DIEL CHANGES IN THE COMPOSITION OF PHOTOSYNTHETIC PIGMENTS AND CELLULAR CARBON AND NITROGEN IN PYRAMIMONAS PARKEAE (PRASINOPHYCEAE)1

Diel changes in mean cell volume, cellular carbon (carbon content per cell), cellular Chl a, C/N ratio, Chl a/carbon ratio and pigment composition were determined for an axenic clonal culture of Pyramimonas parkeae Norris et Pearson through three 12:12 h LD cycles in a laboratory culture tank of 1 m³. Mean cell volume and cellular C, N and most pigments increased during the light period as a result of photosynthesis and decreased with an increase in cell density by phased cell division during the dark period. Chi a and Chi b increased in a parallel manner during the light period. Increases in the diel synthesis pattern of carotenoids varied. Violaxanthin and lutein content increased for a few hours at the beginning of the light period and preceeded that of neoxanthin. The diel synthesis pattern of neoxanthin was similar to that of Chi a. Increases of loroxanthin and its ester form were slower than that of Chi a at the beginning of the light period. A net increase of α-carotene was observed during the dark period. Mass spectroscopy of carotenoid structure showed a new xanthophyll, loroxanthin dodecenoate, in this species.     

INTERACTIONS OF LIGHT/DARK CYCLES AND NITROGEN PULSES ON THE TIMING OF CELL DIVISION IN THE NITROGEN-LIMITED MARINE DIATOM CYLINDROTHECA FUSIFORMIS (BACILLARIOPHYCEAE)1

Cell division in most eukaryotic algae grown on alternating periods of light and dark (LD) is synchronized or phased so that cell division occurs only during a restricted portion of the LD cycle. However, the phase angle of the cell division gate, the time of division relative to the beginning of the light period, is known to be affected by growth conditions such as nutrient status and temperature. In this study, it is shown that the phase angle of cell division in a diatom, Cylindrotheca fusiformis Reimann and Lewin, is affected by the N-limited growth rate; cell division occurred later in the dark period (12:12 h LD cycle) when the growth rate was infradian (D = 0.42 d^?1) than when it was ultradian (D = 1.0 d^?1). Nitrogen-pulses did not affect the phase angle of the division gate, but could shift the time of peak cell division activity within the division gate. The effects, if any, of N-pulses were dependent upon the growth rate and the time of day that the pulses were administered. These responses indicate that the timing of cell division in this diatom is not determined solely by the zeitgeber from the LD cycle, but rather that a LD cycle control mechanism and a N-mediated control mechanism are both involved and are somewhat interdependent. In addition, an increase in protein was observed immediately after administering a N-pulse to C. fusiformis in the ultradian growth mode indicating that the accumulation of protein can be uncoupled from the cell division cycle.     

Variations in biochemical parameters of Heterocapsa sp. and Olisthodiscus luteus grown in 12:12 light:dark cycles I. Cell cycle and nucleic acid composition

The division cycle of two phytoplankton species, Olisthodiscus luteus and Heterocapsa sp. was studied in relation to a 12:12 light:dark cycle. Batch cultures in exponential phase were sampled every three hours during 48 hours. Cell number, cellular volume and DNA and RNA concentrations were measured. Microscopic observations of the nuclei of Heterocapsa sp. were also performed. In both species, cell division took place in the dark. In Heterocapsa sp., DNA and RNA showed a similar diel variability pattern, with synthesis starting at the end of the light period, previously to mitosis and cytokinesis. In O. luteus. Major RNA synthesis occurred during darkness, and DNA was produced almost continuously. Both species presented different values and diel rhythmicity on the RNA/DNA ratios.     

