Ribosome Production and Protein Synthesis in the Preimplantation Rabbit Embryo

Biochemical and radioautographic data show that protein synthesis is increased markedly at the morula stage of rabbit development (60 h embryo). In the late morula an increase in cytoplasmic ribosomes is observed, suggesting that ribosome availability may be rate-limiting for protein synthesis during cleavage. Incorporated ³H-amino acids become highly localized within the nucleoli of late morulae which have been pulse-labelled for 10 min. This localization suggests that ribosomal protein synthesis is increased at the same time as ribosomal RNA synthesis has been shown to increase. Changes in both the incorporation of ³H-amino acids and cytoplasmic ribosome density were found to occur 'synchronously' in all embryonic cells during the cleavage and early blastocyst period (84 h of development). Between 84 h and 108 h, considerable differences in the number of ribosomes per unit area of cytoplasm become apparent among the cells of the blastocyst.     

STUDIES ON DROSOPHILA EMBRYONIC CELLS IN VITRO II. TISSUE- AND TIME-SPECIFICITY OF A LETHAL GENE, DEEP ORANGE

Embryonic cells which were homozygous or hemizygous for an X -linked (1-0.3) recessive lethal gene, deep orange ( dor ), were cultured in T-5 flasks. Muscle cells, epithelial cells, fibroblastic cells and nerve cells were maintained in a functionally active state for longer than the effective lethal phase of the embryos. Some defects in syncytium formation of muscle cells, formation of cellular spheres, and droplet formation on nerve fibers were observed. Addition of an extract of unfertilized wild-type eggs to the culture medium resulted in repair of these defective characters of dor embryonic cells.
To examine the time specificity of the paternal dor ⁺ gene, extracts of embryos obtained from matings of dor sn ³/ dor sn ³ females and dor ⁺ sn ³⁺/ Y males were tested for their ability to repair the defects of dor embryonic cells. The extract from embryos at the stage of blastoderm formation was not effective in repairing dor defects, but extracts from embryos at stages after gastrulation were effective for repair of various defects of dor embryonic cells. Various substances in the pteridine metabolic pathway were tested for their ability to repair defects of dor embryonic cells. The effective lethal phase of dor embryos and the pleiotropic effects of the dor gene are discussed.     

Polypeptide Elongation Factors of the Developing Chick Brain

Abstract: The polypeptide elongation factors (EF-1_L, EF-1_H, and EF-2) of the developing chick brain were separated and purified by means of a combination of gel chromatographic methods. The molecular weight of EF-1_H of the chick brain ranged from 5 to 10 × 10⁵, and was different from that of the chick liver (about 7 × 10⁵). The molecular weight of other purified factors was about 5 × 10⁴ for EF-1_L. and 9.4 × 10⁴ for EF-2. High activities of polyphenylalanine (poly-Phe) synthesis per mg protein in the developing chick brain were observed between the 3rd embryonic week and the 1st post-hatch week and declined afterwards. On the other hand, the levels of both EF-1 and EF-2 per mg protein in the brain were observed to be high in an early embryonic stage, gradually declining afterwards to the adult level. The brain EF-1_L was a major component of EF-1 in an early embryonic stage, while EF-1_H became recognizable in the 3rd embryonic week. Moreover, the EF-1_H activities were found to be more than double with regard to the binding reaction and to be more than 10-fold as active in respect to poly-Phe synthesis in comparison with the activities of EF-1_L. It is proposed that the brain EF-1_H could be due to aggregates consisting of EF-1_L, a stimulatory factor, and other components.     

Morphological development and allometric growth patterns in the juvenile seahorse Hippocampus kuda Bleeker

The morphological development and allometric growth patterns in the juvenile spotted seahorse Hippocampus kuda were studied under hatchery rearing conditions. Newborn spotted seahorses [mean ± s.d . standard length ( L _S) 9·33 ± 0·79 mm] were raised till the age of 124 days (119·35 ± 6·04 mm). Growth was characterized by three stages with two inflexion points occurring at day 21 and 76. The mean growth rates in the first, second and third stages were 0·68, 1·16 and 0·71 mm day⁻¹, respectively. The growth rate was most rapid in the second stage and was probably influenced by a behavioural shift from pelagic to benthic form. The mass ( M ) and L _S relationship was exponential ( M = 7·14 × 10⁻⁶ L _S^2·76), but the slope, b = 2·76, reflected negative allometric growth. Sexes could be distinguished at c. 110 days, and the sex ratio was unbiased. The L _S in males and females did not differ significantly. Morphological stageing series is proposed, which divides H. kuda juvenile development into eight stages based on the development of coronet, cheek and eye spines, keel and pigmentation. The morphometric ratios for all the body parts, except trunk length, showed considerable changes at a transition point occurring at c. 25 mm L _S. The high proportional growth in head length, head depth, pectoral fin base length, dorsal fin base length, snout length, snout depth and eye diameter at the initial stages, and the abrupt increase in tail length only after the first 2 weeks, possibly reflect development priorities during early development where important organs are being developed first for the enhancement of juvenile survival.     

