DARK ADAPTATION OF HORIZONTAL CELLS IN THE TELEOST FISH RETINA

The dark adaptation behaviors of rod-driven and cone-driven horizontal cells were exam-ined by analyzing their light responses recorded intracellularly in the intact, immobilizedcarp, and compared with that of the electroretinographic b--wave recorded simultaneously.Like the b--wave, the light responsiveness of rod horizontal cells increased gradually withtime in the dark and attained a steady level at 60 min. On the other hand, cone horizontalcells initially increased in light responsiveness in the first 10 min, but thereafter decreasedsteadily so that the response amplitudes of these cells to bright light flashes were only 3--5 mV.The results suggest that cone horizontal cells are strongly suppressed in prolonged darkness.     

Changes in the Inner Retinal Cells after Intense and Constant Light Exposure in Sprague-Dawley Rats

Light insult causes photoreceptor death. Few studies reported that continuous exposure to light affects horizontal, Müller and ganglion cells. We aimed to see the effect of constant light exposure on bipolar and amacrine cells. Adult Sprague-Dawley rats were exposed to 300 or 3000 lux for 7 days in 12-h light: 12-h dark cycles (12L:12D). The latter group was then exposed to 24L:0D for 48 h to induce significant damage. The same animals were reverted to 300 lux and reared for 15 days in 12L:12D cycles. They were sacrificed on different days to find the degree of retinal recovery, if any, from light injury. Besides photoreceptor death, continuous light for 48 h resulted in downregulation of parvalbumin in amacrine cells and recoverin in cone bipolar cells (CBC). Rod bipolar cells (RBC) maintained an unaltered pattern of PKC-α expression. Upon reversal, there were increased expressions of parvalbumin in amacrine cells and recoverin in CBC, while RBC showed an increasing trend of PKC-α expression. The data show that damage in bipolar and amacrine cells after exposure to intense, continuous light can be ameliorated upon reversal to normal LD cycles to which the animals were initially acclimated to.     

TRIFLUOPERAZINE: CORNEAL ENDOTHELIAL PHOTOTOXICITY

Abstract— Trifluoperazine is a commonly used agent for the treatment of psychiatric disorders. Perfusion of corneal endothelial cells with trifluoperazine-HCI concurrent with 3 min of exposure to long wavelength ultraviolet light resulted in a corneal swelling rate which was greater than that found in corneas where endothelial cells were perfused with trifluoperazine-HCI and not exposed to ultraviolet light. Exposure of endothelial cells to 25 W incandescent light for 5 min during perfusion with trifluoperazine-HCI did not result in a corneal swelling rate in excess of that found during perfusion with trifluoperazine in the dark. The increased corneal swelling rate could be produced by pre-exposure of the trifluoperazine-HCI perfusing solution to ultraviolet light suggesting the production of toxic photo-products during exposure of trifluoperazine HCI to ultraviolet light. Perfusion of corneal endothelial cells with non-ultraviolet illuminated trifluoperazine HCI had no effect on endothelial cell membranes or ultra-structure. This is in contrast to cells perfused with trifluoperazine HCI that had been exposed to ultraviolet light in which there was an alteration of mitochondria and a loss of cytoplasmic homogeneity. The data imply that the trifluoperazine HCI photoproduct had an adverse effect on cellular transport mechanisms. The study also further demonstrates that value of the corneal endothelial cell model for identifying the physiological and anatomical changes occurring in photo-induced toxic reactions.     

Damage to the albino rat retina produced by low intensity light.

Abstract—Continuous exposure of albino rats to low intensity light causes progressive deterioration of the photoreceptor cells. Losses in ERG (b-wave) sensitivity which accompany this ‘light damage’ are directly related to lowered rhodopsin levels in the retina. The same relationship has been observed in light adaptation studies in which the retina is not damaged. Thus, it appears that the production of b-wave responses is not seriously altered when photoreceptor morphology is disrupted.     

