Quantitative ultrastructural study of nucleolus-organizing regions at some stages of the cell cycle (G0 period, G2 period, mitosis)

The quantitative characteristics of chromosomal nucleolus-organizing regions (NORs) and some other nucleolar components were studied on ultra-thin sections of pig embryo kidney cells (PK cells). It was shown that: 1) nucleoli-per-cell volumes were 3 times smaller in the G0 period than in the G2 period; 2) the number of fibrillar centers (FCs) per cell in the G0 period, the G2 period, and at metaphase was equal to 7, 33.7, and 8, respectively; 3) mean volumes of individual FCs in the G0 period (0.033 +/- 0.005 micron3), G2 period (0.014 +/- 0.001 micron3), and at metaphase (0.025 +/- 0.002 micron3) were significantly different; 4) the total volumes of FCs calculated per haploid set of chromosomes were practically the same in the G0 (0.105 micron3) and G2 (0.107 micron3) periods, but were twice as large as those at metaphase (0.04-0.05 micron3). These data show that partial activation and inactivation of ribosomal genes in interphase PK cells are not accompanied by a considerable change in the total volume of FCs and may be due to the fragmentation and fusion of individual FCs. Complete inactivation of ribosomal genes in mitosis results in a decrease of total volumes of FCs per cell; 5) in G0 and G2 periods the total volume of the dense fibrillar component per nucleolus is practically proportional to the nucleolus volume (r = 0.99); 6) in the G2 period, the nucleolus volume is also proportional to the number of FCs (r = 0.99; 7) the volume of the dense fibrillar component within individual fibrillar complexes is not a constant one.(ABSTRACT TRUNCATED AT 250 WORDS)     

Morphometric study of nucleolus-organizing regions of chromosomes from swine embryo kidney cells during Go-, G2-period and mitosis

Quantitative characteristics of nucleolus-organizing regions of chromosomes (NORs, or fibrillar centers, FCs) and some other nucleolar components have been studied with the aid of complete series of ultrathin sections of PK-cells. It has been found that: 1) the number of FCs per cell in the G0-period, in the G2-period and at metaphase is equal to 7.0, 33.7 and 8.0, respectively; 2) volumes of individual FCs in the G0-period (0.033 micron 3), G2-period (0.014 micron 3) and at metaphase (0.025 micron 3) are different; 3) the total volume of FCs, calculated for a haploid set of chromosomes, do not differ in the G0 (0.105 micron 3) and G2 (0.107 micron 3) periods, but exceed twice the FCs volume at metaphase (0.04-0.05 micron 3). These data show that the activation and inactivation of ribosomal genes in interphase PK-cells are not accompanied by a change in the total volumes of FCs and are probably connected with the "fragmentation" and fusion of FCs. Complete inactivation of ribosomal genes at mitosis leads to a decrease in the total volumes of FCs; 4) the nucleolus volume is proportional to the volume of the dense fibrillar RNP-component; in the G2-period the nucleolus volume also correlates with the number of FCs (r = 0.99); 5) the volume of the dense fibrillar component within individual fibrillar complexes--the structures corresponding to one nucleolus-organizing region--is not constant. This is an indirect evidence for the differences in the functional activity of NORs of different chromosomes.     

Characterization of Photosystem II Activity and Heterogeneity during the Cell Cycle of the Green Alga Scenedesmus quadricauda

The photosynthetic activity of the green alga Scenedesmus quadricauda was investigated during synchronous growth in light/dark cycles. The rate of O₂ evolution increased 2-fold during the first 3 to 4 h of the light period, remained high for the next 3 to 4 h, and then declined during the last half of the light period. During cell division, which occurred at the beginning of the dark period, the ability of the cells to evolve O₂ was at a minimum. To determine if photosystem II (PSII) controls the photosynthetic capacity of the cells during the cell cycle we measured PSII activity and heterogeneity. Measurements of electron-transport activity revealed two populations of PSII, active centers that contribute to carbon reduction and inactive centers that do not. Measurements of PSII antenna sizes also revealed two populations, PSII_α and PSII_β, which differ from one another by their antenna size. During the early light period the photosynthetic capacity of the cells doubled, the O₂-evolving capacity of PSII was nearly constant, the proportion of PSII_β centers decreased to nearly zero, and the proportion of inactive PSII centers remained constant. During the period of minimum photosynthetic activity 30% of the PSII centers were insensitive to the inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, which may be related to reorganization of the thylakoid membrane. We conclude from these results that PSII does not limit the photosynthetic activity of the cells during the first half of the light period. However, the decline in photosynthetic activity observed during the last half of the light period can be accounted for by limited PSII activity.     

