Cinematographic Observation of “Post-Fertilization waves” (PFW) on the zygote ofXenopus laevis

Summary In fertilizedXenopus eggs, shortly after egg rotation but well before the occurrence of the cleavage-associated surface contraction waves, two circular dark zones originate consecutively from the pigment spot marking the site of sperm entrance. They expand and travel centrifugally over the egg surface.     

Effects of cycloheximide on the “autocatalytic” nature of the maturation promoting factor (MPF) in oocytes of Xenopus laevis

It is shown that a factor arising during the course of maturation in amphibian oocytes is by itself capable of inducing maturation when injected into recipient oocytes even after a series of 10 transfers. The mechanism of action of this phenomenon is shown to be under translational control. Experiments using cycloheximide suggest that MPF does not need protein synthesis for germinal vesicle breakdown (GVBD), but does require a translational step when serially transferred in order to sustain its renewal (“autocatalytic”) capacity. It is further shown that oocytes of Xenopus laevis lend themselves to an in vitro system, since when matured under the action of some steroids (progesterone or hydrocortisone), they are capable of supporting functional maturation with cleavage and development after having received a transplanted blastula nucleus.     

Identification and functional analysis of an ovarian form of the egg activation factor phospholipase C zeta (PLCζ) in pufferfish

Recent studies suggest that egg activation in mammals is triggered by a sperm-specific phospholipase C, PLCzeta. In other vertebrate species such as medaka fish, chickens, and quail, PLCzeta is also expressed as a testis-specific mRNA. Functional studies suggest that PLCzeta plays a similar role as a trigger of egg activation in these species. Here, we report the identification of PLCzeta orthologues in pufferfish species Takifugu rubripes (Fugu) and Tetraodon nigroviridis (Tetraodon). Unexpectedly in these species PLCzeta is expressed not in the testis, but in ovary and brain. Injection of pufferfish PLCzeta copy ribonucleic acid (cRNA) into mouse eggs failed to trigger calcium oscillations, unlike medaka PLCzeta cRNA. Our findings provide the first evidence that PLCzeta may be expressed in the egg, rather than the sperm, in some vertebrate species, and that its mechanism of action and physiologic role at fertilization may differ in different vertebrate species.     

Effects of 17α-ethynylestradiol on sexual differentiation and development of the African clawed frog (Xenopus laevis)

Several studies have shown that exposure of amphibians, including the African clawed frog (Xenopus laevis), to potent estrogens at critical times during development results in feminization and/or demasculinization. However, genotyping of X. laevis has only recently become possible, so studies performed in the past were rarely able to make explicit linkages between genetic and phenotypic sex. Therefore, to further characterize this relationship, X. laevis tadpoles were exposed during development to 0.09, 0.84, or 8.81μg/L 17α-ethynylestradiol (EE2), which is the estrogen analog commonly used in oral contraceptives. Exposure to all concentrations of EE2 tested resulted in significant delays in time to metamorphosis. Genotyping showed that genetic sex ratios were similar among treatments. However, morphological evaluation revealed that a significant number of individuals with a male genotype displayed mixed sex and abnormal phenotypes. Additionally, both genetic males and females exposed to EE2 exhibited greater presence of vitellogenin protein relative to the respective controls. Since estrogens function downstream of the initial molecular signals of sexual differentiation, it is likely that genetic male animals received mixed endogenous male and exogenous female signals that caused disordered sexual development. The production of vitellogenin was probably temporally separated and independent from primary effects on sexual differentiation, and might have contributed to delays in metamorphosis observed in individuals exposed to EE2.     

Cytology of the outer penis epithelium ofBuccinum undatum (L.) from the arctic region — an observation of the imposex phenomenon

TBT (tributyltin)-induced imposex is observed in femaleBuccinum undatum from the arctic region. The fine structure of the male penis epithelium was investigated and the results were compared between specimens from non-polluted (Spitsbergen, Norway) and highly-polluted (Brittany, France) areas. Differences in the arrangement and structure of the apical microvilli-border, the intercellular spaces, RER and SER content, as well as a high rate of parasitism in the tissue of individuals from Brittany can be detected between the species of both areas. The results indicate a higher degree of pathological changes in the penis epithelial cells of individuals from TBT-polluted areas than in those from Spitsbergen.     

