‘Milk’ secretion for embryogenesis in a viviparous cockroach

The brood sac of viviparous Diploptera punctata is a typical insect integumentary gland which secretes a ‘milk’ containing protein and carbohydrate to nourish the developing embryos. During gestation the secretory cells proliferate organelles of protein synthesis and secretion and brood sac wet weight, protein content, synthetic activity and secretory output increase five- to six-fold ; a maximum of 0.4 mg protein was collected in 24 hr from one brood sac in a later stage of gestation. Following parturition, when secretory activity ceases, these parameters fall markedly, and the secretory cells decrease their mass by autophagic regression. Acid phosphatase has been located histochemically in autolysomes and assayed in brood sac homogenates; activity reaches a maximum five days after parturition.     

Organic,inorganic, and caloric composition of eggs,pentaculae, and adults of the brooding sea cucumber Cucumaria curata cowles (Echinodermata: Holothuroidea)

Levels (percentage composition) of water, ash, carbohydrate, lipid, protein, and calories were determined for eggs, pentaculae, and adults of the sea cucumber Cucumaria curata Cowles. Component contents (μg/individual) were calculated for eggs and pentaculae. During the 28 days of development to hatching, the large yolky eggs gain water and ash, the total dry weight increasing from 169 to 190 μg/egg during embryogenesis. There were no statistically significant changes in lipid, protein, and caloric contents during embryogenesis, but carbohydrate decreased by 0.82 μg/egg.The decrease in carbohydrate is sufficient to account for estimated embryonic energy requirements. Based on the utilization of carbohydrate, embryos of C. curata show a nutritional pattern similar to that of the planktonic embryos of sea urchins and different from that of embryos developing from terrestrial eggs, freshwater eggs, and planktonic and demersal marine eggs.Although broods varied widely in egg number and mean egg dry weight, C. curata gives eggs which contain a constant proportion of organic components.Levels of ash, water, and protein in the adults exceeded those in the pentacula, and lipid comprises a much smaller proportion of the adult body than it did of the pentacula.     

Water uptake at ecdysis in the western rock lobster

Water ingestion at ecdysis by the western rock lobster. Panulirus longipes (Milne Edwards) was investigated using the reference markers ⁵¹Cr-EDTA and ⁵⁸Co-EDTA. Two possible mechanisms controlling water absorption were examined: first, changes in osmolarity of blood and muscle and secondly, the effects of extracts of central nervous system.Water ingestion was 16.071 ± 2.365 mlkg^?1h^?1 during swelling just before ecdysis (stage D₄(S)) and 23.099 ± 1.238 mlkg^?1h^?1 during stage A. There was no significant absorption in the foregut or hindgut and the digestive gland appeared to be the site of major absorption. Total water ingested during stages D₄(S) and A was 13.7% of the proecdysis weight. Calculating total water uptake by wet weight differences plus wet weight of exuviae gave a value that was too high and instead weight increases were calculated from a carapace length-weight formula. Allowing for postecdysis increase in weight the net increase at ecdysis was 18.4–21.4% which was 4.7–7.7% more than the water ingested. It was concluded from this that water enters the body at ecdysis both by ingestion and by absorption through the external surface. It is suggested that water ingestion provides the main source of swelling of the cephalothorax in stage D₄(S) and after ecdysis both ingested water and external absorption enables the flaccid abdomen and appendages to swell rapidly.Statistically significant differences were found in the concentrations of total cations and chloride in leg muscle during the transition from stage C₄ to late D₄ but the trends were not consistent and probably have no functional significance. There were no changes in the concentration of osmotically active organic constituents. The freezing-point depression of the blood in stage D₄ was significantly higher than that in stage C₄(P < 0.02) but the mean difference was only 1.8%. It was concluded that osmoticchanges were unlikely to be an important mechanism of water uptake.Water-soluble extract (WSE) and acetone-soluble extract (ASE) of brains and first ventral ganglia were without significant effect when compared together with controls. There was a barely significant decrease, however, in water in the proventriculus of WSE-treated animals compared with that of controls (P < 0.05). and further investigation on the effects of such extracts on water uptake at ecdysis is warranted.     

