A histological study on the development of the digestive system of Pseudosciaena crocea larvae and juveniles

The ontogenetic development of the gut and accessory organs in large yellow croaker Pseudosciaena crocea was investigated using light microscopy from hatching up to the juvenile stage (40 days post hatch, dph). At 3 dph (mean ±  s . d ., 4·1 ± 0·1 mm total length, L _T), coinciding with the buccopharynx opening, larvae started to feed exogenously, and the gut consisted of a well‐developed buccopharynx, a partially‐differentiated oesophagus and an intestine divided in three regions (anterior intestine, intermediate intestine and rectum). Yolk reserves were not completely depleted at the onset of exogenous feeding, and a period of mixed nutrition was observed up to 6 dph (4·3 ± 0·1 mm L _T), when yolk was definitively exhausted. Important morphological changes occurred at the end of the larval period, coinciding with metamorphosis. At 17 dph (6·8 ± 0·6 mm L _T), pyloric caeca differentiated at the junction of the pyloric stomach and the anterior intestine. Gastric glands were first observed at 21 dph (9·2 ± 1·2 mm L _T), coinciding with the morphological development of the stomach in three different regions (cardiac, fundic and pyloric) according to the histological characteristics of their mucosa. At this age, large longitudinal folds appeared in the median and posterior oesophageal mucosa. These morphological and histological features suggested the achievement of a digestive system characteristic of large yellow croaker juveniles and adults.     

Ontogeny of the Digestive System of Atlantic Salmon (Salmo salar L.) and Effects of Soybean Meal from Start-Feeding

Despite a long history of rearing Atlantic salmon in hatcheries in Norway, knowledge of molecular and physiological aspects of juvenile development is still limited. To facilitate introduction of alternative feed ingredients and feed additives during early phases, increased knowledge regarding the ontogeny of the digestive apparatus in salmon is needed. In this study, we characterized the development of the gastrointestinal tract and accessory digestive organs for five months following hatch by using histological, biochemical and molecular methods. Furthermore, the effects of a diet containing 16.7% soybean meal (SBM) introduced at start-feeding were investigated, as compared to a fishmeal based control diet. Salmon yolk sac alevins and fry were sampled at 18 time points from hatch until 144 days post hatch (dph). Histomorphological development was investigated at 7, 27, 46, 54 and 144 dph. Ontogenetic expression patterns of genes encoding key digestive enzymes, nutrient transporters, gastrointestinal peptide hormones and T-cell markers were analyzed from 13 time points by qPCR. At 7 dph, the digestive system of Atlantic salmon alevins was morphologically distinct with an early stomach, liver, pancreas, anterior and posterior intestine. About one week before the yolk sac was internalized and exogenous feeding was started, gastric glands and developing pyloric caeca were observed, which coincided with an increase in gene expression of gastric and pancreatic enzymes and nutrient transporters. Thus, the observed organs seemed ready to digest external feed well before the yolk sac was absorbed into the abdominal cavity. In contrast to post-smolt Atlantic salmon, inclusion of SBM did not induce intestinal inflammation in the juveniles. This indicates that SBM can be used in compound feeds for salmon fry from start-feeding to at least 144 dph and/or 4-5 g body weight.     

Expression and activity of trypsin and pepsin during larval development of the spotted rose snapper Lutjanus guttatus

The present study aimed to describe and understand the development of the digestive system in spotted rose snapper (Lutjanus guttatus) larvae from hatching to 40 days post-hatch (dph). The mouth opened between 2 and 3 dph, at that moment the digestive tract was barely differentiated into the anterior and posterior intestine, although the liver and pancreas were already present. Gastric glands were observed until 20 dph, followed by the differentiation of the stomach between 20 and 25 dph. Trypsinogen expression and trypsin activity were detected at hatching, increasing concomitantly to larval development and the change in the type of food. Maximum levels of trypsinogen expression were observed at 25 dph, when animals were fed with Artemia nauplii, and maximum trypsin activity was detected at 35 dph, when larvae were fed with an artificial diet. On the other hand, pepsinogen gene expression was detected at 18 dph, two days before pepsin enzymatic activity and appearance of gastric glands. Maximum pepsin activity was also observed at 35 dph. These results suggest that in this species weaning could be initiated at an earlier age than is currently practiced (between 28 and 30 dph), since larvae of spotted rose snapper develop a functional stomach between days 20 and 25 post-hatch.     