Three-dimensional distribution of Ag-NOR proteins in rat superior cervical ganglia nucleoli according to circadian rhythm

A three-dimensional reconstruction of the distribution of Ag-NOR proteins in nucleoli of sympathetic neurons of a rat killed during the dark period of its light-dark cycle was compared with previously reported analyses on the three-dimensional distribution of fibrillar centers, the high-resolution localization of these proteins, and the morphometric results. The domain occupied by these proteins appeared to far exceed that of the fibrillar centers and included the dense fibrillar RNP component. In the present material this component in turn provided partial bridging between the units consisting of the fibrillar centers plus their surrounding dense fibrillar component.     

Nuclear ultrastructure of the oocytes from human antral follicles. Formation of the karyosphere

The organization of the nucleus in the oocytes from human antral follicles was examined at the electron microscopic level. At this time all the chromosomes are aggregated around an inactivated nucleolus forming a karyosphere 5-7 micron in diameter. The nucleolus bears no granular component and consists of densely packed delicate fibrillar material. The peripheral zone resembling a ring 0.5 micron thick is separated in the nucleolus. Nucleolus-like bodies (NLB), consisting of granules 20 nm in diameter embedded in finely fibrillar material, are constantly observed in contact with the chromatin. The eventually formed karyosphere is a complex of intimately interconnecting structures--the nucleolus, chromosomes and NLB. However, the chromatin surrounding the nucleolus does not form a continuous (compact) mass as it is observed at the light microscopic level. It is determined that the human karyosphere is formed during the preovulatory period when the connection between oocyte and follicular cells of cumulus oophorus is lost. The duration of karyosphere existence in the human oocytes, and relation of the karyosphere to the processes of antral follicle atresia are discussed.     

Light induced larval release of a colonial ascidian

Larval release and photobehavior were studied in the colonial ascidian Polyandrocarpa zorritensis. The test hypothesis was that if larval release is induced by light, then larvae should be attracted to settlement areas where light is sufficient for larval release. Light induced larval release but the time course varied with light intensity. As the intensity of either sunlight or blue-green light decreased (1) the time until the beginning of larval release (latency) became longer, (2) the mean time of larval release increased, and (3) the time interval over which larvae were released increased. The threshold light intensity to induce larval release in blue-green light (8.75x10(12) photons cm(-2) s(-1)) was lower than that in sunlight (3.6x10(13) photons cm(-2) s(-1)). Light induced larval release was not affected by currents up to 15 cm s(-1). Larvae aggregate in light when given a choice between light and dark. This response did not vary with larval age. The lowest light intensity, at which larvae could distinguish between light and dark was 5.0x10(12) photons cm(-2) s(-1) in blue-green light and 2.9x10(14) photons cm(-2) s(-1) in sunlight. Thus, the hypothesis is supported because larvae are attracted to areas where light intensity is sufficient for larval release.     

