Administration Of Pregnant Mare Serum Gonadotropin Research Articles

Effect of Pregnant Mare Serum Gonadotropin and Antidopamine on the gonads of female bileh fish (Rasbora maninjau)

Bileh fish (Rasbora maninjau) still have difficulty spawning in a controlled manner in aquaculture containers because the maturation of female fish gonads is slow and takes a long time. Internal factors, including the hormonal system, strongly influence the maturation of fish gonads. This study aims to determine the effect of Pregnant Mare Serum Gonadotropin (PMSG) and antidopamine on female gonads of bileh fish. The research used an experiment with a CRD (Completely Randomized Design) experimental design. The experiment consisted of four treatments and three repetitions. The treatments were the administration of PMSG and antidopamine hormones at different doses through feed, namely: 0 ml/kg feed (P1), 0.5 ml/kg feed (P2), 1 ml/kg feed (P3) and 2 ml/kg feed (P4). The research procedures included container and fish preparation, hormone, and feed preparation, fish rearing, and data collection. The test parameters evaluated were weight gain (Wg), gonadosomatic index (GSI), and gonadal maturity level (GML). Data on test parameters were analyzed with statistics (Anova). The research results showed that the administration of PMSG hormones and antidopamine through feed had a significant effect on weight gain, gonadosomatic index and gonad maturity level of bileh fish (P<0.05). Effective and best treatment at P3 (1 ml/kg feed) with Wg 0.22grams, GSI 8.58%, and GML IV 60%.

Quantitative Histomorphometry of Pre-ovulatory Follicles and Uterine Glands of Ongole-grade Heifer in Response to the Low Doses of PMSG Administration

Injection of female mammalian animals with gonadotropin prior to mating using superovulation dose increases estradiol 17beta (E2) synthesis and secretion and improves the growth and development of the uterus to support prenatal growth with the side effect of increasing litter size. This experiment was designed to study the effects of low doses of PMSG injection in heifers on the growth of granulose cells (GC) of pre-ovulatory follicles, E2 synthesis, and uterine glands growth and development. Nine Ongole-grade heifers were divided into three groups of doses of PMSG injection, i.e., 0 IU/kg BW, 0.5 IU/kg BW, and 1.0 IU/kg BW. The injection of pregnant mare serum gonadotropin (PMSG) was conducted at the early stage of the second follicle wave. Blood samples were collected at estrus (day 0) to measure plasma E2 concentrations. The ovary and uterus were collected for histological observation after slaughtering the animal. The results showed that non-superovulation doses of PMSG injections at the second follicle wave significantly increased the growth of GC that eventually produced a significantly higher plasma E2 concentration by 39.47% and 113.57%, in heifers receiving 0.5 IU/kg BW and 1.0 IU/kg BW (p<0.05), respectively. The increase in E2 synthesis and secretion significantly stimulated the growth and development of uterine glands (p<0.05), which was reflected in the improved measures of uterine glands. In conclusion, injections of heifers with low doses of PMSG at the second follicle wave could increase the growth of GC to produce a higher secretion of E2 that eventually improves the growth and development of uterine glands in the endometrium without the risk of multiple calving.

Gonadotropin regulates NUCB2/nesfatin-1 expression in the mouse ovary and uterus

NUCB2/nesfatin-1 is expressed in the hypothalamus and regulates food intake and energy metabolism. Recent studies showed that NUCB2/nesfatin-1 also plays a role in other organs. However, its expression pattern and function in female reproductive organs are unclear. Therefore, we investigated NUCB2/nesfatin-1 expression in the ovary and uterus of mice and determined whether it is regulated by gonadotropins and sex steroid hormones. NUCB2 mRNA and nesfatin-1 protein were detected in the ovary and uterus of mice. NUCB2/nesfatin-1 expression in both organs was highest in the estrus period of the estrus cycle. Administration of pregnant mare serum gonadotropin (PMSG) dose-dependently increased mRNA expression of NUCB2 in the ovary and uterus of mice. On the other hand, mRNA expression of NUCB2 in the uterus was dramatically decreased after ovariectomy and was not increased upon administration of PMSG. Injection of 17β-estradiol upregulated mRNA expression of NUCB2 in the uterus of ovariectomized mice, whereas injection of progesterone did not. These results suggest that NUCB2/nesfatin-1 expression in the ovary and uterus of mice is regulated through the hypothalamus-pituitary-ovary axis and that NUCB2/nesfatin-1 is a local regulator of ovarian steroidogenesis and uterine function.