Ontogeny of the gastrointestinal tract and selected digestive enzymes in cobia Rachycentron canadum (L.)

The ontogeny of the digestive system of cobia Rachycentron canadum from hatching to 22 days post-hatch (dph) (20·1 mm standard length) was examined with light microscopy. The activities of selected pancreatic enzymes were also determined during this period in order to optimize current rearing methods for this species. At hatching (3·6 mm), the digestive tract consisted of a relatively undifferentiated, straight tube positioned dorsally to the yolk sac. The major morphological changes in the digestive tract primarily occurred over the first 1–4 dph (3·6–4·4 mm). During this time, larvae began exogenous feeding (3 dph) and the digestive tract differentiated into five histologically distinct regions: buccopharynx, oesophagus, stomach anlage, anterior intestine and posterior intestine. Yolk reserves were exhausted by 5 dph (4·5 mm) and the oil globule began rapidly decreasing in size disappearing entirely by 9–10 dph (6·3–6·8 mm). Gastric glands differentiated at this time, and by 12 dph (8·1 mm) surface mucous cells of the stomach anlage stained positive for neutral mucosubstances. By 16 dph (11·6 mm), the blind sac (fundic region) of the stomach formed as did the pyloric caecae which initially appeared as a single protrusion of the anterior intestine just ventral to the pyloric sphincter. Generally, enzyme activities (U larva⁻¹) for amylase (0·0–1·8), chymotrypsin (0·0–7902·4), trypsin (0·2–16·6) and lipase (9·3–1319·0) were measurable at or soon after hatching and increased steadily from c. 8–22 dph (5·7–20·1 mm). The results of this study are discussed in terms of current and future weaning practices of this species.     

贝氏高原鳅消化系统胚后发育的形态及组织结构

应用解剖学、组织学和组织化学方法,对贝氏高原鳅(Triplophysa bleekeri)消化系统的胚后发育进行观察.结果表明,贝氏高原鳅仔、稚鱼呈线性生长趋势.仔鱼出膜后1～2d为内源性营养阶段,3d进入混合营养阶段,15 d进入外源性营养阶段.初孵仔鱼口凹已经出现,出膜后3d与外界相通,9d口咽腔基本发育完成.8d食道发育基本完成.初孵仔鱼消化道雏形已现,但胃肠未明显分化.出膜后64 d胃小凹处出现胃腺,胃消化功能基本完备.初孵仔鱼肠道已经分化,出膜后27 d肠基本发育完成.初孵仔鱼具有肝前体,出膜后2d肝细胞开始分化,7d肝中出现明显的中央静脉和肝细胞索,肝组织结构与成体差异不大.3d肝前端出现胰组织,4d具有胰雏形,5d完整胰出现,胰腺细胞之间具有大量嗜曙红酶原颗粒物质;9d胰岛出现,胰组织基本发育完成.64 d消化系统各部分组织结构发育基本完成.贝氏高原鳅消化道的形态发育需要很长的时间,出膜后64 d胃肠仅前端膨大,无任何弯曲;85 d胃与食道呈直角弯曲后下行,但胃肠无明显分界;120 d胃弯曲为“Z”形后笔直下行,胃肠仍无明显分界,肝为一整体,未见分叶.1龄幼鱼,消化系统解剖结构与成鱼相似,但肝缺少右叶,肠缺少胃背面的圆环形弯曲.贝氏高原鳅消化系统的胚后发育特点和仔鱼的营养方式可能体现了长江以南地区冬天繁殖鱼类消化系统胚后发育的一般规律和仔鱼的营养趋势.     

