Preimplantation development in vivo and in vitro in bank voles, Clethrionomys glareolus, treated with PMSG and HCG.
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Ovulation can be induced in the bank vole by PMSG and HCG and takes place 9 1/2 to 11 1/2 hr after administration of HCG. The number of eggs obtained varied from 1 to 30 (average 9.3). The highest ovulation rates were observed when PMSG and HCG were administered 41 to 43 1/2 hr apart. The embryos attained the blastocyst stage 96 hr after HCG injection. Bank vole eggs can be cultured in chemically defined media beginning with the eight-cell stage; only 25% of four-cell eggs developed to blastocysts in vitro. (+info)
Inhibition of L-methionine uptake and incorporation by chlorpromazine in preimplantation mouse embryos.
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These studies indicate that chlorpromazine causes a rapid inhibition of the uptake and incorporation of L-[Me-3H]methionine in preimplantation mouse embryos in vitro. Concentrations of chlorpromazine from 10-5 M to 10-4 M inhibit L-[Me-3H]methionine uptake and incorporation in late four-cell embryos in 15 min. Concentrations of chlorpromazine from 2 times 10-5 M to 10-4 M inhibit uptake and incorporation in early blastocysts in 15 min to comparable degrees, suggesting that the effect of chlorpromazine on the early blastocyst is primarily on methionine transport, and not on protein synthesis. Lineweaver-Burk plots constructed from 15-min uptake values of methionine in the presence of various concentrations of chlorpromazine indicate that 5 times 10-5 M-chlorpromazine competitively inhibits methionine uptake in blastocysts, while 10-4 M-chlorpromazine non-competitively inhibits transport. Efflux experiments support the idea that chlorpromazine acts as an inhibitor of the active methionine influx processes, and not through acceleration of efflux. It is suggested that chlorpromazine may produce delayed implantation by directly affecting the preimplantation embryo, as well as through its known inhibitory effects on the hormonal functions of the maternal organism. (+info)
Ovarian tumorigenesis in mice transgenic for murine inhibin alpha subunit promoter-driven Simian Virus 40 T-antigen: ontogeny, functional characteristics, and endocrine effects.
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We previously reported formation of ovarian granulosa cell tumors with 100% penetration in a transgenic mouse model with murine inhibin alpha subunit promoter-driven (inhalpha)/Simian Virus 40 T-antigen (Tag). The tumor-bearing inhalpha/Tag mice showed highly elevated serum levels of immunoreactive inhibin. To investigate the onset of tumorigenesis and related endocrine consequences, 6-8 female mice of two inhalpha/Tag lines and their mating control littermates were killed monthly between 1 and 6 mo of age. We also investigated tumorigenesis-related fertility aspects of these two mouse lines. The ontogeny and progression of tumors could be monitored in both inhalpha/Tag lines by alterations of ovarian weights and serum hormone levels. Serum progesterone levels increased in both inhalpha/Tag lines in an age-dependent manner as ovarian tumorigenesis progressed, and a reciprocal decrease occurred in serum LH and FSH. Neither serum estradiol (E(2)) nor uterine weights were significantly altered during tumorigenesis, suggesting that the ovarian tumors represented late stages of granulosa cell differentiation. In conclusion, the present findings show in the inhalpha/Tag TG mice a relation between endocrine consequences of granulosa cell tumorigenesis, and a connection of onset of tumor formation with aberrant steroidogenesis and gonadotropin secretion. These findings indicate that tumors are endocrinologically active and able to exert enhanced negative feedback effects on pituitary function. The tumors provide a good model for endocrinologically active hormone-dependent tumors. (+info)
Intervals from norgestomet withdrawal and injection of equine chorionic gonadotropin or P.G. 600 to estrus and ovulation in ewes.