Impacts of GnRH, PMSG and HCG Treatments on Follicular Diameter, Conception and Lambing Rates of Egyptian Ewe Lambs Using Intravaginal Sponges

ABSTRACTThe influence of gonadotropin releasing hormone (GnRH), Pregnant Mare SerumGonadotropin (PMSG) and human Chronic Gonadotropin (hCG) on some reproductive responses ofEgyptian ewe lambs was evaluated. Twenty five animals (aged 8 – 12 months and weighed 27 - 33Kg) were divided into 5 groups ( 5 ewe lambs each). Group (1) served as a control, Group (2),received vaginal sponges impregnated with MAP (60 mg medroxy progesterone acetate) for 14 days,during this period, animals fed 3 kg Berseem, (Trifolium alexandrinum) plus 500 gm concentrate /head/ day. Group (3), received vaginal sponges impregnated with (60 mg, MAP) for 14 days, on theday of sponge removal; each animal injected by PMSG (250 IU, I/M). Group (4) animals received1.25 ml Receptal (0.5 μg, GnRH) I/M, on day 0, seven days later, ewe lambs injected with 0.5 mlEstromate (125 μg. PGF2α) I/M, after 48 hours, animals treated with a second dose of GnRH. Group(5) received two injections of (0.5 ml Estromate) at days 0 and 7, respectively, after 72 hours from thesecond dose of PGF2α ewe lambs were injected 0.2 ml hCG. Heat detection was performed and transrectalultrasound scanning also performed to confirm estrus, pregnancy and follicular diameter. Theresults revealed that, administration of PMSG (group 3) reduced the interval to estrus (the onset ofheat /hours) significantly (P< 0.01) compared with control and other groups. Regarding conceptionand lambing rates, groups 2, 3, & 4 had affected significantly (P˂ 0.01) compared to groups 1 & 5.Follicular diameter (mm) tended to increase significantly in treated groups than the control one.Results suggest that, tested hormones have effect on selected reproductive responses of Egyptian ewelambs while administration of PMSG (Group 3) has the best response compared to other hormonaltreatments.

Co-administration of insulin with a gonadotropin partly improves ovulatory responses of estrogen-deficient mice

Administration of 17-βestradiol (E2) with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) can induce ovulation in estrogen-deficient (ArKO) mice; nevertheless, ovulatory efficiency and rate are low. In this study, effects of insulin on the ovulatory responses were investigated. In ArKO ovary, hCG signal was found to be transmitted in an uncoordinated manner when phosphorylation levels of signaling molecules are examined. Co-administration of insulin with hCG improved the transmission of hCG signal as well as the ovulatory efficiency in ArKO mice. It also improved the ovulatory rate but far below the wild-type rate. Gene expression analysis demonstrated that Cyp11a1 and Cyp17a1 mRNAs were significantly induced 4 h after PMSG administration in the wild-type ovary, but not in ArKO ovary. Collectively, these results suggest that insulin improves ovulatory responses of ArKO mice, but it fails to ameliorate follicular dysfunctions caused possibly by an inappropriate intraovarian milieu during follicular maturation.

Cloprostenol and pregnant mare serum gonadotropin promote estrus synchronization, uterine development, and follicle-stimulating hormone receptor expression in mice.

