Ovarian Maturation Stages Research Articles

Effects of spirulina diet on the oogenesis of zebrafish: morphological analysis and immunohistochemical determination of the vitellogenin

Morphological analysis and immunohistochemical determination of vitellogenin (Vtg) were carried out in zebrafish ovaries (Danio rerio). The goal was to determine the nutraceutical effect of Spirulina feed on ovarian maturation stages of adult zebrafish. After 30 d, in the stages of advanced maturation a prevalence of oocytes was found in the experimental group. Immunohistochemical investigations revealed a significant increase in Vtg, precursor of vitelline (Vn) in egg yolk, whose expression increased in the liver in relation to the stages of maturation. In the zebrafish liver, the expression of Vtg was extremely low before vitellogenesis, and then increased and it was maintained at a high level until the final stage of ovarian maturation. The results showed that food additives such as Spirulina can improve fertility in laboratory farmed fish. The data obtained from this study may be a contribution to wider research aimed at improving reproduction in all vertebrates, including humans.

Gonadal recrudescence and annual reproductive hormone pattern of captive female Spotted Scats (Scatophagus argus)

The Spotted Scat (Scatophagus argus), an important euryhaline fish inhabiting mangrove and coastal regions of Indo-Pacific waters, is both an ornamental and food fish in India. Detailed insight into maturation of Spotted Scat when maintained in aquaculture systems, therefore, needs to be elucidated. Lack of information on annual maturation dynamics of female scat collected from their natural habitat and reared in earthen ponds is the basis of this study. Oocytes were classified into five developmental stages: pre-vitellogenic, vitellogenic, late-vitellogenic, ripe, and follicular atresia. Ovarian maturity stages were subsequently categorized as immature (Stage 1), vitellogenesis (Stage 2), maturing (Stage 3), mature (Stage 4), and spent (Stage 5). In oocytes in primary, secondary and tertiary yolk stages, there are greater concentrations of E2 in vitellogenic females between March and June. Significant increases of E2, T, and 17-OHP paralleled the increase of diameter of late–vitellogenic oocytes in maturing females during July. The completion of vitellogenesis and initiation of germinal vesicle migration in the cytoplasm were evident in mature females (Stage 4) with a decreasing trend of sex steroids in and subsequent to the month of August. There were 50 % of oocytes in the final oocyte maturation stage (FOM) (490–620 μm) until completion of Stage 4 in September. The results of this study indicate there is complete ovarian maturation in female scats captured in their natural habitat and maintained in an earthen pond, which may be important information for hatchery management for induction of spawning of Spotted Scat in aquaculture systems.

The reproductive biology of female crimson sea bream Evynnis tumifrons off the southwestern coast of Kyushu, Japan

Ovarian maturation, size at sexual maturity and the annual reproductive cycle of female crimson sea bream Evynnis tumifrons were studied using samples collected from April 2012 to June 2014 off the southwestern coast of Kyushu, Japan. A total of 801 ovaries were examined histologically to estimate the degree of ovarian maturation. E. tumifrons showed an asynchronous ovary signifying multiple spawning in a single reproductive season. Ovarian maturity stages were classified into six categories based on the appearance of the most advanced oocytes, post-ovulatory follicles and atretic oocytes in the ovary as follows: immature, maturing, mature, spawned, spent and resting. Females with ovaries at maturing, mature, spawned or spent stages were defined as sexually mature individuals, and the size at sexual maturity was estimated to be 179-mm fork length based on 50% maturity size. Monthly changes in the gonadosomatic index and the occurrence of mature or spawned maturity stages showed that the spawning season lasts from November to May with an intermission in March 2013. The factor responsible for this intermission was considered to be the low water temperature that occurred in the preceding month.

