The 1-acylglycerol-3-phosphate acyltransferase (AGPAT) acts as a crucial enzyme in the process of triacylglycerol (TAG) synthesis, enabling the acylation of lysophosphatidic acid (LPA) into phosphatidic acid (PA). In order to decode the distinctive roles of AGPAT isoforms in the TAG production pathway, three AGPAT isoforms were detected for the first time in the Chinese mitten crab Eriocheir sinensis (Es-agpat2, Es-agpat3, and Es-agpat4). The mRNA levels of Es-agpat2 and Es-agpat4 demonstrated a conspicuous presence in the hepatopancreas, with subsequent high levels in the heart, muscle, and thoracic ganglion. On the other hand, the thoracic ganglion exhibited abundant levels of Es-agpat3, while other tissues recorded relatively low expression levels. Observing the molting cycle of E. sinensis, the hepatopancreas showed minimum expression levels of Es-agpat2 and Es-agpat4 at stage A/B. A peak at stage C was noted, which was then followed by a gradual drop until stage E. For the ovarian development cycle, stage II witnessed the maximum expression level of Es-agpat2 and Es-agpat4, succeeded by a sharp fall in stage III. After this, there was an increasing trend from stage III up to stage V. Expression of Es-agpat3 in the hepatopancreas was consistently lower than Es-agpat2 and Es-agpat4 during either the molting or ovarian development. However, in terms of ovarian expression, Es-agpat3 outperformed Es-agpat2 and Es-agpat4. It exhibited a steep increase in expression, peaking at stage II and subsequently diminishing. In situ hybridization (ISH) revealed that in stages II and IV hepatopancreas, Es-agpat4-mRNA was primarily located in fibrillar cells (F cell) and resorptive cells (R cell), with no signal from Es-agpat3. During stage II of ovarian development, both Es-agpat3-mRNA and Es-agpat4-mRNA were located in the cytoplasm of previtellogenic oocyte (PRO) and endogenous vitellogenic oocyte (EN), with no expression at stage IV. Additionally, the silencing of Es-agpat2 and Es-agpat4 caused a downward trend in the expression levels of all subsequent genes in the E. sinensis TAG synthesis pathway. To sum up, these findings suggest that the three Es-agpats may have unique functions in TAG synthesis during either the molting process or ovarian maturation of E. sinensis.
Interacting effects of feeding and stress on corticoid responses in fish were investigated in common carp fed 3.0% or 0.5% body mass (BM) which received no implant, a sham or a cortisol implant (250 mg/kg BM) throughout a 168 hour post-implant period (168 h-PI). At 12h-PI, cortisol implants elevated plasma cortisol, glucose and lactate. Plasma osmolality and ions remained stable, but cortisol increased gill and kidney Na(+)/K(+) ATPase (NKA) and H(+) ATPase activities. Gill NKA activities were higher at 3%-BM, whereas kidney H(+) ATPase activity was greater at 0.5%-BM. Cortisol induced liver protein mobilization and repartitioned liver and muscle glycogen. At 3%-BM, this did not increase plasma ammonia, reflecting improved excretion efficiency concomitant with upregulation of Rhesus glycoprotein Rhcg-1 in gill. Responses in glucocorticoid receptors (GR1/GR2) and mineralocorticoid receptor (MR) to cortisol elevation were most prominent in kidney with increased expression of all receptors at 24 h-PI at 0.5%-BM, but only GR2 and MR at 0.5%-BM. In the liver, upregulation of all receptors occurred at 24 h-PI at 3%-BM, whilst only GR2 and MR were upregulated at 0.5%-BM. In the gill, there was a limited upregulation: GR2 and MR at 72 h-PI and GR1 at 168 h-PI at 3%-BM but only GR2 at 72 h-PI at 0.5%-BM. Thus cortisol elevation led to similar expression patterns of cortisol receptors in both feeding regimes, while feeding affected the type of receptor that was induced. Induction of corticoid receptors occurred simultaneously with increases in Rhcg-1 mRNA expression (gill) but well after NKA and H(+) ATPase activities increased (gill/kidney).