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Synchronization of estrus and ovulation is essential for AI of ewes during a predetermined time frame, and progestogen-eCG treatments are typically used to prepare the ewes. However, eCG is not readily available in the United States, but P.G. 600 (400 IU of eCG and 200 IU of hCG) is available. Thus, we conducted a study to determine the effects of eCG and P.G. 600 on the timing of estrus and ovulation after progestogen withdrawal. Ewes were assigned to two replicates of an experiment with the following treatments: 1) 3-mg norgestomet implant (i.e., one-half of a Syncro-Mate-B [SMB] implant) for 10 d, plus 2 mL of saline i.m. at SMB removal (n = 11); 2) 3-mg SMB implant for 10 d, plus 400 IU of eCG i.m. at SMB removal (n = 13); and 3) 3-mg SMB implant for 10 d, plus P.G. 600 i.m. at implant removal (n = 9). On d 6 after SMB insertion, PGF2alpha was used to induce luteolysis. Beginning 12 h after implant removal, vasectomized rams were used at 12-h intervals to check for estrus. When a ewe was detected in estrus, each ovary was evaluated ultrasonically. Ovaries were evaluated again 16 h later and then at 8-h intervals until ovulation. Treatment altered the interval from implant removal to estrus (less [P < 0.05] in SMB + eCG than in the other two groups) and to ovulation (greatest [P < 0.05] in SMB). However, the treatment x replicate interaction was significant for the intervals from implant removal to estrus (P < 0.03) and from implant removal to ovulation (P < 0.05). An inconsistent response in the SMB-treated ewes seemed to be primarily responsible for the interaction. The intervals to estrus and to ovulation for the SMB-treated ewes were shorter (P < 0.05) in Replicate 1 than in Replicate 2. Also, both intervals seemed to be less consistent between replicates for the SMB + P.G. 600- than for the SMB + eCG-treated ewes; that is, eCG seemed to increase the predictability of the intervals to estrus and to ovulation. Neither the main effects of treatment and replicate nor their interaction were significant for the interval from estrus to ovulation (38.4 /- 3.3 h), size of the ovulatory follicle (7.7 +/- 0.8 mm), or ovulation rate (1.6 +/- 0.2). We concluded from this experiment that eCG is a better choice than P.G. 600 as the gonadotropin to use at the time of progestogen withdrawal to prepare ewes for AI during a predetermined interval. (+info)
Induction and inhibition of implantation in lactating rats.
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The interrelationship between prolactin and LH in the maintenance of pregnancy during lactation was studied. The reduction of suckled young from eight to two or less, as late as on the morning of Day 4 of pregnancy, resulted in normal implantation. Reintroduction of eight young on Day 4 to lactating pregnant rats deprived of their litters on Day 1 resulted in an inhibition of implantation, but reintroduction on Days 5 or 6 did not inhibit implantation. If oestrogen, HCG or PMSG was given on Day 4 of pregnancy, implantation was induced at the normal time in rats suckling large litters. When LH antiserum was given on the morning of Day 4 or Day 8 to pregnant rats suckling two young each, it blocked implantation and postimplantation survival of blasto-cysts, respectively. When the number of suckling young was increased from two to eight on Day 6, however, LH antiserum blocked pregnancy only to the extent of 70%. Prolactin administered during the preimplantation phase inhibited implantation in pregnant rats suckling a minimum number of young, but had no effect when given during the postimplantation phase. Progesterone failed to block implantation. Prolactin had no inhibitory effect on implantation in the absence of the suckling stimulus or in non-lactating pregnant rats. The inhibition of implantation by prolactin in rats suckling two young could be effectively reversed by the administration of oestrogen, PMSG or HCG on Day 4 of pregnancy. (+info)
Administration of p.g. 600 to sows at weaning and the time of ovulation as determined by transrectal ultrasound.