Growth performance and biochemical composition dynamics of ovary, hepatopancreas and muscle tissues at different ovarian maturation stages of female mud crab, Scylla paramamosain

Mud crab (Scylla paramamosain) is an economically important crustacean species due to its high market price, especially females with mature ovaries. Therefore, it is vital to better understand the growth performance and biochemical composition dynamics of ovary at different maturation stages. To accomplish this, one hundred female mud crabs were obtained and their morphometric characters, ovary morphology and weight were analysed. Further, the biochemical composition dynamics of ovary, hepatopancreas and muscle tissues at various ovarian maturation stages were compared. The ovarian maturation process of S. paramamosain is divided into five developmental stages based on coloration and size. Gonadosomatic index (GSI) and six growth traits increased with ovarian maturation stages. Protein content was found constant in ovary throughout all stages, and the lipid content was similar in all three tissues. Total carotenoid content in ovary tissues increased with maturation stages whereas no significant difference was observed in the hepatopancreas and muscle tissues across stages. Highly-unsaturated fatty acid (HUFA) was the dominant group in all three tissues. Ovarian saturated fatty acid (SFA), mono-unsaturated fatty acid (MUFA) and HUFA increased with maturation stages whereas ovarian essential amino acids (EAA) increased significantly during initial vitellogenesis and oogenesis stage (stage II and III). Glutamate was the dominant amino acids in all three tissues throughout maturation stages. There was no significant difference in the individual amino acid composition, flavor amino acids (FAA) and EAA profiles of hepatopancreas and muscle tissues across maturation stages. The knowledge of growth performance and biochemical composition dynamics across ovarian maturation stages should be helpful for the establishment of suitable broodstock diet that would promote ovarian maturation and better larval quality.

Ontogeny of the specificity of gonadotropin receptors and gene expression in carp.

The pituitary gonadotropins, luteinizing hormone (LH) and follicle-stimulating hormone (FSH), are the principle endocrine drivers of reproductive processes in the gonads of jawed vertebrates. Canonically, FSH recruits and maintains selected ovarian follicles for maturation and LH induces the stages of germinal vesicle breakdown and ovulation. In mammals, LH and FSH specifically activate cognate G-protein-coupled receptors that affect the proteins involved in steroidogenesis, protein hormone synthesis, and gametogenesis. This dual-gonadotropin model also exists in some fish species, but not in all. In fact, due to their diverse number of species, extended number of ecological niches, and remarkably flexible reproductive strategies, fish are appropriate as models to understand the co-evolution of gonadotropins and their receptors. In this study, we cloned and characterized the expression profile over the final stages of ovarian maturation of carp (Cyprinus carpio) LHCGR and FSHR. Expression of both gonadotropin receptors increased in the later stage of early vitellogenesis, suggesting that both LH and FSH play a role in the development of mature follicles. We additionally tested the activation of cLHCGR and cFSHR using homologous and heterologous recombinant gonadotropins in order to gain insight into an evolutionary model of permissive gonadotropin receptor function. These data suggest that carp (Cyprinus carpio) gonad development and maturation depends on a specific gonadotropin profile that does not reflect the temporally distinct dual-gonadotropin model observed in salmonids or mammals, and that permissive gonadotropin receptor activation is a specific feature of Ostariophysi, not all teleosts.

Distinctive reproductive tactics between sympatric specimens of Astyanax aff. fasciatus in the wild and in captivity

Astyanax fasciatus is a group of neotropical fish with two different diploid chromosome numbers, and in the wild, these individuals are found in sympatry. The annual reproductive cycles of adult females of species with different numbers of chromosomes (G1: 46 chromosomes, low responsiveness to induced spawning; G2: 48 chromosomes, responsive to induced spawning) were investigated in the wild and in captivity. Plasma estradiol (E2) levels, relative fecundity (RF), oocyte diameter, gonadosomatic index, and ovarian morpho-histology were evaluated to identify the ovarian maturation stage. The G1 females in both environments began the vitellogenic phase during winter, concurrent with increased plasma E2 levels. In wild G1 females, large numbers of vitellogenic oocytes were observed during spring and summer, when plasma E2 levels remained high. In contrast, captive G1 females showed higher E2 levels during the winter. They also retained significantly higher RF and vitellogenic oocyte numbers than wild females, but showed no spawning and slow yolk reabsorption. Wild G2 females began the vitellogenic phase in autumn and had progressively increasing E2 levels that peaked in the spring, with spawning occurring during the summer. In captive G2 females, the E2 levels and RF remained high throughout the year. These data suggest that A. aff. fasciatus with different numbers of chromosomes differ in their reproductive performance in the wild and show altered sensitivity to environmental cues in captivity. Our results indicate that G1 females likely respond best to artificial spawning just after winter, whereas G2 females should respond year-round.