DIEL CHANGES IN THE COMPOSITION OF PHOTOSYNTHETIC PIGMENTS AND CELLULAR CARBON AND NITROGEN IN CHATTONELLA ANTIQUA (RAPHIDOPHYCEAE)1

An axenic clonal culture of Chattonella antiqua (Hada) Ono was grown on a 12: 12 h LD cycle in a laboratory culture tank containing 1 m³ of f/2 medium. Diel changes in mean cell volume, cellular carbon (carbon content per cell), C/N ratio, cellular Chl a, Chl a/c ratio and carotenoid composition were observed. Mean cell volume and cellular C, N and pigments increased during the light period as a result of photosynthesis and decreased with increase of cell concentration by phased cell division during the dark period. These changes indicated that carbon assimilation and pigment synthesis occurred together during the light period. However, the patterns of increase were not the same since different diel patterns were also found in the ratios of C/N and chl a/c. Photosynthetic pigments were analyzed by reversed-phase high-performance liquid chromatography with ion-pairing solution. This analysis showed that the dominant carotenoids in C. antiqua were fucoxanthin, violaxanthin and β-carotene. Diel patterns of Chls a and c were similar to that of fucoxanthin but different from those of violaxanthin and β-carotene. The cellular contents of Chl a, fucoxanthin and carbon increased in a parallel manner during the light period. On the other hand, the increase of violaxanthin was restricted to only a few hours at the beginning of the light period during cell division cycles.     

Effects of photoperiod and temperature on the rhythm and free‐running of adult eclosion in the Indian meal moth Plodia interpunctella

The rhythm of adult eclosion in the Indian meal moth Plodia interpunctella Hübner (Lepidoptera: Pyralidae) is investigated under various photoperiods and temperatures aiming to determine the nature of the temperature compensation and the free‐running period. Insects that are committed to a nondiapause larval development show diel rhythms of adult eclosion at 30, 25 and 20 °C. At 30 °C, the eclosion peak (i.e. the mean time of eclosion) occurs approximately 20 h after lights off under an LD 4 : 20 h photocycle, and at approximately 15 h under an LD 20 : 4 h photocycle. At 25 °C, the peak of eclosion occurs approximately 19 h after lights off under an LD 2 : 20 h photocycle and at approximately 16 h under an LD 20 : 4 h photocycle. At 20 °C, the eclosion peak is significantly advanced under long days of >12 h (i.e. approximately 20 h after lights off under an LD 4 : 20 h photocycle and approximately 9 h under an LD 20 : 4 h photocycle), indicating an effect of both lights‐off and lights‐on signals on the timing of the adult eclosion. To determine the involvement of a self‐sustained oscillator, the rhythm of adult eclosion is examined under darkness at different temperatures (30 to 21 °C). The mean free‐running periods are 22.4, 22.8, 22.0 and 22.5 h at 30, 24, 23 and 22 °C, respectively, indicating that the eclosion rhythm is temperature‐compensated. However, this rhythm does not free‐run under constant darkness at 21 °C. Because a clear diel rhythm is observed under 24‐h photocycles at 20 °C, the oscillator might be damped out within 24 h at the lower temperature.     

Synchronous division and the pattern of diel vertical migration of Heterosigma akashiwo (Hada) Hada (Raphidophyceae) in a laboratory culture tank

Heterosigma akashiwo (Hada) Hada (Raphidophyceae) causes red tides in Osaka Bay (Japan). A clonal culture of the alga was grown in a 2 m tall culture tank on a 12: 12 LD cycle to determine patterns of vertical migration and cell division. A specific growth rate of 0.43 ln unit · day⁻¹ was obtained during complete mixing conditions. Under weakly stratified conditions (≈ΔT = 3–4°C/1.5 m), H. akashiwo in the tank grew and showed a similar pattern of vertical migration to that observed in the field for at least 6 days. Cell concentration, mean cell volume, and photosynthetic capacity, estimated by DCMU-induced fluorescence increase of H. akashiwo, were monitored in the stratified tank at 2-h intervals over 24 h at three levels in the water column. Cell ascent began shortly before the light period and vertically swimming cells were smaller in size than those sampled near the bottom of the tank. The cell division cycle and the pattern of vertical migration were phased individually by the light regime and were well synchronized with each other. This synchrony must be due to the interrelation between these two processes or the existence of a clock which controlled endogenous rhythms of both processes and was entrained by a light: dark cycle. The relative increase of fluorescence with DCMU was higher for migrating cells than for non-migrating cells.     

