Increased production of ovarian thromboxane in gonadotropin-treated immature rats: relationship to the ovulatory process

Thromboxane (TX) B2, a stable metabolic product of hydrolysis of TXA2, was measured by radioimmunoassay in tissue extracts of ovaries of immature rats pretreated with pregnant mare's serum gonadotropin and human chorionic gonadotropin. Ovarian concentrations of TXB2 increased before, and remained elevated after, the time of ovulation. In a subsequent study, ovulation was inhibited in a dose-dependent fashion by a reported TXA2 receptor antagonist, AH23848. Nevertheless, inhibition of the preovulatory rise in synthesis of TXB2 by furegrelate (a thromboxane synthetase inhibitor) did not prevent ovulation. Nor was the blockade of ovulation caused by indomethacin (a cyclooxygenase inhibitor) reversed by a TXA2 mimetic (U-46619). It does not appear that a preovulatory increase in ovarian thromboxane is an obligatory component of the ovulatory mechanism of gonadotropin-primed immature rats.     

Kinetic and structural evidence for the identification of 11-hydroxythromboxane B2 dehydrogenase as cytosolic aldehyde dehydrogenase

We have recently purified 11-hydroxythromboxane B₂ dehydrogenase from porcine kidney and identified it as cytosolic aldehyde dehydrogenase (EC 1.2.1.3) based on amino acid analysis and other protein characteristics. In the present paper we have studied the catalytic interaction of thromboxane B₂ (TXB₂) with different aldehyde substrates and a potent aldehyde dehydrogenase inhibitor, disulfiram. TXB₂ was a competitive inhibitor of the aldehyde dehydrogenase reaction in assays with 3,4-dihydroxyphenylacetaldehyde, a high affinity substrate. The conversion of TXB₂ to 11-dehydro-TXB₂ was also inhibited by propanal and disulfiram.

The protein characteristics of the enzyme have also been further studied. The native enzyme is a tetramer and has an isoelectric point of 7.0 which is comparable with that of cytosolic aldehyde dehydrogenases from other species. Taken together the present data further indicate that 11-hydroxythromboxane B₂ dehydrogenase is identical with cytosolic aldehyde dehydrogenase and that substrates and inhibitors of aldehyde dehydrogenase interact with thromboxane metabolism in vitro.     

An improved malondialdehyde assay for estimation of thromboxane synthase activity in washed human blood platelets

After stimulation of the washed human blood platelets by arachidonic acid (AA), the concurrent evaluations for formed malondialdehyde (MDA) measured by the common photometrical thiobarbituric acid (TBA) method, and for thormboxane B₂ (TXB₂ measured by gas chromatography, revealed that the formed MDA exceeded the amount of TXB₂ on a molar base. However, MDA and TXB₂ originating from thromboxane synthesis activity should be produced in approximately equimolar amounts. By treatment of the stimulated platelet samples with stannous chloride it is possible to reduce all peroxidized products of AA which generate MDA otherwise during the TBA reaction and to estimate MDA and TXB₂ in a ratio of nearly 1:1. the stannous chloride treatment does not destroy the MDA and does not influence the TBA reaction with MDA. Therefore the simple and quick TBA method can be used after stannous chloride treatment for estimation of thromboxane synthase activity in AA stimulated washed human platelets.     

Role of progesterone in the modulation of the preovulatory surge of gonadotropins and ovulation in the pregnant mare's serum gonadotropin-primed immature rat and the adult rat

The role of progesterone in the regulation of the preovulatory surge in gonadotropins and ovulation was examined in this study by use of a potent antagonist of progesterone, RU 486 (17 beta-hydroxy-11 beta-[4-dimethyl-aminophenyl]-17 alpha- [prop-1-ynyl]estra-4,9-diene-3-one). The immature rat primed with pregnant mare's serum gonadotropin (PMSG) and the cycling adult animal were the models used to verify the role of progesterone. When RU 486 (200 micrograms/rat) was given as a single dose on the morning of proestrus, there was a significant reduction in the preovulatory surge levels of gonadotropins and ovulation in both animal models. Serum progesterone levels in both models at the time of death on the evening of proestrus were unaltered upon treatment with RU 486. RU 486 did not have any effect on gonadotropin levels in immature rats 7 days after castration. These results show that the actin of RU 486 on the preovulatory gonadotropin surge is due to an antagonism of the action of progesterone on the hypothalamic-pituitary axis. Thus, a role for progesterone in modulating the preovulatory surge of gonadotropins and, consequently, ovulation is strongly suggested.     