Effect of Eyestalk Ablation on the Ovarian Maturation Stages of Blue Swimming Crab, Portunus pelagicus

Effect of Eyestalk Ablation on the Ovarian Maturation Stages of Blue Swimming Crab, Portunus pelagicus

Hepatopancreas and ovarian transcriptome response to different dietary soybean lecithin levels in Portunus trituberculatus

Ovaries (O) are specialized tissues that play critical roles in producing oocytes and hormones. The crustacean hepatopancreas (H) is a metabolic organ that plays important functions including absorption, storage of nutrients and vitellogenesis during growth and ovarian development. However, genetic information on the biological functions of the crustacean ovaries and hepatopancreas are limited. This study compared the transcriptome in the ovary and the hepatopancreas of female P. trituberculatus fed two different diets containing 0% (SL0) and 4% soybean lecithin (SL4), respectively during the growth and ovarian maturation stages by Illumina HiSeq4000 sequencer. The differences between ovary and hepatopancreas of P. trituberculatus were also compared at transcriptional level. A total of 55,667 unigenes were obtained with mean length of 962 bps across the four treatment groups (SL0_O, SL4_O, SL0_H and SL4_H). In ovary, there were 257 differentially expressed genes (DEGs) between SL0_O and SL4_O, with 145 down- and 112 up-regulated genes in the SL4_O group. Candidate genes involved in ovarian development were detected in SL4_H group. In hepatopancreas, 146 DEGs were found between SL0_H and SL4_H, including 43 down- and 103 up-regulated genes in the SL4_H group. The specific DEGs were mainly involved with lipid related metabolism pathways, including fat digestion and absorption, PPAR signaling pathway and insulin resistance. 14,725 DEGs were found in the comparison between SL0_O and SL4_H, including 7250 up- and 7475 down-regulated genes in the SL4_H group. The specific DEGs were mainly involved with lipid (fat digestion and absorption, linoleic acid metabolism), hormone (steroid hormone biosynthesis, ovarian steroidogenesis, etc), and amino acid (phenylalanine metabolism, arginine biosynthesis, tyrosine) related metabolism pathways. Crabs fed the SL4 diet exhibited higher gene expression of cryptocyanin 1 (cc1), cryptocyanin 2 (cc2) and neuroparsin 1 (np1) in hepatopancreas and ovarian than those fed the SL0 diet, however, crab fed SL4 diet showed higher gene expression of fatty acid-binding protein 1 (fabp1), vitellogenin (vtg) and Delta-6 desaturase-like protein (fadsd6) in hepatopancreas than those fed the SL0 diet. Moreover, crabs fed the SL0 diet had lower gene expression of vtg, extracellular copper‑zinc superoxide dismutase (cuznsod) and estrogen sulfotransferase (ests) in ovary compared to those fed the diet containing 4% soybean lecithin. These results might provide important clues with respect to elucidating the molecular mechanisms underlying the regulation of phospholipid on the gonadal development and lipid metabolism of P. trituberculatus.

Reproductive dynamics of the white shrimp Litopenaeus schmitti (Burkenroad 1936) in a beach seine fishery in northeastern Brazil

ABSTRACTThe aim of the present study was to investigate the reproductive dynamics of the Litopenaeus schmitti through macroscopic and microscopic analyses. Specimens were collected monthly from December 2016 to November 2017 through artisanal beach seine fishery in the municipality of Lucena, state of Paraíba, Brazil. A total of 1710 shrimp were caught and the proportion of males was significantly lower than that of females (43.51% and 56.49%). The ovarian maturation stages were classified as immature, initial development, advanced development, mature and spent. These classification criteria added an additional stage than those previously reported for this species on the Brazilian coast. The proportion of ovarian maturation stages per month suggest a seasonal reproductive cycle that extends from September to June, with no reproductive activity occurring in July and August. The reproductive activity was directly related to the precipitation and salinity in the area of the present study. Mean cephalothorax length at first gonad maturity for females (2.20 cm) was larger than values reported 13 years ago for this species in the same area. This finding may indicate that the prohibition of motorized trawler fishing in the area since 1990 is contributing to the more sustainable exploitation of this resource.