This study investigated the effects of pregnant mare se-rum gonadotropin (PMSG) and cloprostenol (CLO) on estrus induc-tion and synchronization, uterine development, and follicle-stimulating hormone receptor (FSHR) expression in mice. A total of 105 Kunming pre-puberty mice were divided into seven subgroups. Three PMSG sub-groups were injected intraperitoneally with 10, 20, and 40 IU PMSG twice (on days 0 and 4), and three CLO subgroups were injected intra-peritoneally with 10, 15, and 20 μg cloprostenol acetate twice (on days 0 and 4). The results showed that 93.33 and 66.67% of synchronized mice displayed estrus within 18.68-37.59 h following CLO and PMSG exposure, respectively. Estrus numbers, estrus onset time, and estrus rates in CLO and PMSG groups were greater than in control groups (CG) (P < 0.05). Uterine weights of the PMSG group were higher than that of CLO and CG groups, and the uterine horn longitudinal diameters in experimental mice were greater than CG. Expression levels of FSHR proteins in CLO and PMSG groups increased slightly when compared to CG. In conclusion, CLO and PMSG administration did not clearly af-fect the expression of uterine FSHR proteins in mice. Moreover, PMSG and CLO treatments synchronized estrus and enhanced the uterine de-velopment of mice. The efficacy of CLO on estrus synchronization was greater than PMSG, and the effects of PMSG on uterine development were stronger than CLO. These results have important significance re-garding the modulation of animal reproductive functions.

Lithium induces follicular atresia in rat ovary through a GSK‐3β/β‐catenin dependent mechanism

Lithium chloride (LiCl) is a drug used to treat bipolar disorder, but has side effects in the female reproductive system. Although lithium is known to decrease folliculogenesis and induce follicular atresia in rodent ovaries, its cellular and molecular effects in the ovary have not yet been addressed. To investigate these effects, 23-day-old immature female rats were injected with 10 IU pregnant mare serum gonadotropin (PMSG), followed by injections of 250 mg/kg LiCl every 12 hr for four doses. Ovaries were removed 40 and 48 hr after PMSG administration and prepared for histology, immunohistochemistry, Western blotting, and DNA laddering analysis. Our results showed that in the ovaries of LiCl-treated rats, few antral but more atretic follicles were present compared to those of the control rats. The induction of atresia by LiCl was further confirmed by the presence of DNA fragmentation, accompanied by a reduced level of 17β-estradiol in the serum. At the cellular level, lithium significantly decreased the number of proliferating cell nuclear antigen (PCNA)-positive cells and conversely increased the number of TUNEL-positive cells in the granulosa layer of the antral follicles. At the molecular level, lithium increased the level of phosphorylated glycogen synthase kinase-3β, and unexpectedly decreased the expression of active (stabilized) β-catenin. Altogether, our results indicate that lithium disrupts the balance between proliferation and apoptosis in granulosa cells, leading to follicular atresia possibly through the reduction in both the stabilized β-catenin and 17β-estradiol synthesis.

The effects of time and dose of pregnant mare serum gonadotropin (PMSG) on reproductive efficiency in hair sheep ewes

The aim of this study was to evaluate the effects of dose and application time of pregnant mare serum gonadotropin (PMSG) on reproductive performance of hair sheep ewes synchronized with fluorogesterone acetate (FGA) under tropical conditions of Northeastern Mexico. Ninety-nine hair ewes (63 Blackbelly and 36 Pelibuey) were treated with intravaginal sponges during 10 days. After insertion of FGA sponges, ewes were divided into four groups, and PMSG was injected intramuscularly at doses of 100, 200, and 400 IU. Relative to FGA sponge removal, PMSG was administrated at -48 h, -24 h, and at sponge removal. PMSG was not administered to the control group. Control ewes had similar (P > 0.05) lambing rate, fertility, and fecundity than those treated with 100 IU of PMSG, but lower (P < 0.05) percentages to these variables than those treated with 200 and 400 IU of PMSG. Time to estrus decreased linearly, and ovulation rate increased quadratically as PMSG dose increased (0 to 400 IU). Administration of PMSG before sponge removal increased (P < 0.01) response to estrus and decreased (P < 0.01) interval to estrus compared with control. Ovulation rate, lambing rate, fertility, and fecundity were not affected (P > 0.05) by administration time of PMSG. Both dose and time of PMSG application did not affect (P > 0.05) pregnancy rate, percentage of single and multiple lambing, and prolificacy. In conclusion, results show that the dose of 400 IU of PMSG administered before sponge withdrawal in an estrus synchronization protocol improved reproductive efficiency of hair sheep ewes.