Antiovulatory effect of a single injection of pure antiestrogen ZK 191703 at early stage of rat estrus cycle

The effects of ZK 191703 (ZK), a pure antiestrogen, on ovulation, follicle development and peripheral hormone levels were investigated in rats with 4-day estrus cycle and gonadotropin-primed immature rats in comparison to tamoxifen (TAM)-treatment. In adult rats, a single s.c. injection of ZK (5 mg/kg) or TAM (5 mg/kg) at an early stage of the estrus cycle (diestrus 9:00) inhibited ovulation, and was associated with suppression of the surge of preovulatory LH, FSH and progesterone. In rats treated with ZK or TAM at a late stage of the estrus cycle (proestrus 9:00), no inhibitory effects on ovulation, the gonadotropin and progesterone surge were detected. ZK treatment at diestrus 9:00, in contrast to TAM, increased the baseline LH level. When immature rats were treated with antiestrogens in the earlier stage of follicular development, 6 and 30 h but not 48 h or later after injection of gonadotropin (PMSG), ovulation was attenuated, associated with a lowered progesterone level. Unruptured preovulatory follicles were found in most of the ovaries from anovulatory animals treated with ZK or TAM. Antiestrogens, ZK and TAM administered at an early phase of the estrus cycle delay the follicular development functionally and inhibit ovulation in rats and suppression of the preovulatory progesterone surge.     

孕康口服液对肾虚-黄体抑制型先兆流产大鼠的安胎作用

目的:孕康口服液为已上市中成药,为进一步评价其药效,本实验通过建立肾虚-黄体抑制型先兆流产模型,观察孕康口服液的安胎作用。方法:60只妊娠大鼠随机分为正常对照组(NC),模型组(MG),地屈孕酮组(DT,3.02 mg/kg),孕康口服液低剂量组(YK-L,4 ml/kg)、中剂量组(YK-M,6 ml/kg)、高剂量组(YK-H,9 ml/kg),每组10只。自妊娠第1日,每日上午各给药组按规定剂量灌予受试药,NC组、MG组给予等体积的纯化水,连续10 d;每天下午灌胃造模,除NC组给予纯化水外,其余各组按450 mg/kg体质量灌胃羟基脲,连续9 d,第10日按4.0 mg/kg体质量灌胃米非司酮。妊娠第9日,测定各组大鼠背温、抓力、痛阈、自主活动等行为体征;妊娠第11日,各组腹主动脉取血,测定血清雌二醇(E₂)、孕酮(P)、血栓素B₂(TXB₂)水平;摘取卵巢、连胎子宫,观察胚胎个数和直径,计算卵巢、连胎子宫指数。结果:与NC组比较,MG组背温、抓力、痛阈、自主活动次数、胚胎个数、胚胎直径、连胎子宫指数和血清E₂、P、TXB₂水平均显著降低(P＜0.05,0.01)。与MG组比较,孕康口服液各剂量组背温、抓力、胚胎个数、胚胎直径和血清E₂、P水平均显著升高(P＜0.05,0.01);YK-M、YK-H组痛阈、自主活动、连胎子宫指数显著升高(P＜0.05);YK-H组血清TXB₂水平明显升高(P＜0.05)。结论:孕康口服液对肾虚-黄体抑制导致的先兆流产大鼠具有明确的补肾安胎作用,其机制可能与升高血清E₂、P、TXB₂水平,改善肾虚体征和提高胚胎质量有关。     