Proliferation and differentiation sequence of osteoblast histogenesis under physiological conditions in rat periodontal ligament

To define the mechanism of osteoblast histogenesis, nuclear morphometry was utilized as a marker for precursor cell differentiation. One hour after 3H-thymidine injection, groups of 7-week-old rats were killed at hourly intervals over one complete 24-hr photoperiod (LD 12:12). S-phase and mitosis were assessed in autoradiographs of 3-micron sections of molar periodontal ligament (PDL) adjacent to a physiological bone-forming surface. Labeled nuclei were divided into four categories according to morphometry of nuclear size: A (40-79 micron3), B (80-119 micron3), C (120-169 micron3), and D (greater than or equal to 170 micron3) cells. C and D cells synthesize DNA during the light and divide in the following dark phase; the rhythm for A cells is the opposite. B cells demonstrated no preference and were subsequently determined to be nonosteogenic. Compared to A cells the S-phase photoperiod of C and D cells (combined) is approximately a one-to-one reciprocal relationship, suggesting two proliferating progenitors in series. Based on arrest points in the histogenesis sequence, five compartments are defined: 1) A cells, less differentiated, self-perpetuating precursors; 2) A' cells, committed osteoprogenitors; 3) C cells, G1 stage preosteoblasts; 4) D cells, G2 stage preosteoblasts; and 5) Ob cells, morphologically distinct osteoblasts. Minimal elapsed time for the A----A'----C----D----Ob sequence is about 60 hr (five alternating dark/light cycles). A stress/strain-mediated increase in nuclear volume (A'----C) is an important, rate-limiting step in osteoblast differentiation.     

Diurnal variation in uptake and xylem contents of inorganic and assimilated N under continuous and interrupted N supply to Phleum pratense and Festuca pratensis

Compensation by dark-period uptake of NH(4)(+) and NO(3)(-) in the grasses Phleum pratense L. and Festuca pratensis Huds. following N deprivation during the preceding light period was investigated in flowing solution culture under an artificial 10/14 h light/dark cycle. N was supplied as either NO(3)(-), NH(4)(+) or NH(4)NO(3) at 20+/-5 mmol m(-3), available continuously or only during the dark period, for 5-10 d. Intermittent N supply did not affect total daily N uptake, growth rate or net partitioning of dry matter. Net uptake and influx of NO(3)(-) varied similarly throughout the diurnal cycle when NO(3)(-) was supplied continuously, with a marginal contribution by NO(3)(-) efflux. Influx was significantly higher and efflux slightly higher following interruption of NO(3)(-) supply during the light period. Nitrate accounted for 80% of N in xylem exudate except between hours 6-9 of the light period when the amino acid concentration increased 3-fold, primarily as glutamine. Diurnal variation in relative NO(3)(-) uptake exhibited five phases of constant acceleration/deceleration, described reasonably well assuming NO(3)(-) influx was subject to metabolic co-regulation by NO(3)(-) and amino acid levels in the cytoplasmic compartment of the roots. Accordingly, influx is determined by variation in root NO(3)(-) levels throughout the dark period and the first half of the light period, but is down-regulated by increased amino acid levels during the second half of the light period. The sharp light/dark transitions affect transpiration rate and hence xylem N flux which, in turn, affect NO(3)(-) levels in the cytoplasmic compartment of the roots and the rate of NO(3)(-) assimilation in the shoot.     

Coccidia (Apicomplexa: Eimeriidae) from the subterranean rodent Ctenomys opimus Wagner (Ctenomyidae) from Bolivia, South America

Of 35 tuco-tucos (Ctenomys opimus) collected in Bolivia, South America, 31 (88%) had eimerian oocysts in their feces at the time they were examined. Eighteen (58%) of the 31 infected animals were concurrently infected with 2 or 3 eimerian species. Four species of Eimeria were recovered and are described as new species based on the characteristics of sporulated oocysts. Oocysts of Eimeria granifera n. sp. were ellipsoidal, 21.1 x 17.2 (15-26 x 11-20) micron with sporocysts ovoidal, 11.3 x 7.1 (8-14 x 5-9) micron. Oocysts of Eimeria montuosi n. sp. were spheroidal, 24.2 x 22.0 (21-28 x 18-25) micron with sporocysts ovoidal, 10.5 x 7.3 (8-14 x 6-9) micron. Oocysts of Eimeria opimi n. sp. were spheroidal to subspheroidal, 24.3 x 21.8 (18-29 x 15-26) micron with sporocysts ovoidal, 11.6 x 7.6 (10-13 x 6-9) micron. Oocysts of Eimeria oruroensis n. sp. were spheroidal to subspheroidal, 27.3 x 23.6 (23-32 x 20-28) micron with sporocysts ovoidal, 13.2 x 8.6 (10-16 x 8-11) micron.     