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This study determined whether the interval from estrus to ovulation was altered by giving P.G. 600 to sows at weaning. Mixed-parity sows received P.G. 600 i.m. (n = 72) or no treatment (n = 65) at weaning (d 0). Beginning on d 0, sows were observed for estrus twice daily. At the onset of estrus and thereafter, ultrasound was performed twice daily to determine the average size of the largest follicles and time of ovulation. Weaning age (20.1+/-0.4 d) did not differ (P > 0.10) between treatments. More P.G. 600 sows expressed estrus within 8 d (P < 0.01) than controls (94.4% vs 78.4%, respectively). Parity was associated with expression of estrus (P < 0.02), with 78% of first-parity and 93% of later-parity sows exhibiting estrus. However, no treatment x parity effect was observed (P > 0.10). The interval from weaning to estrus was reduced (P < 0.0001) by P.G. 600 compared with controls (3.8+/-0.1 d vs 4.9+/-0.1 d). Follicle size at estrus was not affected by treatment (P > 0.10). The percentage of sows that ovulated did not differ (P > 0.10) for P.G. 600 and control sows (90.3% vs 81.5%, respectively). Time of ovulation after estrus was not affected by treatment and averaged 44.8 h. However, univariate analysis indicated that the interval from weaning to estrus influenced the interval from estrus to ovulation (r = 0.43, P < 0.0001). Further, multivariate analysis showed an effect of treatment on the intervals from weaning to estrus, weaning to ovulation (P < 0.0001), and estrus to ovulation (P < 0.04). Within 4 d after weaning, 81% of the P.G. 600 sows had expressed estrus compared with 33% of controls. However, this trend reversed for ovulation, with only 35% of P.G. 600 sows ovulating by 36 h after estrus compared with 40% of controls. The estrus-to-ovulation interval was also longer for control and P.G. 600 sows expressing estrus < or = 3 d of weaning (45 h and 58 h, respectively) than for sows expressing estrus after 5 d (39 h and 32 h, respectively). Farrowing rate and litter size were not influenced by treatment. However, the interval from last insemination to ovulation (P < 0.02) indicated that more sows farrowed (80%) when the last insemination occurred at < or = 23 to > or = 0 h before ovulation compared with insemination > or = 24 h before ovulation (55%). In summary, P.G. 600 enhanced the expression of estrus and ovulation in weaned sows but, breeding protocols may need to be optimized for time of ovulation based on the interval from weaning to estrus. (+info)
Fertilization of rat eggs in vitro at various times before and after ovulation with special reference to fertilization of ovarian oocytes matured in culture.
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Oocytes recovered at various times from immature rats treated with PMSG and HCG were incubated with capacitated epididymal spermatozoa of mature rats. In the presence of follicular cells, sperm penetration was not observed 4 hr after incubation in the oocytes at stages from the intact germinal vesicle to the chromatin mass, but 7 to 55% of oocytes were penetrated at stages from the condensed germainal vesicle to metaphase II. After the removal of follicular cells, 15 to 72% of the oocytes at any stage were penetrated. After further incubation for 15 hr, the proportion of penetrated oocytes increased from 8 to 98% from early to late stages and that of penetrated oocytes with a male and female pronucleus increased from 9 to 100% as maturation progressed. Although the average number of spermatozoa/oocyte was not correlated with its maturation, transformation of the sperm head into a male pronucleus was retarded or failed, especially in the younger oocytes. Following incubation in a defined medium for 13 hr, 85% of oocytes at the intact germinal vesicle stage matured to the stage of the first polar body formation, but only 18 to 22% of these mature oocytes were penetrated by spermatozoa and only a few of the penetrated oocytes cleaved into normal two-cell eggs. When eggs recovered from oviducts 14 to 20 hr after ovulation were exposed to capacitated spermatozoa, the proportion of penetrated eggs (86 to 98%) and that of polyspermic eggs (11 to 27%) were not related to the ages of the eggs, but failure of transformation of the sperm head and the proportion of abnormal eggs increased 14 to 20 hr after ovulation. (+info)
Kallikrein gene expression in the gonadotrophin-stimulated rat ovary.
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The kallikreins (KLKs) are a highly conserved multi-gene family of serine proteases that are expressed in a wide variety of tissues and act on a diverse range of substrates. KLK-like enzyme activity has variously been reported to increase or decrease during the period leading up to ovulation in the equine chorionic gonadotrophin (eCG)primed, human chorionic gonadotrophin (hCG)-stimulated immature rat ovary. These earlier studies, which used biochemical assays to detect enzyme activity, lacked the specificity and sensitivity necessary to characterise conclusively the activity of the individual KLK gene family members. In this study, we have used a gene-specific RT-PCR/Southern hybridisation strategy to delineate the expression patterns of six of the individual KLK genes expressed in the rat ovary (rKLK1-3 and rKLK7-9). We have identified three broad patterns of expression in the eCG/hCG-stimulated ovary in which there is either a post-eCG increase/pre-ovulatory decrease in rKLK expression (rKLK1, rKLK3), a peri-ovulatory decrease in expression (rKLK2, rKLK8) or a relatively unchanged pattern of expression (rKLK7, rKLK9). In addition to clarifying the earlier biochemical studies, these findings support a differential role for the individual KLKs in the ovulatory process. (+info)