Blood eicosanoids and immune indices during fasciolosis in water buffaloes

The effects of trickle infections of water buffaloes with Fasciola hepatica (60 metacercariae daily during a period of 20 days) on the blood plasma levels of prostaglandin E₂ (PGE₂), 6-keto-prostaglandin F₁ (6-keto-PG F₁) and thromboxane B₂ (TXB₂) were assessed. F. hepatica specific IgG and T- and B-lymphocyte ratios were evaluated as indicators of the immune response. Although the applied mode of infection did not result in clinical disease, changes in the plasma eicosanoid pattern were observed. Plasma PGE₂ values were significantly elevated in the infected water buffaloes 11 weeks post-infection (w.p.i.). In contrast, transiently but significantly lower TXB₂ values than in the uninfected controls were recorded in the phase of chronic fasciolosis. Plasma 6-keto-PGF₁ values were not considerably altered by the infection throughout the study period. F. hepatica-specific IgG were detected from 4 to 21 w.p.i. The proportion of peripheral T- and B-lymphocytes shifted towards B-cells from 2 to 12 w.p.i., gradually returning to control values afterwards. Although the water buffaloes appeared to be rather resistant to trickle infection with F. hepatica, moderate changes in plasma eicosanoid patterns were observed, indicating tissue damage and/or inflammation. Induction of the immune response could be monitored by an increase of F. hepatica-specific IgG, which was paralleled by a relative increase of the B-lymphocyte population.     

Effects of the antiandrogen hydroxyflutamide on progesterone secretion by preovulatory rat follicles in vivo and in vitro.

Serum and ovarian progesterone levels and in vitro production of progesterone by preovulatory follicles were measured on proestrus in pregnant mare's serum gonadotropin (PMSG) primed immature rats in which the luteinizing hormone (LH) surge and ovulation were blocked by administration of the antiandrogen hydroxyflutamide. Serum progesterone levels observed at 12:00 on proestrus were significantly elevated, twofold above those observed in vehicle-treated controls, by in vivo administration of 5 mg hydroxyflutamide 4 h earlier. In control rats, proestrous progesterone did not increase until 16:00, in parallel with rising LH levels of the LH surge. No LH surge occurred in the hydroxyflutamide-treated rats, ovulation was blocked, and serum progesterone declined throughout the afternoon of proestrus, from the elevated levels present at 12:00. Administration of human chorionic gonadotropin (hCG) at 11:00 advanced the elevation of serum progesterone by 2 h in vehicle-treated controls and prevented the decline in progesterone levels in hydroxyflutamide-treated rats. The patterns of change in ovarian tissue concentrations with time and treatment were essentially similar to those observed for serum progesterone. In in vitro experiments, progesterone secretion during 24 h culture of preovulatory follicles obtained on PMSG-induced proestrus was significantly increased, sixfold, by addition to the culture media of 370 microM but not of 37 microM hydroxyflutamide. Testosterone (50 nM) and hCG (20 mIU/mL) caused 26- and 14-fold increases, respectively, in progesterone secretion by cultured follicles. Hydroxyflutamide significantly reduced the stimulatory effect of testosterone but not of hCG on progesterone secretion in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)     

Acetyl glyceryl ether phosphorylcholine (platelet-activating factor) mediates heightened metabolic activity in macrophages: Studies on PGE, TXB2 and O2 production, spreading, and the influence of calmodulin-inhibitor W-7

The phospholipid mediator AGEPC (acetyl glyceryl ether phosphorylcholine) was examined for its effects on guinea pig peritoneal macrophages. At a concentration of 10⁻⁹ -10⁻⁶ M, AGEPC evoked release of prostaglandin E (PGE) and thromboxane B₂ (TXB₂) from albumin-elicited macrophages. It also triggerd generation of O⁻₂ by Corynebacterium parvum-induced cells. Moreover, it caused augmented spreading of macrophages. The calmodulin antagonis W-7 attenuated AGEPC-mediated O⁻₂ production and cell spreading whereas prostanoid synthesis was enhanced. These novel actions of AGEPC on the major cellular component of the inflammatory process attest to its role as a potent mediator of immunoinflammatory responses.     

