Inhibition of ovulation in rabbits by intrafollicular injection of indomethacin and prostaglandin F antiserum

Indomethacin, an inhibitor of prostaglandin biosynthesis, was 100% effective in blocking luteinizing hormone (LH)-induced ovulation when administered via microinjection directly into 22 follicles in twelve rabbits, 5 hours after intravenous injection of the gonadotropin. Ovulation was similarly blocked in 24 of 25 follicles injected with antiserum prepared against prostaglandin F_2α. Antiserum against prostaglandin E₂, at the same dosage (100 μg lyophilized serum per follicle), was considerably less effective, preventing ovulation in only 6 of 14 follicles. Control follicles injected with the phosphate buffer vehicle, or with normal rabbit serum, underwent normal ovulation and luteinization. LH injection caused a striking increase in concentration of F-type prostaglandins in follicles shortly before ovulation, an increase which was prevented by i.v. or intrafollicular injection of ovulation-blocking blocking dosages of indomethacin. These findings provide evidence in support of a role for prostaglandins, acting at the follicular level, in the process of ovulation.     

Disorders in the thrombocyte link of hemostasis during endotoxinemia and their correction with the inhibitor of prostaglandin biosynthesis, indomethacin

After 10 minutes of endotoxin Salmonella typhimurium (1 mg/kg) injection into rabbits thrombocytopenia appeared, the aggregation and secretory function of circulating platelets reduced, the transformation of platelet forms from disk-shaped into spheroidal took place. On the surface of plasmatic membranes the pseudopodies and aggregates examining samples of PRP were observed. Indomethacin, blockator of biosynthesis of prostaglandins results in normalisation of morphofunctional properties of platelets.     

Cycloheximide inhibition of ovulation, prostaglandin biosynthesis and steroidogenesis in rabbit ovarian follicles

Cycloheximide (5 mg/kg, i.v.) significantly inhibited ovulation in the rabbit when it was administered as early as 20 h before the ovulation process was initiated by hCG, and as late as 1 h after hCG. The ovulation rate was significantly reduced, but follicular biosynthesis of prostaglandins E and F was only partly inhibited. The biosynthesis of progesterone and oestradiol in follicles during the early stages of the ovulation process was also inhibited. Cycloheximide may therefore inhibit ovulation by a mechanism which is different from the action of indomethacin, and this mechanism may involve the suppression of ovarian steroidogenesis.     

Effect of human chorionic gonadotrophin and indomethacin on ovulation, steroidogenesis and prostaglandin synthesis in preovulatory follicles of PMSG-primed immature rats

Immature rats were treated with PMSG followed 56 h later by 10 i.u. hCG. Follicles were removed at intervals after hCG injection. Transient increases in progesterone, testosterone and oestradiol synthesis were first evident 1 h after hCG, but values peaked at 3-5 h and returned to control levels by 10 h. Increased synthesis of PGE-2 and PGF-2 alpha was not evident until 3 h and peaked at more than 10 h after hCG. Ovulation began between 8 and 10 h after hCG and 83% of animals had ovulated within 12 h. Doses of 90 or 1800 micrograms indomethacin given together with hCG substantially inhibited ovulation and PG synthesis, but only the higher dose inhibited the hCG-induced elevation of progesterone and testosterone synthesis; hCG-induced oestradiol synthesis was not affected by either dose of indomethacin. We conclude that the peak of PG synthesis after hCG treatment related closely to the timing of ovulation; the steroidogenic response to hCG was not blocked by doses of indomethacin sufficient to inhibit synthesis of PGE-2 and PGF-2 alpha by more than 80%.     

