diazotrophically under conditions of continuous illumination by the segregation of photosynthesis and nitrogen fixation within a cell division cycle.     

Light and Dark Controls of Nitrate Reduction in Wheat (Triticum aestivum L.) Protoplasts

Protoplasts were isolated from the leaves of nitrate-cultured wheat (Triticum aestivum L. var. Frederick) seedlings. When incubated in the dark, protoplasts accumulated nitrite under anaerobic, but not under aerobic, conditions. The assimilation of [¹⁵N]nitrite by protoplasts was strictly light-dependent, and no loss of nitrite from the assay medium was observed under dark aerobic conditions. Therefore, the absence of nitrite accumulation under dark aerobic conditions was the result of an O₂ inhibition of nitrate reduction and not a stimulation of nitrite reduction. In the presence of antimycin A, protoplasts accumulated nitrite under dark aerobic conditions. The oxygen inhibition of nitrate reduction was apparently due to a competition between nitrate reduction and dark respiration for cytoplasmic-reducing equivalents.     

Ascorbic acid oxidase is dynamically regulated by light and oxygen. A tool for oxygen management in plants?

Ascorbic acid oxidase (AAO) is a plant blue-copper protein catalyzing dioxygen reduction to water using ascorbic acid as the electron donor. In spite of extensive molecular characterization the physiological role of AAO is still uncertain. Abundant mRNA, protein and activity of AAO were observed in illuminated leaves of Cucurbita pepo. AAO activity was found to be proportional to light intensity. The light effect was rapidly reversed in dark and activity remained low throughout the dark period. Activity was elicited in dark by increased oxygen concentration. AAO activity increased in the facultative CAM Kalancho? blossfeldiana upon induction of the CAM cycle and decreased during germination of C. pepo and Zea mays under hypoxic conditions. These results strongly suggest that AAO activity could be part of a dynamic system for oxygen management in plants.     

Reactive oxygen species can modulate circadian phase and period in Neurospora crassa

Reactive oxygen species (ROS) may serve as signals coupling metabolism to other cell functions. In addition to being by-products of normal metabolism, they are generated at elevated levels under environmental stress situations. We analyzed how reactive oxygen species affect the circadian clock in the model organism Neurospora crassa. In light/dark cycles, an increase in the levels of reactive oxygen species advanced the phase of both the conidiation rhythm and the expression of the clock gene frequency. Our results indicate a dominant role of the superoxide anion in the control of the phase. Elevation of superoxide production resulted in the activation of protein phosphatase 2A, a regulator of the positive element of the circadian clock. Our data indicate that even under nonstress conditions, reactive oxygen species affect circadian timekeeping. Reduction of their basal levels results in a delay of the phase in light/dark cycles and a longer period under constant conditions. We show that under entrained conditions the phase depends on the temperature and reactive oxygen species contribute to this effect. Our results suggest that the superoxide anion is an important factor controlling the circadian oscillator and is able to reset the clock most probably by activating protein phosphatase 2A, thereby modulating the activity of the White Collar complex.     

The Ventral Photoreceptor Cells of Limulus : III. A voltage-clamp study

In the dark, the ventral photoreceptor of Limulus exhibits time-variant currents under voltage-clamp conditions; that is, if the membrane potential of the cell is clamped to a depolarized value there is an initial large outward current which slowly declines to a steady level. The current-voltage relation of the cell in the dark is nonlinear. The only ion tested which has any effect on the current-voltage relation is potassium; high potassium shifts the reversal potential towards zero and introduces a negative slope-conductance region. When the cell is illuminated under voltage-clamp conditions, an additional current, the light-induced current, flows across the cell membrane. The time course of this current mimics the time course of the light response (receptor potential) in the unclamped cell; namely, an initial transient phase is followed by a steady-state phase. The amplitude of the peak transient current can be as large as 60 times the amplitude of the steady-state current, while in the unclamped cell the amplitude of the peak transient voltage never exceeds 4 times the amplitude of the steady-state voltage. The current-voltage relations of the additional light-induced current obtained for different instants of time are also nonlinear, but differ from the current-voltage relations of the dark current. The ions tested which have the greatest effect on the light-induced current are sodium and calcium; low sodium decreases the current, while low calcium increases the current. The data strongly support the hypothesis that two systems of electric current exist in the membrane. Thus the total ionic current which flows in the membrane is accounted for as the sum of a dark current and a light-induced current.     

