Dopaminergic regulation of hatching in fish embryos

Enveloped medaka embryos and denuded zebrafish embryos were exposed to agents that are known to modify the activity of dopaminergic systems. Precocious emergence of medaka embryos was found in the presence of pimozide, salsolinol, and alpha-methyl-rho-tyrosine, whereas delayed hatching occurred with bromocriptine and apomorphine. Moreover, the hatching rate in the light period of medaka eggs, exposed to a 12-hr light/12-hr dark cycle, is significantly higher than in the dark period. Precocious hatching enzyme secretion from denuded zebrafish embryos is caused by salsolinol, whereas dopamine has an opposite effect. At the same time it turned out that in controls hatching enzyme release from denuded zebrafish embryos is well correlated with hatching of enveloped zebrafish embryos. These results do not support the hypothesis proposed by several authors that hatching enzyme is solely mechanically released, but suggest a controlling influence of dopamine receptors, probably located in the developing central nervous system. Assuming a stimulating effect of prolactin on teleostean hatching enzyme secretion, the present data indicate that hypothalamic-hypophyseal tracts are functional at the time of hatching.     

Cardiac rhythms in chick embryos during hatching

Avian embryos develop within a hard eggshell which permits the measurement of heart rate while maintaining an adequate gas exchange through the chorioallantoic membrane. Heart rate has been determined from cardiogenic signals detected either noninvasively, semi-invasively or invasively with various transducers. Firstly, we reviewed these previously-developed methods and experimental results on heart rate fluctuations in prenatal embryos. Secondly, we presented new findings on the development of heart rate fluctuations during the last stages of incubation, with emphasis on the perinatal period, which remained to be studied. Three patterns of acceleration of the instantaneous heart rate were unique to the external pipping period: irregular intermittent large accelerations, short-term repeated large accelerations and relatively long-lasting cyclic small accelerations. Besides these acceleration patterns, respiratory arrhythmia, which comprimised oscillating patterns with a period of 1-1.5 s, appeared during the external pipping period. Furthermore, additional oscillating patterns with a period of 10-15 min were found in some externally pipped embryos.     

Genetic variation in pathogen-induced early hatching of toad embryos

Accelerated hatching is one of few defences available to embryos, and is effective against many egg-stage risks. We present the first analysis of genetic variation in hatching plasticity, examining premature hatching of American toad embryos in response to pathogenic water moulds. We reared eggs from half- and full-sib families in the presence and absence of water mould. Hatching age and hatchling size showed low cross-environment genetic correlations, suggesting that early-induced hatching can evolve largely independently of spontaneous hatching. We found less phenotypic and additive genetic variation for early-induced hatching than spontaneous hatching, and a stronger correlation between egg and induced hatchling sizes. Directional selection by the pathogen may have eroded variation in early-induced hatching, pushing it against the constraint of hatching gland development. Later hatching has a second, muscular component. This pattern of variation may characterize defences based on developmental transitions, although other inducible defences show more variation in induced phenotypes.     

Corticosterone stimulates hatching of late-term tree lizard embryos

The regulation of hatching in oviparous animals is important for successful reproduction and survival, but is poorly understood. We unexpectedly found that RU-486, a progesterone and glucocorticoid antagonist, interferes with hatching of viable tree lizard (Urosaurus ornatus) embryos in a dose-dependent manner and hypothesized that embryonic glucocorticoids regulate hatching. To test this hypothesis, we treated eggs with corticosterone (CORT) or vehicle on Day 30 (85%) of incubation, left other eggs untreated, and observed relative hatch order and hatch time. In one study, the CORT egg hatched first in 9 of 11 clutches. In a second study, the CORT egg hatched first in 9 of 12 clutches, before vehicle-treated eggs in 10 of 12 clutches, and before untreated eggs in 7 of 9 clutches. On average, CORT eggs hatched 18.2 h before vehicle-treated eggs and 11.6 h before untreated eggs. Thus, CORT accelerates hatching of near-term embryos and RU-486 appears to block this effect. CORT may mobilize energy substrates that fuel hatching and/or accelerate lung development, and may provide a mechanism by which stressed embryos escape environmental stressors.     