Surface phenotype and rapid quantification of blood dendritic cell subsets in the rhesus macaque

Background  The study of dendritic cell (DC) biology in the rhesus macaque is becoming increasingly important but is limited by incomplete characterization and the lack of a rapid assay to quantify cells.
Methods  We characterized the surface phenotype of myeloid (mDC) and plasmacytoid DC (pDC) subsets in healthy rhesus macaque blood and developed a flow cytometry-based assay for absolute DC determinations.
Results  Rhesus CD11c⁺ mDC were CD16⁺ CD11b⁺ CD56^lo CD8⁻ CD1c⁻ whereas CD123⁺ pDC lacked expression of these markers. Precise DC determinations were performed using a rapid two-step assay combining the analysis of whole blood and peripheral blood leukocytes (PBL).
Conclusions  Antibodies to CD11b, CD56 and CD16 must be omitted from the lineage antibody cocktail to prevent inadvertent gating-out of DC when analyzing rhesus blood. The combined whole-blood/PBL quantification assay will be invaluable for the rapid and repeated monitoring of blood DC counts in this species.     

Effect of water-borne zinc on osmoregulation in the freshwater amphipod Gammarus pulex (L.) from populations that differ in their sensitivity to metal stress

1. Although there is some evidence that exposure to heavy metals can disrupt osmoregulation in crustaceans, most studies have been carried out on relatively pollution-tolerant, marine or estuarine species. Consequently the effects of water-borne zinc (Zn) on osmoregulation by the freshwater amphipod, Gammarus pulex (L.), from two populations that differ in their heavy metal sensitivity, have been compared.
2. 'Clean' site animals (Clowne, Derbyshire) exhibited a marked haemoconcentration (after 4 days at 37·0 μmol Zn l^–1, 5 days at 18·2 μmol Zn l^–1) shown by an increase in haemolymph osmotic pressure (OP_h) and [Na⁺] and [K⁺]. However, after 5 days at 37·0 μmol Zn l^–1, haemolymph of survivors exhibited an OP_h significantly less than controls. 'Contaminated' site animals showed a reduction in OP_h (but not ions) only after 5 days at 76·2 μmol Zn l^–1.
3. There were differences in the threshold and nature of osmoregulatory response to Zn between animals from 'clean' and 'contaminated' sites, but only at concentrations in excess of those (a) known to affect growth and reproduction in 'clean' site animals and (b) occurring at the 'contaminated' site. Clearly population differences in physiological capacity and tolerance do exist but their ecological significance is unclear.     

Achromosomal Division of Early Starfish Embryos Cultured in the Presence of Actinomycin D

Embryos of the starfish Asterina pectinifera were examined for their ability to undergo the early events of embryonic development in the presence of actinomycin D, a most widely used inhibitor of RNA synthesis. Fertilized eggs continued to divide eight or nine times in the presence of 25 μg ml⁻¹actinomycin D, although delay of development was observed. Chromatin disintegrated in the blastomeres of actinomycin D-treated embryos specifically at the 32-cell stage and the nucleus was undetectable at later stages. Before the 32-cell stage, RNA synthesis was not affected by the presence of actinomycin D whereas DNA synthesis was severely inhibited. The stage when achromosomal divisions cease and embryos begin to die corresponds to the period just before onset of blastulation, suggesting that the presence of the nucleus and chromosomes is a prerequisite for blastula formation and development beyond the 512-cell stage in this species.     

Geographical variation in egg cold hardiness: a study on the adaptation strategies of the migratory locust Locusta migratoria L.

Abstract. 1. For many species of insect, cold hardiness is an important trait that enables a population to develop in the next season and to extend its range. To elucidate the role of cold hardiness of the migratory locust Locusta migratoria L. in its outbreak and distribution areas, egg cold hardiness was examined in locusts derived from four locations from latitude 18°23'N to latitude 41°10'N in eastern China.
2. The supercooling points of eggs from different geographic populations did not differ significantly for the first development stage, with an average ± SE of −24.5 ± 0.51 °C, or for the second stage, −22.06 ± 0.68 °C, however there was a significant difference for the embryonic development phase among the four geographical populations. The egg supercooling point increased gradually from neonatal egg to old egg; eggs prior to hatching always had a much higher supercooling point.
3. Comparisons of the cold hardiness of four populations were carried out by validating the close correlation between latitude and the effects of cold on hatching, low lethal temperature (Ltemp₅₀), and low lethal time (Ltime₅₀). There were significant differences among the four populations; the northern population was more cold hardy than the southern population, and the two mid-latitude populations were intermediately cold hardy.
4. The cold hardiness of all populations was enhanced to various degrees by short-term cold acclimation at 0 °C and 5 °C. For most populations, a 2-day acclimation period seemed to be optimal.     