Hypericin-mediated photocytotoxic effect on HT-29 adenocarcinoma cells is reduced by light fractionation with longer dark pause between two unequal light doses

The present study demonstrates the in vitro effect of hypericin-mediated PDT with fractionated light delivery. Cells were photosensitized with unequal light fractions separated by dark intervals (1 or 6 h). We compared the changes in viability, cell number, survival, apoptosis and cell cycle on HT-29 cells irradiated with a single light dose (12 J/cm(2)) to the fractionated light delivery (1 + 11 J/cm(2)) 24 and 48 h after photodynamic treatment. We found that a fractionated light regime with a longer dark period resulted in a decrease of hypericin cytotoxicity. Both cell number and survival were higher after light sensitization with a 6-h dark interval. DNA fragmentation occurred after a single light-dose application, but in contrast no apoptotic DNA formation was detected with a 6-h dark pause. After fractionation the percentage of cells in the G1 phase of the cell cycle was increased, while the proportion of cells in the G2 phase decreased as compared to a single light-dose application, i.e. both percentage of cells in the G1 and G2 phase of the cell cycle were near control levels. We presume that the longer dark interval after the irradiation of cells by first light dose makes them resistant to the effect of the second illumination. These findings confirm that the light application scheme together with other photodynamic protocol components is crucial for the photocytotoxicity of hypericin.     

Phytochrome-mediated Phototropism in Adiantum Protonemata II. Participation of Phytochrome Dark Reversion

A microbeam irradiation technique was used to analyze phytochrome-mediated phototropism of the protonema of the fern Adiantum capillus-veneris. One side of the sub-apical zone of a dark-adapted protonema was irradiated with a red-light (R) microbeam to induce phototropic curvature toward the irradiated side. Except the cases where fluence-response relationships were examined, the protonema was stimulated with the microbeam for about 30 s to provide a fluence (5.3–5.5 mmol m^-2) optimal for curvature. When the whole protonema was pretreated with a high fluence of R (about 9 mmol m^-2), no significant curvature could be induced by immediately subsequent one-sided microbeam stimulation. It was found, however, that curvature became inducible progressively as the time of microbeam stimulation was delayed after the R pretreatment. The protonema gained nearly full responsiveness within 40 min after the pretreatment. Moreover, the inductive effect of microbeam R escaped rapidly from the reversing effect of far-red light. These results indicated that most of the far-red-absorbing form of phytochrome (Pfr), which mediates the phototropic response, undergoes relatively fast dark reversion to the red-absorbing form. The dark reversion kinetics was analyzed further by taking into account that the phototropic responsiveness after the R pretreatment was measured in the presence of background Pfr over the protonema. This analysis revealed a rate constant of about 0.002 s^-1 (t_1/2? 6 min) for the dark reversion. It is considered that the Pfr dark reversion plays a role in establishing a lateral Pfr gradient in the unilaterally irradiated protonema.     

Cytotoxicity of All‐Trans‐Retinal Increases Upon Photodegradation†

All‐trans‐retinal (AtRal) can accumulate in the retina as a result of excessive exposure to light. The purpose of this study was to compare cytotoxicity of AtRal and photodegraded AtRal (dAtRal) on cultured human retinal pigment epithelial cells in dark and upon exposure to visible light. AtRal was degraded by exposure to visible light. Cytotoxicity was monitored by imaging of cell morphology, propidium iodide staining of cells with permeable plasma membrane and measurements of reductive activity of cells. Generation of singlet oxygen photosensitized by AtRal and dAtRal was monitored by time‐resolved measurements of characteristic singlet oxygen phosphorescence. Photodegradation of AtRal resulted in a decrease in absorption of visible light and accumulation of the degradation products with absorption maximum at ～330 nm. Toxicity of dAtRal was concentration‐dependent and was greater during irradiation with visible light than in dark. DAtRal was more cytotoxic than AtRal both in dark and during exposure to visible light. Photochemical properties of dAtRal indicate that it may be responsible for the maximum in the action spectra of retinal photodamage recorded in animals. In conclusion, photodegradation products of AtRal may impose a significant threat to the retina and therefore their roles in retinal pathology need to be explored.     