Immature rats show ovulatory defects similar to those in adult rats lacking prostaglandin and progesterone actions

Gonadotropin-primed immature rats (GPIR) constitute a widely used model for the study of ovulation. Although the equivalence between the ovulatory process in immature and adult rats is generally assumed, the morphological and functional characteristics of ovulation in immature rats have been scarcely considered. We describe herein the morphological aspects of the ovulatory process in GPIR and their response to classical ovulation inhibitors, such as the inhibitor of prostaglandin (PG) synthesis indomethacin (INDO) and a progesterone (P) receptor (PR) antagonist (RU486). Immature Wistar rats were primed with equine chorionic gonadotropin (eCG) at 21, 23 or 25 days of age, injected with human chorionic gonadotropin (hCG) 48 h later, and sacrificed 16 h after hCG treatment, to assess follicle rupture and ovulation. Surprisingly, GPIR showed age-related ovulatory defects close similar to those in adult rats lacking P and PG actions. Rats primed with eCG at 21 or 23 days of age showed abnormally ruptured corpora lutea in which the cumulus-oocyte complex (COC) was trapped or had been released to the ovarian interstitum, invading the ovarian stroma and blood and lymphatic vessels. Supplementation of immature rats with exogenous P and/or PG of the E series did not significantly inhibit abnormal follicle rupture. Otherwise, ovulatory defects were practically absent in rats primed with eCG at 25 days of age. GPIR treated with INDO showed the same ovulatory alterations than vehicle-treated ones, although affecting to a higher proportion of follicles. Blocking P actions with RU486 increased the number of COC trapped inside corpora lutea and decreased ovulation. The presence of ovulatory defects in GPIR, suggests that the capacity of the immature ovary to undergo the coordinate changes leading to effective ovulation is not fully established in Wistar rats primed with eCG before 25 days of age.     

Prostanoid formation in primary astroglial cell cultures: Ca-dependency and stimulation by A 23187, melittin and phospholipases A2 and C

Prostaglandin (PG) and thromboxane B₂ (TXB₂) biosynthesis was studied in cultured astrocytes from neonatal rat brain hemispheres. After two weeks of cultivation, prostanoids were formed with the spectrum: PGD₂ > TXB₂ > PGF₂ > PGE₂, as measured by specific radioimmunoassays. Under basal conditions PGD₂ biosynthesis (9.55 ng/mg protein/15 min) was in the same order of magnitude as the sum of the other prostanoids. The formation of prostanoids was stimulated in a concentration dependent manner (up to 6–10 fold) by the calcium ionophore A 23187 (0.01–10 μM) as well as by melittin (0.01–5 μg/ml), phospholipase A₂ (10–40 U/ml) and phospholipase C (0.01–1 U/ml). Basal and evoked PG and TXB₂ biosynthesis depended on the availability of Ca²⁺, as demonstrated in Ca²⁺ free incubation medium containing Na₂EDTA (1 μM), or with verapamil (100 μM) and 3,4,5-trimethoxybenzoic acid-8-(diethylamino)-octylester-HCl (TMB-8, 1–100 μM). Indomethacin (10 μM), mepacrine (100 μM) and p-bromophenacylbromide (50 μ M) inhibited basal and evoked PG formation. Thin-layer chromatography (TLC) detection after incubation of the cells with [³H]arachidonic acid (1 μCi/ml, for 60 min) confirmed the results obtained by radioimmunoassay. Incubation of [³H]arachidonic acid labelled cells with inonophore or phospholipases, followed by lipid extraction and TLC, showed that A 23187 liberated [³H]arachidonic acid predominantly from phosphatidylethanolamine, whereas phospholipase A₂ and C reduced mainly the labelling of the phosphatidyl-inositol/-choline fraction. Potassium depolarization of the cells did not enhance prostanoid formation. Similarly, drugs with affinity to - or β-adrenoceptors, or to dopamine-, 5-hydroxytryptamine-, muscarine-, histamine-, glutamate-, aspartate-, GABA, adenosine- and opioid-receptors failed to stimulate prostanoid biosynthesis. Also compounds like angiotensin, bradykinin and thrombin were ineffective in this respect.