Histological development of the digestive system of Mayan cichlid Cichlasoma urophthalmus (Günther 1862)

The ontogeny of the digestive tract in Cichlasoma urophthalmus was studied by means of optical microscopy from hatching to 30 days post‐hatching (dph; 855 degree days, dd). The development of the digestive system in this precocial species was a very intense and asynchronous process, which proceeded from both distal ends interiorly. At hatching, the digestive tract consisted of a straight tube with a smooth lumen dorsally attached to the yolk‐sac. The digestive accessory glands were already differentiated and eosinophilic zymogen granules were visible in the exocrine pancreas. At the onset of exogenous feeding between 5 and 6 dph (142.5–171.0 cumulative thermal units, CTU), the buccopharynx, oesophagus, intestine, liver and pancreas were almost completely differentiated, with the exception of the gastric stomach that completed its differentiation between 11 and 14 dph (313.5–399.0 CTU). The development of gastric glands at 14 dph and the differentiation of the stomach in the fundic, cardiac and pyloric regions at 19 dph (541.5 CTU) were the last major events in digestive tract development and designated the onset of the juvenile period. Remnants of yolk were still detected until 16 dph (456.0 CTU), indicating a long period of mixed nutrition that lasted between 10 and 11 days (285.0–313.5 CTU). The results of the organogenesis of larvae complement previous data on the functionality of the digestive system and represent a useful tool for establishing the functional systemic capabilities and physiological requirements of larvae to ensure optimal welfare and growth under aquaculture conditions, which might be useful for improving current larval rearing practices for this cichlid species.     

Differentiations of 5-HT and GAS cells in the digestive canals of Rana chensinensis tadpoles

In the current study, 5-nydroxytryptamine(5-HT) and gastrin(GAS) cells in the digestive canals of Rana chensinensis tadpoles at different developmental stages were investigated by immunohistochemistry. Results showed that the 5-HT cells were only detected in the duodenum before metamorphosis began, and were extensively distributed in the stomach, duodenum, small intestine, and rectum thereafter, with the highest counts found in the duodenum and rectum when metamorphosis was completed. The GAS cells were only distributed in the stomach and duodenum, and only rarely detected in the duodenum before metamorphosis began, but increased in the stomach during metamorphosis and showed zonal distribution in the gastric mucosa when metamorphosis was completed. Metamorphosis is a critical period for amphibians, during which structural and functional physiological adaptations are required to transition from aquatic to terrestrial environments. During metamorphosis, the differentiations of 5-HT cells in the gastrointestinal canals of tadpoles could facilitate mucus secretion regulation, improve digestive canal lubrication, and help watershortage food digestion in terrestrial environments. Conversely, GAS cell differentiations during metamorphosis might contribute to the digestive and absorptive function transition from herbivore to omnivore.     

Ontogeny of gene expression of group IB phospholipase A₂ isoforms in the red sea bream, Pagrus (Chrysophrys) major

The red sea bream (Pagrus major) was previously found to express mRNAs for two group IB phospholipase A₂ (PLA₂) isoforms, DE-1 and DE-2, in the digestive organs, including the hepatopancreas, pyloric caeca, and intestine. To characterize the ontogeny of the digestive function of these PLA₂s, the present study investigated the localization and expression of DE-1 and DE-2 PLA₂ genes in red sea bream larvae/juveniles and immature adults, by in situ hybridization. In the adults, DE-1 PLA₂ mRNA was expressed in pancreatic acinar cells. By contrast, DE-2 PLA₂ mRNA was detected not only in digestive tissues, such as pancreatic acinar cells, gastric glands of the stomach, epithelial cells of the pyloric caeca, and intestinal epithelial cells, but also in non-digestive ones, including cardiac and lateral muscle fibers and the cytoplasm of the oocytes. In the larvae, both DE-1 and DE-2 PLA₂ mRNAs first appeared in pancreatic tissues at 3 days post-hatching (dph) and in intestinal tissue at 1 dph, and expression levels for both gradually increased after this point. In the juvenile stage at 32 dph, DE-1 PLA₂ mRNA was highly expressed in pancreatic tissue, and DE-2 PLA₂ mRNA was detected in almost all digestive tissues, including pancreatic tissue, gastric glands, pyloric caeca, and intestine, including the myomere of the lateral muscles. In conclusion, both DE-1 and DE-2 PLA₂ mRNAs are already expressed in the digestive organs of red sea bream larvae before first feeding, and larvae will synthesize both DE-1 and DE-2 PLA₂ proteins.     

