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PubMed2000

Effects of X chromosome number and parental origin on X-linked gene expression in preimplantation mouse embryos.

Diploid androgenetic mouse embryos, possessing two sets of paternally inherited chromosomes, and control fertilized embryos were used to examine the relative effects of X chromosome number and parental chromosome origin on androgenone viability and X-linked gene expression. A significant difference in efficiency of blastocyst formation was observed between XX and XY androgenones in some experiments, but this difference was not uniformly observed. Significant effects of both X chromosome number and parental origin on X-linked gene expression were observed. Male and female control embryos expressed the XIST: RNA initially. This expression was followed by a preferential reduction in XIST: RNA abundance in male embryos, indicating that dosage compensation for the X chromosome may normally require the downregulation of XIST: RNA expression in male embryos, in conjunction with the production of stable XIST: transcripts in female embryos. By the late blastocyst stage, XX control embryos expressed significantly more XIST: RNA than did XY embryos. Unlike their normal counterparts, XX androgenones did not express significantly more XIST: RNA than did XY androgenones at the late blastocyst stage. Androgenones exhibited severe repression of the Pgk1 gene, but during development to the late blastocyst stage Pgk1 mRNA expression increased in XX androgenones and decreased in XY androgenones. Thus, the initial repression of the Pgk1 gene in XX androgenones was lost as the XIST: RNA declined in abundance, and this loss was correlated with a failure of XX androgenones to express significantly more XIST: RNA than did XY androgenones. These results indicate that androgenones may lack a factor that is expressed from the maternal genome and required for dosage compensation in preimplantation embryos. The results also indicate that early dosage compensation in preimplantation embryos may normally be reversible, thus providing flexibility to meet different developmental requirements of the embryonic and extraembryonic lineages.

Animals↗
PubMed2001

Ratio of proliferating cell nuclear antigen-positive nuclei to total cell number is higher in day 7 than in day 8 vitrified in vitro-produced bovine embryos.

The aim of the present study was to find a reliable functional criterion for the evaluation of the proliferation potential of bovine in vitro-produced embryos. We used immunocytochemical detection of proliferating cell nuclear antigen (PCNA) combined with propidium iodide (PI) staining and subsequent confocal laser scanning microscopy together with routine morphological evaluation under a stereomicroscope to study fresh Day 7, 8, and 9, and cryopreserved Day 7 and 8 embryos. The ratio of PCNA/PI-positive nuclei was equal in fresh Day 7 and Day 8 embryos and significantly lower in Day 9 embryos. In general, Day 7 embryos tolerated the cryopreservation treatments better than Day 8 embryos. Vitrification in normal straws was especially detrimental to Day 8 embryos. In fresh Day 7 and 8 embryos, the PCNA results were in agreement with stereomicroscopic evaluation. However, in Day 9 fresh and in Day 7 and 8 treated embryos, the missing PCNA revealed disorders that were not observed under morphological evaluation. PCNA immunocytochemistry is an effective method to obtain information about the functional state of nuclei. The ratio of PCNA-positive nuclei can provide more information and numerical data about the developmental potential of bovine embryos after cryopreservation.

Animals↗
PubMed2002

Mosaic gene expression in nuclear transfer-derived embryos and the production of cloned transgenic pigs from ear-derived fibroblasts.

Genetically modified domestic animals have many potential applications ranging from basic research to production agriculture. One of the goals in transgenic animal production schemes is to reliably predict the expression pattern of the foreign gene. Establishing a method to screen genetically modified embryos for transgene expression before transfer to surrogates may improve the likelihood of producing offspring with the desired expression pattern. In order to determine how transgene expression may be regulated in the early embryo, we generated porcine embryos from two distinct genetically modified cell lines by using the nuclear transfer (NT) technique. Both cell lines expressed the enhanced green fluorescent protein (eGFP); the first was a fibroblast cell line derived from the skin of a newborn pig that expressed eGFP, whereas the second was a fetal derived fibroblast cell line into which the eGFP gene was introduced by a retroviral vector. The reconstructed embryos were activated by electrical pulses and cultured in NCSU23. Although the in vitro developmental ability of each group of NT embryos was not different, the eGFP expression pattern was different. All embryos produced from the transduced fetal cell line fluoresced, but only 26% of the embryos generated from the newborn cell line fluoresced, and among those that did express eGFP, more than half had a mosaic expression pattern. This was unexpected because the fetal cell line was not clonally selected, and each cell had potentially different sites of integration. Embryos generated from the newborn cell line were surgically transferred to five surrogate gilts. One gilt delivered four female piglets, all of which expressed eGFP, and all had microsatellites identical to the donor. Here we demonstrate that transgene expression in all the blastomeres of an NT embryo is not uniform. In addition, transgene expression in a genetically manipulated embryo may not be an accurate indicator of expression in the resulting offspring.