Diel Division Cycle and Growth Rates of Synechococcus in Lakes Huron and Michigan1

A clone of Synechococcus isolated from Lake Huron and natural populations of Synechococcus from lakes Huron and Michigan were studied in 1989 to examine the diel division cycle and to provide estimates of the in situ growth rate based on the frequency of dividing cells (FDC) method. Cultured populations of Synechococcus exhibited a consistent diel division pattern with a midday/afternoon (1100–1800 h) peak in the percent of dividing cells. The maximum percent of dividing cells varied among cultures (8-27%) and was related to the growth rate. A small fraction of dividing cells (3-5%) remained throughout the dark period, suggesting that some cells were arrested in the doublet stage prior to division. The duration of division (t_d) ranged from 2.6-4.9 h, with a 3.7 h mean for cultures with growth rates ≥0.34 d⁻¹ but increased to 8 h at a lower growth rate of 0.20 d⁻¹. The diel division pattern for natural populations was very similar to the laboratory clone; an afternoon peak (1400-2100 h) in dividing cells and a small fraction of dividing cells (2-5%) remained during the dark period. The maximum percent of dividing cells for natural populations ranged from 6-10%. In situ growth rates, determined from the FDC and assuming a constant t_d of 3.7 h, ranged from 0.30-0.42 d⁻¹. The FDC method may provide accurate estimates of in situ growth, particularly in environments where the growth rate is >0.34 d⁻¹, but in lakes Huron and Michigan where growth rates can be lower and t_d values may increase, FDC-growth rates must be viewed with caution.     

ROLE OF INORGANIC IONS IN CONTROLLING SEDIMENTATION RATE OF A MARINE CENTRIC DIATOM DITYLUM BRIGHTWELLI1,2

The settling rates and intracellular levels of K⁺, Na⁺, Cl^-, Mg²⁺ and Ca²⁺ were measured in Ditylum bright-welli (West) Grunow, grown axenically in an enriched seawater medium at 20 C at 4,000 lx on an 8:16 LD schedule. Cells at the end of the dark period have high Na⁺ (118 mM), low K⁺ (64 mM) and low Cl^- (117 mM) relative to levels at the end of the light period when K⁺ (126 mM) and Cl^- (154 mM) are high and Na⁺ (101 mM) is low. There is no significant change in Mg²⁺ (16–18 mM) or Ca²⁺ (3–4 mM) with time. The net result of the ion changes during the light period is to increase cell density by about 3.4 mg ml^-1. This change can account for the increase in settling rate of ca. 0.3 day^-1 during the same interval. The density of the cell contents, calculated from observed ion concentrations, is 15–18 mg.ml^-1 less than that of the seawater medium. The ion and settling rate changes are light-dependent and do not persist in the dark or under constant light (ca. 850 lx), but cells do exhibit a free-running circadian rhythm in cell division under continuous dim illumination. The cell vacuole expands during the light period and contracts during the dark, apparently in response to the net ion fluxes. D. brightwelli appears to regulate its density by active ion selectivity accompanied by trans-vacuolar water movement.     