In the present study, the combined effects of hypoxia and nutritional status were examined in common carp (Cyprinus carpio), a relatively hypoxia tolerant cyprinid. Fish were either fed or fasted and were exposed to hypoxia (1.5-1.8mg O2L(-1)) at or slightly above their critical oxygen concentration during 1, 3 or 7days followed by a 7day recovery period. Ventilation initially increased during hypoxia, but fasted fish had lower ventilation frequencies than fed fish. In fed fish, ventilation returned to control levels during hypoxia, while in fasted fish recovery only occurred after reoxygenation. Due to this, C. carpio managed, at least in part, to maintain aerobic metabolism during hypoxia: muscle and plasma lactate levels remained relatively stable although they tended to be higher in fed fish (despite higher ventilation rates). However, during recovery, compensatory responses differed greatly between both feeding regimes: plasma lactate in fed fish increased with a simultaneous breakdown of liver glycogen indicating increased energy use, while fasted fish seemed to economize energy and recycle decreasing plasma lactate levels into increasing liver glycogen levels. Protein was used under both feeding regimes during hypoxia and subsequent recovery: protein levels reduced mainly in liver for fed fish and in muscle for fasted fish. Overall, nutritional status had a greater impact on energy reserves than the lack of oxygen with a lower hepatosomatic index and lower glycogen stores in fasted fish. Fasted fish transiently increased Na(+)/K(+)-ATPase activity under hypoxia, but in general ionoregulatory balance proved to be only slightly disturbed, showing that sufficient energy was left for ion regulation.
The objective of this study was to investigate the interaction between feeding, exercise and cortisol on metabolic strategies of common carp over a 168h post-implant period. Feeding provided readily available energy and clearly increased muscle and liver protein and glycogen stores. Swimming, feeding and cortisol all induced aerobic metabolism by increasing oxygen consumption, and stimulated protein metabolism as demonstrated by the increased ammonia and urea excretion and ammonia quotient. Hypercortisol stimulated ammonia self-detoxifying mechanisms by enhancing ammonia and urea excretion, especially during severe exercise. At high swimming level, higher branchial clearance rates in cortisol treated fish succeeded in eliminating the elevation of endogenous ammonia, resulting in reduced plasma Tamm levels compared to control and sham implanted fish. Carp easily induced anaerobic metabolism, both during routine and active swimming, with elevated lactate levels as a consequence. Both feeding and cortisol treatment increased this dependence on anaerobic metabolism. Hypercortisol induced both glycogenesis and gluconeogenesis resulting in hyperglycemia and muscle and liver glycogen deposition, most likely as a protective mechanism for prolonged stress situations and primarily fuelled by protein mobilization.
The foraging patterns of termites are strongly related to physiological limits in overcoming desiccation stress. In this study, we examined moisture preferences and physiological characteristics of Macrotermes carbonarius (Hagen) and M. gilvus (Hagen) as both exhibit conspicuous patterns of foraging activity. Despite both species showing no significant differences in calculated cuticular permeability, and percentage of total body water, they differed greatly in rate of water loss and surface area to volume ratio. For example, M. carbonarius which had a lower surface area to volume ratio (29.26-53.66) showed lower rate of water loss and percentage of total body water loss. This also resulted in higher LT(50) when exposed to extreme conditions (≈2% RH). However, contrasting observations were made in M. gilvus that has smaller size with higher surface area to volume ratio of 40.28-69.75. It is likely that the standard equation for calculating insect surface areas is inadequate for these termite species. The trend was further supported by the result of a moisture preference bioassay that indicated M. carbonarius had a broader range of moisture preference (between 5% and 20%) than M. gilvus which had a relatively narrow moisture preference (only 20%). These results explain why M. carbonarius can tolerate desiccation stress for a longer period foraging above-ground in the open air; while M. gilvus only forages below ground or concealed within foraging mud tubes.