Animals↗
PubMed2004

Distribution patterns of immunocompetent cells in the pregnant mouse uteri carrying allogeneic mouse and xenogeneic vole embryos.

Mouse and vole embryos were allogeneically and xenogeneically transferred into pseudopregnant CD-1 and immunodeficient (scid) female mice, and we investigated the distribution of immunocompetent cells, uterine natural killer (uNK) cells, mast cells and macrophages, in the implantation sites on days 6, 7 and 8 of gestation. The survival rate of the vole embryos decreased gradually with increased gestation, but the rate was higher in the scid uteri than in the CD-1 mice. The number of uNK cells increased markedly at the mesometrial triangle and the outer decidual area in the CD-1 uteri containing vole embryos; by contrast, scid uteri having vole embryos showed almost the same number as those having mouse embryos. Mast cells were present in large numbers at the myometrium, but rarely in the decidua in all types of pregnant uteri. Cells at the myometrium were more numerous in xenogeneic than in allogeneic transfer. Many mast cells appeared in the inner decidua where xenogeneically transferred vole embryos were dead and aborted. Macrophages were present in the outer decidua and myometria in all types of pregnant uteri, and their distribution pattern did not change even in aborted uterine sites. These results suggest: (1) the response of macrophages to dead embryos is completely inhibited, (2) uNK cells and mast cells increase near dead and aborted embryos, and (3) the increment in uNK cells responding to xenogeneic embryos is suppressed in scid mice, and the suppression may contribute partly to survival of the embryos.

Animals↗
PubMed2005

The effect of paternal heat stress on protein profiles of pre-implantation embryos in the mouse.

The study was undertaken to compare the protein profiles of [35S]-methionine-labelled control-sired embryos with heat-sired embryos at 7, 14 or 21 days after mature fertile B6CBF F1 male mice were kept at 36 +/- 0.3 degrees C and 62 +/- 2.7% relative humidity for 24 h. One-dimensional gel electrophoresis and autoradiographs were used to examine the protein profiles between the two-cell embryos and the blastocysts. The results obtained demonstrate that paternal heat stress 7 or 14 days earlier did not apparently affect protein patterns of two-cell embryos, four-cell to eight-cell embryos, morulae or blastocysts. However, 21 days earlier, there were changes in protein patterns of two-cell embryos and abnormal embryos, but not the morulae. To further support and extend these results, two-dimensional gel electrophoresis and phosphorimaging were employed and the results obtained show that paternal heat stress 21 days before mating affected protein profiles of two-cell embryos and morulae in the mouse. Together, these findings have indicated that paternal heat stress affects most but not all protein patterns of pre-implantation embryos, which strongly supports our previous results demonstrating that paternal heat stress significantly reduced the developmental proportion of pre-implantation embryos in the mouse.

Animals↗
PubMed1998

TNF-alpha expression in embryos exposed to a teratogen.

