Embryonic Nutrition Research Articles

BETWEEN BUCCAL PUMPING AND BITING - EVIDENCE OF INTRAUTERINE MOUTH MOVEMENTS AND REPRODUCTIVE ASPECTS OF NEOTROPICAL FRESHWATER STINGRAYS (Elasmobranchii: Potamotrygoninae)

Preliminary findings regarding the reproduction of three neotropical freshwater stingray species, Potamotrygon henlei, P. motoro, and P. orbignyi, are presented herein. These species are characterized by the presence of a singular functional left ovary and two operational uteri. Two distinct fertility models have been identified: fecundity dependent, and fecundity independent of maternal size. Furthermore, this study briefly outlines the process of capture-induced parturition and its implications for the fate of the litter. Additionally, it sheds light on novel embryonic behavioral patterns, particularly highlighting the active swimming exhibited by embryos within the uterine environment, notably during the latter stages of development. Consequently, it is suggested that these movements, coupled with oral activities and active biting, facilitate branchial ventilation and potentially intrauterine foraging - a phenomenon previously documented only in sharks belonging to the order Lamniformes. Keywords: Abortion, Histotroph, Capture stress, Embryonic nutrition.

Aberrant methylation of placental development genes in chorionic villi of spontaneous abortions with trisomy 16.

In humans, aneuploidy is incompatible with the birth of healthy children and mainly leads to the death of embryos in the early stages of development in the first trimester of pregnancy. Trisomy 16 is the most common aneuploidy among spontaneous abortions of the first trimester of pregnancy. However, the mechanisms leading to the death of embryos with trisomy 16 remain insufficiently investigated. One of these potential mechanisms is abnormal placental development, including aberrant remodeling of spiral arteries. Spiral artery remodeling involves the migration of trophoblast cells into the maternal spiral arteries, replacing their endothelium and remodeling to ensure a stable embryonic nutrition and oxygen supply. This is a complex process which depends on many factors from both the embryo and the mother. We analyzed the methylation level of seven genes (ADORA2B, NPR3, PRDM1, PSG2, PHTLH, SV2C, and TICAM2) involved in placental development in the chorionic villi of spontaneous abortions with trisomy 16 (n = 14), compared with spontaneous abortions with a normal karyotype (n = 31) and the control group of induced abortions (n = 10). To obtain sequencing libraries, targeted amplification of individual gene regions using designed oligonucleotide primers for bisulfite-converted DNA was used. The analysis was carried out using targeted bisulfite massive parallel sequencing. In the group of spontaneous abortions with trisomy 16, the level of methylation of the PRDM1 and PSG2 genes was significantly increased compared to induced abortions (p = 0.0004 and p = 0.0015, respectively). In the group of spontaneous abortions, there was no increase in the level of methylation of the PRDM1 and PSG2 genes, but the level of methylation of the ADORA2B gene was significantly increased compared to the induced abortions (p = 0.032). The results obtained indicate the potential mechanisms of the pathogenetic effect of trisomy 16 on the placental development with the participation of the studied genes.

Exploring the Confluence of In-Ovo Mineral Supplementation and Hatching Attributes in Broiler Chick Progeny: A Symphony of Nutritional Enrichment and Broiler Chicks Development

Embryonic nutrition is a key factor that influences broiler progeny performance with long-lasting nutritional adequacies or insufficiencies in the developmental phase. Objective: To explores the effects of in-ovo mineral supplementation or day in-ovo feeding on the hatching attributes and chick quality in broiler breeders. Methods: About 1400 hatching eggs of broiler breeders were selected, categorized into seven experimental groups: one control group, three macro mineral (P, Mg, and Ca) groups, and three micro-mineral (Zn, Mn, and Cu) groups. During the 12th and 18th day of incubation, eggs received in-ovo injections of their respective mineral (5% of the total mineral content in a 50g egg) solutions, prepared by dissolving the corresponding salts in deionized water. Post-injection, eggs were placed in the setter, and upon completion of incubation, hatching trays were removed to evaluate hatchability performance and chick quality parameters for each group. Results: The results showed that Zn, Ca, and Mg-supplemented eggs demonstrated the highest (P<0.05) hatchability rates for settable and fertile eggs at day 12 of in-ovo feeding. The chick weight, hatch window, and hatchling size were significantly influenced (P<0.05) by in-ovo feeding and the day of in-ovo feeding. However, no effect of in-ovo mineral feeding and day of in ovo feeding was observed on chick quality (chick grades, navel area, retracted yolk, and membrane score) and post-hatch chick activities (activity, down-appearance, vitality, eye score, and gait score). Conclusions: In conclusion, Zn, Ca, and Mg in ovo feeding improve the hatchability traits when administered at day 12 of incubation.

