Termination Of Embryonic Diapause Research Articles

Signaling Transduction Pathways and G-Protein-Coupled Receptors in Different Stages of the Embryonic Diapause Termination Process in Artemia.

Artemia is a widely distributed small aquatic crustacean, renowned for its ability to enter a state of embryonic diapause. The embryonic diapause termination (EDT) is closely linked to environmental cues, but the precise underlying mechanisms remain elusive. In this study, ATAC-seq and RNA-seq sequencing techniques were employed to explore the gene expression profiles in Artemia cysts 30 min after EDT. These profiles were compared with those during diapause and 5 h after EDT. The regulatory mechanisms governing the EDT process were analyzed through Gene Ontology (GO) enrichment analysis of differentially expressed genes. Furthermore, the active G-protein-coupled receptors (GPCRs) were identified through structural analysis. The results unveiled that the signaling transduction during EDT primarily hinges on GPCRs and the cell surface receptor signaling pathway, but distinct genes are involved across different stages. Hormone-mediated signaling pathways and the tachykinin receptor signaling pathway exhibited heightened activity in the '0-30 min' group, whereas the Wnt signaling pathway manifested its function solely in the '30 min-5 h' group. These results imply a complete divergence in the mechanisms of signal regulation during these two stages. Moreover, through structural analysis, five GPCRs operating at different stages of EDT were identified. These findings provide valuable insights into the signal regulation mechanisms governing Artemia diapause.

Extrinsic and intrinsic factors drive the timing of gestation and reproductive success of Scandinavian brown bears

IntroductionClimate change is altering the reproductive phenology of many organisms, but the factors that influence the timing of gestation in ursids are still poorly understood. Higher temperatures in spring are already causing an earlier den exit in some brown bear populations, and a temporal mismatch between hibernation and reproduction could have dramatic consequences for reproductive success. Therefore, understanding the factors that control the timing of these events is important to forecast the consequences of climate change on population growth rates.MethodsIn this study, we used abdominal temperature loggers and GPS collars with acceleration sensors on 23 free-ranging pregnant female brown bears living in two areas in Sweden (61°N and 67°N latitude) to pinpoint hibernation and reproductive events. We investigated how intrinsic and extrinsic factors influence the termination of embryonic diapause and parturition, as well as their impact on reproductive success.ResultsThe termination of embryonic diapause was later in the northern area compared to the southern area and occurred earlier when ambient temperature at den entry was higher in both areas. In the southern area, young adults (i.e., females = 7 years old) had a delayed parturition when bilberry abundance was low the year of mating. Additionally, young adults had a lower reproductive success than adults and their probability to reproduce successfully was dependent on bilberry abundance, whereas adult females were not affected by this parameter.DiscussionAs den exit occurs later in the northern study area, we suggest that a later parturition might ensure that females lactate their cubs in the den for a reasonable amount of time while fasting. Similarly, a later parturition combined with an earlier emergence could allow young adults to spend less time in the den lactating if they could not reach an optimal body condition prior to hibernation. But as a result, their cubs are younger and more vulnerable when they leave the den leading to lower survival rates. Our results suggest that a decreased berry abundance in the fall could impact the reproductive and hibernation phenology of Scandinavian brown bear females and lead to a lower cub survival with potential consequences on the population dynamics.

Interactive effect of photoperiod and temperature on the induction and termination of embryonic diapause in the migratory locust.

Diapause plays a vital role in the survival and population reproduction of the migratory locust in temperate regions. Although some studies have suggested that photoperiod and temperature are two important environmental factors involved in the induction and termination of embryonic diapause in locusts, the independent or synergetic roles of these two factors are still unclear. We designed crossover experiments to investigate the effects of photoperiod, temperature and treatment time on the induction and termination of embryonic diapause in locusts by optical coherence tomography (OCT), which can track the entire embryonic developmental process via noninvasive 3D real-time imaging. Diapause induction experiments showed that a short photoperiod (in adults) and a low egg treatment temperature are the most important prerequisites for inducing egg diapause. The diapause rate reached 90.7% when the adults were reared under a short photoperiod (8 L:16 D) and the eggs were treated at 22 °C. However, the adult rearing temperature had no significant effect on diapause induction. Analysis of variance also confirmed that there was a significant interactive effect between parental photoperiod and egg treatment temperature. Moreover, diapause termination experiments showed that 13 °C (near the developmental threshold temperature of 15 °C) for 20 days is the most effective condition for terminating diapause. Our study provides accurate photoperiod and temperature thresholds for inducing and terminating diapause in the migratory locust, suggesting potential applications in the prediction and control of locust plagues. © 2021 Society of Chemical Industry.

