Polypolar spindle formation during first cell cycle in rainbow trout <Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis> embryos after heat-shock treatment

ABSTRACT:   Heat shock has been used to inhibit cleavage for the induction of monogenic diploids or tetraploids in animals, but usually the success rate is low. Heat-shocked rainbow trout Oncorhynchus mykiss embryos were used in this histological study to clarify the causes of this low success rate. Embryos treated with hydrostatic pressure were used for comparison. After heat shock had disorganized the spindles, polypolar (tripolar or tetrapolar) spindles in addition to bipolar spindles were often reassembled soon after treatment. The embryos then completed tripolar or tetrapolar division at the first mitosis, and directly turned into three- or four-cell embryos as a result of the first cleavage. During the second mitosis, a monopolar spindle was formed in each blastomere of four-cell embryos and approximately 60% of three-cell embryos. In the remaining three-cell embryos, two of the three blastomeres formed a monopolar spindle, and the third one formed a bipolar spindle. The formation of polypoles is assumed to be caused by insufficient disorganization of daughter centrioles and splitting from the mother centriole by heat shock. Polypolar division is considered to be the cause of aneuploidy and the low success rate of chromosome set doubling. In the case of hydrostatic pressure treatment, the regenerated spindles were bipolar in almost all embryos.     

Wind‐induced stock variation of the neon flying squid (Ommastrephes bartramii) winter–spring cohort in the subtropical North Pacific Ocean

Our examination of the neon flying squid (Ommastrephes bartramii) winter–spring cohort catch per unit effort (CPUE, an index of stock) revealed significant positive correlations with the interannual variations of observed chlorophyll‐a (Chl‐a) concentration and autumn–winter mixed layer depth (MLD) in the winter–spring feeding grounds of paralarvae and juveniles (130–170°E, 20–27°N). These correlations suggest the importance of integrated bottom‐up effects by the autumn–winter MLD for the neon flying squid stocks. However, the influence of autumn–winter MLD interannual variation in the forage availability for paralarvae and juveniles, i.e., particulate organic matter and zooplankton, has still been unclear. In this study, we use the lower trophic ecosystem model NEMURO, which uses the physical environmental data from the ocean reanalysis dataset obtained by the four‐dimensional variational (4DVAR) data assimilation method. The model‐based investigation enables us to clarify how the autumn–winter MLD controls the particulate organic matter and zooplankton abundance in the feeding grounds. Further, our investigation of the autumn–winter MLD interannual variation demonstrates that the stronger autumn wind in the feeding grounds develops a deeper mixed layer. Therefore, the deep mixed layer entrains nutrient‐rich water and enhances photosynthesis, which results in good feeding conditions for paralarvae and juveniles. Our results underline that the wind system interannual variation has critical roles on the winter–spring cohort of the neon flying squid stock.     

Effects of alcohol extract of defatted soybean meal on growth performance and digestive physiology of yellowtail <Emphasis Type="Italic">Seriola quinqueradiata</Emphasis>

The effects of alcohol extract of soybean meal on growth performance, digestibility, and pancreatic enzyme and bile acid status of yellowtail Seriola quinqueradiata were studied. Fish were fed one of the following for 8 weeks: a fish meal (FM) control diet (FMD), a defatted soybean meal diet (SBMD), an alcohol-extracted SBM diet (ExSBMD), an ExSBMD with alcohol-extract diet (ExSBM + ExtD), and an FMD with alcohol-extract diet (FM + ExtD). Growth performance was significantly inferior in fish fed with SBM, ExSBM + ExtD, and FM + ExtD compared to FMD- or ExSBMD-fed fish. Total bile acid levels, and trypsin and lipase activities in the anterior intestine were significantly lower in fish fed with SBM, ExSBM + ExtD, and FM + ExtD than in fish fed with ExSBMD and FMD, despite similar values in gallbladder or pyloric caeca. Cholecystokinin mRNA levels of the former diet-fed fish were significantly lower than those of FMD- or ExSBMD-fed fish. Lipid and protein digestibility of fish fed with SBMD, ExSBM + ExtD, and FM + ExtD was significantly reduced in comparison with that of fish fed with FMD or ExSBMD. These findings indicate that the alcohol extract of soybean meal inhibited the secretion of bile acids and pancreatic enzymes by a decrease of cholecystokinin stimulation. This inhibition seemed to be responsible for a low growth performance through impairment of the assimilation of lipid and protein.     

Response of cholecystokinin and digestive enzyme mRNA levels to various feed ingredients in yellowtail Seriola quinqueradiata

Cholecystokinin (CCK) is the key regulator hormone that stimulates the secretion of digestive pancreatic enzymes in vertebrates. In fish, little is known about the mechanism of induction of CCK in the digestive tract by feed ingredients. To investigate the response of CCK and digestive enzymes to fish feed ingredients in yellowtail Seriola quinqueradiata, we performed a number of experiments in which we measured the mRNA levels of CCK, trypsin, and lipase after oral administration of a single bolus of the ingredients. We administered fish meal and fish oil in experiment 1; high and low concentrations of fish meal in experiment 2; and five different dietary protein sources (fish meal, soybean meal, soy protein concentrate, corn gluten meal, and glutamic acid fermentation by-products) in experiment 3. In experiments 1 and 3, only fish meal significantly increased the mRNA levels of CCK and digestive enzyme. In experiment 2, a high concentration of fish meal [20?% (w/v)] significantly increased CCK and trypsin mRNA levels, but a low concentration of fish meal [1?% (w/v)] did not. These results suggest that high concentrations of fish meal (the protein source in fish feed) has the most potent effect on stimulation of CCK synthesis and secretion of digestive enzymes in yellowtail.     