Migration of the monarch butterfly, Danaus plexippus: energy sources,

The origin and nature of the metabolic reserves of the monarch butterfly during its autumn migration through Missouri were examined. Information was obtained about the butterfly's protein, glycogen, lipid, and lipid-soluble pigment composition. The principal results showed that lipids, which made up ca. 45 per cent of the insect's dry weight, were the major reserve. Analyses of the lipid classes showed that triglycerides were the dominant class in each sex. Separate analyses of the thorax and abdomen showed that >90 per cent of the lipids were present in the massive fat body which is restricted to the abdomen. The adult female rapidly incorporated U-¹⁴C-d-glucose into abdominal glycerides. The rôle of larval reserves and nectar consumption in migration and ovarial maturation was discussed.     

Biochemical Alterations of Dermatophytes during Growth

Alterations in the biochemical constituents of mycelia were studied during the growth, development, and starvation of Microsporum quinckeanum. On the basis of dry weight, growth of this dermatophyte could be divided into four phases: lag, log, stationary, and death. The percentage of total nitrogen, inorganic phosphorus, ribonucleic acid (RNA), and protein increased rapidly during the lag phase. The percentage of protein remained constant after the initial increase; however, inorganic phosphate and RNA decreased in older mycelia. Acid-soluble materials in the cells increased in concentration as the organism aged. Chitin was present in the spores at a much higher concentration than in the mycelia. The percentage of this compound decreased rapidly until the end of the lag phase. An increase and subsequent decrease in per cent chitin occurred during the log phase. Inorganic phosphorus in the mycelia increased from the value in the spore stage to a maximum in the early log phase, and then decreased rapidly during the remainder of the growth cycle. Compounds involved in protein synthesis increased rapidly during the lag phase of growth. Changes in chemical composition of the mold during starvation indicate that carbohydrate does not form the principal endogenous reserve of M. quinckeanum, whereas lipids may represent the primary reserve material.     

The developmental profiles of two major haemolymph proteins from Drosophila melanogaster.

There are four major protein species in the haemolymph of the late 3rd instar of Drosophila. Two of these, LSP-1 and LSP-2, have been studied in detail. Larvae, pupae, and flies of different ages were measured for wet weight, total extractable protein and using an immunoassay, the amounts of LSP-1 and LSP-2. The synthesis of both proteins begins after the 2nd larval ecdysis and at puparium formation they represent 9% and 1.5% of total extractable protein. This value remains constant during the first part of metamorphosis, then falls rapidly. The function of these proteins and their suitability as systems for the study of gene control and protein synthesis in Drosophila are discussed.     

Vitellogenin fluctuations in haemolymph and fat body and dynamics of uptake into oöcytes during the reproductive cycle of Diploptera punctata

Haemolymph and fat body soluble protein titres have been examined during the reproductive cycle of Diploptera punctata, with particular emphasis on the occurrence of vitellogenin and its uptake into the developing oöcytes. Vitellogenin was first detected in the haemolymph of mated females 2 days after adult eclosion at about the same time that vitellin deposition in basal oöcytes began. Peak haemolymph titres of vitellogenin occurred on day 6, correlated with the completion of yolk uptake. Thereafter vitellogenin levels declined and were generally undetectable throughout most of gestation, rising again shortly before parturition in association with the second gonotrophic cycle. Total haemolymph protein levels were not correlated with vitellogenesis.Soluble fat body vitellogenin titres of mated females remained low during the first oöcyte growth period but then rose several-fold at its completion and remained high throughout pregnancy and the second gonotrophic cycle. Total fat body soluble proteins decline after adult eclosion in association with oöcyte growth.Vitellin accumulation in basal oöcytes was related linearly to increase in volume until the onset of chorion formation. Thus no post-vitellogenic growth period was detected.     

