A New Lyngbyatoxin from the Hawaiian Cyanobacterium Moorea producens

Lyngbyatoxin A from the marine cyanobacterium Moorea producens (formerly Lyngbya majuscula) is known as the causative agent of “swimmer’s itch” with its highly inflammatory effect. A new toxic compound was isolated along with lyngbyatoxin A from an ethyl acetate extract of M. producens collected from Hawaii. Analyses of HR-ESI-MS and NMR spectroscopies revealed the isolated compound had the same planar structure with that of lyngbyatoxin A. The results of optical rotation and CD spectra indicated that the compound was a new lyngbyatoxin A derivative, 12-epi-lyngbyatoxin A (1). While 12-epi-lyngbyatoxin A showed comparable toxicities with lyngbyatoxin A in cytotoxicity and crustacean lethality tests, it showed more than 100 times lower affinity for protein kinase Cδ (PKCδ) using the PKCδ-C1B peptide when compared to lyngbyatoxin A.     

Nitrergic inhibition of tachykininergic neuro-muscular transmission via cyclic GMP in the hamster ileum

The present study was designed to explore the inhibitory mechanism by nitric oxide (NO) of the tachykininergic neuro-muscular transmissions in the hamster ileum. In the presence of guanethidine (1 μM), atropine (0.5 μM), nifedipine (0.1 μM) and apamin (100 nM), electrical field stimuli (EFS; 0.5 ms duration, 15 V) evoked non-adrenergic, non-cholinergic excitatory junction potentials (EJPs) in circular smooth muscle cells. The EJPs were markedly inhibited by the tachykinin NK1 receptor antagonists [D-Pro(4), D-Trp(7,9)]-SP(4-11) (3 μM). Both the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME, 200 μM) and the soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ, 10 μM), did not affect on the resting membrane potentials, but enhanced the tachykininergic EJPs. In the presence of L-NAME (200 μM), exogenously applied NO (10 μM) and the membrane permeable analogue of guanosine 3',5'-cyclic monophosphate (cGMP), 8-bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP, 3 mM), significantly inhibited the tachykininergic EJPs. Application of EFS (0.5 msec duration, 15 V) with trains of 20 pulses at 20 Hz increased amount of released substance P (SP). The release of SP was further increased by the treatment of L-NAME or ODQ, but markedly reduced by exogenously applied NO and 8-Br-cGMP. These results suggest that the endogenous NO may inhibit the tachykininergic neuro-muscular transmissions by the decrease of SP release from the tachykininergic neurons, possibly through a guanylate cyclase-cGMP-dependent mechanism in the hamster ileum.     

Isolation, cloning, sequencing of brain type aromatase and its expression in male and female Wrasse, Pseudolabrus sieboldi

Pseudolabrus sieboldi, wrasse being a diurnal spawner provides a good opportunity to study the endocrine mechanism of estrogen formation in brain and gonads. Moreover, an extremely large amount of E2 was produced in serum and testis of wrasse. It is assumed that the presence of E2 may play a major role in diurnal gametogenesis in male fish. In this study brain type aromatase have been isolated, cloned and sequenced from the brain of wrasse. Further, the expression pattern of brain type aromatase in gonads and adult tissue of male and female fish have been analyzed. In addition, the diurnal expression pattern of brain type aromatase in both male and female fish brain during spawning season have been analyzed. The P450arom (br) was isolated, cloned and sequenced from both male and female bamboleaf wrasse. The P450arom (br) gene (1877 sequenced nucleotide) contains an ORF of 1470 bp, a 5′-UTR of 18 bp and at least 407 bp in 3′-UTR. The amino acid sequence homology in the coding region of wrasse P450arom (br) is high compared to that of medaka, Oryzias latipes (80%), rainbow trout type 2, Oncorhynchu mykiss (78.2%), fugu, Takifugu ribripes (78%) rainbow trout type 1, (76%), goldfish, Carassius auratus (66.8%) and zebrafish, Danio rerio (66.2%). Expression study reveals that P450arom (br) mRNA were most abundant in brains of both male and female fish throughout the day during the spawning season. RT-PCR study revealed that P450arom (br) was expressed in skin, anal fin and tail fin of both male and female wrasse. P450arom (br) was not detected at any time of the spawning day in either ovary or testis of wrasse.     