Expression of SDF-1α and leptin, and their effect on expression of angiogenic factors in mouse ovaries

ObjectiveOvarian angiogenesis plays an important role in folliculogenesis. However, little is known about the expression of angiogenic factors during follicular development according to female age. Stromal cell derived factor-1α (SDF-1α) plays a role in granulosa cell survival and embryo quality as an angiogenic chemokine. Leptin is also involved in folliculogenesis and angiogenesis. This study examined expression of SDF-1α and leptin, and their effects on the expression of angiogenic factors in the ovary during follicular development according to female age.MethodsOvaries were collected from C57BL mice of two age groups (6-9 weeks and 24-26 weeks) at 6, 12, 24, and 48 hours after 5 IU pregnant mare's serum gonadotropin (PMSG) injection. The expression of ovarian SDF-1α and leptin mRNA was evaluated by RT-PCR. In the organ culture experiment, the ovaries were cultured in transwell permeable supports with Waymouth's medium treated with various doses of SDF-1α (50-200 ng/mL) or leptin (0.01-1 µg/mL) for 7 days. Then, mRNA expression of vascular endothelial growth factor (VEGF), endothelial nitric oxide synthase (eNOS), and visfatin were examined in the cultured ovaries.ResultsExpression of SDF-1α and leptin in the ovary was significantly lower in the aged mouse group compared to the young mouse group (p<0.05). Expression of these two factors increased with follicular development after PMSG administration. SDF-1α treatment stimulated visfatin expression in a dose-dependent manner, while leptin treatment significantly increased eNOS expression.ConclusionThese results suggest that decrease of ovarian SDF-1α and leptin expression may be associated with aging-related reduction of ovarian function. SDF-1α and leptin may play a role in follicular development by regulating the expression of angiogenic factors in mouse ovaries.

Expression of cystic fibrosis transmembrane conductance regulator in rat ovary

The protein expression of cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-activated Cl(-) channel, in ovarian stimulated premature female rat ovary during a cycle of follicle development and corpus luteum formation was investigated. Animals were injected with 10 U pregnant Mare's serum gonadotropin (PMSG) and subsequently 10 U hCG 48 h later. Time-dependent immunohistochemistry and Western blotting experiments were performed before and 24, 48, 72 h after hCG treatment. The immunohistochemistry revealed that administration of PMSG stimulated the CFTR expression in thecal cell layer and granulosa cell layer of mature follicles 48 h post injection, coincident with the PMSG-induced peak in follicular estradiol. However, the expression of CFTR in the granulose lutein cell layer and thecal lutein cell layer was time-dependently reduced following hCG injection, in accordance with the gradually increased progestogen level during luteum corpus formation. Western blotting analysis demonstrated that rat ovarian tissue expressed the special CFTR band at 170 kD. It is concluded that cAMP-dependent Cl(-) channels are involved in regulation of follicle development and luteum formation.

Concentrations de l’hormone lutéinisante et de la progestérone chez des vaches zébus Azawak soumises à différents protocoles de synchronisation de l’oestrus