Ovarian changes and development of the branchial placenta occurring in Jenynsia lineata (Cyprinodontiformes, Anablepidae).

In viviparous teleosts, intraovarian gestation occurs intrafollicularly, as in poeciliids, or intraluminally, as in goodeids and anablepids. Furthermore, there are two different forms of embryonic nutrition: lecithotrophy and matrotrophy; depending on the species, these can be exclusive or coexist during gestation. In matrotrophic species, nutrients are transmitted from the mother to the embryo and are especially important in species with intraluminal gestation. Jenynsia lineata is a South American viviparous teleost with intraluminal gestation, characterized by eggs with scarce yolk, which is resorbed when embryos are 6 mm long, thus developing a branchial placenta. Using histological, histochemical, and immunohistochemical techniques, the present study describes the characteristics and changes of the ovarian mucosa in J. lineata during gestational and nongestational phases, and analyzes the embryonic pharyngeal epithelium in the branchial placenta. The ovaries of 30 adult female specimens were processed using histological techniques and stained with hematoxylin-eosin, Masson's trichrome, and Alcian Blue pH 2.5/periodic acid Schiff reagent. To detect cell proliferation, we used antiproliferating cell nuclear antigen antibody. In nonpregnant females, eosinophilic granular cells (EGCs) and lymphocytes were identified in the lamina propria of the tunica mucosa, and melanomacrophage centers (MMCs) and fibroblasts were identified adjacent to tissue debris in the ovarian folds'. In the cellular debris, an embryo in resorption was observed. In pregnant females, the ovarian mucosa has thin vascularization branches entering the opercular chamber of the embryos, in close contact with the forming gill processes, thereby establishing a branchial placenta. Active cell replacement was observed in these ovarian branches. The identification of fibroblasts, lymphocytes,EGCs, and MMCs adjacent to tissue debris could indicate that these cell types are involved in the embryonic resorption process. Considering the new data obtained in this study on the branchial placenta of J. lineata, we conclude that cell proliferation could be involved in the development of maternal-embryonic interaction.

Distinguishing Between Embryonic Provisioning Strategies in Teleost Fishes Using a Threshold Value for Parentotrophy.

The source of embryonic nutrition for development varies across teleost fishes. A parentotrophy index (ratio of neonate: ovulated egg dry mass) is often used to determine provisioning strategy, but the methodologies used vary across studies. The variation in source and preservation of tissue, staging of embryos, and estimation approach impedes our ability to discern between methodological and biological differences in parentotrophy indices inter- and intra-specifically. The threshold value used to distinguish between lecithotrophy and parentotrophy (0.6-1) differs considerably across studies. The lack of a standardised approach in definition and application of parentotrophy indices has contributed to inconsistent classifications of provisioning strategy. Consistency in both methodology used to obtain a parentotrophy index, and in the classification of provisioning strategy using a threshold value are essential to reliably distinguish between provisioning strategies in teleosts. We discuss alternative methods for determining parentotrophy and suggest consistent standards for obtaining and interpreting parentotrophy indices.

Measuring of the heifers ovaries and probiotic defence of the mucous membranes

The results of the influence of complex probiotic defence of the mucous membranes of the reproductive system and intestines on the development of the corpus luteum of the cycle in heifers have been presented. By the method of differential palpation diagnostics in vivo, it was reliably examined that in the experimental group after carrying out procedures for the normalization of mucous membranes with the multicomponent probiotic drug “Multibacterin Veterinary Ba + La”, the number of heifers with high-quality of corpus luteum the ovaries was 48.21% more, and the number of cases of premature luteolysis was 12.50 % less (P < 0.01). The level of pregnancy in heifers in the experiment was on average 60.38 %, in the control – 46.94 %, which confirmed the positive effect of the probiotic protection of the mucous membranes on increasing the embryo survival by optimizing the conditions for embryonic nutrition on the surface of the endometrium.