In conclusion, our results confirm that cultured astrocytes possess the complete pattern of enzymes necessary for prostanoid formation and hence might play a crucial role in brain prostanoid biosynthesis. Stimulation of prostanoid biosynthesis involves Ca²⁺-dependent activation of phospholipase A₂, cyclooxygenase reaction and further PG metabolism. However, the endogenous stimulus for enhanced prostanoid synthesis in the brain still has to be established.     

Modulation of ovarian ornithine decarboxylase activity during follicular differentiation/maturation

The role of gonadotropins and estrogen on the regulation of ovarian ornithine decarboxylase was studied during follicular differentiation/maturation. In intact immature rats follicular differentiation/maturation was initiated with sequential administration of estrogen and follicle stimulating hormone. Ornithine decarboxylase activity in response to human chorionic gonadotropin was markedly enhanced (2-fold) in rats with preovulatory antral follicles when compared to rats with non-ovulatory follicles. This increase could be attributed to the alteration in the turnover of the enzyme. Following follicle maturation the half life of the human chorionic gonadotropin stimulated ornithine decarboxylase was increased from 18 to 62 min. This increase in half life was associated with differentition of follicles. In the estrogen treated group (which does not induce follicular differentiation), the half life of the enzyme remained unaltered. The regulation of ornithine decarboxylase through the formation of protein inhibitor antizyme induced by diamino hexane, was unaltered during follicular differentiation.     

Abundant expression of thromboxane synthase in rat macrophages

The cloned cDNA for rat thromboxane (TX) synthase with a size of 1851 bp contained a 1599-bp open reading frame which encoded a 533-amino acid protein sharing 79.7% identity with human TX synthase. RNA blot analysis was carried out with rat cells and tissues. Rat peritoneal macrophages most abundantly expressed mRNA for TX synthase, and its level was not changed by in vivo stimulation of casein. Bone marrow, spleen, lung and thymus also expressed the TX synthase gene. These findings suggest the possibility that TXA₂ plays a role in the immune system.     

Ovulation in the isolated perfused rat ovary as documented by intravital microscopy

Surface cell changes at the apices of preovulatory follicles and ovulations were documented in isolated perfused ovaries from immature rats treated with pregnant mare serum gonadotropin (20 IU) and 48 h later with human chorionic gonadotropin (hCG) (10 IU). A video camera coupled to an inverted microscope and a video recorder captured the preovulatory and ovulatory events at a cellular level. At around 8 h post-hCG, the follicular apex changed from a smooth and optically homogeneous appearance into a rough surface with bleb formation and extrusions of single cells through minute perforations (early stigma formation). At approximately 10 h, a sticky material formed a basketlike structure with trapped cells (late stigma formation). At 12 to 15 h, ovulation took place at a constant speed and with no contractions of the follicular wall. This indicates that ovulation can occur with no visible circumfollicular muscular activity. Furthermore, the observations of a leakage of cells over an extended period of time indicates that the follicular wall is partly digested several hours before ovulation occurs.     

Physiological peripubertal activation of the ovary is not reproduced by pregnant mare serum gonadotropin (PMSG) administration