Physiological responses to social isolation stress were compared in 56-day-old male Japanese quail. Birds were fed pretreated diets for 3 days as follows: (i) Basal diet (control); (ii) Basal diet+1500 mg/kg metyrapone (BM); (iii) Basal diet+30 mg/kg corticosterone (BCO); (iv) Basal diet+250 mg/kg ascorbic acid (BC); (v) Basal diet+250 mg/kg α-tocopherol (BE); (vi) Basal diet+250 mg/kg ascorbic acid and 250 mg/kg α-tocopherol (BCE). The birds were subsequently socially isolated in individual opaque brown paper box for 2 hours. Plasma corticosterone (CORT) concentration and heart and brain heat shock protein 70 (Hsp 70) expressions were determined before stress and immediately after stress. Two hours of isolation stress elevated CORT concentration significantly in the control and BE birds but not in the BC, BCE and BM birds. There was a significant reduction in CORT concentration after isolation stress in the BCO group. Isolation stress increased Hsp 70 expression in the brain and heart of control and BM birds. However, brain and heart Hsp 70 expressions were not significantly altered in the isolated BC, BCE and BE birds. Although, the CORT concentration of BM birds was not affected by isolation stress, Hsp70 expression in both brain and heart were significantly increased. Moreover, exogenous corticosterone supplementation did not result in elevation of Hsp 70 expression. It can be concluded that, although Hsp 70 induction had not been directly affected by CORT concentration, it may be modulated by the HPA axis function via activation of ACTH.
There is very little information on the capacity of freshwater carnivorous fish to biosynthesize highly unsaturated fatty acids (HUFA). The striped snakehead fish (Channa striata) is a carnivorous species cultured inland of several Southeast Asian countries due to its pharmaceutical properties in wound healing enhancement. We described here the full-length cDNA cloning of a striped snakehead fatty acid desaturase (fads), which is responsible for desaturation of unsaturated fatty acids in the HUFA biosynthesis. Bioinformatics analysis reveals a protein coding region with length of 445 amino acids containing all characteristic features of desaturase enzyme, including a cytochrome b5-domain with the heme-binding motif, two transmembrane domains and three histidine-rich regions. The striped snakehead fads amino acid sequence shares high similarity with known fads of other teleosts. The mRNA expression of striped snakehead fads also showed an ontogenic-related increase in expression in 0-20 days after hatch larva. Using ISH, we localized the presence of fads in larva brain, liver and intestinal tissues.
Heat production (HP) of male and female mouse deer during eating, standing and sitting was determined using the open circuit respiration chamber (RC). The time taken for similar activities was also determined in an outdoor enclosure (OD). The animals were fed kangkong (Ipomoea aquatica), sweet potato (Ipomoea batatas) and rabbit pellet ad libitum. Male mouse deer consumed more dry matter (DM), organic matter (OM) and gross energy (GE) than female. The time for each activity of male and female mouse deer kept in RC and OD was similar. The average time spent in RC and OD for both male and female, respectively, for sitting (956 and 896 min/day) was significantly (P<0.01) longer than standing (463 and 520 min/day) and eating (21 and 24 min/day). Heat production for male and female mouse deer, respectively, during eating was the highest (0.44 and 0.43 kJ/kg W(0.75)/min) followed by standing (0.37 and 0.33 kJ/kgW(0.75)/min) and sitting (0.26 and 0.26 kJ/kg W(0.75)/min). The difference in HP per min during standing between male and female was significant (P<0.05). The HP for 08.00-14.00 h and 14.00-20.00 h periods were higher than 20.00-02.00 h and 02.00-08.00 h periods. The overall HP for males during 08.00-14.00 h and 14.00-20.00 h periods were significantly (P<0.05) higher (114.8 and 119.2 kJ/kg W(0.75)) than female (107.5 and 110.4 kJ/kg W(0.75)), respectively.