Review of extra-embryonic tissues in the closest arthropod relatives, onychophorans and tardigrades.

The so-called extra-embryonic tissues are important for embryonic development in many animals, although they are not considered to be part of the germ band or the embryo proper. They can serve a variety of functions, such as nutrient uptake and waste removal, protection of the embryo against mechanical stress, immune response and morphogenesis. In insects, a subgroup of arthropods, extra-embryonic tissues have been studied extensively and there is increasing evidence that they might contribute more to embryonic development than previously thought. In this review, we provide an assessment of the occurrence and possible functions of extra-embryonic tissues in the closest arthropod relatives, onychophorans (velvet worms) and tardigrades (water bears). While there is no evidence for their existence in tardigrades, these tissues show a remarkable diversity across the onychophoran subgroups. A comparison of extra-embryonic tissues of onychophorans to those of arthropods suggests shared functions in embryonic nutrition and morphogenesis. Apparent contribution to the final form of the embryo in onychophorans and at least some arthropods supports the hypothesis that extra-embryonic tissues are involved in organogenesis. In order to account for this role, the commonly used definition of these tissues as 'extra-embryonic' should be reconsidered. This article is part of the theme issue 'Extraembryonic tissues: exploring concepts, definitions and functions across the animal kingdom'.

Effect of maternal sleep on embryonic development

The concept of developmental origin of health and disease has ignited a search for mechanisms and health factors influencing normal intrauterine development. Sleep is a basic health factor with substantial individual variation, but its implication for early prenatal development remains unclear. During the embryonic period, the yolk sac is involved in embryonic nutrition, growth, hematopoiesis, and likely in fetal programming. Maternal body measures seem to influence its size in human female embryos. In this prospective, longitudinal observational study of 190 healthy women recruited before natural conception, we assessed the effect of prepregnant sleep duration (actigraphy) on the fetal crown-rump-length (CRL) and yolk sac size (ultrasound). All women gave birth to a live child. The prepregnancy daily sleep duration had an effect on the male yolk sac and CRL at the earliest measurement only (7 weeks). I.e., the yolk sac diameter decreased with increasing sleep duration (0.22 mm·h−1d−1, 95%CI [0.35–0.09], P < 0.01), and CRL increased (0.92 mm·h−1d−1, 95%CI [1.77–0.08], P = 0.03). Since there was no association at the second measurement (10 weeks), and in the group of female fetuses at any measure point, we suggest a sex- and time-dependent embryonic adaptation to sleep generated differences in the intrauterine environment in normal pregnancies.

Aquatic Walking and Swimming Kinematics of Neonate and Juvenile Epaulette Sharks.

The epaulette shark, Hemiscyllium ocellatum, is a small, reef-dwelling, benthic shark that-using its paired fins-can walk, both in and out of water. Within the reef flats, this species experiences short periods of elevated CO2 and hypoxia as well as fluctuating temperatures as reef flats become isolated with the outgoing tide. Past studies have shown that this species is robust (i.e., respiratory and metabolic performance, behavior) to climate change-relevant elevated CO2 levels as well as hypoxia and anoxia tolerant. However, epaulette shark embryos reared under ocean warming conditions hatch earlier and smaller, with altered patterns and coloration, and with higher metabolic costs than their current-day counterparts. Findings to date suggest that this species has adaptations to tolerate some, but perhaps not all, of the challenging conditions predicted for the 21st century. As such, the epaulette shark is emerging as a model system to understand vertebrate physiology in changing oceans. Yet, few studies have investigated the kinematics of walking and swimming, which may be vital to their biological fitness, considering their habitat and propensity for challenging environmental conditions. Given that neonates retain embryonic nutrition via an internalized yolk sac, resulting in a bulbous abdomen, while juveniles actively forage for worms, crustaceans, and small fishes, we hypothesized that difference in body shape over early ontogeny would affect locomotor performance. To test this, we examined neonate and juvenile locomotor kinematics during the three aquatic gaits they utilize-slow-to-medium walking, fast walking, and swimming-using 13 anatomical landmarks along the fins, girdles, and body midline. We found that differences in body shape did not alter kinematics between neonates and juveniles. Overall velocity, fin rotation, axial bending, and tail beat frequency and amplitude were consistent between early life stages. Data suggest that the locomotor kinematics are maintained between neonate and juvenile epaulette sharks, even as their feeding strategy changes. Studying epaulette shark locomotion allows us to understand this-and perhaps related-species' ability to move within and away from challenging conditions in their habitats. Such locomotor traits may not only be key to survival, in general, as a small, benthic mesopredator (i.e., movements required to maneuver into small reef crevices to avoid aerial and aquatic predators), but also be related to their sustained physiological performance under challenging environmental conditions, including those associated with climate change-a topic worthy of future investigation.

