Effects of Beauveria bassiana on embryos of the inland silverside fish (Menidia beryllina).

A chemical toxicity and teratogenicity test was adapted to assess potential adverse effects of a microbial pest control agent on a nontarget fish. Developing embryos of the inland silverside, Menidia beryllina, were exposed to conidiospores of the insect-pathogenic fungus Beauveria bassiana. Embryo rupture and death were observed. Embryo rupture did not always result in death, nor was death always associated with embryo rupture. Adherence of spores to the chorion, followed by germination and penetration by the germ tube, probably caused the embryos to rupture. Statistically significant (P less than or equal to 0.05) responses were observed in tests in which conidiospore concentrations were greater than or equal to 8.3 x 10(4) or less than or equal to 1.5 x 10(6)/ml. Conidiospores treated with a dispersant (biological detergent) showed significantly less binding (P less than or equal to 0.01) to embryos than did untreated spores. Both detergent-treated and heat-killed spores failed to cause significant adverse effects.     

Relative developmental abilities of hamster 2- and 8-cell embryos cultured in hamster embryo culture medium-1 and -2

The relative developmental abilities of hamster 2- and 8-cell embryos in culture were compared using two versions of hamster embryo culture medium (HECM). These media differed primarily in the number of amino acids present, i.e., 20 amino acids in HECM-1 and four amino acids in HECM-2. When 2-cell embryos were cultured for 24 h, the percentages of greater than or equal to 4-cell embryos obtained in both HECM-1 and HECM-2 were comparable (congruent to 93%); at the end of 48 h, the proportion of greater than or equal to 8-cell embryos obtained in HECM-1 (82.5%) was significantly (P less than or equal to 0.001) more than that obtained in HECM-2 (67.9%). Interchange of media, after 24 h culture, did not enhance the ability of cultured 2-cell embryos to become blastocysts. When 8-cell embryos were cultured for 18 h in HECM-1 and -2, there was no appreciable difference in the proportion of total blastocysts formed (89-91%). However, there were significantly (P less than or equal to 0.001) more late blastocysts in HECM-2 than in HECM-1 (68.2% vs. 38.4%). Embryo development from 2- and 8-cell stages was compared in media that differed by the presence and absence of phenol red and penicillin-G. There was no difference in embryo development when these compounds were present or absent. Similarly, the difference in pyruvate concentration between HECM-1 and -2 (0.5 and 0.2 mM, respectively) did not affect embryo development.(ABSTRACT TRUNCATED AT 250 WORDS)     

Teratogenicity of Ni2+ inXenopus laevis,assayed by the FETAX procedure

The teratogenicity of Ni2+ was tested by the FETAX (Frog Embryo Teratogenesis Assay: Xenopus) procedure in the South African frog, Xenopus laevis. In seven assays, beginning at 5 h postfertilization, groups of Xenopus embryos were incubated for 96 h in media that contained Ni2+ (added as NiCl2) at concentrations ranging from 1 x 10(-7) to 3 x 10(-3) mol/L; control groups were incubated in the same medium without added NiCl2. At 101 h postfertilization, surviving embryos were counted, fixed in formalin, and examined by microscopy to determine their developmental stages, malformations, and head-to-tail lengths. In control embryos, survival was greater than or equal to 95% and malformations were less than or equal to 7%. Malformations were found in greater than 95% of embryos exposed to Ni2+ concentrations greater than or equal to 5.6 mumol/L. The most frequent malformations in Ni(2+)-exposed embryos were ocular, skeletal, and intestinal deformities; less common malformations included facial, cardiac, and integumentary deformities. Other abnormalities, not categorized as malformations, included stunted growth, dermal hypopigmentation, and coelomic effusions or hemorrhages. The median embryolethal concentration (LC50) of Ni2+ was 365 (SE +/- 9) mumol/L; the median teratogenic concentration (EC50) was 2.5 (SE +/- 0.1) mumol/L; the Teratogenic Index (TI = LC50/EC50) was 147 (SE +/- 5), indicating that Ni2+ is a potent teratogen for Xenopus laevis. Experiments in which Ni(2+)-exposures were limited to specific 24 h periods showed that Xenopus embryos were most susceptible to Ni(2+)-induced malformations on the second and third days of life, during the most active period of organogenesis.     