Non-specific carboxylic esterase activity in the digestive tract of the perch, Perca fluviatilis L.

The distribution of non-specific carboxylic esterases (Ec 3.1.1) in the digestive tract of perch, Perca fluviatilis L., was investigated histochemically using 1-naphthyl acetate as the substrate. Strong enzymatic activity was present in the gastric glands and surface cells of the stomach, intestinal mucosa of the pyloric caeca, upper and middle intestine, pancreas (exocrine cells) and liver. The enzymatic activity in the lower intestine and rectum was weak. The activity was not demonstrated in the oesophagus or pyloric sphincter. In the intestine, the activity was localized in the columnar cells especially in the supranuclear cytoplasm. The enzymatic activity demonstrated in the digestive tract of perch using 1 -naphthyl acetate represents combined esterolytic and lipoproteolytic activity.     

A histological study of the organogenesis of the digestive system in bay snook Petenia splendida Günther, 1862 from hatching to the juvenile stage

In bay snook (Petenia splendida) larvae the histological development of the digestive system and swim bladder, and their relative timing of differentiation were studied from hatching to 45 days post‐hatch (dph) at 29°C. Newly hatched larvae showed a simple digestive tract, which appeared as a straight undifferentiated tube lined by a single layer of columnar epithelial cells (future enterocytes). The anatomical and histological differentiation of the digestive tract and accessory glands was a very intense, asynchronous process, proceeding from the distal to the anterior part. The intestine was the first region to differentiate (9 days post‐hatch – dph, 6.5 mm SL), and the oesophagus the last (21 dph, 8.4 mm SL). At the onset of feeding, the digestive system was organized into different functional and histologically differentiated sections, such as the buccopharynx, oesophagus, glandular stomach, and anterior and posterior intestine. This organization resembled that of the juveniles, with the exception of pharyngeal teeth and buccopharyngeal as well as oesophageal goblet cells, which proliferated later during the mixed feeding period. Histological observations revealed that bay snook larvae retained endogenous yolk reserves until 24 dph (8.9 ± 0.4 mm SL), which might be helpful for weaning this species onto a compound diet. The important lipidic accumulation observed in the intestinal mucosa, liver, and pancreas in fish fed a compound trout diet indicated that although fish were able to digest and absorb lipids, the diet formulation did not fit the nutritional requirements of early juveniles of this species. The ontogeny of the digestive system followed the same general pattern as in most cichlid species described to date. However, we detected species‐specific differences in the timing of differentiation that were related to their reproductive guild. According to the histological results, some recommendations regarding the intensive culture of this species are also provided.     

The ontogeny of the alimentary tract of larval pandora, Pagellus erythrinus L.

The ontogenesis of the alimentary tract and its associated structures (liver, pancreas, gall bladder) was studied in common pandora Pagellus eythrinus L., a promising species for diversification in Mediterranean aquaculture. Mass production of pandora has been limited so far by high larval and juvenile mortalities, which appear to be related to nutritional deficiencies. The development of the larval digestive system was studied histologically from hatching (0 DAH) until day 50 (50 DAH) in reared specimens, obtained by natural spawning from a broodstock adapted to captivity. At first feeding (3–4 DAH) both the mouth and anus had opened and the digestive tract was differentiated in four portions: buccopharynx, oesophagus, incipient stomach and intestine. The pancreas, liver and gall bladder were also differentiated at this stage. Soon after the commencement of exogenous feeding (5–6 DAH), the anterior intestinal epithelium showed large vacuoles indicating the capacity for absorption of lipids, whereas acidophilic supranuclear inclusions indicating protein absorption were observed in the posterior intestinal epithelium. Both the bile and main pancreatic ducts had opened in the anterior intestine, just after the pyloric sphincter, at this stage. Intestinal coiling was apparent since 4 DAH, while mucosal folding began at 10 DAH. Scattered mucous cells occurred in the oral cavity and the intestine, while they were largely diffused in the oesophagus. Gastric glands and pyloric caeca were firstly observed at 28 DAH and appeared well developed by 41 DAH, indicating the transition from larval to juvenile stage and the acquisition of an adult mode of digestion.     