The effects of egg yolk,the low density lipoprotein fraction of egg yolk,and three monosaccharides on the survival of cat (Felis catus) spermatozoa stored at 5°C

The survival of cat spermatozoa at 5°C was studied in the presence of egg yolk, the low density fraction (LDF) of egg yolk, glucose, fructose and galactose each made up to the desired concentration in 325 mOsm Tes-Tris buffer at pH 7.5.Increasing the egg yolk concentration (v/v) from 2 to 20% significantly decreased both the percentage of motile cells and the quality of motility score, but did not significantly increase the number of cells staining with a supravital stain. In contrast, increasing concentrations of LDF neither significantly decreased the percentage of motile cells nor increased the number of cells staining with a supravital stain, and significantly increased the motility score. However, the protection provided by the higher concentration of LDF (20%) was no more effective than that provided by the lower concentration of egg yolk (2%). It is contended that the currently high levels of egg yolk (20%) used in the preservation of cat spermatozoa may have contributed to the very poor fertility reported for preserved semen.The effect of glucose, fructose and galactose was examined at concentrations of 0.5 and 1.5 mg/ml. All three sugars significantly decreased the percentage of motile spermatozoa when used at the higher concentration, although for glucose alone the quality of motility was not significantly poorer than that in either the lower concentration or the control.In all experiments, motility declined significantly, and, where it was studied, the number of cells staining with a vital stain increased significantly, as a result of cooling and storage for up to 1 week. There was also significant differences between ejaculates in all experiments.     

Effects of incubation temperature on hatchling phenotypes in an oviparous lizard with prolonged egg retention: are the two main hypotheses on the evolution of viviparity compatible?

Females of several lizard species modify their body temperature during pregnancy, probably in connection with the optimisation of hatchling phenotypes. We studied variations in the temperature selected by gravid females compared with those selected by males and non-gravid females in an oviparous population of Zootoca vivipara (Jacquin, 1797) (Squamata: Lacertidae) of Northern Spain and examined the effects of incubation temperature on the phenotypic variation of hatchlings. Cloacal temperatures of gravid females active in the field were lower than those of males and non-gravid females, as well as the temperatures selected in a thermal gradient created in the laboratory (mean±s.d.: 32.33±1.27 °C for gravid females; 34.05±1.07 °C for males and non-gravid females). Effects of temperature were assessed by incubating eggs at five constant temperatures (21, 25, 29, 32 and 34 °C). Incubation time decreased as temperature increased, following a negative exponential function. Incubation temperatures also affected the hatchlings’ morphology: hatchlings incubated at 34 °C had shorter heads than those from other temperatures. Survival at 34 °C (58%) was significantly lower than at the other temperatures (mean 93%). Pregnant females select lower body temperature, approaching the temperatures that optimise hatchling phenotypes, according to predictions of the maternal manipulation hypothesis on the evolution of viviparity. The shift in preferred temperature by pregnant females would result in only a very short delay, if any, of hatching time and, because the temperature selected by pregnant females is much higher than average temperatures recorded in natural nests of Z. vivipara, egg retention considerably shortens incubation time, according to predictions of the cold-climate hypothesis. Our experimental results indicate that the two main hypotheses on the evolution of viviparity are compatible in our study model.     

The long-term (1979–2005) effects of the North Atlantic Oscillation on wind-induced wave mixing in Loch Leven (Scotland)

We report on long-term covariation (1979–2005) between indices of the North Atlantic Oscillation (NAO) and wind speed and direction in Loch Leven. The effects of the observed variations in wind speed and direction were combined to produce modelled wave-mixed depths (Zc). Positive correlations were observed between seasonal and annual wind speeds and westerly frequency and indices of the NAO that are in line with general perception: positive NAO was correlated with stronger, more westerly winds and these correlations were strongest in winter and spring. Correlations between NAO and estimates of Zc were strongest in the most westerly exposed site in spring (r ² = 0.701; Zc_spring versus spring NAO index). On average, over a 25-year period Zc was deeper in spring and shallower in summer. Major anomalies from the 25-year seasonal means were observed in 1982, 1979, and 1991. Annual average Zc was low in the late 1970s and early 1980s (shallowest average annual Zc of 1.0 m (1984)), high in the late 1980s and early 1990s (deepest average annual Zc of 1.9 m (1990)) and moderate in recent years (up to 2005). This study has major implications for our understanding of potential climate change drivers and the related responses of shallow lake ecosystems, including alterations to littoral habitat quality and benthic–pelagic coupling.     