DIEL PATTERNS OF GROWTH AND DIVISION IN MARINE PICOPLANKTON IN CULTURE

The effect of a 12:12-h light:dark (LD) cycle on the phasing of several cell parameters was explored in a variety of marine picophytoplanktonic strains. These included the photosynthetic prokaryotes Prochlorococcus (strains MED 4, PCC 9511, and SS 120) and Synechococcus (strains ALMO 03, ROS 04, WH 7803, and WH 8103) and five picoeukaryotes (Bathycoccus prasinos Eikrem et Throndsen, Bolidomonas pacifica Guillou et Chrétiennot-Dinet, Micromonas pusilla Manton et Parke, Pelagomonas calceolata Andersen et Saunders, and Pycnococcus provasolii Guillard et al.). Flow cytometric analysis was used to determine the relationship between cell light scatter, pigment fluorescence, DNA (when possible), and the LD cycle in these organisms. As expected, growth and division were tightly coupled to the LD cycle for all of these strains. For both Prochlorococcus and picoeukaryotes, chl and intracellular carbon increased throughout the light period as estimated by chl fluorescence and light scatter, respectively. In response to cell division, these parameters decreased regularly during the early part of the dark period, a decrease that either continued throughout the dark period or stopped for the second half of the dark period. For Synechococcus, the decrease of chl and scatter occurred earlier (in the middle of the light period), and for some strains these cellular parameters remained constant throughout the dark period. The timing of division was very similar for all picoeukaryotes and occurred just before the subjective dusk, whereas it was more variable between the different Prochlorococcus and Synechococcus strains. The burst of division for Prochlorococcus SS 120 and PCC 9511 was recorded at the subjective dusk, whereas the MED 4 strain divided later at night. Synechococcus ALMO 03, ROS 04, and WH 7803, which have a low phycourobilin to phycoerythrobilin (PUB:PEB) ratio, divided earlier, and their division was restricted to the light period. In contrast, the high PUB:PEB Synechococcus strain WH 8103 divided preferentially at night. There was a weak linear relationship between the FALS_max:FALS_min ratio and growth rate calculated from cell counts (r = 0.83, n = 11, P < 0.05). Because of the significance of picoplanktonic populations in marine systems, these results should help to interpret diel variations in oceanic optical properties in regions where picoplankton dominates.     

DIEL IN SITU PICOPHYTOPLANKTON CELL DEATH CYCLES COUPLED WITH CELL DIVISION1

The diel variability in picophytoplankton cell death was analyzed by quantifying the proportion of dead cyanobacteria Prochlorococcus and Synechococcus cells along several in situ diel cycles in the open Mediterranean Sea. During the diel cycle, total cell abundance varied on average 2.8 ± 0.6 and 2.6 ± 0.4 times for Synechococcus and Prochlorococcus populations, respectively. Increasing percentages of dead cells of Prochlorococcus and Synechococcus were observed during the course of the day reaching the highest values around dusk and decreasing as the night progressed, indicating a clear pattern of diel variation in the cell mortality of both cyanobacteria. Diel cycles of cell division were also monitored. The maximum percentage of dead cells (Max % DC) and the G2 + M phase of the cell division occurred within a period of 2 h for Synechoccoccus and 4.5 h for Prochlorococcus, and the lowest fraction of dead cells occurred at early morning, when the maximum number of cells in G1 phase were also observed. The G1 maximum corresponded with the maximal increase in newly divided cells (minimum % dead cells), and the subsequent exposure of healthy daughter cells to environmental stresses during the day resulted in the progressive increase in dying cells, with the loss of these cells from the population when cell division takes place. The discovery of diel patterns in cell death observed revealed the intense dynamics of picocyanobacterial populations in nature.     