Cloning, expression pattern, and potential role of apoptosis inhibitor 5 in the termination of embryonic diapause and early embryo development of Artemia sinica

During the embryonic development of Artemia sinica, the diapause phenomenon can be induced by high salinity or low temperature conditions. The diapause embryo at the gastrula stage is maintained under the threat of apoptosis to guarantee the embryo's normal development. In this process, apoptosis inhibitor proteins play vital roles in protecting embryos against apoptosis. Apoptosis inhibitor5 (API5) plays a pivotal role in regulating the cell cycle and preventing programmed cell death after growth factor starvation. In the present study, we cloned the full-length cDNA representing the api5 gene from A. sinica (As-api5), which encodes a 372-amino acid protein. In situ hybridization experiments revealed that As-api5 expression is not tissue or organ specific. Quantitative real-time PCR analyses of the developmental expression of As-api5 showed that it reached its highest level at 10h, after which its expression decreased. High salinity and low temperature treatments increased the expression of As-api5. Western blotting was used to assess the abundance of As-API5 and related proteins (As-CyclinA, As-CyclinE, As-E2F1, As-CDK2, As-APAF1, and As-Caspase9). Downregulation of As-api5 expression using a short interfering RNA resulted in increased mortality and embryo malformation of A. sinica. Taken together, the results indicated that API5 plays a crucial role in embryonic diapause termination and early embryo development of A. sinica.

Endocrine control of embryonic diapause in the Australian sharpnose shark Rhizoprionodon taylori.

The reproductive cycle of the Australian sharpnose shark, Rhizoprionodon taylori, includes a temporary suspension of development at the commencement of embryogenesis termed embryonic diapause. This study investigated levels of 17β-estradiol (E2), testosterone (T) and progesterone (P4) in plasma samples of mature wild female R. taylori captured throughout the reproductive cycle and correlated them with internal morphological changes. Levels of T were elevated through most of the embryonic diapause period, suggesting a role of this hormone in the maintenance of this condition. Increasing plasma T concentrations from late diapause to early active development were associated with a possible role of androgens in the termination of embryonic diapause. As in other elasmobranchs, a concomitant increase of E2 with ovarian follicle size indicated a direct role of this hormone in regulating vitellogenesis, while a peak in P4 suggested this hormone is associated with preovulation and ovulation. Additionally, significant correlations between photoperiod or water temperature and maximum follicular diameter and hepatosomatic index suggest that these abiotic factors may also play a role triggering and regulating the synchrony and timing of reproductive events.

Effect of Photoperiod on Gonadal Steroid Hormone Levels and Reproductive Cycles of Northern Fur Seals (Callorhinus ursinus)