PHOTOREACTIVATION OF ULTRAVIOLET LIGHT-INDUCED DAMAGE IN CULTURED FISH CELLS AS REVEALED BY INCREASED COLONY FORMING ABILITY AND DECREASED CONTENT OF PYRIMIDINE DIMERS

Abstract— Cultured cells derived from a goldfish were irradiated with 254nm ultraviolet light. Cell survival and splitting of pyrimidine dimers after photoreactivation treatment with white fluorescent lamps were examined by colony forming ability and by a direct dimer assay, respectively. When UV-irradiated (5 J/m²) cells were illuminated by photoreactivating light, cell survival was enhanced up to a factor of 9 (40min) followed by a decline after prolonged exposures. Exposure of UV-irradiated (15 J/m²) cells to radiation from white fluorescent lamps reduced the amounts of thymine-containing dimers in a photoreactivating fluence dependent manner, up to about 60% reduction at 120 min exposure. Keeping UV-irradiated cells in the dark for up to 120min did not affect either cell survival or the amount of pyrimidine dimers in DNA, indicating that there were not detectable levels of a dark-repair system in the cells under our conditions. Correlation between photoreactivation of colony forming ability and photoreactivation of the pyrimidine dimers was demonstrated, at least at relatively low fluences of photoreactivating light.     

Cobalt(III) Chaperone Complexes of Curcumin: Photoreduction,Cellular Accumulation and Light‐Selective Toxicity towards Tumour Cells

Light‐activated prodrugs offer the potential for highly selective tumour targeting. However, the application of many photoactivated chemotherapeutics is limited by a requirement for oxygen, or for short activation wavelengths that can damage surrounding tissue. Herein, we present a series of cobalt(III)‐curcumin prodrugs that can be activated by visible light under both oxygenated and hypoxic conditions. Furthermore, the photoproduct can be controlled by the activation wavelength: green light yields free curcumin, whereas blue light induces photolysis of curcumin to a phototoxic product. Confocal fluorescence microscopy and phototocytotoxicity studies in DLD‐1 and MCF‐7 tumour cells demonstrated that the cobalt(III) prodrugs are nontoxic in the dark but accumulate in significant concentrations in the cell membrane. When cells were treated with light for 15 min, the cytotoxicity of the cobalt complexes increased by up to 20‐fold, whereas free curcumin exhibited only a two‐fold increase in cytotoxicity. The nature of the ancillary ligand and cobalt reduction potential were found to strongly influence the stability and biological activity of the series.     

Fractionated illumination after topical application of 5-aminolevulinic acid on normal skin of hairless mice: the influence of the dark interval

We have previously shown that light fractionation during topical aminolevulinic acid based photodynamic therapy (ALA-PDT) with a dark interval of 2h leads to a significant increase in efficacy in both pre-clinical and clinical PDT. However this fractionated illumination scheme required an extended overall treatment time. Therefore we investigated the relationship between the dark interval and PDT response with the aim of reducing the overall treatment time without reducing the efficacy. Five groups of mice were treated with ALA-PDT using a single light fraction or the two-fold illumination scheme with a dark interval of 30 min, 1, 1.5 and 2h. Protoporphyrin IX fluorescence kinetics were monitored during illumination. Visual skin response was monitored in the first seven days after PDT and assessed as PDT response. The PDT response decreases with decreasing length of the dark interval. Only the dark interval of 2h showed significantly more damage compared to all the other dark intervals investigated (P<0.05 compared to 1.5h and P<0.01 compared to 1h, 30 min and a single illumination). No relationship could be shown between the utilized PpIX fluorescence during the two-fold illumination and the PDT response. The rate of photobleaching was comparable for the first and the second light fraction and not dependent of the length of dark interval used. We conclude that in the skin of the hairless mouse the dark interval cannot be reduced below 2h without a significant reduction in PDT efficacy.     