Zika virus infection drives epigenetic modulation of immunity by the histone acetyltransferase CBP of Aedes aegypti.

Epigenetic mechanisms are responsible for a wide range of biological phenomena in insects, controlling embryonic development, growth, aging and nutrition. Despite this, the role of epigenetics in shaping insect-pathogen interactions has received little attention. Gene expression in eukaryotes is regulated by histone acetylation/deacetylation, an epigenetic process mediated by histone acetyltransferases (HATs) and histone deacetylases (HDACs). In this study, we explored the role of the Aedes aegypti histone acetyltransferase CBP (AaCBP) after infection with Zika virus (ZIKV), focusing on the two main immune tissues, the midgut and fat body. We showed that the expression and activity of AaCBP could be positively modulated by blood meal and ZIKV infection. Nevertheless, Zika-infected mosquitoes that were silenced for AaCBP revealed a significant reduction in the acetylation of H3K27 (CBP target marker), followed by downmodulation of the expression of immune genes, higher titers of ZIKV and lower survival rates. Importantly, in Zika-infected mosquitoes that were treated with sodium butyrate, a histone deacetylase inhibitor, their capacity to fight virus infection was rescued. Our data point to a direct correlation among histone hyperacetylation by AaCBP, upregulation of antimicrobial peptide genes and increased survival of Zika-infected-A. aegypti.

Functional complexity in the chorioallantoic membrane of an oviparous snake: Specializations for calcium uptake from the eggshell.

The chorioallantoic membrane of oviparous reptiles forms a vascular interface with the eggshell. The eggshell contains calcium, primarily as calcium carbonate. Extraction and mobilization of this calcium by the chorioallantoic membrane contributes importantly to embryonic nutrition. Development of the chorioallantoic membrane is primarily known from studies of squamates and birds. Although there are pronounced differences in eggshell structure, squamate and bird embryos each mobilize calcium from eggshells. Specialized cells in the chicken chorionic epithelium transport calcium from the eggshell aided by a second population of cells that secrete protons generated by the enzyme carbonic anhydrase. Calcium transporting cells also are present in the chorioallantoic membrane of corn snakes, although these cells function differently than those of chickens. We used histology and immunohistology to characterize the morphology and functional attributes of the chorioallantoic membrane of corn snakes. We identified two populations of cells in the outer layer of the chorionic epithelium. Calbindin-D28K , a cellular marker for calcium transport expressed in squamate chorioallantoic membranes, is localized in large, flattened cells that predominate in the chorionic epithelium. Smaller cells, interspersed among the large cells, express carbonic anhydrase 2, an enzyme not previously localized in the chorionic epithelium of an oviparous squamate. These findings indicate that differentiation of chorionic epithelial cells contributes to extraction and transport of calcium from the eggshell. The presence of specializations of chorioallantoic membranes for calcium uptake from eggshells in chickens and corn snakes suggests that eggshell calcium was a source of embryonic nutrition early in the evolution of Sauropsida.

Molecular mechanism of nutrient uptake in developing embryos of oviparous cloudy catshark (Scyliorhinus torazame).