The C-Terminal Domain of the mGluR1 Metabotropic Glutamate Receptor Affects Sensitivity to Agonists

Abstract: The metabotropic glutamate receptor (mGluR) subtype 1 exists as at least three variants (−1a, −1b, and −1c) generated by alternative splicing at the C-terminal domain. Fluorometric Ca²⁺ measurements were used to compare the concentration dependency of agonist-induced rises in intracellular free Ca²⁺ concentration ([Ca²⁺]_i) in human embryonic HEK 293 cells transiently expressing rat mGluR1a, mGluR1b, or mGluR1c. The rank order of agonist potencies was quisqualate ≫ (2 S, 1' S, 2' S )-2-(carboxycyclopropyl)glycine (L-CCG-I) > (1 S, 3 R )-1-aminocyclopentane-1,3-dicarboxylic acid [(1 S, 3 R )-ACPD] and did not differ among the splice variants. However, agonists were consistently more potent at mGluR1a than at mGluR1c and mGluR1b. In the same system, we characterized the agonist pharmacology of two chimeric rat mGluR3/1 receptors where the first and/or the second intracellular loop(s) and the C-terminal domain were exchanged with the corresponding mGluR1a or mGluR1c sequences and that were previously shown to mediate elevations in [Ca²⁺]_i in response to agonists. The potency of agonists was higher at the chimera having the C-terminus of mGluR1a as compared with those having the mGluR1c C-terminus. Both chimeric mGluR3/1 receptors had the same rank order of agonist potencies: L-CCG-I ≫ (1 S, 3 R )-ACPD ∼ quisqualate. These data support the hypothesis that the C-terminal domain of mGluRs plays a role in determining the potency of agonists for inducing mGluR-mediated functional responses.     

Female control of the embryonic environment in a terrestrial amphipod, Mysticotalitrus cryptus (Crustacea)

1. In amphipod crustaceans the ventral chamber plays an integral role in a number of physiological processes and in the female forms the marsupium in which eggs are brooded. The ventral chamber can be viewed as a pre-adaptation to the colonization of land by the family Talitridae. The hypothesis that the female of the terrestrial species, Mysticotalitrus cryptus , can control the osmotic concentration of the marsupial fluid ([MF]) bathing the eggs, thereby buffering the brood from potential physiological stresses presented by the terrestrial environment, is examined.
2. [MF] was maintained significantly higher than the concentration of the external medium ([Medium]) on both dechlorinated tap-water and 400 mOsm kg^–1 media. In each case, [MF] was intermediate to [Medium] and the concentration of the female haemolymph ([Haem]): when [Medium] = 40 mOsm kg^–1, [MF] = 277 mOsm kg^–1 and [Haem] = 590 mOsm kg^–1, respectively, and when [Medium] = 413 mOsm kg^–1, [MF] = 516 mOsm kg^–1 and [Haem] = 722 mOsm kg^–1, respectively.
3. Evidence is provided that females produce urine that is isosmotic with the haemolymph and that the urine is directed, by capillarity, into the marsupium via cuticular channels. It is suggested that this urine plays a role in controlling [MF] in combination with other behavioural mechanisms.
4. Some preliminary observations are presented on the ontogeny of embryonic osmoregulation in M. cryptus which suggest that osmoregulatory ability improves with developmental stage. There is also limited evidence for the ability of the late embryonic stages to hypo-osmoregulate on concentrated media, even though adults lack this capacity.
5. The results are discussed in relation to the colonization of the terrestrial environment by the Talitridae.     

Acute toxicity of copper, zinc and manganese in single and mixed salt solutions to juvenile longfin dace, Agosia chrysogaster

The acute toxicity of copper, zinc and manganese and copper-zinc and copper-manganese mixtures were determined for juvenile longfin dace, Agosia chrysogaster in hard water bioassays (mean=218 mg 1⁻¹ CaCO₃). Copper-zinc was the most lethal toxicant (96-h L.c.₅₀= 0.21 mg 1⁻¹ copper and 0.28 mg 1⁻¹ zinc) and exhibited a more than additive toxicity which was in contrast to the additive toxicity of copper-manganese mixtures (96-h L.c.₅₀= 0.45 mg 1⁻¹ copper and 64.0 mg 1⁻¹ manganese). The toxicity of copper (96-h L.c.₅₀= 0.86 mg 1⁻¹) and zinc (96-h L.c.₅₀= 0.79 mg 1⁻¹) to the fish was similar but both were considerably more lethal than manganese (96-h L.c.₅₀= 130 mg 1⁻¹).     