Reproductive biology and histological studies of ovarian development ofSchizothorax plagiostomusin river Lidder from Kashmir Himalaya

The present study investigated the reproductive biology and histological study of gonads of Schizothorax plagiostomus female and its relationship with gonadosomatic index. The study period was from July, 2013 to June, 2014. Fish samples within the range of 25.5 cm to 40.33 cm in total length and weight ranging from 250 g to 580 g were used during the study. The mean GSI value of female fish fluctuated between minimum 1.059 ± 0.62 in August to maximum 14.95 ± 2.69 in May. The results showed that S. plagiostomus breeds during May, June and July confirmed by the gonadosomatic index and egg diameter measurements. The ova diameter ranged from 0.3 to 2.02 mm with minimum in July and maximum in June. The breeding status confirmed by gonadosomatic index (GSI) and ovarian histology showed six stages of ovarian maturation (viz. stage I: chromatin nucleaolar stage; II: perinucleolar stage; III: cortical alveolar stage; IV: vitellogenic yolk stage; V: ripe mature stage; and VI: atretic stage). Results show that S. plagiostomus is a seasonal breeder with unimodal spawning activity.

Gonadal steroids and energy availability during ovarian maturation stages of the Amazonian pirarucu Arapaima gigas (Teleostei: Osteoglossidae) in the wild

The Amazonian pirarucu, Arapaima gigas, belongs to a phylogenetically basal group of teleosts (family Osteoglossidae). The commercial importance and the overexploited status of natural stocks of A. gigas brought the interest to rear it in the Amazon region, but rearing techniques are not fully understood to obtain reproduction success. Gonadal steroids (plasma and cephalic liquid), lipids and proteins (muscles, liver and ovary) of wild A. gigas females were analyzed during one year on Mexiana Island (Amazon Basin, Brazil) and ovarian maturation stages were described. The sexual gender determination method based on 11-ketostestosterone (11KT) and estradiol (E2) ratio was applied to check if the value is in agreement of the sex previously determined by direct observation. Adult females were classified into four reproductive stages according to ovary development stage: previtellogenic, maturation, mature, and regression/spawning. Immature females were also analyzed. All mature females were sampled during the rainy season. Lipids stored in the ventrecha (belly) are a possible source of this substrate for the developing ovary. Plasma steroid profile throughout A. gigas female reproductive stages is similar to that found in most teleosts, but plasma 11KT levels peaked at the mature stage, suggesting an important role of this androgen in female ovulation. The 11KT/E2 method was 100% efficient when sexing juveniles and adults at the beginning of their reproductive cycle and can successfully be applied to identify the sexual gender of animals. The steroid profile of the cephalic liquid suggests a possible role of this fluid as a pheromone, due to the high level of 17α-hydroxyprogesterone at the final stages of the reproductive cycle.

Development of ovarian maturations in orange mud crab, Scylla olivacea (Herbst, 1796) through induction of eyestalk ablation and methyl farnesoate

Many crustacean species including mud crab, genus Scylla are incapable of natural maturation under captivity, thus putting high pressure on the wild catch. Therefore, to increase the availability of mature broodstocks in captivity, this study determined the effect of Eyestalk Ablation (EA) and Methyl Farnesoate (MF) administration on ovarian maturation stages of the orange mud crab, Scylla olivacea. The study was conducted using a control group (T1) consisting of 95% alcohol (widely used as a chemical solvent), and three treatment groups consisting of: 5 µl/g oral administrated MF in the diet (T2), EA (T3) and a combination of both treatments of MF and EA (T4). The highest percentage of ovarian maturation Stage 4 was found in the T4 treatment (20.8%) compared to the other treatments which were T1 (0%), T2 (8.33%) and T3 (12.5%). Ovarian development of the treated groups (T2, T3, and T4) was significantly different compared to the control group (T1) (p < 0.05). However, there was no significant difference observed in the mean Gonadosomatic Index (GSI) in orally administrated MF (T2) compared to the EA group (T3) (p > 0.05), but it was significantly different when compared to the combination group of MF and EA (T4) (p < 0.05). It can be concluded that the combination method of oral administration of MF through diet and EA (T4) is the best technique for producing mature ovaries.