Effects of 11-ketotestosterone and gonadotropin-releasing hormone on follicle-stimulating hormone and luteinizing hormone gene expression in castrated and sham-operated male red seabream <Emphasis Type="Italic">Pagrus major</Emphasis>

ABSTRACT:   In order to clarify the roles of androgen and gonadotropin-releasing hormone (GnRH) on gonadotropin (GTH; luteinizing hormone [LH] and follicle stimulating hormone [FSH]) synthesis, effects of castration and implantation of GnRH analog (GnRHa) or 11-ketotestosterone (11-KT) on expression of GTH subunit, α-glycoprotein subunit (αGSU), FSHβ, and LHβ genes, during the early spermatogenic stage in male red seabream Pagrus major were examined. Male red seabream underwent castration or sham-operation and were subsequently implanted with cholesterol pellets containing GnRHa, silicone capsules filled with 11-KT, or blank capsules (control). FSHβ mRNA levels increased due to castration, and it was reversed by treatment with 11-KT. 11-ketotestosterone treatment also decreased FSHβ mRNA levels in sham-operated fish. These results suggest that 11-KT acts on the pituitary to suppress FSH synthesis in male red seabream. On the other hand, neither castration nor replacement of 11-KT in castrated fish had effects on LHβ mRNA levels, whereas 11-KT treatment had slightly but significantly decreased LHβ mRNA in sham-operated fish. αGSU mRNA levels were not changed by castration or 11-KT treatment in both sham-operated and castrated fish. Meanwhile, treatment with GnRHa significantly decreased FSHβ mRNA levels in sham-operated fish, but not in castrated fish. This suggests that GnRHa may down-regulate expression of FSHβ mRNA through the production of 11-KT in testis. LHβ and αGSU mRNA levels in sham-operated fish, but not in castrated fish, were significantly elevated by treatment with GnRHa.     

Diurnal rhythm of steroid biosynthesis in the testis of terminal phase male of protogynous wrasse, Pseudolabrus sieboldi, a daily spawner

The wrasse, Pseudolabrus sieboldi, is a diandric protogynous labrid fish. Spawning is performed by a terminal phase (TP) male and an initial phase (IP) female between 6:00 and 9:00 h daily during two-month-long spawning season. In the present study, to investigate the roles of steroid hormones in the diurnal spermatogenesis of the P. sieboldi TP male, all steroid hormones produced in the testis were identified and the synthetic pathways of these steroids were determined. Furthermore, the circulating levels of the major steroids produced were analyzed throughout a day at 3-hour intervals during spawning season. In the testis, 11-ketotestosterone (11-KT), estradiol-17β (E2), 17,20β-dihydoxy-4-pregnane-3-one (17,20β-P) and 17,20β,21-trihydroxy-4-pregnen-3-one (20β-S) were synthesized as the major metabolites. In vitro steroid biosynthesis experiments showed similar results to the circulation profiles of the major steroids. This study is the first to clarify the complete steroidogenic pathways in the gonads of a diandric protogynous species throughout its life, when combined with the results of the steroidogenesis in the ovarian follicles. This is also the first report of a clear diurnal rhythm of the steroid production corresponding to the spermatogenic process in the testis of a male teleost.     

Milk production,nutrient digestibility and nitrogen balance in lactating cows fed total mixed ration silages containing steam‐flaked brown rice as substitute for steam‐flaked corn,and wet food by‐products

The objective of this study was to evaluate the effect of substituting brown rice grain for corn grain in total mixed ration (TMR) silage containing food by‐products on the milk production, whole‐tract nutrient digestibility and nitrogen balance in dairy cows. Six multiparous Holstein cows were used in a crossover design with two dietary treatments: a diet containing 30.9% steam‐flaked corn (corn TMR) or 30.9% steam‐flaked brown rice (rice TMR) with wet soybean curd residue and wet soy sauce cake. Dietary treatment did not affect the dry matter intake, milk yield and compositions in dairy cows. The dry matter and starch digestibility were higher, and the neutral detergent fiber digestibility was lower for rice TMR than for corn TMR. The urinary nitrogen (N) excretion as a proportion of the N intake was lower for rice TMR than for corn TMR with no dietary effect on N secretion in milk and fecal N excretion. These results indicated that the replacement of corn with brown rice in TMR silage relatively reduced urinary N loss without adverse effects on feed intake and milk production, when food by‐products such as soybean curd residue were included in the TMR silage as dietary crude protein sources.     

Body growth and plasma concentrations of metabolites and metabolic hormones during the pubertal period in female Shiba goats

It has been shown in various species that the onset of puberty is closely associated with body growth and nutritional state rather than age. The present study was conducted to determine the timing of puberty and to clarify body growth and metabolic changes around the pubertal period in female Shiba goats. Blood samples were collected between 10 to 38 weeks of age from 12 female goats, and plasma concentrations of progesterone, metabolites (glucose, nonesterified fatty acid, ketone body and acetic acid) and metabolic hormones (insulin and insulin-like growth factor-I (IGF-I)) were analyzed. Physical parameters (body weight, withers height and body length) were also measured at the blood sampling. The week when plasma progesterone concentrations first exceeded 1.0 ng/ml was designated as the onset of puberty. The results showed that the average age of the onset of puberty was 27.0 +/- 0.9 (mean +/- SEM) weeks in female Shiba goats. When the goats reached puberty, the average values of body weight and goat body mass index ((body weight (kg)/withers height (cm)/body length (cm)) x 10(3)) were 12.2 +/- 0.5 kg and 5.7 +/- 0.2, respectively. No particular change associated with puberty was apparent for plasma concentrations of the metabolites examined. Plasma insulin concentrations were maintained at lower levels until the onset of puberty, and then they began to gradually increase. Plasma IGF-I concentrations began to gradually increase 1 to 4 weeks before the onset of puberty and this increase continued throughout the peripubertal period. These results imply that IGF-I acts as a peripheral nutritional signal to trigger the onset of puberty in Shiba goats.     