The northern fur seal (Callorhinus ursinus) is a migratory marine mammal belonging to the family Otariidae that breeds on the islands in the North Pacific Ocean, Bering Sea, Sea of Okhotsk, and near the coast of California during summer (Peterson 1968; Gentry 1998). Their reproduction is characterized by extreme polygyny and highly synchronized annual cycles that are concentrated around the boreal summer (Peterson 1968; Trites 1992; Gentry 1998). Females arrive at breeding sites in early July, give birth within 36 hours, and then enter estrus and copulate within a few days (Peterson 1968). Although active gestation persists for approximately eight months, the total duration of gestation period is approximately one year because of the embryonic diapause (delayed implantation), which accurately synchronizes the annual reproductive cycle (Craig 1964; Boyd 1991). Males return to the breeding sites approximately one month before females, establish their territories in the breeding area, and maintain breeding activities for approximately two months (Peterson 1968). Photoperiod is commonly cited as the main trigger for the synchronization of the annual reproductive cycles of pinnipeds (Boyd 1991; Atkinson 1997; Boyd et al. 1999), and this theory is supported by several studies. Temte (1985) compared the mean dates of parturition between the two wild populations of northern fur seals in Alaska (57°N) and California (33°N) and suggested that a mean photoperiod of 12.5 h/day in autumn acts as a trigger for the termination of embryonic diapause and initiation of implantation to synchronize parturition (also see Spotte and Adams 1981). Kiyota et al. (2009) also observed a significant latitudinal variation in the parturition of captive northern fur seals in Japan, which supported the theory of photoperiodic control of the reproductive cycle. The effect of photoperiod on reproductive cycles has been only experimentally demonstrated in harbour seals (Phoca vitulina) (Bigg and Fisher 1975). Bigg and Fisher (1975) demonstrated that estrus occurred earlier than usual during extended photoperiods and was delayed during shortened photoperiods. However, Daniel (1981) did not detect any effect of a shortened photoperiod on the rate of blastocyst development in northern fur seals. Boyd (1991) interpreted the finding of Daniel’s study to be the result of the relatively insignificant shortened photoperiod and/or the short study period. Seasonal changes that occurred in the testicular activity of male fur seals in captivity corresponded with those that occurred in males in the wild (Kohyama et al. 1999; Tsubota et al. 2001). These studies indicate that photoperiodic control of the male reproductive cycle is similar to that of female reproductive cycle, but this photoperiodic effect has not been extensively studied (Boyd 1991). Photoperiod has been reported to control the timing of implantation and parturition in pinnipeds (Temte 1985). However, this interpretation has not been applied to the timing of estrous and ovulation of non-pregnant females or the testicular activities of males. Other mammals, such as Siberian hamsters (Phodopus sungorus) and gray mouse lemurs (Microcebus murinus), are defined as long-day breeders because their reproductive activities increase in response to day length as the seasons change from spring to summer (Goldman 1991, 1999; Perret and Aujard 2001). Therefore, we aimed to determine whether the reproductive cycles of both female and male northern fur seals would be altered under experimentally induced conditions of extended photoperiods. Seasonal changes in gonadal steroid hormone levels and body weight were used as indicators of the reproductive cycle indices, because changes in these factors in relation to the reproductive cycle have previously been described (Kiyota et al. 1999; Kohyama et al. 1999). The effects of photoperiod on seasonal changes in reproductive cycles

Reversible change in embryonic diapause intensity by mild temperature in the Chinese rice grasshopper, Oxya chinensis

AbstractThe effects of temperature on maintenance and termination of embryonic diapause were investigated in Jining (35.4°N, 116.6°E) and Sihong (33.5°N, 118.2°E) strains of the Chinese rice grasshopper, Oxya chinensis Thunberg (Orthoptera: Catantopidae). Eggs of both strains entered diapause when incubated at 30, 25, or 20 °C. Chilling at 8 °C had an evident effect on diapause termination and almost all eggs chilled for 60 days ended diapause development. Chilling of eggs at 8 °C for only 20 days failed to result in any hatching at 20 °C, suggesting that such level of chilling was not enough to induce diapause termination. However, the treatment combining incubation of eggs at 30 °C for varying lengths of time with subsequent incubation to 20 °C had a distinct effect on the completion of diapause of the eggs. The results indicate that there were two temperature optima, that is, low temperature (chilling) and high temperature, for diapause development in this grasshopper species. Incubation of chilled eggs at 20 °C for 5–15 days followed by further incubation at 25 °C reduced termination of diapause significantly compared with the eggs only chilled at 8 °C. Exposure of eggs chilled at 8 °C to a pulse of 25 °C from 1 to 7 days, separated by a 20‐day interval at 8 °C, resulted in a decrease in the percentage of successfully hatched eggs as the length of the pulse of 25 °C increased. The results suggest that diapause intensity may be restored at moderately high temperatures. This reversible change in diapause intensity would play an important role in maintaining diapause before winter.