Semiaerobic induction of bacteriochlorophyll synthesis in the green bacterium Chloroflexus aurantiacus.

Comparison of Chloroflexus aurantiacus J-10-fl cells by freeze-fracture electron microscopy showed that cell shape and dimensions did not depend on oxygen tension or light intensity during growth. The major morphological difference between cells cultured anaerobically in the light and aerobically in the dark was the absence of chlorosomes in aerobically grown cells. C. aurantiacus cells cultured aerobically in the dark began bacteriochlorophyll synthesis immediately when shifted to either phototrophic or semiaerobic conditions. Cells adapting to phototrophic conditions grew to the same density and synthesized as much bacteriochlorophyll as nonadapting phototrophic cultures grown at the same light intensity. Cells adapting to reduced oxygen tension (semiaerobic conditions) in the dark entered an 8- to 12-h growth lag during which the bacteriochlorophyll content increased significantly. Despite variations in the initial bacteriochlorophyll content and in the length of the growth lag, the amounts of bacteriochlorophyll a and c were constant at the end of the semiaerobic growth lag. At later times during adaptation to semiaerobic conditions, after growth resumed, variations in the ratio of bacteriochlorophyll c/bacteriochlorophyll a were observed and suggested independent regulation of the two bacteriochlorophylls.     

Induction of benzo[a]pyrene Mono-oxygenase in liver cell culture by the photochemical generation of active oxygen species. Evidence for the involvement of singlet oxygen and the formation of a stable inducing intermediate.

1. The photochemical generation of excited states of oxygen in liver cell culture by the mild ilumination of culture medium containing riboflavin, results in stimulation of benzo[a]pyrene 3-mono-oxygenase, a cytochrome P-450-linked mono-oxygenase. 2. The same large increase in mono-oxygenase activity was found when medium containing riboflavin was illuminated in the absence of cells and then stored in the dark for 24h before contact with the cells. From this it may be inferred that stimulation is due to the formation of a stable inducer in the culture medium. Further experiments indicate that the stable inducer is due to the photo-oxidation of an amino acid. 3. Evidence that singlet oxygen is responsible for initiating the stimulation of the mono-oxygenase is based on the use of molecules that scavenge particular active oxygen species. Of all the scavengers tested, only those that scavenge single oxygen inhibited the stimulation. 4. A hypothesis is developed to relate the stimulation of the mono-oxygenase by singlet oxygen in cultured cells to the regulation of the cytochrome P-450 enzyme system in vivo. It is suggested that single oxygen generation within cells may be a common factor linking the many structurally diverse inducers of the enzyme system.     

Redox changes of cytochrome b(559) in the presence of plastoquinones

We have found that short chain plastoquinones effectively stimulated photoreduction of the low potential form of cytochrome b(559) and were also active in dark oxidation of this cytochrome under anaerobic conditions in Triton X-100-solubilized photosystem II (PSII) particles. It is also shown that molecular oxygen competes considerably with the prenylquinones in cytochrome b(559) oxidation under aerobic conditions, indicating that both molecular oxygen and plastoquinones could be electron acceptors from cytochrome b(559) in PSII preparations. alpha-Tocopherol quinone was not active in the stimulation of cytochrome photoreduction but efficiently oxidized it in the dark. Both the observed photoreduction and dark oxidation of the cytochrome were not sensitive to 3-(3,4-dichlorophenyl)-1, 1-dimethylurea. It was concluded that both quinone-binding sites responsible for the redox changes of cytochrome b(559) are different from either the Q(A) or Q(B) site in PSII and represent new quinone-binding sites in PSII.     