Inhibition of prostaglandin biosynthesis during postluteolysis and effects on CL regression, prolactin, and ovulation in heifers

The beginning of postluteolysis (progesterone, <1 ng mL⁻¹) in heifers was targeted by using 8 h after ultrasonic detection of a 25% decrease in CL area (cm²) and was designated Hour 0. Flunixin meglumine (FM; n = 10) to inhibit PGF_2α secretion or vehicle (n = 9) were given intramuscularly at Hours 0, 4, 8, 16, 24, 32, and 40. The dose of FM was 2.5 mg/kg at each treatment. Blood sampling and measurement of the CL and dominant follicle were done every 8 h beginning 14 days postovulation in each group. Blood samples for detection of pulses of PRL and pulses of a metabolite of PGF_2α (PGFM) were obtained every hour for 24 h beginning at Hour 0. Pulse concentrations of both PGFM and PRL were lower in the FM group than in the vehicle group. Concentration of PRL was greatest at the peak of a PGFM pulse. Neither CL area (cm²) nor progesterone concentration differed between groups during Hours 0 to 48 (postluteolysis). Ovulation occurred in nine of nine heifers in the vehicle group and in three of 10 heifers in the FM group. The anovulatory follicles in the FM group grew to 36.2 ± 2.9 mm, and the wall became thickened from apparent luteinization. The hypothesis that PGF_2α was involved in the continued P4 decrease and structural CL regression during postluteolysis was not supported. However, the hypotheses that pulses of PGFM and PRL were temporally related and that systemic FM treatment induced an anovulatory follicle were supported.     

Effects of prostaglandin E2 and DL204 IT,an inhibitor of prostaglandin degradation,on ovulation and ovum transport in the hamster

The effects of 2(3-ethoxyphenyl)-5,6-dihydro-s-triazole-[5,1-a] isoquinoline (L-11204 or DL 204 IT) and PGE₂ on ovulation and ova transport were studied. DI 204 IT was administered in doses of 0.2–25 mg/Kg s.c. on the day of estrus. A small reduction in ovulating follicles was observed 96 hours later, but only at the 5 mg/Kg dose level. At all dose levels, however DL 204 IT caused a dose-related reduction in the number of ova in the oviducts. PGE₂ at a total dose of 2 mg/animal s.c., administered in 4 divided doses over the second and third day of the cycle did not affect ovulation or ova transport. PGE₂ plus DL 204 IT (5 mg/Kg), however, completely blocked ovulation in all but one animal. The animal had one ovulated follicle and a single ova was recovered from its oviduct.     

Differential effects of indomethacin on the sheep ovary: Prostaglandin biosynthesis, intracellular calcium, apoptosis, and ovulation

Cells of the apical wall of the dominant follicle and contiguous ovarian surface epithelium become apoptotic with the approach of ovulation in the sheep. It was hypothesized that indomethacin, an established inhibitor of prostaglandin biosynthesis and ovulation, would protect apical ovarian cells from programmed death. The anovulatory potencies of two systemic doses of indomethacin (200 and 800 mg) were tested in gonadotropin-stimulated ewes. A complete blockade of ovulation occurred at the higher dose of indomethacin. Ovulation was not inhibited by 200 mg indomethacin. Both doses of drug suppressed follicular prostaglandin production below pregonadotropin levels. Immunofluorescence detection of digoxigenin end-labeled (fragmented) DNA was used as a marker of apoptosis among ovarian surface epithelial and granulosa cells recovered from the apical hemisphere of preovulatory ovine follicles. Cellular DNA fragmentation was averted in animals given 800 mg indomethacin, whereas apoptosis ensued after 200 mg. A sustained increase in cytosolic calcium is generally a prerequisite to apoptotic DNA fragmentation and cell death. Indeed, intracellular calcium, detected by fluorescence of fura-2, was elevated in ovarian cells of animals destined to ovulate (controls, 200 mg indomethacin) in comparison to (safeguarded) cells of anovulatory ewes (800 mg indomethacin). These observations provide circumstantial evidence that apical ovarian cell degeneration by calcium-mediated apoptosis is a determinant of follicular instability and rupture, but that these events are unrelated to the gonadotropin-induced rise in prostanoid production characteristic of preovulatory follicles.     