A STUDY OF LIGHT INTENSITY,PERIODICITY, AND WAVELENGTH ON ZOOSPORE PRODUCTION BY PROTOSIPHON BOTRYOIDES KLEBS12

When mature Protosiphon cells were placed in darkness, zoospore production was more extensive and was completed in a shorter time at a temperature of 27 C than at 22 or 15 C. Cool-white fluorescent (Sylvania) light inhibited the process measurably at a radiation intensity of 0.6±10³ ergsjcm²-sec; inhibition was 96% complete at 14±10³ ergs/cm²-sec. For mature cells previously grown under repeated 12-12 hr light-dark cycles, a dark period of approximately 2 hr at 22 C allowed cell division to proceed to a stage such that reillumination did not inhibit continued development of zoospores. Monochromatic light from 402 to approximately -494 nm, as compared to darkness, inhibited zoospore formation; maximal inhibition was at 432-461 nm. In contrast, monochromatic light from 522 to 726 nm stimulated zoospore formation relative to darkness. Synchronous zoospore production was obtained using the following regimes: (A) 12 hr cool-white alternated with 12 hr yellow, (B) 12 hr cool-white alternated with 12 hr blue. Under regime A synchronous zoospore release (following synchronous production) occurred near the end of the yellow irradiation period, while under regime B it occurred near the end of the cool-white irradiation period. The significance of this in terms of photoprocesses and possible photoreceptors is discussed.     

A study of spatial and temporal variability in the adenine nucleotides of lake phytoplankton during a diel cycle (Lac Pavin — France)

A study of diel variations in the adenine nucleotide concentrations of natural phytoplanktonic populations led to the following conclusions:There are significant diel variations in cellular ATP concentrations. However, this temporal variability is not as great as the spatial variability between the various sampling depths, characterized by different phytoplankton populations. Within a globally stable pool of adenine nucleotides, diel variations inside the algal cell mainly take place at the expense of ATP and AMP, while ADP concentrations remain relatively stable.The statistical relationship between algal biomass obtained from cell counts and from adenine nucleotides also confirms maximum stability for ADP and (ATP + ADP + AMP). The diel variations in adenine nucleotide cell concentrations gradually become smaller with depth. This probably reflects the importance of light in causing such variability.     

PHOTOCONTROL OF THE ORIENTATION OF CELL DIVISION IN ADIANTUM. I. EFFECTS OF THE DARK AND RED PERIODS IN THE APICAL CELL OF GAMETOPHYTES

When protonemata of Adiantum capillus-veneris L. which had been grown filamentously under continuous red light were transferred to continuous white light, the apical cell divided transversely twice, but the 3rd division was longitudinal. An intervening period of darkness lasting from 0 to 90 hr either between the 1st and the 2nd cell division or between the 2nd and the 3rd one did not affect the number of protonemata in which the 3rd cell division was longitudinal. The insertion of red light instead of darkness greatly decreased the percentage of 1st longitudinal divisions occurring at the 3rd division, and increased the number of transverse divisions. Fifty percent reduction of induction of 1st longitudinal division was caused by ca. 50 hr exposure to red light between 1st and 2nd division and by ca. 20 hr between 2nd and 3rd division, and total loss was induced by an exposure of ca. 100 hr or longer to red light in the former and by ca. 40 hr longer in the latter. Thus, by using an appropriate intervening dark period or exposure to red light, the orientation and timing of cell division could be controlled in apical cell of the fern protonemata.     