Dynamics of fatty acid regulatory genes during ovarian development in Penaeus monodon.

The delay in ovarian maturation in farmed black tiger shrimp Penaeus monodon has resulted in the widespread practice of feeding broodstock with the polychaete Perinereis nuntia and their unilateral eyestalk ablation. Although this practice alters fatty acid content in shrimp ovaries and hepatopancreas, its effects on fatty acid regulatory genes are yet to be systematically examined. Here, microarray analysis was performed on hepatopancreas and ovary cDNA collected from P. monodon at different ovarian maturation stages, revealing that 72 and 58 genes in fatty acid regulatory pathways were differentially expressed in hepatopancreas and ovaries respectively. Quantitative real-time PCR analysis revealed that ovarian maturation was associated with higher expression levels of acetyl-CoA acetyltransferase, acyl-CoA dehydrogenase, acyl-CoA oxidase 3 and long-chain fatty acid transport protein 4 in hepatopancreas, whereas the expression levels of 15 fatty acid regulatory genes were increased in shrimp ovaries. To distinguish the effects of different treatments, transcriptional changes were examined in P. monodon with stage 1 ovaries before polychaete feeding, after 1 month of polychaete feeding and after eyestalk ablation. Polychaete feeding resulted in lower expression levels of enoyl-CoA hydratase and acyl-CoA synthetase medium-chain family member 4, while the expression level of phosphatidylinositide phosphatase SAC1 was higher in shrimp hepatopancreas and ovaries. Additionally, eyestalk ablation resulted in a higher expression level of long-chain fatty acid-CoA ligase 4 in both tissues. Together, our findings describe the dynamics of fatty acid regulatory pathways during crustacean ovarian development and provide potential target genes for alternatives to eyestalk ablation in the future.

Morphological, biochemical and histological analysis of mud crab ovary and hepatopancreas at different stages of development

This study describes the fatty acids, total carotenoids, and cell diameter characteristics of the female ovary and hepatopancreas of the mud crab, Scylla olivacea, with comparisons at different ovarian maturation stages. Seventy-one S. olivacea individuals at all stages of ovarian maturation were sampled from the Setiu wetlands, Terengganu, Malaysia. The ovary and hepatopancreas of each crab were used for morphological studies, histological and biochemical analyses (fatty acid composition and total carotenoids). Morphological observations indicated there was an increase in ovarian gonado-somatic index (GSI), with color changes from translucent to dark red; however, a relatively consistent hepato-somatic index (HSI) in the hepatopancreas, with the color ranging from yellow to yellowish-brown. Histological analysis indicated that oocyte diameter was positively correlated with GSI. Hepatopancreatic tubules had a relatively constant diameter from Stage 2 to 4, with increased proportions of R- and B-cells. Biochemical analysis indicated there was a significant increase in total carotenoids in the ovary during maturation. The hepatopancreas, however, had relatively consistent total carotenoid concentrations that were greater than those of the ovary. Overall, the lipid analysis results indicated there were lesser concentrations of fatty acids in the hepatopancreas, while in the ovary there were increasing concentrations during maturation. The lesser concentrations of fatty acids in the hepatopancreas than ovary suggested that energy was transferred to the ovary for future embryonic and larval development. The relationship between the hepatopancreas and the ovary in nutrient content is an important finding in providing a baseline to formulate an optimal diet for improved mud crab hatchery practices.

