Fine structure of the intestine development in cultured sea bream larvae

At hatching, the gut cells of Sparus aurata are quite undifferentiated; however, slight ultrastructural differences can already be distinguished between the presumptive intestinal regions. The hindgut cells are more differentiated than midgut cells and the rectal cells show rather particular ultrastructural features. During days 1 (D1) and 2 (D2) after hatching, major changes occur that lead to full differentiation of the epithelial cells. Shortly before the onset of exogenous feeding (D3), the anterior intestine enterocytes can synthesize lipoprotein particles (LP) from endogenous lipids. The posterior intestine enterocytes show morphological features indicating a role in absorption and intracellular digestion of nutrients, whereas the rectal cells do not. Transient ciliated cells occur at hatching (D0) in the presumptive intestine, except in the caudal rectum, and disappear at the start of the late endotrophic phase about 3 days after hatching (D3). At hatching, very scarce enteroendocrine and leucocyte-like cells are found at the base of the gut epithelium. Their number increases throughout development. At D3 (late endotrophic phase), LP synthesized mainly in the periblast invade the circulatory system, interstitial spaces of the subepithelial tissue and intercellular spaces of the gut epithelium. When the endo-exotrophic phase begins (D4), the enterocytes can absorb exogenous food. Acid phosphatase activity was detected in microvilli, pical vacuoles and Golgi complex in both anterior and posterior enterocytes, as well as in supranuclear vacuoles (SNV) of posterior enterocytes, but not in the apical tubulovesicular system (TVS). During the exotrophic phase, large lipid droplets (LD) are found in anterior enterocytes, and the SNV occupy a large cell volume in posterior enterocytes. LP accumulate first in extracellular spaces and then are transferred to the circulatory system. Mucous and rodlet cells appear in the intestinal epithelium during the exotrophic phase, from D15.     

Development of the digestive tract in larval summer flounder

Histological changes of the digestive system and its associated glands, and structures of the jaw were studied in summer flounder Paralichthys dentatus from hatching (day 0) until day 44. Specimens for this study were hatched from artificially spawned broodstock and maintained in the laboratory (20 ± 1° C). During the first 3 days after hatching, the formation of the oral jaw apparatus, lengthening of the digestive tube, yolk resorption, and mucosae differentiation are the most conspicuous elements of development. The larval digestive system is morphologically ready to process external food at the time of mouth opening (3–4 days after hatching). Epithelial cells of the anteromedian and the posterior intestine show evidence of lipid and protein absorption, respectively, after first feeding. The most noticeable events occurring during the next month of independent life are an increase in mucosal folding, cellular differentiation in the luminal epithelia, gut segmentation and looping, and liver growth. Gastric glands and pyloric caeca appear by day 31 and complete the morphological digestive features characteristic of the juvenile stage.     

Electrophoretic characterization of glycosaminoglycans during development of the chick embryo skin

Glycosaminoglycans extracted by CPC precipitation from chick embryo skin at 9, 12, 15, 18, 21 days of incubation were separated by three different electrophoretic methods on acetate cellulose strips. We observed the presence of Hyaluronic acid, Dermatan Sulfate and Chondroitin-4-Sulfate during the whole period considered and of Heparan Sulfate only after the 9th day. Dermatan Sulfate increases until the 15th day then decreases progressively; on the contrary Hyaluronic acid and Chondroitin-4-Sulfate decrease during days 9 to 15 then increase until hatching. Heparan Sulfate appear the 9th day then increases progressively until hatching.     

Post-hatching dynamics of plasma biochemistry in free-living European starlings (Sturnus vulgaris)

This is the first study of plasma biochemical parameters in free-living altricial birds during an entire developmental period in a nest, represented by European starling (Sturnus vulgaris). Dynamics of postnatal changes from hatching until close to fledging (days 1 to 15) were registered. Parameters of protein metabolism represented by total proteins, albumin and globulin concentrations increased continuously during the observed developmental period. There were two peaks in uric acid concentration on days 5 and 11. To the contrary, the creatinine content did not change throughout the observed period and increased only on day 15. Creatine kinase activity gradually increased until day 11 and then fell before fledging. Parameters of lipid metabolism (concentration of total lipids, triacylglycerols and nonesterified fatty acids) in plasma increased gradually reaching a plateau between days 8 and 11 and then declined on day 15. The cholesterol concentration pattern was similar to maximum value on day 11, then consecutively decreased. Concentration of glucose increased until day 8 and remained unchanged until fledging. Whereas calcium reached the highest concentration during days 8 and 11, phosphorus peaked earlier on day 5. The activity of alkaline phosphatase was similar to the pattern found in calcium concentration. Presented data showed an increase in both protein and lipid metabolism during the phase of rapid growth. A remarkable decrease in parameters of lipid metabolism before fledging may reflect increased physical activity and changes in nutrition.     