Exogenous gamma-glutamyl cycle compounds supplemented to in vitro maturation medium influence in vitro fertilization, culture, and viability parameters of porcine oocytes and embryos

High concentrations of intracellular glutathione (GSH) enhance in vitro production of porcine embryos. Objectives were: (1) to determine the effects of gamma-glutamyl cycle compound supplements to the IVM medium on IVF and IVC; and (2) to evaluate embryo viability. Porcine oocytes were matured in NCSU 23 medium supplemented with either l-cysteine (3.3 mM), l-cysteamine (150 P < 0.05microM), l-cysteine and l-cystemaine, l-glycine (1, 2.5, or 5 mM), l-glutamate (1, 2.5, or 5 mM), l-alpha-aminobutyrate (3.3mM), beta-mercaptoethanol (BME) (25 microM), l-cysteine and BME, or l-alpha-aminobutyrate and BME. Increases (P < 0.05) in GSH concentrations were observed using l-cysteine, 1.0 mM l-glutamate, l-alpha-aminobutyrate, and l-alpha-aminobutyrate with BME. Oocytes matured with l-alpha-aminobutyrate and BME had a lower (P < 0.05) occurrence of polyspermy during IVF compared to controls and a greater percentage (P < 0.05) of embryos reaching the blastocyst stage compared to other treatment groups. For Objective 2, oocytes were matured in NCSU 23 or NCSU 23 supplemented with l-alpha-aminobutyrate with BME. Embryo cell death was determined using an Annexin V-FITC assay. Supplementation had no effect on the time of cell death. Embryo mortality was increased (P < 0.05) from 24 to 42 h post-IVF, with the greatest occurrence around 36 h. In conclusion, supplementing l-alpha-aminobutyrate and BME into the IVM medium increased intracellular GSH concentrations, decreased the occurrence of polyspermy during IVF, and increased embryo development parameters during IVC, but did not affect cell death during embryo development. The onset of cell death occurred from 24 to 42 h post-IVF, with the greatest occurrence around 36 h post-IVF.     

Effect of Salicylhydroxamic Acid on Endosperm Strength and Embryo Growth of Lactuca sativa L. cv Waldmann''s Green Seeds

Salicylhydroxamic acid (SHAM) stimulated germination of photosensitive lettuce (Lactuca sativa L. cv Waldmann's Green) seeds in darkness. To determine whether SHAM acts on the embryo or the endosperm, we investigated separately effects of SHAM on growth potential of isolated embryos as well as on endosperm strength. Embryo growth potential was quantified by incubating decoated embryos in various concentrations of osmoticum and measuring subsequent radicle elongation. Growth potential of embryos isolated from seeds pretreated with 4 millimolar SHAM was equal to that of untreated controls. Rupture strength of endosperm tissue excised from seeds pretreated with SHAM was 33% less than that of controls in the micropylar region. To determine if the embryo must be in contact with the endosperm for SHAM to weaken the endosperm, some endosperms were incubated with SHAM only after dissection from seeds. Rupture strength of SHAM-treated, isolated endosperms in the micropylar region was 25% less than that of untreated controls. There was no difference in rupture strength in the cotyledonary region of endosperm isolated from seeds treated with SHAM in buffer or buffer alone. SHAM therefore stimulates germination not by enhancing embryo growth potential, but by weakening the micropylar region of the endosperm enclosing the embryo.     