PROBLEM: The role of tumor necrosis factor (TNF)-alpha produced by embryonic cells in normal and abnormal development is poorly understood. To assess to what extent TNF-alpha may be involved in the process of induced dysmorphogenesis, the expression of TNF-alpha and TNF-alpha receptor (TNFRI) mRNA as well as TNF-alpha protein was evaluated in embryos responding to a cyclophosphamide (CP)-induced teratogenic insult. The effect of maternal immunostimulation increasing the embryo's tolerance to CP on TNF-alpha expression was also investigated. METHOD OF STUDY: ICR female mice were treated intraperitoneally with 40 mg/kg CP on day 12 of pregnancy. The immunostimulator, xenogeneic rat splenocytes, was injected intrauterine 21 days before mating. Embryos were collected on days 13, 14, or 15 of pregnancy. TNF-alpha mRNA, TNFRI mRNA, and TNF-alpha protein expression were evaluated by in situ hybridization and immunostaining techniques in control, teratogen-treated, and immuno-stimulated teratogen-treated embryos. RESULTS: CP-treated embryos showed severe external brain and craniofacial anomalies already visible on day 14 of pregnancy. TNF-alpha mRNA transcripts were detected in cells of the brain and the head of 13-day embryos, which preceded the occurrence of CP-induced external craniofacial anomalies. On day 15 of pregnancy, when severe craniofacial anomalies increased, a significant increase in the intensity of TNF-alpha, TNFR1 mRNA transcripts, and TNF-alpha protein expression were observed in cells of the malformed regions of the head and the brain. In other nonmalformed organs of CP-treated embryos such as the liver (not macroscopically different from controls), neither TNF-alpha nor TNFR1 transcripts were detected. Immunostimulation substantially diminished the severity of CP-induced brain and craniofacial anomalies, decreased the resorption rate, and was associated with decreased intensity of TNF-alpha mRNA transcripts detected on day 15 of pregnancy in the head and the brain of CP-treated embryos. CONCLUSIONS: TNF-alpha expressed in the embryo may be one of the molecules promoting the formation of CP-induced brain and craniofacial anomalies. The decrease of TNF-alpha expression in embryos of immunostimulated females may be one of the mechanisms responsible for the increased tolerance to the teratogenic insult.

Animals↗
PubMed1999

Interleukin-6 levels in co-culture of human in vitro fertilization embryos with Vero cells are not predictive of future successful development.

PROBLEM: In an attempt to predict successful embryo transfer and implantation, we measured interleukin (IL)-6 levels in culture supernatants of co-cultured preimplantation human embryos. We tested whether all in vitro fertilized human embryos in co-cultures do secrete IL-6, and whether there was any difference in such production between embryos that successfully reached the blastocyst stage and blocked embryos. We also addressed the question of IL-6 secretion by co-culture support cells, namely Vero cells themselves. METHOD OF STUDY: Each fertilized oocyte was cultured individually and transferred in culture wells supplemented with a feeder layer of Vero cells at day 2. In vitro IL-6 production was measured by bioassay of the culture media. RESULTS: Because Vero cells themselves secrete IL-6, it became impossible, in co-culture, to quantify production of IL-6 by the sole embryos. On the other hand, the co-culture technique has shown us that embryos are likely to consume IL-6. There was no difference between blastocysts and blocked embryos. CONCLUSIONS: IL-6 levels in human embryo co-cultures do not correlate with future successful embryo transfer.

Adult↗
PubMed1984

Comparison of properties of calcium channels between the differentiated 1-cell embryo and the egg cell of ascidians.

In the ascidians Halocynthia roretzi and H. aurantium the Ca channels in the differentiated embryo whose cleavage was arrested with cytochalasin B at the 1-cell stage and in the unfertilized egg were studied using the voltage-clamp technique. In the cleavage-arrested 1-cell embryo, which differentiates into a cell of epidermal type after culturing until the time of hatching of the control larvae, Ca channel and Ca-induced K channel currents were observed upon depolarization of the membrane. Inward current through Ca channels in the embryo was analysed after suppressing Ca-induced K current by intracellular injection of EGTA. Sr or Ba ions could substitute for Ca ions as the charge carrier through Ca channels both in the cleavage-arrested embryo and in the egg. The selectivity ratios among these cations at their respective maximum inward currents were 1.0 (Ca):2.0 (Sr):4.5 (Ba) for the Ca channel in the embryo and 1.0 (Ca):1.9 (Sr):1.1 (Ba) for that in the egg. The time course of inactivation of Ca channels in Ca artificial sea water (ASW) was different from that in Sr or Ba ASW in the cleavage-arrested embryo. Fast inactivation was observed only in Ca ASW, and slight and slow inactivation was seen in Ba or Sr solution. In the egg, Ca, Sr and Ba currents through Ca channels all showed a similar time course of inactivation. The time course and voltage dependence of inactivation in Ca ASW were studied by measuring Ca tail current at a constant potential level of -28 mV. In the cleavage-arrested embryo the inactivation became slower and smaller in accordance with the decrease in inward Ca current when the potential level of the command pulse was increased in the positive direction from 10 to 80 mV. In the egg the time course of inactivation became faster when the potential level was similarly increased. The experimental results in (4) and (5) above suggest that the inactivation of the Ca channel in the cleavage-arrested embryo was dependent on Ca inward current while that in the egg was potential dependent. The developmental changes of Ca channels from egg type to epidermal type were studied in the cleavage-arrested 1-cell embryo. The epidermal-type Ca channels appeared at about 40 h after fertilization at 9 degrees C. The Ca channels in those blastomeres which differentiated to a cell of muscular type in the cleavage-arrested 8- or 16-cell embryo were studied after suppressing the outward current by tetraethylammonium and by intracellular injection of both Cs ions and EGTA.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1986