Ontogenetic development of the digestive tract in Cuban gar (Atractosteus tristoechus) larvae

Histological method was used to describe the development of the digestive tract in Atractosteus tristoechus larvae reared under culture conditions. The larvae were kept at 28 ± 1 °C in three 15 L circular tanks for 18 days and they were fed with Artemia. According to the structural changes in the digestive system, three significant stages were established: (1) lecithotrophic, (2) lecithoexotrophic and (3) exotrophic. The first stage spanned from hatching to 3 days after hatching (DAH), the digestive system started to differentiate and larvae depended entirely on the endogenous nutrition from the yolk sac. During second stage (4–10 DAH), considered critical since it is the transition period to exotrophic feeding, the digestive tract was fully differentiated into buccopharynx, esophagus, non-glandular and glandular stomach, anterior and posterior intestine. First periodic Schiff reagent-positive goblet cells also appeared, interdispersed within the epithelium of the digestive tract, increasing substantially in numbers and distribution as development continued. At this early stage, gastric glands were only observed in the fundic stomach and not in the cardiac and pyloric region. Pyloric caeca, spiral valve and rectum were also clearly distinguishable in the intestine. After the onset of the exogenous feeding (11–18 DAH), the organization and differentiation of the digestive tract did not undergo any noticeable modification, only the increase in size and complexity of the structures, and it attained the four tissue layer arrangement characteristic of adult vertebrates.     

Larval organogenesis of Pagrus pagrus L., 1758 with special attention to the digestive system development

Organogenesis of the red porgy (Pagrus pagrus L., 1758) was examined from hatching until 63 days post-hatching (dph) using histological and histochemical techniques. At hatching, the heart appeared as a tubular structure which progressively developed into four differentiated regions at 2 dph: bulbus arteriosus, atrium, ventricle and sinus venosus. First ventricle and atrium trabeculae were appreciated at 6 and 26 dph, respectively. Primordial gill arches were evident at 2 dph. Primordial filaments and first lamellae were observed at 6 and 15 dph, respectively. At mouth opening (3dph), larvae exhausted their yolk-sac reserves. The pancreatic zymogen granules appeared at 6 dph. Glycogen granules, proteins and neutral lipids (vacuoles in paraffin sections) were detected in the cytoplasm of the hepatocytes from 4-6 dph. Hepatic sinusoids could be observed from 9 dph. Pharyngeal and buccal teeth were observed at 9 and 15 dph, respectively. Oesophageal goblet cells appeared around 6 dph, containing neutral and acid mucosubstances. An incipient stomach could be distinguished at 2 dph. The first signs of gastric gland development were detected at 26 dph, increasing in number and size by 35-40 dph. Gastric glands were concentrated in the cardiac stomach region and presented a high content of protein rich in tyrosine, arginine and tryptophan. The intestinal mucous cells appeared at 15 dph and contained neutral and acid glycoconjugates, the carboxylated mucins being more abundant than the sulphated ones. Acidophilic supranuclear inclusions in the intestinal cells of the posterior intestine, related to pynocitosis of proteins, were observed at 4-6 dph.     

Why is grouper larval rearing difficult?: an approach from the development of the feeding apparatus in early stage larvae of the grouper,Epinephelus coioides

The osteological development of elements forming the oral cavity was examined in early stage larvae of the grouper,Epinephelus coioides, from hatching to 242.5 hours after hatching. By the time of initial mouth opening, at 54 hours after hatching, the fundamental elements, composed of the trabecula, some components of the lower branchial and hyoid arches, the quadrate and symplectic-hyomandibular cartilages, maxilla and Meckel's cartilage, had appeared. No further elements were observed until 165 hours after initial mouth opening, except some components in the lower branchial arch and head region. The appearance of new elements and initial ossification of existing cartilage occurred thereafter, but all elements related to feeding either had not appeared or had not started ossifying until 188.5 hours after initial mouth opening. Based on the morphology and developmental modes of these elements, the feeding mode of grouper larvae was considered to be “sucking/grasping.” However, the appearance and ossification of elements occurred slowly, with no transitional phase from sucking to grasping modes of feeding being observed during the study; such delayed development of the feeding-related bony elements was considered to be a cause of the difficulty in rearing early stage grouper larvae.     

Development of the cerebrospinal fluid in chick embryos. Physicochemical properties.