SPONTANEOUS AND STIMULATED BIOLUMINESCENCE OF THE DINOFLAGELLATE CERATOCORYS HORRZDA (PERIDINIALES)1

This is the first report of spontaneous bioluminescence in the autotrophic dinoflagellate Ceratocorys horrida von Stein. Bioluminescence was measured, using an automated data acquisition system, in a strain of cultured cells isolated from the Sargasso Sea. Ceratocorys horrida is only the second dinoflagellate species to exhibit rhythmicity in the rate of spontaneous flashing, flash quantum flux (intensity), and level of spontaneous glowing. The rate of spontaneous flashing was maximal during hours 2–4 of the dark phase [i.e. circadian time (CT)16–18 for a 14:10 h LD cycle (LD14:10)], with approximately 2% of the population flashing-min^?1, a rate approximately one order of magnitude greater than that of the dinoflagellate Gonyaulax polyedra. Flash quantum flux was also maximal during this period. Spontaneous flashes were 134 ms in duration with a maximum flux (intensity) of 3.1×10⁹ quanta-s^?1. Light emission presumably originated from blue fluorescent microsources distributed in the cell periphery and not from the spines. Values of both spontaneous flash rate and maximum flux were independent of cell concentration. Isolated cells also produced spontaneous flashes. Spontaneous glowing was dim except for a peak of 6.4× 10⁴quanta-s^?1 cell^?1, which occurred at CT22.9 for LD14:10 and at CT22.8 for LD12:12. The total integrated emission of spontaneous flashing and glowing during the dark phase was 4×10⁹ quantacell^?1, equivalent to the total stimulable luminescence. The rhythms for C. horrida flash and glow behavior were similar to those of Gonyaulax polyedra, although flash rate and quantum flux were greater. Spontaneous bioluminescence in C. horrida may be a circadian rhythm because it persisted for at least three cycles in constant dark conditions. This is also the first detailed study of the stimulated bioluminescence of C. horrida, which also displayed a diurnal rhythm. Cultures exhibited >200 times more mechanically stimulated bioluminescence during the dark phase than during the light phase. Mechanical stimulation during the dark phase resulted in 6.7 flashes. cell^?1; flashes were brighter and longer in duration than spontaneous flashes. Cruise-collected cells exhibited variability in quantum flux with few differences in flash kinetics. The role of dinoflagellate spontaneous bioluminescence in the dynamics of near-surface oceanic communities is unknown, but it may be an important source of natural in situ bioluminescence.     

Effect of temperature on circadian rhythm controlling the crepuscular activity of the burying beetle Nicrophorus quadripunctatus Kraatz (Coleoptera: Silphidae)

Locomotor activity rhythm was examined at various temperatures under a 16 h light : 8 h dark photoperiod (LD 16:8) or LD 12:12 using adults of the burying beetle Nicrophorus quadripunctatus. At 20°C, the locomotor activity of the beetles showed a bimodal daily pattern with two peaks around lights on and lights off under both photoperiods. This bimodal activity rhythm persisted under constant darkness; therefore, the activity of adult N. quadripunctatus is controlled by a circadian clock. Adults showed a bimodal activity pattern for temperatures ranging from 15 to 25°C. The evening peak of the daily activity was earlier at lower temperatures. These findings suggest that in the field, N. quadripunctatus shows crepuscular activity, and is active earlier in the afternoon in cooler seasons. In this species, therefore, temperature appears to play an important role in the determination of daily activity patterns.     