Day of embryo collection, quality and pregnancy rates in cattle.

In 1300 donor cows, total embryos increased from 8.5 to 15.3, (P less than 0.001), the mean number of transferable embryos from 3.1 to 6.5 (P = 0.067) and pregnancies increased from 1.3 to 3.3 (P = 0.584) as the day of collection increased from six to 7.5. Most of the embryos recovered on day 6, 7.5 and 8 were morulae, early blastocysts and late blastocysts respectively. Morulae formed the majority of the embryos collected on days 6 and 6.5. Pregnancy rates in early and late blastocysts were highest on days 7 (54.4 and 60.2 per cent) and 7.5 (53.6 and 53.1 per cent, P = 0.009 and 0.004). There were significant differences in pregnancy rates between embryo stages on days 7 and 7.5 (P less than 0.001), embryo grades on days 7, 7.5 and 8 (P less than 0.001), and within embryo grades 1 (P = 0.015) and 3 and 4 (pooled, P = 0.009). On all days combined there were significant differences in pregnancy rates between both embryo stages (P = 0.007) and embryo grades (P less than 0.001). It appears that the concept of embryo fitness may be applied to blastocysts but not to morulae.

Animals↗
PubMed1988

Embryo transfer: its uses and recent developments.

The technique of embryo transfer is now routinely employed in several species and with several objectives. In laboratory animals the technique is used mainly in investigations of reproductive biology, whereas in human beings it is used to overcome specific forms of impaired fertility. Because embryo transfer combined with superovulation of the donor can significantly increase the female reproductive rate its greatest application to date has been in farm animals, in which it is widely used in both research and commercial production. Within the past few years there have been many advances in the techniques used in farm animals, particularly in the area of embryo manipulation. The supply of embryos can now be increased by repeated superovulation and by embryo bisection and there has been significant progress in in vitro fertilisation technology. Deep freeze (-196 degrees C) storage of embryos is now routine and may be combined with direct one-step thawing and removal of cryoprotectant. This technique allows the routine non-surgical transfer of embryos in the field. There is also evidence that a routine non-traumatic procedure for sexing embryos may soon be developed. Identical multiplets or clones have been produced by the microdissection of embryos and more recently by the transfer of nuclei from embryos into unfertilized oocytes. Transgenic animals have been produced by the microinjection of recombinant DNA into one of the pronuclei of single-cell unfertilized ova.

Animal Husbandry↗
PubMed1995

Relationship between associations of NOR and chromosomal anomalies in the abnormal embryos of nonobese diabetic and STZ-diabetic mouse.

Associations of nucleolar organizing regions (NORs) in the postimplantation stage embryos of nonobese diabetic (NOD) mice, and diabetic ICR mice induced by streptozotocin (ST), were studied to investigate the possible cause of the numerical anomalies of the chromosomes in their abnormal embryos. The incidence of NOR associations in abnormal embryos from diabetic NOD (NOD-DM) and STZ-diabetic mice was 11.7 and 7.7%, respectively. This incidence was significantly higher than that (1.2%, p < 0.05) of normal embryos from ICR mice which were used as control. From the results analyzed cytogenetically it was suggested that the higher incidence of chromosomal numerical anomalies in the embryos from NOD-DM and STZ-diabetic mice were caused by the chromosomal nondisjunction induced by associations of NORs. Furthermore, it was suggested that NOD-DM embryos have a tendency to increase the associations of NOR in a diabetic condition together with other factors such as autoimmune disease, however a diabetic condition alone induced chromosomal anomalies. Regarding relationships between the incidence of associations of NOR and the types of malformed embryos, it was also clear that all of the abnormal embryos from NOD-DM and STZ-diabetic mice had a high incidence of associations of NOR, and that the incidence was not related to the types of congenital anomalies. Furthermore, in the mal-developed tissue of embryos from STZ-diabetic mice, many chromosomal anomalies were found (26.6%), and the incidence was similar to that of whole embryos (22.6%).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Relation of early hemodynamic changes to final cardiac phenotype and survival after neural crest ablation in chick embryos.