Embryo survival and conceptus growth after reciprocal embryo transfer between Chinese Meishan and Landrace x Large White gilts

Embryos were transferred between Meishan and Landrace x Large White (control) gilts on Day 4 or 5 to establish approximately equal numbers of all four possible combinations of donor breed and recipient breed. The breed of the donor gilt significantly (P less than 0.01) affected embryo survival with 44.5% of transferred Meishan embryos and 69.6% of transferred control embryos surviving to Day 30 +/- 1. There was no influence of the breed of the recipient gilt on the proportion of embryos which survived. These differences in embryo survival between the two breeds could not be explained by differences in (1) the number of embryos transferred, (2) the stage of development of the embryos transferred, (3) the interval between ovulation and transfer or (4) the degree of asynchrony between donor and recipient gilt. On Day 30 +/- 1 embryos from control donors developed into longer fetuses (P less than 0.01) with larger allantoic sacs (P less than 0.05) than did embryos from Meishan donors. Fetuses in control recipients were longer (P less than 0.01), heavier (P less than 0.001) and had larger allantoic sacs (P less than 0.05) than fetuses occupying Meishan uteri. The interaction between breed of donor gilt and breed of recipient gilt did not significantly affect conceptus growth. These results suggest that Meishan pig embryos may be less tolerant to routine embryo transfer procedures than those of control gilts, that the genotype of the dam does not affect the proportion of embryos surviving to Day 30 +/- 1, and that both fetal and maternal factors affect conceptus growth.     

The effect of freezer type, cryoprotectant, and processing methods on viability of frozen embryos

Two hundred forty excellent-quality blastocysts flushed from 53 superovulated Holstein heifers were frozen by 1 of 16 procedures in a 2 x 2 x 2 x 2 factorial design. The main effects included a simple, inexpensive, portable mechanical freezing unit instead of a programmable Liquid Nitrogen (LN) freezer for freezing bovine embryos, cryoprotective agents dimethyl sulfoxide (DMSO) and glycerol, addition rates of the cryoprotectants and freezing rates. Embryo viability was assessed morphologically and by fluorescein diacetate (FDA) evaluation. Neither the type of freezer, the cryoprotectant nor the rate of cryoprotectant addition affected embryo viability. Embryo survival after 12 h of incubation was higher (P < 0.05) using a conventional freezing rate than a two-step method (37.2 vs 16.5%). The correlation coefficient between viability evaluation methods (morphology vs FDA) was influenced by cryoprotectant and embryo processing methods and ranged from -0.13 to +0.70. This study indicates that more simplified embryo freezing equipment and handling procedures may provide protection equal to that of more complicated, expensive equipment and more time-consuming methods. Economical on-farm embryo freezing is feasible.     

Donor and recipient genotype and heterosis effects on survival and prenatal growth of transferred mouse embryos

Reciprocal embryo transfers amongst two inbred strains (C3HeB/FeJ and SWR/J) and their F1 cross (C3SWF1) were used to examine donor and recipient genotype and heterosis effects on survival and prenatal growth of mouse embryos. Among inbred strains, significant recipient genotype effects were detected for both embryo survival (P less than 0.01) and prenatal growth (P less than 0.05), while no donor genotype effects were observed. The recipient effect on overall embryo survival was due to a higher proportion of C3H recipients maintaining pregnancy to term than SWR recipients (P less than 0.01), rather than survival within litters. Irrespective of their own genotype, embryos developing in C3H uteri achieved larger body weights (P less than 0.01) and longer tail lengths (P less than 0.05) at birth than did embryos developing in SWR uteri. Recipient heterosis was not significant, while donor heterosis was significant for prenatal growth traits (P less than 0.001).     

Sucrose dilution of glycerol from mouse embryos frozen rapidly in liquid nitrogen vapour

The toxic effects of sucrose and the conditions of in-straw glycerol removal after freezing and thawing were studied using Day-3 mouse embryos. At 20 degrees C, exposure to less than or equal to 1.0 M-sucrose for periods up to 30 min had no adverse effects on freshly collected embryos. At 25 and 36 degrees C, however, greater than or equal to 1.0 M-sucrose significantly reduced the developmental potential (P less than 0.001). In the freezing experiments the embryos were placed in 0.5 ml straws containing 40 microliters freezing medium separated by an air bubble from 440 microliters sucrose solution. The straws were frozen rapidly in the vapour about 1 cm above the surface of liquid nitrogen. The post-thaw viability was substantially better after sucrose dilution at 20 degrees C than at 36 degrees C. Mixing the freezing medium with the sucrose diluent immediately after thawing further improved the rate of survival relative to mixing just before freezing (P less than 0.001). The best survival was obtained when the freezing medium contained 3.0 M-glycerol + 0.25 M-sucrose; it was mixed with the diluent after thawing and the glycerol was removed at 20 degrees C. Under such conditions the sucrose concentration in the diluent had no significant effect on the rate of development (0.5 M, 69%; 1.0 M, 73%; 1.5 M, 64%). The results show that during sucrose dilution the temperature should be strictly controlled and suggest that intracellular and extracellular concentrations of glycerol are important in the cryoprotection of embryos.     