The life cycle of Eimeria falciformis var. pragensis (Sporozoa: Coccidia) in the mouse, Mus musculus

The life cycle of Eimeria falciformis var. pragensis, established from a single oocyst, is described in experimentally infected mice (Mus musculus). The coccidium had a prepatent period of 7 days and a patent period of 10--16 days. Oocysts were spherical to ellipsoidal in shape and measured 21.2 x 18.3 micron. Sporulation time was 3 to 3.5 days. Sporocysts measured 12.2 x 7.2 micron and contained a circular to avoid granular sporocyst residuum measuring 5.5 X 5.0 micron. One, 2 or 3 circular to rectangular polar granules were observed within each sporulated oocyst. The endogenous stages developed primarily in the cecum and colon and only occasionally in the lower ileum. Four generations of schizonts were found. Mature 1st-generation schizonts, first observed 48 hr postinfection (PI), measured 17.8 x 12.3 micron and had 12 merozoites that measured 13.3 x 2.0 micron. Mature 2nd-generation schizonts appeared 78 hr PI. They measured 10.2 x 9.3 micron and had 8 merozoites measuring 5.0 x 1.6 micron. Mature 3rd-generation schizonts appeared first at 114 hr PI and measured 17.5 x 10.2 micron and had 10 merozoites that measured 12.4 x 1.8 micron. Mature 4th-generation schizonts appeared first at 144 hr PI. They measured 18.2 x 15.3 micron and had 18 merozoites. The merozoites of the 4th-generation schizont were 4.5 x 1.2 micron. Mature macrogamonts and microgamonts developed simultaneously appearing at 156 hr PI. Macrogamonts measured 16 x 14.5 micron and microgamonts were 18.2 x 15.3 micron. In experimentally infected rats (Rattus norvegicus), development of E. falciformis var. pragensis progressed only as far as mature 1st-generation schizonts.     

Different circadian rhythms regulate photoperiodic flowering response and leaf movement in Pharbitis nil (L.) Choisy

The phase shifting effect of red light on both the leaf movement rhythm, and on the rhythm of responsiveness of photoperiodic flower induction towards short light breaks (10 min red light), has been studied in Pharbitis nil, strain Violet, and comparisons between the two rhythms have been made. The phase angle differences between the rhythms after a phase shift with 2 or 6 h of red light given at different times during a long dark period were not constant. The results indicate the involvement of two different clocks controlling leaf movement and photoperiodic flower induction.Abbreviations DD continuous darkness - l:D x:y light/dark cycles with x hours of light and y hours of darkness - PPR rhythm of photoperiodic responsiveness towards light break     

Interleukin-15 and interleukin-2 enhance non-REM sleep in rabbits

Interleukin (IL)-15 and -2 share receptor- and signal-transduction pathway (Jak-STAT pathway) components. IL-2 is somnogenic in rats but has not been tested in other species. Furthermore, the effects of IL-15 on sleep have not heretofore been described. We investigated the somnogenic actions of IL-15 in rabbits and compared them with those of IL-2. Three doses of IL-15 or -2 (10, 100, and 500 ng) were injected intracerebroventriculary at the onset of the dark period. In addition, 500 ng of IL-15 and -2 were injected 3 h after the beginning of the light period. IL-15 dose dependently increased non-rapid eye movement sleep (NREMS) and induced fever. IL-15 inhibited rapid eye movement sleep (REMS) after its administration during the light period; however, all doses of IL-15 failed to affect REMS if given at dark onset. IL-2 also dose dependently increased NREMS and fever. IL-2 inhibited REMS, and this effect was observed only in the light period. IL-15 and -2 enhanced electroencephalographic (EEG) slow waves during the initial 9-h postinjection period, then, during hours 10-23 postinjection, reduced EEG slow-wave activity. Current data support the notion that the brain cytokine network is involved in the regulation of sleep.     