Forms of embryonic nutrition are highly diverse in cartilaginous fishes, which contain oviparity, yolk-sac viviparity and several types of matrotrophic viviparity (histotrophy, oophagy, and placentotrophy). The molecular mechanisms of embryonic nutrition are poorly understood in these animals as few species are capable of reproducing in captivity. Oviparous cartilaginous fishes solely depend on yolk nutrients for their growth and development. In the present study, we compared the contribution to embryonic nutrition of the embryonic intestine with the yolk sac membrane (YSM). RNA-seq analysis was performed on the embryonic intestine and YSM of the oviparous cloudy catshark Scyliorhinus torazame to identify candidate genes involved in nutrient metabolism to further the understanding of nutrient utilization of developing embryos. RNA-seq discovery was subsequently confirmed by quantitative PCR analysis and we identified increases in several amino acid transporter genes (slc3a1, slc6a19, slc3a2, slc7a7) as well as genes involved in lipid absorption (apob and mtp) in the intestine after ‘pre-hatching’, which is a developmental event marked by an early opening of the egg case about 4 months before hatching. Although a reciprocal decrease in the nutritional role of YSM was expected after the intestine became functional, we observed similar increases in gene expression among amino acid transporters, lipid absorption molecules, and lysosomal cathepsins in the extraembryonic YSM in late developmental stages. Ultrastructure of the endodermal cells of YSM showed that yolk granules were incorporated by endocytosis, and the number of granules increased during development. Furthermore, the digestion of yolk granules in the YSM and nutrient transport through the basolateral membrane of the endodermal cells appeared to be enhanced after pre-hatching. These findings suggest that nutrient digestion and absorption is highly activated in both intestine and YSM after pre-hatching in catshark embryos, which supports the rapid growth at late developmental stages.

Mode of uterine milk secretion in the white shark.

Examination of the uterus of a dead female white shark (Carcharodon carcharias), which contained the earliest known white shark embryos, revealed that the uterine wall produces lipid-rich secretion (histotroph or "uterine milk") for embryonic nutrition. Uterine tissue was processed for light and electron microscopy, and immunohistochemical techniques to identify its secretory mechanism. Our results indicate that the white shark uterus secretes lipids via holocrine secretion. This type of secretion is characterized by the release of large lipid droplets accumulated in the epithelial cells into the uterine lumen through cell disintegration. The secretory epithelium of the uterus is stratified, and new surface epithelial cells are continuously supplied from deeper epithelial layers to replace the dead secretory cells at the surface. This vertical replacement possibly facilitates the active renewal of the surface epithelium, which is necessary for maintaining holocrine secretory mechanisms. These secretory mechanisms are different from those of myliobatiform stingrays, another elasmobranch taxon that exhibits lipid histotrophy. This may reflect the different origins of lipid histotrophy between these taxa.

Morphological basis for maternal nutrient provision to embryos in the viviparous fish Ataeniobius toweri (Teleostei: Goodeidae).

In viviparous Mexican fishes of the family Goodeidae, embryos develop in the maternal ovarian lumen. They typically absorb maternal nutrients during gestation by means of "trophotaeniae," that is, specialized, elongated extensions of the hindgut that are exposed to the fluids, which occupy the ovarian lumen. The sole exception is Ataeniobius toweri, whose embryos lack trophotaeniae but are nevertheless matrotrophic. Thus, how its embryos obtain maternal nutrients is unclear. We studied a series of non-pregnant and pregnant ovaries of A. toweri using histology to identify the mechanism of maternal-embryo nutrient transfer. By early-gestation, embryos have depleted their yolk supplies. Yolks are released into the ovarian lumen and are ingested by the developing embryos, as shown by yolk material in their digestive tracts. The embryonic gut is lined by an epithelium consisting of columnar cells with apical microvilli, providing a means for nutrient absorption. Contrary to statements in the literature, embryos develop minuscule trophotaenial rudiments that extend slightly into the ovarian lumen. These structures are formed of an absorptive epithelium that overlies a vascular stroma, similar to the trophotaeniae of other goodeids. Through late gestation, vitellogenic follicles form and oocytes are discharged into the ovarian lumen, contributing to embryonic nutrition. Thus, histological evidence suggests that embryos chiefly obtain nutrients from ingestion of yolk and maternal secretions released into the ovarian lumen. This function possibly is supplemented by uptake via the small hindgut protrusions and other absorptive surfaces (e.g., the skin and the gill epithelium). Our observations are consistent with two evolutionary interpretations of the hindgut protrusions: (a) that they are rudimentary, evolutionary precursors of trophotaeniae formed by exteriorized hindgut; and (b) that they are vestigial remnants of trophotaeniae that were lost during a switch to a form of matrotrophy involving nutrient ingestion.