During the days preceding the first ovulation the ovary of the rat exhibits a remarkable increase in estradiol (E2) and progesterone (P) release in response to gonadotropins. No such increase is observed in the case of androgens (A, testosterone + dihydrotestosterone). The present experiments were undertaken to examine the possibility of reproducing these developmental events by stimulating the ovary with a gonadotropin that has substantial FSH-like activity. In vivo administration of pregnant mare serum gonadotropin (PMSG) to juvenile 29-day-old rats greatly increased the in vitro E2 and A response to human chorionic gonadotropin (hCG) measured 2 days later in the morning. The magnitude of the A response was significantly larger than that of ovaries from juvenile animals or rats in first proestrus. The E2 response was much greater than that of juvenile ovaries but similar to that of ovaries from late proestrous rats. In contrast, the P response to hCG was not enhanced by PMSG. In fact the response was similar to that of juvenile ovaries and markedly less than that of first proestrous rats. This decreased P response was not due to a greater conversion of P to its less active metabolite 20 alpha-hydroxy-4-pregnen-3-one (20 alpha-OH-P). The results suggest that PMSG enhances the E2 and A response of immature ovaries to hCG at the expense of that of P. Treatment of immature rats with PMSG may represent a useful model to study E2 release from preovulatory ovaries, but it cannot be used to reproduce in its entirety the developmental changes in steroidal response to gonadotropins associated with normal puberty.     

Blood flow changes and vascular appearance in preovulatory follicles and corpora lutea in immature, pregnant mare's serum gonadotropin-treated rats

In the present study, synchronized follicular growth, ovulations, and luteogenesis were prematurely induced in 26-day-old immature rats by the s.c. injection of 4 IU of pregnant mare's serum gonadotropin (PMSG) at 2100 h. Relative blood flow of follicles/corpora lutea, fallopian tube, and uterus was measured with radioactive microspheres during the periovulatory period (Day 28, 1700 h-Day 31, 1300 h). Also, follicular/corpus luteal light microscopy and plasma progesterone were studied at the same intervals after PMSG injection. It was found that the relative follicular blood flow did not increase after the endogenous gonadotropin surge (Day 29, 0300-0500 h) and toward ovulation (Day 29, 1300-1500 h). During the same time period, light microscopy showed an interstitial edema and extravasation of erythrocytes appearing in the follicular wall near the time of ovulation. The relative blood flow reached its nadir in the young corpus luteum (21 h after ovulation) and increased thereafter (i.e., 48 h after ovulation). Plasma progesterone showed a preovulatory increase and then declined just prior to the ovulatory period. Between 24 and 48 h after ovulation, parallel increases in relative blood flow, morphological vascularization, morphological luteinization, and plasma progesterone levels were observed in the growing corpus luteum. These data indicate that a functional relationship between blood flow and steroid output may exist within the ovarian follicle and corpus luteum.     

Pharmacological profile of MEN 11066, a novel potent and selective aromatase inhibitor

MEN 11066 is a new non-steroidal compound which potently inhibits human placenta (K_i=0.5 nM) and rat ovarian (K_i=0.2 nM) aromatase in vitro. In vivo, a single oral dose of 0.3 mg kg⁻¹ significantly decreased uterus weight in immature rats after stimulation of uterus growth by androstenedione. MEN 11066 reduced in a dose-dependent manner plasma estradiol levels in adult female rats treated with pregnant mare serum gonadotropin (PMSG). After 2 weeks of repeated daily treatment in adult rats, a significant decrease in uterine weight was observed together with a 65% decrease in plasma estradiol, whereas plasma levels of testosterone, progesterone, aldosterone, corticosterone, cholesterol, LH and FSH were not affected. The lack of any effect by MEN 11066 on adrenal steroids was confirmed by the unchanged plasma corticosterone and aldosterone levels in immature rats and also in adult rats when the repeated treatment with MEN 11066 (15 days) was followed by the administration of a synthetic ACTH analogue. No change in 11β-hydroxylase or 21-hydroxylase activities was produced in vitro by the addition of 10 μM MEN 11066. Fifteen-day treatment with MEN 11066 did not produce changes in several rat hepatic enzymatic activities involved in the metabolism of xenobiotics. These results demonstrated that MEN 11066 is a potent inhibitor of aromatase which does not interfere with the cytochrome P450 involved in the synthesis of other steroids or in the metabolism of xenobiotics.     