Critical factors affecting the permeabilization of Drosophila embryos by alkanes.

Because of waxes in the vitelline membrane, the Drosophila egg is effectively impermeable to liquid water and to aqueous solutes, and consequently it cannot be cryopreserved unless it can be permeabilized. The more successful of the few published permeabilization procedures involve the removal of the chorion mechanically or by hypochlorite solution, the removal of all surrounding water by air drying or alcohol, the exposure of eggs to pure alkanes like octane or hexane for some 30 s, the removal of the alkane and the transfer of the eggs to aqueous culture medium without their desiccation, and lastly incubation of the permeabilized embryos under mineral oil. In following these procedures we opted for a somewhat different approach to applying hypochlorite, water, alcohol, and alkane; namely, eggs were placed between two Nucleopore filters, and the fluids drawn sequentially through the filters by vacuum. Extensive initial attempts were mystifying and discouraging in that although permeabilization was good, survivals were poor, and modifications that increased the latter reduced the former. The explanation turned out to be that permeabilization and survival depended critically on the amount of carry-over alcohol that contaminated the alkane. To determine the effects of alcohol concentration in the alkane, it was essential first to effectively eliminate carry-over contamination and then re-add precise amounts of alcohol (isopropanol) to the alkane (n-hexane, heptane, or octane). When the alcohol concentration is less than or equal to 0.2%, permeabilization is poor; when it is greater than or equal to 0.5%, permeabilization is good but survival (hatching) is poor. There are strong interactions between alcohol concentration and exposure time to alkane/alcohol mixtures with respect to the fraction of embryos that become permeabilized and the percentage that survive. There are also significant but less critical effects from the type of alcohol and alkane. The best results for 12-h embryos (greater than or equal to 90% permeabilization and 70-80% hatching) were achieved with eggs exposed to 0.3 or 0.4% 1-butanol in n-heptane for 90 s. High survivals of permeabilized 12-h embryos did not require incubation under mineral oil. Permeabilized embryos are permeable to water, ethylene glycol, glycerol, and the stain rhodamine B (which was used to assess permeabilization). They are effectively impermeable to sucrose. Embryo age is important. Between 14 and 16 h the above permeabilization procedures become dramatically less effective.(ABSTRACT TRUNCATED AT 400 WORDS)     

In vitro and in vivo survival of bisected sheep embryos derived from frozen-thawed unsorted, and frozen-thawed sex-sorted and refrozen-thawed ram spermatozoa

Ovine IVP embryos were derived from frozen-thawed unsorted and frozen-thawed sex-sorted spermatozoa that had been refrozen and thawed. The embryos were bisected and cultured in vitro, or transferred to recipient ewes to determine their survival in vitro and in vivo. Oocyte progression to the blastocyst stage was similar for unsorted (97/232, 41.8%) and sex-sorted spermatozoa (113/286, 39.5%; P > 0.05). Embryo survival in vitro post-bisection was similar for demi-embryos derived from unsorted and sex-sorted sperm, and embryos bisected at the blastocyst and expanded blastocyst stage (P > 0.05). A higher proportion of recipient ewes were pregnant at Day 63 after transfer of two intact embryos derived from unsorted (17/21, 80.9%) than two demi-embryos derived from unsorted (5/15, 33.3%) or sex-sorted spermatozoa (7/17, 41.2%). The number of fetuses per original embryo at Day 63 did not differ among groups (unsorted intact: 23/42, 54.8%; unsorted demi: 7/15, 46.7%; sex-sorted demi: 10/17, 58.8%) and twin pregnancies were observed in all groups. Embryo survival to term was high, and was not significantly different among intact (unsorted: 22/42, 52.4%) and demi-embryos (unsorted: 4/15, 26.7%; sex-sorted spermatozoa: 7/17, 41.2%; P > 0.05). Dizygotic twins (n = 6 sets) were born after the transfer of two intact embryos derived from unsorted spermatozoa, but only singleton lambs resulted from the transfer of demi-embryos. In conclusion, bisected IVP embryos successfully developed into morphologically normal lambs. However, embryo survival to term was neither increased nor decreased by embryo bisection.     