Influence of darkness on embryonic diapause termination in dormant Artemia cysts with no experience of desiccation

One of the environmental cues for Artemia embryonic diapause termination (EDT), the nature of which is barely understood, is the absence of light, although light-dark cycles (photoperiod) have been examined in inducing embryonic diapause. The influence of constant darkness (DD) on EDT was investigated in laboratory-produced cysts of Artemia franciscana. In a previous report (Nambu et al., 2008. J Exp Zool 309A:17-24 ), we described a facilitated effect of pretreatment of DD (5-14 days) on EDT in Artemia cysts exposed to a standard rearing condition. In this article, the influence of pretreatment of DD (1-150 days) was investigated in detail by bioassay of hatching. It was found that all the DD conditions significantly promoted EDT. Either the plateau value of the percent hatch increased in response to each DD condition, or the rearing period to attain the half value of the plateau value became shorter. It was also found that DD (1-100 days) led to cyclic changes of EDT: the logistic functions of hatching were cyclically regulated and 30 days-DD cycles of relative values of their parameters were observed synchronously. The effect of DD-duration was not cumulative, and a couple of optimum conditions for higher EDT levels were obtained. The probability of Artemia's encountering DD and the possible meaning of cyclic regulation of EDT by DD in Artemia are discussed.

Influence of temperature and darkness on embryonic diapause termination in dormant Artemia cysts that have never been desiccated

Environmental cues for embryonic diapause termination (EDT) were investigated in the laboratory-produced encysted dormant embryos of the brine shrimp, Artemia franciscana. The cysts were spawned and kept throughout in a 2% sea salt solution. They were activated by various temperatures of the temperate zones or by continuous dark condition (DD), resulting in a state of EDT, and were thereafter able to resume their subsequent development and hatch under appropriate conditions. The level of EDT was conveniently assayed by a hatch test observed within 2 days at 28 degrees C under continuous light condition (LL). A cold treatment of the newly spawned dormant cysts, at 4 degrees C under DD for more than 100 days, resulted in more than 95% hatch of the dormant cysts. Similar treatments of the dormant cysts but at room temperature or 28 degrees C led to significantly different results (30-40% hatch). Almost all the residual non-hatched cysts derived from the above could hatch after an additional cold treatment (at 4 degrees C under DD for about 100 days). This might prove to be latent partial bivoltine in Artemia. Meanwhile, a rearing condition (28 degrees C under LL) induced the newly spawned cysts to hatch scatteredly at and after 1 month, resulting in 22% cumulative hatch on the 92nd day after spawning. When the newly spawned dormant cysts were pretreated at 28 degrees C under DD for 5 or 14 days and then reared at 28 degrees C under LL, the cumulative hatch significantly increased (60%). These results are discussed with respect to probable diapause regulator(s) involved in EDT.

20.P1. Effects of temperature and light on embryonic diapause termination in Artemia dormant cysts that have never been desiccated

20.P1. Effects of temperature and light on embryonic diapause termination in Artemia dormant cysts that have never been desiccated

20.5. A comprehensive study of Artemia genes activated in a cold treatment to induce embryonic diapause termination

20.5. A comprehensive study of Artemia genes activated in a cold treatment to induce embryonic diapause termination

Temperature-dependent activation of ERK/MAPK in yolk cells and its role in embryonic diapause termination in the silkworm Bombyx mori

The silkworm Bombyx mori requires 2–3 months of low temperature (5 °C) to terminate embryonic diapause. The molecular mechanisms, however, are unknown. Extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) is temperature-dependently activated in the yolk cells of diapausing eggs after 45 days at 5 °C, coincident with the acquisition of developmental competence of the embryos at 25 °C. Yolk cell granulation and dissociation also begin in diapause eggs incubated at 5 °C for 45 days. We used dechorionated egg culture as a model system of diapause termination and observed that both yolk cell dissociation and embryonic development are inhibited by MAPK–ERK kinase (MEK) inhibitor U0126. Therefore, we suggest that ERK in yolk cells has a role in regulating changes in yolk morphology and termination of embryonic diapause in B. mori.

Changes in uterine expression of leukemia inhibitory factor receptor gene during pregnancy and its up-regulation by prolactin in the western spotted skunk.