Chromosome doubling of Lychnis spp. by in vitro spindle toxin treatment of nodal segments

Lychnis (Caryophyllaceae) consists of about 30 species distributed throughout the temperate regions of the Northern Hemisphere, from East Asia to Europe. Many Lychnis spp. have high ornamental value and cultivated as pot or garden plants. In the present study, in vitro chromosome doubling of several Lychnis spp. was examined in order to widen their variability in horticultural traits. Initially effect of various spindle toxin treatments [100, 500 or 1000 mg l⁻¹ colchicine (COL), 10, 20 or 50 mg l⁻¹ oryzalin (ORY), or 1, 5, 10 mg l⁻¹ amiprophos-methyl (APM)] of nodal segments of a triploid genotype of L. senno (3x) was investigated on survival of nodal segments and chromosome doubling in nodal segment-derived plantlets. Significantly higher percentage (75.0%) of surviving segments after spindle toxin treatment was obtained in 10 mg l⁻¹ ORY treatment. Flow cytometry (FCM) analysis of leaf tissues showed that 9.4–13.8% of plantlets, which were derived from 10 to 20 mg l⁻¹ ORY, or 5 mg l⁻¹ APM treatments, were hexaploid (6x) or ploidy chimera (3x + 6x, 4x + 6x, 5x + 6x, 3x + 4x + 6x). The results obtained by FCM analysis were confirmed by chromosome observation in root tip cells. Thus 10 mg l⁻¹ ORY treatment of nodal segments is suitable for in vitro chromosome doubling of triploid L. senno. Efficient chromosome doubling was also achieved in diploid L. fulgens (2x) and L. sieboldii (2x) by treating nodal segments with 10 mg l⁻¹ ORY: 68.9–88.7% of nodal segments survived after ORY treatment, and 24.7–26.5% of plantlets derived from ORY-treated nodal segments were tetraploid (4x) or ploidy chimera (2x + 4x) in both species. These results indicate that the in vitro chromosome doubling method established for triploid L. senno may be applicable to a wide range of Lychnis spp. Tetraploid L. fulgens and L. sieboldii showed a compact plant form, and had thick stems and deep green leaves compared with the diploid mother plants. On the other hand, hexaploid L. senno showed very poor growth and died before flowering.     

The TLR Expression Pattern on Monocyte-Derived Macrophages for Lipopolysaccharid Stimulation of Calves

In this paper, toll-like receptor expression pattern in monocytes-derived macrophages by lipopolysaccharid （LPS） stimulation was examined. Jugular venous blood samples from 4 Japanese calves were obtained and the peripheral blood mononuclear cells （PBMC） were isolated. The PBMC were cultured for 7 d so as to collect monocytes-derived macrophages in Repcell. The PBMC were stimulated by LPS for 24 h and the mRNA expression pattern of TLR and cytokines in monocytes-derived macrophages （Mod-Mφ） was analyzed. Results showed that LPS stimulation of Mod-Mφ could increase the mRNA levels of the genes of TNF-α, IL-6, and IL-8. In addition, the mRNA levels of the genes of TNF-α and IL-6 in the group of LPS stimulation were most significantly （P 〈 0.01） higher than those in control group and the mRNA levels of TLR1, 3, 5, 8, and 10 were significantly （P 〈 0.05） decreased after LPS stimulation. There was no difference in the mRNA expressions of TLR2, 4, 6, and 7 between the groups of the control and LPS stimulation. Besides, expression of TLR9 was not found. It suggested that monocytes-derived macrophages could respond to LPS and they might take an important role in the innate immunity. The important function of the cells might contribute to better disease treatment.     

Molecular cloning and tissue distribution of uncoupling protein 1 (UCP1) in plateau pika (Ochotona dauurica)

Uncoupling protein 1 (UCP1) is present exclusively in brown adipose tissue, and contributes to body temperature control during cold exposure. We cloned UCP1 cDNA of plateau pika (Ochotona dauurica), a small, non-hibernating, diurnal lagomorph that inhabits in relatively cold climates and at high altitudes in Mongolia and in northern China. The nucleotide sequence of pika UCP1 was highly homologous to UCP1 of other species, and the deduced amino acid sequence had some common domains for UCP, including six mitochondrial carrier protein motifs and a putative purine-nucleotide binding site. RT-PCR and Western blot analyses revealed that both UCP1 mRNA and protein were expressed exclusively in the interscapular adipose tissue. These results suggest that pika UCP1 contributes to heat production in brown adipose tissue, as do those in other species.     

Variations in the proximate composition of muscle in cultured yellowtail Seriola quinqueradiata at different anatomical portions

In this study, we examined differences in the proximate composition of five major muscles of the yellowtail, as well as the relation between seasonal variations in the levels of crude lipids and texture of these muscles. The differences in content among these muscles and the variations in each muscle were greater for crude lipid and moisture content than for crude protein content. The crude lipid content of Musculus latero-dorsalis correlated with that of M. carinatus dorsalis, M. latero-ventralis, M. carinatus ventralis, and red muscle. In M. latero-dorsalis, the crude lipid content started to increase in October, remained high from November to January, started to decrease in March, and was low from April to June. The firmness of M. latero-dorsalis increased from October to November and started to decrease in March. Firmness decreased concurrently with decreases in crude lipid content and increased concurrently with increases in crude lipid content. The results of this study suggest that it is possible to predict the proximate compositions of other yellowtail muscles from the proximate composition of M. latero-dorsalis. Seasonal variation in crude lipid content of the M. latero-dorsalis was found, and variations in crude lipid tended to parallel those in firmness.