Expression patterns of the BRCA1 splicing variants in canine normal tissues and mammary gland tumors

Human BRCA1 is familial breast cancer susceptibility gene. Recently, decreased BRCA1 mRNA and protein expression has been identified in sporadic breast tumors. In the reported human BRCA1 splicing variants, delta11b lacks the majority of exon11 and is suspected to have a distinct function in normal tissues. The splicing variants display a variety of expression pattern in breast cancer samples. Although mammary gland tumor is important disease in dog, there are few reports for BRCA1 in the canine tumors. In this study, we examined the relative amounts of BRCA1 splicing variants mRNA in canine normal and mammary tumor samples by RT-PCR to investigate whether there is the altered expression of variant mRNAs in the canine tumor as reported in human. The exon11b-defecting RT-PCR products were observed in all the normal tissues examined and the nucleotide sequence was quite similar to that of human BRCA1 delta11b. In some tumor samples, we did not detect the products targeted for exon10-13 and exon14-15, while these products were observed in all the normal samples examined. Especially, the relative amounts of the exon11-defecting products were remarkably decreased in most of the tumors (11/16).     

Differential responses to high temperature during maturation in heat-stress-tolerant cultivars of Japonica rice

High-temperature stress during the grain-filling stage reduces grain quality of rice, and this is a serious problem in Japan, especially in the Kyushu region. To solve this problem, various heat-tolerant cultivars have been bred, such as ‘Nikomaru’, ‘Kumasannochikara’, ‘Genkitsukushi’, ‘Sagabiyori’, and ‘Otentosodachi’. When cultivated under high temperature after flowering, these heat-tolerant cultivars had lower percentages of chalky grains than in the heat-sensitive cultivar ‘Hinohikari’. All the heat-tolerant cultivars markedly decreased the nonstructural carbohydrate content in the stem under the high temperature compared to control condition during early grain-filling stage, which is considered to be a common trait of heat tolerance. Notably, ‘Sagabiyori’, ‘Genkitsukushi’, and ‘Nikomaru’ maintained a nucellar epidermis at 17 days after flowering (DAF) under high temperature, whereas the nucellar epidermis disappeared in ‘Hinohikari’. In addition, the expression of AGPS2b, thought to be a rate-limiting enzyme in starch synthesis, in ‘Kumasannochikara’, ‘Otentosodachi’, and ‘Nikomaru’ did not decrease under high temperature, whereas ‘Hinohikari’, ‘Sagabiyori’, and ‘Genkitsukushi’ could not maintain expression of the gene at 17 DAF. Moreover, the expression of Amy3E, a starch-degradation-related gene considered to induce grain chalkiness, in ‘Kumasannochikara’ at 17 DAF was not increased by high temperature. These results suggest that the heat-stress-tolerant cultivars have various mechanisms for dealing with high-temperature stress.     

Effect of low salinity on the expression profile of Na+/K+‐ATPase and the growth of juvenile chub mackerel (Scomber japonicus Houttuyn)

Chub mackerel (Scomber japonicus Houttuyn) is one of the most commercially important scombroid fish used as a food resource. Recently, there has been a demand for efficient rearing methods of this fish for a full‐life cycle aquaculture. In the present study, we evaluated the physiological responses in the juvenile S. japonicus to different ambient salinities. A significantly higher gain of the body mass was observed in the juveniles reared in 24 g/L and 13 g/L seawater than in those reared in natural seawater (34 g/L) within 40 days of the experimental period without affecting mortality. A principal enzyme for osmoregulation, Na⁺/K⁺‐ATPase, was expressed in the ionocytes located in the gill filaments of the juveniles. The number and the cell size of ionocytes and the enzymatic activity of Na⁺/K⁺‐ATPase in the gills decreased within 10 days after the low‐salinity challenge, which implies the reduction of the energy‐consuming active ion secretion under the low‐salinity environment. The physiological capacity for adaptation to low‐salinity seawater in chub mackerel could be basic knowledge to carry out culturing of these fish in coastal sea pens where ambient salinity fluctuates. The improvement of the growth performance by rearing in low‐salinity seawater will contribute to the efficient production of the seed juveniles for aquaculture.