BACKGROUND: Microcinephotography was used to study a model of persistent truncus arteriosus created in chick embryos by ablation of premigratory neural crest destined for the third and fourth aortic arch arteries as well as the septum of the cardiac outflow tract. METHODS AND RESULTS: Twenty-five control embryos and 105 of 202 experimental embryos were filmed on day 3 of incubation and then reincubated. The remaining 97 experimental embryos were not filmed because of twisting of the embryos, but they were reincubated. There was no difference in either the survival rate (p greater than 0.23) from day 3 to day 11 of incubation or the incidence of persistent truncus arteriosus (p greater than 0.08) between the filmed and the nonfilmed embryos. Incomplete looping of the cardiac tube observed in experimental embryos during early cardiogenesis correlated with a right ventricular origin of the outflow vessels in the definitive heart. Hemodynamic measurements indicated that there was no difference in heart rate, ejection fraction, systolic and diastolic areas, stroke volume, and cardiac output between controls and the experimental group as a whole. However, embryos that did not survive to day 11 had decreased stroke volume (p less than 0.001) and cardiac output (p less than 0.001), whereas embryos that survived to day 11 with cardiac malformations had increased stroke volume and cardiac output in early embryogenesis. CONCLUSIONS: Increased stroke volume and cardiac output may be necessary factors for survival in embryos with cardiac dysmorphogenesis and probably are associated with dilation of the ventricular portion of the cardiac tube, which leads to malalignment of the outflow vessel or vessels.

Animals↗
PubMed1994

Developmentally regulated expression of the preproinsulin gene in the chicken embryo during gastrulation and neurulation.

Despite the absence of a pancreas, which develops between embryonic day 3 (E3) to E4, previous studies showed that insulin receptors are widely expressed in chicken embryos from the blastoderm stage (unincubated embryo, E0) through gastrulation (E0.5-E1), neurulation (E1.5-E2), and organogenesis. We now characterize prepancreatic preproinsulin gene expression and its regulation, using a highly sensitive modification of the polymerase chain reaction. We found preproinsulin messenger RNA (mRNA) expression at all stages, from the unincubated chicken blastoderm through early organogenesis, with the highest expression in embryos undergoing gastrulation. In situ hybridization analysis of E1-E1.5 embryos in toto showed widespread distribution of preproinsulin mRNA in a pattern similar to that of insulin receptor mRNA. In contrast, insulin-like growth factor-I mRNA expression appeared later than preproinsulin mRNA in the embryo; it was first demonstrable in the head portion of E3 and was found in head, trunk, and caudal regions by E4. With a novel culture system for chicken embryos during neurulation, we examined whether glucose regulated prepancreatic preproinsulin mRNA expression. Embryos cultured in glucose-free medium had increased preproinsulin mRNA with respect to the value in ovo, but the addition of 17 mM glucose had no stimulatory effect. In marked contrast, in organ cultures of E13 pancreas, insulin mRNA expression decreased in glucose-free medium by 50% relative to that in ovo. The addition of glucose restored the levels to a concentration similar to that found in ovo. Exogenous insulin added to cultured E1.5 embryos increased protein and DNA synthesis. We conclude that the preproinsulin gene is widely expressed in chicken embryo structures throughout gastrulation and neurulation. This prepancreatic preproinsulin mRNA is differentially regulated compared to the pancreatic mRNA. Preproinsulin gene products may have a role in cell proliferation, differentiation, or survival in very early avian embryos at a time when insulin-like growth factor-I expression is absent or undetectable.