Effects of total parenteral nutrition on rat bile canaliculi: a stereologic analysis

OBJECTIVE: To quantitate, in a stereologic manner, changes in bile canalicular morphology before and after choleretic infusion of total parenteral nutrition (TPN) and to determine whether TPN produces changes in localized regions within the hepatic lobule. STUDY DESIGN: Livers were obtained from sham-operated on normal adult male rats (control) and from rats that received intravenous TPN solution containing 20% glucose and 3.5% Molipron F. The tissues, obtained by a rigorous sampling procedure, were systematically subjected to stereologic analysis. Measurements were made on electron micrographs at two levels of magnification by point, intersection and profile counts, and then volume, surface area and length were estimated per unit parenchymal volume. RESULTS: The surface area of the canalicular wall per parenchymal volume increased significantly (from 5.33 x 10(-2) to 6.73 x 10(-2) microns 2/micron 3) after TPN treatment, as did the length of microvilli (from 0.241 to 0.267 microns/micron 3). However, the volume of bile canalicular lumina per parenchymal volume (0.306% and 0.320%), total length of bile canaliculi (1.05 x 10(-2) and 1.06 x 10(-2) microns/micron 3) and diameter of microvilli (8.73 x 10(-2) and 8.94 x 10(-2) microns) remained constant. CONCLUSION: These results indicate that changes in canalicular shape and microvillus hypertrophy may cause lowering efficiency of the bile flow rate.     

Diurnal variation in uterine estrogen receptors in immature female rats--inhibition by arginine vasotocin

A diurnal variation in uterine estrogen receptors from immature female rats was observed with the peak occurring during the mid-light phase (noon) of the cycle. Although no dose response was noted, all concentrations of arginine vasotocin ranging from 5 x 10(-7) to 5 x 10(-9) significantly inhibited binding of 3H-estradiol to estrogen receptors in the cytosol fraction of uteri obtained from immature female rats either during the light or dark phase of the photoperiod.     

Hypoxic ventilatory response during light and dark periods and the involvement of histamine H1 receptor in mice

Ventilation oscillates throughout a day in parallel with oscillations in metabolic rate. Histamine affects ventilation and the balance of the energy metabolism via H1 receptors in the brain. We tested the hypothesis that the ventilatory response to hypoxia varies between light and dark periods and that histamine H1 receptors are required for the circadian variation, using wild-type (WT) and histamine H1 receptor knockout (H1RKO) mice. Mice were exposed to hypoxic gas (7% O(2) + 3% CO(2) in N(2)) during light and dark periods. Ventilation initially increased and then declined. In WT mice, minute ventilation (.Ve) during hypoxia was higher in the dark period than in the light period, which was an upward shift along with the baseline ventilation. Hypoxia decreased the metabolic rate, whereas O2 consumption (.VO(2)) and CO(2) excretion were higher in the dark period than in the light period. However, in H1RKO mice, changes in Ve during hypoxia between light and dark periods were minimal, because .Ve was increased relative to .VO(2), particularly in the light period. In H1RKO mice, the HCO(3)(-) concentration and base excess values were increased in arterial blood, and the level of ketone bodies was increased in the serum, indicating that metabolic acidosis occurred. Respiratory compensation takes part in the .Ve increase relative to .VO(2) during hypoxia. These results suggested that changes in .Ve during hypoxia vary between light and dark periods and that H1 receptors play a role in circadian variation in .Ve through control of the acid-base status and metabolism in mice.     

Effect of Low-intensity Red and Far-red Light and High-intensity White Light on the Flowering Response of the Long-day Plant Lemna gibba G3

The long-day plant Lemna gibba L., strain G3 exhibits a relatively low sensitivity to short, white-light interruptions given during the dark period of a short-day cycle. However, the plants are fairly sensitive to low-intensity red light treatments given during a 15-hour dark period on the third day of a 2LD-(9L:15D)-2LD-7SD schedule. Far-red light is almost as effective as red light, and attempts to reverse the red light response with subsequent far-red light treatments have not been successful. Blue light proved to be without effect. When plants were grown on a 48-hour cycle with 15 minutes of red light every 4 hours during the dark period, the critical daylength was reduced from about 32 hours to slightly less than 12 hours.

