Adenosine as substrate and receptor agonist in the ovary.

In the present study the possible dual effects of adenosine as substrate and adenosine receptor agonist in rat granulosa cells, cumulus-oocyte complexes, luteal cells and ovarian membranes are discussed. Adenosine is an indispensable compound in cell energy metabolism, as precursor to cofactors, second messenger and nucleic acids. Adenosine is also an agonist to adenosine receptors.

Effects of inhibitors of protein synthesis on the ovulatory process of the perfused rat ovary.

The effects of two different protein synthesis inhibitors (cycloheximide and puromycin) on the ovulatory process were examined in vitro using a perfused rat ovary model. Ovaries of PMSG (20 i.u.

Sphingosine and psychosine, suggested inhibitors of protein kinase C, inhibit LH effects in rat luteal cells.

The possible involvement of protein kinase C on luteinizing hormone (LH) effects in dispersed rat luteal cells was investigated using two substances that have been reported to be protein kinase C inhibitors, sphingosine and psychosine. Sphingosine efficiently inhibited protein kinase C activity both in brain and luteal cytosol fractions. Both substances inhibited LH-stimulated cyclic adenosine monophosphate (cAMP) accumulation in a dose-dependent fashion with an LD50 at 3-7 microM (sphingosine) and 40 microM (psychosine).

Histamine stimulates progesterone synthesis and cyclic adenosine 3',5'-monophosphate accumulation in isolated preovulatory rat follicles.

The effect of histamine on progesterone synthesis and cyclic adenosine 3',5'-monophosphate (cAMP) accumulation was studied in superfused and incubated follicles dissected free from immature rats treated with pregnant mare serum gonadotrophin (PMSG). Histamine, like LH, increased the progesterone synthesis, but to a smaller extent. The H2-antagonist, cimetidine, inhibited completely the histamine-induced progesterone increase while the H1-antagonist, pyrilamine, as well as propranolol and atropine did not affect the initial response but modified its duration.

Effects of forskolin, luteinizing hormone and prostaglandin F2 alpha on isolated rat corpora lutea.

The diterpene forskolin increased in a dose-dependent way cyclic AMP (cAMP) accumulation in isolated corpora lutea from immature rats injected with an ovulatory dose of pregnant mare's serum gonadotropin (PMSG). The cAMP increase was significant already after 1 min of incubation with forskolin, and cAMP continued to rise to a maximum at about 30-60 min, with a clear decrease after 240 min of incubation. The forskolin effect was more pronounced in very young corpora lutea (1-day-old) than in older corpora lutea.

Adenosine potentiates the effects of LH and isoproterenol on luteal cells but not on isolated intact corpora lutea.

Adenosine potentiated the stimulatory effect of luteinizing hormone (LH) in a dose-dependent way on the production of cyclic AMP (cAMP) in isolated cells from heavily luteinized rat ovaries and from individual rat corpora lutea of various ages (2- and 5-day-old). Such an effect has earlier been reported only for cells from heavily luteinized ovaries (Behrman et al. 1983).

Studies on the morphology of the isolated perfused rabbit ovary. II. Ovulation in vitro after HCG-treatment in vivo.

Ovulation was induced in rabbits by intravenous administration of human chorionic gonadotrophin (HCG), and 4-5 h later the ovaries were isolated and introduced into an in-vitro perfusion system containing synthetic medium with albumin. Rupture of follicles occurred in vitro within the physiological time range (mean 11.3 h after injection of HCG), although with a reduced frequency.

Studies on the morphology of the isolated perfused rabbit ovary. I. Effect of long-term perfusion.

Isolated ovaries from untreated, sexually mature rabbits were introduced into an in vitro perfusion system and perfused with a chemically defined medium containing albumin. The ovaries were perfused for up to 15 h (mean 11.5 h) and then processed for morphological investigation.

Effects of PGF2 alpha and indomethacin on ovulation and steroid production in the isolated perfused rabbit ovary.

Both ovaries of 31 rabbits were perfused with a chemically defined medium in vitro in a recirculation system. In one series of experiments, hCG (100 IU) was injected iv 5-6 h prior to anaesthesia and surgery. Approximately 1 h later the perfusion was started.

Prostaglandin F2 alpha inhibition of epinephrine stimulated cyclic AMP and progesterone production by rat corpora lutea of various ages.

Epinephrine can mimic the stimulatory effects of LH in vitro on cyclic AMP (cAMP) and progesterone production by isolated rat corpora lutea. The aim of the present study was to test whether the effects of epinephrine in vitro on the rat corpus luteum, as with LH, can be inhibited by prostaglandin F2 alpha (PGF2 alpha). The stimulatory effect of epinephrine on tissue levels of cAMP in 1-day-old corpora lutea was not inhibited by PGF2 alpha.

Development of catecholamine responsiveness in granulosa cells from preovulatory rat follicles--dependence on preovulatory luteinizing hormone surge.

Factors responsible for development of catecholamine (CA) responsiveness in granulosa cells (Gc) of preovulatory follicles from immature rats injected with 10 IU of pregnant mare's serum gonadotropin (PMSG) on Day 26 were studied. CA did not stimulate cyclic AMP (cAMP) production in whole follicles isolated before (morning) or after (evening) the preovulatory gonadotropin surge on Day 28, while newly formed corpora lutea found on Day 29 responded to CA. Gc from the preovulatory follicles did not respond to CA when tested immediately after isolation.

Prostaglandin levels in preovulatory follicles from rabbit ovaries perfused in vitro.