Control of nitrate and nitrite assimilation by carbohydrate reserves,adenosine nucleotides and pyridine nucleotides in leaves of Zea mays L. under dark conditions

The rate of in-vivo nitrate reduction by leaf segments of Zea mays L. was found to decline during the second hour of dark anaerobic treatment. On transfer to oxygen the capacity to reduce nitrate under dark conditions was restored. These observations led to the proposal that nitrate reductase is a regulatory enzyme with ADP acting as a negative effector. The effect of ADP on the invitro activity of nitrate reductase and the changes in the in-vivo adenylate pool under dark-N₂ and dark-O₂ were investigated. It was found that ADP inhibited the activity of partially purified nitrate reductase. Similarly, the in-vivo anaerobic inhibition of nitrate reduction was associated with a build-up of ADP in the leaf tissue. Under anaerobic conditions nitrite accumulated and on transfer to oxygen the accumulated nitrite was reduced. To explain this phenomenon the following hypothesis was proposed and tested. Under anaerobic conditions the supply of reducing equivalents for nitrite reduction in the plastid becomes restricted and nitrite accumulates as a consequence. On transfer to oxygen this restriction is removed and nitrite disappears. This capacity to reduce accumulated nitrite was found to be dependent on the carbohydrate status of the leaf tissue.     

Synthesis of bacteriochlorophyll a by the purple nonsulfur bacterium Rhodobacter capsulatus

The ability of purple nonsulfur bacteria Rhodobacter capsulatus B10 to synthesize bacteriochlorophyll under phototrophic and dark conditions was studied. The modes for cultivation in the dark with oxygen limitation in a continuous culture at D = 0.1 h(-1) were selected. The yield of biomass reached 20 g/l; the bacteriochlorophyll a output of the process amounted to 16.6 mg/l h(-1).     

Participation of oxygen in the reduction of methylviologen, photosensitized by chlorophyll, in an aqueous solution of detergent]

Dependence of chlorophyll "a" photosensitized reduction of methylviologene with tiourea on the temperature of reaction mixture was studied in aerobic conditions in triton X-100 aqueous solution. It was found that the reaction consisted of two stages: the light and dark ones. Photosensitized oxidation of tiourea with air oxygen proceeds at the temperatures up to -70 degrees C. Reduction of methylviologen is a dark stage for which diffusion processes are necessary. The role of hydrogen peroxide in the reaction studied has been investigated. It has been shown that hydrogen peroxide is not the "initiator" of the reaction which results in the reduction of methylviologen. Reduced glutation and the mixture of reduced and oxidized glutations were used as electron donors in photosensitized reaction in the presence of air oxygen. An increase of the depth and rate of the reduction of methylviologen under aerobic conditions as compared to anaerobic ones points to the formation of more active reducers than the initial electron donor.     

Some effects of light on the viability of rhodopsin-containing halobacteria

Under starvation conditions, rhodopsin-containing halobacteria show a light-accelerated death under aerobic conditions. This is attributed to photooxidative processes. Under anaerobic conditions, halobacteria die rapidly in the dark, and light prevents death. Since it has been shown by others that lightdriven ATP synthesis can occur under anaerobic conditions, it is postulated that rhodopsin-mediated photophorylation is of survival value for this organism in the brines in which it lives, especially because the solubility of oxygen is low in highly saline waters and anaerobic conditions can often develop.     

On the role of oxygen for nitrogen fixation in the marine cyanobacterium Trichodesmium sp

The marine, non-heterocystous, filamentous cyanobacterium Trichodesmium shows a distinct diurnal pattern of nitrogenase activity. In an attempt to reveal the factors that control this pattern, a series of measurements were carried out using online acetylene reduction assay. Light response curves of nitrogenase were recorded applying various concentrations of oxygen. The effect of oxygen depended on the irradiance applied. Above a photon irradiance of 16 mumol m(-2) s(-1) nitrogenase activity was highest under anoxic conditions. Below this irradiance the presence of oxygen was required to achieve highest nitrogenase activity and in the dark 5% oxygen was optimal. At any oxygen concentration a photon irradiance of 100 mumol m(-2) s(-1) was saturating. When Trichodesmium was incubated in the dark, nitrogenase activity gradually decreased and this decline was higher at higher levels of oxygen. The activity recovered when the cells were subsequently incubated in the light. This recovery depended on oxygenic photosynthesis because it did not occur in the presence of DCMU [3-(3,4-dichlorophenyl)-1,1-dimethylurea]. Recovery of nitrogenase activity in the light was faster at low oxygen concentrations. The results showed that under aerobic conditions nitrogenase activity was limited by the availability of reducing equivalents suggesting a competition for electrons between nitrogenase and respiration.