The development of the physicochemical properties of the cerebrospinal fluid (CSF) was studied in chick embryos from the 9th day of incubation up to hatching. Some of these properties were compared with the corresponding blood or blood plasma properties. During the second half of incubation the CSF pressure rose from 13.2 plus or minus 0.18 mm H2O in 9-day-old embryos to 80.7 plus or minus 0.48 mm H2O just prior to hatching. The critical stages of this development were the 13th to 15th and the 19th to 21st day of incubation. In 13- and 15-day-old embryos, CSF pressure fell sharply after the intracerebral injection of ouabain, but in 19-day embryos it was unaffected. Except for the 15th and 19th incubation day, the CSF pH was always lower than the plasma pH. From the 11th day of incubation up to hatching, the CSF pH fell from 7.36 plus or minus 0.002 to 7.2 plus or minus 0.005. On the 11th and 13th day, specific CSF resistance was higher than plasma resistance, whereas from the 17th incubation day it was significantly lower than the plasma value. During the second half of incubation, specific CSF resistance fell from 1.059 times 10(6) to 0.824 times 10(6) omega mm.m(-1). A difference between the D.C. potential of the venous blood and the CSF appeared for the first time in 15-day-old embryos, the CSF being negative in relation to the blood. By the end of the incubation period this potential difference rose to 10.82 times 0.07 mv.     

Ultrastructural studies on the pheromone-producing cells in the silkmoth, Bombyx mori: formation of cytoplasmic lipid droplets before adult eclosion

In Bombyx mori, pheromone-producing cells accumulate a number of lipid droplets in the cytoplasm preceding the production of the sex pheromone, bombykol. The process of lipid droplet formation in the pheromone-producing cells was investigated by using light and electron microscopy. Light microscopy revealed that the lipid droplets appeared from 2 days before adult eclosion and dramatic accumulation took place between 2 days and 1 day before eclosion. Electron microscopical studies revealed that smooth endoplasmic reticulum and numerous vesicles, their sizes being less than 1 microm, were detectable 2 days before eclosion, and some vesicles were fused with mitochondria at this stage. These characteristic changes in the pheromone-producing cells suggest that fatty acyl-CoA synthesis following de novo fatty acid synthesis takes place at this time. Involutions in the basal plasma membrane of the cells occurred throughout the observed period, which were extensive on the day before adult eclosion. Besides extensive basal involutions, immature lipid droplets appeared and then mature fully electron-dense lipid droplets were observed on the day of adult eclosion. These ultrastructural observations, combined with recent physiological studies suggest, that the basal involutions presumably reflect the uptake of lipidic components required for the construction of lipid droplets, the function of which is to store the bombykol precursor and to provide it for bombykol biosynthesis in response to pheromonotropic stimuli by pheromone biosynthesis activating neuropeptide (PBAN).     

Embryonic development of the sea bass Dicentrarchus labrax

The embryonic development of the sea bass Dicentrarchus labrax during the endotrophic period is discussed. An 8 cells stage, not reported for other studied species, results from two rapid successive cleavages. Blastula occurs at the eighth division when the embryo is made of 128 cells. During gastrulation, the infolded blastoderm creates the endomesoblastic layer. The Kupffer??s vesicle is reported to drive the left/right patterning of brain, heart and digestive tract. Heart formation starts at 8 pairs of somites, differentiation of myotomes and sclerotomes starts at the stage 18 pairs of somites; main parts of the digestive tract are entirely formed at 25 pairs of somites. At 28 pairs of somites, a rectal region is detected, however, the digestive tube is closed at both ends, the jaw appears the fourth day after hatching, but the mouth is not opened before the fifth day. Although cardiac beating and blood circulation are observed, gills are not reported in newly hatched individuals; eye melanization appears concomitant with exotrophic behavior.     

Ontogeny of enzymatic activities in fed and fasting turbot, Scophthalmus maximus L.

The presence and localization of acid and alkaline phosphatase, non-specific proteases, aminopeptidase, amylase, non-specific esterase and lipase was investigated by histoenzymologic methods in fed and fasting turbot from day 1 to day 40 post-hatching and compared with published data. Alkaline phosphatase and aminopeptidase activities were delected at day 1 in the distal region of the developing digestive tube. At day 3 (opening of the mouth) aminopeptidase and alkaline phosphatase activities were found all along the intestine. Sites of non-specific esterase and protease activities became apparent in the digestive tract at days 2 and 3 respectively. Amylase was present in the exocrine pancreas at day 3 and in the lumen of the intestine at day 4. Acid phosphatase was active in the cellular structure surrounding the yolk stores and in the lipid droplets at day 1 and in the intestinal epithelium at day 3. Lipase was found at day 15 when the larvae metamorphose into juveniles.
All the investigated enzymes were detected in fasting animals, except for lipase. However, the intensities of the enzymatic activities were weaker in the fasting specimens relative to the fed specimens between days 7 and 10.     