Animals↗
PubMed1992

Evidence of a role for endogenous electrical fields in chick embryo development.

We have tested directly the hypothesis that the endogenous electrical field in the chick embryo plays a causal role in development. Conductive implants, which shunt currents out of the embryo and thus alter the internal field, were placed under the dorsal skin at the mid-trunk level of stage 11-15 embryos. Currents leaving the posterior intestinal portal (p.i.p.) of these embryos were reduced by an average of 30%. Control embryos receiving non-conductive implants showed no change in p.i.p. currents. In the group receiving current shunts, 92% of the embryos exhibited some developmental abnormality. Only 11% of the control group displayed defects. The most common defect in the experimental group (81%) was in tail development. Tail defects ranged from complete absence to the formation of a normal length, but morphologically abnormal tail. Internally, tail structures (neural tube, notochord and somites) were frequently absent or aberrantly formed. In 33% of the experimental embryos, the notochord continued lengthening in the absence of any other tail development. This led to the formation of ourenteric outgrowths from the hindgut. Defects in limb bud and head development were also found in experimentally treated embryos, but at a much lower frequency than tail defects. The abnormalities observed in experimental embryos were very similar to those produced naturally in rumpless mutant chicks. A vibrating probe analysis of these mutants (from both dominant and recessive strains) showed that currents leaving the p.i.p. were significantly lower in phenotypically abnormal mutants than in wild-type and phenotypically normal mutant embryos from both strains. There was no apparent correlation between the average transepithelial potential (TEP) of these mutants and the development of tail abnormalities. The possible role of endogenous electrical fields in chick tail development is discussed.

Animals↗
PubMed2006

Stepwise in-straw dilution and direct transfer using open pulled straws (OPS) in the mouse: a potential model for field manipulation of vitrified embryos.

In the present study, mouse blastocysts were employed to investigate the feasibility and efficiency of stepwise in-straw dilution and direct transfer using the open pulled straw (OPS) method. In experiment I, the effects of various vitrification solutions (VS) on embryo survival were examined. After thawing, the expanded blastocyst rates (97.59 and 95.05%) and hatching rates (80.48 and 78.95%) achieved in the EDFS30 [15% ethylene glycol (EG), 15% dimethyl sulfoxide (DMSO), Ficoll, and sucrose] and EFS40 [40% EG, Ficoll, and sucrose] groups were no different from those (96.15% and 83.33%) of the control group. However, the rates in the EFS30 [30% EG, Ficoll, and sucrose] (87.80 and 55.43%) and EDFS40 [20% EG, 20% DMSO, Ficoll, and sucrose] (95.69 and 70.97%) groups were significantly lower than those (96.15 and 83.33%) of the control group (P<0.05). In the experiment II, the effects of the volume of VS in the OPS on the survival of embryos after in-straw thawing were investigated. When the length of the VS in the column was less than 1 cm, the in vitro viability of embryos thawed by stepwise in-straw dilution was no different among the experimental and control groups. The embryos could be successfully thawed by immersing the OPS in 0.5 M sucrose for 3 min and then 0.25 M sucrose for 2 min. In experiment III, the effect of immersion time of the OPS in diluent (PBS) on the viability of vitrified embryos was investigated. After in-straw thawing, OPSs were immersed immediately in 1 ml PBS for 0 to 30 min. When the immersion time of the OPSs in PBS was less than 12 min, in vitro development of the in-straw thawed embryos was no different from that of the controls. In experiment IV, in-straw thawed blastocysts were directly transferred to pseudopregnant mice to examine their in vivo developmental viability. The pregnancy (91.67%) and birth rates (42.42%) of embryos in-straw thawed and directly transferred were no different from those of the unvitrified controls (90.90 and 40%) and embryos thawed by the conventional method (84.61 and 46.94%). These results demonstrate that mouse embryos vitrified with OPS could be successfully thawed by stepwise in-straw dilution and transferred directly to a recipient and that this method might be a model for field manipulation of vitrified embryos in farm animals.

Animals↗
PubMed2003

Effects of BSA and fetal bovine serum in culture medium on development of rat embryos.