Cleavage beyond the block stage and survival after transfer of early bovine embryos cultured with trophoblastic vesicles

Early bovine embryos (1- to 8-cell stages) were recovered from superovulated heifers at slaughter on Days 2 or 3. Embryos were cultured for 3-4 days in Medium B2 supplemented with 15% (v/v) fetal calf serum in the absence (B2SS, 106 embryos) or presence of trophoblastic vesicles (B2SS + TV, 190 embryos). At the end of culture, there were more (P less than 0.001) morulae (greater than or equal to 16 cells) in B2SS X TV (46%) than in B2SS alone (18%) irrespective of the initial cell stage. More 8-cell embryos reached the 16-cell stage than did embryos with less than 8 cells (30% vs 15% in B2SS, P greater than 0.05; 70% vs 41% in B2SS + TV, P less than 0.005). After culture, 102 morulae were transferred non-surgically to temporary recipient heifers (84 embryos cultured in B2SS + TV and 18 in B2SS). After 2 or 3 days, 14 out of 58 embryos from the B2SS + TV group and 3 out of 10 embryos from the B2SS group were recovered as blastocysts. Most blastocysts were deep-frozen and stored for several weeks. After thawing, 10 apparently normal embryos from the B2SS + TV group were transferred non-surgically into 10 recipient heifers. Four pregnancies were induced, but only one embryo survived to term (birth of a normal male calf). It is concluded that trophoblastic vesicles release one or several unknown compound(s) normally present in vivo, promoting the cleavage of early bovine embryos.     

Genotype,plant, bud size and media factors affecting anther culture of cauliflowers (Brassica oleracea var. botrytis)

Summary Eleven F₁ hybrid cultivars of cauliflower, representing a range of maturity types, were examined for their responsiveness to anther culture. Embryos were produced from each of the cultivars tested, and the mean embryo yield varied from 82.2 embryos per 100 anthers cultured for cv Dova to 0.6 embryos for cv Serrano. Variation between genotypes and between plants within a genotype was significant, both in terms of embryo yield and percentage responsive anthers. Autumn and winter maturing cauliflowers were generally more responsive than summer types. Embryo yields were enhanced by culturing anthers on solid rather than on liquid media. An increase in concentration of 2,4-Dichlorophenoxyacetic acid (2,4-D) from 0.1 to 0.3 mg/l also increased embryo yield. Embryo yield was doubled when anthers were cultured on solid media containing 0.3 mg/l 2,4-D compared to liquid media containing 0.1 mg/l 2,4-D. Although bud size alone did not have a significant effect on embryo production, genotype x bud size and plant x bud size (within genotype) interactions were significant. Estimation of the variance components demonstrated that, apart from the residual plate-to-plate variation, variation between plants was the largest source of variation, accounting for approximately 30% of total variance. Plant x bud size (within genotype) interaction accounted for 18% of total variance and genotypic differences for approximately 8%.     

The Aggregation of Four Reconstructed Zygotes is the Limit to Improve the Developmental Competence of Cloned Equine Embryos