Ontogeny of the digestive tract in sharpsnout sea bream Diplodus puntazzo (Cetti, 1777)

The ontogeny of the digestive tract was studied histologically and histochemically in sharpsnout sea bream Diplodus puntazzo from hatching (0 DAH, Days After Hatching) until day 57 (57 DAH). At hatching, the digestive tract appeared as a histologically undifferentiated straight tube lying dorsally to the yolk sac. When the mouth opened at 3 DAH, the digestive tract was differentiated into buccopharynx, oesophagus, incipient stomach and intestine. The pancreas, liver and gall bladder were also differentiated at this stage and both the bile and pancreatic duct had opened into the anterior intestine. Active feeding began in 50% of larvae at 4 DAH, although permanence of yolk reserves until 7 DAH suggests a period of both endogenous and exogenous feeding. Nutrient absorption was first visible from 5 DAH, as colourless supra- and infranuclear vacuoles in the anterior intestinal mucosa, suggesting a lipid content, as well as supranuclear, eosinophilic vacuoles, containing protein, in the posterior intestinal mucosa. Early caecal development could be detected from 10 DAH, whereas gastric glands appeared at 30 DAH, indicating the transition from larval to juvenile stage and the acquisition of an adult mode of digestion. Goblet cells appeared in the digestive tract of sharpsnout sea bream larvae shortly after first feeding. The mucus content of goblet cells varied with the digestive region and, in the buccal cavity and oesophagus, also with the developmental phase. This study provides knowledge for better husbandry practices in the aquaculture industry, as well as for the implementation of future nutritional studies.     

Morphological development of the swim bladder in hatchery-reared striped trumpeter Latris lineata

This study examined swim bladder morphogenesis in three cohorts of striped trumpeter (Latris lineata), a euphysoclist species with physostomous larvae. The swim bladder was first discernible 1–2 days after hatching as an evagination on the dorsal surface of the incipient digestive tract. It comprised a cluster of mesenchymal cells surrounding an inner primordium of epithelial cells. At mouth opening in larvae of 5.3 mm standard length (SL), the swim bladder was noticeably enlarged. Histologically, the swim bladder lumen was dilated and liquid filled. The pneumatic duct was first seen during the dilation stage and the rete mirabile began forming among the connective tissue surrounding the swim bladder. Initial swim bladder inflation occurred on day 11 post‐hatching in Cohort 1, at 14°C, and day 9 post‐hatching, in Cohorts 2 and 3, at 16°C. Histologically, the lumens of inflated swim bladders were ellipsoid and the epithelium was squamous, except for cuboidal gas gland cells at the anterio‐ventral and anterio‐lateral regions of the swim bladder. During the initial inflation interval the pneumatic duct was dilated in larvae both with and without swim bladder inflation. The pneumatic duct began to regress in some larvae over 7.5 mm SL. The swim bladder of striped trumpeter was similar to larvae of other altricial perciform marine fish in respect to organ derivation, tissue differentiation, luminal dilation and initial gaseous inflation. However, variations, particularly the delay in initial swim bladder inflation until after the start of feeding, were observed that could be fundamental to problems encountered during larval rearing.     

Energy metabolism in eggs and larvae of the Senegal sole

Oxygen consumption in Solea senegalensis increased during the egg stage reaching values close to 4 nmol O₂ ind⁻¹ at hatching. After hatching, larval oxygen consumption continued to increase, reaching a maximum rate of 9.97⁻¹±87 nmol O₂ ind⁻¹ h⁻¹ 2 days after the opening of the mouth. Body nitrogen content decreased mainly after exhaustion of yolk reserves. Carbon content decreased during the whole endogenous feeding phase, although it decreased twice as quickly after yolk-sac absorption. The free amino acid (FAA) depletion rate was higher during egg development and the yolk-sac period. Complete yolk absorption coincided with the consumption of the 90% of initial FAA content in the eggs and the remaining FAA were consumed at a lower rate. Based on stoichiometrical calculations, FAA appears to be the most important energy substrate during the egg stage (86%) in the Senegal sole. During the period from hatching to the mouth opening, contributions of FAA and lipids as metabolic fuels were similar (41 and 47%, respectively). The decrease in larval protein content during starvation indicates that amino acids from body protein are used as energy substrates under food deprivation.     