Growth of organ systems of Dentex dentex(L) and Psetta maxima(L) during larval development

Growth in volume of common dentex Dentex dentex and turbot Psetta maxima during larval development was studied by means of a quantitative histological method. A two‐phase pattern of volume increase was recorded for both species, turbot volume being always higher than dentex volume. During the first phase, the increase was small but during the second phase volume rose sharply from 22 days post hatch (dph) and 17 dph onwards in dentex and turbot, respectively. In dentex, the specific growth rate ( G ) of the whole larva as well as that of all the structures studied (nervous tissue, trunk musculature, digestive tract, liver, pancreas, spleen and thymus) was always higher during the second phase, whereas in turbot, only total volume of the larva, trunk musculature and nervous tissue had a higher G during the same period. The pattern of allometric growth of digestive organs was similar for both species. These organs showed an initial positive allometric growth that later became near‐isometric (digestive tract and liver) or negative (pancreas). In dentex, nervous tissue and trunk musculature showed near‐isometry throughout the period studied. In turbot, nervous tissue exhibited negative allometry and trunk musculature changed from negative to positive allometry. In both species studied, the highest allometry coefficients were recorded for digestive organs before the larva switched to strict exotrophy. This would indicate the importance of the development of these organs for survival.     

Morphology of the digestive system in carnivorous freshwater dourado Salminus brasiliensis

The digestive system of teleost shows remarkable functional and morphological diversity. In this study, the digestive tract and accessory organs of dourado Salminus brasiliensis are characterized using anatomical, histological, histochemical and immunohistochemical analyses. The existence of taste buds bordered by microridges in the oesophagus of dourado was recorded for the first time, thus showing that the species drives food intake by either swallowing or rejecting the food item. The Y-shaped stomach of dourado consisted of cardiac, cecal and pyloric regions with tubular gastric glands registered solely in the cardiac and cecal segments. The intestine is a short N-shaped tube with two loops, an intestinal coefficient of 0.73. The structure of pyloric caeca is similar to that of the intestine wall, comprising tunica mucosa, tela submucosa, tunica muscularis and tunica serosa layers. Histochemical analyses revealed an increased incidence of goblet cells from the midgut to the hindgut segment. A well-developed enteric plexus of scattered nerve cell and fibres are found along the digestive tract, and the calcitonin gene-related peptide (CGRP) immunoreactive neurons and fibres were identified in the myenteric plexus from the oesophagus to the hindgut. The exocrine pancreas appears diffuse in the mesentery around the stomach, intestine and also reaches the liver, and the endocrine pancreas is organized as a few islets of Langerhans. The liver comprises three distinct, asymmetric lobes, and the portal triad arrangement was registered in this tissue.     

Effects of diets containing soybean meal on trypsin mRNA expression and activity in Atlantic salmon (Salmo salar L)

Atlantic salmon develop subacute enteritis in the distal intestine (DI) when fed diets containing soybean meal (SBM) at high levels, a condition accompanied by increased trypsin activity in the DI intestinal content compared to fish fed conventional fishmeal (FM) based diets. To further investigate the responses of Atlantic salmon to dietary SBM, we measured trypsin activity in intestinal contents, quantified pancreatic trypsin mRNA expression, surveyed trypsin mRNA expression in selected tissues and characterized active forms of trypsin in the intestinal wall and brain. Enzyme measurements showed that trypsin activity in the intestinal content of SBM fed fish was lower in the proximal segments of the intestine, but higher in the DI compared to FM fed fish. The difference in enzyme activity was not reflected in a differential expression of pancreatic trypsin mRNA between fish fed the different diets (FM or SBM). Trypsin mRNA was expressed in 18 different tissues (esophagus, stomach, pancreas, pyloric tissue, midintestine, distal intestine, liver, head kidney, kidney, heart, spleen, thymus, brain, eye, gills, gonads, muscle and skin) but was most prominently expressed in tissues of the gastrointestinal (GI) tract and brain. We report for the first time an upregulation of trypsin-like activity in the DI wall using an in-gel trypsin activity assay, as well as modulated activity in the brain of fish fed SBM. The increased activity in the DI wall may contribute to disease severity and higher trypsin activity in the intestinal content.     