The multifunctional cytokine leukemia inhibitory factor (LIF) is presumed to participate in preparing the uterus for blastocyst implantation. Increased production of LIF is positively correlated with termination of embryonic diapause and preparation for implantation in the spotted skunk. This study examined changes in the expression, localization, and hormonal regulation of LIF receptor (LIFRbeta) gene expression in the uterus of the skunk. Changes in the uterine concentration of LIFRbeta mRNA during pregnancy or in response to hormones after ovariectomy were determined by Northern hybridization analysis and reverse-transcriptase polymerase chain reaction. The skunk uterus produces two LIFRbeta transcripts, the levels of which increase in concentration when the blastocysts resume their development but then decline somewhat during the latter stage of blastocyst activation. Ovariectomy significantly reduced uterine LIFRbeta expression. Estradiol and/or progesterone failed to significantly elevate LIFRbeta mRNA levels in ovariectomized animals. Prolactin significantly increased uterine concentrations of LIFRbeta mRNA to greater than those of ovariectomized controls, but these levels were not comparable to those observed during preimplantation. The LIFRbeta mRNA was predominately localized to stromal cells surrounding the uterine glands and in yolk sac endoderm, syncytiotrophoblast, and cytotrophoblast of postimplantation embryos.

Luteotropic hormone receptors in the ovary of the mink (Mustela vison) during delayed implantation and early-postimplantation gestation.

The reproductive cycle of the mink displays rigid seasonality and obligate embryonic diapause. After ovulation, the corpus luteum (CL) involutes, and it secretes basal progesterone until activated prior to implantation. To study changes in the relative abundance of luteal prolactin and LH receptor mRNA through gestation, ovaries and serum were collected from pregnant female mink at 2-day intervals (n = 3 per date) through embryonic diapause and CL activation (March 19-31) and at 5-day intervals through implantation and early-postimplantation gestation (March 31-April 15). To determine the effect of endogenous prolactin, mink received Alzet osmotic minipumps releasing 2 mg/day 2-bromo-alpha-ergocryptine (bromocriptine) or saline on March 19. Ovaries and serum were taken from 3 animals every 2 days until March 31. Prolactin receptor mRNA in ovaries was low during CL activation but increased 3-fold through embryo implantation. Its abundance correlated with prolactin binding to ovarian membranes and with circulating prolactin. Bromocriptine suppressed endogenous prolactin levels and prevented the increase in prolactin receptor mRNA. There was a transient peak in LH receptor mRNA in the ovaries at March 19-23, which declined to basal levels by March 25 and remained constant through midgestation. Bromocriptine prevented the preimplantation peak in LH receptor mRNA and reduced its abundance below pretreatment levels. The results suggest that prolactin up-regulates its receptor and maintains the LH receptor in the mink CL. The pattern of LH receptor mRNA argues for a role for LH in CL reactivation and termination of embryonic diapause in mink.

Cloning of leukemia inhibitory factor (LIF) and its expression in the uterus during embryonic diapause and implantation in the mink (Mustela vison).

Leukemia inhibitory factor (LIF) is essential for embryo implantation in mice. Whether LIF plays a role in termination of embryonic diapause and initiation of implantation in carnivores, especially in species with obligate delayed implantation such as the mink, is not known. The objectives of this study were to clone the LIF coding sequence in the mink and determine its mRNA abundance in the uterus through embryonic diapause, implantation, and early postimplantation. We show that the mink LIF cDNA contains 609 nt encoding a deduced protein of 203 amino acids. The homologies are 80.6, 90, 88.2, 87.6, and 86.8% in coding sequence and 79.2, 90.1, 91, 90.1 and 85.4% in amino acid sequence with mouse, human, pig, cow, and sheep respectively. Glycosylation sites and disulfide bonds present in other species are generally conserved in the mink LIF sequence. Quantitation by polymerase chain reaction amplification indicates that LIF mRNA is expressed in mink uterus just prior to implantation and during the first two days after implantation, but not during diapause or later after implantation pregnancy. The abundance of LIF mRNA was significantly higher in the uterus at the embryo expansion stage (P < 0.05) than at days 1-2 of postimplantation. By immunohistochemical localization it was shown that LIF is expressed in the uterine epithelial glands at time of embryonic expansion and in early postimplantation. The coincidence of LIF expression with implantation in this species suggests that LIF is involved in the implantation process, and may be a maternal signal which terminates obligate embryonic diapause.