Embryo aggregation has been demonstrated to improve cloning efficiency in mammals. However, since no more than three embryos have been used for aggregation, the effect of using a larger number of cloned zygotes is unknown. Therefore, the goal of the present study was to determine whether increased numbers of cloned aggregated zygotes results in improved in vitro and in vivo embryo development in the equine. Zona-free reconstructed embryos (ZFRE''s) were cultured in the well of the well system in four different experimental groups: I. 1x, only one ZFRE per microwell; II. 3x, three per microwell; III. 4x, four per microwell; and IV. 5x, five ZFRE''s per microwell. Embryo size was measured on day 7, after which blastocysts from each experimental group were either a) maintained in culture from day 8 until day 16 to follow their growth rates, b) fixed to measure DNA fragmentation using the TUNEL assay, or c) transferred to synchronized mares. A higher blastocyst rate was observed on day 7 in the 4x group than in the 5x group. Non-aggregated embryos were smaller on day 8 compared to those aggregated, but from then on the in vitro growth was not different among experimental groups. Apoptotic cells averaged 10% of total cells of day 8 blastocysts, independently of embryo aggregation. Only pregnancies resulting from the aggregation of up to four embryos per microwell went beyond the fifth month of gestation, and two of these pregnancies, derived from experimental groups 3x and 4x, resulted in live cloned foals. In summary, we showed that the in vitro and in vivo development of cloned zona-free embryos improved until the aggregation of four zygotes and declined when five reconstructed zygotes were aggregated.     

Effects of melatonin and undernutrition on the viability of ovine embryos during anestrus and the breeding season

This study examined the effects of melatonin and level of nutrition on embryo yield during anestrous and breeding season. Adult Rasa Aragonesa ewes were assigned randomly to one of the four treatment groups in two experiments using a 2x2x2 factorial design. Individuals were treated (+MEL) or not treated (-MEL) with a subcutaneous implant of melatonin for 42d (Melovine, CEVA) and fed 1.5 (control, C) or 0.5 (low, L) times the daily maintenance requirements for 20d. Ewes were mated at oestrus (Day=0) and embryos were recovered on Day 5. Level of nutrition and melatonin supplements did not have a significant effect on ovulation rate or the number of recovered ova per ewe in the Reproductive Season (RS) and the Anestrous Season (AS). During the RS, undernutrition reduced the number of viable embryos per ewe (C: 1.1+/-0.2; L: 0.6+/-0.2; P<0.05); however, the number of viable embryos per ewe in the L+MEL group (0.2+/-0.15) was significantly lower than it was in the L, C+MEL and C groups (0.9+/-0.3, 1.2+/-0.3, 1.0+/-0.4, respectively; P<0.05). In the AS, nutrition did not have a significant effect on the number of viable embryos per ewe, although melatonin supplements might have improved rates slightly. Embryo viability rate (% viable embryos/embryos recovered) was unaffected by melatonin supplements or level of nutrition in the RS and the AS. Season had a strong effect on the number of viable embryos per functional corpus luteum among ewes in the L+MEL group, only (RS: 0.2+/-0.1; AS: 0.6+/-0.2; P<0.05). In conclusion, undernutrition impaired the viability of sheep embryos in the RS, particularly among ewes that were given melatonin supplements subcutaneously, but melatonin appeared to improve embryo quality in the AS, which suggests that the mechanisms involved in the interactive effects of melatonin and nutrition on embryo development are influenced by season.     

The effect of external medium composition on membrane water permeability of zebrafish (Danio rerio) embryos

The effect of external medium composition on chorion and plasma membrane permeability of zebrafish (Danio rerio) embryos was investigated in this study. Initially, survival of embryos spawned into varying strengths (10-40%) of Hank's solution (HBSS) was assessed. Development and hatching rates for embryos spawned into 30% and 40% HBSS were significantly lower than those obtained with embryos spawned into system water. The effect of embryo survival in 30% HBSS with different calcium levels was then investigated. Embryo survival in calcium free 30% HBSS or 30% HBSS with 10x the standard calcium concentration was similar to survival in standard 30% HBSS. Membrane water permeability was determined by measuring the floatation time of embryos in test solutions made up with heavy water (D2O) instead of deionized water. Intact embryos at early developmental stages were less permeable than later stages irrespective of the external medium that they were spawned into. In system water, the floatation time of embryos at one-cell and two-cell stages were 1323+/-83 and 1189+/-55 s, respectively, compared to 432+/-6 and 353+/-10 s at the high and 50% epiboly stages. Change of external medium composition had no effect on membrane permeability of intact embryos at early developmental stages. However, at later stages embryos spawned into 30% HBSS were less permeable than embryos spawned into system water, irrespective of calcium concentration. The flotation time of embryos at the high stage increased from 432+/-6s in system water to 468+/-10s in 30% HBSS. The study on dechorionated embryos showed that change of external medium composition had no effect on plasma membrane permeability.     