Exploring the effectiveness of in ovo feeding of vitamin C based on the embryonic vitamin C synthesis and absorption in broiler chickens

BackgroundMany researches about in ovo feeding (IOF) of vitamin C (VC) are gradually carried out to explore physiological development in chicken, but little studies focus on VC synthesis capacity of the embryo itself, the selection of injection site and the effectiveness of IOF of VC. This study aims to explore the above problems.ResultsKidney and yolk sac were the main organs for VC synthesis and L-gulonolactone oxidase (GLO) expression was lower during pre-hatch development than that during post-hatch development. Sodium-dependent vitamin C transporter 1 (SVCT1) expression was increased continuously in yolk sac from embryonic age 19 (E19) to post-hatch day 1 (D1) and in intestine (duodenum, jejunum and ileum) from E17 to D1. Plasma VC content was higher at D1 than that at D21 and D42. IOF of VC significantly reduced GLO expression in liver, kidney and yolk sac as well as SVCT1 expression in duodenum, jejunum and ileum, but increased the VC content in plasma, brain, kidney and liver. In addition, IOF of VC obviously reduced the embryonic morality and increased the hatchability under heat stress.ConclusionsThis study suggested that IOF of VC at E11 in yolk was effective for embryonic VC supplementation. These findings provide a theoretical reference about the method of embryonic VC supplementation and effective methodology on embryonic VC nutrition in broiler chickens.

Morphology of yolk and pericardial sacs in lecithotrophic and matrotrophic nutrition in poeciliid fishes.

We used histological techniques to describe the morphology of the yolk and pericardial sacs in developing embryos of the lecithotrophic species Girardinus creolus, Gambusia puncticulata, Limia vittata, and Quintana atrizona, in comparison with the extreme matrotrophic Heterandria formosa. In lecithotrophic species, the yolk sac was enlarged and lasted until the final stages of development, while in H. formosa it was completely absorbed soon after fertilization. Lecithotrophic poeciliids showed a pericardial sac with a single layer of blood vessels covering the dorsal surface of the cephalic region only, while H. formosa showed a more complex largely vascularized pericardial sac covering the entire dorsal surface, except the caudal region. In advanced gestation of G. creolus, a vascular plexus of the yolk sac reaches the pharyngeal region, behind the gills, suggesting that the pharynx may play a role in embryonic nutrition in lecithotrophic species. These morphological evidences suggest that matrotrophy derives from lecithotrophy.

Metabolomic analysis of the egg yolk during the embryonic development of broilers

The chicken egg yolk, which is abundant with lipids, proteins, and minerals, is the major nutrient resource for the embryonic development. In fact, the magnitude and type of yolk nutrients are dynamically changed during the chicken embryogenesis to meet the developmental and nutritional requests at different stages. The yolk nutrients are metabolized and absorbed by the yolk sac membrane and then used by the embryo or other extraembryonic tissues. Thus, understanding the metabolites in the yolk helps to unveil the developmental nutritional requirements for the chicken embryo. In this study, we performed ultra high performance liquid chromatography/tandem mass spectrometry (UHPLC-MS/MS) analysis to investigate the change of metabolites in the egg yolk at embryonic (E) 07, E09, E11, E15, E17, and E19. The results showed that 1) the egg yolk metabolites at E07 and E09 were approximately similar, but E09, E11, E15, E17, and E19 were different from each other, indicating the developmental and metabolic change of the egg yolk; and 2) most of the metabolites were annotated in amino acid metabolism pathways from E11 to E15 and E17 to E19. Especially, arginine, lysine, cysteine, and histidine were continuously increased during the embryonic development, probably because of their effects on the growth promotion and oxidative stress amelioration of the embryo. Interestingly, the ferroptosis was found as one of major processes occurred from E15 to E17 and E17 to E19. Owing to the upregulated expression of acyl-CoA synthetase long-chain family member 4 detected in the yolk sac, we assumed that the ferroptosis of the yolk sac was perhaps caused by the accumulation of reactive oxygen species, which was induced by the large amount of polyunsaturated fatty acids and influx of iron in the yolk. Our findings might offer a novel understanding of embryonic nutrition of broilers according to the developmental changes of metabolites in the egg yolk and may provide new ideas to improve the health and nutrition for prehatch broiler chickens.