Effects of Physical and Chemical Stimuli on Termination of Embryonic Diapause in the Spider Mite, Panonychus bambusicola Ehara et Gotoh(Acari: Tetranychidae).

Gotoh, T., T. Kamoto and H. Oku, 1994. Effects of physical and chemical stimuli on termination of embryonic diapause in the spider mite, Panonychus bambusicola Ehara et Gotoh(Acari: Tetranychidae). J. Acarol. Soc. Jpn., 3(2): 53-58. Physical and chemical stimuli were effective in breaking diapause in eggs of the spider mite, Panonychus bambusicola Ehara et Gotoh. A 60-day or longer exposure to 5°C promoted hatching in some diapause eggs. However, more eggs terminated diapause when stimulated with a needle or exposed to ammonia-gas after chilling, although such artificial stimuli were not effective in terminating diapause if the eggs had not been chilled. Eggs stimulated with a needle or ammoniagas after chilling hatched more rapidly at 25°C than did those experienced chilling alone. Because such physical and chemical treatments were effective in terminating diapause only when they were applied to pre-chilled eggs, these stimuli seemed to influence the final phase of diapause development.

A time-interval activation of esterase A4 by cold: Relation to the termination of embryonic diapause in the silkworm Bombyx mori

The inactive esterase A4 (Ease A4) purified from the diapausing eggs of the silkworm, Bombyx mori, was chilled in vitro. The enzyme activity was very low during the early chilling period and it was suddenly elevated at a certain time of the chilling (2 weeks or less after chilling), depending upon when the chilling period began, and was followed by a rapid fall. The sudden elevation of the Ease A4 activity in vitro is equivalent to that observed in vivo and is coincident with the chilling period, the latter being indispensable for diapause termination. Data are also presented that suggest that the cold-induced activation of the Ease A4 may result from an autonomous structural change of the enzyme molecule which proceeds gradually in the cold.

Control of luteal function and implantation in the mink by prolactin.

Mink (Mustela vison) were treated during the period of embryonic diapause with prolactin or ergocryptine (CB-154). Prolactin advanced implantation time and hastened onset of luteal phase progesterone secretion. Duration of gestation in prolactin-treated adult mink was shorter than that of control mink. Ergocryptine had the opposite effects, prolonging gestation and inhibiting onset of luteal phase progesterone secretion. Prolactin is suggested to be the luteotrophin necessary for termination of embryonic diapause in mink.

Photoperiod and the termination of embryonic diapause in the marsupial macropus eugenii.

The effects of the photoperiod on embryonic diapause in tammar wallabies (Macropus eugenii) were investigated. The animals were subjected to the following photoperiods: A) normal daylight photoperiod B) 15 hours light:9 hours dark and C) 12 hours light:12 hours dark at the vernal equinox. On January 19th the photoperiods for Groups B and C were reversed. Reducing the photoperiod by 3 hours caused a termination of embryonic diapause though this was independent of the previous light regimen. The results suggest that the reactivation of embryonic development may be related to the time of the full moon in December relative to the summer solstice.

Esterases in relation to embryonic diapause in the field cricket, Teleogryllus commodus

Fourteen esterase isozymes were detected by polyacrylamide gel electrophoresis in homogenates of eggs of the field cricket Teleogryllus commodus, at various stages of development. One esterase isozyme appeared during embryonic diapause termination effected by prolonged chilling of the eggs. Esterase activity towards methyl butyrate in T. commodus egg homogenates increased with time. Sonication generated the maximum levels of esterase activity observed. Methyl butyrate esterase activity in T. commodus eggs was high during the first few days of embryogenesis; fell markedly during the period of rapid water uptake, and then remained fairly constant during diapause, diapause development, and during resumed embryogenesis.