Quantitative changes and synthesis of cyanoprotein in whole body and tissues during development of the bean bug,Riptortus clavatus

Changes in biliverdin-binding cyanoprotein content in whole body and tissue extracts during development of nymphal and adult (non-diapause) bean bugs, Riptortus clavatus were analyzed by rocket immunoelectrophoresis (RIE). RIE using anti-CPegg serum can be used to determine the content of CP-A (Cp-1, 2 and 3) and CP-B (CP-4) separately. During the nymphal stage CP content of whole body changes cyclically in each instar. In the first nymphal instar, CPegg is the main CP which disappears during the first-second instar ecdysis. In nymphal bugs from the 2nd to 4th instars only CP-B (CP-4) is detected, and at the beginning of each instar the CP content is very low but increases toward the next ecdysis, after which CP decreases and disappears very rapidly. In the 5th nymphal instar, CP-B is the major CP but CP-A (CP-1, 2 and 3) is also detected. These changes in whole body CP content of 5th instar nymphs are observed in both females and males. The content of total CPs in the 5th instar nymph reaches about 1000 μg in the whole insect. During nymphal-adult ecdysis, nymphal CPs decrease and disappear at day 2 after emergence. In female adults CP-A (CP-1 only) increases rapidly after day 4 of adult emergence, while no CP is detected in male adults. In females CPs were detected only in the fat body, hemolymph and ovary. In the mid-5th-instar nymphs, CPs (CP-A and B) are mainly distributed in the hemolymph. CPs in the Hemolymph decrease during nymphal-adult ecdysis, whereas they increase in the fat body. CPs disappear from both the hemolymph and fat body by 2 days after ecdysis. Subsequently in the adult stage only CP-A increases again in the fat body and ovary. By tracer experiments using [³⁵S]-methionine, the fat body was shown to be the site of CP synthesis. CP-A and B synthetic activity was detected in nymphal females whereas, only CP-A synthesis was observed in adult females, while no CP synthesis was seen in adult males.     

Matrotrophic viviparity constrains microbiome acquisition during gestation in a live‐bearing cockroach,Diploptera punctata

The vertical transmission of microbes from mother to offspring is critical to the survival, development, and health of animals. Invertebrate systems offer unique opportunities to conduct studies on microbiome‐development‐reproduction dynamics since reproductive modes ranging from oviparity to multiple types of viviparity are found in these animals. One such invertebrate is the live‐bearing cockroach, Diploptera punctata. Females carry embryos in their brood sac, which acts as the functional equivalent of the uterus and placenta. In our study, 16S rRNA sequencing was used to characterize maternal and embryonic microbiomes as well as the development of the whole‐body microbiome across nymphal development. We identified 50 phyla and 121 classes overall and found that mothers and their developing embryos had significantly different microbial communities. Of particular interest is the notable lack of diversity in the embryonic microbiome, which is comprised exclusively of Blattabacteria, indicating microbial transmission of only this symbiont during gestation. Our analysis of postnatal development reveals that significant amounts of non‐Blattabacteria species are not able to colonize newborn D. punctata until melanization, after which the microbial community rapidly and dynamically diversifies. While the role of these microbes during development has not been characterized, Blattabacteria must serve a critical role providing specific micronutrients lacking in milk secretions to the embryos during gestation. This research provides insight into the microbiome development, specifically with relation to viviparity, provisioning of milk‐like secretions, and mother–offspring interactions during pregnancy.     

The biochemical composition and calorie density of the walleye pollock <Emphasis Type="Italic">Theragra chalcogramma</Emphasis> in the Sea of Okhotsk