Viviparous stingrays avoid contamination of the embryonic environment through faecal accumulation mechanisms

In viviparous (live-bearing) animals, embryos face an embryo-specific defecation issue: faecal elimination in utero can cause fatal contamination of the embryonic environment. Our data from the viviparous red stingray (Hemitrygon akajei) reveals how viviparous elasmobranchs circumvent this issue. The exit of the embryonic intestine is maintained closed until close to birth, which allows the accumulation of faeces in the embryonic body. Faecal accumulation abilities are increased by (1) the large intestine size (represents about 400–600% of an adult intestine, proportionally), and (2) the modification in the intestinal inner wall structure, specialized to increase water uptake from the faecal matter. According to the literature, faecal accumulation may occur in embryos of the lamniform white shark as well. The reproductive biology of myliobatiform stingrays and lamniform sharks is characterized by the onset of oral feeding before birth (i.e. drinking of uterine milk and eating of sibling eggs, respectively), which is expected to result in the production of large amounts of faeces during gestation. The strong ability of faecal accumulation in these lineages is therefore likely an adaptation to their unique embryonic nutrition mechanism.

Embryonic/Fetal Development, Placentation and Glycosaminoglycans in the Female Reproductive Tract and Placenta

Background: Fetal attachments, placentation and embryonic development have been widely discussed in rodents such as agoutis and cavies, as well as research on glycosaminoglycans (GAGs) in rats and rabbits. Moreover, studies on buffalo, cattle and sheep are described in ruminants, and work has also been reported in sheep with GAGs in placentoma. However, further studies are needed in this regard, since there are reports of economic losses associated with reproductive failures described for cattle such as changes in the chorion and allantois, and in sheep in which changes between the transition from vitelline to allantois circulation have been discussed. Review: In relation to embryonic development, detailed studies have been described in rodents such as rats (12 days old), desert mouse (15 days old) and agoutis at 30 days. Macroscopic structures such as the cephalic region, nose, optic vesicle, cervical curvature, thoracic and pelvic limbs were observed, as well as microscopic structures such as the pituitary, lung, heart, brain cavity, liver, retina, and ossification regions. There are reports of buffalo and cattle studies in ruminants describing early embryonic development. However, the research in the case of sheep is limited, meaning there is only the ultrasound examination, such as gestational diagnosis and morphometric measurement of the embryonic vesicle. Still, studies with umbilical funicular and placental development of sheep with different gestational ages can be highlighted. Regarding extraembryonic annexes, four important structures which contribute to embryonic maintenance have been reported. These are called the chorion, amnion, allantois and yolk sac, respectively, and are responsible for originating the placenta, embryonic protection, collecting metabolic waste and early embryonic nutrition. In addition, correlating the annexes gives rise to the placentation process, which were described two models; the first is transient, called chorioviteline, and the second is called chorioallantoid, which represents the definitive model. Allied to the gestational process, the importance of glycosaminoglycans (GAGs) and proteoglycans are worth mentioning, as they are essential components of the extracellular matrix. They are related in the implantation process, tissue organization during gestation, and also in placental angiogenesis, as described in ruminants (i.e. involved in the vascular growth that accompanies the development of the placenta), which in turn causes an increase in blood flow in this organ, and constitutes a determining factor for fetal development. Conclusion: Such studies regarding the embryonic development of ruminants are still limited to sonographic description and some information is only available in the context of extraembryonic membranes. For glycosaminoglycans, chondroitin sulfate, dermatan sulfate, heparan sulfate and hyaluronic acid are identified in the reproductive tract and placenta. We intend to produce important information for the reproductive and sanitary management of ruminants with the information in this article, providing data to stimulate new studies aiming to minimize the occurrence of embryonic death and economic losses. In addition, further studies on GAGs are needed to better understand their true correlation with gestation, so they can intercede through supplementation and minimize reproductive losses.

Reptile Embryology and Regeneration.

Reptiles (lizards, snakes, turtles, and crocodilians) are becoming increasingly popular as models for developmental investigations. In this review the leopard gecko, Eublepharis macularius, is presented as a reptilian model for embryonic and tissue regeneration studies. We provide details of husbandry and breeding and discuss aspects of embryonic nutrition, egg anatomy, and sex determination. We provide comprehensive protocols for transcardial perfusion, short-term anesthesia using the injectable anesthetic Alfaxan, and full-thickness cutaneous biopsy punches, used in geckos for the study of scar-free wound healing. We also provide modifications to three popular histological techniques (whole-mount histochemistry, immunohistochemistry, and double-label immunofluorescence) and provide details on bromodeoxyuridine (BrdU) labeling and immuno-detection.