Quinze vaches zébus Azawak ont été soumises à deux protocoles de synchronisation de l’oestrus. Les concentrations plasmatiques de LH et le pourcentage des femelles ayant exhibé un comportement d’oestrus ont été répertoriés. Le premier traitement de synchronisation (T1, n = 9) a été effectué avec des implants sous-cutanés de norgestomet placés sous la peau de l’oreille pendant 10 jours. Le traitement a été complété par des injections de valérate d’oestradiol, de prostaglandine F2α et de pregnant mare serum gonadotropin (PMSG). Le deuxième protocole (T2, n = 6) a consisté en deux injections de prostaglandine F2α à 11 jours d’intervalle, suivies d’une injection de PMSG deux jours après la deuxième injection de prostaglandine. Des échantillons de sang ont été prélevés à trois heures d’intervalle pendant cinq jours après le retrait de l’implant (T1) ou l’injection de PMSG (T2) pour mesurer les taux de LH. Le taux d’oestrus induit a été plus élevé chez les vaches du groupe T2, mais cette différence n’a pas été significative. En général, le temps observé entre la fin du traitement et le début de l’oestrus a été plus court chez les femelles traitées au norgestomet (35,9 ± 3,9 h) que chez celles traitées à la prostaglandine (49,5 ± 5,8 h). Sept sur les neuf femelles traitées au norgestomet et seulement deux sur les six traitées à la prostaglandine ont présenté un pic de LH. L’intervalle moyen entre la fin des traitements et le pic de LH a eu tendance à être plus long chez les femelles traitées à la prostaglandine que chez celles traitées au norgestomet. Une femelle a présenté des concentrations de LH anormalement élevées après le retrait de l’implant. Toutefois, aucun pic de LH n’a été observé pendant la période de suivi.

Local production of the gonadotropic hormones in the rat ovary

The gonadotropic hormones, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) are synthesized by and released from the anterior pituitary in response to the hypothalamic gonadotropin-releasing hormone (GnRH) signaling. In the female, LH and FSH affect folliculogenesis, ovarian steroid production, oocyte maturation, ovulation and corpus luteum formation. We have recently studied the expression of GnRH and its receptor in the rat ovary and found organ-specific, estrous cycle-dependant, fluctuations. Subsequently, we wished to determine whether rat ovaries also express gonadotropic hormones. Using RT-PCR, we detected LHβ, FSHβ and the common α-subunit mRNA's in intact follicles, theca cells, corpora lutea and in meiotically competent and incompetent oocytes. Granulosa cells, however, express mRNA's for LHβ and the common α-subunit, but not for FSHβ. We cloned and sequenced the ovarian LHβ transcript and found it to be longer (2.3 kb) than the one produced by pituitary gonadotropes (0.8 kb), due to a longer 5′-UTR. We studied the regulation of ovarian LHβ mRNA in sexually immature female rats administered with pregnant mare serum gonadotropin (PMSG) and in adult cyclic rats. PMSG administration caused a significant decrease in LHβ mRNA expression, detected by real-time PCR. Similarly, LHβ mRNA levels were lower on estrous morning versus proestrous evening. Interestingly, ovarian content of LH remained unchanged following hypophysectomy, although ovarian weight was immensely reduced. Taken together, it seems probable that ovarian LH is heterologously/homologously regulated by pituitary, and possibly also by local gonadotropins. Thus, these findings may imply the existence of a local GnRH–gonadotropin axis in the mammalian ovary that may be involved in the management of processes that lead to ovulation.

Hormonal induction of ovulation stimulates atresia of antral follicles in a vespertilionid bat, Scotophilus heathi

Scotophilus heathi is a seasonally monoestrous subtropical vespertilionid bat found at Varanasi, India. Although the antral follicles remain present in the ovaries of S. heathi from November till March, ovulation is delayed in this species until early March. In order to understand the mechanism of ovulation suppression during this period of delayed ovulation, the effects of human chorionic gonadotropin (hCG), pregnant mare's serum gonadotropin (PMSG), follicle stimulating hormone (FSH) and gonadotropin releasing hormone agonist (GnRH agonist) on ovarian morphology and steroid concentration were investigated. Hormonal treatments were given as a single i.p. dose 24 h after capture. The bats were sacrificed 48 h after the injection. Treatment with hCG, PMSG, FSH and GnRH agonist failed to induce ovulation in S. heathi, although these hormones produced a high degree of ovarian stimulation. The administration of hCG and PMSG induced ovarian enlargement, intense hyperemia, marked changes in the interstitial cells (ICs), development of several antral follicles and a varying degree of abnormalities in the oocytes of most of the antral follicles. In the bats treated with hCG, PMSG and GnRH agonist, androstenedione concentration increased significantly to extraordinarily high levels, whereas estradiol concentration decreased. Administration of FSH caused regression of ICs and pyknosis of granulosa cells in the majority of antral follicles. FSH did not enhance androstenedione concentration. The results of the present study suggest that the failure of hormonal treatments to induce ovulation during the period of delayed ovulation might be due to a seasonal desensitization of ovarian follicles in S. heathi. The hormonal treatment instead stimulated the ICs to produce a high level of androstenedione resulting in atretic changes of the antral follicles.