Diet and internal physiological changes of female orange mud crabs, Scylla olivacea (Herbst, 1796) in different ovarian maturation stages

Recently, there has been a growing interest in the ovarian maturation of mud crabs, genus Scylla. Studies regarding the factors that affect ovarian maturation in mud crabs, however, are still lacking. This study, therefore, evaluates the relationship between diet and internal physiological changes of female orange mud crabs, Scylla olivacea. Sixty female adult S. olivacea were sampled from Setiu Wetland, Malaysia. Foreguts were sampled to study fullness and content. The hepatopancreas was sampled to study digestive enzyme activity, biochemical composition, and histology. Ovaries were sampled to study ovarian biochemical composition and histology. Foregut fullness data suggest that there is an increase in feeding with advancing ovarian maturation in mud crabs. Data for foregut contents indicated that when the ovary was in Stage 3 of maturation the diet was animal-based. The activity of amylase, cellulase, lipase, and trypsin during Stages 1 and 4 of ovarian maturation were less than during Stage 3 of ovarian maturation. Biochemical composition (protein and lipid) was greater during Stage 3 of ovarian maturation. Histological analysis of the hepatopancreas indicated an increase in hepatopancreas tubules, B and R cells during Stage 3 as compared with Stage 1 of ovarian maturation. Histological analysis of the ovary indicated increases in oocyte diameter due to concentrated large yolk globules. Based on these results, it is concluded that vitellogenesis mainly occurred during Stage 3 of ovarian maturation, based on evidence of increased feeding with the diet being predominantly animal-based. Likewise, digestive enzyme activities, proteins, lipids, B cells, and R cells were all greater during Stage 3 of ovarian maturation.

Distribution and dynamic expression of serotonin and dopamine in the nervous system and ovary of Holothuria scabra during ovarian maturation.

In the present study, the distribution and dynamic expression of serotonin and dopamine in the nervous system and ovary of the sea cucumber, Holothuria scabra, during different ovarian stages were investigated. We found that serotonin-immunoreactivity was more intense in the neurons and neuropils of the outer ectoneural part, the inner hyponeural part, and the wall of hyponeural canal of radial nerve cord during the mature stages of ovarian cycle, whereas dopamine-immunoreactivity was detected at a higher intensity in these tissues during the early stages. Both neurotransmitters were detected in the ectoneural part of the nerve ring. In the ovary, serotonin intensity was more intense in the cytoplasm of late oocytes, while dopamine-immunoreactivity was more intense in the early stages. The changes in the levels serotonin in the radial nerve cord and oocytes are incremental towards the late stages of ovarian maturation. In contrast, dopamine levels in the nervous tissues and oocytes were more intense in early stages and became decremental towards the late stages. These findings suggest that serotonin and dopamine may have opposing effects on ovarian development in this sea cucumber species.

Seasonal Reproductive Biology of Drosophila suzukii (Diptera: Drosophilidae) in Temperate Climates.

Drosophila suzukii (Matsumura; Diptera: Drosophilidae) is a key pest of sweet cherry and small fruits worldwide. The present studies were designed to describe the reproductive physiology in both sexes, through dissections of their reproductive organs. We extensively dissected female D. suzukii throughout the season from 2013 to 2016 and classified the reproductive status flies based on five recognizable ovarian maturation stages: 1) no ovaries; 2) unripe ovaries 3) ripening eggs in ovarioles; 4) mature eggs in ovarioles; and 5) mature eggs in the abdomen. Development was examined as a function of calendar days as well as degree-days (DD). Results obtained from winter collections revealed that females collected from November to March contained a lower percentage of mature eggs than females collected from April to September. These data suggest that environmental conditions during the dormant period induce reproductive diapause. Oogenesis likely increased with an increase in mean monthly temperatures and DD. The first overwintered females with mature eggs were dissected as early as 21 February 2014 in Trento (7 DD). Additionally, we found that a low proportion of males (less than 50%) had sperm in their testes between January and March, yet during the same period females already have sperm stored in their spermathecal. Ivy berries was an alternative but unfavorable non-crop host during the late dormant period, as evidenced by emergence of smaller adults when compared to individuals emerging from cherry fruits. This study showed that D. suzukii females have great potential for oviposition early in the season, posing a risk to early season maturing crop hosts.