Superovulation treatments and embryo transfer in Angora goats

A high incidence of early luteal regression after PMSG superovulation was associated with low recovery of embryos from reproductive tracts of Angora goats flushed later than Day 5 after onset of oestrus. Embryos were successfully recovered (mean 7.9/female) by flushing on Days 2-5. Mean ovulation rate after an FSH regimen (16.1 +/- 0.8) was significantly higher than that after a single injection of PMSG (10.8 +/- 1.2). Fertilization rate and survival of embryos following transfer to naturally synchronized recipient feral goats did not differ between the two gonadotrophin regimens: the mean number of kids born to 47 donors treated with FSH (7.5 +/- 0.6) was significantly greater than that to 28 donors treated with PMSG (4.8 +/- 0.6). Irrespective of hormonal treatment, the numbers of embryos recovered and of kids born were correlated with ovulation rate (r = 0.82, P less than 0.001 for both). Embryo survival was influenced by ovulation rate in recipients, with 52%, 63% and 75% of transferred embryos being carried to term by recipients with 1,2 and 3 CL, respectively (P less than 0.01). More embryos survived (65%) when 2 embryos were transferred to each recipient than when 1 (51%) or 3 (48%) were transferred. In recipients receiving 2 embryos, survival was significantly improved by transfer of both embryos to the same oviduct (70%) than when one was transferred to each oviduct (62%). The percentage survival of embryos was optimal when oestrus of recipients was synchronized within +/- 1 day of oestrus in donors.     

Protective effect of ouabain on adriamycin-induced cardiovascular anomalies in chick embryos

Adriamycin (2.5-10.0 micrograms) was administered to 4 1/2 and 5 day embryonic chicks (Hamburger-Hamilton developmental stages 24-26) to investigate the effect of the drug on cardiovascular morphogenesis. The drug produced dose-related increases in both mortality rate and malformation frequency with a maximum incidence of 82% cardiovascular anomalies following a dose of 10.0 micrograms/egg (P less than .001 relative to saline controls). Frequencies of embryos with ventricular septal defect (P less than .005), dextroposition of the aorta (P less than .005), or aortic arch anomalies (P less than .05) were significantly higher than among controls. In a second study, embryos were pretreated with ouabain (12.2 micrograms), verapamil (0.5 micrograms), coenzyme Q10 (100 micrograms, 200 micrograms), or vitamin E (1.0 mg, 5.0 mg)--agents previously shown to protect against adriamycin-induced cardiotoxicity. Pretreatment of embryos with ouabain significantly reduced the incidence of cardiovascular malformations induced by adriamycin from 55 to 21% (P less than .05). A major protective effect was observed relative to the induction of ventricular septal defect, the frequency of which was reduced from 45 to 14% (P less than .05). However, administration of verapamil, coenzyme Q10, or vitamin E did not have an appreciable effect on adriamycin-induced frequencies of cardiovascular malformations. Negative inotropism is suggested as a mechanism for adriamycin-induced cardiac anomalies but warrants further study.     

IN OVULO EMBRYO DEVELOPMENT AND PLANT FORMATION FROM STENOSPERMIC GENOTYPES OF VITIS VINIFERA

Embryo enlargement and plant formation via in ovulo embryo culture was studied in stenospermic grapes. Genotypes, culture media and their interactions significantly affected embryo enlargement, whereas culture dates did not. Genotype P60-58, when compared to cv. Thompson Seedless, had the greatest number of enlarged embryos after 3 months in culture. P60-58 ovules cultured on Cain's basal medium plus L-serine or L-glutamine and cv. Thompson Seedless ovules cultured on Cain's basal medium plus L-cysteine or L-asparagine produced the greatest number of enlarged embryos. Viable embryos and morphologically normal-appearing plants were obtained. Polyembryony was observed in cultured zygotic embryos.