Real-time quantification of Pseudomonas fluorescens cell removal from glass surfaces due to bacteriophage varphiS1 application

Aims:  To study the efficacy of the lytic phage φS1 in eliminating Pseudomonas fluorescens in the early stage of biofilm formation, using an in situ and real time methodology for cell quantification.
Methods and Results:  Cell adhesion and phage infection studies were carried out in a parallel plate flow chamber under laminar conditions. Cells were allowed to adhere until reaching 1·7–1·8 × 10⁶ cells cm⁻² and phage infection was performed with two different phage concentrations (2 × 10⁹ PFU ml⁻¹ and 1 × 10¹⁰ PFU ml⁻¹). Phage concentration clearly affects the speed of infection. The less concentrated phage solution promoted a three times slower rate of cell removal but did not affect the overall percentage of cell removal. In fact, after a longer infection period the less concentrated phage solution reached the same 93% cell removal value.
Conclusions:  Phages are efficient in the eradication of bacterial cells at the early stage of biofilm formation and their presence at the surface did not allow bacterial recolonization of the surface.
Significance and Impact of the Study:  To date, no published studies have been made concerning in situ and real time quantification of cell removal from surfaces due to phage action.     

Concentrations and annual loading of dissolved organic matter in a small moorland stream

SUMMARY. 1. Dissolved organic matter (DOM) was measured at 8h intervals over a 1 year period in a stream draining 51 ha of moorland with peaty soils.
2. DOM concentrations increased with increasing stream discharge from low flow values of 0–3 mg 1⁻¹ to maximum values of 30 mg 1⁻¹. There were also seasonal differences of up to 13mgl⁻¹ between August (maximum) and February, and differences of about 2.5 mg l⁻¹ between rising and falling stage samples.
3. Seasonal variation was closely related to mean temperature.
4. Total loss over the year was 168 kg ha⁻¹ DOM (84 kg ha⁻¹ C), larger than previous estimates for upland sites.     

Accumulation and Spatial Distribution of Poly(A)+RNA in Oocytes and Early Embryos of Drosophila melanogaster

We describe the accumulation and distribution of poly (A)⁺RNA during oogenesis and early embryogenesis as revealed by in situ hybridization with a radio-labeled poly (U) probe. The amount of poly (A)⁺RNA in nurse cell cytoplasm continuously increased untill mid-vitellogenic stage (st. 10), then decreased with the rapid increase of poly (A)⁺RNA in the oocyte (st. 11). The localization of poly (A)⁺RNA at stage 10 was in the anterior region of the oocyte, where it is connected by cytoplasmic bridge to the nurse cells. These observations indicate that most of the poly (A)⁺RNA synthesized in the nurse cells is transferred to the oocyte through the cytoplasmic bridges at stage 10–11. During the remainder of oogenesis (st. 11–14) and during preblastodermal embryogenesis, poly (A)⁺RNA was evenly distributed over the cytoplasm of oocytes and embryos. At blastoderm stage, poly(A)⁺RNA became concentrated in the peripheral region of embryos. Though the somatic nuclei of the blastoderm contained a detectable amount of poly (A)⁺ RNA, the pole cell nuclei did not. The cytoplasmic RNA visualised by acridine orange staining and the poly (A)⁺RNA detected by hybridization with [³H]poly (U) exhibited identical distributions during oogenesis and early embryogenesis. These observations provide a basis to assess the unique distributions of specific RNA sequences involved in early development.     

Differentiation of 3T3-F442A Cells into Adipocytes is Inhibited by Retinoic Acid

The break of dormancy and the early development of Artemia are known to occur in the absence of any DNA and RNA synthesis. The presence and function of preformed messengers in the developing embryos were studied using ³²PO₄ to track the RNA species that turnover. The rapid labelling of the poly(A) tails of the particulate RNA by ³²PO₄ is found to be the predominant metabolic event accompanying initiation of development. Although these RNA populations represent a meagre percentage of the total poly(A) RNA of the cells, they nevertheless constitute more than 60% of the labelled poly(A) populations at early stages of development. Moreover the rise in the poly(A) RNA levels of the embryos observed during the first four hours of development could be attributed to the increase in the particulate poly(A) RNA. Prelabelled RNA of this fraction remained rather firmly associated with this fraction in chase experiments, indicating that once processed these RNA species function in association with membranes. The observed shift in the size of these RNAs from low to high molecular weight species further implies that they are being activated to take part in the early developmental programme.