Cloning and expression of xP1-L, a new marker gene for larval surface mucous cells of tadpole stomach in Xenopus laevis

The amphibian gastrointestinal tract is remodeled from a larval-type to an adult-type during metamorphosis. In the present study, we examined the products of subtractive hybridization between tadpole and frog stomach cDNAs of Xenopus laevis in order to identify genes expressed specifically in the larval stomach epithelium. A new gene homologous to xP1 was obtained and named xP1-L. In the genome database of Silurana tropicalis, we found a homologue of xP1-L and named it stP1-L. RT-PCR showed that the expression of xP1-L was detected in stage 41/42 tadpoles. In addition, in situ hybridization showed that xP1-L was localized to surface mucous cells of the larval stomach. The H(+)/K(+)-ATPase beta subunit, a marker gene for manicotto gland cells in the tadpole stomach, was also detected at the same time. However, adult marker genes such as xP1 for surface mucous cells and pepsinogen C (PgC) for oxynticopeptic cells were not expressed in the tadpole stages. The expression of xP1-L gradually decreased towards the metamorphic climax and disappeared after stage 61 when larval-type gastric epithelium is replaced by adult-type. We found that xP1-L was never expressed in surface mucous cells of the adult-type stomach, and xP1, instead of xP1-L, was expressed. During T3-induced metamorphosis, xP1-L expression decreased in the same manner as during natural metamorphosis. Thus, xP1-L is a useful marker for larval surface mucous cells in tadpole stomach. This is the first demonstration of a marker gene specific for the surface mucous cells of the larval stomach.     

Studies on the Diet,Feeding Mechanism and Alimentary Tract in Two Closely Related Teleosts,the Freshwater Cottus gobio L. and the Marine Parenophrys bubalis Euphrasen

The diet and feeding mechanism in Cottus gobio and Parenophrys bubalis are described, together with the morphology and histology of the alimentary tract. Both species are sluggish bottom dwelling, carnivorous fish, and are capable of catching and swallowing relatively large prey. The gut is fully differentiated into esophagus, stomach, intestine with pyloric ceca, and rectum. The liver is morphologically separate from the pancreas, and separate bile and pancreatic ducts open into the base of one of the pyloric ceca. The organisation of the gut is well suited to the fishes' mode of life, showing adaptations for taking large meals which may be at irregular intervals.     

Thyroid hormone-induced expression of Sonic hedgehog correlates with adult epithelial development during remodeling of the Xenopus stomach and intestine

Sonic hedgehog (Shh) was isolated from the Xenopus laevis intestine as an early thyroid hormone (TH) response gene. To investigate possible roles of TH-upregulated expression of Shh during metamorphosis, we raised a polyclonal antibody against Xenopus Shh and immunohistochemically examined the relationship between Shh expression and the larval-to-adult intestinal remodeling at the cellular level. Our results indicate that the epithelial-specific expression of Shh in the intestine spatiotemporally correlates well with active proliferation and/or initial differentiation of the secondary (adult) epithelial primordia that originate from stem cells, but not with apoptosis of the primary (larval) epithelium. Given the similar transformations of the stomach during metamorphosis, we also analyzed Shh expression in this organ and found similar correlations in the stomach, although the position of the adult epithelial primordia and their final differentiation in the stomach are different from those in the intestine. Furthermore, we show here that Shh expression is organ-autonomously induced by TH and its correlation with the adult epithelial development is reproduced in vitro in both the intestine and the stomach. More importantly, addition of recombinant Shh protein to the culture medium results in developmental anomalies of both organs. However, differentiation of the adult epithelium is more severely inhibited by exogenous Shh in the intestine than in the stomach. These results suggest that TH-upregulated expression of Shh plays important roles in the postembryonic gastrointestinal remodeling, but its roles are at least partially different between the intestine and the stomach.