EVIDENCE FOR DARK REPAIR OF FAR ULTRAVIOLET LIGHT DAMAGE IN THE BLUE-GREEN ALGA, GLOEOCAPSA ALPICOLA

Abstract— The inactivating effect of far UV light on the unicellular blue-green alga Gloeocapsa alpicola could be totally reversed by exposure to blue light immediately after irradiation. However, if the irradiated cells were held in the dark before exposure to blue light, reversal became progressively less efficient and almost disappeared after 60–80 h holding. Caffeine and acriflavine inhibited loss of photoreversibility, suggesting an involvement of excision functions. Chloramphenicol and rifampicin slightly increased the rate of loss of photoreversibility, indicating that inducible functions play only a minor role. Split UV dose experiments indicated that light-dependent repair remained operational during dark liquid holding. These results provide preliminary evidence for dark repair in G. alpicola .     

Dark Color ZnO Quasi-One-Dimensional Nanostructures Grown by Hydrothermal Method and Modulation of their Optical Properties

Zinc oxide has a large energy gap and thus it has potential application in the field of solar cells by tuning the absorption of sunlight. In order to enhance its absorption of sunlight, dark color zinc oxides have been prepared by traditional hydrothermal method directly using a zinc foil as both source and substrate. We found that we could tune the optical properties of ZnO samples by changing the temperature. In particular, increasing temperature could significantly reduce the reflectivity of solar energy in the visible range. We speculate that the phenomenon is relevant to the sharp cone morphology of the ZnO nanorods grown on the surface of Zn foils, which furthermore enhance refraction and reflection of light in the nanorods. The capacity to improve the light absorption of ZnO may have a bright application in raising the efficiency of solar cells.     

REVERSIBLE BLEACHING OF Chlamydomonas reinhardtii RHODOPSIN in vivo

Abstract— The effect of hydroxylamine on the phototactic activity of Chlamydomonas reinhardtii was investigated. The following results were obtained: (1) wild type cells, irradiated for 10 min with green light immediately after addition of 1 mM hydroxylamine, showed a 20 min transient loss of phototactic activity, (2) irradiation of cells, preincubated in the dark with 4 mM. hydroxylamine for 30 min, diminished the phototactic sensitivity permanently by more than 100-fold without loss of cell motility. (3) The phototactic sensitivity completely recovered within 3(1 min of the removal of hydroxylamin from carotenoid-containing cells or from carotenoid-negative cells upon addition of 11- cis or all- trans retinal. Our explanation is bleaching of rhodopsin by more than 99% and reconstitution by de novo synthesized or by added retinal.     

MODIFYING FACTORS OF THE CELLULAR CONCENTRATION OF PHOTOLYASE MOLECULES IN Saccharomyces cerevisiae CELLS–I. EFFECTS OF TEMPERATURE AND LIGHT

Abstract— The effects of temperature and light on the cellular concentration of photoreactivating enzyme (PRE) molecules in haploid Saccharomyces cerevisiae cells were investigated. (1) Temperature effect: The number of active PRE molecules per cell (N_PRE) in cells grown at 37°C was about 13% of that grown at 23°C, although the amount of proteins per cell remained the same. (2) Light effect: N_PRE in cells grown in light was about 2.8 times larger than that grown in the dark. The value of N_PRE in cells grown in the light decreased more rapidly during holding in buffer in the dark than in the light. The N_PRE decrease during holding in buffer in the dark was more rapid in cells grown in the light than grown in the dark. A comparable decrease was observed after holding in buffer in the presence of cycloheximide. (3) In cells harboring a plasmid containing the gene PHR1, N_PRE was larger in cells grown at 23 than at 30°C.     