Tertiary Follicular Growth Wave Dynamics after Oestrus Synchronization and Superovulation in Sheep and Goats

Tertiary Follicular Growth Wave Dynamics after Oestrus Synchronization and Superovulation in Sheep and Goats

Expression and regulation of gonadotropin‐re leasing hormone (GNRH) and its receptor MRNA transcripts during the mouse ovarian development

The present study examines the expression and regulation of gonadotropin‐releasing hormone (GnRH) and its receptor (GnRH‐R) mRNA levels during mouse ovarian development. A fully processed, mature GnRH mRNA together with intron‐containing primary transcripts was expressed in the immature mouse ovary as determined by Northern blot analysis and reverse transcription‐polymerase chain reaction (RT‐PCR). The size of ovarian GnRH mRNA was similar to that of hypothalamus, but its amount was much lower than that in the hypothalamus. Quantitative RT‐PCR procedure also revealed the expression of GnRH‐R mRNA in the ovary, but the estimated amount was a thous and‐fold lower than that in the pituitary gland. We also examined the regulation of ovarian GnRH and GnRH‐R mRNA levels during the follicular development induced by pregnant mare's serum gonadotropin (PMSG) and/or human chorionic gonadotropin (hCG). Ovarian luteinizing hormone receptor (LH‐R) mRNA was abruptly increased at 48 h after the PMSG administration and r...

Serum progesterone and estradiol-17beta concentrations, and lapaloscopic observations of the ovary in the cheetah (Acinonyxjubatus) with pregnant mare serum gonadotropin and human chorionic gonadotropin treatments.

In 3 adult female cheetahs, induced-superovulation treatment was conducted, by means of 200 IU of pregnant mare serum gonadotropin (PMSG) and 100 IU of human chorionic gonadotropin (hCG) 80 hr after PMSG. The administration of PMSG created a sharp increase in the estradiol-17beta concentration, resulting in 232 pg/ml 8 hr later in one specimen out of three. The hCG administration showed an increase in the progesterone concentration of 2.29 ng/ml 46 hr later. In addition, after direct observation of the ovary surface by laparoscopy, 5 follicles in the right ovary over 2 mm in diameter, and 7 corpora lutea (5 in the right ovary and 2 in the left) were found. It is assumed that ovulation can be induced with hCG after 80 hr on PMSG during a cheetah's diestrus or proestrus.

Concentrations of Tissue-Type Plasminogen Activator and Relaxin in Normal and Induced-Cystic Follicles of Gilts

Manipulation of one ovary in prepubertal gilts treated with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) results in cysts on the manipulated ovary and corpora lutea (CL) on the non-manipulated (control) ovary. Because tissue-type plasminogen activator (tPA) might play a role in follicular rupture and because relaxin might increase tPA production, concentrations of tPA and relaxin in manipulated and control follicles were measured at different stages of development. Prepubertal gilts were treated with 1000 IU PMSG followed by 750 IU hCG at 72 hr later. Follicles on one ovary in each gilt were manipulated at laparotomy 48 hr after PMSG administration. Gilts were ovariectomized at 72, 90, 108, 114, 144, and 216 hr after PMSG. Concentrations of tPA and relaxin were determined for follicular fluid from follicles dissected free of ovarian stroma and snap frozen in liquid nitrogen and media from follicles cultured for 48 hr. Relaxin did not differ between treatment groups (manipulated and control) at any time (P > 0.05); whereas, tPA was greater in control follicles at 114 hr after PMSG than in manipulated follicles (P < 0.01). The effect of pyrilamine, a histamine-1 receptor antagonist, on tPA concentrations was determined in manipulated and control follicles collected at 3, 12, 24, 42, and 66 hr after manipulation. Concentrations of tPA were similar in control and manipulated follicles for gilts treated with pyrilamine, but again control follicles had greater (P < 0.05) tPA concentrations at 114 hr after PMSG. Thus, tPA seems to be involved in ovulation, and blockage of ovulation and subsequent cyst formation results from inadequate tPA activity in manipulated follicles.