Rat 1-cell embryos, recovered from naturally mated females, were cultured in a chemically defined medium (mR1ECM) or in mR1ECM supplemented with BSA (4 mg/ml; mR1ECM-BSA) or fetal bovine serum (FBS; 10%, v:v; mR1ECM-FBS) instead of polyvinylalcohol. There was no difference in percentages of embryos that developed to the 2-cell to blastocyst stages between mR1ECM and mR1ECM-BSA, but in mR1ECM-FBS, no development beyond the 2-cell stage was observed. When embryos were transferred to mR1ECM-FBS from mR1ECM after 24 to 64 h of culture, development of embryos to and beyond the 4-cell stage was inhibited. However, when transferred after 80 h of culture, more embryos developed to blastocysts and hatching or hatched blastocysts than in embryos cultured in mR1ECM. When 8-cell embryos and early morulae obtained after 72 and 80 h of culture in mR1ECM, respectively, were cultured in mR1ECM-FBS, a higher proportion of early morulae developed to the blastocyst stage than did 8-cell embryos. When morulae obtained after culture in mR1ECM or mR1ECM-BSA were transferred to recipient females, there was no difference in proportions of fetuses obtained. However, a higher proportion of blastocysts cultured in mR1ECM-FBS developed to fetuses compared with those obtained in mR1ECM. These results indicate that BSA has neither deleterious nor beneficial effects on development of rat 1-cell embryos. In contrast, FBS has deleterious effects on early cleavage of embryos but it promotes more rapid development of morulae to blastocysts, resulting in better quality blastocysts.

Animals↗
PubMed1978

The parental source of heteroploidy in chick embryos determined with chromosomally marked gametes.

Males homozygous for chromosomal translocations were mated to karyologically normal females. The resulting embryos, after incubation for 16-18 h, were prepared for cytogenetic analysis. Of the 62 embryos analysed, 54 (7.8%) were heteroploid or contained a major heteroploid cell line. Each of the 638 normal 2n embryos contained one marker and one normal chromosome, indicating that none arose from gynogenesis. Thirteen homogeneous haploid embryos were identified; 24 embryos were chimaeric 1n/2n and one was 1n/3n. All haploid cell lines contained a marker chromosome indicating androgenetic origins. The 9 homogeneous triploid (3n) embryos and the 3n cell line in a single 1n/3n embryo contained a single marker. All resulted from fertilization by single spermatozoon of eggs that were diploid as a result of suppression of the second meiotic division. The 3n lines of two 2n/3n embryos were derived from other mechanisms. A single homogeneous tetraploid (4n) embryo and the 4n cell lines of three 2n/4n mosaic embryos each contained two marker chromosomes and presumable resulted from failure of cytokinesis in an early cleavage division.

Animals↗
PubMed1986

Chlorpromazine inhibits the mitotic index, cell number, and formation of mouse blastocysts, and delays implantation of CBA mouse embryos.

Chlorpromazine, administered to pregnant CBA mice 56 h after copulation in single doses of 10 or 15 mg/kg bodyweight, inhibited the compaction of embryos, formation of blastocysts, and reduced the mitotic index and cell number of embryos 86 h after copulation but did not adversely influence their viability or induce structural chromosomal aberrations. Blastocyst formation was more severely affected than embryo compaction. When 86-h embryos were treated with chlorpromazine (10 or 15 mg/kg) and subsequently cultured for 120 h, there was delayed hatching from the zona pellucida, delayed attachment to the culture dish, outgrowth of the trophoblast and expansion of the inner cell mass. Mice treated identically and evaluated on the 18th day of gestation had fewer implanted embryos than did controls, and the fetuses weighed less. No resorptions, malformations or significant differences in intrauterine deaths were found. Chlorpromazine given in the same manner but at 0.5 mg/kg did not affect any of the aforementioned criteria. When 56 h embryos were cultured in vitro in the presence of 50 microM-chlorpromazine for a further 40 h, embryo compaction, blastocyst formation, the mitotic index and the total cell number were significantly reduced compared with controls. Blastocyst formation was again more severely affected than embryo compaction. The inhibition of embryo compaction, blastocyst formation, and reduction in mitotic index and cell number associated in this study with chlorpromazine in vivo and in vitro indicate that the drug inhibits the development of cleavage-stage embryos in the mouse. These effects might be mediated by antagonistic effects of calmodulin.

Animals↗