There is a lack of understanding on how the environment and trophic niche affect the capability of long-chain polyunsaturated fatty acids (LC-PUFA) in freshwater carnivorous teleost. In this present study, we isolated and functionally characterised a fatty acyl desaturase (Fads) from the striped snakehead Channa striata. Sequence comparison and phylogenetic analysis suggested a Fads2 protein that is closely related to previously characterised Fads2 proteins from freshwater carnivorous and marine herbivorous fish species. We further demonstrated the capacity of Δ6 and Δ5 desaturation activities for this particular desaturase, with highest activities towards the conversion of omega-3 (n-3) polyunsaturated fatty acids (PUFA). Low Δ4 desaturation activity was also detected, although the significance of this at a physiological level remains to be studied. The expression of this striped snakehead Δ6/Δ5 fads2 gene was highest in brain, followed by liver and intestine. In liver, diet fortified with high LC-PUFA concentration impeded the expression of Δ6/Δ5 fads2 gene compared to vegetable oil (VO) based diets. The discovery of Δ6/Δ5 Fads2 desaturase here complements the previous discovery of a Δ4 Fads2 desaturase and an Elovl5 elongase, lending proof to the existence of all the required enzymatic machinery to biosynthesise LC-PUFA from C18 PUFA in a freshwater carnivorous species.
Red tilapia has become more popular for aquaculture production in China in recent years. However, the pigmentation differentiation that has resulted from the process of genetic breeding and skin color variation during the overwintering period are the main problems limiting the development of commercial culture. The genetic basis of skin color differentiation is still not understood. Solute carrier family 7 member 11 (slc7a11) has been identified to be a critical genetic regulator of pheomelanin synthesis in the skin of mammals. However, little information is available about its molecular characteristics, expression, location and function in skin color differentiation of fish. In this study, three complete cDNA sequences (2159 bp, 2190 bp and 2249 bp) of slc7a11 were successfully isolated from Malaysian red tilapia, encoding polypeptides of 492, 525 and 492 amino acids respectively. Quantitative real-time PCR demonstrated that slc7a11 mRNA expression is high in the ventral skin of PR (pink with scattered red spots) fish. Immunofluorescence analysis revealed that xCT (the protein encoded by slc7a11) was concentrated mainly in the cytoplasm and nucleus of both the dorsal and ventral skin cells of fish. After RNA interference of slc7a11, slc7a11 and cbs mRNA expressions decreased, but the tyr mRNA expression increased in the skin of fish. Results suggest that slc7a11 plays an important role in skin color formation and differentiation of red tilapia through the melanogenesis pathway.
Crustacean molting is highly related to energy and lipid metabolism. This study was conducted to detect the changes of total lipids (TL), triacylglyceride (TAG), phospholipid (PL) and lipid droplets in hepatopancreas, and then to investigate the gene expression patterns related to hepatopancreatic lipid metabolism during the molting cycle of Chinese mitten crab Eriocheir sinensis. Hepatopancreatic TL and TAG increased significantly from post-molt stage to pre-molt stage, then decreased significantly from pre-molt stage to ecdysis stage, which is consistent to the changes of neutral lipid-rich adipocytes in hepatopancreas. By transcriptomic analysis, 65,325 transcripts were sequenced and assembled, and 28,033 transcripts were annotated. Most genes were related to energy metabolism, and the enriched genes were involved in carbohydrate and lipid metabolism and biosynthesis, especially in de novo synthesis of fatty acids and TAG, and ketone body production. Compared to the inter-molt stages, acetyl-CoA carboxylase, fatty acid synthase and other genes related to the synthesis of fatty acids were upregulated in the pre-molt stage. TAG synthesis related genes, including Glycerol-3-phosphate acyltransferase and 1-acylglycerol-3-phosphate acyltransferases, were upregulated in the post-molt stage compared to the inter-molt stage. The expression of ketone body-related genes had no significant changes during the molting cycle. Compared to the TAG synthetic pathway, ketone body biosynthesis may contribute less/secondarily to fatty acid metabolic processes, which could be involved in the other physiological processes or metabolism. In conclusion, these results showed that TAG is the major lipid deposition during inter- and pre-molt stages, and the most genes are related to the fatty acids and TAG metabolism in the hepatopancreas during the molting cycle of E. sinensis.