Locomotor activity rhythm in four wrasse species under varying light conditions

Under controlled laboratory conditions, the locomotor activity rhythms of four species of wrasses (Suezichthys gracilis, Thalassoma cupido, Labroides dimidiatus andCirrhilabrus temminckii) were individually examined using an actograph with infra-red photo-electric switches in a dark room at temperatures of 21.3–24.3°C, for 7 to 14 days. The locomotor activity ofS. gracilis occurred mostly during the light period under a light-dark cycle regimen (LD 12:12; 06:00-18:00 light, 18:00-06:00 dark). The locomotor activity commenced at the beginning of the light period and continued until a little before the beginning of dark period. The diel activity rhythm of this species synchronizes with LD. Under constant illumination (LL) this species shows distinct free-running activity rhythms varying in length from 23 hrs. 39 min. to 23 hrs. 47 min. Therefore,S. gracilis appears to have a circadian rhythm under LL. However, in constant darkness (DD), the activity of this species was greatly suppressed. All the fish showed no activity rhythms in DD conditions. After DD, the fish showed the diel activity rhythm with the resumption of LD, but this activity began shortly after the beginning of light period. The fish required several days to synchronize with the activity in the light period. Therefore,S. gracilis appeared to continue the circadian rhythm under DD. InT. cupido, the locomotor activity commenced somewhat earlier than the beginning of the light period and continued until the beginning of the dark period under LD. The diel activity rhythm of this species synchronizes with LD. Under LL, four of the five specimens of this species tested showed free-running activity rhythms for the first 5 days or longer varying in length from 22 hrs. 54 min. to 23 hrs. 39 min. Although the activity of this species was suppressed under DD, two of five fish showed free-running activity rhythms throughout the experimental period. The lengths of such free-running periods were from 23 hrs. 38 min. to 23 hrs. 50 min. under DD. Therefore, it was ascertained thatT. cupido has a circadian rhythm. InL. dimidiatus, the locomotor activity rhythm under LD resembled that observed inT. cupido. The diel activity rhythm of this species synchronizes with LD. Under LL, four of seven of this species showed free-running activity rhythms throughout the experimental period. The lengths of such free-running periods were from 23 hrs. 07 min. to 25 hrs. 48 min. Although the activity of this species was suppressed under DD, three of five fish showed free-running activity rhythms throughout the experimental period. The lengths of such free-running periods were from 23 hrs. 36 min. to 23 hrs. 41 min. under DD. Therefore, it was ascertained thatL. dimidiatus has a circadian rhythm. Almost all locomotor activity of C.temminckii occurred during the light period under LD. The diel activity rhythm of this species coincides with LD. Under LL, two of four of this species showed free-running activity rhythms throughout the experimental period. The lengths of such free-running periods were from 23 hrs. 32 min. to 23 hrs. 45 min. Although the activity of this species was suppressed under DD, one of the four fish showed free-running activity rhythms throughout the experimental period. The length of the free-running period was 23 hrs. 21 min. under DD. Therefore,C. temminckii appeared to have a circadian rhythm. According to field observations,S. gracilis burrows and lies in the sandy bottom whileT. cupido, L. dimidiatus, andC. temminckii hide and rest in spaces among piles of boulders or in crevices of rocks during the night. It seems that the differences in nocturnal behavior among the four species of wrasses mentioned above are closely related to the intensity of endogenous factors in their locomotor activity rhythms.     

PRESENCE OF CIRCADIAN RHYTHMS IN THE LOCOMOTOR ACTIVITY OF A CAVE-DWELLING MILLIPEDE GLYPHIULUS CAVERNICOLUS SULU (CAMBALIDAE,SPIROSTREPTIDA)

The locomotor activity of the millipede Glyphiulus cavernicolus (Spirostreptida), which occupies the deeper recesses of a cave, was monitored in light-dark (LD) cycles (12h light and 12h darkness), constant darkness (DD), and constant light (LL) conditions. These millipedes live inside the cave and are apparently never exposed to any periodic factors of the environment such as light-dark, temperature, and humidity cycles. The activity of a considerable fraction of these millipedes was found to show circadian rhythm, which entrained to a 12:12 LD cycle with maximum activity during the dark phase of the LD cycle. Under constant darkness (DD), 56.5% of the millipedes (n = 23) showed circadian rhythms, with average free-running period of 25.7h ± 3.3h (mean ± SD, range 22.3h to 35.0h). The remaining 43.5% of the millipedes, however, did not show any clear-cut rhythm. Under DD conditions following an exposure to LD cycles, 66.7% (n = 9) showed faint circadian rhythm, with average free-running period of 24.0h ± 0.8h (mean ± SD, range 22.9h to 25.2h). Under constant light (LL) conditions, only 2 millipedes of 11 showed free-running rhythms, with average period length of 33.3h ± 1.3h. The results suggest that these cave-dwelling millipedes still possess the capacity to measure time and respond to light and dark situations. (Chronobiology International, 17(6), 757–765, 2000)     

SYNCHRONIZATION OF CHLOROPLAST DIVISION IN THE ULTRAMICROALGA CYANIDIOSCHYZON MEROLAE (RHODOPHYTA) BY TREATMENT WITH LIGHT AND APHIDICOLIN