In tissues of the walleye pollock Theragra chalcogramma the dry matter content averages 18.5%. The lipid content of the raw material is 0.7%, the protein content is 15.3%, carbohydrates are 0.6%, and ash is 1.3%. The average calorie density is 940 cal/g wet weight and 5080 cal/g dry weight. The dry matter content of gonads varies within 14.9–28.0% in females and 14.5–17.0% in males. The lipid content of the raw material is 0.9–3.0% in females and 1.3–1.8% in males; the protein content is 10.2–21.5% and 10.7–13.4%, respectively. The calorie density of female gonads is 702–1537 cal/g wet weight and 4426–5482 cal/g dry weight; for the male gonads it is 760–960 cal/g wet weight and 4952–5641 cal/g dry weight. The dry matter content of the liver varies within 42.2–62.2% for females and 34.4–62.4 for males. The lipid content of the raw material is 25.6–44.5% for females and 16.6–41.3% for males; the protein content is 6.3–9.8% and 8.1–12.3%, respectively. The calorie density of the liver in females varies within 2918–4601 cal/g wet weight and 6370–7395 cal/g dry weight; in males it is 2291–4357 cal/g wet weight and 6392–7492 cal/g dry weight. The minimum calorie density of the liver is observed in juvenile pollock: 963 cal/g wet weight and 2045 cal/g dry weight. The dry-matter content of feces in different size groups varies within 15.0–18.4%. Values of the average lipid content of raw material range from 1.1 to 1.6%; the protein content is from 1.8 to 3.8% and carbohydrates are from 0.9 to 1.4%. The calorie density of feces from variously-sized walleye pollock varies within a narrow range, from 308 to 362 cal/g wet weight. The energy equivalent ranges, depending on body size, from 259 to 2377 cal. The share of energy concentrated in the somatic (muscle) tissue of variously-sized walleye pollock during ontogenesis constitutes 56.5–93.9%; in female gonads it is 0.9–26.6%; in male gonads it is 0.4–7.3%, in the female liver it is 7.9–27.2%, and in the male liver it is 5.7–26.9%. The amount of energy (cal), concentrated in the female liver and gonads is on average 1.5 and 3 times as high as that in the male liver and gonads, respectively. The maximum total energy loss (15–30%) in mature walleye pollock of various-sizes occurs in the spawning period, during the transition from the maturity stage 5 to stage 6. The total amount of energy accumulated during the lifecycle from small juveniles (<17 cm) to very large individuals (>60 cm) averages 1964 kcal for females and 1465 kcal for males. The difference in the amount of energy is explained by the fact that oogenesis requires more energy than spermatogenesis.     

The inhibition of milk synthesis by juvenile hormone in the viviparous cockroach, Diploptera punctata

The brood sac of the viviparous cockroach, Diploptera punctata, synthesizes a protein rich milk which nourishes developing embryos. Milk is first detected in the brood sac (by immunoelectrophoresis) when the embryos begin drinking and continues to increase in parallel with total protein of the brood sac. When embryos cease drinking, both total protein and milk decline in the brood sac. Premature decline in protein and milk content of the brood sac has been observed after treatment with juvenile hormone (from implanted active corpora allata) or a juvenile hormone analogue (ZR 512 applied topically). The fine structure of the brood sac 7 days after corpora allata implant is consistent with that of gland cells which are not actively synthesizing milk. The effect of ZR 512 is detected in decreased milk content of the brood sac after 24 hr of treatment.     

Physiological roles of a yeast-like symbiote in reproduction and embryonic development of the brown planthopper,Nilaparvata lugens Stål

The physiological roles of a yeast-like symbiote in oviposition and embryonic development of its host, the brown planthopper, Nilaparvata lugens Stål, were studied using heat treatment, lysozyme injection and ligation of eggs. The eggs laid by the heat-treated females harboured only a few of the symbiotes, and their symbiote ball through embryonic development was free of symbiotes. The embryos of subsymbiotic eggs could not undergo blastokinesis and dorsal closure, and failed to hatch due to lack of differentiation of the abdominal segments. Electrophoretic profile of the eggs laid by the heat-treated females indicated the absence of several minor proteins which are usually found in the fat body of normal females. A protein (Y) of 131 kD was barely detectable in the heat-treated insects, and could not be found in the ligated eggs in which the symbiote ball was completely separated from the developing germ band. It is suggested that the symbiote supplies its host with proteins for normal embryonic and postembryonic development. The number of yeast-like symbiotes in female adults was reduced after injection with lysozyme solution, and some of the eggs were unable to hatch due to failure in blastokinesis; this was similar to the heat-treated insects. The embryos of ligated eggs could complete segmentation and differentiation normally before 110 h, but the abdominal segments failed to differentiate after dorsal closure, and regressed leaving only the head. Partially ligated eggs harboured some symbiotes and could produce normal larvae. It is concluded that the yeast-like symbiote is significant in abdominal segmentation and differentiation of the planthopper embryo.     