Squaraine dyes for photodynamic therapy: study of their cytotoxicity and genotoxicity in bacteria and mammalian cells

Halogenated squaraine dyes are characterized by long wavelength absorption (>600 nm) and high triplet yields and therefore represent new types of photosensitizers that could be useful for photodynamic therapy. We have analyzed the cytotoxicity and genotoxicity of the bromo derivative 1, the iodo derivative 2 and the corresponding nonhalogenated dye 3 in the absence and presence of visible light. At concentrations of 1-2 microM, 1 and 2 reduced the cloning efficiency of AS52 Chinese hamster ovary cells to less than 1% under conditions that were well tolerated in the dark. Similarly, the proliferation of L5178Y mouse lymphoma cells was inhibited by photoexcited 1 and 2 with high selectivity. The squaraine 3 was much less efficient. Both 1 and 2 induced only few mutations in the gpt locus of the AS52 cells in the presence of light and were not mutagenic in the dark. No mutagenicity with and without irradiation was observed in Salmonella typhimurium TA100 and TA2638. However, both 1 and 2 plus light increased the frequency of micronuclei in AS52 cells. The results indicate that halogenated squaraines exhibit photobiological properties in vitro that are favorable for photodynamic therapeutical applications.     

PHOTOSENSITIVITY IN THE SKIN OF THE LIZARD, Anolis carolinensis

Skin of the lizard, Anolis carolinensis, is remarkably sensitive to light. Illuminated in vitro with visible light from a tungsten source (110 W (m-2)), skin changes from brilliant green to dark brown (50% reduction in reflectance) within 2-4 min as a result of dispersion of melanin from a perinuclear position within dermal melanophores into superficial dendritic processes. Reversal of the process, reaggregation of pigment, will occur within 2.0 min upon return to darkness. This photic response can be initiated with light levels as low as 5.0 W m(-2) and is maximized by light levels only 5% that of midwinter sunshine. Pigment dispersion in response to both melanocyte simulating hormone and to light is inhibited by cytochalasin-B, indicating that microfilaments may be the motor element for pigment movement in that direction. Colchicine, but not cytochalasin-B, totally blocks pigment reaggregation following melanocyte stimulating hormone exposure and partially blocks it in the dark phase of the photic response. The results of this study are consistent with a model for pigment movement in A. carolinensis that provides microfilaments for pigment dispersion and microtubule involvement in both dispersion and aggregation. Finally, because it is readily visible, easily quantified, rapid and reversible, photic response in the skin of A. carolinensis is recommended as a valuable model system for the study of saltatory movement of organelles within cells.     

Microtubules and actin filaments in teleost visual cone elongation and contraction.

Teleost retinal cones contract in light and elongate in darkness. This paper describes the disposition of microtubules and cytoplasmic filaments in cone cells of 2 species of fish (Haemulon sciurus and Lutjanus griseus). In Haemulon, the neck-like "myoid" region of the cone changes in length from 5 mu to 75 mu. Maximal observed rates of elongation and contraction are comparable to that of chromosome movement in mitosis (2-3 mu/min). Microtubules presumably participate in cone elongation, since numerous longitudinal microtubules are present in the myoid region, and colchicine blocks dark-induced elongation. Myoid shortening, on the other hand, appears to be an active contractile process. Disruption of microtubules in dark-adapted cones does not produce myoid shortening in the absence of light, and light-induced myoid shortening is blocked by cytochalasin-B. Cone cells possess longitudinally-oriented thin filaments which bind myosin subfragment-1 to form arrowhead complexes typical of muscle actin. Myoid thin filaments are clearly observed in negatively stained preparations of isolated cones which have been disrupted with detergent after attachment to grids. These myoid filaments are not, however, generally preserved by conventional fixation, though bundles of thin filaments are preserved in other regions of the cell. Thus, actin filaments are poorly retained by fixation in precisely the region of the cone cell where contraction occurs. Cone cells also possess longitudinally-oriented thick filaments 130-160 A in diameter. That these thick filaments may be myosin is suggested by the presence of side-arms with approximately 150 A periodicity. The linear organization of the contractile apparatus of the retinal cone cell makes this cell a promising model for morphological characterization of the disposition of actin and myosin filaments during contraction in a nonmuscle cell.     