Cathepsin L involved in the freezing resistance of murine normal hatching embryos and dormant embryos

The cryopreservation of mammalian embryos is an important technology in embryo engineering. The discovery and application of the embryo's own high freezing resistance factors are the main methods to improve the utilization of mammalian embryos in cryopreservation. Cathepsin L gene expression in the frozen and thawed dormant embryos displayed a significant difference from those normal hatched ones. The aim of the present study was to dig out the potential role of Cathepsin L in anti-freezing capacity of murine blastocysts by investigating the location and expression of Cathepsin L in frozen and thawed both activated and dormant hatching blastocysts. Different concentrations of Cathepsin L recombinant protein and E-64d were then respectively added into the embryo cryoprotectant and pre-cryo culture medium. Our results found that down-regulation of Cathepsin L improves the freezing resistance of murine normal hatching embryos by reducing apoptosis. Cathepsin L inhibitors can be used to improve the efficiency of cryopreservation and recovery of blastocysts in vitro. Our study provides a theoretical basis for the further development and application of Cathepsin L.     

Observations on the hatching process and salt and water regulation in the embryos of the freshwater snail Indoplanorbis exustus

The freshwater snail Indoplanorbis exustus laid its eggs in triple layers in a gelatinous, ribbon shaped matrix. Hatching was by mechanical means. The capsular membrane of the embryos was more permeable to water than to other organic and inorganic ions.     

Liposome-mediated gene transfer in fish embryos

Liposome-mediated gene transfer was used to introduce large DNA constructs into zygotes of African catfish. The technique is based on the delivery of recombinant bacteriophage lambda particles (or DNA-protamine complexes) into the cytoplasm of target cells by negatively charged, large unilamellar liposomes. Dechorionated zygotes and early fish embryos were treated with the transforming liposomes. Expression of the introduced reporter genes during the first three weeks of the development of the larvae was followed by measuring the activity of corresponding enzymes. These assays have indicated very efficient DNA uptake into the embryos.     

The functioning of mammalian mitochondria injected into fish embryos

The possibility of mammalian mitochondria functioning in fish embryos has been studied. Suspension of mitochondria isolated from the mouse fibroblast B-82/cap (chloramphenicol-resistant) and B-82 (chloramphenicol sensitive) cell cultures, were injected into the fertilized loach eggs. These embryos with an artificially increased number of mouse mitochondria developed and lived till the larval stages. Activity of cytochrome oxidase in these embryos was 1.5-2 times that in the control several hours after the injection, decreased during development and reached the control level by the gastrula stage. If these embryos with artificially increased number of mouse mitochondria were incubated in presence of chloramphenicol, only embryos that contained mitochondria from chloramphenicol-resistant cells survived, thus suggesting that the injected mitochondria do not degrade but are preserved and function in the cytoplasm of developing loach embryos.     

Live imaging of development in fish embryos

Understanding the cellular and molecular mechanisms that drive the development of embryos requires a detailed knowledge of the way cells divide, move, change shape, interact with one another and die during embryogenesis. Ideally this should be analysed in intact embryos using minimally invasive techniques. Because of their easy accessibility, external development and excellent transparency the teleost embryo has emerged as probably the premier vertebrate model for this type of study. This review will discuss some of the recent advances in this field including attempts to image every cell and their movements during the first 24 h of development as well as other studies that focus on the development of specific organs or high resolution analyses of the behaviour of individual cells.     

Thermally induced torpor in fullterm lizard embryos synchronizes hatching with ambient conditions

Eggs inside an underground nest have limited access to information about above-ground conditions that might affect the survival of emerging hatchlings. Our measurements of heart rates of embryos inside the intact eggs of montane lizards (Bassiana duperreyi, Scincidae) show that low temperatures induce torpor in fullterm embryos, but do not do so during earlier embryogenesis or later, post-hatching. Because above-ground conditions affect soil temperatures, this stage-dependent torpor effectively restricts hatching to periods of high ambient temperatures above ground. Torpor thus can function not only to synchronize activity with suitable environmental conditions during post-hatching life (as reported for many species), but also can occur in embryos, to synchronize hatching with above-ground conditions that facilitate successful emergence from the nest.