On the mechanism of activation of protein kinase FA (an activating factor of ATP·Mg-dependent protein phosphatase) in brain myelin

Protein kinase F_A (an activating factor of ATP·Mg-dependent protein phosphatase) has been characterized to exist in two forms in the purified brain myelin. One form of kinase F_A is spontaneously active and trypsin-labile, whereas the other form of kinase F_A is inactive and trypsin-resistant, suggesting a different membrane topography with active F_A exposed on the outer face of the myelin membrane and inactivu F_Q buried within the myelin membrane. When myelin was solubilized in 1% Triton X-100, all kinase F_A became active and trypsin-labile. Phospholipid reconstitution studies further indicated that when kinase F_A was reconstituted in acidic phospholipids, such as phosphatidylinositol and phosphatidylserine, the enzyme activity was inhibited in a dose-dependent manner, suggesting that kinase F_A interacts with acidic phospholipids which inhibit its activity. Furthermore, when myelin was incubated with exogenous phospholipase C, the inactive/trypsin-resistant F_A could be converted to the active/trypsin-labile F_A in a time- and dose-dependent manner. Taken together, it is concluded that membrane phospholipids play an important role in modulating the activity of kinase F_A in the brain myelin. It is suggested that phospholipase C may mediate the activation-sequestration of inactive/trypsin-resistant kinase F_A in the brain myelin through the phospholipase C-katalyzed degradation of acidic membrane phospholipids. The activation-sequestration of protein Kinase F_A may represent one mode of control modulating the activity of kinase F_A in the central nervous system myelin.     

A review on the “in vitro” culture of freshwater mussels (Unionoida)

Many Unionoida are considered to be extinct, endangered, or of special concern. These bivalves have complex life cycle stages that limit successful culture. In nature, the larvae (glochidia) of these bivalves must successfully parasitize a host (mainly fish) in order to metamorphose into juveniles. The two artificial methods used to obtain juvenile freshwater mussels in laboratory are either by induced attachment to host fish or by in vitro culture of glochidia. This article is focused on the in vitro method that represents a novel and alternative process to fish infestation, offering the ability to obtain larger numbers of juveniles without the need for host fishes and reducing the overall costs of propagation. In vitro culture requires a medium which fulfills the nutritional needs of each glochidia species and avoids microbial contamination. Recently, this methodology has presented excellent results with survival and transformation rates up to 94% using host fish plasma. High efficiencies on growth, and survival rates (84%) of juvenile freshwater bivalve Hyriopsis myersiana (Lea, 1856) up to 120 days were obtained when reared in adequate recirculating aquacultural systems using a very specific diet. More research is still needed to demonstrate successful propagation, mainly concerning the media nutritional composition to increase glochidia transformation and juvenile quality.     

Response of the gut neuroendocrine system of Leuciscus cephalus (L.) to the presence of Pomphorhynchus laevis Müller, 1776 (Acanthocephala)

Immunohistochemical tests were applied to sections of intestine of uninfected and Pomphorhynchus laevis Muller-infected chub, Leuciscus cephalus (L.) using 15 different antisera. Nerve cell bodies and fibres immunoreactive (IR) to the anti-bombesin, -Cholecystokinin-8 (CCK-8), -galanin, -Gastrin-Releasing Peptide (-GRP), -Nitric Oxide Synthase (-NOS), -Substance P (-SP), and -Vasoactive Intestinal Peptide (-VIP) sera were observed in the myenteric plexus of uninfected chub. The density of nerve components immunoreactive to these antisera was high in the intestine of the infected fish, especially near the site of attachment. Moreover, numerous nerve fibres, immunoreactive to anti-bombesin, -GRP, -galanin, -SP, and -VIP sera, were encountered in the connective tissue capsule surrounding the bulb and proboscis of P. laevis. The occurrence of P. laevis in the chub gut significantly increased the number of endocrine cells per intestinal fold immunoreactive to galanin, met-enkephalin and leu-enkephalin antisera. CCK-8, Neuropeptide Y and glucagon-like immunoreactive cells were less numerous in the intestine of infected chub. A large number of cells in the tunica propria-submucosa of L. cephalus infected with P. laevis were immunoreactive to anti-serotonin and -leu-enkephalin sera.