A system of highly synchronized chloroplast divisions was developed in the unicellular red alga Cyanidioschyzon merolae De Luca, Taddei, & Varano. Chloroplast divisions were examined by epifluorescence microscopy following treatments with light and inhibitors. When the cells during stationary phase were transferred into a new medium under a 12:12 h LD cycle, chloroplasts, mitochondria, and cell nuclei divided synchronously in that order soon after the initiation of dark periods. More than 40% of the cells contained dividing chloroplasts. To obtain a system of highly synchronized cell division and chloroplast division, the cells synchronized by a 12:12 h LD cycle were treated with various inhibitors. Nocodazole and propyzamide did not affect cell and organelle divisions, whereas aphidicolin markedly inhibited cell-nuclear divisions and cytokinesis and induced a delay in chloroplast division. More than 80% of the cells contained dividing chloroplasts when cells synchronized by light were treated with aphidicolin for 12 h. This synchronized system will be useful for studies of the molecular and cellular mechanisms of organelle divisions .     

BEHAVIOR OF CHLOROPLAST NUCLEOIDS DURING THE CELL CYCLE OF CHLAMYDOMONAS REINHARDTII (CHLOROPHYTA) IN SYNCHRONIZED CULTURE1

Cells of Chlamydomonas reinhardtii Dangeard were synchronized under a 12:12 h light: dark regimen. They increased in size during the light period, while nuclear division, chloroplast division and cytokinesis occurred during the dark period. Zoospores were liberated toward the end of the dark period. Changes in profile and distribution of chloroplast nucleoids were followed with a fluorescence Microscope after fixation with 0.1%(w/v) glutaraldehyde followed by staining with 4′.6-diamidino-2-phenylidole (DAPI), a DNA fluorochrome. About ten granular nucleoids were dispersed in the chloroplast at the beginning of the light period (0 h). Within 4 h the nucleoids aggregated around the pyrenoid giving a compact profile. The formation of the compact aggregate of cp-nucleoids around the pyrenoid occurred with maximal frequency twice during the light period. Toward the end of the light period the nucleoids were transformed into the form of threads interconnected with fine fibrils spreading throughout the chloroplast. Initially the thread-like nucleoids fluoresced only faintly. The fluorescence of some parts of the threadlike form became brighter over a period of 6 h; these nucleoids were divided into daughter chloroplasts during chloroplast division. Soon after chloroplast division, these thread-like nucleoids were transformed into about 20 granular forms, which were gradually combined to form about ten larger granular bodies in zoospores immediately prior to liberation from mother cells. Fixation of cells with glutaraldehyde at high concentrations or treatment of cells with protease significantly modified the profiles of DAPI-stained nucleoids. The different morphologies of chloroplast nucleoids are discussed in relation to changes in configuration of their protein components.     

Application of the frequency of dividing cells technique to estimate the in situ growth rate of Microcystis (Cyanobacteria)

1. Diel patterns of the frequency of dividing cells (FDC) of the bloom‐forming cyanobacteria Microcystis were investigated using both culture strains and natural populations. 2. In laboratory experiments, diel division cycles were examined twice in a 24‐h light/dark cycle during time‐course batch incubations of six culture conditions using two strains (morphospecies) of Microcystis (M. aeruginosa and M. wesenbergii). While both strains clearly showed phased cell division in the light period during the logarithmic growth phase, the peaks of FDC became unclear towards the stationary phases. Some dividing cells were always found in the dark period regardless of whether or not division had paused at the same time. 3. This result implied the inadequacy of applying the model of McDuff & Chisholm [Limnology and Oceanography (1982) vol. 27 , pp. 783–787] directly to calculate the duration of cell division. Modified equations are proposed to calculate the duration of cytokinesis as a terminal event, in which the FDC values at night are regarded as 0% and all FDC values are subtracted by the minimum FDC value. 4. The diel FDC in natural populations of M. aeruginosa and M. wesenbergii were examined at five sites from a harbour to several distances offshore in Lake Biwa. While both species showed phased cell division patterns in the daytime at the harbour, no peaks in FDC were discernible in the samples taken from the offshore sites. These results strongly suggested that Microcystis growth was higher inshore than offshore. The in situ growth rates were estimated using the new equations.