The effect of polarized versus nonpolarized light on melatonin regulation in humans

The aim of this study was to compare the effects of polarized light versus nonpolarized light on melatonin secretion in healthy, humans (mean age, 25 years; N = 6). On separate evenings, each subject was exposed to four different light intensities (20, 40, 80 and 3200 lx) of both polarized and nonpolarized light, as well as to a control, dark exposure. Each evening experiment consisted of a 120 min dark exposure (0000-0200 h) followed by a 90 min light exposure (0200-0330 h). Subjects' pupils were dilated prior to exposures. Blood samples were drawn at the start and end of each light-exposure period and later assayed for melatonin by radioimmunoassay. When compared to control exposures, both polarized and nonpolarized light elicited significant suppression of plasma melatonin at each illuminance (P < 0.03 to P < 0.0001), There were no significant differences between the effects of polarized light and nonpolarized light at any illuminance. The two light stimuli modalities demonstrated very similar fluence-response relationships between illuminance and melatonin suppression. Thus, the human pineal gland is responsive to ocular exposure with polarized light in a dose-dependent manner similar to that of nonpolarized light, although no significant differences were detected between polarized and nonpolarized light on melatonin regulation.     

Peculiar properties of chlorophyll thermoluminescence emission of autotrophically or mixotrophically grown Chlamydomonas reinhardtii

The microalgae Chlamydomonas reinhardtii and Chlorella sp. CCAP 211/84 were grown autotrophically and mixotrophically and their thermoluminescence emissions were recorded above 0 °C after excitation by 1, 2 or 3 xenon flashes or by continuous far-red light. An oscillation of the B band intensity according to the number of flashes was always observed, with a maximum after 2 flashes, accompanied by a downshift of the B band temperature maximum in mixotrophic compared to autotrophic grown cells, indicative of a dark stable pH gradient. Moreover, new flash-induced bands emerged in mixotrophic Chlamydomonas grown cells, at temperatures higher than that of the B band. In contrast to the afterglow band observed in higher plants, in Chlamydomonas these bands were not inducible by far-red light, were fully suppressed by 2 μM antimycin A, and peaked at different temperatures depending on the flash number and growth stage, with higher temperature maxima in cells at a stationary compared to an exponential growth stage. These differences are discussed according to the particular properties of cyclic electron transfer pathways in C. reinhardtii.     

Dynamics of FLCs with a tilted bookshelf structure using time-resolved FTIR spectroscopy

Time-resolved FTIR spectroscopy with microsecond resolution was applied to the switching process of FLCs using an IR polarizer attached to a microscope whose axis was set along the aligning direction (direction of SiO flux). A tilted bookshelf geometry with a layer tilt angle of approximately 34° was obtained by alignment using obliquely evaporated SiO layers in a CaF₂ cell with about 1.7 μm gap. Six kinds of FLC mixtures were prepared by mixing 5 wt % of a chiral molecule with host mixtures of phenylpyrimidines. By monitoring an absorption band at 1432 cm^-1, assigned to vibrational modes of the pyrimidine skeleton, it was found that the molecular long axis either rotates along the upper side or along the lower side of the cone during switching. Moreover, we investigated the relationship between the rotational direction of the directors and the stability of the dark level transmission during multiplexing. The dark level transmission during rotation along the upper side of the cone may fluctuate less than that during rotation along the lower side of the cone. We found that the bias stability of the dark level transmission is closely related to the direction of the rotation along the cone. The direction of the rotation of FLCs along the cone was therefore found to play an important role in realizing high contrast ratios during multiplexing.