Cyclic guanosine 5'-monophosphate-dependent protein kinase II is induced by luteinizing hormone and progesterone receptor-dependent mechanisms in granulosa cells and cumulus oocyte complexes of ovulating follicles

Cyclic GMP (cGMP)-dependent protein kinase II (Prkg2, cGK II) was identified as a potential target of the progesterone receptor (Nr3c3) in the mouse ovary based on microarray analyses. To document this further, the expression patterns of cGK II and other components of the cGMP signaling pathway were analyzed during follicular development and ovulation using the pregnant mare serum gonadotropin (PMSG)-human chorionic gonadotropin (hCG)-primed immature mice. Levels of cGK II mRNA were low in ovaries of immature mice, increased 4-fold in response to pregnant mare serum gonadotropin and 5-fold more within 12 h after hCG, the time of ovulation. In situ hybridization localized cGK II mRNA to granulosa cells and cumulus oocyte complexes of periovulatory follicles. In progesterone receptor (PR) null mice, cGK II mRNA was reduced significantly at 12 h after hCG in contrast to heterozygous littermates. In primary granulosa cell cultures, cGK II mRNA was induced by phorbol 12-myristate 13-acetate enhanced by adenoviral expression of PR-A and blocked by RU486 and trilostane. PR-A in the absence of phorbol 12-myristate 13-acetate was insufficient to induce cGK II. Expression of cGK I (Prkg1) was restricted to the residual tissue and not regulated by hormones. Guanylate cyclase-A (Npr1; GC-A) mRNA expression increased 6-fold by 4 h after hCG treatment in contrast to pregnant mare serum gonadotropin alone and was localized to granulosa cells of preovulatory follicles. Collectively, these data show for the first time that cGK II (not cGK I) and GC-A are selectively induced in granulosa cells of preovulatory follicles by LH- and PR-dependent mechanisms, thereby providing a pathway for cGMP function during ovulation.     

Involvement of cytosolic phospholipase A2 in the ovulatory process in gonadotropin-primed immature rats

The preovulatory LH surge induces a remarkable increase in ovarian prostaglandins (PGs) which help to mediate the ovulatory process. We investigated whether cytosolic phospholipase A2 (cPLA2) has a role in this PG production in PMSG/hCG-primed immature rats. The immunoreactive signal for cPLA2 was localized in both thecal and granulosa layers of mature follicles and became evident in response to gonadotropins. The PLA2 activity in the whole ovarian cytosol rose slightly after PMSG stimulation, persisted relatively constant until 24 h after hCG injection and thereafter increased gradually. Intra-ovarian bursal injection of arachidonyl trifluoromethyl ketone, a specific inhibitor for cPLA2 ( 1.0-3.0 mg/ovary), significantly reduced ovarian PGE2 content and the ovulation rate. These results suggest that cPLA2 exists in periovulatory follicles and functions in PG production related to the ovulation process.     

Participation of the lower brain stem in induction of preovulatory gonadotropin surges in female rats

The role of the lower brain stem in controlling preovulatory gonadotropin surges was investigated in female rats under acute experimental conditions. Electrolytic lesions or diethyldithiocarbamate implantations in the ventrolateral part of the medulla oblongata (VLMO), which were carried out at 1100-1330 h on the day of proestrus, resulted in a blockade of the preovulatory surges of LH, FSH and PRL as well as subsequent ovulation. Such treatments in the dorsomedial part of the medulla oblongata did not affect gonadotropin surges or ovulation. By means of electrolytic lesions in the VLMO, norepinephrine concentrations were significantly reduced in the preoptic-anterior hypothalamic area at 1700-1800 h on proestrus, though they did not change in the mid-posterior hypothalamus. Electrochemical stimulations of the suprachiasmatic part of the preoptic area or norepinephrine injections into the third ventricle at 1400-1500 h on proestrus in animals with VLMO lesions succeeded in induce gonadotropin surges and ovulation. These results suggest that the lower brain stem is involved in the induction of preovulatory gonadotropin surges and that the process may be mediated by the ascending noradrenergic system which originates in the VLMO.