Inhibition of LH-induced and spontaneous meiotic maturation in mouse oocytes by N alpha-tosyl-L-lysine chloromethylketone

The effect of N alpha-tosyl-L-lysine chloromethylketone (TLCK), an inhibitor of trypsin-type proteases, on luteinizing hormone (LH)-induced and spontaneous meiotic maturation and follicular production of cAMP in mice was determined. When follicle-enclosed mouse oocytes were incubated with LH (1 micron/ml), they underwent the breakdown of the germinal vesicle (GVBD). TLCK (0.02-0.5 mM) inhibited LH-induced GVBD in folliculated oocytes. The concentration (0.5 mM) of TLCK that inhibited LH-induced GVBD did not significantly suppress LH-induced cAMP production by follicle cells. The effect of TLCK on spontaneous maturation in cumulus cell-enclosed and denuded oocytes was also determined. TLCK strongly inhibited spontaneous maturation in denuded oocytes only if it was added to the incubation medium for 1-3 h before oocytes were liberated from the follicular tissue. The inhibition of oocyte maturation by TLCK was significantly greater in cumulus cell-enclosed oocytes than in denuded oocytes, either with or without preincubation with TLCK. These results suggest that trypsin-type protease in oocytes participates in the process of meiotic maturation in mouse oocytes.     

Reversal of indomethacin blockade of ovulation in gilts by prostaglandins

Prepubertal gilts given 750 IU pregnant mares′ serum gonadotropin (PMSG) followed 72 h later by 500 IU human chorionic gonadotropin (hCG) to induce follicular growth and ovulation fail to ovulate when 10 mg/kg indomethacin (INDO) is injected 24 h after hCG administration. This study examines the effects of administration of exogenous prostaglandins F_2α and E₂ (PGF_2α and PGE₂) alone or in combination, and at various times prior to the expected time of ovulation, on the INDO blockade of ovulation in PMSG/hCG-treated gilts. Occurrence of ovulation was determined by visual observation at laparotomy 48 h after hCG. When 5 mg or 10 mg PGF_2α was injected at each of 38, 40 and 42 h after hCG injection, 63% and 79%, respectively, of preovulatory follicles ovulated. In contrast, injection of 5 mg PGE₂ or 5 mg PGE₂ plus 5 mg PGF_2α induced ovulation in 0% and 24% of preovulatory follicles, respectively. In control groups, 100% of folicles in PMSG/hCG-treated gilts ovulated whereas none did so in PMSG/hCG/INDO-treated animals. These results indicate that administration of PGF_2α can induce ovulation in the PMSG/hCG/INDO-treated prepubertal gilt and suggest that PGE₂ is ineffective and may be antagonistic to PGF_2α in overcoming the ovulation blocking effect of INDO.     

Pulvomycin,an inhibitor of prokaryotic protein biosynthesis

Antibiotic 1063-Z isolated from culture fluids of Streptoverticillium mobaraense was identified as pulvomycin. Pulvomycin was observed to inhibit protein biosynthesis in growing cells of Bacillus brevis. The poly(U)-directed poly(Phe) synthesis in cell-free systems of Bacillus brevis and Escherichia coli was highly susceptible to the antibiotic. Pulvomycin did not affect the transfer of Phe to tRNA. The results suggest that the target of pulvomycin action is the polypeptide chain elongation.     

The metabolism of inhibitor of flowering and prostaglandin biosynthesis, acetylsalicylic acid, in Pharbitis nil cotyledons

Acetylsalicylic acid, which applied to cotyledons of the short day plant Pharbitis nil prior to an inductive 16-h dark period inhibits flowering by 90 %, is converted to salicylic acid and to a lesser extent to gentisic acid in the cotyledons during this 16-h dark period. Our results confirmed that salicylic acid and gentisic acid are responsible for the inhibition of flowering. They also inhibit prostaglandin biosynthesis. This revised version was published online in July 2006 with corrections to the Cover Date.