Proximate body composition of the larval,metamorphosing and downstream migrant stages in the life cycle of the Southern Hemisphere lamprey,Geotria australis

Synopsis The water, total lipid, protein and ash content have been measured in larval, metamorphosing (stages 1–7) and downstream migrantGeotria australis caught in Western Australia between October 1977 and August 1979. The total lipid content of ammocoetes changed markedly with season and increased with body size. Although, unlike other species, the ammocoetes ofG. australis continue to increase in length during the latter part of larval life, the relative amount of total lipid still rose during this period, eventually reaching levels equivalent to approximately 14% of the wet body weight at the commencement of metamorphosis. During the six months between the onset of metamorphosis and the downstream migration, total lipid declined to approximately 8%. Assays for phospholipid of larval and metamorphosingG. australis indicated that changes in total lipid were almost entirely due to variations in neutral lipid. Changes in the percentage amount of total lipid were accompanied by an inverse but slightly greater amount of change in percentage water. During metamorphosis, the absolute amount of total lipid in a standard animal declined from 122 mg at stage 1 to 53 mg at stage 7, whereas water rose initially from 597 mg at stage 1 to 638 mg at stage 3, before declining to 442 mg by stage 7. Although the percentage amount of protein and ash tended to increase slightly during larval life, neither showed conspicuous seasonal changes. In both relative and absolute terms, protein declined during metamorphosis. A comparison of the data on the proximate body composition inG. australis and holarctic lampreys shows that different strategies have been employed to accumulate large amounts of fat by the end of larval life and to utilize protein during metamorphosis.     

Changes in lipid and protein during starvation and the moulting cycle in the tiger prawn, Penaeus esculentus Haswell

Tiger prawns, Penaeus esculentus Haswell (mean wt 20.8 ± 0.3 g, range 13.9–27.7 g) contained 1–2% extractable lipid, 13% protein (biuret method) and 71–74% water (wet wt). In 21 days, the weight of fed prawns increased by 3% and that of starved prawns decreased by 4.4%. Protein was the major energy source during 14 days of starvation, with a loss of 550 mg of total protein compared with a loss of 84 mg of total lipid. The absolute amount of water present remained constant. Of three different tissue compartments, abdomen, cephalothorax, and digestive gland, the abdomen contributed the most protein (330 mg) and lipid (35 mg) during 14 days of starvation. Digestive gland, although containing the largest percentage wet wt of lipid, accounted for only 8.3% of the total lipid in the prawn, and contributed only 18 mg of lipid in 14 days of starvation. Lipid concentration in the digestive gland increased during early premoult (stage D₄) and dropped in late premoult (stage D₄). Resting oxygen consumption rate remained constant at ≈0.1 ml · g^?1 · h^?1 at 25°C during 21 days of starvation.     

Regulation of juvenile hormone synthesis during pregnancy in the cockroach, Diploptera punctata

Regulation of juvenile hormone synthesis during pregnancy was investigated after determining the normal rates of synthesis in pregnancy and the second gonadotrophic cycle in Diploptera punctata by direct in vitro radiochemical assay.The low rate of juvenile hormone synthesis during early pregnancy is maintained by three factors: (1) the small ovary which is incapable of eliciting increased rates of juvenile hormone synthesis (2) an inhibitory centre in the brain acting via intact nerves to the corpora allata (similar to that in virgin females) and (3) an inhibitory centre in the brain acting via the haemolymph (elicited by embryos in the brood sac).The existence of two inhibitory centres in the brain is supported by the additive effect of denervating the corpora allata and removing embryos. Whereas these operations alone activated the corpora allata in 54 and 31% of the females, respectively, together they activated 87%, similar to the 91% activated by denervation alone in late pregnancy.The inhibition which remains after denervation of the corpora allata can be removed by decapitation and restored by implantation of the protocerebrum from a pregnant female but not from one developing oöcytes.The inhibition elicited by embryos in the brood sac can be overcome by introduction of a stimulatory ovary and/or substitution of active corpora allata.     