Characterization of Gnrh/Gnih elements in the olfacto-retinal system and ovary during zebrafish ovarian maturation

Gonadotropin releasing hormone (GnRH) is one of the key players of brain-pituitary-gonad axis, exerting overall control over vertebrate reproduction. In zebrafish, two variants were characterized and named as Gnrh2 and Gnrh3. In this species, Gnrh3, the hypohysiotropic form, is expressed by neurons of the olfactory-retinal system, where it is related with food detection, intra/interspecific recognition, visual acuity and retinal processing modulation. Previous studies have reported the presence of Gnrh receptors in the zebrafish retina, but not yet in the zebrafish olfactory epithelium. The current study analyzed the presence of gnrh2 and gnrh3, their receptors (gnrhr 1,2,3 and 4) and gnih (gonadotropin inhibitory hormone) transcripts, as well as the Gnrh3 protein in the olfactory epithelium (OE), olfactory bulb (OB), retina and ovary during zebrafish ovarian maturation. We found an increase of gnrh receptors transcripts in the OE at the final stages of ovarian maturation. In the OE, Gnrh3 protein was detected in the olfactory receptor neurons cilia and in the olfactory nerve fibers. Interestingly, in the OB, we found an inverse expression pattern between gnih and gnrh3. In the retina, gnrhr4 mRNA was found in the nuclei of amacrine, bipolar, and ganglion cells next to Gnrh3 positive fibers. In the ovary, gnrh3, gnrhr2 and gnrhr4 transcripts were found in perinucleolar oocytes, while gnih in oocytes at the cortical alveolus stage. Our results suggested that Gnrh/Gnih elements are involved in the neuromodulation of the sensorial system particularly at the final stages of maturation, playing also a paracrine role in the ovary.

Ovarian Maturation Stages of Wild and Captive Mud Crab, Scylla olivacea Fed with Two Diets

This study was aimed to determine the ovarian maturation stages of wild and captive orange mud crab, Scylla olivacea fed with different diets via gonadosomatic status, oocyte diameter and histological examinations. Captive crabs were fed with blood cockle, Anadara granosa, or fish, Decapterus spp. Through the histological examinations, ovarian maturation stages of wild and captive S. olivacea was classified into four stages: Immature (Stage 1), Early maturing (Stage 2), Pre-maturing (Stage 3) and Fully matured (Stage 4). Gonadosomatic Index of wild and captive crabs remained low during immature and 2 but increased significantly (p<0.05) in pre-maturing and 4 ovaries. Oocytes size were significantly different (p<0.05) in all ovarian maturation stages of wild and captive crabs. Follicle cells surround the oocyte of immature ovary and small yolk globules start to appear in early maturing ovary with large nucleus size. Oocyte size increased significantly (p<0.05) and yolk globule obviously appeared in pre-maturing ovary. Large and fused yolk globules appeared in the oocytes of fully matured ovary with nucleus was barely visible. The present study revealed that, ovarian maturation stages of S. olivacea are closely related to its morphological appearance and cellular development.

Molecular cloning, expression and functional analysis of three subunits of protein phosphatase 2A (PP2A) from black tiger shrimps (Penaeus monodon).

Protein phosphatase 2A (PP2A) is a cellular serine-threonine (Ser/Thr) phosphatase that plays a crucial role in regulating most cellular functions. In the present study, the full-length cDNAs of three subunits of PmPP2A (PmPP2A-A, PP2A-B and PP2A-C) were cloned from Penaeus monodon, which are the first available for shrimps. Sequence analysis showed that PmPP2A-A, PmPP2A-B and PmPP2A-C encoded polypeptides of 591, 443, and 324 amino acids, respectively. The mRNAs of three subunits of PmPP2A were expressed constitutively in all tissues examined, and predominantly in the ovaries. In ovarian maturation stages, the three subunits of PmPP2A were continuously but differentially expressed. Dopamine and 5-hydroxytryptamine injection experiments were conducted to study the expression profile of three subunits of PmPP2A, and the results indicated that PmPP2A played a negative regulatory role in the process of ovarian maturation. In addition, the recombinant proteins of three subunits of PmPP2A were successfully obtained, and the phosphatase activity of PmPP2A was tested in vitro. The results of this study will advance our understanding about the molecular mechanisms of PmPP2A in Penaeus monodon.