Histochemistry of mucosubstances in the gallbladder epithelium of the chick embryo

Mucosubstances of epithelial cells of the chick embryo gallbladder were investigated by histochemical methods from days 12-21 of incubation (stages 38-46). On incubation day 14, only neutral mucins were detected; on day 15, neutral and sulfated mucosubstances were observed in the epithelial cells that invaginate the underlying mesenchyme. In the same sites, at day 16 of incubation, neutral, carboxylated and sulfated mucins were seen. From the 17th day of incubation until hatching, neutral, carboxylated and sulfated mucosubstances were present in the surface cells and in the cells lining the epithelial invaginations. During this period, the chemical characteristics of the secretory material are similar to those observed by Yamada and Hoshino (1972) in the fowl.     

Physiological status and change of cytoplasmic lipid droplets in the pheromone-producing cells of the silkmoth, Bombyx mori (Lepidoptera, Bombycidae)

Changes in size and number of cytoplasmic lipid droplets were quantified in the pheromone gland (PG) of Bombyx mori before and after adult eclosion. Two days before eclosion, size and number of droplets are small (diameter is 2-7 microm) and few. The formation and significant proliferation of larger droplets (5-12 microm) take place between 2 days and 1 day before eclosion. From the day of emergence until day 3 a fluctuation in size and number of lipid droplets during the photophase (4h intervals) is observed. The changes are more characteristic and dramatic on the day of emergence and first day, while attenuation of these changes can be observed from the second day and seems to disappear by day 4. Bombykol content, at each respective time, is in good correlation with the observed fluctuation in lipid droplet parameters. Highest bombykol production daily is observed towards the early evening, when lipid droplets are the smallest (2-4 microm) and most numerous. By day 4, however, this regularity also ceases. In 24h old mated females PG cell structure is quite similar to newly emerged ones. In glands of 72 h old decapitated females the formation of 'extra' large lipid droplets is remarkable. In vivo pheromone biosynthesis activating neuropeptide (PBAN) treatment, however, induced the formation of many small droplets, although numerous larger ones also remained. The morphological changes in lipid droplets and cellular dynamics associated with the external signal of PBAN in the PG suggest a storage-pool function of the lipid droplets.     

Assessment of digestive enzymes activity during the fry development of the endangered Caspian brown trout Salmo caspius

The study of digestive enzymes activity at Salmo caspius fry showed that enzymes were available at the moment of mouth opening on the first day post hatching (dph) and the activity of enzymes showed no significant difference from the hatching day 28 dph. An increased activity was seen between 32 and 43 dph and this activity was significantly higher than the activity during the first 28 days. In the primary stages after yolk sac resorption (43–58 dph), enzymes activity showed an increased profile, however none of them showed a significant difference between 43 and 58 dph.     

Histochemistry of mucosubstances in the gallbladder epithelium of the chick embryo

Summary Mucosubstances of epithelial cells of the chick embryo gallbladder were investigated by histochemical methods from days 12–21 of incubation (stages 38–46). On incubation day 14, only neutral mucins were detected; on day 15, neutral and sulfated mucosubstances were observed in the epithelial cells that invaginate the underlying mesenchyme. In the same sites, at day 16 of incubation, neutral, carboxylated and sulfated mucins were seen. From the 17th day of incubation until hatching, neutral, carboxylated and sulfated mucosubstances were present in the surface cells and in the cells lining the epithelial invaginations. During this period, the chemical characteristics of the secretory material are similar to those observed by Yamada and Hoshino (1972) in the fowl.Dedicated to Professor Dr. T.H. Schiebler on the occasion of his 65th birthday     

Early development of the digestive tract of cod larvae, Gadus morhua L., during start-feeding and starvation

Cod larvae, Gadus morhua L., were reared in the laboratory and released to a large marine enclosure 4 to 5 days after hatching (6–8° C). The development of the digestive system was studied until day 24 after hatching. Morphological investigations of the jaw apparatus and the digestive tract showed that the larvae are able to absorb ingested food well before exhaustion of the yolk sac. The foregut, and especially the midgut, were particularly active in lipid absorption, and the hindgut was characterized by pinocytotic activity. Duhng the first days of feeding, no distinct prey organisms were observed in the gut, and signs of food absorption in the epithelial cells of the gut were sparse.A distinct red fluorescence, restricted to the hindgut, was observed from day 11 to day 19. On the basis of changes in absorptive pattern in the gut we suggest that changes in digestive and absorptive abilities, as well as in nutritional needs, take place around days 15–17 after hatching.
In starved larvae, signs of degeneration of the gut tissue were first visible in the foregut. By day 9 after hatching, microvilli was degenerated to such an extent that the ability to absorb food must have been severely restricted. If larvae are starved longer than this, they will probably not survive.