Ovarian and haemolymph titres of ecdysteroid during the gonadotrophic cycle in Diploptera punctata

The free (non-conjugated) ecdysteroid in the ovaries during the first gonadotrophic cycle of Diploptera punctata was identified as 20-hydroxyecdysone. The hormone, quantified by radioimmunoassay and by ultraviolet absorbance, was detectable in the ovary toward the end of vitellogenesis; the quantity increased rapidly during chorion formation. Ovaries with chorionated eggs contained 67 μg of 20-hydroxyecdysone per g fresh weight. The haemolymph free-ecdysteroid, not identified physicochemically, was quantified by radioimmunoassays. The highest concentration was observed at adult emergence; the titre declined between days 1–3 and then remained at a relatively constant level through oviposition (which occurs between day 7 and 8); titres in pregnant females were higher. Ovariectomized females exhibited the same pattern of ecdysteroid titres in the haemolymph as the sham operated controls throughout the period corresponding to the first gonadotrophic cycle. Thus the ovary may not be the only source of haemolymph ecdysteroid related to reproduction in adult females.     

Hypertrehalosaemic and hyperlipaemic responses to adipokinetic hormone in fifth larval instar locusts,Locusta migratoria

In fifth instar larvae of Locusta migratoria the haemolymph lipid concentration is elevated after injection of adipokinetic hormone (AKH). This hyperlipaemic response in larvae remains substantially lower than in adults; over 75% of the mobilized lipid consists of diacylglycerol. In addition, unlike adult locusts, fifth instar larvae also exhibit a consistent, though moderate, hypertrehalosaemic response to AKH. The increases of both lipid and carbohydrate concentrations in larvae are dose-dependent, showing a significant linear regression on log dose in the range 0.2–20 pmol AKH.Glycogen phosphorylase in the fat body of fifth instar larvae as well as young adults is activated on injection of AKH, the percentage active phosphorylase increasing linearly with log dose in the range 0.04–20 pmol AKH. For a given response, a somewhat higher dose of AKH is needed in larvae than in young adults.Fat body glycogen phosphorylase is strongly activated during the period of the larval-adult ecdysis, when active phosphorylase accounts for almost half of the total enzyme, which is approximately ten times more than it is two days before, and two days after the ecdysis.The corpora cardiaca of fifth larval instar locusts already possess the potencies to elevate carbohydrate and lipid concentrations in larval haemolymph, and to activate fat body glycogen phosphorylase.     

Biochemical composition of the Atlantic bonito Sarda sarda from the Aegean Sea (eastern Mediterranean Sea) in different stages of sexual maturity

The content (% wet mass) in water, ash, lipid, crude protein, DNA and RNA of different tissues was determined during sexual maturation of bonitos Sarda sarda from the Aegean Sea. A total of 220 specimens were collected in the following stages of sexual maturity: immature, resting, developing, mature, spawning and spent. Highest lipid levels in the white muscle, red muscle and liver were measured in immature specimens, while lowest levels were found in spawning bonitos. The gradual percentage of lipid reduction from immature to spawning bonitos was relatively higher in the liver (females 71·2% and males 64·4%) than in the white (females 59·2% and males 53·5%) and red (females 62·1% and males 51·7%) muscle. Lipid levels in the gonads increased gradually from the immature to spawning stage. The decrease of lipid in the somatic tissues was more intense in females than in males, and gonadal lipid content was higher in females than in males. There was a strong reverse correlation between water and lipid percentage in all tissues. Protein content decreased significantly only in spawning bonitos. The percentage of protein reduction from immature to spawning stage was relatively higher in males than in females in both white (females 3·4% and males 4·6%) and red (females 4·6% and males 5·1%) muscles. Protein content in the liver was significantly lower than in the other tissues, being highest in mature females. Gonadal protein content in females increased with maturation and decreased after spawning. The content in ash exhibited considerable stability. The RNA:DNA ratio exhibited a similar pattern of variation in both muscles. The RNA:DNA ratio increased during gonadal development gradually from the developing to spent stage. It was concluded that in S. sarda during gonadal development, there was an increase in gonadal lipid accompanied by a decrease in somatic tissue lipid reserves. Thus, reproductive inactive bonitos have more lipid in their edible part and a higher nutritional value than active ones.     