Expression and Localization of Inducible Nitric Oxide Synthase in the Rat Ovary: A Possible Involvement of Nitric Oxide in the Follicular Development

In the present study, using Northern blot analysis, we demonstrated that the level of iNOS mRNA in the ovary of immature rats decreased after 6 h of pregnant mare serum gonadotropin administration and recovered gradually up to the untreated level 48 h after the administration. Bothin situhybridization and immunohistochemistry revealed that iNOS mRNA and protein was predominantly localized in granulosa cells in most of immature follicles, but not in mature follicles with an antrum, which was a consistent finding regardless of gonadotropin treatment. Furthermore, we found that cultured granulosa cells had the ability to express iNOS mRNA in the presence of cytokines such as tumor necrosis factor-α, interleukin-1β and interferon-γ, which are inherently detectable in the ovary. These results raise the possibility that locally produced NO synthesized by iNOS may be involved in the developmental status of ovarian follicles in concert with gonadotropin.

Evidence for an inverse relationship between apoptosis and inducible nitric oxide synthase expression in rat granulosa cells: a possible role of nitric oxide in ovarian follicle atresia.

Ovarian follicle atresia is thought to be induced through apoptosis of granulosa cells. This study was designed to investigate the possible involvement of nitric oxide (NO) in granulosa cell apoptosis. In immature rat ovaries obtained 48 h after pregnant mare serum gonadotropin administration, immunohistochemistry and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), a method to detect apoptotic cells, revealed that inducible NO synthase (iNOS) was predominantly localized in granulosa cells in most healthy immature follicles with TUNEL-negative granulosa cells. In contrast, all atretic follicles with TUNEL-positive granulosa cells were iNOS-negative whatever the developmental stage of the follicle. In cultured granulosa cells, the addition of S-nitroso-N-acetyl-DL-penicillamine (SNAP), an NO generator, directly inhibited spontaneously occurring apoptosis. These results suggest that NO produced by iNOS in granulosa cells of immature follicles may prevent ovarian follicle atresia by inhibiting granulosa cell apoptosis in an autocrine/paracrine manner.

Expression of Fas ligand in murine ovary.

Corresponding to the expression of Fas in the ovarian oocytes as previously reported (Guo et al., Biochem Biophys Res Commun 1994; 203:1438-1446; Mori et al., JSIR 1995; 9:49-50), the expression of Fas ligand (FasL) in the ovarian follicle was found to be restricted in the area of granulosa cells by the indirect immunofluorescence (IIF) test. Reverse transcriptase/polymerase chain reaction (RT/PCR) technique coupled with Southern blot hybridization analysis showed that the highest level of FasL mRNA was demonstrated in murine ovaries and granulosa cells 1 day after the administration of pregnant mare's serum gonadotropin (PMSG), while the level of FasL mRNA became very weak on the day 5, respectively. The observed gradual decrease in FasL mRNA could not be attributed to a generalized degradation of cellular RNA during atresia, as evidenced by the presence of constitutive expression of elongation factor 1 alpha (EF-1 alpha) mRNA in murine ovaries and granulosa cells treated with PMSG. Furthermore, in situ hybridization analysis with a FasL-specific probe confirmed that FasL was specifically localized in the granulosa cells of most follicles and its expression was regulated by PMSG administration. FasL localized in granulosa cells might possibly play an important role in the formation of the ovarian atretic follicles, most likely depending on PMSG administration.