Continuous red light induced a fairly good flowering response. However, as little as 1 hour of white light each day gave a significant improvement in the flowering response over that of the continuous red light control. White light of 600 to 700 ft-c was more effective than white light of 60 to 70 ft-c. The white light was much more effective when divided into 2 equal exposures given 8 to 12 hours apart. These results suggest an increase in light sensitivity with regard to flower induction about 8 to 10 hours after the start of the light period.

     

The ontogeny of physiological response to light intensity in early stage spiny lobster (Jasus edwardsii) larvae

Early stage phyllosoma larvae of the spiny rock lobster Jasus edwardsii were examined for swimming speed, feeding, oxygen consumption and nitrogen excretion as instantaneous performance indicators when exposed to different irradiance levels. Swimming speed was measured in recently hatched Stage I larvae while all other parameters were measured in larvae from hatch to mid-Stage V. The swimming speed of recently hatched Stage I phyllosoma increased logarithmically between light intensities of 2.9 x 10(14) and 1.8 x 10(16) quanta s(-1) cm(-2) indicating that, within this range, swimming activity was only suppressed at the lowest irradiance level. Larvae examined under dark (no light) conditions showed lower feed intake, oxygen consumption and nitrogen excretion than larvae under low (7.7 x 10(12) q s(-1) cm(-2)) and high (3.9 x 10(14) q s(-1) cm(-2)) light intensities, and this was a consistent pattern observed throughout development from hatch to Stage V. There was no difference in feeding, oxygen consumption and nitrogen excretion between larvae exposed to low and high light intensities. However, from mid-Stage I to mid-Stage V, the metabolic feeding efficiency (feed intake:oxygen consumption ratio) was consistently higher in larvae exposed to low light intensity than in phyllosoma assessed in the dark and under high irradiance. A light intensity of about 7.7 x 10(12) quanta s(-1) cm(-2) and no higher than 3.9 x 10(14) quanta s(-1) cm(-2) is recommended to stimulate feeding and optimise metabolic feeding efficiency in early larval stages of J. edwardsii.     

Laser microdissection of actinomycin D segregated nucleoli

Nucleoli of tissue culture cells were segregated into their fibrillar (light) and granular (dark) components by treatment with actinomycin D. Following this segregation, the cells were treated with quinacrine hydrochloride, an agent which selectively sensitizes the nucleoli to argon laser light. The actinomycin D-segregated, quinacrine-sensitized nucleolar components (dark and light) were selectively irradiated with the laser microbeam and subsequent uridine uptake assayed. The data indicate that selective damage to the light (fibrillar) area is generally more damaging than damage to the dark (granular) area. These results support the idea that DNA is closely associated with the nucleolar fibrillar component.     

An investigation of the presence of ultramicrocells in natural mineral water

The presence of 'ultramicrocells' in natural mineral water, capable of passing through a 0.2 micron filter, has been demonstrated. Filters allowing the greatest proportion of viable (culturable) cells to pass ranked in the order, 0.4 micron polycarbonate (5.02%) > 0.2 micron polycarbonate (0.02%) > or = 0.45 micron cellulose nitrate (0.02%) > 0.2 micron cellulose acetate (< 0.002%). Following incubation for 4 d at 22 degrees C, viable counts in filtered mineral water increased from < 2-8.7 x 10(2) cfu ml-1(-2).8 x 10(4)-1.9 x 10(6) cfu ml-1. Successive filtration/incubation cycles of mineral water increased the proportion of cells passing through a 0.2 micron cellulose acetate filter from < 0.003% to 0.11% and 0.69%, suggesting selection for 'ultramicrocells'. Cells isolated from this process and grown on liquid R2A medium were thin, Gram-negative rods, of 0.15-0.40 micron wide and 0.50-6.20 microns long. Membrane filtration techniques used for pathogen detection in mineral waters will not retain all the cells present. If pathogens are able to form ultramicrocells, these may go undetected.