Prostaglandin (PG) levels in follicular fluid from preovulatory follicles of rabbit ovaries perfused in vitro were measured in order to compare PG changes in this model system with those that occur in vivo and in isolated, LH-treated follicles in vitro. One ovary from each rabbit was perfused without further treatment (control). The other ovary was exposed to LH (0.

Involvement of cyclic AMP and protein synthesis in the desensitization of the prepubertal rat ovary.

Luteinizing hormone (LH) stimulates cyclic AMP (cAMP) production in ovarian cells. After the initial stimulation there is a development of refractoriness (desensitization) to a subsequent LH stimulation. The present investigation was designed to study the role of cAMP in this desensitization process and the influence of protein-synthesis inhibitors.

Effect of gonadotrophin-releasing hormone (GnRH) and GnRH agonists upon accumulation of progesterone, cAMP and prostaglandin in isolated preovulatory rat follicles.

To study the acute and direct effects of GnRH agonists preovulatory follicles were isolated from PMSG-treated immature rats and incubated for 15-360 min in modified Kreb's bicarbonate buffer. The levels of cAMP, prostaglandin E, and progesterone were analysed in the tissue and/or incubation media. GnRH and two GnRH agonists produced a dose-dependent stimulation of progesterone production with maximal levels 5-6-fold higher than the control group.

The preovulatory decline in follicular oestradiol is not required for ovulation in the rabbit.

Using a method of in vitro perfusion of the rabbit ovary with a chemically defined medium in a recirculation system, normal appearing follicular ruptures occurred following exposure of the ovaries to hCG in vivo (100 IU) or following addition of LH (0.25 microgram/ml of NIH B9) to the perfusate. The addition of oestradiol-17 beta (10 micrograms/ml) to the perfusate did not inhibit these follicular ruptures, although the follicular fluid oestradiol contents were increased more than 100-fold as compared to the control side not receiving the addition of oestradiol.

Catecholamine stimulation of cyclic AMP and progesterone production in rat corpora lutea of different ages.

In vitro effects of catecholamines (adrenaline and noradrenaline) and adrenergic antagonists on adenosine 3',5'-cyclic monophosphate (cAMP) and progesterone production by rat corpora lutea (CL) of different ages (1-8 days old) were studied. To obtain defined ages of CL a pregnant mare's serum gonadotrophin (PMSG) model was used. The effect of catecholamines on cAMP decreased with luteal age while the effect on progesterone production was maximal on 5 day old CL.

The study of ovulation in the isolated perfused rabbit ovary. II. Photographic and cinematographic observation.

Using the model of in vitro perfusion of the isolated rabbit ovary, in which follicular ruptures can be observed, in this paper we report on the photographic and cinematographic recording of the gross anatomy of ovulation. The observed sequence of events and their timing were very similar to that which has been reported for ovulation observed in vivo. In addition, the use of infrared film made it possible to visualize intrafollicular events prior to rupture.

The study of ovulation in the isolated perfused rabbit ovary. 1. Methodology and pattern of steroidogenesis.

A previously described technique for perfusion of the isolated rabbit ovary in a recirculating system (Ahrén et al., 1975) was modified for studies of preovulatory follicular development and ovulation. Follicular ruptures were induced either by injection of human chorionic gonadotropin (hCG) to the animal 5-6 h prior to perfusion or by adding luteinizing hormone (LH) directly to the medium during perfusion.

Luteal blood flow and plasma steroids in rats with corpora lutea of different ages.

Ovarian and luteal blood flow rates were measured at different stages of luteal development in anaesthetized rats using 15 +/- 5 micron radioactive microspheres. Ovulations were induced by injection of 8 IU of PMSG at 28 days of age. Steroid concentrations in peripheral plasma were determined using radioimmunoassays.

Different patterns of cyclic AMP accumulation in rat corpora lutea after LH stimulation in vitro and in vivo.

Tissue levels of cyclic AMP were measured in rat corpora lutea at various times after administration of LH in vitro and in vivo. In-vitro addition of LH produced more pronounced accumulation of cyclic AMP in very young corpora lutea (1-day-old) than in the older corpora lutea (3- and 7-day-old). When LH was administered in vivo the highest accumulation of cyclic AMP was seen in the older corpora lutea (7- and 3-day-old).

Desensitization of the cyclic AMP system in rat corpora lutea. Comparison between the effects of hCG and LH.

The changes in hormonal responsiveness of corpora lutea produced by a single injection of hCG or ovine LH were studied in PMSG-treated immature rats with well-characterized corpora lutea of different ages. The hormonal response was measured as LH-stimulated accumulation of cyclic AMP in isolated, whole corpora lutea and, in some experiments, as LH stimulated luteal adenylate cyclase activity. Injection of hCG produced a complete desensitization (refractoriness) of very long duration (4-5 days) as has earlier been found in rats with heavily luteinized ovaries, while the injection of LH in equivalent doses, produced only a partial desensitization of much shorter duration (12-48 h).

Prolongation of the effects of gonadotrophins and prostaglandin E2 on ovarian cyclic AMP formation by inhibitors of protein synthesis.

The effects of LH, FSH and PGE2 on the accumulation of cyclic AMP by isolated whole ovaries from 23-24 day-old rats were studied in time-course experiments in presence and absence of two inhibitors of protein synthesis, puromycin and cycloheximide. In absence of these inhibitors ovarian cyclic AMP levels reached peak levels within 30 min for all three stimulators and returned towards pre-stimulation levels within 2-4 h. When puromycin (500 microgram/ml) or cycloheximide (5 microgram/ml) was present together with LH, FSH or PGE2, respectively, ovarian cyclic AMP levels did not decrease after the initial peak but remained high for the entire incubation period (4-5 h).