Sfrp1a and Sfrp5 function as positive regulators of Wnt and BMP signaling during early retinal development

Axial patterning of the developing eye is critically important for proper axonal pathfinding as well as for key morphogenetic events, such as closure of the optic fissure. The dorsal retina is initially specified by the actions of Bone Morphogenetic Protein (BMP) signaling, with such identity subsequently maintained by the Wnt-β catenin pathway. Using zebrafish as a model system, we demonstrate that Secreted frizzled-related protein 1a (Sfrp1a) and Sfrp5 work cooperatively to pattern the retina along the dorso-ventral axis. Sfrp1a/5 depleted embryos display a reduction in dorsal marker gene expression that is consistent with defects in BMP- and Wnt-dependent dorsal retina identity. In accord with this finding, we observe a marked reduction in transgenic reporters of BMP and Wnt signaling within the dorsal retina of Sfrp1a/5 depleted embryos. In contrast to studies in which canonical Wnt signaling is blocked, we note an increase in BMP ligand expression in Sfrp1a/5 depleted embryos, a phenotype similar to that seen in embryos with inhibited BMP signaling. Overexpression of a low dose of sfrp5 mRNA causes an increase in dorsal retina marker gene expression. We propose a model in which Sfrp proteins function as facilitators of both BMP and Wnt signaling within the dorsal retina.     

In vivo development of vitrified rabbit embryos: Effects on prenatal survival and placental development

The aim of this work is to study the effect of the vitrification procedure on prenatal survival and on placental development at the end of gestation in rabbits (Oryctolagus cuniculus). One hundred eighty-one females were slaughtered at 72 h of gestation. Morphologically normal embryos recovered at 72 h of gestation were kept at room temperature until transfer or vitrification. Vitrified embryos (320 embryos) were transferred into a total of 24 does and fresh embryos (712 embryos) were transferred into a total of 43 does. Females were induced to ovulate 72 h before transfer when fresh embryos were transferred and 60 to 63 h before transfer when vitrified embryos were transferred. Each recipient doe received eight embryos into the left oviduct and eight embryos into the right oviduct. The number of implanted embryos was estimated by laparoscopy as number of implantation sites at Day 14 of gestation. Recipient females were slaughtered by stunning and exsanguination 25 d after the transfer, and fetuses were classified according to their status. Live fetuses and fetal and maternal placenta were weighed Pregnancy rate was defined as the total number of females having at least one live fetus at Day 28 of gestation divided by the total number of females. Prenatal survival was estimated as live fetuses at Day 28 of gestation divided by the number of transferred embryos. The pregnancy rate after transfer of vitrified embryos (92%) was similar to that achieved with fresh embryos (86%), but prenatal survival was lower for vitrified than for fresh embryos (53% vs. 34%). We did not find differences in embryo survival from 72 h to implantation. Transfer of vitrified embryos reduced fetal survival from implantation to Day 28 (57% vs. 82%). Differences in the number of live fetuses at Day 28 of gestation were mainly due to the higher fetal mortality observed soon after implantation in pregnancies resulting from the transfer of vitrified embryos. A higher percentage of decidual reactions and atrophic maternal placentas (27.5% vs. 8.3%) and also of atrophic fetal and maternal placentas (12.1% vs. 5.3%) were observed after transfer of vitrified embryos. Both treatments showed similar percentage of dead fetuses (3.3% vs. 4%). Maternal placenta of the fetuses from fresh embryos was 15% heavier than maternal placenta of fetuses from vitrified embryos.